DYSFUNCTIONAL ANTIGEN-SPECIFIC CD8 T CELLS IN THE TUMOR

MICROENVIRONMENT

CROSS-REFERENCE TO RELATED APPLICATIONS The present invention claims priority to U. S. Provisional Patent Application Serial

Number 62/447, 199, filed January 17, 2017, which is incorporated by reference in its entirety.

STATEMENT OF GOVERNMENT SUPPORT

This invention was made with government support under Grant No. R01 CA161005 awarded by the National Institutes of Health. The government has certain rights in the invention.

FIELD

Provided herein are compositions and methods for detecting and/or targeting dysfunctional tumor antigen-specific CD8 ⁺ T cells in the tumor microenvironment for diagnostic, therapeutic and/or research applications. In particular, dysfunctional tumor antigen-specific CD8 ⁺ T cells are detected and/or targeted via their expression of cell surface receptors described herein, such as 4- IBB, LAG-3, or additional markers that correlate with 4-1BB and LAG-3 expression, such as markers differentially expressed on the surface of the T cells.

BACKGROUND

The immune system plays a critical role in protecting the host from cancer (Vesely et al., 2011 ; incorporated by reference in its entirety). Innate sensing of tumors leads to an adaptive T cell response through the presentation of tumor-associated antigens (TAAs) derived from mutations and epigenetic changes that contribute to carcinogenesis (Gajewski et al., 2013; incorporated by reference in its entirety). Spontaneously -primed CD8+ T cells home to tumor sites in mouse tumor models (Harlin et al, 2009; Fuertes et al., 2011 ;

incorporated by reference in their entireties) and in a subset of patients with advanced cancer (Harlin et al, 2006; incorporated by reference in its entirety). These tumor-infiltrating lymphocytes (TIL) have the ability to recognize tumor antigens and are believed to contribute to tumor control in cancer patients, based on the correlation between activated CD 8+ T cell infiltration with improved prognosis and response to immunotherapy (Fridman et al, 2012; Tumeh et al, 2014; incorporated by reference in their entireties). However, without additional manipulation, this endogenous anti-tumor response is usually not sufficient to mediate complete rejection of an established tumor (Gajewski, 2007b; Pardoll, 2012; Baitsch et al, 2011; Gajewski et al., 2006; Larkin et al., 2015). Data accumulated over the past several years have indicated that tumors with spontaneous anti-tumor T cell responses have high expression of immune-inhibitory pathways that subvert the effector phase of the response. These include PD-Ll/PD-1 interactions (Pardoll, 2012; incorporated by reference in its entirety), recruitment of CD4+Foxp3+ regulatory T (Treg) cells (Gajewski, 2007a; incorporated by reference in its entirety), and metabolic dysregulation by indoleamine-2,3- di oxygenase (IDO) (Spranger et al, 2013; incorporated by reference in its entirety).

However, even when CD8+ T cells specific for tumor antigens are isolated from tumors, away from these extrinsic immune inhibitory factors, they still show altered functional properties ex vivo (Harlin et al, 2006; Baitsch et al, 2011 ; incorporated by reference in their entireties).

Expression of PD-1 has been described to identify tumor-specific exhausted T cells (Ahmadzadeh et al, 2009; Fourcade et al, 2012; Wu et al, 2014; Gros et al, 2014;

incorporated by reference in their entireties). However, T cells expressing PD-1 in the context of chronic infection can still retain effector function (Wherry and Kurachi, 2015; incorporated by reference in its entirety), and PD-1 is not required for the induction of T cell exhaustion (Odorizzi et al, 2015; incorporated by reference in its entirety). In addition to PD-1, several additional co-inhibitory receptors, including CD223 (LAG-3), CD244 (2B4), T-cell immunoreceptor with Ig and ITIM domains (TIGIT), hepatitis A virus cellular receptor 2 (TIM-3), and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), are also be expressed on dysfunctional T cells and expression of a greater number of inhibitory receptors has been correlated with diminished cytokine secretion (in particular IFN-g and TNF-a) as well as proliferative capacity (Blackburn et al, 2009; incorporated by reference in its entirety). Expression of these receptors has been observed in both viral and cancer models, however, a complete analysis of both co-inhibitory and co-stimulatory receptors on the same population is lacking in the tumor setting.

SUMMARY

Provided herein are compositions and methods for detecting and/or targeting dysfunctional tumor antigen-specific CD8 ⁺ T cells in the tumor microenvironment for diagnostic, therapeutic and/or research applications. In particular, dysfunctional tumor antigen-specific CD8 ⁺ T cells are detected and/or targeted via their expression of cell surface receptors described herein, such as 4- IBB, LAG-3, or additional markers that correlate with 4-1BB and LAG-3 expression, such as markers differentially expressed on the surface of the T cells (e.g., PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nml, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM and Sema7a).

In some embodiments, provided herein are methods of treating a subject with cancer comprising administering an agent that specifically targets dysfunctional tumor antigen- specific CD8 ⁺ T cells. In some embodiments, the subject suffers from a solid tumor cancer. In some embodiments, the tumor allows T cell infiltration, but is resistant to

immunotherapies. In some embodiments, the tumor environment comprises dysfunctional tumor antigen-specific CD8 ⁺ T cells. In some embodiments, contacting the dysfunctional tumor antigen-specific CD8 ⁺ T cells with an anti-4-lBB and/or anti-LAG3 agent. In some embodiments, the anti-4-lBB and/or anti-LAG3 agent is an antibody, antibody fragment, or antibody mimetic molecule. In some embodiments, methods further comprise coadministration of an additional therapeutic agent. In some embodiments, the additional therapeutic agent is a chemotherapeutic or an immunotherapeutic agent. In some

embodiments, the additional therapeutic agent is an immunotherapeutic agent selected from the list consisting of cell-based therapies, monoclonal antibody (mAb) therapy, cytokine therapy, and adjuvant treatment. In some embodiments, the immunotherapeutic agent is a mAb therapy selected from the list consisting of anti-CTLA-4 monoclonal antibodies and/or anti-PD-Ll monoclonal antibodies. In some embodiments, the immunotherapeutic agent is a cell-based therapy selected from the list consisting of dendritic-cell therapy and T-cell therapy. In some embodiments, the additional therapeutic agent targets one of the markers/receptors listed in Table 2. In some embodiments, the additional therapeutic targets a marker/receptor expressed on the surface of the T cells. In some embodiments, the additional therapeutic targets PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nml, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM or Sema7a. In some embodiments, the additional therapeutic agent targets Nrnl, Sema7a, or CRTAM.

In some embodiments, provided herein are methods of treating a subject with cancer comprising administering a therapeutic agent that specifically targets dysfunctional tumor antigen-specific CD8 ⁺ T cells, wherein the agent targets one of the receptors listed in Table 2. In some embodiments, the therapeutic targets a marker/receptor expressed on the surface of the T cells. In some embodiments, the therapeutic targets PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nrnl, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM or Sema7a. In some embodiments, the therapeutic agent targets Nrnl, Sema7a, or CRTAM. In some embodiments, the therapeutic agent is an antibody, antibody fragment, or antibody mimetic molecule that binds the target marker/receptor. In some embodiments, the therapeutic agent is an anti-Nrn antibody, antibody fragment, or antibody mimetic molecule. In some embodiments, the therapeutic agent is an anti-Sema7a antibody, antibody fragment, or antibody mimetic molecule. In some embodiments, the therapeutic agent is an anti-CRTAM antibody, antibody fragment, or antibody mimetic molecule.

In some embodiments, provided herein are compositions comprising: (a) one or more of an anti-4-lBB agent, an anti-LAG-3 agent, an anti-Nrnl agent, an anti-Sema7a agent, and an anti-CRTAM agent; and (b) an immunotherapeutic agent, said composition formulated for therapeutic delivery to a subject. In some embodiments, the anti-4-lBB agent, anti-LAG-3 agent, anti-Nrnl agent, anti-Sema7a agent, and/or anti-CRTAM agent is an antibody, antibody fragment, or antibody mimetic molecule.

In some embodiments, provided herein are compositions comprising: (a) an agent that targets and/or binds one of PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nrnl, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A; and (b) an immunotherapeutic agent, said composition formulated for therapeutic delivery to a subject.

In some embodiments, provided herein are methods comprising: (a) testing CD8 ⁺ T cells from a cell population to determine whether the CD8+ T Cells co-express LAG-3 and 4- 1BB; and (b) administering one or more agents that target and/or bind one of PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nrnl, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A. In some embodiments, the agent is an anti-Nrnl agent, an anti-Sema7a agent, and an anti-CRTAM agent. In some embodiments, the anti-Nrnl agent, anti-Sema7a agent, and/or anti-CRTAM agent is an antibody, antibody fragment, or antibody mimetic molecule. In some embodiments, testing is performed in vitro.

In some embodiments, provided herein are methods of identifying dysfunctional T cells by testing said cells for co-expression of 4-1BB and LAG-3. In some embodiments, provided herein are methods of identifying dysfunctional T cells by testing said cells for expression of one or more of the markers/receptors of Table 2 (e.g., a T-cell surface marker/receptor (e.g., PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nrnl, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, TMEM126A). BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 A-J. Co-expression of 4- IBB and LAG-3 identifies a significant fraction of the CD8 ⁺ TIL compartment found in progressing tumors. (A) Representative analysis of 4- IBB and LAG-3 expression on CD8 ⁺ T cells from B16.SIY tumors and the spleen and TdLN from tumor bearing mice on day 7, 14 and 21 after s.c. tumor inoculation. (B-D)

Longitudinal summary of the composition, n=5; four to five independent experiments per time point, (C) absolute cell number, n=5; seven to nine independent experiments per time point, and (D) cellular density of the CD8 ⁺ 4-1BB/LAG-3 TIL subpopulations, n=5; two to five independent experiments per time point. Absolute cell numbers were determined by acquiring the complete tumor sample by flow cytometery. (E) Day 14 summary of the proportion of the CD8 ⁺ 4-1BB/LAG-3 TIL subpopulations that are Ki67 ⁺ . n=3-5; two independent experiments. (F) Summary of BrdU uptake on day 13 in the CD8 ⁺ 4-1BB/LAG- 3 TIL subpopulations after a 24 hour BrdU pulse. n=5; three independent experiments. (G-I) Representative flow plots (G and H) and summary (I) of the 4-1BB/LAG-3 populations in other tumor models. Mice were inoculated with 2xl0 ⁶ C1498.SIY, MC38.SIY, EL4.SIY, B16 Parental, MC57.SIY or 1969.SIY subcutaneously and analyzed for 4-1BB and LAG-3 expression on day 14 after tumor inoculation. n=3-5; two to 5 independent experiments for each time point. (J) Mice were inoculated on both flanks with 2xl0 ⁶ MC57.SIY or B16.SIY, at indicated time points tumors from each mouse were pooled and analyzed for co-expression of 4-1BB and LAG-3 in the CD8 ⁺ TIL compartment. n=3-5; two independent experiments for each time point. All error bars indicate ± SEM. *:P < 0.05, **:P<0.01, ***:P<0.001. A two- way ANOVA with Bonferroni post-hoc test was used for (B, C, D, H) longitudinal studies and Kruskal-Wallis (non-parametric) test was used for (E and F) analysis at one time-point.

Figure 2A-G. Egr2 and a component of the Egr2 -transcriptional network are enriched in 4-lBB ⁺ LAG-3 ⁺ CD8 ⁺ TILs. (A) Representative flow plot and summary of Egr2 ^EGFP expression. Egr2 ^EGFP mice were inoculated with 2xl0 ⁶ B16.SIY tumors s.c. CD8 ⁺ T cells from the tumor, TdLN and spleen were analyzed for Egr2 ^EGFP expression on day 7 and day 14. n=4-5; two-independent experiments. (B) Expression of Egr2 target genes (Zheng et al, 2013). CD8 ⁺ TILs from day 14 tumor bearing mice were sorted based on high or low expression of Egr2 ^EGFP and analyzed directly for expression of Egr2 targets by qRT-PCR. Two tumors on opposite flanks pooled per mouse. n=3; two independent experiments. (C) Representative flow plots and summary of the 4-1BB/LAG-3 subpopulations in CD8 ⁺ Egr2GFP ^hl and Egr2GFP ^l0 TILs on day 7 and 14. n=4-5. Two-independent experiments per time point. (D) Expression of Egr2 targets in the 4- IBB LAG-3 and 4- IBB LAG-3 subpopulations. The subpopulations were sorted and analyzed directly for the expression of targets by qRT-PCR. Two tumors on opposite flanks pooled per mouse. n=4; two- independent experiments. (E) Egr2 ^flox/flox x pLCKCreERT2 x YFP-Rosa26 mice given 5 doses of tamoxifen by gavage and inoculated 3 days later with 2xl0 ⁶ B16.SIY cells. YFP ⁺ or YFP CD8 ⁺ TILs were sorted and analyzed for Egr2 transcript directly and after in vitro stimulation. Two tumors on opposite flanks pooled per mouse. n=3; two independent experiments. (F) Representative flow plots and summary of 4-1BB/LAG-3 co-expression in YFP ⁺ or YFP CD8 ⁺ TILs on day 7 and 14. n=3; two independent experiments. (G)

Expression of Egr3 and Hifl in Egr2GFP ^hi and Egr2GFP ^l0 from day 7 CD8 ⁺ TILs isolated from Egr2GFP mice. n=5; two-independent experiments. Error bars indicate ± SEM. *:P < 0.05, **:P<0.01, ***:P<0.001. A two-way ANOVA with Bonferroni post-hoc test was used for longitudinal studies (A and C) and a Mann- Whitney test was used to compute significance in (B, D, E, F and G).

Figure 3A-H. Co-expression of 4-lBB and LAG-3 identifies tumor antigen-specific

TILs in progressing tumors. (A) Representative CDR3 distributions from the different 4- 1BB/LAG-3 subpopulations and CD8 ⁺ T cells isolated from the spleen. Boxed regions represent dominant peaks in the 4-lBB ⁺ LAG-3 ⁺ CD8 ⁺ TIL subpopulation. (B) As a measure of skewness, the Hamming Distance (HD) for each νβ spectratype was calculated between each TIL subpopulation and CD8 ⁺ T cell spleen population within the same mouse. As a control the HDs from CD8 ⁺ splenocyte populations between mice (grey bar) were calculated. n=3; one independent experiment. (C-D) Representative flow analysis of the 4-1BB/LAG-3 subpopulation in H-2K ^b /SIY ⁺ and H-2K ^b /SIY CD8 ⁺ TILs on day 14 after B16.SIY and MC38.SIY or (D) MC57.SIY and 1969.SIY tumor inoculation. n=3-4; three to five independent experiments. (E) Summary of the composition of H-2K ^b /SIY ⁺ and H-2K ^b /SIY CD8 ⁺ TILs co-expressing 4-lBB and LAG-3 comparing B16.SIY, MC38.SIY, MC57.SIY and 1969. SIY tumors on day 14 after tumor inoculation. n=5; three to four independent experiments. (F-H) On day 7 after tumor inoculation lxlO ⁶ P14/CD45.2 and 2C/CD45.1/2 Tg T cells were adoptively transferred, via tail vein, into CD45.1 congenic tumor bearing hosts and analyzed for the (F) total number of recovered cells in the tumor, (G and H) profile of 4-lBB and LAG-3 expression in 2C, P14 and host CD8 ⁺ TILs. n=5; two-independent experiments. All error bars indicate ± SEM. *:P < 0.05, **:P<0.01, ***:P<0.001. A Kruskal- Wallis (non-parameteric) test was used for (B) spectratype analysis and (E and F) H-2K ^b /SIY analysis. A two-way ANOVA with Bonferroni post-hoc test was used for (H) 2C, Host and P14 composition analysis.

Figure 4A-G. Co-expression of 4-1BB and LAG-3 but not PD-1 define dysfunctional CD8 ⁺ TILs with diminished IL-2. (A and B) Sorted cells from day 14 B16.SIY tumor bearing mice were stimulated in vitro with anti-CD3s and anti-CD28 for 12 hours and analyzed for (A) 11-2 transcript by qRT-PCR and (B) IL-2 protein by ELISA. Two tumors on opposite flanks pooled per mouse. n=4-5; three independent experiments. (C) Egr2GFP ^hi and

Egr2GFP ^l0 TILs were sorted from day 14 B16.SIY tumor bearing Egr2 mice and stimulated in vitro for 12 hours and analyzed for 11-2 transcript by qRT-PCR. Two tumors on opposite flanks pooled per mouse. n=5; two independent experiments. (D) On day 7 after tumor inoculation lxlO ⁶ 2C/CD45.1/2 Tg T cells were transferred into mice, 7 days later host 4-lBB ⁺ LAG-3 ⁺ T cells sorted from the tumor and 2C T cells sorted from the tumor or TdLN were stimulated in vitro and analyzed for expression of 11-2 transcript by qRT-PCR. Two tumors on opposite flanks pooled per mouse. n=3; two independent experiments. (E and F) Representative flow analysis of PD-1 expression on 4-1BB/LAG-3 CD8 ⁺ TIL subpopulations and (F) summary of the composition of the 4-1BB AG-3 PD-1 ⁺ subpopulation in the CD8 ⁺ TIL compartment on day 14 and 21. n=5; three independent experiments. (G) 4-1BB LAG- 3 PD-r and LAG-3 ⁺ 4-lBB ⁺ CD8 ⁺ TILs were sorted from day 14 tumor bearing mice, stimulated in vitro and analyzed for 11-2 transcript by qRT-PCR. Two tumors on opposite flanks pooled per mouse. n=3; two independent experiments. All error bars indicate ± SEM. *:P < 0.05, **:P<0.01, ***:P<0.001 ****:P<0.0001. A Kruskal-Wallis (non-parametric) test was used for analysis of multiple comparisons (A, B, and D) and a Mann-Whitney test was used for pair-wise comparisons (C and G).

Figure 5A-E. Dysfunctional CD8 ⁺ TILs retain IFN-γ production, cytolytic capacity and produce Treg-recruiting chemokines. (A) Longitudinal analysis CD8 ⁺ TIL subpopulation cytokine production capacity. CD8 ⁺ TIL subpopulations were sorted and stimulated with anti- CD3s and anti-CD28 for 10-12 hours and the concentration of IL-2, IFN-γ and TNF-a was measured. Concentration was normalized to cell number. Two tumors on opposite flanks pooled for day 7 and 14. n=4-5; two-independent experiments. (B) Ifn-y Tnf-a and Gzmb transcript levels in the 4-1BB/LAG-3 subpopulations analyzed directly ex vivo. Two tumors on opposite flanks pooled per mouse. n=3-5; three-independent experiments. (C)

Representative flow plot and summary of IFN-γ production analyzed directly ex vivo.

Briefly, 100 μΐ of PBS containing 2 mg/mL GolgiPlug was injected intratumorally on day 14 after tumor inoculation. 8 hours later TILs were isolated. All steps were performed on ice with media containing 1 mg/mL GolgiStop until fixation. n=5; two independent experiments. (D) CD8 ⁺ TIL subpopulations at indicated time points were sorted and plated with 50,000 P815 target cells and 1 μg/mL anti-CD3s. Lysed target cells were measured by positive staining for propidium iodide and/or live/dead fixable viability dye. P815 target cells plated without CTLs were used as a negative control (black bar). Primed OTI cells were used as a positive control. Tumors from 10 mice with 2 tumors on opposite flank were pooled to obtain sufficient quantities of CD8 ⁺ TILs. Data are representative of three independent experiments. (E) Cell and Ccl22 transcript levels in the 4-1BB/LAG-3 subpopulations analyzed directly ex vivo by qRT-PCR. n=4; two independent experiments. *:P < 0.05, **:P<0.01, ***:P<0.001, ****:P<0.0001. A Kruskal-Wallis (non-parametric) test was used for (A-C, E) cytokine/chemokine analysis and a two-way ANOVA with Bonferroni post-/roc test was used for (D) cytolytic assay.

Figure 6A-D. Dysfunctional CD8 ⁺ TILs express a wide range of co-inhibitory and co-stimulatory receptors. (B) Gene expression profile of cell surface receptors in the 4- 1BB/LAG-3 CD8 ⁺ TIL subsets. Probe sets that revealed a 1.5-fold increase in the 4- lBB ⁺ LAG-3 ⁺ population relative to the 4-1BB AG-3TD-1 population are displayed. Columns show the log ₂ -transformed signal intensity. (C) Longitudinal study of selected up- regulated cell surface receptors. Flow plots are representative of the CD8 ⁺ TIL subsets on day 14. n=5; two to five independent experiments for each time point. (D) Representative flow plot and summary of KLRG-1 and IL-7Ra expression among the 4-1BB/LAG-3 subpopulations on day 14 after tumor inoculation. n=5; two independent experiments. *:P < 0.05, **:P<0.01, ***:P<0.001, ****:P<0.0001. A two-way ANOVA with Bonferroni post- hoc test was used for all analyses.

Figure 7A-G. Anti-4-lBB and anti-LAG-3 acts synergistically to control tumor outgrowth and restore TIL function. (A) Tumor outgrowth measured in mm ² . Arrows indicate on which days mice received antibody therapy. Statistical significance at indicate time points is in comparison to anti-4-lBB + anti-LAG-3 treatment. n=5; two independent experiments. (B) Composition of H-2K ^b /SIY ⁺ CD8 ⁺ TILs on day 14. Mice received antibody doses (100 μg each) on days 7,10, 13 and 16. n=5; two independent experiments. (C-F) Representative flow plot and summary of NRP1/2B4 (C and E) and KLRG-l/IL-7Ra (D and F) expression in H-2K ^b /SIY ⁺ CD8 ⁺ TILs without FTY720 (C and D) and with FTY720 (E and F) on day 14 after tumor inoculation. Mice received antibody treatment as in (A and B) and FTY720 was administered at a dose of 25 μg/mouse by gavage starting one day before treatment and continuing one dose per day until analysis (day 6 to day 13). n=5; two-independent experiments. (G) IL-2 production after treatment. Sorted cells from treated or untreated day 14 B16.SIY tumor bearing mice were stimulated in vitro for 12 hours and analyzed for 11-2 transcript by qRT-PCR. Protein concentration was determined by the bead-based

LEGENDplex immunoassay and normalized to cell number. Two tumors on opposite flanks pooled per mouse. n=2-3; two independent experiments. A two-way ANOVA with

Bonferroni post-/roc test was used for all analyses. *:P < 0.05, **:P<0.01, ***:P<0.001.

Figure 8. Spectratype graphs used in the analysis in Figure 3B.

Figure 9. CD3+ T cells on day 14 after FTY720 administration.

Figure 10A-B. Statistical analysis of the cross-study comparison of gene expression profiles. (A) Rank-Rank Hypergeometric plots of each pair-wise comparison. (B) Pair-wise correlation of expression values between each data set. Rho (p) is the spearman rank correlation coefficient.

Figure 11A-E. Nrnl, CRT AM and Sema7a are regulators of anti-tumor immunity. (A) Tumor growth measured in mm2. Nrnl ^_/~ or Sema7a and littermate control mice were engrafted with 2 x 106 B16.SIY cells subcutaneously. (B) Gene expression analysis of Nrnl in T cell subsets of the spleen, TdLN and Tumor. (C) Representative flow plot and summary of IFN-g production of WT, Nrnl ^"7" or (D) CRTAM ⁷ 2C T cells on day 7. Briefly, on the same day as tumor inoculation, lxlO ⁶ Cell Trace Violet-labeled 2C T cells were transferred into mice by tail vein injection. On day 7, whole TdLN suspensions were restimulated with SIY peptide for 12 hours and analyzed for cell trace dilution and IFN-g production. (E) Mice that received lxl 0 ⁶ Nrnl ^{_ ~} 2C T cells are more likely to exhibit complete tumor control compared to mice that received the same number of WT 2C T cells. Adoptive transfer of T cells was performed the same way as in (C).

Figure 12. Exemplary experimental protocol and data.

DEFINITIONS

Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of embodiments described herein, some preferred methods, compositions, devices, and materials are described herein. However, before the present materials and methods are described, it is to be understood that this invention is not limited to the particular molecules, compositions, methodologies or protocols herein described, as these may vary in accordance with routine experimentation and optimization. It is also to be understood that the terminology used in the description is for the purpose of describing the particular versions or embodiments only, and is not intended to limit the scope of the embodiments described herein.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. However, in case of conflict, the present specification, including definitions, will control. Accordingly, in the context of the embodiments described herein, the following definitions apply.

As used herein and in the appended claims, the singular forms "a", "an" and "the" include plural reference unless the context clearly dictates otherwise. Thus, for example, reference to "an antibody" is a reference to one or more antibodies and equivalents thereof known to those skilled in the art, and so forth.

As used herein, the term "comprise" and linguistic variations thereof denote the presence of recited feature(s), element(s), method step(s), etc. without the exclusion of the presence of additional feature(s), element(s), method step(s), etc. Conversely, the term "consisting of and linguistic variations thereof, denotes the presence of recited feature(s), element(s), method step(s), etc. and excludes any unrecited feature(s), element(s), method step(s), etc., except for ordinarily-associated impurities. The phrase "consisting essentially of denotes the recited feature(s), element(s), method step(s), etc. and any additional feature(s), element(s), method step(s), etc. that do not materially affect the basic nature of the composition, system, or method. Many embodiments herein are described using open "comprising" language. Such embodiments encompass multiple closed "consisting of and/or "consisting essentially of embodiments, which may alternatively be claimed or described using such language.

As used herein, the term "subject" broadly refers to any animal, including but not limited to, human and non-human animals (e.g., dogs, cats, cows, horses, sheep, poultry, fish, crustaceans, etc.). As used herein, the term "patient" typically refers to a subject that is being treated for a disease or condition (e.g., cancer, solid tumor cancer, etc.).

As used herein, an "immune response" refers to the action of a cell of the immune system (e.g., T lymphocytes, B lymphocytes, natural killer (NK) cells, macrophages, eosinophils, mast cells, dendritic cells, neutrophils, etc.) and soluble macromolecules produced by any of these cells or the liver (including antibodies, cytokines, and complement) that results in selective targeting, binding to, damage to, destruction of, and/or elimination from a subj ect of invading pathogens, cells or tissues infected with pathogens, or cancerous or other abnormal cells. As used herein, the term "immunoregulator" refers to a substance, an agent, a signaling pathway or a component thereof that regulates an immune response. "Regulating," "modifying" or "modulating" an immune response refers to any alteration in a cell of the immune system or in the activity of such cell. Such regulation includes stimulation or suppression of the immune system which may be manifested by an increase or decrease in the number of various cell types, an increase or decrease in the activity of these cells, or any other changes which can occur within the immune system. Both inhibitory and stimulatory immunoregulators have been identified, some of which may have enhanced function in the cancer microenvironment.

As used herein, the term "immunotherapy" refers to the treatment or prevention of a disease or condition by a method comprising inducing, enhancing, suppressing or otherwise modifying an immune response.

As used herein, "potentiating an endogenous immune response" means increasing the effectiveness or potency of an existing immune response in a subject. This increase in effectiveness and potency may be achieved, for example, by overcoming mechanisms that suppress the endogenous host immune response or by stimulating mechanisms that enhance the endogenous host immune response.

As used herein, the term "antibody" refers to a whole antibody molecule or a fragment thereof (e.g., fragments such as Fab, Fab', and F(ab')2), unless otherwise specified (e.g., "whole antibody," "antibody fragment"). An antibody may be a polyclonal or monoclonal antibody, a chimeric antibody, a humanized antibody, a human antibody, etc.

A native antibody typically has a tetrameric structure. A tetramer typically comprises two identical pairs of polypeptide chains, each pair having one light chain (in certain embodiments, about 25 kDa) and one heavy chain (in certain embodiments, about 50-70 kDa). In a native antibody, a heavy chain comprises a variable region, VH, and three constant regions, CHI, CH2, and CH3- The VH domain is at the amino-terminus of the heavy chain, and the CH3 domain is at the carboxy -terminus. In a native antibody, a light chain comprises a variable region, VL, and a constant region, CL. The variable region of the light chain is at the amino-terminus of the light chain. In a native antibody, the variable regions of each light/heavy chain pair typically form the antigen binding site. The constant regions are typically responsible for effector function.

In a native antibody, the variable regions typically exhibit the same general structure in which relatively conserved framework regions (FRs) are joined by three hypervariable regions, also called complementarity determining regions (CDRs). The CDRs from the two chains of each pair typically are aligned by the framework regions, which may enable binding to a specific epitope. From N-terminus to C-terminus, both light and heavy chain variable regions typically comprise the domains FR1 , CDR1 , FR2, CDR2, FR3, CDR3 and FR4. The CDRs on the heavy chain are referred to as HI , H2, and H3, while the CDRs on the light chain are referred to as LI , L2, and L3. Typically, CDR3 is the greatest source of molecular diversity within the antigen-binding site. H3, for example, in certain instances, can be as short as two amino acid residues or greater than 26. The assignment of amino acids to each domain is typically in accordance with the definitions of Kabat et al. (1991) Sequences of Proteins of Immunological Interest (National Institutes of Health, Publication No. 91-3242, vols. 1 -3, Bethesda, Md.); Chothia, C, and Lesk, A. M. (1987) J. Mol. Biol. 196:901-917; or Chothia, C. et al. Nature 342: 878-883 (1989). In the present application, the term "CDR" refers to a CDR from either the light or heavy chain, unless otherwise specified.

As used herein, the term "heavy chain" refers to a polypeptide comprising sufficient heavy chain variable region sequence to confer antigen specificity either alone or in combination with a light chain.

As used herein, the term "light chain" refers to a polypeptide comprising sufficient light chain variable region sequence to confer antigen specificity either alone or in combination with a heavy chain.

As used herein, when an antibody or other entity "specifically recognizes" or

"specifically binds" an antigen or epitope, it preferentially recognizes the antigen in a complex mixture of proteins and/or macromolecules, and binds the antigen or epitope with affinity which is substantially higher than to other entities not displaying the antigen or epitope. In this regard, "affinity which is substantially higher" means affinity that is high enough to enable detection of an antigen or epitope which is distinguished from entities using a desired assay or measurement apparatus. Typically, it means binding affinity having a binding constant (K _a ) of at least 10 ⁷ M ^"1 (e.g., >10 ⁷ M ^"1 , >10 ⁸ M ^"1 , >10 ⁹ M ^"1 , >10 ¹⁰ M ^"1 , >10 ⁿ M ^"1 , >10 ¹² M ^"1 , >10 ¹³ M ^"1 , etc.). In certain such embodiments, an antibody is capable of binding different antigens so long as the different antigens comprise that particular epitope. In certain instances, for example, homologous proteins from different species may comprise the same epitope.

As used herein, the term "anti-4-lBB antibody" or "4-1BB antibody" refers to an antibody which specifically recognizes an antigen and/or epitope presented by 4-1BB.

Similarly, the terms "anti-LAG-3 antibody" and "LAG-3 antibody" refer to an antibody which specifically recognizes an antigen and/or epitope presented by LAG-3, the terms "anti- Nrnl antibody" and "Nrnl antibody" refer to an antibody which specifically recognizes an antigen and/or epitope presented by Nrnl, the terms "anti-CRTAM antibody" and "CRTAM antibody" refer to an antibody which specifically recognizes an antigen and/or epitope presented by CRTAM, and the terms "anti-Sema7a antibody" and "Sema7a antibody" refer to an antibody which specifically recognizes an antigen and/or epitope presented by Sema7a. Antibodies that recognize epitopes on other molecular entities may be referred to according to a similar scheme (e.g., anti-CTLA-4, anti-PD-Ll, etc.).

As used herein, the term "monoclonal antibody" refers to an antibody which is a member of a substantially homogeneous population of antibodies that specifically bind to the same epitope. In certain embodiments, a monoclonal antibody is secreted by a hybridoma. In certain such embodiments, a hybridoma is produced according to certain methods known to those skilled in the art. See, e.g., Kohler and Milstein (1975) Nature 256: 495-499; herein incorporated by reference in its entirety. In certain embodiments, a monoclonal antibody is produced using recombinant DNA methods (see, e.g., U.S. Pat. No. 4,816,567). In certain embodiments, a monoclonal antibody refers to an antibody fragment isolated from a phage display library. See, e.g., Clackson et al. (1991) Nature 352: 624-628; and Marks et al.

(1991) J. Mol. Biol. 222: 581-597; herein incorporated by reference in their entireties. The modifying word "monoclonal" indicates properties of antibodies obtained from a

substantially -homogeneous population of antibodies, and does not limit a method of producing antibodies to a specific method. For various other monoclonal antibody production techniques, see, e.g., Harlow and Lane (1988) Antibodies: A Laboratory Manual (Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.); herein incorporated by reference in its entirety.

As used herein, the term "antibody fragment" refers to a portion of a full-length antibody, including at least a portion antigen binding region or a variable region. Antibody fragments include, but are not limited to, Fab, Fab', F(ab')2, Fv, scFv, Fd, diabodies, and other antibody fragments that retain at least a portion of the variable region of an intact antibody. See, e.g., Hudson et al. (2003) Nat. Med. 9: 129-134; herein incorporated by reference in its entirety. In certain embodiments, antibody fragments are produced by enzymatic or chemical cleavage of intact antibodies (e.g., papain digestion and pepsin digestion of antibody) produced by recombinant DNA techniques, or chemical polypeptide synthesis.

For example, a "Fab" fragment comprises one light chain and the CHI and variable region of one heavy chain. The heavy chain of a Fab molecule cannot form a disulfide bond with another heavy chain molecule. A "Fab"' fragment comprises one light chain and one heavy chain that comprises additional constant region, extending between the CHI and Cm domains. An interchain disulfide bond can be formed between two heavy chains of a Fab' fragment to form a "F(ab')2" molecule.

An "Fv" fragment comprises the variable regions from both the heavy and light chains, but lacks the constant regions. A single-chain Fv (scFv) fragment comprises heavy and light chain variable regions connected by a flexible linker to form a single polypeptide chain with an antigen-binding region. Exemplary single chain antibodies are discussed in detail in WO 88/01649 and U.S. Pat. Nos. 4,946,778 and 5,260,203; herein incorporated by reference in their entireties. In certain instances, a single variable region (e.g., a heavy chain variable region or a light chain variable region) may have the ability to recognize and bind antigen.

Other antibody fragments will be understood by skilled artisans.

As used herein, the term "chimeric antibody" refers to an antibody made up of components from at least two different sources. In certain embodiments, a chimeric antibody comprises a portion of an antibody derived from a first species fused to another molecule, e.g., a portion of an antibody derived from a second species. In certain such embodiments, a chimeric antibody comprises a portion of an antibody derived from a non-human animal fused to a portion of an antibody derived from a human. In certain such embodiments, a chimeric antibody comprises all or a portion of a variable region of an antibody derived from a non-human animal fused to a constant region of an antibody derived from a human.

A "humanized" antibody refers to a non-human antibody that has been modified so that it more closely matches (in amino acid sequence) a human antibody. A humanized antibody is thus a type of chimeric antibody. In certain embodiments, amino acid residues outside of the antigen binding residues of the variable region of the non-human antibody are modified. In certain embodiments, a humanized antibody is constructed by replacing all or a portion of a complementarity determining region (CDR) of a human antibody with all or a portion of a CDR from another antibody, such as a non-human antibody, having the desired antigen binding specificity. In certain embodiments, a humanized antibody comprises variable regions in which all or substantially all of the CDRs correspond to CDRs of a non- human antibody and all or substantially all of the framework regions (FRs) correspond to FRs of a human antibody. In certain such embodiments, a humanized antibody further comprises a constant region (Fc) of a human antibody.

The term "human antibody" refers to a monoclonal antibody that contains human antibody sequences and does not contain antibody sequences from a non-human animal. In certain embodiments, a human antibody may contain synthetic sequences not found in native antibodies. The term is not limited by the manner in which the antibodies are made. For example, in various embodiments, a human antibody may be made in a transgenic mouse, by phage display, by human B-lymphocytes, or by recombinant methods.

As used herein, the term "natural antibody" refers to an antibody in which the heavy and light chains of the antibody have been made and paired by the immune system of a multicellular organism. For example, the antibodies produced by the antibody -producing cells isolated from a first animal immunized with an antigen are natural antibodies. Natural antibodies contain naturally -paired heavy and light chains. The term "natural human antibody" refers to an antibody in which the heavy and light chains of the antibody have been made and paired by the immune system of a human subject.

Native human light chains are typically classified as kappa and lambda light chains. Native human heavy chains are typically classified as mu, delta, gamma, alpha, or epsilon, and define the antibody's isotype as IgM, IgD, IgG, IgA, and IgE, respectively. IgG has subclasses, including, but not limited to, IgGl, IgG2, IgG3, and IgG4. IgM has subclasses including, but not limited to, IgMl and IgM2. IgA has subclasses including, but not limited to, IgAl and IgA2. Within native human light and heavy chains, the variable and constant regions are typically joined by a "J" region of about 12 or more amino acids, with the heavy chain also including a "D" region of about 10 more amino acids. See, e.g., Fundamental Immunology (1989) Ch. 7 (Paul, W., ed., 2nd ed. Raven Press, N.Y.); herein incorporated by reference in its entirety.

The term "neutralizing antibody" or "antibody that neutralizes" refers to an antibody that reduces at least one activity of a polypeptide comprising the epitope to which the antibody specifically binds. In certain embodiments, a neutralizing antibody reduces an activity in vitro and/or In vivo. In some embodiments, by neutralizing the polypeptide comprising the epitope, the neutralizing antibody inhibits the capacity of the cell displaying the epitope.

As used herein, the term "glycoengineered", as used herein, includes any manipulation of the glycosylation pattern of a naturally occurring or recombinant protein, polypeptide or a fragment thereof.

The term "antigen-binding site" refers to a portion of an antibody capable of specifically binding an antigen. In certain embodiments, an antigen-binding site is provided by one or more antibody variable regions. The term "epitope" refers to any polypeptide determinant capable of specifically binding to an immunoglobulin or a T-cell or B-cell receptor. In certain embodiments, an epitope is a region of an antigen that is specifically bound by an antibody. In certain embodiments, an epitope may include chemically active surface groupings of molecules such as amino acids, sugar side chains, phosphoryl, or sulfonyl groups. In certain embodiments, an epitope may have specific three dimensional structural characteristics (e.g., a

"conformational" epitope) and/or specific charge characteristics.

An epitope is defined as "the same" as another epitope if a particular antibody specifically binds to both epitopes. In certain embodiments, polypeptides having different primary amino acid sequences may comprise epitopes that are the same. In certain embodiments, epitopes that are the same may have different primary amino acid sequences. Different antibodies are said to bind to the same epitope if they compete for specific binding to that epitope.

A "conservative" amino acid substitution refers to the substitution of an amino acid in a polypeptide with another amino acid having similar properties, such as size or charge. In certain embodiments, a polypeptide comprising a conservative amino acid substitution maintains at least one activity of the unsubstituted polypeptide. A conservative amino acid substitution may encompass non-naturally occurring amino acid residues, which are typically incorporated by chemical peptide synthesis rather than by synthesis in biological systems. These include, but are not limited to, peptidomimetics and other reversed or inverted forms of amino acid moieties. Naturally occurring residues may be divided into classes based on common side chain properties, for example: hydrophobic: norleucine, Met, Ala, Val, Leu, and He; neutral hydrophilic: Cys, Ser, Thr, Asn, and Gin; acidic: Asp and Glu; basic: His, Lys, and Arg; residues that influence chain orientation: Gly and Pro; and aromatic: Trp, Tyr, and Phe. Non-conservative substitutions may involve the exchange of a member of one of these classes for a member from another class; whereas conservative substitutions may involve the exchange of a member of one of these classes for another member of that same class.

As used herein, the term "sequence identity" refers to the degree to which two polymer sequences (e.g., peptide, polypeptide, nucleic acid, etc.) have the same sequential composition of monomer subunits. The term "sequence similarity" refers to the degree with which two polymer sequences (e.g., peptide, polypeptide, nucleic acid, etc.) have similar polymer sequences. For example, similar amino acids are those that share the same biophysical characteristics and can be grouped into the families (see above). The "percent sequence identity" (or "percent sequence similarity") is calculated by: (1) comparing two optimally aligned sequences over a window of comparison (e.g., the length of the longer sequence, the length of the shorter sequence, a specified window, etc.), (2) determining the number of positions containing identical (or similar) monomers (e.g., same amino acids occurs in both sequences, similar amino acid occurs in both sequences) to yield the number of matched positions, (3) dividing the number of matched positions by the total number of positions in the comparison window (e.g., the length of the longer sequence, the length of the shorter sequence, a specified window), and (4) multiplying the result by 100 to yield the percent sequence identity or percent sequence similarity. For example, if peptides A and B are both 20 amino acids in length and have identical amino acids at all but 1 position, then peptide A and peptide B have 95% sequence identity. If the amino acids at the non-identical position shared the same biophysical characteristics (e.g., both were acidic), then peptide A and peptide B would have 100% sequence similarity. As another example, if peptide C is 20 amino acids in length and peptide D is 15 amino acids in length, and 14 out of 15 amino acids in peptide D are identical to those of a portion of peptide C, then peptides C and D have 70% sequence identity, but peptide D has 93.3% sequence identity to an optimal comparison window of peptide C. For the purpose of calculating "percent sequence identity" (or "percent sequence similarity") herein, any gaps in aligned sequences are treated as mismatches at that position.

The term "effective dose" or "effective amount" refers to an amount of an agent, e.g., an antibody, that results in the reduction of symptoms in a patient or results in a desired biological outcome. In certain embodiments, an effective dose or effective amount is sufficient to treat or reduce symptoms of a disease or condition.

As used herein, the terms "administration" and "administering" refer to the act of giving a drug, prodrug, or other agent, or therapeutic to a subject or in vivo, in vitro, or ex vivo cells, tissues, and organs. Exemplary routes of administration to the human body can be through space under the arachnoid membrane of the brain or spinal cord (intrathecal), the eyes (ophthalmic), mouth (oral), skin (topical or transdermal), nose (nasal), lungs (inhalant), oral mucosa (buccal), ear, rectal, vaginal, by injection (e.g., intravenously, subcutaneously, intratumorally, intraperitoneally, etc.) and the like.

The term "treatment" encompasses both therapeutic and prophylactic/preventative measures unless otherwise indicated. Those in need of treatment include, but are not limited to, individuals already having a particular condition as well as individuals who are at risk of acquiring a particular condition or disorder (e.g., those having a genetic or epigenetic predisposition; based on age, gender, lifestyle, etc.). The term "treating" refers to

administering an agent to a subject for therapeutic and/or prophylactic/preventative purposes.

A "therapeutic agent" refers to an agent that may be administered In vivo to bring about a therapeutic and/or prophylactic/preventative effect.

A "therapeutic antibody" refers to an antibody that may be administered In vivo to bring about a therapeutic and/or prophylactic/preventative effect.

As used herein, the terms "co-administration" and "co-administering" refer to the administration of at least two agent(s) or therapies to a subject. In some embodiments, the coadministration of two or more agents or therapies is concurrent. In other embodiments, a first agent/therapy is administered prior to a second agent/therapy. Those of skill in the art understand that the formulations and/or routes of administration of the various agents or therapies used may vary. The appropriate dosage for co-administration can be readily determined by one skilled in the art. In some embodiments, when agents or therapies are coadministered, the respective agents or therapies are administered at lower dosages than appropriate for their administration alone. Thus, co-administration is especially desirable in embodiments where the co-administration of the agents or therapies lowers the requisite dosage of a potentially harmful (e.g., toxic) agent(s), and/or when co-administration of two or more agents results in sensitization of a subject to beneficial effects of one of the agents via co-administration of the other agent.

As used herein, the term pharmaceutical composition" refers to the combination of an active agent (e.g., binding agent) with a carrier, inert or active, making the composition especially suitable for diagnostic or therapeutic use in vitro, in vivo or ex vivo.

The terms "pharmaceutically acceptable" or "pharmacologically acceptable," as used herein, refer to compositions that do not substantially produce adverse reactions, e.g., toxic, allergic, or immunological reactions, when administered to a subj ect.

As used herein, the term "pharmaceutically acceptable carrier" refers to any of the standard pharmaceutical carriers including, but not limited to, phosphate buffered saline solution, water, emulsions (e.g., such as an oil/water or water/oil emulsions), and various types of wetting agents, any and all solvents, dispersion media, coatings, sodium lauryl sulfate, isotonic and absorption delaying agents, disintigrants (e.g., potato starch or sodium starch glycolate), and the like. The compositions also can include stabilizers and

preservatives. For examples of carriers, stabilizers and adjuvants, see, e.g., Martin,

Remington's Pharmaceutical Sciences, 15th Ed., Mack Publ. Co., Easton, Pa. (1975), incorporated herein by reference in its entirety. As used herein, a "diagnostic" or "diagnostic test" includes the detection,

identification, or characterization of a disease state or condition of a subject. For example, a disease or condition may be characterized to determine the likelihood that a subject with a disease or condition will respond to a particular therapy, determine the prognosis of a subject with a disease or condition (or its likely progression or regression), determine the effect of a treatment on a subject with a disease or condition, or determine a future treatment course of action.

DETAILED DESCRIPTION

Provided herein are compositions and methods for detecting and/or targeting dysfunctional tumor antigen-specific CD8 ⁺ T cells in the tumor microenvironment for diagnostic, therapeutic and/or research applications. In particular, dysfunctional tumor antigen-specific CD8 ⁺ T cells are detected and/or targeted via their expression of cell surface receptors described herein, such as 4-lBB,LAG-3, or additional markers that correlate with 4-1BB and LAG-3 expression, such as markers differentially expressed on the surface of the T cells (e.g., PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nml, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM and Sema7a).

Experiments conducted during development of embodiments herein identified markers/receptors that correlate and/or are responsible for tumor antigen-specific CD8 ⁺ T cell dysfunction. In some embodiments, the markers/receptors are overexpressed in dysfunctional tumor antigen-specific CD8 ⁺ T cells. In such embodiments, detecting the level (e.g., above a threshold) of such markers provides a diagnostic for detecting tumor antigen-specific CD8 ⁺ T cell dysfunction. Further, in such embodiments, targeting (e.g., inhibiting (e.g., expression and/or activity of)) such markers/receptors provides a therapeutic. In other embodiments, the markers/receptors are underexpressed in dysfunctional tumor antigen-specific CD8 ⁺ T cells. In such embodiments, detecting the level (e.g., below a threshold) of such markers provides a diagnostic for detecting tumor antigen-specific CD8 ⁺ T cell dysfunction. Further, in such embodiments, targeting (e.g., enhancing (e.g., expression and/or activity of)) such markers/receptors provides a therapeutic. Transcription factor Egr2 is a critical regulator of the anergic state in CD4 ⁺ T cell clones manipulated in vitro (Zheng et al, 2013; 2012; incorporated by reference in their entireties). Egr2 has also been shown to be involved in negative regulation of T cell activation in several in vivo model systems (Sumitomo et al, 2013; incorporated by reference in its entirety). Egr2 contributes to upregulation of DGKa and -z which act to blunt TCR- mediated Ras pathway activation (Zha et al, 2006; incorporated by reference in its entirety). By comparing gene expression profiling of anergized cells along with Egr2 ChlP-Seq analysis multiple additional Egr2-driven gene targets were identified (Zheng et al, 2013; incorporated by reference in its entirety). These gene targets include 4-1BB (Tnfrsf9 or CD137), Lag3, Nml, Sema7a, Crtam, and Rankl, which encode cell surface proteins.

4-1BB is a co-stimulatory molecule transiently expressed after TCR engagement. Lag3 (lymphocyte-activation gene 3 or CD223) is a CD4 homologue and functions as an inhibitory receptor. Expression of 4- IBB and Lag3 is regulated following TCR engagement and continues throughout differentiation. In humans, 4-1BB and LAG-3 are expressed on CD8+ TILs from human melanoma tumors (Gros et al, 2014; Baitsch et al, 2012;

incorporated by reference in their entireties). In both mice and humans, either molecule alone are expressed on populations of activated T cells. However, co-expression is more limited and is rarely observed in circulating T cells. The function of CD8+ TILs co-expressing these markers is unknown.

Experiments were conducted during development of embodiments herein to investigate the detailed characteristics of CD8+ TILs expressing 4-1BB and LAG-3 using mouse tumor models. It was found that the co-expression of 4-1BB and LAG-3 was sufficient to identify tumor antigen-specific dysfunctional CD8+ TILs enriched in the expression of Egr2 target genes. These CD8+ TILs failed to make IL-2 following in vitro stimulation, yet still produced IFN-g and Treg-recruiting chemokines and lysed target cells ex vivo, indicating they are not completely functionally inert. Combinatorial treatment with anti-LAG-3/anti-4-lBB restored the function of this population and promoted in situ acquisition of KLRG-lhi effector cells. Additional gene expression profiling provided a complete phenotyping of this T cell subset, which revealed expression of a broad panel of both inhibitory receptors and co-stimulatory receptors (e.g., receptors of Table 2 (e.g. Nrnl , Sema7a, CRTAM, etc.)). Inhibitory receptors and co-stimulatory receptors idientified in this profiling that are displayed on the surface of T cells include PD-1 , TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF 18, Nrnl, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A. These approaches have thus enabled the characterization of the population of tumor antigen-specific CD8+ T cells that arise specifically within the tumor

microenvironment having altered functional properties. In some embodiments, this population is a target for immunotherapeutic approaches to restore desired functionality and promote tumor regression. In some embodiments, the receptors/markers identified herein (e.g., 4-1BB, LAG-3, receptors/markers of Table 2 (e.g., surface markers/receptors (e.g. Nrnl, Sema7a, CRTAM, etc.), etc.) etc.) are targeted (e.g., via immunotherapeutic approaches) to restore desired immunoresponsiveness, to promote tumor regression, and/or for the treatment of cancer.

Experiments conducted during development of embodiments herein applied knowledge of Egr2 targets to evaluate applicability of these markers toward understanding dysfunctional T cells within tumors in vivo. The data indeed confirm that co-expression of LAG-3 and 4- IBB is sufficient to identify the majority of tumor antigen-specific CD8 ⁺ T cells within the tumor microenvironment. Co-expression of these markers was not observed within peripheral lymphoid organs in tumor-bearing mice, indicating that a property unique to the tumor context drives 4- IBB and LAG-3 expression. In addition, acquisition of LAG-3 and 4- IBB expression was not observed within tumors that were undergoing successful rejection, indicating that the acquisition of this phenotype occurs under conditions of incomplete antigen clearance.

In some embodiments, cancer treatment methods described herein comprise administration (or co-administration with one or more additional therapies/therapeutics) of one or more anti-4-lBB and/or anti-LAG-3 agents (e.g., antibodies, antibody fragments, antibody mimetic molecules (e.g., DARPins, affibodies, aptamers, nanobodies, etc.), etc.). In some embodiments, an anti-4-lBB and/or anti-LAG-3 agents is administered to render cancer cells, tumor(s), and/or the tumor microenvironment accessible or susceptible to treatment with additional therapies/therapeutics (e.g., immunotherapeutics). Anti-4-lBB and/or anti- LAG-3 agents that find use in embodiments described herein are not limited by their mechanism of action. Agents may be small molecules, peptide, polypeptides, proteins, nucleic acids (e.g., antisense, RNAi, etc.), antibodies, antibody fragments, etc.

In some embodiments, cancer treatment methods described herein comprise enhancing the activity or expression of a marker/receptor identified herein that negatively correlates with tumor antigen-specific CD8 ⁺ T cell dysfunction.

Experiments conducted during development of embodiments herein identified receptors/markers that are differentially expressed in dysfunctional CD8+ TILs (See Table 2). Testing of targets of interest identified in that screen demonstrate that at least neuritin 1

(Nrnl), cytotoxic and regulatory t-cell molecule (CRTAM), and Semaphorin 7A (Sema7a) are regulators of anti-tumor immunity, with Nrnl and CRTAM blockade correlating with increased tumor area, and Sema7a blockade correlating with decreased tumor area. In some embodiments, cancer treatment methods described herein comprise administration (or co-administration with one or more additional therapies/therapeutics) of agents (e.g., antibodies, antibody fragments, antibody mimetic molecules (e.g., DARPins, affibodies, aptamers, nanobodies, etc.), etc.) that target one or more receptors/markers of Table 2 (e.g. PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nml, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, Nml, CRTAM, Sema7a, etc.). In some embodiments, an agent is administered to render cancer cells, tumor(s), and/or the tumor microenvironment accessible or susceptible to treatment with additional

therapies/therapeutics (e.g., immunotherapeutics). Agents targeting one or more

receptors/markers of Table 2 (e.g. PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nml, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, Nml, CRTAM, Sema7a, etc.) that find use in embodiments described herein are not limited by their mechanism of action. Agents may be small molecules, peptide, polypeptides, proteins, nucleic acids (e.g., antisense, RNAi, etc.), antibodies, antibody fragments, etc. In some embodiments, an antagonist of Nml is administered. In some embodiments, an antagonist of CRTAM is administered. In some embodiments, an agonist of Sema7a is administered.

In some embodiments, antibodies, antibody fragments, antibody mimetic molecules (e.g., DARPins, affibodies, aptamers, nanobodies, etc.) targeting 4-1BB, LAG-3 and/or one or more receptors/markers of Table 2 (e.g. PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nml, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM, Sema7a, etc.), or fragments thereof, are provided. Such agents may be naked, deriving their effect by target binding (e.g., neutralizing the target), or may be conjugated to a functional moiety (e.g., drug, toxin, effector moiety, etc.).

In some embodiments, a subject is treated with (i) one or more agents (e.g., antibodies, antibody fragments, antibody mimetic molecules (e.g., DARPins, affibodies, aptamers, nanobodies, etc.), etc.) that target 4-1BB, LAG-3 and/or one or more

receptors/markers of Table 2 (e.g. PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nml, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM, Sema7a, etc.), as well as (ii) one or more additional cancer therapies. Such therapies include chemotherapy, immunotherapy, radiation, surgery, etc. In some embodiments, agents targeting the receptors/markers described herein are co-administered with one or more additional agents for the treatment of cancer.

In some embodiments, exemplary anticancer agents suitable for use in compositions and methods described herein include, but are not limited to: 1) alkaloids, including microtubule inhibitors (e.g., vincristine, vinblastine, and vindesine, etc.), microtubule stabilizers (e.g., paclitaxel (Taxol), and docetaxel, etc.), and chromatin function inhibitors, including topoisomerase inhibitors, such as epipodophyllotoxins (e.g., etoposide (VP-16), and teniposide (VM-26), etc.), and agents that target topoisomerase I (e.g., camptothecin and isirinotecan (CPT-1 1), etc.); 2) covalent DNA-binding agents (alkylating agents), including nitrogen mustards (e.g., mechlorethamine, chlorambucil, cyclophosphamide, ifosphamide, and busulfan (MYLERAN), etc.), nitrosoureas (e.g., carmustine, lomustine, and semustine, etc.), and other alkylating agents (e.g., dacarbazine, hydroxymethylmelamine, thiotepa, and mitomycin, etc.); 3) noncovalent DNA-binding agents (antitumor antibiotics), including nucleic acid inhibitors (e.g., dactinomycin (actinomycin D), etc.), anthracyclines (e.g., daunorubicin (daunomycin, and cerubidine), doxorubicin (adriamycin), and idarubicin (idamycin), etc.), anthracenediones (e.g., anthracycline analogues, such as mitoxantrone, etc.), bleomycins (BLENOXANE), etc., and plicamycin (mithramycin), etc. ; 4)

antimetabolites, including antifolates (e.g., methotrexate, FOLEX, and MEXATE, etc.), purine antimetabolites (e.g., 6-mercaptopurine (6-MP, PURINETHOL), 6-thioguanine (6- TG), azathioprine, acyclovir, ganciclovir, chlorodeoxyadenosine, 2-chlorodeoxyadenosine (CdA), and 2'-deoxycoformycin (pentostatin), etc.), pyrimidine antagonists (e.g.,

fluoropyrimi dines (e.g., 5-fluorouracil (ADRUCIL), 5 -fluorodeoxy uridine (FdUrd)

(floxuridine)) etc.), and cytosine arabinosides (e.g., CYTOSAR (ara-C) and fludarabine, etc.); 5) enzymes, including L-asparaginase, and hydroxyurea, etc. ; 6) hormones, including glucocorticoids, antiestrogens (e.g., tamoxifen, etc.), nonsteroidal antiandrogens (e.g., flutamide, etc.), and aromatase inhibitors (e.g., anastrozole (ARIMIDEX), etc.); 7) platinum compounds (e.g., cisplatin and carboplatin, etc.); 8) monoclonal antibodies (e.g., conjugated with anticancer drugs, toxins, and/or radionuclides, etc. ; neutralizing antibodies; etc.); 9) biological response modifiers (e.g., interferons (e.g., IFN-. alpha., etc.) and interleukins (e.g., IL-2, etc.), etc.); 10) adoptive immunotherapy; 11) hematopoietic growth factors; 12) agents that induce tumor cell differentiation (e.g., all-trans-retinoic acid, etc.); 13) gene therapy techniques; 14) antisense therapy techniques; 15) tumor vaccines; 16) therapies directed against tumor metastases (e.g., batimastat, etc.); 17) angiogenesis inhibitors; 18) proteosome inhibitors (e.g., VELCADE); 19) inhibitors of acetylation and/or methylation (e.g., HDAC inhibitors); 20) modulators of NF kappa B; 21) inhibitors of cell cycle regulation (e.g., CDK inhibitors); and 22) modulators of p53 protein function.

In some embodiments, agents targeting 4-1BB, LAG-3 and/or one or more receptors/markers of Table 2 (e.g. Nml, Sema7a, CRT AM, etc.) are administered to overcome immune invasion of the cancer cells, tumor, tumor microenvironment, etc. In some embodiments, one or more additional cancer immunotherapies are employed (e.g., concurrently or serially) to make use of the immune-responsiveness of the treated

cells/tumor. Suitable immunotherapies may include, but are not limited to: cell-based therapies (e.g., dendritic cell or T cell therapy, etc.), monoclonal antibody (mAb) therapy (e.g., naked mAbs, conjugated mAbs), cytokine therapy (e.g., interferons, interleukins, etc.), adjuvant treatment (e.g., polysaccharide-K), etc.

In some embodiments, agents targeting 4-1BB, LAG-3 and/or one or more receptors/markers of Table 2 (e.g. PD-1 , TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nrnl, Nrpl, KLRG1, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM, Sema7a, etc.) are co-administered with agents (e.g., small molecules, peptides, antibodies, antibody fragments, etc.) that target one or more cancer cell or tumor) markers or

components. In some embodiments, such co-administration renders the cancer cells, tumor, and/or tumor microenvironment susceptible and/or accessible to the treatment with the additional agent.

In some embodiments, agents for use in the methods and compositions described herein target and/or binds a cancer or tumor cell marker or component, selected from the group including but not limited to, epidermal growth factor receptor (EGFR, EGFRl , ErbB-1 , HERl). ErbB-2 (HER2/neu), ErbB-3/HER3, ErbB-4/HER4, EGFR ligand family; insulin-like growth factor receptor (IGFR) family, IGF-binding proteins (IGFBPs), IGFR ligand family (IGF-1R); platelet derived growth factor receptor (PDGFR) family, PDGFR ligand family; fibroblast growth factor receptor (FGFR) family, FGFR ligand family, vascular endothelial growth factor receptor (VEGFR) family, VEGF family; HGF receptor family: TRK receptor family; ephrin (EPH) receptor family: AXL receptor family; leukocyte tyrosine kinase (LTK) receptor family; TIE receptor family, angiopoietin 1 , 2; receptor tyrosine kinase-like orphan receptor (ROR) receptor family; discoidin domain receptor (DDR) family; RET receptor family; KLG receptor family; RYK receptor family; MuSK receptor family; Transforming growth factor alpha (TGF-a), TGF-a receptor; Transforming growth factor-beta (TGF-β), TGF-β receptor; Interleukin β receptor alpha2 chain (IL13Ralpha2), Interleukin-6 (IL-6), 1L- 6 receptor, interleukin-4, IL-4 receptor, Cytokine receptors, Class I (hematopoietin family) and Class II (interferon/1 L- 10 family) receptors, tumor necrosis factor (TNF) family, TNF-a, tumor necrosis factor (TNF) receptor superfamily (TNTRSF), death receptor family, TRAIL- receptor; cancer-testis (CT) antigens, lineage-specific antigens, differentiation antigens, alpha-actinin-4, ARTC1 , breakpoint cluster region-Abelson (Bcr-abl) fusion products, B- RAF, caspase-5 (CASP-5), caspase-8 (CASP-8), beta-catenin (CTN B1), cell division cycle 27 (CDC27), cyclin-dependent kinase 4 (CDK4), CDK 2A, COA-1, dek-can fusion protein, EFTUD-2, Elongation factor 2 (ELF2), Ets variant gene 6/acute myeloid leukemia 1 gene ETS (ETC6-AML1) fusion protein, fibronectin (FN), GPNMB, low density lipid

receptor/GDP-L fucose: beta-Dgalactose 2-alpha-Lfucosyltraosferase (LDLR/FUT) fusion protein, HLA-A2, MLA-A11, heat shock protein 70-2 mutated (HSP70-2M), KIAA0205, MART2, melanoma ubiquitous mutated 1, 2, 3 (MUM-1, 2, 3), prostatic acid phosphatase (PAP), neo-PAP, Myosin class 1, NFYC, OGT, OS-9, pml-RARalpha fusion protein, PRDX5, PTPRK, K-ras (KRAS2), N-ras (NRAS), HRAS, RBAF600, SIRT12, SNRPD1, SYT-SSXl or -SSX2 fusion protein, Triosephosphate Isomerase, BAGE, BAGE-1, BAGE-2, 3, 4, 5, GAGE-1, 2, 3, 4, 5, 6, 7, 8, GnT-V (aberrant N-acetyl glucosaminyl transferase V, MGAT5), HERV-K MEL, KK-LC, KM-HN-1, LAGE, LAGE-1, CTL-recognized antigen on melanoma (CAMEL), MAGE-A1 (MAGE-1). MAGE-A2, MAGE- A3, MAGE-A4, MAGE- AS, MAGE-A6, MAGE-A8, MAGE-A9, MAGE-A10. MAGE-A11, MAGE-A12, MAGE-3, MAGE-B1, MAGE-B2, MAGE-B5. MAGE-B6, MAGE-C1, MAGE-C2, mucin 1 (MUC1), MART-l/Melan-A (MLANA), gplOO, gpl00/Pmel l7 (S1LV), tyrosinase (TYR), TRP-1, HAGE, NA-88, NY-ESO-1, NY-ESO-l/LAGE-2, SAGE, Spl7. SSX-1, 2, 3, 4, TRP2-1NT2, carcino-embryonic antigen (CEA), Kallikrein 4, mammaglobin-A, OA1, prostate specific antigen (PSA), prostate specific membrane antigen, TRP-1/, 75. TRP-2 adipophilin, interferon inducible protein absent in melanoma 2 (AIM-2). BING-4, CPSF, cyclin Dl, epithelial cell adhesion molecule (Ep-CAM), EpbA3, fibroblast growth factor-5 (FGF-5), glycoprotein 250 (gp250intestinal carboxyl esterase (iCE), alpha-feto protein (AFP), M-CSF, mdm-2, MUCI, p53 (TP53), PBF, PRAME, PSMA, RAGE-1, RNF43, RU2AS, SOX10, STEAP1, survivin (BIRCS), human telomerase reverse transcriptase (hTERT), telomerase, Wilms' tumor gene (WT1), SYCP1, BRDT, SPANX, XAGE, ADAM2, PAGE-5, LIP1, CTAGE-1, CSAGE, MMA1, CAGE, BORIS, HOM-TES-85, AF15ql4, HCA66I, LDHC, MORC, SGY-1, SPOl l, TPX1, NY-SAR-35, FTHLI7, NXF2 TDRD1, TEX 15, FATE, TPTE, immunoglobulin idiotypes, Bence- Jones protein, estrogen receptors (ER), androgen receptors (AR), CD40, CD30, CD20, CD19, CD33, CD4, CD25, CD3, cancer antigen 72-4 (CA 72-4), cancer antigen 15-3 (CA 15-3), cancer antigen 27-29 (CA 27-29), cancer antigen 125 (CA 125), cancer antigen 19-9 (CA 19-9), beta-human chorionic gonadotropin, 1-2 microglobulin, squamous cell carcinoma antigen, neuron-specific enolase, heat shock protein gp96. GM2, sargramostim, CTLA-4, 707 alanine proline (707-AP), adenocarcinoma antigen recognized by T cells 4 (ART-4), carcinoembryogenic antigen peptide-1 (CAP-1), calcium- activated chloride channel-2 (CLCA2), cyclophilin B (Cyp-B), human signet ring tumor-2 (HST-2), etc.

Examples of antibodies which can be incorporated into compositions and methods disclosed herein include, but are not limited, to antibodies such as trastuzumab (anti- HER2/neu antibody); Pertuzumab (anti-HER2 mAb); cetuximab (chimeric monoclonal antibody to epidermal growth factor receptor EGFR); panitumumab (anti-EGFR antibody); nimotuzumab (anti-EGFR antibody); Zalutumumab (anti-EGFR mAb); Necitumumab (anti- EGFR mAb); MDX-210 (humanized anti-HER-2 bispecific antibody); MDX-210 (humanized anti-HER-2 bispecific antibody); MDX-447 (humanized anti-EGF receptor bispecific antibody); Rituximab (chimeric murine/human anti-CD20 mAb); Obinutuzumab (anti-CD20 mAb); Ofatumumab (anti-CD20 mAb); Tositumumab-1131 (anti-CD20 mAb); Ibritumomab tiuxetan (anti-CD20 mAb); Bevacizumab (anti-VEGF mAb); Ramucirumab (anti-VEGFR2 mAb); Ranibizumab (anti-VEGF mAb); Aflibercept (extracellular domains of VEGFR1 and VEGFR2 fused to IgGl Fc); AMG386 (angiopoietin-1 and -2 binding peptide fused to IgGl Fc); Dalotuzumab (anti-IGF-lR mAb); Gemtuzumab ozogamicin (anti-CD33 mAb);

Alemtuzumab (anti-Campath-l/CD52 mAb); Brentuximab vedotin (anti-CD30 mAb):

Catumaxomab (bispecific mAb that targets epithelial cell adhesion molecule and CD3); Naptumomab (anti-5T4 mAb); Girentuximab (anti-Carbonic anhydrase ix); or Farletuzumab (anti-folate receptor). Other examples include antibodies such as Panorex™ (17-1A) (murine monoclonal antibody); Panorex (@(17-1A)) (chimeric murine monoclonal antibody); BEC2 (ami-idiotypic mAb, mimics the GD epitope) (with BCG); Oncolym (Lym-1 monoclonal antibody); SMART M195 Ab, humanized 13' 1 LYM-1 (Oncolym). Ovarex (B43.13, anti- idiotypic mouse mAb); 3622W94 mAb that binds to EGP40 (17-1 A) pancarcinoma antigen on adenocarcinomas; Zenapax (SMART Anti-Tac (IL-2 receptor); SMART M195 Ab, humanized Ab, humanized); NovoMAb-G2 (pancarcinoma specific Ab); TNT (chimeric mAb to histone antigens); TNT (chimeric mAb to histone antigens); Gliomab-H

(Monoclonals— Humanized Abs); GNI-250 Mab; EMD-72000 (chimeric-EGF antagonist); LymphoCide (humanized IL.L.2 antibody); and MDX-260 bispecific, targets GD-2, ANA Ab, SMART IDIO Ab, SMART ABL 364 Ab, or ImmuRAIT-CEA.

In some embodiments, an agent that finds use in embodiments herein specifically binds a component of a regulatory T cell, myeloid suppressor cell, or dendritic cell. In another aspect, the targeting moiety specifically binds one of the following molecules: CD4; CD25 (IL-2a receptor; IL-2aR); cytotoxic T-lymphocyte antigen-4 (CTLA-4; CD152); Interleukin-10 (IL-10); Transforming growth factor-beta receptor (TGF- R); Transforming growth factor-beta (TGF-β); Programmed Death-1 (PD-1); Programmed death-1 ligand (PD- Ll or PD-L2); Receptor activator of nuclear factor-κΒ (RANK); Receptor activator of nuclear factor-κΒ (RANK) ligand (RANKL); LAG-3; glucocorticoid-induced tumor necrosis factor receptor family -related gene (GITR; TNFRSF18); or Interleukin-4 receptor (IL-4R). In some embodiments, the agent is an agonist that increases the function of the targeted molecule. In other embodiments, the agent is an antagonist that inhibits the function of the targeted molecule.

In some embodiments, an agent that finds use in embodiments herein binds a specific cytokine, cytokine receptor, co-stimulatory molecule, co-inhibitory molecule, or

immunomodulatory receptor that modulates the immune system. In another aspect, the targeting moiety specifically binds one of the following molecules: tumor necrosis factor (TNF) superfamily; tumor necrosis factor-a (TNF-a); tumor necrosis factor receptor (TNFR) superfamily; Interleukin-12 (IL-12); IL-12 receptor; 4-1BB (CD137); 4-1BB ligand (4- 1BBL; CD137L); OX40 (CD134; TNR4); OX40 ligand (OX40L; CD40; CD40 ligand (CD40L); CTLA-4; Programmed death-1 (PD-1); PD-1 ligand I (PD-L1 : B7-H1); or PD-1 ligand 2 (PD-L2; B7-DC); B7 family; B7-1 (CD80); B7-2 (CD86); B7-H3; B7-H4;

GITR/AITR: GITRL/AITRL; BTLA; CD70; CD27; LIGHT; HVEM: Toll-like receptor (TLR) (TLR 1, 2, 3, 4, 5, 6, 7, 8, 9, 10). In some embodiments, the agent is an agonist that increases the function of the targeted molecule. In other embodiments, the agent is an antagonist that inhibits the function of the targeted molecule.

In some embodiments, agents (e.g., immunotherapeutics) targeting 4-1BB, LAG-3 and/or one or more receptors/markers of Table 2 (e.g. PD-1, TIM-3, OX-40ICOS, TIGIT, CD244, TNFRSF18, Nrnl , Nrpl , KLRG1 , GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM, Sema7a, etc.) are co-administered (e.g., serially or sequentially) with one or more adjuvants. Suitable adjuvants include, but are not limited to, one or more of: oil emulsions (e.g., Freund's adjuvant); saponin formulations; virosomes and viral-like particles; bacterial and microbial derivatives; immunostimulatory oligonucleotides; ADP-ribosylating toxins and detoxified derivatives; alum; BCG; mineral-containing compositions (e.g., mineral salts, such as aluminium salts and calcium salts, hydroxides, phosphates, sulfates, etc.); bioadhesives and/or mucoadhesives; microparticles; liposomes; polyoxyethylene ether and poly oxy ethylene ester formulations; polyphosphazene; muramyl peptides; imidazoquinolone compounds; and surface active substances (e.g. lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, keyhole limpet hemocyanin, and dinitrophenol). Adjuvants may also include immunomodulators such as cytokines, interleukins (e.g., IL-1, IL-2, IL-4, IL-5, IL-6, IL-7, IL-12, etc.), interferons (e.g., interferon-. gamma.), macrophage colony stimulating factor, and tumor necrosis factor. In addition to variant B7- DC polypeptides, other co-stimulatory molecules, including other polypeptides of the B7 family, may be administered. Proteinaceous adjuvants may be provided as the full-length polypeptide or an active fragment thereof, or in the form of DNA, such as plasmid DNA.

Pharmaceutical and immunotherapeutic compositions described herein may be delivered by any suitable route of administration (e.g., oral delivery, parenteral delivery, mucous membrane delivery, pulmonary delivery, intravenous delivery, etc.). Appropriate formulations for such delivery routes are understood in the field.

Non-limiting examples of cancers that may be treated with the compositions and methods described herein include, but are not limited to: melanoma (e.g., metastatic malignant melanoma), renal cancer (e.g. clear cell carcinoma), prostate cancer (e.g. hormone refractory prostate adenocarcinoma), pancreatic cancer (e.g., adenocarcinoma), breast cancer, colon cancer, lung cancer (e.g. non-small cell lung cancer), esophageal cancer, squamous cell carcinoma of the head and neck, liver cancer, ovarian cancer, cervical cancer, thyroid cancer, glioblastoma, glioma, leukemia, lymphoma, and other neoplastic malignancies. In some embodiments, the cancer is a solid tumor cancer.

Some embodiments described herein are particularly useful for the treatment of tumors that do not otherwise respond to immunotherapeutic approaches. In some embodiments, provided herein is the treatment of cancers that are non-responsive (or have a reduced response) to T cells or antigen presenting cells (e.g., dendritic cells (e.g.,

CD103 DCs, etc.), etc.). In some embodiments, provided herein is the treatment of cancers that are non-responsive to treatments, despite T cell infiltration. In some embodiments, compositions and methods described herein find use in the treatment of cancers in which T cells are not appropriately primed against tumor-associated antigens. In some embodiments, compositions and methods described herein find use in the treatment of cancers comprising tumors or cells that are defective in recruitment of dendritic cells (e.g., CD 103 ⁺ DCs, etc.). In some embodiments, compositions and methods described herein find use in the treatment of cancers comprising tumors or cells that are defective in production of the chemokine CCL4.

In some embodiments, the therapeutic compositions and methods herein find use with those described in, for example WO 2016/141312; incorporated by reference in its entirety.

In some embodiments, methods are provided for testing sample (e.g., cell, tissue, population of cells, tumor, blood, urine, saliva, etc.) from a subject for one or more biomarkers (e.g., biomarkers of dysfunctional tumor antigen-specific CD8 T cells). Such biomarkers may comprise nucleic acids, small molecules, proteins, peptides, etc., and may be detected using any suitable assay of technique. In some embodiments, provided herein are DNA-, RNA-, small molecule, and/or protein-based diagnostic methods that either directly or indirectly detect the biomarkers of the evasion of immune response or immunotherapy by cancer cells or tumors. The present invention also provides compositions, reagents, and kits for such diagnostic purposes.

In some embodiments, biomarkers are detected at the nucleic acid (e.g., RNA) level. For example, the presence or amount of biomarker nucleic acid (e.g.,mRNA) in a sample is determined (e.g., to determine the presence or level of biomarker expression). Biomarker nucleic acid (e.g., RNA, amplified cDNA, etc.) may be detected/quantified using a variety of nucleic acid techniques known to those of ordinary skill in the art, including but not limited to nucleic acid sequencing, nucleic acid hybridization, nucleic acid amplification (e.g., by PCR, RT-PCR, qPCR, etc.), micorarray, Southern and Northern blotting, sequencing, etc. Non-amplified or amplified nucleic acids can be detected by any conventional means. For example, in some embodiments, nucleic acids are detected by hybridization with a detectably labeled probe and measurement of the resulting hybrids. Nucleic acid detection reagents may be labeled (e.g., fluorescently) or unlabeled, and may by free in solution or immobilized (e.g., on a bead, well, surface, chip, etc.).

In some embodiments, biomarkers are detected at the protein level. For example, the presence or amount of biomarker protein in a sample is determined (e.g., to determine the presence or level of biomarker expression or localization). In some embodiments, reagents are provided for the detection and/or quantification of biomarker proteins. Suitable reagents include primary antibodies (e.g., that bind to the biomarkers), secondary antibodies (e.g., that bind primary antibodies), antibody fragments, aptamers, etc. Protein detection reagents may be labeled (e.g., fluorescently) or unlabeled, and may by free in solution or immobilized (e.g., on a bead, well, surface, chip, etc.).

In some embodiments, biomarker capture reagents are provided to localize, concentrate, aggregate, etc. a biomarker. For example, in some embodiments a biomarker capture reagent that interacts with the biomarker is linked to a solid support (e.g., a bead, surface, resin, column, and the like) that allows manipulation by the user on a macroscopic scale. Often, the solid support allows the use of a mechanical means to isolate and purify the biomarker from a heterogeneous solution. For example, when linked to a bead, separation is achieved by removing the bead from the heterogeneous solution, e.g., by physical movement. In embodiments in which the bead is magnetic or paramagnetic, a magnetic field is used to achieve physical separation of the capture reagent (and thus the target) from the

heterogeneous solution. Magnetic beads used to isolate targets are described in the art, e.g., as described in European Patent Application No. 87309308, incorporated herein in its entirety for all purposes.

Compositions for use in the diagnostic methods or testing steps described herein include, but are not limited to, probes, amplification oligonucleotides, and antibodies. Any of the detection and/or diagnostic reagents used in embodiments described herein may be provided alone or in combination with other compositions in the form of a kit. Kits may include any and all components necessary or sufficient for assays including, but not limited to, the detection reagents, buffers, control reagents (e.g., tissue samples, positive and negative control sample, etc.), solid supports, labels, written and/or pictorial instructions and product information, inhibitors, labeling and/or detection reagents, package environmental controls (e.g., ice, desiccants, etc.), and the like. In some embodiments, the kits provide a sub-set of the required components, wherein it is expected that the user will supply the remaining components. In some embodiments, the kits comprise two or more separate containers wherein each container houses a subset of the components to be delivered.

In some embodiments, a computer-based analysis program is used to translate the raw data generated by the detection assay (e.g. , the presence, absence, or amount of expression a biomarker) into data of predictive value for a clinician. In some embodiments, computer analysis combines various data into a single score or value that is predictive and/or diagnostic. The clinician can access the predictive data using any suitable means. Thus, in some preferred embodiments, the present invention provides the further benefit that the clinician, who is not likely to be trained in genetics or molecular biology, need not understand the raw data. The data is presented directly to the clinician in its most useful form. The clinician is then able to immediately utilize the information in order to optimize the care of the subject. Contemplated herein are any methods capable of receiving, processing, and transmitting the information to and from laboratories conducting the assays, information providers, medical personal, and subjects. For example, in some embodiments of the present invention, a sample (e.g. , a biopsy, cell, or blood sample) is obtained from a subject and submitted to a profiling service (e.g. , clinical lab at a medical facility, third-party testing service, genomic profiling business, etc. to generate raw data. Where the sample comprises a tissue or other biological sample, the subject may visit a medical center to have the sample obtained and sent to the profiling center, or subjects may collect the sample themselves and directly send it to a profiling center. In some embodiments, a report is generated (e.g., by a clinician, by a testing center, by a computer or other automated analysis system, etc.). A report may contain test results, diagnoses, and/or treatment recommendations. EXPERIMENTAL

Materials and Methods

Mice and tumor inoculation

Female C57BL/6 mice ranging from 6 to 8 weeks were purchased from Taconic Farms. CD45.1 and Rag2 ^mice on the C57BL/6 background were obtained from Taconic Farms and bred at the University of Chicago. 2C/Rag2 ^~μ and P14/Rag2 ^ mice have been previously described (Brown et al, 2006; incorporated by reference in its entirety). pLCK- CreERT2 x ROSA-YFP mice were generated and have been described (Evaristo et al, 2016; incorporated by reference in its entirety). B16.SIY.dsRed (Kline et al, 2012; incorporated by reference in its entirety), C1498.SIY.GFP (Zhang et al, 2009; incorporated by reference in its entirety), and MC57.SIY.GFP (Spiotto et al, 2002; incorporated by reference in its entirety) tumor cells were engineered to express either dsRed or GFP in frame with the H2-K ^b - restricted model antigen SIYRYYGL. The 1969.SIY.GFP cell line was engineered by retroviral transduction of the 1969 cell line (Diamond et al, 2011; incorporated by reference in its entirety) using the pLEGFP plasmid expressing cDNA for SIYRYYGL (Spiotto et al, 2002; incorporated by reference in its entirety). For experiments, mice 6 to 9 weeks of age and received 2x10 ⁶ tumor cells subcutaneously on either the left flank or both the left and right flank. All mice were maintained according to the National Institute of Health Animal Care guidelines and studied under IACUC-approved protocols.

To generate the targeting construct for the Egr2 ^EGFP knock-in reporter mice, a 12.6kb mouse genomic DNA fragment including the egr2 gene was excised with SacII and cloned into a pEasy-Flox vector adjacent to the thymidine kinase (TK) selection marker. A cassette containing IRES2-eGFP and a LoxP-flanked neomycin selection marker was inserted into an Nhel site between the translation stop codon (TGA) and the polyadenylation signal of the egr2 gene. ES cell clones from 129 mice were electroporated and selected for Neomycin resistance. ES cell clones were verified for homologous insertion in the endogenous locus by PCR and southern blot with 5' and 3' probes. Mice were backcrossed to C57BL/6 for over 8 generations. TIL isolation

Tumors were harvested from mice at the indicated time points. Tumors were dissociated through a 50 μηι filter and washed with PBS. TILs were further enriched by layering Ficoll-Hypaque beneath the cell suspension followed by centrifugation without breaks for 30 min at 400 x g. The buffy-layer was isolated and washed twice with PBS before staining. For isolating specific cell populations by FACS, tumors were pooled when indicated and the cell layer was re-purified by Ficoll-Hypaque centrifugation twice. For day 28 tumors, after Ficoll-Hypaque separation, T cells were further purified by negative bead selection according to manufacturer's instructions (MAGNISORT, eBiosciences). Cells were then washed with PBS, stained at 4°C for 15 minutes before resuspending in complete

DMEM (cDMEM: 10% FBS, lOOU/mL Penicillin-Streptomycin, 1% MEM Non-Essential Amino Acids, 50 μΜ β-ΜΕ, 0.01M MOPS), and were sorted into either RLT lysis buffer (QIAGEN) or cDMEM depending on the experimental assay. Cells sorted into RLT buffer were put directly on dry ice as soon as the sort was finished.

Flow cytometry and antibodies

Cell suspensions were washed twice in PBS before staining an FACS buffer (10% FBS, 2mM EDTA, 0.001 % NaN ₃ ). Cells were stained for 30 min on ice and fixed in 1% PFA. Antibodies against the following molecules were used: CD3 (17A2, AX700), 2B4 (2B4, FITC), CD127 (A7R34, PE), OX-40 (OX-86, PE), 4-lBB (17B5, Biotin, APC), CD160 (7H1, PE-Cy7), LAG- 3 (C9B7W, PerCPeFluor710), PD-1 (RMP1 -30, PE-Cy7), NRP1 (3E12, BV421), GITR (DTA-1 , FITC), ICOS (7E.17G9, BV421), KLRG-1 (2F1, eF450, BV605), TIGIT (1G9, APC), TIM-3 (RMT3-23, PE), CD4 (RM4-5, BV605), CD45.1 (A20, FITC), CD45.2 (104, PE), CD8a (53-6.7, BV711). Fixable Viability Dye 506 (eBioscience) was used for live/dead discrimination. Staining of SIY-specific T cells was performed utilizing the SIYRYYGL-Pentamer (PE) (Proimmune); a SIINFEKL-pentamer (PE) was used as a non-specific control. All flow cytometric analysis was conducted on an LSRFortessa (BD) and analyzed using Flow Jo software (Tree Star).

Quantitative real-time PCR

Total RNA was extracted from sorted cell populations using the RNEasy Micro Kit (QIAGEN) following the manufacturer's protocol. cDNA was synthesized using the High Capacity cDNA Reverse Transcription kit (Applied Biosy stems) according to manufacturer's instructions. Transcript levels were determined using primer-probe sets (Tables la and lb) developed through the online ProbeFinder Software and the Universal Probe Library (Roche) with the exception of IL-2 (Mm00434256_ml) and 18S (Hs99999901_sl). To minimize batch effect, when possible, all samples probed for a gene were run on the same 96-well qRT- PCR plate. All primer-probe sets either contained a primer spanning an exon-exon boundary or primers spanning an intron. Expression levels of transcripts were normalized to 18S expression.

Table la. Primer Sequences

Table lb. Primer/Probe

Arl3 30 ctggcagatccagtcctgtt acccagttcatgccatcct 49 100

Exph5 31 atgagggaggagageggtat cagcttgttgtccaaatcgtc 50 67

Fhl2 32 agaaaaccatcatgccaggt acaggtgaagcaggtctcgt 51 74

Nrn1 33 atcctcgcggtgcaaata gcccttaaagactgcatcaca 52 108

Ptgfrn 34 ccggggagatctcatcaaa tcgaaggccatgtcatctg 53 12

Rankl 35 tgaagacacactacctgactcctg cccacaatgtgttgcagttc 54 88

Hifla 36 gctgctcactgtgaaggaagt tggggaatgeattttaccat 55 2

Eqr3 37 caatctgtaccccgaggaga ccgatgtccatcacattctct 56 74

Tnfa 38 ctgtag cccacgtcgtag c ttgagatccatgccgttg 57 25

Gzmb 39 gctgctcactgtgaaggaagt tggggaatgeattttaccat 58 2

Cell 40 tcaccatgaaacccactgc agcagcagctattggagacc 59 71

Ccl22 41 tcttgctgtggcaattcaga gcagagggtgacggatgtag 60 74

In vivo proliferation assay

In vivo proliferation was measured by a BrdU pulse 24 hours prior to flow cytometric analysis. Each mouse received 0.8 mg BrdU injected i.p. (intraperitoneal) on day 12 after tumor inoculation. TILs were isolated and surface stain was performed as described above. Following surface staining, cells were fixed and permeabilized using the Foxp3 staining kit (BD), according to manufacturer's protocol, and incubated with 100 μΐ PBS/DNase solution (300 μg/ml) for 30 minutes at 37 °C. Cells were washed and incubated for 30 minutes at room temperature with anti-BrdU (FITC, Bu20a) and then washed with and resuspended in PBS.

In vitro stimulation assays

Tissue culture-treated 96-well round bottom plates were coated with anti-CD3s (1 μζΐντλ; 2C1 1) in DPBS overnight at 4°C or for 2 hours at 37°C. Cells were sorted into cold cDMEM media and put on ice as soon as the sort was finished. Cells were then pelleted, resuspended in 50 μΐ cDMEM and incubated with soluble anti-CD28 (2 μ^ητΐ; PV-1) for 10- 12 hours for a final volume of 100 μΐ. After stimulation supernatants were removed for ELISA or bead-based immunoassay (LegendPlex), and cells were washed once with DPBS and resuspended in 15 μΐ of RNAlater Stabilization Solution (QIAGEN) or 300 μΐ of RLT buffer. Cells were stored at -80°C until RNA isolation was performed.

Protein quantification

Measurement of protein concentration was determined either by a standard ELISA or bead-based immunoassay (LEGENDplex, BioLegend). ELISAs were performed according to manufacturer's protocol (Ready-SET-Go ELISA; eBioscience) on supernatants from in vitro stimulations. Absorbance values were obtained at 450 nm using an Emax microplate reader (Molecular Devices) and IL-2 concentration was determined by standard curve. Protein concentration values were normalized to the number of sorted cells plated. LEGENDplex assays were performed according to manufacturer's protocols. IL-2 concentration (Figure 4B) was confirmed by both methods in separate experiments with no significant difference in IL- 2 concentration between the two methods.

Spectratype analysis and sequencing

Mice were injected with 2x10 ⁶ B16.SIY.dsRed tumor cells. 14 days later, tumors were harvested and specific CD8 ⁺ TIL subpopulations were sorted into RLT buffer

(QIAGEN) and immediately frozen. cDNA was synthesized from sorted cell populations and CDR3 regions were amplified by PCR with 21 different νβ-5' primers paired with a FAM- Οβΐ.1 primer (Table 1). Three νβ PCR reactions did not reach significant amplification for analysis and were removed from the analysis. For sequencing, Οβ-νβ PCR products were purified using the QIAquick PCR purification kit (QIAGEN) and sequenced at the University of Chicago Genomics Core Facility. Cβ-Vβ PCR products were analyzed by capillary electrophoresis at the University of Chicago Genomics core and CDR3 peaks were aligned using the Liz500 ladder. Spectratype graphs were displayed using the GeneiousR9 software (Kearse et al, 2012). To generate the frequency profile for each νβ spectratype, the area under each peak was measured using peak studio (fodorlab.uncc.edu/software/peakstudio). The Hamming Distance (Currier and Robinson, 2001; incorporated by reference in its entirety) was calculated between each νβ spectratype from each CD8 ⁺ spleen and TIL population within a given mouse. To determine significance between the HD from each comparison the HDs for each νβ from mice were averaged and a One-Way ANOVA with Dunn's correction for multiple comparisons was performed.

TCR transgenic T cell transfer experiments

Cell suspensions were generated from spleens and lymph nodes from congenic 2C/Rag2 ^~/ 7CD45.1/2 and/or P14/Rag2 ^" 7CD45.2 mice and T cells were purified by CD8 ⁺ negative selection (Miltenyi Biotechnologies) over magnetic columns according to the manufacturer's protocol. TCR Transgenic (Tg) T cells were washed with PBS, resuspended at a concentration of 10x10 /ml and 1x10 TCR Tg cells were adoptively transferred into CD45.1 tumor bearing mice by tail vein transfer in a volume of 0.1 mL. After indicated times, 2C T cells and corresponding host CD8 ⁺ T cells were sorted and stimulated as described above.

In vitro cytotoxicity assay

Per individual experiment, 10 C57BL/6 mice were injected s.c. (subcutaneous) with 2 x 10 ⁶ B16.SIY cells on both left and right flanks. On day 14, all 20 tumors were pooled and dissociated using the Tumor Dissociation Kit (Miltenyi Biotec) following the manufacturer's protocol. Tumor cell suspensions were washed 3-5 times with PBS and TILs were enriched for by Ficoll-Hypaque gradient centrifugation. TILs were stained, sorted and put directly on ice. TILs were titrated and added directly to a 96-well plate containing 50,000 P815 mastocytoma cells and 1 μg/mL anti-CD3. For a positive control, OT-I cells were isolated from OT-I/Rag2 ^ mice and stimulated with plate-bound anti-CD3 (0.25 μg/mL), anti-CD28 (2 μg/mL) and 100 U/mL IL-2 for 2-3 days. For a negative control, P815 cells were cultured alone or cultured with naive CD8 ⁺ T cells isolated from lymph nodes. After 12 hours of incubation, cells were stained for Thyl, CD45, CD8a, Fixable Viability Dye 450

(eBioscience) and/or propidium Iodide.

Gene expression analysis

Total RNA for the CD8 ⁺ TIL subpopulations was isolated following the

manufacturer's protocol (RNEasy Micro Kit: QIAGEN) from sorted cells pooled from 10 mice. Samples were analyzed by the University of Chicago Genomics Facility using Illumina MouseRef8 microarray chips. Two experimental replicates were performed, and the results were log ₂ transformed and averaged. Probe sets that revealed a 1.5-fold difference abs(log ₂ (ratio)> 1.5)) relative to CD8 ⁺ 4-lBB AG-3 PD-1 cells were identified and used for subsequent analysis. The microarray data are available in the Gene Expression Omnibus database (ncbi.nlm.nih.gov/gds) under accession number GSE79919. For cross-study comparisons, log2-fold change values were extracted using the GE02R online software from the hypofunctional CD8 ⁺ TIL data set, GSE79858 ((GSM2107353, GSM2107353 and GSM2107355) versus (GSM2107350, GSM2107351, GSM210732)) and the CD8 ⁺ T cell exhausted data set, GSE41870 ((GSM 1026819, GSM1026820, GSM1026821) versus (GSM1026786, GSM1026787, GSM1026788, GSM1026789)). Upregulated genes showing a 2-fold difference were used for analysis. Multiple genes names with from the GE02R extracted data were identified and matched to gene names from the Illumina data set. The rank-rank hypergeometric overlap (RRHO) analysis (Plaisier et al, 2010; incorporated by reference in its entirety) was conducted at systems.crump.ucla.edu/rankrank/index.php and the associated Bioconductor package "RRHO" (Rosenblatt and Stein, 2014; incorporated by reference in its entirety).

Gene Ontology enrichment analysis

In a pair-wise fashion, shared upregulated genes were used as the input for the ClueGO software with the Cytoscape application (Shannon et al., 2003; incorporated by reference in its entirety). Both the Biological Process and Immune System Process Gene Ontology Annotations were used for analysis. Only pathways with a Bonferroni step down correction p-value > 0.01 were considered when generating pathway nodes. Non-redundant pathways with the greatest number of genes found within each node were used as examples in Figure 6A.

Antibody and FTY720 treatments

Mice were treated i.p. with 100 μg/mouse of anti-4-lBB (Bio-X-Cell; LOB12.3) antibody and/or 100 μg/mouse anti-LAG-3 (Bio-X-Cell; C9B7W). For tumor outgrowth experiments, mice were treated on day 7, 10, 13 and 16 after tumor inoculation. For ex vivo functional experiments mice were treated on day 7, 10 and 13 and cells were sorted on day 14. For experiments blocking lymph node egress, 25 μg of FTY720 was given by gavage one day prior to first antibody treatment (day 6) and continued every day until endpoint on day 14.

Results

4- IBB and LAG-3 identify a major population of CD8 ⁺ TILs

To determine whether 4- IBB and LAG-3 could identify dysfunctional CD8 ⁺ TILs, the expression partem of LAG-3 and 4- IBB was examined using the well-characterized B16.SIY model of melanoma. On day 7 following tumor inoculation, the 4-lBB ⁺ LAG-3 ⁺ population comprised 15.8% of all CD8 ⁺ TILs. The frequency of this population significantly increased to 44% by day 21. The frequency of 4-lBB AG-3 ⁺ (4 ⁺ ) population also increased 1.9- fold from day 7 to day 14 to comprise 25% of the CD8 ⁺ TIL compartment. In contrast, the frequency of the 4- IBB LAG-3 (4 ) population decreased by 2.7-fold by day 21. There was no significant increase in the proportion or number of 4-lBB ⁺ LAG-3 CD8 ⁺ TILs within the time frame of the experiment (Figure 1 A and B). Similar patterns were seen when analyzing absolute numbers of cell subsets (Figure 1 C and D). Acquisition of these phenotypes was specific for the tumor microenvironment, as they were not observed in the spleen or tumor-draining lymph node (TdLN) (Figure 1 A). These data indicate that the tumor microenvironment preferentially supports the induced co-expression of LAG-3 and 4-1BB.

The selective increase in cell numbers and proportional shift towards the 4-1BB LAG-3 ⁺ and 4-lBB ⁺ LAG-3 ⁺ populations during tumor progression indicated that expansion of these populations was occurring within the tumor microenvironment. CD8 ⁺ TILs were stained for Ki67 at day 14 after tumor inoculation and analyzed by flow cytometry. 81% of 4-1BB LAG-3 ⁺ cells and 85% of 4-lBB ⁺ LAG-3 ⁺ cells were Ki67 ⁺ compared to only 32% of the 4- IBB LAG-3 TILs (Figure IE). Mice were pulsed with BrdU on day 12, and 24 hours later the CD8 ⁺ TIL subpopulations were analyzed for BrdU incorporation. Indeed, the 4- lBB AG-3 ⁺ and 4-lBB ⁺ LAG-3 ⁺ populations incorporated more BrdU compared to the 4- 1BB AG-3 population (Figure IF). These data indicate that once CD8 ⁺ T cells arrive at the tumor site, a fraction of TILs expands within the tumor, and that these expanding TILs are identified by increased expression of 4- IBB and LAG-3.

To determine if upregulation of LAG-3 and 4- IBB was simply a product of the B16. SIY tumor model or if it is a more general feature of CD8 ⁺ T cells within tumors, T cells from three additional progressively growing tumor models, C1498. SIY, MC38. SIY, EL4.SIY and B16F 10 parental were analyzed. TILs were analyzed for expression of 4-1BB and LAG- 3 at day 14. A partem of expression was found that is similar to that seen in CD8 ⁺ TILs isolated from B16. SIY tumors (Figure 1G and I). The results from the B 16F10 parental tumor confirm that presence of SIY is not required to see co-expression of 4- IBB and LAG- 3. In order to determine whether the 4-lBB ⁺ LAG-3 ⁺ TIL subset was generated only in progressing tumors or also in tumors that were rejected, T cell phenotypes in the 1969. SIY and MC57. SIY fibrosarcoma tumor models were analyzed, which are more immunogenic and undergo spontaneous rejection. Distinctly fewer 4-lBB ⁺ LAG-3 ⁺ cells were found among the CD8 ⁺ TIL compartment in the 1969. SIY and MC57.SIY tumors (Figure H and I). Over time, co-expression of 4-1BB and LAG-3 was maintained in B 16. SIY tumors but not MC57. SIY tumors (Figure 1 J). These data indicate that the acquisition of the LAG-3 ⁺ 4-lBB ⁺ TIL phenotype preferentially occurs within the tumor microenvironment and only upon conditions of tumor progression rather than regression. CD8 4-1BB LAG-3 TILs express Egr2 and multiple Egr2 gene targets

Experiments conducted during development of embodiments herein to determine whether Egr2 expression itself was also characteristic of T cells within the CD8 ⁺ TIL compartment; an Egr2-IRES-GFP (Egr2 ^GFP ) knock-in reporter mouse was utilized.

Approximately 14% of all CD8 ⁺ TILs were GFP ⁺ on both day 7 and day 14 (Figure 2A). To confirm that Egr2 is faithfully reported, CD8 ⁺ TILs expressing high and low levels of EGFP were sorted and screened for Egr2 and several Egr2 targets by qRT-PCR. The Egr2-GFP ^hl population expressed greater levels of Egr2 and many Egr2-target genes previously defined using in vitro anergy models. These include Tnfrsf9, Lag3, Ngn, Sema7a, Crtam, Cell and Nrnl (Figure 2B). Expression of 4-1BB and LAG-3 in the Egr2-GFP ^hi CD8 ⁺ TILs was confirmed by flow cytometry. The majority of Egr2-GFP ^hl cells expressed LAG-3 and/or 4- 1BB. The Egr2GFP ^l0 cells also showed expression of 4- IBB and LAG-3 on a subpopulation at day 14 (Figure 2C). This result indicates either that CD8 ⁺ TILs expressing Egr2 encompass only a subset of the TILs expressing LAG-3 and/or 4-1BB, or that Egr2 is transiently expressed and is subsequently downregulated after the induction of LAG-3 and 4-1BB.

Using Egr2 target genes from in vitro anergic CD4 ⁺ T cell clones (Zheng et al., 2013; incorporated by reference in its entirety), the Egr2-driven transcriptional program was examined in sorted 4- IBB LAG-3 and 4-lBB ⁺ LAG-3 ⁺ cells by qRT-PCR. Of the 43 Egr2 target genes examined, 10 showed detectably increased expression in 4-lBB ⁺ LAG-3 ⁺ population, while expression of a similar subset of genes was increased in the 4-1BB LAG-3 ⁺ population (Figure 2D). Collectively, these data demonstrate that Egr2 is expressed in a subpopulation of CD8 ⁺ TILs expressing LAG-3 and/or 4-1BB, and that a subset of known Egr2 targets was detected in these larger T cell populations as a whole.

It was next examined whether Egr2 was required for expression of LAG-3 and 4- IBB among CD8 ⁺ TIL in vivo. To this end Egr2 ^flox/flox X pLCK-CreERT2 x ROSA- YFP mice were utilized, in which oral tamoxifen administration results in a fraction of the CD8 ⁺ T cells deleting Egr2 and expressing YFP (Figure 2E). This allowed comparison of both Egr2- sufficient (YFP ) and Egr2-deficient (YFP ⁺ ) CD8 ⁺ within the same tumor. To determine that Egr2 was in fact deleted from the YFP ⁺ fraction, both YFP ⁺ and YFP CD8 ⁺ TILs were sorted and Egr2 transcripts were measured directly ex vivo and upon ex vivo stimulation. The YFP ⁺ CD8 ⁺ TILs expressed substantially less Egr2 transcripts compared to the YFP counterparts (Figure 2E). To determine if Egr2 is required for 4-1BB and LAG-3 expression, CD8 ⁺ TILs were analyzed at day 7 and 14 after tumor inoculation and compared the YFP ⁺ and YFP populations to mice not treated with tamoxifen. At day 7, the YFP ⁺ fraction expressed less 4- IBB and LAG-3 compared to the YFP population and the WT CD8 TILs. However, expression of 4- IBB and LAG-3 was not significantly different at day 14 (Figure 2F). This indicates that other transcriptional regulators compensate and contribute to the expression of LAG-3 and 4- IBB, especially at later time points.

Egr3 has been shown to have overlapping function with Egr2 (Safford et al, 2005; incorporated by reference in its entirety) and HIFla can contribute to 4- IBB expression (Palazon et al., 2012). To investigate whether these transcription factors may compensate for 4-lBB and/or LAG-3 expression we sorted Egr2GFP ^hi and Egr2GFP ^l0 CD8 ⁺ TILs expressing 4-lBB and LAG-3 on day 7 and analyzed expression of Egr3 and HIFla by qRT-PCR. Egr3 and HIFla were indeed expressed in both the Egr2GFP ^hl and Egr2GFP ^l0 populations. It was confirmed differential expression of Egr2 and CCL1 to between the Egr2GFP ^hl and

Egr2GFP ^l0 populations to assure sort purity (Figure 2G). Together, these data indicate that Egr2 contributes to upregulation of 4-lBB and LAG-3 expression at early time points, but that other transcriptional regulators compensate and drive expression of LAG-3 and 4-lBB as the T cell-tumor interaction progresses.

CD8 ⁺ 4-lBB ⁺ LAG-3 ⁺ TILs are oligoclonal and enriched for tumor antigen specificity

Not all T cells in the tumor microenvironment are specific for tumor-associated antigens, as memory T cells specific for irrelevant antigens are often found among TIL, and non-specific T cell trafficking has been documented in vivo (Harlin et al, 2006; incorporated by reference in its entirety). Experiments conducted during development of embodiments herein to determine whether 4-lBB ⁺ LAG-3 ⁺ CD8 ⁺ TILs are tumor-antigen specific. LAG-3, 4- IBB and Egr2 are upregulated after TCR stimulation and experiments indicate that this population expands within the tumor microenvironment in situ. Three complementary techniques were employed. First, the CD8 ⁺ TILs were isolated based on LAG-3 and 4- IBB expression by cell sorting and performed TCR spectratype analysis. Compared to the 4- 1BB LAG-3 TILs and CD8 ⁺ splenocytes, the 4-lBB ⁺ LAG-3 ⁺ TILs had a non-Gaussian distribution and shared one or two dominant peaks (Figure 3A). Analysis of several νββ displaying one dominant peak revealed that νβ7 contained a single CDR3 sequence shared between the 4-lBB LAG-3 ⁺ and 4-lBB ⁺ LAG-3 ⁺ populations, indicating a clonal relationship (Figure 3A). To measure the oligoclonality of the CDR3 repertoires the Hamming Distance (HD) was calculated for each νβ between the CD8 ⁺ TIL subpopulations and the splenic CD8 ⁺ population within three separate mice (Figure 8). By transforming each spectratype into area under the curve frequency profiles the Hamming Distance computes the changes in frequency and reports a value of comparison between 0 and 1, with 0 indicating a completely identical frequency profile and 1 signifying a completely discordant profile. As a control, the HD of the splenic CD8 ⁺ populations between different mice was calculated (Figure 3B, black bar). Since the splenic CD8 ⁺ spectratypes are largely Gaussian this value represents the HD between two similar distributions. Analysis of the HD between the CD8 ⁺ TIL subpopulations revealed that the 4-lBB ⁺ LAG-3 ⁺ and 4-1BB LAG-3 ⁺ but not the 4- IBB LAG-3 CDR3 distributions are significantly different (less Gaussian) compared to the splenic CD8 ⁺ population (Figure 3B). These data indicate that the 4-lBB ⁺ LAG-3 ⁺ and 4-1BB LAG-3 ⁺ populations are oligoclonal expanded subsets of TILs, indicating antigen specificity in these subpopulations.

As a second approach, the B16.SIY melanoma and MC38.SIY adenocarcinoma models were utilized. CD8 ⁺ T cells specific for the H-2K ^b -restricted SIY epitope

(SIYRYYGL) were monitored. SIYRYYGL/K ^b pentamer ⁺ (H-2K ^b /SIY) cells were found in expanded numbers within B16.SIY and MC38.SIY tumors at day 14 after tumor inoculation (Figure 3C). Nearly 47% of the H-2K ^b /SIY ⁺ cells expressed both 4-1BB and LAG-3, in contrast to 32% of the H-2K ^b /SIY population (Figure 3C and E). This enrichment of antigen-specific CD8 ⁺ TILs in the 4-lBB ⁺ LAG-3 ⁺ populations indicates that these markers identify tumor antigen-specific TILs. The H-2K ^b /SIY cells also contained significant numbers of 4-lBB ⁺ LAG-3 ⁺ cells, which is consistent with the notion that tumor antigens other than SIY are also recognized by subsets of CD8 ⁺ TILs in vivo (Figure 3C). H-2K ^b /SIY ⁺ cells in the spleen or TdLN did not co-express 4- IBB and LAG-3, indicating that this phenotype is acquired within the tumor microenvironment.

These features were also analyzed in the context of tumor-antigen specific CD8 ⁺ TILs in two spontaneously rejected tumor models. To this end, H-2K ^b /SIY-specific CD8 ⁺ TILs cells were evaluated from MC57.SIY and 1969.SIY tumors. At day 14 after tumor inoculation, approximately 5% of the H-2K ^b /SIY-specific CD8 ⁺ TILs were found in the 4- lBB ⁺ LAG-3 ⁺ fraction. As with the B16.SIY tumors, no H-2K ^b /SIY-specific CD8 T cells co- expressed 4- IBB and LAG-3 in the TdLN or spleen (not shown) (Figure 3D). Unlike the B16.SIY and MC38.SIY tumors, no significant enrichment of 4-lBB ⁺ LAG-3 ⁺ H-2K ^b /SIY- specific CD8 ⁺ TILs was observed (Figure 3D and E). These data indicate that tumor antigen specificity per se does not determine dysfunctionality, and that this is a feature unique to the microenvironment of progressing tumors.

As a third measure to determine if tumor-antigen specific CD8 ⁺ T cells acquire the 4- lBB ⁺ LAG-3 ⁺ phenotype, congenically marked 2C and P14 transgenic (Tg) T cells, isolated from 2C/Rag2 ^{~ ~} and P14/Rag2 ^{~ ~} mice, were transferred into tumor-bearing hosts. The 2C TCR is specific for the SIY model antigen expressed by B16.SIY tumor cells, while P14 is an irrelevant TCR specific for the LCMV-derived gp33-4i epitope; both TCRs are H-2K ^b - restricted. 2C and P14 Tg CD8 ⁺ T cells were transferred via tail vein 7 days after tumor inoculation. Seven days after transfer, tumors and TdLNs were extracted and the phenotypic profile of the transferred populations was analyzed. This system allowed for the analysis of two T cell populations with defined antigen specificities within the same tumor

microenvironment, as well as the polyclonal host CD8 ⁺ T cells. The 2C T cells were more efficiently recruited and expanded within the tumor microenvironment compared to the P 14 T cells and encompassed a large fraction of the total CD8 ⁺ TIL population (Figure 3F). Of the 2C T cells, nearly all expressed LAG-3 and or 4- IBB while this was true for only a small percentage of the P14 cells (Figure 3G and H). Consistent with the SIY-K ^b pentamer analysis, the co-expression of LAG-3 and 4-lBB on 2C T cells was not observed in the TdLN. Together, these results demonstrate that the 4-lBB ⁺ LAG-3 ⁺ phenotype is a property of tumor antigen-specific TIL under conditions of tumor progression.

CD8 ⁺ TILs expressing LAG-3 and 4-1 BB exhibit defective IL-2 production yet produce IFN-y and Treg-recruiting chemokines

Based on the characteristics of the in vitro T cell anergy model that led to the identification of Egr2 as an important regulator, experiments conducted during development of embodiments herein to determine whether the tumor-antigen specific 4-lBB ⁺ LAG-3 ⁺ CD8 ⁺ TIL population is dysfunctional in their capacity to produce IL-2. To this end each subpopulation was sorted and stimulated with anti-CD3 and anti-CD28 mAb and analyzed IL-2 production by qRT-PCR and ELISA. Since nearly all CD8 ⁺ TILs displayed an activated phenotype, CD8 ⁺ CD44 ⁺ splenocytes wer used as a positive control. Indeed, the 4-lBB ⁺ LAG- 3 ⁺ cells showed a 100-fold reduction in 11-2 mRNA and as much as a 40-fold reduction in IL- 2 protein levels compared to the 4-lBB ^~ LAG-3 ^~ population (Figure 4A and 4B). As a second approach, Egr2 ^hl TIL (which are also largely 4-lBB ⁺ LAG-3 ⁺ ) was examined by utilizing the Egr2-GFP reporter mice. Indeed, ex vivo stimulated Egr2-GFP ^hi CD8 ⁺ TILs also exhibited reduced 11-2 transcript compared to Egr2-GFP ^l0 cells (Figure 4C). As a final approach, congenically marked 2C T cells were adoptively transferred intravenously into tumor-bearing hosts and recovered the 2C T cells 7 days later from the tumor and TdLN. 2C T cells isolated from tumors exhibited a reduced capacity to produce 11-2 transcripts, at a level equivalent to 4-lBB ⁺ LAG-3 ⁺ TILs, compared to 2C CD44 ⁺ T cells isolated from the TdLN (Figure 4D). In chronic infection models, expression of PD-1 has been suggested to identify intrinsically dysfunctional or "exhausted" CD8 ⁺ T cells. To determine if PD-1 alone might be sufficient to identify cells that lack the capacity to produce IL-2, CD8 ⁺ TILs that lacked expression of LAG-3 and 4- IBB were isolated and tested for the ability of the PD-1 ⁺ fraction to produce IL-2. Approximately -10% of CD8 ⁺ TILs were 4-1BB LAG-3 PD-1 ⁺ on day 14 and 21

(Figure 4E and F). Upon ex vivo stimulation, this population retained the capacity to produce 11-2 mRNA at a level comparable to the 4- IBB LAG-3 cells (Figure 4G). These results indicate that PD-1 expression alone is not sufficient to identify dysfunctional TIL in the tumor microenvironment.

To further examine functional alterations during tumor progression protein levels of

IL-2, IFN-γ and TNF-a wer tested after TCR stimulation. As the loss of the ability of CD8 ⁺ TILs to produce cytokines is suggested to be a temporal process reported initiated following entry into the tumor microenvironment (Waugh et al, 2016; Schietinger et al, 2016;

incorporated by reference in their entireties) or progressively after 30 days in the chronic LCMV model (Wherry et al, 2007; incorporated by reference in its entirety), cytokine production was tested on day 7, 14, 21 and 28. The 4-lBB ⁺ LAG-3 ⁺ population lost the capacity to produce IL-2 as early as day 7 while the 4-1BB LAG-3 ⁺ population lost IL-2 production between day 7 and day 14 (Figure 5A). The 4- IBB LAG-3 population did not lose the ability to produce IL-2 at any time point tested (Figure 5A), supporting the notion that this population is not tumor antigen specific and that differentiation into the

dysfunctional state is an antigen-dependent process (Schietinger et al, 2016; incorporated by reference in its entirety). The 4-lBB ⁺ LAG-3 ⁺ population produced more IFN-γ at all time points after day 7 compared to their negative counterparts, albeit with a slight decrease in IFN-γ production over time. While the increase in IFN-γ was maintained until later time points, TNF-a production was lost by day 28 (Figure 5A).

Experiments were conducted during development of embodiments herein to evaluate production of cytokines directly in the tumor without in vitro restimulation, which may more closely reflect which T cells were receiving TCR stimulation in situ. Each T cell population was sorted directly ex vivo without any culturing and mRNA levels were measured by qRT- PCR. Elevated Ifn-y and Gzmb transcripts were observed from the 4-lBB ⁺ LAG-3 ⁺ subpopulation, along with a slight decrease in Tnf-a levels, compared to the 4- IBB LAG-3 cells (Figure 5B). Production of IFN-γ in primary TILs was confirmed by injecting tumors with Brefeldin A prior to analysis by intracellular cytokine staining. Consistent with the mRNA expression, the 4-lBB ⁺ LAG-3 ⁺ population produced significantly greater amounts of IFN-γ protein (Figure 5C). Thus, the 4-1BB LAG-3 TIL are not completely devoid of functionality, as they continue to produce IFN-γ despite defective production of IL-2. This phenotype is consistent with in vitro T cell anergy models (Jenkins et al, 1987; incorporated by reference in its entirety).

To test whether the 4-lBB ⁺ LAG-3 ⁺ population still retains cytotoxic capacity, redirected lysis was performed by co-culturing anti-CD3 bound P815 mastocytoma target cells with the different CD8 ⁺ TIL subpopulations directly after sorting. 4-lBB ⁺ LAG-3 ⁺ CD8 ⁺ TILs isolated from day 14 tumors were able to lyse target cells at a comparable efficacy to in vitro primed OT-I cells. 4-lBB ⁺ LAG-3 ⁺ TILs isolated from day 21 tumors were still able to lyse target cells, albeit to a lesser extent compared to primed OT-I cells (Figure 5D).

CD8 ⁺ T cells in the tumor can be the source of the chemokine CCL22 that recruits FoxP3 ⁺ regulatory T cells (Tregs) to the tumor microenvironment (Spranger et al, 2013; incorporated by reference in its entirety). In addition, the chemokine Cell was an Egr2 target in anergic T cells (Zheng et al, 2013; incorporated by reference in its entirety), and it has been suggested that CCL1 also contribute to Treg recruitment in the tumor context in vivo (Hoelzinger et al, 2010; incorporated by reference in its entirety). However, whether all CD8 ⁺ T cells in the tumor produce these chemokines or if they are only produced by subpopulations of T cells had not been determined. To address this the CD8 ⁺ TIL phenotypic subpopulations were analyzed for Cell and Ccl22 mRNA expression directly ex vivo by qRT- PCR. Indeed, the 4-lBB ⁺ LAG-3 ⁺ TIL population produced substantially greater Cell and Ccl22 compared to their negative counterparts or to splenic CD8 ⁺ CD44 ⁺ T cells (Figure 4K). As a control, expression of a distinct chemokine Ccl5 was found not to be differentially expressed.

Together, these data show that co-expression of 4- IBB and LAG-3 delineates tumor antigen-specific CD8 ⁺ TIL that lack the ability to produce IL-2 yet retain the ability to produce IFN-γ, kill target cells in vitro, and secrete chemokines capable of Treg recruitment. Given the fact that IFN-γ is responsible for the upregulation of PD-L1 and IDO in the tumor microenvironment, and that chemokines produced by CD8 ⁺ TIL contribute to Treg recruitment (Spranger et al, 2013; incorporated by reference in its entirety), these data indicate that the 4-lBB ⁺ LAG-3 ⁺ population contributes to the network of immune suppressive mechanisms within the tumor microenvironment that limit the efficacy of antitumor immunity. Gene expression profiling reveals that CD8 4-1BB LAG-3 TILs express an extensive array of additional co-stimulatory and co-inhibitory receptors

Having in hand surface markers that define tumor antigen-specific dysfunctional CD8 ⁺ TILs, experiments conducted during development of embodiments herein to compare the gene expression profile of this population to other published profiles of dysfunctional CD8 ⁺ T cells to determine genes that regulate or are differentially expressed in cells in this dysfunctional state. To this end, a cross-study comparison was conducted of the

transcriptional profiles of the "dysfunctional" 4-lBB ⁺ LAG-3 ⁺ CD8 ⁺ TILs, "hypofunctional" CD8 ⁺ TILs from a study utilizing the murine CT26 tumor model (Waugh et al, 2016;

incorporated by reference in its entirety) and LCMV "exhausted" GP33 specific CD8 ⁺ T cells (Doering et al., 2012; incorporated by reference in its entirety). The results are depicted in Table 2. Only genes with a 2-fold increase over controls from each study independently were considered. Over a 2-fold greater number of genes was found to be shared between the dysfunctional TIL dataset and the previously published hypofunctional CD8 ⁺ TIL data, than with the exhausted T cell profile (Figure 6A). In addition, a rank-rank hypergeometric overlap (RRHO) analysis indicated a greater statistically significant overlap (Figure 10A) and a greater correlation (Figure 10B) between the current dysfunctional TIL and the published hypofunctional CD8 ⁺ TIL gene expression profiles compared to the virally -induced exhausted CD8 ⁺ T cell profile, indicating a more similar molecular program between CD8 ⁺ T cells isolate from tumors compared to chronic viral infection.

Table 2. Differentially regulated genes in CD8 4-1BB LAG-3 TILs

0037

GZME granzyme E 7.16037 ITFG3 Description Not Found

5687 1.55016

2812

RPL6 ribosomal protein L6 7.14210 HAAO 3-hydroxyanthranilate 3,4- 7057 dioxygenase 1.55055

3207

NRN1 neuritin 1 7.08799 RNF138 ring finger protein 138

3146 1.55144

9524

LPL lipoprotein lipase 7.00450 UNC93B1 unc-93 homolog B 1 (C.

1392 elegans) 1.55176

7491

CLGN calmegin 6.93369 ANKZF1 ankyrin repeat and zinc

0655 finger domain containing 1 1.55221

4097

CD70 CD70 molecule 6.90689 IFITM3 interferon induced

0596 transmembrane protein 3 1.55264

4542

AREG amphiregulin 6.71287 TXNIP thioredoxin interacting

0868 protein 1.55278

5452

ZRANB3 zinc finger RANBP2- 6.59544 LMAN1L lectin, mannose binding 1

type containing 3 3985 like 1.55458

8852

ASNS asparagine synthetase 6.59496 ALDH3B1 aldehyde dehydrogenase 3

(glutamine-hydrolyzing) 878 family member B 1 1.55471

1558

FANCD2 Fanconi anemia 6.35314 GIP gastric inhibitory

complementation group 6826 polypeptide 1.55551 D2 1104

GM156 predicted gene 6.29370 COX7A2L cytochrome c oxidase

156(Gml56) 1542 subunit 7A2 like 1.55557

2553

ACAA1B acetyl-Coenzyme A 6.29370 APPL2 adaptor protein,

acyltransf erase 1542 phosphotyrosine 1.55559 lB(Acaalb) interacting with PH 8704 domain and leucine zipper 2

IGF2BP3 insulin like growth factor 6.18685 KLHL22 kelch like family member

2 mRNA binding protein 7067 22 1.55592

3 9583

GZMG granzyme G 6.09381 OLFR272 olfactory receptor

3673 272(01fr272) 1.55748

2156

CIB2 calcium and integrin 6.00786 LRRC29 leucine rich repeat

binding family member 8243 containing 29 1.55936 2 6716

ATG9B autophagy related 9B 5.98641 A630095E1 Description Not Found

0935 3RIK 1.56071

4954

XKR8 XK related 8 5.97727 OLFR194 olfactory receptor

9924 194(01frl94) 1.56071

4954

EPDR1 ependymin related 1 5.95652 OLFR1013 olfactory receptor

1363 1013(Olfrl013) 1.56071

4954

SPP1 secreted phosphoprotein 5.79776 GLRA4 glycine receptor alpha 4

1 9502 1.56071 4954

RGS8 regulator of G-protein 5.75380 P2RY6 pyrimidinergic receptor

signaling 8 5672 P2Y6 1.56071

4954

MDFIC MyoD family inhibitor 5.73063 RASGEF1B RasGEF domain family

domain containing 9956 member IB 1.56071

4954

DMWD dystrophia myotonica, 5.68720 IL22RA2 interleukin 22 receptor

WD repeat containing 0695 subunit alpha 2 1.56071

4954

KIF11 kinesin family member 5.66959 LIN7C lin-7 homolog C, crumbs

11 3751 cell polarity complex 1.56071 component 4954

LGI2 leucine rich repeat LGI 5.65535 DMRT1 doublesex and mab-3

family member 2 1829 related transcription factor 1.56071

1 4954

ZFP41 ZFP41 zinc finger 5.61544 T SPAN 12 tetraspanin 12

protein 5725 1.56071

4954

MLKL mixed lineage kinase 5.60584 PAK3 p21 (R AC 1) activated

domain-like 9867 kinase 3 1.56071

4954

CENPH centromere protein H 5.56376 COL2A1 collagen type II alpha 1

8278 chain 1.56071

4954

SERPINF1 serpin family F member 5.53605 SLC37A1 solute carrier family 37

1 29 member 1 1.56071

4954

UNC13B unc-13 homolog B (C. 5.50303 PSD3 pleckstrin and Sec7

elegans) 0646 domain containing 3 1.56071

4954

ML ANA melan-A 5.49665 RDH5 retinol dehydrogenase 5

4083 1.56071

4954

PES1 pescadillo ribosomal 5.48437 ABC A3 ATP binding cassette

biogenesis factor 1 6709 subfamily A member 3 1.56126

3453

2900026A0 Description Not Found 5.47735 PLA2G4E phospholipase A2 group

2RIK 3527 IVE 1.56165

0879

OSR2 odd-skipped related 5.41616 DDIT3 DNA damage inducible

transciption factor 2 4165 transcript 3 1.56356

6526

MPP6 membrane palmitoylated 5.40850 ZFP12 zinc finger protein

protein 6 6442 12(Zfpl2) 1.56430

8646

HIST1H3C histone cluster 1, H3c 5.39746 PIGYL phosphatidylinositol

0726 glycan anchor 1.56458 biosynthesis, class Y- 5219 like(Pigyl)

PI4K2B phosphatidylinositol 4- 5.37503 CCDC97 coiled-coil domain

kinase type 2 beta 9431 containing 97 1.56535

5117

SH3YL1 SH3 and SYLF domain 5.37503 OLFR1112 olfactory receptor

containing 1 9431 1112(01frl l l2) 1.56589

319

RAD51 RAD51 recombinase 5.37155 ACTN2 actinin alpha 2

8863 1.56693

1646 ZBTB32 zinc finger and BTB 5.31831 POLG polymerase (DNA)

domain containing 32 6841 gamma, catalytic subunit 1.56726

5595

MSC musculin 5.28540 FBX032 F-box protein 32

2219 1.56728

1905

TG thyro globulin 5.25927 MRPL15 mitochondrial ribosomal

2487 protein L 15 1.57072

2678

RSPH1 radial spoke head 1 5.23649 FCHSD2 FCH and double SH3

homolog 2618 domains 2 1.57182

1211

ARL11 ADP ribosylation factor 5.21916 RECQL RecQ like helicase

like GTPase 11 852 1.57288

9668

NUDT11 nudix hydrolase 11 5.21529 NDUFBl l NADH : ubiquinone

0306 oxidoreductase subunit 1.57288

Bl l 9668

APBB1 amyloid beta precursor 5.19770 SOX8 SRY-box 8

protein binding family B 8158 1.57334 member 1 1535

SPINK2 serine peptidase 5.18982 1700030J22 Description Not Found

inhibitor, Kazal type 2 4559 RIK 1.57662

394

HMGN3 high mobility group 5.16892 EMB embigin

nucleosomal binding 2782 1.57789 domain 3 0585

FAM20B family with sequence 5.12722 CELSR1 cadherin EGF LAG seven- similarity 20 member B 055 pass G-type receptor 1 1.57820

1987

CDC25C cell division cycle 25C 5.11997 COL1A2 collagen type I alpha 2

861 chain 1.58068

2782

FAM20A family with sequence 5.10852 1700080E1 Description Not Found

similarity 20 member A 4457 IRK 1.58104

6002

PPP1R16B protein phosphatase 1 5.09592 GALNT12 polypeptide N- regulatory subunit 16B 442 acetylgalactosaminyltransf 1.58136 erase 12 3645

SBNOl strawberry notch 5.05093 RMND5B required for meiotic

homolog 1 (Drosophila) 6965 nuclear division 5 1.58396 homolog B 0816

ST14 suppression of 5.02680 LRRC28 leucine rich repeat

tumorigenicity 14 0059 containing 28 1.58398

7499

LRRC49 leucine rich repeat 5.02470 OLFR622 olfactory receptor

containing 49 4311 622(01fr622) 1.58496

2501

ΉΑΜ1 T-cell lymphoma 5.00450 OLFR339 olfactory receptor

invasion and metastasis 1392 339(01fr339) 1.58496 1 2501

APLF aprataxin and PNKP like 4.95186 NEIL3 nei like DNA glycosylase

factor 7504 3 1.58496

2501

PGPEP1 pyroglutamyl-peptidase I 4.92718 SNX24 sorting nexin 24

5358 1.58496

2501

ALCAM activated leukocyte cell 4.90929 SLC7A11 solute carrier family 7

adhesion molecule 3086 member 11 1.58496 containing 77 662 subunit A 1.59958

8488

RHBDF1 rhomboid 5 homolog 1 4.62643 OLFR952 olfactory receptor

9137 952(01fr952) 1.59967

9175

REEP3 receptor accessory 4.59991 1700021F05 Description Not Found

protein 3 2842 RIK 1.60162

3253

ITGA3 integrin subunit alpha 3 4.59096 CCDC79 Description Not Found

1241 1.60219

5565

SCCPDH saccharopine 4.59096 FAM134B family with sequence

dehydrogenase (putative) 1241 similarity 134 member B 1.60271

5966

MY ADM myeloid associated 4.58796 SEMA3B semaphorin 3B

differentiation marker 4989 1.60288

4409

FAM132A family with sequence 4.58195 FA2H fatty acid 2-hydroxylase

similarity 132 member A 3751 1.60449

4406

FOXRED2 FAD dependent 4.57288 ULK1 unc-51 like autophagy

oxidoreductase domain 9668 activating kinase 1 1.60465 containing 2 3903

CENPK centromere protein K 4.56985 MCOLN1 mucolipin 1

5608 1.60624

2992

DCXR dicarbonyl and L- 4.56224 BMP5 bone morphogenetic

xylulose reductase 2424 protein 5 1.60676

0033

TSPAN6 tetraspanin 6 4.54225 ANKRD50 ankyrin repeat domain 50

805 1.60713

7028

UPP1 uridine phosphorylase 1 4.53838 OLFR560 olfactory receptor

296 560(Olfr560) 1.60880

9243

DOK4 docking protein 4 4.52042 OLFR366 olfactory receptor

2249 366(01fr366) 1.60880

9243

ELOVL4 ELOVL fatty acid 4.50143 OLFR273 olfactory receptor

elongase 4 9145 273(01fr273) 1.60880

9243

KNDCl kinase non-catalytic C- 4.49979 FHIT fragile histidine triad

lobe domain containing 0117 1.60880 1 9243

KRT17 keratin 17 4.49185 AQP11 aquaporin 11

3096 1.60880

9243

CHST2 carbohydrate 4.48731 TMEM176 transmembrane protein

sulfotransferase 2 5031 A 176A 1.60880

9243

TPX2 TPX2, microtubule 4.47573 ENAH enabled homolog

nucleation factor 3431 (Drosophila) 1.60880

9243

DUSP14 dual specificity 4.45614 CLDN6 claudin 6

phosphatase 14 9035 1.60880

9243

BGN biglycan 4.44956 SP1 Spl transcription factor

1375 1.60880

9243 3217 nuclear protein 2 1.63938

3642

IL4I1 interleukin 4 induced 1 3.80219 BLOC1S3 biogenesis of lysosomal

3217 organelles complex 1 1.63958 subunit 3 5785

PHF19 PHD finger protein 19 3.80219 ELL elongation factor for RNA

3217 polymerase II 1.64021

945

CKAP2L cytoskeleton associated 3.79701 GTF3C4 general transcription factor

protein 2 like 2978 IIIC subunit 4 1.64065

8029

GSTT1 glutathione S-transferase 3.79181 MYLPF myosin light chain,

theta 1 4071 phosphorylatable, fast 1.64066 skeletal muscle 0074

ADAM3 a disintegrin and 3.78135 CYP2A12 cytochrome P450, family

metallopeptidase domain 9714 2, subfamily a, 1.64194 3 (cyritestin)(Adam3) polypeptide 12(Cyp2al2) 7141

SLAMF7 SLAM family member 7 3.78135 RNF139 ring finger protein 139

9714 1.64201

0395

MCPT8 mast cell protease 3.77082 C78339 Description Not Found

8(Mcpt8) 9046 1.64357

3868

DGKG diacylglycerol kinase 3.76553 EDEM1 ER degradation enhancing

gamma 4746 alpha-mannosidase like 1.64385 protein 1 619

NLGN2 neuroligin 2 3.71699 UBE2E1 ubiquitin conjugating

0894 enzyme E2 El 1.64585

9791

SERPINE2 serpin family E member 3.69488 PALMD palmdelphin

2 0193 1.64632

2067

IL10 interleukin 10 3.68929 AMICA1 adhesion molecule,

9161 interacts with CXADR 1.64747 antigen 1 (Arnica 1) 8619

SLC6A13 solute carrier family 6 3.68929 KLHL11 kelch like family member

member 13 9161 11 1.65061

1828

STAU2 staufen double-stranded 3.66675 IFNGR2 interferon gamma receptor

RNA binding protein 2 6592 2 (interferon gamma 1.65105 transducer 1) 0175

ARHGDIG Rho GDP dissociation 3.65535 DECR1 2,4-dienoyl-CoA reductase

inhibitor gamma 1829 1, mitochondrial 1.65140

6438

TKl thymidine kinase 1 3.63747 SAMD3 sterile alpha motif domain

7097 containing 3 1.65321

3853

PCYT1A phosphate 3.61772 9130409123 Description Not Found

cy tidy lyltransf erase 1, 8231 RIK 1.65535 choline, alpha 1829

LAMB 3 laminin subunit beta 3 3.60880 2010107G1 Description Not Found

9243 2RIK 1.65535

1829

UBE2N ubiquitin conjugating 3.59096 ZFP354B zinc finger protein

enzyme E2 N 1241 354B(Zfp354b) 1.65535

1829

STARD8 StAR related lipid 3.57893 TAS2R143 taste receptor, type 2,

transfer domain 8713 member 143(Tas2rl43) 1.65535 containing 8 1829 PRR5 proline rich 5 3.57893 OLFR65 olfactory receptor

8713 65(01fr65) 1.65535

1829

BDH2 3 -hy droxybuty rate 3.55458 NRP neural regeneration

dehydrogenase, type 2 8852 protein(Nrp) 1.65535

1829

FAM124B family with sequence 3.54843 DOK3 docking protein 3

similarity 124 member B 6625 1.65535

1829

MGAT3 mannosyl (beta- 1,4-)- 3.54843 HIGD1A HIG1 hypoxia inducible

glycoprotein beta- 1 ,4-N- 6625 domain family member 1A 1.65535 acetylglucosaminyltransf 1829 erase

LAG3 lymphocyte activating 3 3.54234 CCDC13 coiled-coil domain

6309 containing 13 1.65535

1829

GDPD5 glycerophosphodiester 3.53881 ANGPTL2 angiopoietin like 2

phosphodiesterase 2733 1.65535 domain containing 5 1829

RNF168 ring finger protein 168 3.53605 CNGB3 cyclic nucleotide gated

29 channel beta 3 1.65535

1829

LYPLA1 lysophospholipase I 3.52982 HOXD4 homeobox D4

0947 1.65535

1829

TUBGCP4 tubulin gamma complex 3.52356 KIFC3 kinesin family member C3

associated protein 4 1956 1.65535

1829

PYGL phosphorylase, 3.51412 AMACR alpha-methylacyl-CoA

glycogen, liver 226 racemase 1.65535

1829

CCL3 C-C motif chemokine 3.51028 2310014L1 Description Not Found

ligand 3 1539 7RIK 1.65570

7015

BCAT1 branched chain amino 3.50816 BRAP BRCAl associated protein

acid transaminase 1 3667 1.65709

0723

ATP6V0A1 ATPase H+ transporting 3.50143 SLC39A1 solute carrier family 39

V0 subunit al 9145 member 1 1.65763

1089

EIF4E eukaryotic translation 3.49825 OLFR419 olfactory receptor

initiation factor 4E 0868 419(01fr419) 1.65813

796

HIST1H4B histone cluster 1, H4b 3.49185 NHP2L1 NHP2 non-histone

3096 chromosome protein 2-like 1.65829

1 (S. cerevisiae)(Nhp211) 8045

LAD1 ladinin 1 3.49085 STOML2 stomatin like 2

426 1.65935

7735

ITGAV integrin subunit alpha V 3.48542 SAMM50 SAMM50 sorting and

6827 assembly machinery 1.66240 component 0762

MRPL47 mitochondrial ribosomal 3.48542 CCDC91 coiled-coil domain

protein L47 6827 containing 91 1.66322

99

CAMK2N1 calcium/calmodulin 3.48446 ATF3 activating transcription

dependent protein kinase 0783 factor 3 1.66348 II inhibitor 1 3642

UEVLD UEV and lactate/malate 3.46597 RAI1 retinoic acid induced 1 - dehyrogenase domains 4465 1.66388

5989

SFXN4 sideroflexin 4 3.46270 RRAS2 related RAS viral (r-ras)

6751 oncogene homolog 2 1.66582

6896

2810417H1 Description Not Found 3.46163 UROS uroporphyrinogen III

3RIK 4298 synthase 1.66592

3156

RAD51AP1 RAD51 associated 3.45943 SCOC short coiled-coil protein

protein 1 1619 1.66627

2349

FUT4 fucosyltransferase 4 3.45285 DUSP10 dual specificity

8965 phosphatase 10 1.66648

5948

CTN BIP1 cateninbeta interacting 3.44625 CYB5R4 cytochrome b5 reductase 4

protein 1 623 1.66675

6592

ZBTB80S zinc finger and BTB 3.42626 9930104L0 Description Not Found

domain containing 8 4755 6RIK 1.66715 opposite strand 0978

LYSMD4 LysM domain containing 3.42259 ZFP579 zinc finger protein

4 008 579(Zfp579) 1.66902

3741

DIAP3 Description Not Found 3.40599 RGP1 RGP1 homolog, RAB6A

236 GEF complex partner 1 1.66939

3721

PTGIS prostaglandin 12 3.39917 PIAS2 protein inhibitor of

(prostacyclin) synthase 1094 activated STAT 2 1.67213

7196

MO API modulator of apoptosis 1 3.39231 METTLl mefhyltransferase like 1

7423 1.67242

5342

SLC27A3 solute carrier family 27 3.39231 POU5F1 POU class 5 homeobox 1

member 3 7423 1.67385

4965

MRPL39 mitochondrial ribosomal 3.37149 SERPINB6 serine (or cysteine)

protein L39 2175 C peptidase inhibitor, clade 1.67393

B, member 6c(Serpinb6c) 2658

WTAP Wilms tumor 1 3.36457 STXBP4 syntaxin binding protein 4

associated protein 2432 1.67555

2278

RAD54L RAD54-like (S. 3.35658 RIMS3 regulating synaptic

cerevisiae) 9854 membrane exocytosis 3 1.67612

0648

CETN4 centrin 4(Cetn4) 3.33628 XYLT2 xylosyltransferase 2

3388 1.67697

6793

CEP55 centrosomal protein 55 3.32912 TAS2R107 taste receptor, type 2,

3596 member 107(Tas2rl07) 1.67807

1905

CYP4F39 cytochrome P450, family 3.32192 SKP1A S-phase kinase-associated

4, subfamily f, 8095 protein lA(Skpla) 1.67807 polypeptide 39(Cyp4f39) 1905

PTPN5 protein tyrosine 3.31469 OLFR165 olfactory receptor

phosphatase, non6526 165(01frl65) 1.67807 receptor type 5 1905

TUBEl tubulin epsilon 1 3.29278 OLFR111 olfactory receptor

1749 l l l(Olfrl l l) 1.67807

1905 subunit alpha 3675 and SH3 binding motif 1 1.68552

4532

TMEM53 transmembrane protein 3.14159 ATL2 atlastin GTPase 2

53 6278 1.68573

1341

THNSL2 threonine synthase like 2 3.14159 RAD52 RAD52 homolog, DNA

6278 repair protein 1.68952

3672

2810408M0 Description Not Found 3.12928 GPC1 glypican 1

9RIK 3017 1.68964

6894

ADAMDEC ADAM like decysin 1 3.12101 ARHGAP1 Rho GTPase activating

1 5401 5 protein 15 1.69080

4518

ASB2 ankyrin repeat and 3.11879 GPRC5B G protein-coupled receptor

SOCS box containing 2 2343 class C group 5 member B 1.69399

9744

SLC37A4 solute carrier family 37 3.11270 ZBTB1 zinc finger and BTB

member 4 0133 domain containing 1 1.69404

6727

NICNl nicolin 1 3.10847 NARFL nuclear prelamin A

8268 recognition factor like 1.69488

0193

2310067B1 Description Not Found 3.08746 SLC26A6 solute carrier family 26

ORIK 2841 member 6 1.69525

2347

PIGL pho sphatidy lino sitol 3.07723 MAPKBP1 mitogen-activated protein

glycan anchor 9787 kinase binding protein 1 1.69590 biosynthesis class L 8738

1190005106 Description Not Found 3.07038 RAB6B RAB6B, member RAS

RIK 9328 oncogene family 1.69754

1036

DHFR dihydrofolate reductase 3.07038 ARL2 ADP ribosylation factor

9328 like GTPase 2 1.70034

9879

FABP5 fatty acid binding protein 3.06608 ZFP646 zinc finger protein

5 919 646(Zfp646) 1.70043

9718

POMT2 protein O- 3.05579 SELENBP2 selenium binding protein

mannosyltransferase 2 4286 2(Selenbp2) 1.70043

9718

F2RL2 coagulation factor II 3.05311 ACOT3 acyl-CoA thioesterase

thrombin receptor like 2 1336 3(Acot3) 1.70043

9718

GRB7 growth factor receptor 3.04885 REG3G regenerating family

bound protein 7 2907 member 3 gamma 1.70043

9718

SNX21 sorting nexin family 3.04439 GAB1 GRB2 associated binding

member 21 4119 protein 1 1.70043

9718

SUFU SUFU negative regulator 3.04439 LCN10 lipocalin 10

of hedgehog signaling 4119 1.70043

9718

RFC3 replication factor C 3.02928 MTHFD2L methylenetetrahydrofolate

subunit 3 8361 dehydrogenase (NADP+ 1.70043 dependent) 2-like 9718

CLDN12 claudin 12 3.01792 PTCD3 pentatricopeptide repeat

1908 domain 3 1.70043

9718 C1QTNF6 Clq and tumor necrosis 3.01445 NTHL1 nth-like DNA glycosylase factor related protein 6 0679 1 1.70043

9718

PLCXD 1 pho sphatidy lino sitol 2.99095 NUDT3 nudix hydrolase 3

specific phospholipase C 486 1.70043 X domain containing 1 9718

SULT4A1 sulfotransferase family 2.99095 CLEC12A C-type lectin domain

4A member 1 486 family 12 member A 1.70043

9718

CTTNBP2N CTTNBP2 N-terminal 2.98185 ZBTB3 zinc finger and BTB

L like 2653 domain containing 3 1.70043

9718

SNX5 sorting nexin 5 2.97727 AMT aminomethyltransferase

9924 1.70043

9718

HPS5 HPS5, biogenesis of 2.97269 ZDHHC14 zinc finger DHHC-type

lysosomal organelles 2654 containing 14 1.70043 complex 2 subunit 2 9718

WISP1 W T1 inducible 2.96809 NKX2-5 NK2 homeobox 5

signaling pathway 0752 1.70049 protein 1 1519

PTPN9 protein tyrosine 2.96347 FOXA3 forkhead box A3

phosphatase, non4124 1.70281 receptor type 9 5694

USP37 ubiquitin specific 2.95419 WASF1 WAS protein family

peptidase 37 631 member 1 1.70641

2734

SH3BGRL SH3 domain binding 2.93545 OLFR690 olfactory receptor

glutamate rich protein 9748 690(Olfr690) 1.70719 like 2688

NCALD neurocalcin delta 2.93545 ENTPD5 ectonucleoside

9748 triphosphate 1.70776 diphosphohydrolase 5 4551

CDC42EP4 CDC42 effector protein 2.91647 PCDHGA4 protocadherin gamma

4 6644 subfamily A, 4 1.70904

2655

IGFBP7 insulin like growth factor 2.91055 TCF12 transcription factor 12

binding protein 7 3168 1.71030

8209

ABHD4 ab hydrolase domain 2.90886 MTRR 5-methyltetrahydrofolate- containing 4 8748 homocysteine 1.71149 methyltransferase 4907 reductase

CSF1 colony stimulating factor 2.90689 CDKN1C cyclin dependent kinase

1 0596 inhibitor 1C 1.71169

0028

COX7A1 cytochrome c oxidase 2.89724 PRICKLEl prickle planar cell polarity

subunit 7A1 0426 protein 1 1.71341

0822

TTYH2 tweety family member 2 2.89239 ATXN7L1 ataxin 7 like 1

1026 1.71669

984

ACOl aconitase 1 2.87774 SLC03A1 solute carrier organic

425 anion transporter family 1.71923 member 3A1 5762

BARD1 BRCA1 associated 2.86789 TMEM110 transmembrane protein

RING domain 1 6464 110 1.72004

6704

GPN1 GPN-loop GTPase 1 2.86789 KLF2 Kruppel like factor 2 - 6464 1.72137

4729

PTTG1 pituitary tumor- 2.86789 FGG fibrinogen gamma chain

transforming 1 6464 1.72246

6024

2810408A1 Description Not Found 2.85798 ASAH2 N-acylsphingosine

1RIK 0995 amidohydrolase 2 1.72246

6024

BBX BBX, HMG-box 2.85798 LAP3 leucine aminopeptidase 3

containing 0995 1.72246

6024

LTBP3 latent transforming 2.83794 STAB 2 stabilin 2

growth factor beta 3242 1.72246 binding protein 3 6024

ACTG2 acfin, gamma 2, smooth 2.82781 IL22RA1 interleukin 22 receptor

muscle, enteric 9025 subunit alpha 1 1.72246

6024

ISLR immunoglobulin 2.82781 SERINC4 serine incorporator 4

superfamily containing 9025 1.72246 leucine rich repeat 6024

NARS2 asparaginyl-fRNA 2.82308 GPR180 G protein-coupled receptor

synthetase 2, 7408 180 1.72246 mitochondrial (putative) 6024

ICAM4 intercellular adhesion 2.81452 TIPARP TCDD inducible

molecule 4 (Landsteiner- 379 poly(ADP-ribose) 1.72246 Wiener blood group) polymerase 6024

ABCB8 ATP binding cassette 2.81335 USP11 ubiquitin specific

subfamily B member 8 8991 peptidase 11 1.72246

6024

IDI1 isopentenyl-diphosphate 2.81178 TRIP6 thyroid hormone receptor

delta isomerase 1 2922 interactor 6 1.72246

6024

GLS2 glutaminase 2 2.79701 KCNH2 potassium voltage-gated

2978 channel subfamily H 1.72246 member 2 6024

HDAC8 histone deacetylase 8 2.79701 ESR2 estrogen receptor 2

2978 1.72246

6024

BRIP1 BRCA1 interacting 2.79701 FGF13 fibroblast growth factor 13

protein C-terminal 2978 1.72263 helicase 1 9247

USP6NL USP6 N-terminal like 2.79441 KBTBD7 kelch repeat and BTB

5866 domain containing 7 1.72423

7927

TLCD2 TLC domain containing 2.79181 UHRF1BP1 UHRF1 binding protein 1

2 4071 1.72583

5292

GUCY1A3 guanylate cyclase 1 2.78750 BCAM basal cell adhesion

soluble subunit alpha 2763 molecule (Lutheran blood 1.72650 group) 9704

OCA2 OCA2 melanosomal 2.78659 ELOVL6 ELOVL fatty acid

transmembrane protein 6362 elongase 6 1.72656

5554

VAT1 vesicle amine transport 1 2.77250 PPM1K protein phosphatase,

2543 Mg2+/Mn2+ dependent 1.72664

IK 3643

HIST1H2A histone cluster 1, H2ab 2.76791 SPATA6 spermatogenesis

B 4142 associated 6 1.72767

3077 PIGC pho sphatidy lino sitol 2.76022 NAV1 neuron navigator 1

glycan anchor 0946 1.72792 biosynthesis class C 0455

PARG poly(ADP-ribose) 2.75655 ANK3 ankyrin 3, node of Ranvier

glycohydrolase 8208 (ankyrin G) 1.72792

0455

ESC02 establishment of sister 2.75488 KCNAB 1 potassium voltage-gated

chromatid cohesion N- 7502 channel subfamily A 1.72792 acetyltransferase 2 member regulatory beta 0455 subunit 1

HIPK2 homeodomain 2.75488 CYP27A1 cytochrome P450 family

interacting protein 7502 27 subfamily A member 1 1.72792 kinase 2 0455

ΓΜΡΑ1 inositol 2.75294 MAP4K4 mitogen-activated protein

monophosphatase 1 5007 kinase kinase kinase 1.72975 kinase 4 6006

COQ4 coenzyme Q4 2.74416 ANKRD7 ankyrin repeat domain 7

1096 1.73064

6873

ZBTB7A zinc finger and BTB 2.74416 IFRD1 interferon related

domain containing 7A 1096 developmental regulator 1 1.73244

7522

GAMT guanidinoacetate N- 2.74416 ALX3 ALX homeobox 3

methyltransferase 1096 1.73335

4341

BIK BCL2 interacting killer 2.74416 SNURF SNRPN upstream reading

1096 frame 1.73335

4341

PMS1 PMS1 homolog 1, 2.73335 AMZ2 archaelysin family

mismatch repair system 4341 metallopeptidase 2 1.73350 component 053

HAVCR2 hepatitis A virus cellular 2.72976 ROGDI rogdi homolog

receptor 2 9667 1.73419

198

FHL2 four and a half LIM 2.72725 DAGLA diacylglycerol lipase alpha

domains 2 4747 1.73447

1203

CHAF1A chromatin assembly 2.72524 4930432K2 Description Not Found

factor 1 subunit A 8783 IRK 1.73624

3886

2810004N2 Description Not Found 2.72246 KRCC1 lysine rich coiled-coil 1

3RIK 6024 1.73665

741

TBC1D 14 TBC1 domain family 2.72246 OLFR1331 olfactory receptor

member 14 6024 1331(01frl331) 1.73682

6447

EHD2 EH domain containing 2 2.71149 SLC25A25 solute carrier family 25

4907 member 25 1.73690

749

APH1A aph-1 homolog A, 2.70597 CXCR4 C-X-C motif chemokine

gamma-secretase subunit 7902 receptor 4 1.73777

9353

TMEM2 transmembrane protein 2 2.70321 EPB4.1L3 Description Not Found

1467 1.73876

7837

LCAT lecithin-cholesterol 2.70043 CEP 164 centrosomal protein 164

acyltransf erase 9718 1.73879

5736

FBX015 F-box protein 15 2.68929 AGER advanced glycosylation - CRYBG3 crystallin beta-gamma 2.60539 BLK BLK proto-oncogene, Src domain containing 3 3551 family tyrosine kinase 1.74953

4268

DIXDCl DIX domain containing 2.59694 MGAT5 mannosyl (alpha- 1,6-)- 1 0379 glycoprotein beta- 1 ,6-N- 1.74953 acetyl- 4268 glucosaminyltransf erase

TACSTD2 tumor-associated 2.59392 RNF2 ring finger protein 2

calcium signal 6161 1.75089 transducer 2 0228

TRP53RK Description Not Found 2.58806 COL14A1 collagen type XIV alpha 1

6506 chain 1.75209

3722

PDCD 1LG2 programmed cell death 1 2.58496 PLEKHG3 pleckstrin homology and

ligand 2 2501 RhoGEF domain 1.75210 containing G3 9698

SEC23IP SEC23 interacting 2.58496 ARHGEF18 Rho/Rac guanine

protein 2501 nucleotide exchange factor 1.75410

18 0479

ORM1 orosomucoid 1 2.58496 LEF1 lymphoid enhancer

2501 binding factor 1 1.75488

7502

ZFP322A zinc finger protein 2.57502 COMMD9 COMM domain containing

322A(Zfp322a) 4164 9 1.75490

709

4931406C0 Description Not Found 2.56071 SLC20A1 solute carrier family 20

7RIK 4954 member 1 1.75863

7847

ZFP382 zinc finger protein 2.56071 ACTR5 ARP5 actin-related protein

382(Zfp382) 4954 5 homolog 1.75924

4091

CLIP2 CAP-Gly domain 2.56071 UBQLN3 ubiquilin 3

containing linker protein 4954 1.76510 2 9548

TNFAIP8L TNF alpha induced 2.56071 ZFP770 zinc finger protein

1 protein 8 like 1 4954 770(Zfp770) 1.76553

4746

NPvCAM neuronal cell adhesion 2.56071 PCDHB18 protocadherin beta

molecule 4954 18(Pcdhbl8) 1.76553

4746

HPSE heparanase 2.56071 OLFR700 olfactory receptor

4954 700(Olfr700) 1.76553

4746

RTKN rhotekin 2.55898 FOXP4 forkhead box P4

5655 1.76553

4746

DLGAP5 DLG associated protein 2.55012 CDC34 cell division cycle 34

5 5328 1.76553

4746

ENPP2 ectonucleotide 2.54843 HIST1H1E histone cluster 1, Hie

py ropho sphatase/pho sph 6625 1.76553 odiesterase 2 4746

GCNT1 glucosaminyl (N-acetyl) 2.54843 G6PC2 glucose-6-phosphatase

transferase 1, core 2 6625 catalytic subunit 2 1.76553

4746

SASS6 SAS-6 centriolar 2.54843 FUT1 fucosyltransferase 1 (H

assembly protein 6625 blood group) 1.76553

4746

AMIG03 adhesion molecule with 2.54843 ZFP69 ZFP69 zinc finger protein - 13

MX1 MX dynamin like 2.48542 RHOX4B reproductive homeobox

GTPase 1 6827 4B(Rhox4b) 1.78659

6362

SMTN smoothelin 2.48542 OLFR1134 olfactory receptor

6827 1134(01frl l34) 1.78659

6362

PLA2G15 phospholipase A2 group 2.48194 CAR11 carbonic anhydrase

XV 563 l l(Carl l) 1.78659

6362

OLFR192 olfactory receptor 2.47248 LRRIQ4 leucine rich repeats and IQ

192(01frl92) 7771 motif containing 4 1.78659

6362

ITGB5 integrin subunit beta 5 2.47248 C ASP 12 caspase 12

7771 (gene/pseudogene) 1.78659

6362

RAPSN receptor associated 2.46597 ODF3L1 outer dense fiber of sperm

protein of the synapse 4465 tails 3 like 1 1.78659

6362

SNX3 sorting nexin 3 2.45943 CCDC3 coiled-coil domain

1619 containing 3 1.78659

6362

FERMT2 fermitin family member 2.45943 SSPN sarcospan

2 1619 1.78659

6362

CCR5 C-C motif chemokine 2.44441 KLK1 kallikrein 1

receptor 5 0478 1.78659 (gene/pseudogene) 6362

UPK1A uroplakin 1A 2.43962 SENP7 SUMOl/sentrin specific

3138 peptidase 7 1.78689

7131

BCL2L2 BCL2 like 2 2.43629 CAML calcium modulating

512 ligand(Caml) 1.78773

5284

2610002M0 Description Not Found 2.43295 YEATS2 YEATS domain

6RIK 9407 containing 2 1.78862

7083

CENPN centromere protein N 2.43295 SERPINF2 serpin family F member 2

9407 1.79181

4071

HBEGF heparin binding EGF 2.43096 KCNMB1 potassium calcium- like growth factor 254 activated channel 1.79259 subfamily M regulatory 7191 beta subunit 1

TYMS thymidylate synthetase 2.42710 FCH02 FCH domain only 2

3287 1.79266

6489

MGA MGA, MAX 2.42693 BBS9 Bardet-Biedl syndrome 9

dimerization protein 9834 1.79273

4984

RAI14 retinoic acid induced 14 2.42626 OLFR323 olfactory receptor

4755 323(01fr323) 1.79460

9131

CFI complement factor I 2.41953 CD247 CD247 molecule

8892 1.79608

1585

PLK4 polo like kinase 4 2.41953 HIST2H2A histone cluster 2,

8892 Al H2aal(Hist2h2aal) 1.79684

7743 SLC6A9 solute carrier family 6 2.41953 PDK1 pyruvate dehydrogenase member 9 8892 kinase 1 1.80056

3818

TMED2 transmembrane p24 2.41953 NRARP NOTCH-regulated ankyrin

trafficking protein 2 8892 repeat protein 1.80304

9246

TMEM120 transmembrane protein 2.41857 BTBD11 BTB domain containing

B 120B 423 11 1.80479

3263

TRIM36 tripartite motif 2.41785 CSF2RA colony stimulating factor 2

containing 36 2515 receptor alpha subunit 1.80508

9518

CCDC93 coiled-coil domain 2.41616 DEXI Dexi homolog

containing 93 4165 1.80699

8156

SLC25A35 solute carrier family 25 2.40936 OLFR1276 olfactory receptor

member 35 7225 1276(01frl276) 1.80735

4922

BNC1 basonuclin 1 2.40599 TCSTV3 2-cell-stage, variable

236 group, member 3(Tcstv3) 1.80735

4922

FOXL2 forkliead box L2 2.40599 SPRR2D small proline rich protein

236 2D 1.80735

4922

TFPI2 tissue factor pathway 2.40599 SEMA4G semaphorin 4G

inhibitor 2 236 1.80735

4922

NET1 neuroepithelial cell 2.40599 KCNK9 potassium two pore

transforming 1 236 domain channel subfamily 1.80735

K member 9 4922

SLC02A1 solute carrier organic 2.40599 SNAPC3 small nuclear RNA

anion transporter family 236 activating complex 1.80738 member 2A1 polypeptide 3 5513

A730008H2 Description Not Found 2.39927 AXIN2 axin 2

3RIK 5037 1.80842

9403

CDKN2B cyclin dependent kinase 2.39726 PCNXL3 Description Not Found

inhibitor 2B 4578 1.80899

5133

ZFP532 zinc finger protein 2.39313 KLHL7 kelch like family member

532(Zfp532) 8801 7 1.80901

6035

GTSE1 G2 and S -phase 2.39242 ZFP281 zinc finger protein

expressed 1 8431 281(Zfp281) 1.81155

6991

CCDC14 coiled-coil domain 2.39231 CHRNB2 cholinergic receptor

containing 14 7423 nicotinic beta 2 subunit 1.81249

8225

AD ATI adenosine deaminase, 2.39231 TBC1D 15 TBCl domain family

fRNA specific 1 7423 member 15 1.81290

9044

DGKH diacylglycerol kinase eta 2.39231 GALNT9 polypeptide N- 7423 acetylgalactosaminyltransf 1.81340 erase 9 7449

ZRSR1 zinc finger CCCH-type, 2.39231 DYNC1I1 dynein cytoplasmic 1

RNA binding motif and 7423 intermediate chain 1 1.81343 serine/arginine rich 1 4179

NFE2 nuclear factor, erythroid 2.39152 MYH8 myosin heavy chain 8

2 9377 1.81403 224

CD63 CD63 molecule 2.38785 CEP57 centrosomal protein 57

3137 1.81568

4972

MIB1 mindbomb E3 ubiquitin 2.38645 LTK leukocyte receptor tyrosine

protein ligase 1 559 kinase 1.81762

3258

TSN translin 2.38234 COMMD2 COMM domain containing

9023 2 1.81762

3258

2510003E0 Description Not Found 2.37851 MEF2C myocyte enhancer factor

4RIK 1623 2C 1.81762

3258

BC043934 cDNA sequence 2.37851 LONRF2 LON peptidase N-terminal

BC043934(BC043934) 1623 domain and ring finger 2 1.81794

1412

AHCYL1 adenosylhomocysteinase 2.36673 PDCD6IP programmed cell death 6

like 1 4247 interacting protein 1.82057

5529

OLFR731 olfactory receptor 2.36457 DHX16 DEAH-box helicase 16

731(01fr731) 2432 1.82066

1084

CDKN2A cyclin dependent kinase 2.36457 ZFYVE19 zinc finger FYVE-type

inhibitor 2A 2432 containing 19 1.82528

1028

SLC29A4 solute carrier family 29 2.36457 H2-T10 histocompatibility 2, T

member 4 2432 region locus 10(H2-T10) 1.82621

8639

SLC4A10 solute carrier family 4 2.36457 ARID 1 A AT-rich interaction

member 10 2432 domain 1A 1.82704

3205

CYCS cytochrome c, somatic 2.35187 NODI nucleotide binding

2866 oligomerization domain 1.82718 containing 1 5706

COL5A1 collagen type V alpha 1 2.35049 2610318N0 Description Not Found

7247 2RIK 1.82781

9025

UTRN utrophin 2.35049 BC048644 cDNA sequence

7247 BC048644(BC048644) 1.82781

9025

AURKA aurora kinase A 2.34967 CDC42EP2 CDC42 effector protein 2

8136 1.82781

9025

KREMEN2 kringle containing 2.34943 CCL25 C-C motif chemokine

transmembrane protein 2 1709 ligand 25 1.82781

9025

FGL2 fibrinogen like 2 2.34640 TBX6 T-box 6

9407 1.82781

9025

NCAM1 neural cell adhesion 2.34340 PLEKHG4 pleckstrin homology and

molecule 1 7822 RhoGEF domain 1.82781 containing G4 9025

ALG8 ALG8, alpha-1,3- 2.34340 RAD18 RAD18, E3 ubiquitin

glucosyltransferase 7822 protein ligase 1.83064

2494

OLFR703 olfactory receptor 2.33628 SLC12A9 solute carrier family 12

703(Olfr703) 3388 member 9 1.83080

7586

SLC39A10 solute carrier family 39 2.33628 NR1D2 nuclear receptor subfamily - member 10 3388 1 group D member 2 1.83794

3242

HIST1H2A histone cluster 1, H2ali 2.32214 NLK nemo like kinase

H 1712 1.84017

0811

TSGA8 testis specific gene 2.32192 TTC37 tetratricopeptide repeat

A8(Tsga8) 8095 domain 37 1.84046

2743

ELOVL2 ELOVL fatty acid 2.32192 DLG3 discs large MAGUK

elongase 2 8095 scaffold protein 3 1.84150

7525

MLF1 myeloid leukemia factor 2.32192 PCF11 PCF11 cleavage and

1 8095 polyadenylation factor 1.84334 subunit 9827

FZD6 frizzled class receptor 6 2.32192 HIST1H4D histone cluster 1, H4d

8095 1.84638

6944

PLD1 phospho lipase Dl 2.32192 PEX26 peroxisomal biogenesis

8095 factor 26 1.84744

0096

IFRD2 interferon-related 2.32192 CYP2B 10 cytochrome P450, family

developmental regulator 8095 2, subfamily b, 1.84799 2 polypeptide 10(Cyp2blO) 6907

OLA1 Obg-like ATPase 1 2.32192 GDF3 growth differentiation

8095 factor 3 1.84799

6907

ASPA aspartoacylase 2.32192 GPR33 G protein-coupled receptor

8095 33 (gene/pseudogene) 1.84799

6907

TGFB3 transforming growth 2.32192 TDG thymine DNA glycosylase

factor beta 3 8095 1.84799

6907

PKIG protein kinase (c AMP- 2.31469 HIPK3 homeodomain interacting

dependent, catalytic) 6526 protein kinase 3 1.84799 inhibitor gamma 6907

TNFRSF4 tumor necrosis factor 2.30883 PAPOLA poly(A) polymerase alpha

receptor superfamily 2886 1.84799 member 4 6907

IQCB1 IQ motif containing B 1 2.30798 MAPK4 mitogen-activated protein

4443 kinase 4 1.84799

6907

SLC16A11 solute carrier family 16 2.30766 FRAT2 frequently rearranged in

member 11 2797 advanced T-cell 1.84969 lymphomas 2 115

1190002N1 Description Not Found 2.30742 HEXIM1 hexamethylene

5RIK 8525 bisacetamide inducible 1 1.85103

5845

LCE1L late cornified envelope 2.30742 TATDN2 TafD DNase domain

lL(Lcell) 8525 containing 2 1.85143

3223

RGS13 regulator of G-protein 2.30742 KLRB1C killer cell lectin-like

signaling 13 8525 receptor subfamily B 1.85425 member lC(Klrblc) 3843

FBXW8 F-box and WD repeat 2.29998 SLC16A9 solute carrier family 16

domain containing 8 7517 member 9 1.85508

3462

SNCA synuclein alpha 2.29645 ACBD4 acyl-CoA binding domain

7407 containing 4 1.85573

9032 6464

TBX1 T-box 1 2.24792 PCTP phosphatidylcholine

7513 transfer protein 1.86789

6464

DHRS13 dehydrogenase/reductase 2.24792 CALD1 caldesmon 1

13 7513 1.86789

6464

HSPG2 heparan sulfate 2.24792 TREML2 triggering receptor

proteoglycan 2 7513 expressed on myeloid cells 1.86789 like 2 6464

FRMD8 FERM domain 2.24777 RTN4RL1 reticulon 4 receptor like 1

containing 8 312 1.86789

6464

MIOX myo-inositol oxygenase 2.24057 PARVA parvin alpha

9987 1.86847

9018

LYRM1 LYR motif containing 1 2.23266 NPCD neuronal pentraxin chromo

0757 domain(Npcd) 1.87190

2039

ST API signal transducing 2.23266 RFXANK regulatory factor X

adaptor family member 1 0757 associated ankyrin 1.87206 containing protein 109

NAT2 N-acetyltransferase 2 2.23266 MAP3K14 mitogen-activated protein

0757 kinase kinase kinase 14 1.87229

1304

SRGAP3 SLIT-ROBO Rho 2.23266 KLHL9 kelch like family member

GTPase activating 0757 9 1.87452 protein 3 8943

NXT2 nuclear transport factor 2 2.23266 SESN1 sestrin 1

like export factor 2 0757 1.87526

0951

RCOR1 REST corepressor 1 2.23266 ADAMTS7 ADAM metallopeptidase

0757 with thrombospondin type 1.87940

1 motif 7 4807

SRR serine racemase 2.23083 SNAPC1 small nuclear RNA

6503 activating complex 1.88488 polypeptide 1 993

IKBKAP inhibitor of kappa light 2.22617 ADAR adenosine deaminase,

polypeptide gene 7109 RNA specific 1.88529 enhancer in B-cells, 9379 kinase complex- associated protein

AI597479 expressed sequence 2.22581 LCE1C late cornified envelope 1C

AI597479(AI597479) 9675 1.88562

6461

POP1 POP1 homolog, 2.22496 FBX021 F-box protein 21

ribonuclease P/MRP 6365 1.88615 subunit 5099

SLC35E4 solute carrier family 35 2.21723 2610524H0 Description Not Found

member E4 0716 6RIK 1.88752

5271

XAB2 XPA binding protein 2 2.21723 1700016K1 Description Not Found

0716 9RIK 1.88752

5271

MREG melanoregulin 2.21292 ZFP715 zinc finger protein

58 715(Zfp715) 1.88752

5271

FKBP11 FK506 binding protein 2.21072 OLFR446 olfactory receptor

11 1954 446(01fr446) 1.88752 IGF2BP2 insulin like growth factor 2.20778 PTK7 protein tyrosine kinase 7

2 mRNA binding protein 9851 (inactive) 1.88752 2 5271

NUP133 nucleoporin 133 2.20744 TMEM117 transmembrane protein

7199 117 1.88752

5271

OLFR1183 olfactory receptor 2.20163 ITIH2 inter-alpha-trypsin

1183(01frl l83) 3861 inhibitor heavy chain 2 1.88752

5271

IL1F6 interleukin 1 family, 2.20163 TAGLN3 transgelin 3

member 6(11 lf6) 3861 1.88752

5271

OTX1 orthodenticle homeobox 2.20163 IFI203 interferon activated gene

1 3861 203(Ifi203) 1.88764

4112

MSH3 mutS homolog 3 2.20163 ATP1B1 ATPase Na+/K+

3861 transporting subunit beta 1 1.88766

4186

SCN4B sodium voltage-gated 2.20163 BLCAP bladder cancer associated

channel beta subunit 4 3861 protein 1.88859

6201

CROCC ciliary rootlet coiled- 2.20163 IGF1R insulin like growth factor 1

coil, rootletin 3861 receptor 1.89024

137

NSUN2 NOP2/Sun RNA 2.19434 HMG20A high mobility group 20A

methyltransferase family 9986 1.89057 member 2 9593

GAS2L1 growth arrest specific 2 2.19377 WDR24 WD repeat domain 24

like 1 1743 1.89152

7175

3110007F17 Description Not Found 2.19074 CDX4 caudal type homeobox 4

RIK 0399 1.89265

5439

DEFB 15 defensin beta 2.18586 CLDN18 claudin 18

15(Defbl5) 6545 1.89344

9375

C1QTNF2 Clq and tumor necrosis 2.18586 IL4RA interleukin 4 receptor,

factor related protein 2 6545 alpha(I14ra) 1.89536

9594

RAP 1 GAP RAP1 GTPase activating 2.18586 RETNLA resistin like alpha(Retnla)

protein 6545 1.89573

9477

SNTB 1 syntrophin beta 1 2.18586 AA388235 expressed sequence

6545 AA388235(AA388235) 1.89573

9477

FAH fumarylacetoacetate 2.18292 ZC3H6 zinc finger CCCH-type

hydrolase 5576 containing 6 1.89612

7489

AVPI1 arginine vasopressin 2.17439 D930015E0 RIKEN cDNA

induced 1 3775 6RIK D930015E06 1.89965 gene(D930015E06Rik) 6973

RPA2 replication protein A2 2.17275 NPFFR2 neuropeptide FF receptor 2

1912 1.90207

3579

BRCA2 BRCA2, DNA repair 2.16873 IRAKI interleukin 1 receptor

associated 2488 associated kinase 1 1.90243

374

RBM47 RNA binding motif 2.16591 CWF19L2 CWF19-like 2, cell cycle - protein 47 1939 control (S. pombe) 1.90370

4505

MSL3L2 male-specific lethal 3- 2.15906 STK40 serine/threonine kinase 40

like 2 1455 1.90396

(Drosophila)(Msl312) 4448

TNFRSF9 tumor necrosis factor 2.15607 MARS2 methionyl-fRNA

receptor superfamily 1704 synthetase 2, 1.90457 member 9 mitochondrial 1951

TRF transferrin(Trf) 2.15458 RAB5A RAB5A, member RAS

8207 oncogene family 1.90635

0687

ZDHHC15 zinc finger DHHC-type 2.15437 OLFR1037 olfactory receptor

containing 15 2546 1037(Olfrl037) 1.90689

0596

IGJ Description Not Found 2.15380 ARHGAP2 Rho GTPase activating

5336 2 protein 22 1.90689

0596

FBX027 F-box protein 27 2.15380 DENND IB DENN domain containing

5336 IB 1.90689

0596

ZDHHC24 zinc finger DHHC-type 2.15380 EAPP E2F associated

containing 24 5336 phosphoprotein 1.90689

0596

SPCS2 signal peptidase complex 2.15380 ANKRD 13 ankyrin repeat domain

subunit 2 5336 D 13D 1.90689

0596

UCN3 urocortin 3 2.15380 EFCAB2 EF-hand calcium binding

5336 domain 2 1.90689

0596

SLC35A1 solute carrier family 35 2.15380 HOXC9 homeobox C9

member Al 5336 1.90689

0596

PODXL podocalyxin like 2.15380 SENP6 SUMOl/sentrin specific

5336 peptidase 6 1.90795

6932

FAM154B Description Not Found 2.15379 SIDT1 SID 1 transmembrane

2145 family member 1 1.90828

6674

NRP1 neuropilin 1 2.14747 2310057J18 Description Not Found

0553 RIK 1.91647

6644

ERGIC1 endoplasmic reficulum- 2.14710 SPRYD4 SPRY domain containing

golgi intermediate 4727 4 1.91647 compartment 1 6644

RNF26 ring finger protein 26 2.14681 LY6D lymphocyte antigen 6

0011 complex, locus D 1.91647

6644

LCN3 lipocalin 3(Lcn3) 2.13750 PPARGC1B PPARG coactivator 1 beta

3524 1.91729

1956

FMOl flavin containing 2.13750 SH3TC1 SH3 domain and

monooxygenase 1 3524 tetratricopeptide repeats 1 1.91790

6346

RAB20 RAB20, member RAS 2.13750 FOXOl forkhead box 01

oncogene family 3524 1.92020

9106

KATNAL1 katanin catalytic subunit 2.13750 DHX40 DEAH-box helicase 40

Al like 1 3524 1.92062

3917 GPR107 G protein-coupled 2.13642 RECQL5 RecQ like helicase 5

receptor 107 4717 1.92066

4575

MELK maternal embryonic 2.13339 RBM15 RNA binding motif

leucine zipper kinase 9125 protein 15 1.92261

6041

KCTD9 potassium channel 2.13207 EGLN2 egl-9 family hypoxia

tetramerization domain 329 inducible factor 2 1.92407 containing 9 9933

PBK PDZ binding kinase 2.13041 GPR112 Description Not Found

7144 1.92599

9419

ENPP5 ectonucleotide 2.12411 OLFR829 olfactory receptor

py ropho sphatase/pho sph 2676 829(01fr829) 1.92599 odiesterase 5 (putative) 9419

ZDHHC16 zinc finger DHHC-type 2.12361 OLFR684 olfactory receptor

containing 16 008 684(01fr684) 1.92599

9419

OLFR1346 olfactory receptor 2.12101 RETN resistin

1346(01frl346) 5401 1.92599

9419

MILL1 MHC I like leukocyte 2.12101 ST6GALN ST6 N- l(Milll) 5401 AC2 acetylgalactosaminide 1.92599 alpha-2,6-sialyltransferase 9419

2

RHCG Rh family C 2.12101 FES FES proto-oncogene,

glycoprotein 5401 tyrosine kinase 1.92599

9419

CLDN1 claudin 1 2.12101 KIF13A kinesin family member

5401 13A 1.92599

9419

LHX3 LIM homeobox 3 2.12101 TRPT1 tRNA phosphotransferase

5401 1 1.92645

7816

TUBB2A tubulin beta 2 A class Ila 2.12101 PLCB2 phospholipase C beta 2

5401 1.92734

3833

GSG2 germ cell associated 2, 2.11941 NADSYN1 NAD synthetase 1

haspin 2265 1.92967

4394

HYAL2 hyaluronoglucosaminida 2.10734 4833420G1 Description Not Found

se 2 5942 7RIK 1.93060

469

1700003F12 Description Not Found 2.10433 P2RY10 purinergic receptor P2Y10

RIK 666 1.93073

7338

RUSC2 RUN and SH3 domain 2.10433 PPAPDC3 Description Not Found

containing 2 666 1.93545

9748

LRRIQ3 leucine rich repeats and 2.10433 DIP2B disco interacting protein 2

IQ motif containing 3 666 homolog B 1.93545

9748

CHSY1 chondroitin sulfate 2.10433 RHAG Rh-associated

synthase 1 666 glycoprotein 1.93545

9748

DUSP23 dual specificity 2.10433 EMTDl EMI domain containing 1

phosphatase 23 666 1.93545

9748

RRAGB Ras related GTP binding 2.10433 RNF4 ring finger protein 4 - B 666 1.93883

4579

KCNAB3 potassium voltage-gated 2.10433 UBL5 ubiquitin like 5

channel subfamily A 666 1.93895 regulatory beta subunit 3 2478

GRPEL2 GrpE like 2, 2.10312 PROSC proline synthetase

mitochondrial 9681 cotranscribed homolog 1.94016

(bacterial) 675

TRAF2 TNF receptor associated 2.10202 FZD5 frizzled class receptor 5

factor 2 9095 1.94250

3137

COQ7 coenzyme Q7, 2.10020 UBE2D1 ubiquitin conjugating

hydroxylase 5246 enzyme E2 D 1 1.94277

5467

TMEM126 transmembrane protein 2.09918 KLRA7 killer cell lectin-like

B 126B 7297 receptor, subfamily A, 1.94351 member 7(Klra7) 0757

SGPL1 sphingosine- 1 -phosphate 2.09711 TMEM63C transmembrane protein

lyase 1 2667 63 C 1.94425

562

CAPN2 calpain 2 2.09644 2810006K2 Description Not Found

7979 3RIK 1.94485

8446

CHEK2 checkpoint kinase 2 2.08845 OLFR672 olfactory receptor

7439 672(01fr672) 1.94485

8446

GLRP1 glutamine repeat protein 2.08746 OLFR1347 olfactory receptor

l(Glrpl) 2841 1347(01frl347) 1.94485

8446

RTN4R reticulon 4 receptor 2.08746 MTTP microsomal triglyceride

2841 transfer protein 1.94485

8446

TRIM37 tripartite motif 2.08746 MSX1 msh homeobox 1

containing 37 2841 1.94485

8446

NUCB2 nucleobindin 2 2.08746 BSND barttin CLCNK type

2841 accessory beta subunit 1.94485

8446

UBE2T ubiquitin conjugating 2.07361 MARK1 microtubule affinity

enzyme E2 T 6696 regulating kinase 1 1.94485

8446

CREB3L3 cAMP responsive 2.07038 CHRNB 1 cholinergic receptor

element binding protein 9328 nicotinic beta 1 subunit 1.94485 3 like 3 8446

CHRM4 cholinergic receptor 2.07038 CRYL1 crystallin lambda 1

muscarinic 4 9328 1.94641

9425

SLC16A13 solute carrier family 16 2.07038 TEC tec protein tyrosine kinase

member 13 9328 1.94733

0641

OLFML2B olfactomedin like 2B 2.07038 XKR6 XK related 6

9328 1.95031

589

CSNK1G1 casein kinase 1 gamma 1 2.07038 ARC activity-regulated

9328 cytoskeleton-associated 1.95363 protein 6949

S100A14 S100 calcium binding 2.07038 WFDC10 WAP four-disulfide core

protein A14 9328 domain 10(Wfdc 10) 1.95419

631 CDC6 cell division cycle 6 2.01116 CSF1R colony stimulating factor 1

6077 receptor 2.00635

0699

AXL AXL receptor tyrosine 2.00813 ZFP1 ZFP1 zinc finger protein

kinase 1619 2.00790

4843

RBBP7 RB binding protein 7, 2.00674 SFN stratifin

chromatin remodeling 6832 2.00898 factor 8783

PABPC4 poly (A) binding protein 2.00526 COL17A1 collagen type XVII alpha 1

cytoplasmic 4 0152 2.01038

6372

HIST1H2A histone cluster 1, H2ak 2.00330 XKRX XK related, X-linked

K 7679 2.01056

96

MTFMT mitochondrial 2.00175 BRD8 bromodomain containing 8

methionyl-fRNA 4595 2.01346 formyltransferase 226

ZFP449 zinc finger protein 2 ZFP213 zinc finger protein

449(Zfp449) 213(Zfp213) 2.01353

2276

D930020B 1 RIKEN cDNA 2 ZFY2 zinc finger protein 2, Y- 8RIK D930020B18 linked(Zfy2) 2.01565 gene(D930020B18Rik) 7249

LCE1D late cornified envelope 2 MAP3K3 mitogen-activated protein

ID kinase kinase kinase 3 2.01612

652

UCN urocortin 2 ZFP445 zinc finger protein

445(Zfp445) 2.01792

1908

SYT4 synaptotagmin 4 2 MTAP7D3 MAP7 domain containing

3(Mtap7d3) 2.01792

1908

GPR132 G protein-coupled 2 TMPRSSl l transmembrane protease,

receptor 132 A serine 11A 2.01792

1908

SDHD succinate dehydrogenase 2 OLFM2 olfactomedin 2

complex subunit D 2.01792

1908

PANK3 pantothenate kinase 3 2 GRM4 glutamate metabotropic

receptor 4 2.01792

1908

SBSN suprabasin 1.99095 ONECUT2 one cut homeobox 2

486 2.01792

1908

WDR59 WD repeat domain 59 1.98997 HNRNPH3 heterogeneous nuclear

6974 ribonucleoprotein H3 2.01792

1908

MTMR9 myotubularin related 1.98784 ZMYM5 zinc finger MYM-type

protein 9 4644 containing 5 2.02020

4421

IL15RA interleukin 15 receptor 1.98562 RAPGEF6 Rap guanine nucleotide

subunit alpha 8881 exchange factor 6 2.02095

3989

RHBDF2 rhomboid 5 homolog 2 1.98468 CD34 CD34 molecule

1148 2.02671

4044

NHLRC2 NHL repeat containing 2 1.98375 ACVR2B activin A receptor type 2B

117 2.02671 4044

NMRAL1 NmrA-like family 1.98337 RILP Rab interacting lysosomal

domain containing 1 0163 protein 2.02680

0059

OLFR120 olfactory receptor 1.98185 EMR1 Description Not Found

120(Olfrl20) 2653 2.03121

8731

OLFR1051 olfactory receptor 1.98185 DNAJA2 DnaJ heat shock protein

1051(Olfrl051) 2653 family (Hsp40) member 2.03129

A2 1874

PCDHGA9 protocadherin gamma 1.98185 SEMA4B semaphorin 4B

subfamily A, 9 2653 2.03198

5281

FST follistatin 1.98185 1700015E1 Description Not Found

2653 3RIK 2.03562

391

RECQL4 RecQ like helicase 4 1.97661 RHOX1 reproductive homeobox

1605 l(Rhoxl) 2.03562

391

NF BIL1 NFKB inhibitor like 1 1.97096 TCP 11 t-complex 11

9489 2.03562

391

TUBD1 tubulin delta 1 1.96436 FBXW11 F-box and WD repeat

7355 domain containing 11 2.03562

391

FSD1 fibronectin type III and 1.96347 ALX1 ALX homeobox 1

SPRY domain 4124 2.03562 containing 1 391

GDF5 growth differentiation 1.96347 BST1 bone marrow stromal cell

factor 5 4124 antigen 1 2.03562

391

TREML4 triggering receptor 1.96347 GPR83 G protein-coupled receptor

expressed on myeloid 4124 83 2.03562 cells like 4 391

SORD sorbitol dehydrogenase 1.96347 RECK reversion inducing

4124 cysteine rich protein with 2.03611 kazal motifs 2118

HEBP1 heme binding protein 1 1.96347 ABHD14B ab hydrolase domain

4124 containing 14B 2.04046

0993

KDELR2 KDEL endoplasmic 1.96155 GPRC6A G protein-coupled receptor

reticulum protein 465 class C group 6 member A 2.04212 retention receptor 2 2888

TRPV4 transient receptor 1.95884 GRAMD3 GRAM domain containing

potential cation channel 2675 3 2.04229 subfamily V member 4 6131

ABHD5 ab hydrolase domain 1.95738 IMPACT impact RWD domain

containing 5 9419 protein 2.04243

6285

YOD1 YOD 1 deubiquitinase 1.95419 TOPI topoisomerase (DNA) I

631 2.04439

4119

MAGOHB mago homolog B, exon 1.95293 NACC2 NACC family member 2

junction complex core 2368 2.04439 component 4119

TSPAN2 tetraspanin 2 1.95176 PKNOX1 PBX/knotted 1 homeobox

103 1 2.04579

7958

LDB3 LIM domain binding 3 1.94850 TMEM79 transmembrane protein 79 - 842 2.04662

8729

1700067P10 Description Not Found 1.94485 MYCBP2 MYC binding protein 2,

RIK 8446 E3 ubiquitin protein ligase 2.04736

8853

9530091C0 Description Not Found 1.94485 MASl MASl proto-oncogene, G

8RIK 8446 protein-coupled receptor 2.04805

5651

RHOJ ras homolog family 1.94485 GEMIN6 gem nuclear organelle

member J 8446 associated protein 6 2.05311

1336

SFRP1 secreted frizzled related 1.94485 TMEM100 transmembrane protein

protein 1 8446 100 2.05311

1336

XPNPEP2 X-prolyl aminopeptidase 1.94485 FOXI1 forkhead box ll

(aminopeptidase P) 2, 8446 2.05311 membrane-bound 1336

RNASE4 ribonuclease A family 1.93545 OPLAH 5-oxoprolinase (ATP- member 4 9748 hydrolysing) 2.05311

1336

NAPSA napsin A aspartic 1.93158 BC094916 Description Not Found

peptidase 6931 2.05833

7935

TIMM22 translocase of inner 1.93120 GZMM granzyme M

mitochondrial membrane 2999 2.06119 22 homolog (yeast) 3332

MTCH2 mitochondrial carrier 2 1.92977 RCOR2 REST corepressor 2

4464 2.06280

495

ADCK4 aarF domain containing 1.92792 NR2E1 nuclear receptor subfamily

kinase 4 1426 2 group E member 1 2.06366

268

PDSS1 prenyl (decaprenyl) 1.92624 NT5DC1 5'-nucleotidase domain

diphosphate synthase, 5513 containing 1 2.06599 subunit 1 4119

ZFP94 zinc finger protein 1.92599 SCN8A sodium voltage-gated

94(Zfp94) 9419 channel alpha subunit 8 2.06750

099

FABP9 fatty acid binding protein 1.92599 CBX7 chromobox 7

9 9419 2.06750

099

RNF170 ring finger protein 170 1.92599 FHAD1 forkhead associated

9419 phosphopeptide binding 2.06811 domain 1 4527

TLR3 toll like receptor 3 1.92599 KCNQ3 potassium voltage-gated

9419 channel subfamily Q 2.06888 member 3 5643

LIPH lipase H 1.92599 BC025920 zinc finger protein

9419 pseudogene(BC025920) 2.07038

9328

PLEKHA7 pleckstrin homology 1.92599 FCGR1 Fc receptor, IgG, high

domain containing A7 9419 affinity I(Fcgrl) 2.07038

9328

LXN latexin 1.92446 SYN3 synapsin III

06 2.07038

9328

PPCS phosphopantothenoylcys 1.92294 KLHL5 kelch like family member

teine synthetase 738 5 2.07038

9328 BTRC beta-transducin repeat 1.92065 EDA2R ectodysplasin A2 receptor containing E3 ubiquitin 845 2.07038 protein ligase 9328

APIP APAF1 interacting 1.92032 STK38 serine/threonine kinase 38

protein 6443 2.07038

9328

ANK1 ankyrin 1 1.91647 CDKN2D cyclin dependent kinase

6644 inhibitor 2D 2.07220

5467

TOMM70A translocase of outer 1.91310 IL6ST interleukin 6 signal

mitochondrial membrane 7017 transducer 2.07266 70 homolog A 0321 (yeast)(Tomm70a)

ABCB1B ATP -binding cassette, 1.90803 OLFR427 olfactory receptor

sub -family B 3945 427(01fr427) 2.07431 (MDR/TAP), member 8985 lB(Abcblb)

ACN9 Description Not Found 1.90689 BAIAP2 BAIl associated protein 2

0596 2.07895

1341

DLX1AS distal-less homeobox 1, 1.90689 TIMP2 TIMP metallopeptidase

antisense(Dlxlas) 0596 inhibitor 2 2.07980

5224

MRGPRD MAS related GPR 1.90689 CDCP1 CUB domain containing

family member D 0596 protein 1 2.08399

1945

WDHD1 WD repeat and HMG- 1.90689 RGS14 regulator of G-protein

box DNA binding 0596 signaling 14 2.08419 protein 1 8537

USP46 ubiquitin specific 1.90689 VASP vasodilator-stimulated

peptidase 46 0596 phosphoprotein 2.08635

9868

PKN3 protein kinase N3 1.90689 ZFP318 zinc finger protein

0596 318(Zfp318) 2.08746

2841

OSCAR osteoclast associated, 1.90689 PSG25 pregnancy-specific

immunoglobulin-like 0596 glycoprotein 25(Psg25) 2.08746 receptor 2841

CDK2 cyclin dependent kinase 1.90674 PDZD8 PDZ domain containing 8

2 6727 2.08746

2841

TRIM62 tripartite motif 1.90552 DET1 de-etiolated homolog 1

containing 62 0967 (Arabidopsis) 2.08746

2841

SQLE squalene epoxidase 1.90376 CHST3 carbohydrate

7694 sulfotransferase 3 2.08746

2841

MCM10 minichromosome 1.89598 EHHADH enoyl-CoA, hydratase/3- maintenance 10 378 hydroxyacyl CoA 2.08746 replication initiation dehydrogenase 2841 factor

CCDC90B coiled-coil domain 1.89480 FCGRT Fc fragment of IgG

containing 90B 3124 receptor and transporter 2.09073

5607

SPATS 1 spermatogenesis 1.89284 CFP complement factor

associated serine rich 1 8083 properdin 2.09437

407

GPNMB glycoprotein nmb 1.89142 SOCS6 suppressor of cytokine

7809 signaling 6 2.09463 MST1 macrophage stimulating 1.88993 SYT11 synaptotagmin 11 1 148 2.09592

442

LTB4R1 leukotriene B4 receptor 1.88764 MBTPS2 membrane bound

l(Ltb4rl) 4112 transcription factor 2.09592 peptidase, site 2 442

DNAJC5B DnaJ heat shock protein 1.88752 MEFV Mediterranean fever

family (Hsp40) member 5271 2.09705 C5 beta 9135

PCDHGC4 protocadherin gamma 1.88752 SRPK2 SRSF protein kinase 2

subfamily C, 4 5271 2.10044

313

HMX2 H6 family homeobox 2 1.88752 DUSP16 dual specificity

5271 phosphatase 16 2.10274

0277

NDUFAB 1 NADH:ubiquinone 1.88752 SLC6A7 solute carrier family 6

oxidoreductase subunit 5271 member 7 2.10312 AB 1 9681

MGP matrix Gla protein 1.88752 HBB-B1 hemoglobin, beta adult

5271 major chain(Hbb-bl) 2.10433

666

ZKSCAN2 zinc finger with KRAB 1.88752 TNP03 transportin 3

and SCAN domains 2 5271 2.10433

666

CCDC51 coiled-coil domain 1.88752 CSNK2B casein kinase 2 beta

containing 51 5271 2.10433

666

CTSK cathepsin K 1.88752 BCAS1 breast carcinoma amplified

5271 sequence 1 2.10433

666

PRDM9 PR domain 9 1.88752 INO80 INO80 complex subunit

5271 2.10433

666

C8A complement component 1.88752 MPG N-methylpurine DNA

8 alpha subunit 5271 glycosylase 2.10433

666

NEUROG1 neurogenin 1 1.88708 FOXP1 forkhead box PI

2413 2.10755

7734

NUSAP1 nucleolar and spindle 1.88695 USP21 ubiquitin specific

associated protein 1 1242 peptidase 21 2.10765

8353

LZIC leucine zipper and 1.87789 LIMS1 LIM zinc finger domain

CTNNBIP1 domain 9051 containing 1 2.11270 containing 0133

ZFP609 zinc finger protein 1.87774 FXYD1 FXYD domain containing

609(Zfp609) 425 ion transport regulator 1 2.11270

0133

GPR87 G protein-coupled 1.87774 POU3F1 POU class 3 homeobox 1

receptor 87 425 2.11357

4207

GMPPB GDP-mannose 1.87152 OLFR591 olfactory receptor

pyrophosphorylase B 3637 591(01fr591) 2.11449

4844

TMEM115 transmembrane protein 1.87036 GRAMD4 GRAM domain containing

115 4796 4 2.11467

3101

DSN1 DSN1 homolog, MIS12 1.86847 BCL2 BCL2, apoptosis regulator - kinetochore complex 9018 2.11587 component 8669

A530099J1 Description Not Found 1.86789 PELI3 pellino E3 ubiquitin

9RIK 6464 protein ligase family 2.11891 member 3 5146

1700007K0 Description Not Found 1.86789 PPP1CB protein phosphatase 1

9RIK 6464 catalytic subunit beta 2.11923

6221

1810043G0 Description Not Found 1.86789 TFF2 trefoil factor 2

2RIK 6464 2.12101

5401

UCHL1 ubiquitin C -terminal 1.86789 GCA grancalcin

hydrolase LI 6464 2.12101

5401

PTCH2 patched 2 1.86789 LYLl LYLl, basic helix-loop- 6464 helix family member 2.12101

5401

APBB3 amyloid beta precursor 1.86789 ATG4B autophagy related 4B

protein binding family B 6464 cysteine peptidase 2.12101 member 3 5401

PTER phosphodiesterase 1.86789 CCDC102A coiled-coil domain

related 6464 containing 102 A 2.12101

5401

PRKCE protein kinase C epsilon 1.86789 ATP2A1 ATPase

6464 sarcoplasmic/endoplasmic 2.12101 reticulum Ca2+ 5401 transporting 1

PLEKHM3 pleckstrin homology 1.86789 TERF2 telomeric repeat binding

domain containing M3 6464 factor 2 2.12358

5568

HIST1H4C histone cluster 1, H4c 1.86789 LCN5 lipocalin 5(Lcn5)

6464 2.12443

2612

PLS3 plastin 3 1.86789 TM6SF1 transmembrane 6

6464 superfamily member 1 2.12453

3495

DUSP4 dual specificity 1.86768 SSBP2 single stranded DNA

phosphatase 4 6654 binding protein 2 2.12928

3017

SCLY selenocysteine lyase 1.86280 KRTAP6-2 keratin associated protein

2277 6-2 2.13750

3524

RPRD1A regulation of nuclear 1.86177 CRHBP corticotropin releasing

pre-mRNA domain 7838 hormone binding protein 2.13750 containing 1A 3524

CCRL2 C-C motif chemokine 1.86175 TOPBP1 topoisomerase (DNA) II

receptor like 2 579 binding protein 1 2.13750

3524

CCT7 chaperonin containing 1.86163 SLC35A3 solute carrier family 35

TCP1 subumt 7 6037 member A3 2.13750

3524

ZFP217 zinc finger protein 1.86109 CACNB4 calcium voltage-gated

217(Zfp217) 7096 channel auxiliary subunit 2.13750 beta 4 3524

ACTN4 actinin alpha 4 1.85968 TASP1 taspase 1

9938 2.13750

3524

KCNA3 potassium voltage-gated 1.85913 HMBOX1 homeobox containing 1

channel subfamily A 5363 2.14531 member 3 3833

CUL7 cullin 7 1.85859 ZFP62 ZFP62 zinc finger protein

7911 2.14567

7455

LRRC59 leucine rich repeat 1.85754 PCDHB4 protocadherin beta 4

containing 59 3219 2.14866

6128

PHTF2 putative homeodomain 1.85560 SLC35F3 solute carrier family 35

transcription factor 2 2651 member F3 2.15120

644

KDELC1 KDEL motif containing 1.85255 AW549877 expressed sequence

1 6218 AW549877(AW549877) 2.15132

4826

SEC24D SEC24 homolog D, 1.84838 GIMAP9 GTPase, IMAP family

COPII coat complex 41 member 9(Gimap9) 2.15240 component 0921

OLFR222 olfactory receptor 1.84799 ZFP329 zinc finger protein

222(01fr222) 6907 329(Zfp329) 2.15380

5336

OLFR118 olfactory receptor 1.84799 KRT74 keratin 74

118(01frl l8) 6907 2.15380

5336

CASKIN2 CASK interacting 1.84799 REG3A regenerating family

protein 2 6907 member 3 alpha 2.15380

5336

TPK1 thiamin 1.84799 RAB4A RAB4A, member RAS

pyrophosphokinase 1 6907 oncogene family 2.15430

8231

NOL3 nucleolar protein 3 1.84799 CECR5 cat eye syndrome

6907 chromosome region, 2.15568 candidate 5 2653

UBA6 ubiquitin like modifier 1.84738 ESM1 endothelial cell specific

activating enzyme 6 8943 molecule 1 2.15715

6463

RAVER 1 ribonucleoprotein, PTES 1.84615 HS6ST1 heparan sulfate 6-0- binding 1 1947 sulfotransferase 1 2.16482

0712

NAT 10 N-acetyltransferase 10 1.84330 DDB2 damage specific DNA

0131 binding protein 2 2.16833

8824

HIST1H3H histone cluster 1, H3h 1.84205 5430435G2 Description Not Found

5889 2RIK 2.16992

5001

SNX8 sorting nexin 8 1.84098 ALOX12B arachidonate 12- 5134 lipoxygenase, 12R type 2.16992

5001

POLR3K polymerase (RNA) III 1.83953 SLC34A3 solute carrier family 34

subunit K 8616 member 3 2.16992

5001

WDR55 WD repeat domain 55 1.83595 TNS4 tensin 4

7408 2.16992

5001

WDR93 WD repeat domain 93 1.83054 CANX calnexin

1464 2.16992

5001

PLSCR1 phospholipid scramblase 1.82863 BET1 Betl golgi vesicular

1 5636 membrane trafficking 2.16992 protein 5001

ARL6 ADP ribosylation factor 1.82781 BEST2 bestrophin 2 - like GTPase 6 9025 2.16992

5001

NOL9 nucleolar protein 9 1.82781 USP28 ubiquitin specific

9025 peptidase 28 2.17299

8154

PNKD paroxysmal 1.82781 PDE4B phosphodiesterase 4B

nonkinesigenic 9025 2.17361 dyskinesia 4018

TMEM139 transmembrane protein 1.82781 CNOT4 CCR4-NOT transcription

139 9025 complex subumt 4 2.17791

7792

ASPH aspartate beta- 1.82781 NEC API NECAP endocytosis

hydroxylase 9025 associated 1 2.17804

3245

LZTFL1 leucine zipper 1.82781 JUN Jun proto-oncogene, AP-1

transcription factor like 1 9025 transcription factor subunit 2.17856

5309

RHEBL1 Ras homolog enriched in 1.82781 SLC10A7 solute carrier family 10

brain like 1 9025 member 7 2.17990

909

CHCHD5 coiled-coil-helix-coiled- 1.82552 IL17A interleukin 17A

coil-helix domain 849 2.18170 containing 5 2586

GPD2 gly cerol-3 -phosphate 1.82414 ERICH 1 glutamate rich 1

dehydrogenase 2 8697 2.18228

6216

STK39 serine/threonine kinase 1.82360 HN1L hematological and

39 8879 neurological expressed 1- 2.18586 like 6545

MAGED2 MAGE family member 1.82086 SLFNLl schlafen like 1

D2 3253 2.18586

6545

TBC1D9B TBCl domain family 1.81321 MYOD1 myogenic differentiation 1

member 9B 9568 2.18586

6545

LSS lanosterol synthase (2,3- 1.80954 TRIM35 tripartite motif containing

oxidosqualene-lanosterol 0228 35 2.18586 cyclase) 6545

OLFR859 olfactory receptor 1.80735 CHRNE cholinergic receptor

859(01fr859) 4922 nicotinic epsilon subunit 2.18639

7884

OLFR1225 olfactory receptor 1.80735 PHF21A PHD finger protein 21 A

1225(01frl225) 4922 2.19094

3197

IFNA11 interferon alpha 1.80735 HIST1H2A histone cluster 1, H2ae

l l(Ifnal l) 4922 E 2.19669

8179

ARG1 arginase 1 1.80735 SATB 1 SATB homeobox 1

4922 2.19865

9952

ASCL3 achaete-scute family 1.80735 LCN8 lipocalin 8

bHLH transcription 4922 2.20163 factor 3 3861

AGA aspartylglucosaminidase 1.80735 ABCG5 ATP binding cassette

4922 subfamily G member 5 2.20163

3861

MAP3K12 mitogen-activated 1.80653 KRBA1 KRAB-A domain

protein kinase kinase 0545 containing 1 2.20295 kinase 12 9029 1749

EN2 engrailed homeobox 2 1.76553 CXCL2 C-X-C motif chemokine

4746 ligand 2 2.29278

1749

HOOK3 hook micro tubule- 1.76553 SNX7 sorting nexin 7

tethering protein 3 4746 2.29278

1749

MY09A myosin IXA 1.76553 ZDHHC23 zinc finger DHHC-type

4746 containing 23 2.29278

1749

STX7 syntaxin 7 1.76506 GALNT6 polypeptide N- 0364 acetylgalactosaminyltransf 2.29278 erase 6 1749

ATM ATM serine/threonine 1.76350 AMPD 1 adenosine monophosphate

kinase 4031 deaminase 1 2.29784

4157

KCNK6 potassium two pore 1.76338 GIMAP5 GTPase, IMAP family

domain channel 5753 member 5 2.30324 subfamily K member 6 6615

PQLC3 PQ loop repeat 1.75995 ATP5F1 ATP synthase, H+

containing 3 4577 transporting, 2.30539 mitochondrial Fo complex 9163 subunit Bl

KIFAP3 kinesin associated 1.75884 LHFPL2 lipoma HMGIC fusion

protein 3 3168 partner-like 2 2.30742

8525

E2F4 E2F transcription factor 1.75775 KIF1B kinesin family member IB

4 2886 2.31323

1129

ETV5 ETS variant 5 1.75770 TLE6 transducin like enhancer of

9335 split 6 2.32192

8095

GTF2E2 general transcription 1.75666 SHF Src homology 2 domain

factor HE subunit 2 387 containing F 2.33069

1998

GPR150 G protein-coupled 1.75547 NGFR nerve growth factor

receptor 150 927 receptor 2.33143

8521

E130308A1 RIKEN cDNA 1.75488 KLRA4 killer cell lectin-like

9RIK E130308A19 7502 receptor, subfamily A, 2.33448 gene(E130308A19Rik) member 4(Klra4) 5632

DPYSL4 dihydropyrimidinase like 1.75488 ITGAE integrin subunit alpha E

4 7502 2.33594

8972

FNBP1 formin binding protein 1 1.75468 PQLC2 PQ loop repeat containing

902 2 2.33614

1568

TMOD4 tropomodulin 4 1.75406 KLRB1A killer cell lectin-like

4107 receptor subfamily B 2.33628 member lA(Klrbla) 3388

ERLIN1 ER lipid raft associated 1 1.75115 IRF9 interferon regulatory factor

4691 9 2.33630

8285

ENOPH1 enolase-phosphatase 1 1.74844 GATA3 GATA binding protein 3

7442 2.33897

1433

RAB31 RAB31, member RAS 1.74621 RSAD2 radical S-adenosyl

oncogene family 5332 methionine domain 2.33997 containing 2 952 HOXA6 homeobox A6 1.74518 RNF215 ring finger protein 215

4623 2.34197

6415

TAS2R126 taste receptor, type 2, 1.74416 IL7R interleukin 7 receptor

member 126(Tas2rl26) 1096 2.34339

5577

AGXT2 alanine-glyoxylate 1.74416 ACP5 acid phosphatase 5, tartrate

aminotransferase 2 1096 resistant 2.34527

0806

ST 32C serine/threonine kinase 1.74416 STYXL1 serine/threonine/tyrosine

32C 1096 interacting-like 1 2.34695

6889

P2RY2 purinergic receptor 1.74416 NOXOl NADPH oxidase organizer

P2Y2 1096 1 2.35030

956

NWD1 NACHT and WD repeat 1.74416 IGFALS insulin like growth factor

domain containing 1 1096 binding protein acid labile 2.35866 subunit 4554

UQCRQ ubiquinol-cytochrome c 1.74416 STIM1 stromal interaction

reductase complex III 1096 molecule 1 2.35933 subunit VII 5599

PPP1R3A protein phosphatase 1 1.74416 TMEM186 transmembrane protein

regulatory subunit 3 A 1096 186 2.36103

0771

GOLT1A golgi transport 1A 1.74416 OLFR1043 olfactory receptor

1096 1043(Olfrl043) 2.36457

2432

EZH1 enhancer of zeste 1 1.74416 D8ERTD82 DNA segment, Chr 8,

polycomb repressive 1096 E ERATO Doi 82, 2.36457 complex 2 subunit expressed(D8Ertd82e) 2432

MTHFD2 methylenetetrahydrofolat 1.74415 MYOG myogenin

e dehydrogenase 4314 2.36457 (NADP+ dependent) 2, 2432 mefhenyltetrahydrofolate

cyclohydrolase

PGRMC1 progesterone receptor 1.74254 NCLN nicalin

membrane component 1 5062 2.36457

2432

DNAJB12 DnaJ heat shock protein 1.74186 MTSS1 metastasis suppressor 1

family (Hsp40) member 3621 2.36457 B12 2432

DNAJC11 DnaJ heat shock protein 1.73876 TRMU TRNA 5- family (Hsp40) member 7837 methylaminomethyl-2- 2.36457 Cl l thiouridylate 2432 methyltransferase

TOMM6 translocase of outer 1.73844 EMILIN2 elastin microfibril

mitochondrial membrane 8709 interfacer 2 2.36911

6 9767

RPS6KL1 ribosomal protein S6 1.73839 MPV17L MPV17 mitochondrial

kinase like 1 3453 inner membrane protein 2.37155 like 8863

CDC73 cell division cycle 73 1.73665 WWC2 WW and C2 domain

741 containing 2 2.37155

8863

NDC80 NDC80, kinetochore 1.73207 TMEM178 transmembrane protein

complex component 8892 178(Tmeml78) 2.37400

5585

TACC3 transforming acidic 1.73137 TPCN1 two pore segment channel

coiled-coil containing 2884 1 2.37523 protein 3 2208

CPSF3 cleavage and 1.72792 LRRC45 leucine rich repeat

polyadenylation specific 6568 containing 45 2.37720 factor 3 7351

ARID 3 A AT-rich interaction 1.72647 1110059G1 Description Not Found

domain 3 A 1722 0RIK 2.37791

5929

LLPH LLP homolog, long-term 1.72610 MCOLN2 mucolipin 2

synaptic facilitation 7859 2.37851

1623

PCNA proliferating cell nuclear 1.72544 DDX58 DEXD/H-box helicase 58

antigen 1599 2.37851

1623

GJC2 gap junction protein 1.72297 H2-OA histocompatibility 2, O

gamma 2 8517 region alpha locus(H2-Oa) 2.38232

9516

OLFR373 olfactory receptor 1.72246 RARG retinoic acid receptor

373(01fr373) 6024 gamma 2.38882

7772

H2-T24 histocompatibility 2, T 1.72246 SERPINB 1 serine (or cysteine)

region locus 24(H2-T24) 6024 A peptidase inhibitor, clade 2.39231

B, member la(Serpinbla) 7423

AKAP7 A-kinase anchoring 1.72246 GHRL ghrelin/obestatin

protein 7 6024 prepropeptide 2.39231

7423

NDUFB7 NADH:ubiquinone 1.72246 ZMAT4 zinc finger matrin-type 4

oxidoreductase subunit 6024 2.39231 B7 7423

PRR11 proline rich 11 1.72246 BTBD6 BTB domain containing 6

6024 2.39289

7478

TJP1 tight junction protein 1 1.72246 KLRA16 killer cell lectin-like

6024 receptor, subfamily A, 2.39453 member 16(Klral6) 4969

SI 00 A3 S100 calcium binding 1.72246 EPS15L1 epidermal growth factor

protein A3 6024 receptor pathway substrate 2.39701

15 like 1 2831

KRT78 keratin 78 1.71872 VCPIP1 valosin containing protein

9711 interacting protein 1 2.39730

3585

GMDS GDP-mannose 4,6- 1.71790 RRP7A nbosomal RNA processing

dehydratase 4741 7 homolog A 2.40499

2223

PDGFB platelet derived growth 1.71440 IL 1B interleukin 1 beta

factor subunit B 0534 2.40599

236

SLC36A1 solute carrier family 36 1.71429 NAT 14 N-acetyltransf erase 14

member 1 7338 (putative) 2.40599

236

RSU1 Ras suppressor protein 1 1.71264 SLC40A1 solute carrier family 40

7036 member 1 2.40599

236

STX12 syntaxin 12 1.71191 RAB37 RAB37, member RAS

1478 oncogene family 2.40599

236

SLC25A34 solute carrier family 25 1.71149 IL 17RA interleukin 17 receptor A

member 34 4907 2.40599

236

AFG3L2 AFG3 like matrix AAA 1.71105 BACE1 beta-secretase 1 - peptidase subunit 2 7 2.40599

236

RPL24 ribosomal protein L24 1.70919 CTNS cystinosin, lysosomal

3708 cystine transporter 2.40599

236

UBE3C ubiquitin protein ligase 1.70878 IFIT3 interferon induced protein

E3C 9682 with tetratricopeptide 2.41140 repeats 3 4504

CAR12 carbonic anhydrase 1.70867 ZFYVE21 zinc finger FYVE-type

12(Carl2) 626 containing 21 2.41237

8292

ZFP207 zinc finger protein 1.70760 1700016D0 Description Not Found

207(Zfp207) 3009 6RIK 2.41953

8892

XIST X inactive specific 1.70606 STK25 serine/threonine kinase 25

transcript (non-protein 5607 2.41953 coding) 8892

NCAPD2 non-SMC condensin I 1.70501 PLEKHJ1 pleckstrin homology

complex subunit D2 2178 domain containing Jl 2.41953

8892

ZSWIM2 zinc finger SWIM-type 1.70480 TGIF2 TGFB induced factor

containing 2 2998 homeobox 2 2.41953

8892

CASP1 caspase 1 1.70065 SLC25A29 solute carrier family 25

942 member 29 2.41953

8892

OLFR701 olfactory receptor 1.70043 DAPL1 death associated protein

701(Olfr701) 9718 like 1 2.41966

1316

CBLC Cbl proto-oncogene C 1.70043 P2RX4 purinergic receptor P2X 4

9718 2.42574

8008

HIST1H2A histone cluster 1, H2ac 1.70043 170000102 Description Not Found

C 9718 2RIK 2.42626

4755

EPHA10 EPH receptor A10 1.70043 C9 complement component 9

9718 2.42961

5964

NDUFC2 NADH:ubiquinone 1.70043 KLF13 Kruppel like factor 13

oxidoreductase subunit 9718 2.43062 C2 8023

DLG1 discs large MAGUK 1.70043 GADD45A growth arrest and DNA

scaffold protein 1 9718 damage inducible alpha 2.43259

1239

SCN10A sodium voltage-gated 1.70043 OLFR788 olfactory receptor

channel alpha subunit 10 9718 788(01fr788) 2.43295

9407

RGL3 ral guanine nucleotide 1.70043 FADS6 fatty acid desaturase 6

dissociation stimulator 9718 2.43295 like 3 9407

TMC03 transmembrane and 1.70043 CHCHD2 coiled-coil-helix-coiled- coiled-coil domains 3 9718 coil-helix domain 2.43295 containing 2 9407

BCL2L14 BCL2 like 14 1.70043 MPPE1 metallophosphoesterase 1

9718 2.43295

9407

TH0P1 thimet oligopeptidase 1 1.70029 CHAC1 ChaC glutathione specific

0033 gamma- 2.43295 glutamylcyclotransferase 1 9407 MTIF3 mitochondrial 1.69830 2310011J03 Description Not Found translational initiation 5331 RIK 2.43501 factor 3 7448

XDH xanthine dehydrogenase 1.69771 LRSAM1 leucine rich repeat and

7724 sterile alpha motif 2.43747 containing 1 3925

ANXA9 annexin A9 1.69718 SIRPA signal regulatory protein

4071 alpha 2.44312

5132

OLFR1502 olfactory receptor 1.69404 CYP24A1 cytochrome P450 family

1502(Olfrl502) 6727 24 subfamily A member 1 2.44625

623

HCFC2 host cell factor C2 1.69378 NQOl NAD(P)H quinone

0609 dehydrogenase 1 2.44625

623

DIDOl death inducer-obliterator 1.69359 HRH4 histamine receptor H4

1 6948 2.44625

623

PGAM1 phosphoglycerate mutase 1.68984 NUDCD 1 NudC domain containing 1

1 6917 2.44625

623

RASGEF1C RasGEF domain family 1.68929 CCNDl cyclin D 1

member 1C 9161 2.44792

4527

SLC25A42 solute carrier family 25 1.68677 ADAM22 ADAM metallopeptidase

member 42 4817 domain 22 2.45285

8965

CPT2 carnitine 1.68636 MDK midkine (neurite growth- palmitoyltransferase 2 4794 promoting factor 2) 2.45614

9035

MAD2L1 MAD2 mitotic arrest 1.68616 STX1A syntaxin 1A

deficient-like 1 (yeast) 1103 2.45672

9828

NQ02 NAD(P)H quinone 1.68555 HEMK1 HemK methyltransferase

dehydrogenase 2 8757 family member 1 2.45943

1619

HIP1R huntingtin interacting 1.68547 B4GALT7 beta- 1,4- protein 1 related 3307 galactosyltransferase 7 2.45943

1619

ALOX12E arachidonate 1.68437 ASXL2 additional sex combs like

lipoxygenase, 3244 2, transcriptional regulator 2.45943 epidermal(Aloxl2e) 1619

LMAN1 lectin, mannose binding 1.68351 TLR7 toll like receptor 7

1 4205 2.46052

038

ASB3 ankyrin repeat and 1.68014 TDP1 tyrosyl-DNA

SOCS box containing 3 2991 phosphodiesterase 1 2.46446

1869

XKR5 XK related 5 1.67925 1700025G0 Description Not Found

4438 4RIK 2.46930

3076

ZFP235 zinc finger protein 1.67807 SLC16A6 solute carrier family 16

235(Zfp235) 1905 member 6 2.47104

5434

OLFR971 olfactory receptor 1.67807 DOXL2 diamine oxidase-like

971(01fr971) 1905 protein 2(Doxl2) 2.47248

7771

OLFR374 olfactory receptor 1.67807 PKD 1L3 polycystin 1 like 3,

374(01fr374) 1905 transient receptor potential 2.47248 channel interacting 7771

NOS1AP nitric oxide synthase 1 1.67807 ZC3H11A zinc finger CCCH-type

adaptor protein 1905 containing 11 A 2.47248

7771

GALM galactose mutarotase 1.67807 LY6K lymphocyte antigen 6

1905 complex, locus K 2.47248

7771

MEGF9 multiple EGF like 1.67807 KLF7 Kruppel like factor 7

domains 9 1905 2.47475

5307

CCDC66 coiled-coil domain 1.67807 BTLA B and T lymphocyte

containing 66 1905 associated 2.47560

4026

LRRC40 leucine rich repeat 1.67807 CDON cell adhesion associated,

containing 40 1905 oncogene regulated 2.48542

6827

RALA RALA Ras like proto- 1.67807 DDC dopa decarboxylase

oncogene A 1905 2.48542

6827

YIPF4 Yipl domain family 1.67807 GTF2A2 general transcription factor

member 4 1905 IIA subunit 2 2.48542

6827

TAL2 T-cell acute lymphocytic 1.67807 DTX4 deltex E3 ubiquitin ligase

leukemia 2 1905 4 2.48542

6827

LRRC8A leucine rich repeat 1.67807 GSTKl glutathione S -transferase

containing 8 family 1905 kappa 1 2.48619 member A 5934

APOM apolipoprotein M 1.67807 OLFR213 olfactory receptor

1905 213(01fr213) 2.48912

5048

KCNG3 potassium voltage-gated 1.67807 PDE5A phosphodiesterase 5A

channel modifier 1905 2.49057 subfamily G member 3 1469

CN 1 calponin 1 1.67807 TOB1 transducer of ERBB2, 1

1905 2.49676

3907

STAC2 SH3 and cysteine rich 1.67807 1700109H0 Description Not Found

domain 2 1905 8RIK 2.49825

0868

SFRP2 secreted frizzled related 1.67807 LEFTY 1 left-right determination

protein 2 1905 factor 1 2.49825

0868

SERPINB9 serine (or cysteine) 1.67016 SNAPC4 small nuclear RNA

E peptidase inhibitor, clade 9131 activating complex 2.50087

B, member polypeptide 4 8922 9e(Serpinb9e)

TFB 1M transcription factor B 1 , 1.66894 RNF41 ring finger protein 41

mitochondrial 6692 2.50355

1585

SLC25A10 solute carrier family 25 1.66885 KLHL34 kelch like family member

member 10 6925 34 2.50462

0392

BID BH3 interacting domain 1.66799 SSH2 slingshot protein

death agonist 2567 phosphatase 2 2.50549

2762

MRPS27 mitochondrial ribosomal 1.66729 CAMK2B calcium/ calmodulin

protein S27 5766 dependent protein kinase 2.50704

II beta 7355 inhibitor 4954

SLC38A5 solute carrier family 38 1.65535 ZFP667 zinc finger protein

member 5 1829 667(Zfp667) 2.56667

0372

VAT1L vesicle amine transport 1.65535 SH3BP1 SH3 domain binding

Hike 1829 protein 1 2.56673

4604

HOXB7 homeobox B7 1.65535 FFAR2 free fatty acid receptor 2

1829 2.57288

9668

GAN gigaxonin 1.65535 EEF2K eukaryotic elongation

1829 factor 2 kinase 2.57288

9668

MMP28 matrix metallopeptidase 1.65535 SLPI secretory leukocyte

28 1829 peptidase inhibitor 2.57472

1828

METTLIO methyltransf erase like 10 1.65535 CMA1 chymase 1

1829 2.58496

2501

SIX4 SIX homeobox 4 1.65535 ASCL1 achaete-scute family

1829 bHLH transcription factor 2.58496

1 2501

TDRD6 tudor domain containing 1.65535 ACPP acid phosphatase, prostate

6 1829 2.58496

2501

COMMD5 COMM domain 1.65460 CLCNKB chloride voltage-gated

containing 5 4999 channel Kb 2.59693

5142

PRDX4 peroxiredoxin 4 1.65192 FBXW7 F-box and WD repeat

3925 domain containing 7 2.59693

5142

HS3ST3A1 heparan sulfate- 1.64929 OLIG3 oligodendrocyte

glucosamine 3- 8274 transcription factor 3 2.59693 sulfotransf erase 3A1 5142

CALCA calcitonin related 1.64906 WHRN whirlin

polypeptide alpha 7786 2.60678

9951

SLC12A2 solute carrier family 12 1.64844 DNAJC14 DnaJ heat shock protein

member 2 9243 family (Hsp40) member 2.60880

C14 9243

TJP2 tight junction protein 2 1.64414 PIGT phosphatidylinositol

5647 glycan anchor biosynthesis 2.61103 class T 1218

LRRC16B Description Not Found 1.64385 AP1G2 adaptor related protein

619 complex 1 gamma 2 2.61470 subunit 9844

AP3S2 adaptor related protein 1.64385 SAA2 serum amyloid A2

complex 3 sigma 2 619 2.62058 subunit 641

PSMD9 proteasome 26 S subunit, 1.64385 USP30 ubiquitin specific

non-ATPase 9 619 peptidase 30 2.62058

641

PARD6G par-6 family cell polarity 1.64337 RPE65 retinal pigment epithelium

regulator gamma 9419 specific protein 65 2.63226

8216

CIAPIN1 cytokine induced 1.64321 CML1 Description Not Found

apoptosis inhibitor 1 9709 2.63489

1632

CKAP5 cytoskeleton associated 1.64274 SLC6A19 solute carrier family 6 - 1905

ADCY10 adenylate cyclase 10 1.63226 ZCCHC12 zinc finger CCHC-type

(soluble) 8216 containing 12 2.67833

354

PEX12 peroxisomal biogenesis 1.63226 ADCY6 adenylate cyclase 6

factor 12 8216 2.68088

6921

IQCC IQ motif containing C 1.63226 ECM1 extracellular matrix

8216 protein 1 2.68345

512

ENPP1 ectonucleotide 1.63208 AFP alpha fetoprotein

py ropho sphatase/pho sph 6279 2.68929 odiesterase 1 9161

ADAL adenosine deaminase- 1.63066 GP5 glycoprotein V platelet

like 4126 2.68929

9161

SCRN2 secernin 2 1.63056 GAB 3 GRB2 associated binding

6247 protein 3 2.69140

5681

CEP78 centrosomal protein 78 1.62985 USP2 ubiquitin specific

1642 peptidase 2 2.69333

4369

SLC25A15 solute carrier family 25 1.62979 PLXNB1 plexin B 1

member 15 8606 2.70043

9718

ADSSL1 adenylosuccinate 1.62827 PODXL2 podocalyxin like 2

synthase like 1 2149 2.70079

9925

TM6SF2 transmembrane 6 1.62775 RAD9B RAD 9 checkpoint clamp

superfamily member 2 8638 component B 2.70103

836

TUBGl tubulin gamma 1 1.62451 AKAP10 A-kinase anchoring

1879 protein 10 2.70597

7902

FASTK Fas activated 1.62333 PIGW phosphatidylinositol

serine/threonine kinase 6662 glycan anchor biosynthesis 2.71699 class W 0894

RBBP5 RB binding protein 5, 1.62216 COL12A1 collagen type XII alpha 1

histone lysine 3711 chain 2.72246 methyltransferase 6024 complex subunit

1700071K0 Description Not Found 1.62146 GPR137B G protein-coupled receptor 1RIK 5074 137B 2.73335

4341

SLC25A33 solute carrier family 25 1.62128 IMMP2L inner mitochondrial

member 33 2718 membrane peptidase 2.73335 subunit 2 4341

MDM4 MDM4, p53 regulator 1.62058 PIK3CB phosphatidylinositol-4,5- 641 bisphosphate 3 -kinase 2.73732 catalytic subunit beta 0423

T0P2A topoisomerase (DNA) II 1.62037 TGFBI transforming growth factor

alpha 4948 beta induced 2.74027

6443

OLFR139 olfactory receptor 1.61973 ZFP106 zinc finger protein

139(01frl39) 1323 106(Zfpl06) 2.74416

1096

PAPLN papilin, proteoglycan 1.61876 ARNTL aryl hydrocarbon receptor

like sulfated 2248 nuclear translocator like 2.74416 glycoprotein 1096 PACSIN2 protein kinase C and 1.61740 HS3ST3B 1 heparan sulfate- casein kinase substrate in 1771 glucosamine 3- 2.74416 neurons 2 sulfotransferase 3B1 1096

TRDMT1 tRNA aspartic acid 1.61523 OASL2 2'-5' oligoadenylate

methyltransferase 1 9219 synthetase-like 2(Oasl2) 2.74489

2108

4932438H2 Description Not Found 1.61468 PRDX6 peroxiredoxin 6

3RIK 1809 2.75084

462

SPAG9 sperm associated antigen 1.61456 RASA2 RAS p21 protein activator

9 7709 2 2.75120

3108

RPA3 replication protein A3 1.61436 HOXB2 homeobox B2

984 2.75488

7502

GNPTAB N-acetylglucosamine- 1 - 1.61329 TULP3 tubby like protein 3

phosphate transferase 8199 2.75488 alpha and beta subunits 7502

SNX9 sorting nexin 9 1.60925 MFRP membrane frizzled-related

1493 protein 2.75488

7502

OLFR550 olfactory receptor 1.60919 MEN1 menin 1

550(Olfr550) 5813 2.75755

6689

ZFP160 zinc finger protein 1.60880 C330021F2 RIKEN cDNA

160(Zfpl60) 9243 3RIK C330021F23 2.76219 gene(C330021F23Rik) 9201

TAS2R129 taste receptor, type 2, 1.60880 CSTAD CSA-conditional, T cell

member 129(Tas2rl29) 9243 activation-dependent 2.76553 protein(Cstad) 4746

OLFR371 olfactory receptor 1.60880 ALDH5A1 aldehyde dehydrogenase 5

371(01fr371) 9243 family member Al 2.77302

2439

OLFR281 olfactory receptor 1.60880 EPM2AIP1 EPM2A interacting protein

281(01fr281) 9243 1 2.77346

8928

OLFR195 olfactory receptor 1.60880 PDE8B phosphodiesterase 8B

195(01frl95) 9243 2.77610

3988

OLFR142 olfactory receptor 1.60880 DMRTA1 DMRT like family Al

142(01frl42) 9243 2.77618

4379

PRSS3 protease, serine 3 1.60880 LYPD6B LY6/PLAUR domain

9243 containing 6B 2.78004

8768

CX3CL1 C-X3-C motif 1.60880 CD300E CD300e molecule

chemokine ligand 1 9243 2.78659

6362

TMPRSS6 transmembrane protease, 1.60880 NPFF neuropeptide FF-amide

serine 6 9243 peptide precursor 2.78659

6362

ALK anaplastic lymphoma 1.60880 FASTKD l FAST kinase domains 1

receptor tyrosine kinase 9243 2.79376

5229

ITGA9 integrin subunit alpha 9 1.60880 OLFR802 olfactory receptor

9243 802(Olfr802) 2.79701

2978

TIMM13 translocase of inner 1.60880 HIVEP1 human immunodeficiency

mitochondrial membrane 9243 virus type I enhancer 2.79701 13 binding protein 1 2978

MSH5 mutS homolog 5 1.60880 HICl hypermethylated in cancer

9243 1 2.79701

2978

XP04 exportin 4 1.60581 TRIM33 tripartite motif containing

8241 33 2.80200

9226

MED21 mediator complex 1.60330 SELL selectin L

subunit 21 9406 2.80327

4253

CHST12 carbohydrate 1.60261 EPHX1 epoxide hydrolase 1

sulfotransferase 12 2589 2.80375

8579

6030408B1 Description Not Found 1.60219 BCL9 B-cell CLL/lymphoma 9

6RIK 5565 2.80735

4922

SLU7 SLU7 homolog, splicing 1.60154 STAT2 signal transducer and

factor 8066 activator of transcription 2 2.80852

1822

CDK5RAP2 CDK5 regulatory 1.60112 ELM03 engulfment and cell

subunit associated 0229 motility 3 2.81249 protein 2 8225

CASP7 caspase 7 1.60024 HDC histidine decarboxylase

02 2.81516

7456

KIF22 kinesin family member 1.59901 AI317395 Description Not Found

22 1705 2.81762

3258

E2F1 E2F transcription factor 1.59844 RPL14 ribosomal protein L 14

1 9678 2.81762

3258

MXI1 MAX interactor 1, 1.59769 SNAI1 snail family transcriptional

dimerization protein 0116 repressor 1 2.81825

6244

DONSON downstream neighbor of 1.59693 NUPR1 nuclear protein 1,

SON 5142 transcriptional regulator 2.82781

9025

TBX22 T-box 22 1.59693 IGSF8 immunoglobulin

5142 superfamily member 8 2.82781

9025

INPPL1 inositol polyphosphate 1.59630 SLC12A7 solute carrier family 12

phosphatase like 1 0192 member 7 2.82781

9025

CSE1L chromosome segregation 1.59586 RENBP renin binding protein

1 like 273 2.83743

1463

NDFIP2 Nedd4 family interacting 1.59470 ZFP553 zinc finger protein

protein 2 9608 553(Zfp553) 2.83794

3242

LYPD6 LY6/PLAUR domain 1.59296 LRFN2 leucine rich repeat and

containing 6 2293 fibronectin type III domain 2.83794 containing 2 3242

DDX49 DEAD -box helicase 49 1.59219 HP haptoglobin

0323 2.83973

7506

MGLL monoglyceride lipase 1.59094 TOMM40 translocase of outer

8822 mitochondrial membrane 2.84799

40 6907

NR4A3 nuclear receptor 1.59092 GABARAP GABA type A receptor - CREB3L2 cAMP responsive 1.56136 NSG2 neuron specific gene

element binding protein 1122 family member 2(Nsg2) 3.02046 3 like 2 6888

OLFR492 olfactory receptor 1.56071 ID 1 inhibitor of DNA binding

492(01fr492) 4954 1, HLH protein 3.02680

0059

OLFR1312 olfactory receptor 1.56071 CYP2D22 cytochrome P450, family

1312(01frl312) 4954 2, subfamily d, 3.04428 polypeptide 22(Cyp2d22) 2215

UPK2 uroplakin 2 1.56071 H2AFJ H2A histone family

4954 member J 3.04429

7135

RESP18 regulated endocrine 1.56071 TGFBR3 transforming growth factor

specific protein 18 4954 beta receptor 3 3.05311

1336

CRCT1 cysteine rich C-terminal 1.56071 IRS2 insulin receptor substrate 2

1 4954 3.06177

6198

NEUROD4 neuronal differentiation 1.56071 ADCY7 adenylate cyclase 7

4 4954 3.06608

919

SENP1 SUMOl/sentrin specific 1.56071 HYI hydroxypyruvate

peptidase 1 4954 isomerase (putative) 3.07231

5809

MR1 major histocompatibility 1.56071 TRIP4 thyroid hormone receptor

complex, class I-related 4954 interactor 4 3.07895

1341

BIVM basic, immunoglobulin- 1.56071 D730001G1 RIKEN cDNA

like variable motif 4954 8RIK D730001G18 3.08746 containing gene(D730001G18Rik) 2841

KPNA2 karyopherin subunit 1.56071 PRR7 proline rich 7 (synaptic)

alpha 2 4954 3.08746

2841

BAG2 BCL2 associated 1.56071 GFPT2 glutamine-fructose-6- athanogene 2 4954 phosphate transaminase 2 3.09592

442

SLC12A8 solute carrier family 12 1.56071 SCMH1 sex comb on midleg

member 8 4954 homolog 1 (Drosophila) 3.10013

6671

SCN7A sodium voltage-gated 1.56071 ANKRD 12 ankyrin repeat domain 12

channel alpha subunit 7 4954 3.10745

6458

SLC5A7 solute carrier family 5 1.56071 PTPRV protein tyrosine

member 7 4954 phosphatase, receptor type, 3.11270

V(Ptprv) 0133

ENPEP glutamyl aminopeptidase 1.56071 TMEM135 transmembrane protein

4954 135 3.11270

0133

ANGPTL4 angiopoietin like 4 1.56060 AKAP3 A-kinase anchoring

777 protein 3 3.11460

665

OSBPL3 oxysterol binding protein 1.55977 CBR2 carbonyl reductase

like 3 8376 2(Cbr2) 3.12928

3017

MCFD2 multiple coagulation 1.55961 CXCL16 C-X-C motif chemokine

factor deficiency 2 7874 ligand 16 3.12928

3017

MAP2K1 mitogen-activated 1.55855 MBTD1 mbt domain containing 1

protein kinase kinase 1 6708 3.14567 SLC31A1 solute carrier family 31 1.54027 KCNRG potassium channel

member 1 5536 regulator 3.26303

4406

TCTEX1D2 Tctexl domain 1.53833 POU6F1 POU class 6 homeobox 1

containing 2 2378 3.26851

7714

TTF2 transcription termination 1.53787 REV3L REV3 like, DNA directed

factor 2 1953 polymerase zeta catalytic 3.27052 subunit 8942

ZFP7 zinc finger protein 1.53605 TCF7 transcription factor 7 (T- 7(Zfp7) 29 cell specific, HMG-box) 3.27241

9178

G6PD2 gluco se -6 -pho sphate 1.53605 NME4 NME/NM23 nucleoside

dehydrogenase 2(G6pd2) 29 diphosphate kinase 4 3.28355

1423

DEFB 14 defensin beta 1.53605 PLAUR plasminogen activator,

14(Defbl4) 29 urokinase receptor 3.28540

2219

SLC18A3 solute carrier family 18 1.53605 CD4 CD4 molecule

member A3 29 3.28540

2219

AHNAK2 AHNAK nucleoprotein 2 1.53605 ZMYND11 zinc finger MYND-type

29 containing 11 3.29318

6363

HOXC12 homeobox C12 1.53605 ARMCX5 armadillo repeat

29 containing, X-linked 5 3.29840

4158

CEACAM1 carcinoembryonic 1.53605 LPHN1 Description Not Found

6 antigen related cell 29 3.30012 adhesion molecule 16 3725

MOSPD3 motile sperm domain 1.53605 PIK3IP1 phosphoinositide-3 -kinase

containing 3 29 interacting protein 1 3.30742

8525

DCTN1 dynactin subunit 1 1.53605 ERDR1 erythroid differentiation

29 regulator l(Erdrl) 3.31765

1188

MYB MYB proto-oncogene, 1.53605 PLD4 phospholipase D family

transcription factor 29 member 4 3.32844

4792

GLIPR1L2 GLI pathogenesis-related 1.53605 BMF Bcl2 modifying factor

1 like 2 29 3.33628

3388

ALDH1A3 aldehyde dehydrogenase 1.53605 GALNT11 polypeptide N- 1 family member A3 29 acetylgalactosaminyltransf 3.34511 erase 11 8795

SLC2A8 solute carrier family 2 1.53605 LCN2 lipocalin 2

member 8 29 3.37851

1623

SRC SRC proto-oncogene, 1.53605 PAG1 phosphoprotein membrane

non-receptor tyrosine 29 anchor with 3.38543 kinase glycosphingolipid 1037 microdomains 1

ZCCHC17 zinc finger CCHC-type 1.53561 DTX1 deltex E3 ubiquitin ligase

containing 17 8518 1 3.42557

6064

HNRNPUL heterogeneous nuclear 1.53442 RFFL ring finger and FYVE-like

1 ribonucleoprotein U like 0207 domain containing E3 3.42668

1 ubiquitin protein ligase 4082

TRIM68 tripartite motif 1.53305 MAFF MAF bZIP transcription - protein 1 like 1 2169 3.56778

1854

CHGB chromogranin B 1.51924 TNFSF14 tumor necrosis factor

262 superfamily member 14 3.58889

5735

TXNRD2 thioredoxin reductase 2 1.51900 TDRKH tudor and KH domain

8256 containing 3.59096

1241

NCF1 neutrophil cytosolic 1.51887 FIB CD 1 fibrinogen C domain

factor 1 3761 containing 1 3.60865

6121

OAF out at first homolog 1.51743 RBBP9 RB binding protein 9,

1856 serine hydrolase 3.60880

9243

FAM110A family with sequence 1.51726 DERL1 derlin 1

similarity 110 member A 3583 3.61765

1119

ANGEL 1 angel homolog 1 1.51583 LENG9 leukocyte receptor cluster

(Drosophila) 2566 member 9 3.62058

641

RTN4IP1 reticulon 4 interacting 1.51576 TRPC2 transient receptor potential

protein 1 0776 cation channel subfamily 3.62058

C member 2, pseudogene 641

LAMP2 lysosomal associated 1.51570 CCDC134 coiled-coil domain

membrane protein 2 9038 containing 134 3.63226

8216

KRT4 keratin 4 1.51429 OAS2 2'-5'-oligoadenylate

9789 synthetase 2 3.63226

8216

PAFAH1B3 platelet activating factor 1.51429 2410127L1 Description Not Found

acetylhydrolase lb 35 7RIK 3.64673 catalytic subunit 3 8698

STT3A STT3A, catalytic subunit 1.51353 RSAD1 radical S-adenosyl

of the 7695 methionine domain 3.64922 oligosaccharyltransferas containing 1 0471 e complex

PRKAR1B protein kinase cAMP- 1.51340 H2-DMB1 histocompatibility 2, class

dependent type I 003 II, locus Mb l(H2-DMbl) 3.64961 regulatory subunit beta 5459

HIST1H2B histone cluster 1, H2bb 1.51294 IFT81 intraflagellar transport 81

B 1595 3.67383

9056

ZFP39 zinc finger protein 1.51138 MIDI midline 1

39(Zfp39) 5424 3.68369

6454

PLK1 polo like kinase 1 1.51115 DEPDC1B DEP domain containing

1166 IB 3.68369

6454

1700028P14 Description Not Found 1.51096 SMAD3 SMAD family member 3

RIK 1919 3.71629

6166

D10BWG13 Description Not Found 1.51096 UBTD1 ubiquitin domain

79E 1919 containing 1 3.71699

0894

TREM3 triggering receptor 1.51096 FBX044 F-box protein 44

expressed on myeloid 1919 3.73876 cells 3(Trem3) 7837

GM128 predicted gene 1.51096 KCNMB4 potassium calcium- 128(Gml28) 1919 activated channel 3.74195 5679

ZFP282 zinc finger protein 1.50741 ZFP810 zinc finger protein

282(Zfp282) 9453 810(Zfp810) 4.07038

9328

TBC1D7 TBC1 domain family 1.50484 NAGA alpha-N- member 7 7821 acetylgalactosaminidase 4.07467

6686

ARSB arylsulfatase B 1.50484 KLRA20 killer cell lectin-like

5728 receptor subfamily A, 4.07895 member 20(Klra20) 1341

RAD 17 RAD 17 checkpoint 1.50417 STKl lIP serine/threonine kinase 11

clamp loader component 7542 interacting protein 4.08321

3368

CMTM7 CKLF like MARVEL 1.50329 KLF4 Kruppel like factor 4

transmembrane domain 7831 4.08430 containing 7 6687

NFKB2 nuclear factor kappa B 1.50036 INADL Description Not Found

subunit 2 3085 4.08666

7018

TOP3A topoisomerase (DNA) III URM1 ubiquitin related modifier

alpha 1.50007 1 4.09070

357 78

RAB33B RAB33B, member RAS PELI1 pellino E3 ubiquitin

oncogene family 1.50054 protein ligase 1 4.09381

042 3673

LYSMD1 LysM domain containing FBLN1 fibulin 1

1 1.50061 4.09803

4885 2083

POLG2 polymerase (DNA) HR hair growth associated

gamma 2, accessory 1.50070 4.13545 subunit 7646 2784

TGIF1 TGFB induced factor ASB6 ankyrin repeat and SOCS

homeobox 1 1.50119 box containing 6 4.13750

6523 3524

RELL1 RELT like 1 SLC27A5 solute carrier family 27

1.50300 member 5 4.14159 255 6278

CYP26B1 cytochrome P450 family PPP1R3F protein phosphatase 1

26 subfamily B member 1.50439 regulatory subunit 3F 4.14974 1 813 712

PTRH2 peptidyl-tRNA hydrolase AB124611 cDNA sequence

2 1.50467 AB124611(AB124611) 4.17337

8598 3402

ZKSCAN3 zinc finger with KRAB CD40 CD40 molecule

and SCAN domains 3 1.50491 4.18189

6722 7643

SP8 Sp8 transcription factor SMAD5 SMAD family member 5

1.50599 4.18388 9092 3459

SAMD 14 sterile alpha motif COL23A1 collagen type XXIII alpha

domain containing 14 1.50627 1 chain 4.22110

2343 3725

MX2 MX dynamin like ZFP595 zinc finger protein

GTPase 2 1.50726 595(Zfp595) 4.22881

8463 8691

OCRL OCRL, inositol PEC AMI platelet and endothelial

polyphosphate-5- 1.50763 cell adhesion molecule 1 4.23278 phosphatase 8755 9973

SY J2BP synaptojanin 2 binding - TMEM138 transmembrane protein - protein 1.50766 138 4.24122

9173 8289

CPLX4 complexin 4 RFX2 regulatory factor X2

1.50855 4.24412 4002 5943

LGALS9 galectin 9 KCTD12 potassium channel

1.50924 tetramerizafion domain 4.24784

6723 containing 12 6204

TAZ tafazzin TRIM56 tripartite motif containing

1.50926 56 4.26200 9953 8929

2310002L0 Description Not Found EIF4EBP2 eukaryotic translation

9RIK 1.51096 initiation factor 4E binding 4.26303

1919 protein 2 4406

ZFP97 zinc finger protein RALGPS2 Ral GEF with PH domain

97(Zfp97) 1.51096 and SH3 binding motif 2 4.27984

1919 2694

OLFR1494 olfactory receptor TGM2 transglutaminase 2

1494(01frl494) 1.51096 4.29316

1919 1941

BC030867 cDNA sequence ENC1 ectodermal-neural cortex 1

BC030867(BC030867) 1.51096 4.31106

1919 7102

CEACAM9 carcinoembryonic LRIG1 leucine rich repeats and

antigen-related cell 1.51096 immunoglobulin like 4.37503 adhesion molecule 1919 domains 1 9431 9(Ceacam9)

LRIT1 leucine rich repeat, Ig- PRM1 protamine 1

like and transmembrane 1.51096 4.37503 domains 1 1919 9431

KLK5 kallikrein related DUSP7 dual specificity

peptidase 5 1.51096 phosphatase 7 4.38353

1919 8076

KRT27 keratin 27 SERTAD3 SERTA domain containing

1.51096 3 4.39917 1919 1094

CACNG4 calcium voltage-gated KCNC1 potassium voltage-gated

channel auxiliary subunit 1.51096 channel subfamily C 4.40939 gamma 4 1919 member 1 0936

IL13RA1 interleukin 13 receptor UBE2D3 ubiquitin conjugating

subunit alpha 1 1.51096 enzyme E2 D3 4.46270

1919 6751

TMEM121 transmembrane protein SEPP1 selenoprotein P, plasma, 1

121 1.51096 4.46338

1919 3458

HIST1H2A histone cluster 1, H2aa ADRB2 adrenoceptor beta 2

A 1.51096 4.46391

1919 0999

MRZL3 myelin protein zero like PPP1R13B protein phosphatase 1

3 1.51096 regulatory subunit 13B 4.47141

1919 7658

TGFB2 transforming growth ARRDC3 arrestin domain containing

factor beta 2 1.51096 3 4.50462

1919 0392

IFT74 intraflagellar transport GNGT2 G protein subunit gamma

74 1.51096 transducin 2 4.53138

1919 1461

FCRL1 Fc receptor like 1 SIAH1A seven in absentia

1.51096 lA(Siahla) 4.53915 1919 8811

ADRB1 adrenoceptor beta 1 XPC XPC complex subunit,

1.51096 DNA damage recognition 4.56376 1919 and repair factor 8278

MAGI2 membrane associated HIPK1 homeodomain interacting

guanylate kinase, WW 1.51096 protein kinase 1 4.68369 and PDZ domain 1919 6454 containing 2

SCG5 secretogranin V H2-OB histocompatibility 2, O

1.51096 region beta locus(H2-Ob) 4.70043 1919 9718

GCK glucokinase BACH2 BTB domain and CNC

1.51096 homolog 2 4.71699 1919 0894

ASB 10 ankyrin repeat and MAP1LC3 microtubule associated

SOCS box containing 10 1.51096 A protein 1 light chain 3 4.72246

1919 alpha 6024

SELE selectin E LRRFIP1 LRR binding FLU

1.51096 interacting protein 1 4.76155 1919 1232

IGFBP3 insulin like growth factor ATP10D ATPase phospholipid

binding protein 3 1.51096 transporting 10D 4.76658

1919 (putative) 1958

TPT1 tumor protein, IGFBP4 insulin like growth factor

translationally-controlled 1.51096 binding protein 4 4.79099

1 1919 3785

ROCK1 Rho associated coiled- TMEM108 transmembrane protein

coil containing protein 1.51096 108 4.86542 kinase 1 1919 3978

OGFRL1 opioid growth factor PTK2 protein tyrosine kinase 2

receptor-like 1 1.51096 4.87571

1919 9796

TMEM38A transmembrane protein CLEC11A C-type lectin domain

38A 1.51096 family 11 member A 4.89724

1919 0426

RLTPR Description Not Found LRP12 LDL receptor related

1.51227 protein 12 4.95502

339 9571

ITPKC inositol-trisphosphate 3- GCNT2 glucosaminyl (N-acetyl)

kinase C 1.51238 transferase 2, 1-branching 4.95884

9725 enzyme (I blood group) 2675

TLE4 transducin like enhancer F10 coagulation factor X

of split 4 1.51341 4.96578

989 4285

PDE4D phosphodiesterase 4D DBP D-box binding PAR bZIP

1.51366 transcription factor 4.96654 7908 9451

A130010J1 Description Not Found ABCG1 ATP binding cassette

5RIK 1.51429 subfamily G member 1 5.00225

6211 2452

RNF167 ring finger protein 167 WDR78 WD repeat domain 78

1.51476 5.01792 5492 1908

CCBL1 Description Not Found DNAJC6 DnaJ heat shock protein

1.51562 family (Hsp40) member 5.01792 6494 C6 1908

HSD17B1 hydroxysteroid 17-beta AFF4 AF4/FMR2 family

dehydrogenase 1 1.51687 member 4 5.03342

5069 3002 OSM oncostatin M TNFRSF26 tumor necrosis factor

1.51723 receptor superfamily, 5.04001 4668 member 26(Tnfrsf26) 5679

RHPN1 rhophilin, Rho GTPase GFOD2 glucose-fructose

binding protein 1 1.51727 oxidoreductase domain 5.07038

5693 containing 2 9328

TAS2R105 taste receptor, type 2, TYROBP TYRO protein tyrosine

member 105(Tas2rl05) 1.51743 kinase binding protein 5.11478

1856 3447

NIPBL NIPBL, cohesin loading TMEM176 transmembrane protein

factor 1.51756 B 176B 5.11894

9618 1073

CXCR3 C-X-C motif chemokine ZFP710 zinc finger protein

receptor 3 1.51932 710(Zfp710) 5.15987

5267 1337

SMURF1 SMAD specific E3 ENPP4 ectonucleotide

ubiquitin protein ligase 1 1.52026 pyrophosphatase/phosphod 5.18189

3252 iesterase 4 (putative) 7643

RNF208 ring finger protein 208 MAPK8 mitogen-activated protein

1.52126 kinase 8 5.25927

647 2487

ITGA5 integrin subunit alpha 5 TNFRSF25 tumor necrosis factor

1.52351 receptor superfamily 5.28909 7983 member 25 6702

USP18 ubiquitin specific LCN4 lipocalin 4(Lcn4)

peptidase 18 1.52481 5.36632

4077 2214

PIP5K1A phosphatidylinositol-4- CRIM1 cysteine rich

phosphate 5 -kinase type 1.52507 transmembrane BMP 5.36981 1 alpha 4369 regulator 1 5424

STRBP spermatid perinuclear RTP4 receptor transporter

RNA binding protein 1.52561 protein 4 5.44460

213 0814

GRAMD2 GRAM domain PRNP prion protein

containing 2 1.52652 5.49505

805 5528

ZFP101 zinc finger protein ZFP747 zinc finger protein

lOl(ZfplOl) 1.52655 747(Zfp747) 5.49665

5668 4083

RUNDCl RUN domain containing CD7 CD7 molecule

1 1.52656 5.50462

3287 0392

SLC13A3 solute carrier family 13 ARHGAP2 Rho GTPase activating

member 3 1.52848 6 protein 26 5.54843

7927 6625

CCDC94 coiled-coil domain S100A9 SI 00 calcium binding

containing 94 1.52848 protein A9 5.55765

7927 5155

MRPS14 mitochondrial ribosomal AQP9 aquaporin 9

protein S14 1.52896 5.57288

2318 9668

NEU4 neuraminidase 4 CXCR5 C-X-C motif chemokine

(sialidase) 1.52982 receptor 5 5.57364

0947 7187

PCGF1 polycomb group ring CCNO cyclin O

finger 1 1.53059 5.57440

536 4309

PNPLA7 patatin like LYNX1 Ly6/neuro toxin 1

phospholipase domain 1.53207 5.66675 containing 7 883 6592

SPATA19 spermatogenesis CLDN10 claudin 10

associated 19 1.53301 5.78201

4103 5335

AP4B1 adaptor related protein AMIG02 adhesion molecule with

complex 4 beta 1 subunit 1.53382 Ig-like domain 2 5.83541

1865 884

BC068281 cDNA sequence CD79B CD79b molecule

BC068281(BC068281) 1.53605 5.94016

29 675

GK2 glycerol kinase 2 USP53 ubiquitin specific

1.53605 peptidase 53 5.98071 29 0829

PIGM pho sphatidy lino sitol IKBKE inhibitor of kappa light

glycan anchor 1.53605 polypeptide gene enhancer 6.00562 biosynthesis class M 29 in B-cells, kinase epsilon 4549

FKBP6 FK506 binding protein 6 ALOX5AP arachidonate 5-

1.53605 lipoxygenase activating 6.00898 29 protein 8783

EVI5 ecotropic viral GGT1 gamma- integration site 5 1.53605 glutamyltransferase 1 6.01010

29 8453

BCL11A B-cell CLL/lymphoma CAMK2D calcium/ calmodulin

11A 1.53605 dependent protein kinase 6.04766

29 II delta 9251

PERI period circadian clock 1 RAB3D RAB3D, member RAS

1.53727 oncogene family 6.15684 8499 1525

BTBD9 BTB domain containing MAP3K8 mitogen-activated protein

9 1.53745 kinase kinase kinase 8 6.37677

1456 6572

USP38 ubiquitin specific NOTCH4 notch 4

peptidase 38 1.53776 6.49505

3627 5528

LRRC57 leucine rich repeat MACROD 1 MACRO domain

containing 57 1.53808 containing 1 6.58120

3341 0582

5830415F09 Description Not Found RNF144A ring finger protein 144A

RIK 1.53855 6.63226

912 8216

EGR2 early growth response 2 PDE2A phosphodiesterase 2A

1.54003 6.86913 8325 112

GMEB2 glucocorticoid THAI threonine aldolase l(Thal)

modulatory element 1.54112 6.88508 binding protein 2 2795 6225

PIK3R4 phosphoinositide-3 - APP amyloid beta precursor

kinase regulatory subunit 1.54197 protein 6.94075 4 5323 4047

KRR1 KRR1, small subunit FAM109A family with sequence

processome component 1.54225 similarity 109 member A 6.96866 homolog 805 6793

COL9A1 collagen type IX alpha 1 LRG1 leucine rich alpha-2-

1.54225 glycoprotein 1 6.99548 805 4519

POLD4 polymerase (DNA) delta IL11RA1 interleukin 11 receptor,

4, accessory subunit 1.54265 alpha chain 1(111 lral) 7.01625

4605 1155

ACSS2 acyl-CoA synthetase - CNR2 cannabinoid receptor 2 - short-chain family 1.54404 7.21334 member 2 5378 7282

PDLIM1 PDZ and LIM domain 1 NUAK2 NUAK family kinase 2

1.54478 7.36981 5186 5424

A430107P0 Description Not Found GPR146 G protein-coupled receptor 9RIK 1.54492 146 7.57780

1568 6447

SLC38A11 solute carrier family 38

member 11 1.54622

2547

*Log2Fold Change = log2(4+L+/4-L^)

To investigate the molecular pathways between these three populations, gene ontology networks were grouped into nodes and the most significant pathways within each node were determined (Figure 6A). Gene ontology (GO) terms shared between our dysfunctional T cell dataset and the published hypofunctional T cell dataset were greatly enriched in cell cycle genes, consistent with the observation that the dysfunctional population is largely Ki67 ⁺ . GO terms shared between dysfunctional and exhausted gene sets encompassed effector programs such as regulation of cell killing, chemotaxis, interferon-γ production. GO terms shared between hypofunctional and exhausted gene sets consisted of cell cycle pathways, negative regulation of lymphocytes, and interferon-γ production. These data indicate that while some conserved molecular programs likely exist in these

dysfunctional differentiation states, many pathways may be differentially regulated between chronic viral infections and in the tumor context.

While many inhibitory receptors, including Pdcdl (PD-1), Haver 2 (TIM-3), Cd244 (2B4), Klrel, and Lag3 were shared between all data sets; the co-stimulatory receptors Tnfrsf4 (OX- 40) and Tnfrsf9 (4- IBB) were upregulated in dysfunctional and hypofunctional CD8 ⁺ TIL data sets. Therefore, to enrich in potential markers and therapeutic targets on tumor specific CD8 ⁺ TILs, the complete cell surface phenotype of the 4-lBB ⁺ LAG-3 ⁺ CD8 TIL population was characterized. Comparing the different CD8 ⁺ TIL subpopulations, several additional upregulated co-stimulatory receptors were found: Tnfrsfl8 (GITR), Nkg2d (KLRK1) and Cd27. The transcript for Nrpl (neuropilin-1), which encodes for a cell surface receptor protein implicated in CD4 ⁺ Treg function (Sarris et al, 2008; incorporated by reference in its entirety), was also highly expressed. Expression of many of these molecules was confirmed by flow cytometry at day 7, 14 and 21 after tumor inoculation (Figure 6C). The analysis was extended to include the co-stimulatory molecules ICOS and CD 160 and the inhibitory receptor T cell immunoreceptor with Ig and ITIM domains (TIGIT) because ICOS and CD 160 were close to the cutoff value and no probe was present for TIGIT in the gene array. In addition, recent reports indicate that targeting these receptors can be therapeutic in murine models of cancer (Johnston et al, 2014; Fan et al, 2014; incorporated by reference in their entireties). PD-1, TIGIT, TIM-3, CD27 and NRP1 were expressed the majority of the 4- lBB ⁺ LAG-3 ⁺ TIL population and expression was maintained over time. 2B4, CD160, CTLA4, OX-40, and GITR subdivided a lesser fraction of the 4-lBB ⁺ LAG-3 ⁺ population. The expression of several inhibitory receptors, 2B4, TIM3 and CD160 increased over this 3- week time frame while expression of the co-stimulatory receptors, ICOS and OX-40, decreased (Figure 6C).

To address if the dysfunctional CD8 ⁺ TILs are terminally-differentiated short term effector cells or memory -like cells, the expression of KLRG-1 and IL-7Ra (Joshi et al, 2007). Most of the CD8 ⁺ TIL were negative for KLRG-1 expression and there was no difference between the 4-lBB ⁺ LAG-3 ⁺ and 4-1BB LAG-3 populations. However, the majority of the 4-lBB ⁺ LAG-3 ⁺ TIL did not express the IL-7 receptor (IL-7Ra) compared to their negative counter parts (Figure 6D). These results indicate that the 4- IBB LAG-3 TIL, which are not apparently specific for antigens expressed in the tumor microenvironment, are more memory-like, yet at the same time the tumor antigen-specific LAG-3 ⁺ 4-lBB ⁺ subset has not fully acquired a terminal effector phenotype.

Functional relevance of genes that are differentially regulated in CD8 ⁺ 4-lBB ⁺ LAG-3 ⁺ TILs

The gene array results in Table 2 provide a list of genes characterizing CD8 ⁺ 4- lBB ⁺ LAG-3 ⁺ TILs. The list includes therapeutic targets and additional markers of anti-tumor immunity. Experiments conducted during development of embodiments herein to test the functional relevance of these additional targets/markers (Figure 11). Data indicate that the array has identified targets for immunotherapy, using knockout mice (e.g., PD-1, TIM-3, OX- 40ICOS, TIGIT, CD244, TNFRSF18, Nml, Nrpl, KLRGl, GM156, GPNMB, GPR65, TMEM205, and TMEM126A, CRTAM, Sema7a, etc.). Experiments demonstrate that Nrnl and CRTAM are negative regulators of the anti-tumor immune response, as knockout mice lacking either of these molecules showed improved immune-mediated tumor control in vivo. In contrast, Sema7a is a positive regulator of anti-tumor immune responses, as knockout mice lacking this molecule show diminished immune-mediated tumor control in vivo (Figure 11). These experiments indicate that agonists of Sema7a signaling and antagonists of Nrnl and/or CRTAM should be useful therapeutics for the treatment of cancer. Targeting 4- IBB andLAG-3 exerts anti-tumor activity in vivo and normalizes the function and phenotypic composition of CD8 ⁺ TILs

Experiments were conducted during development of embodiments herein to assess whether targeting these receptors might have therapeutic utility. To this end, an agonistic anti-4-lBB mAb was administered alone or in combination with a blocking anti-LAG-3 mAb in mice bearing established B16.SIY tumors. While each antibody treatment alone had some therapeutic effect as reflected by slower tumor growth, the combination was particularly potent (Figure 7A). Analysis of the tumor microenvironment revealed that improved tumor control with the combination therapy was accompanied by an increase in the number of CD8 ⁺ TILs specific for the SIY antigen (Figure 7B), consistent with results reported previously with anti-PD-Ll + anti-CTLA-4 mAb (Spranger et al., 2014b; Twyman-Saint Victor et al, 2015; incorporated by reference in their entireties).

It was next examined whether the therapeutic effect of anti-4-lBB + anti-LAG-3 mAbs was associated with a loss of phenotypic markers defining dysfunctional T cells in the steady state. Due to concern that re-analyzing the T cells for expression of LAG-3 and 4- 1BB might be problematic, as the administered Abs could theoretically modulate the target receptors from the cell surface, the coordinate expression of additional receptors as identified above by gene expression profiling was taken advantage of. Preliminary analyses of the bulk TIL subpopulations revealed decreased expression of NRPl and 2B4 following anti-LAG-3 + anti-4-lBB treatment (data not shown). Co-expression of 2B4 and NRPl on SIY-reactive CD8 ⁺ TILs identified by pentamer staining was analyzed. A 2.7-fold-decrease in the co- expression of 2B4 and NRPl was observed upon anti-4-lBB + and anti-LAG-3 mAb treatment (Figure 7C), indicating a loss of the surface phenotype associated with T cell dysfunction. To determine whether this change was accompanied by a shift towards an effector phenotype, expression of KLGR-1 was examined. Indeed, a marked increase in KLGR-1 expression was observed on the SIY-reactive TIL following treatment, and a 3.7- fold increase in the KLRG-l ^hl IL-7RA ^l0 population was observed (Figure 7D).

To eliminate the possibility that treatment with anti-LAG-3 + anti-4-lBB mAbs was not altering the phenotype of T cells already within the tumor but rather was supporting recruitment of newly primed functional T cells from secondary lymphoid organs, the S1PR inhibitor FTY720, which prevents T cell egress from lymph nodes (Halin et al., 2005;

incorporated by reference in its entirety), was utilized. The efficacy of anti-PD-Ll -based immunotherapies was preserved in the presence of FTY720, arguing for re-functionalization of TIL as the major mechanism of action (Spranger et al, 2014a; incorporated by reference in its entirety). FTY720 administration was started on day 6 after tumor inoculation, 24 hours before the start of anti-LAG-3 + anti-4-lBB treatment, and continued every day until TIL analysis on day 14. Peripheral blood analyzed at the same time point revealed marked depletion of circulating T cells (Figure 9). Despite this loss of circulating T cells, the down regulation of 2B4 and NRP1 and the shift towards the KLRG1 ^I -IRA ¹⁰ phenotype was nonetheless preserved (Figure 7E and F).

To examine functional restoration of the TIL, the KLRG-l ^lo IL-7RA ^l0 and KLRG- 1 ^hi IL-7RA ^l0 CD8 ⁺ TIL populations were sorted from B16.SIY tumors on day 14 following treatment and analyzed for IL-2 after restimulation in vitro. Indeed, the KLRG-l ^lo IL-7RA ^l0 and KLRG- l ^hl IL-7RA ^l0 populations showed an increased capacity to produce IL-2 upon stimulation (Figure 7G). The relative level of 11-2 mRNA was comparable between the two CD8 ⁺ TIL populations and control CD8 ⁺ CD44 ⁺ TdLN T cells. Collectively, these data indicate that anti- 4-lBB/anti-LAG-3 combinatorial treatment induces significant changes in the phenotype profile and promotes functional restoration of tumor antigen-specific CD8 ⁺ T cells already present within the tumor microenvironment.

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