WO/2021/044208 | ANTIBODY-DRUG CONJUGATE COMPRISING ANTIBODY AGAINST HUMAN ROR1, AND USE FOR THE SAME |
WO/2019/022563 | IDURONATE-2-SULFATASE CONJUGATE |
JP2023550287 | SAP FC fusion protein and method of use |
KUDIRKA ROMAS (US)
SAFINA BRIAN (US)
ZHOU MATTHEW (US)
WO2019084060A1 | 2019-05-02 | |||
WO2019222676A1 | 2019-11-21 | |||
WO2019069275A1 | 2019-04-11 | |||
WO2021081407A1 | 2021-04-29 | |||
WO2021067242A1 | 2021-04-08 | |||
WO2021046112A1 | 2021-03-11 | |||
WO2020252294A1 | 2020-12-17 | |||
WO2000069472A2 | 2000-11-23 | |||
WO2011096519A1 | 2011-08-11 | |||
WO2013125654A1 | 2013-08-29 | |||
WO2013125636A1 | 2013-08-29 | |||
WO2013125640A1 | 2013-08-29 | |||
WO2013125630A1 | 2013-08-29 | |||
WO2013018889A1 | 2013-02-07 | |||
WO2013018891A1 | 2013-02-07 | |||
WO2013018883A1 | 2013-02-07 | |||
WO2013018892A1 | 2013-02-07 | |||
WO2014014082A1 | 2014-01-23 | |||
WO2014014086A1 | 2014-01-23 | |||
WO2015020212A1 | 2015-02-12 | |||
WO2018079740A1 | 2018-05-03 | |||
WO2008144891A1 | 2008-12-04 | |||
WO2011145744A1 | 2011-11-24 | |||
WO2011155579A1 | 2011-12-15 | |||
WO2013077458A1 | 2013-05-30 | |||
WO2003074566A2 | 2003-09-12 | |||
WO2011068845A1 | 2011-06-09 | |||
WO2013068946A2 | 2013-05-16 | |||
WO2015098099A1 | 2015-07-02 | |||
WO2020142659A2 | 2020-07-09 | |||
WO2009052249A1 | 2009-04-23 | |||
WO2017156152A1 | 2017-09-14 | |||
WO2015172099A1 | 2015-11-12 |
US20020193311A1 | 2002-12-19 | |||
US20020147138A1 | 2002-10-10 | |||
US20020193311A1 | 2002-12-19 | |||
US6855689B2 | 2005-02-15 | |||
US20160145350A1 | 2016-05-26 | |||
US7416726B2 | 2008-08-26 | |||
US5624821A | 1997-04-29 | |||
US20070014795A1 | 2007-01-18 | |||
US20080286819A1 | 2008-11-20 | |||
US5821337A | 1998-10-13 | |||
US5677171A | 1997-10-14 | |||
US6054297A | 2000-04-25 | |||
US6165464A | 2000-12-26 | |||
US6339142B1 | 2002-01-15 | |||
US6407213B1 | 2002-06-18 | |||
US6639055B1 | 2003-10-28 | |||
US6719971B1 | 2004-04-13 | |||
US6800738B1 | 2004-10-05 | |||
US7074404B2 | 2006-07-11 | |||
US7862817B2 | 2011-01-04 | |||
US6676924B2 | 2004-01-13 | |||
US8642742B2 | 2014-02-04 | |||
US0723288A | 1903-03-24 | |||
US7776330B2 | 2010-08-17 | |||
US9617345B2 | 2017-04-11 | |||
US9982063B2 | 2018-05-29 | |||
US7999083B2 | 2011-08-16 | |||
US20160297890A1 | 2016-10-13 | |||
US7238785B2 | 2007-07-03 | |||
US9797907B2 | 2017-10-24 | |||
US9382329B2 | 2016-07-05 | |||
US7420040B2 | 2008-09-02 | |||
US20120237518A1 | 2012-09-20 | |||
US10227417B2 | 2019-03-12 | |||
US8871908B2 | 2014-10-28 | |||
US9676863B2 | 2017-06-13 | |||
US7521541B2 | 2009-04-21 | |||
US7723485B2 | 2010-05-25 | |||
US20120121615A1 | 2012-05-17 | |||
US20190300513A1 | 2019-10-03 |
STARCHER, J., INVEST. DERM., vol. 107, 1996, pages 159 - 163
LERMAN, I., STEROIDS, vol. 133, 2018, pages 96 - 101
DIAS, A.R.M ET AL., CHEM. EUR. J., vol. 25, 2019, pages 1696 - 1700
HANAHAN D. ET AL., CELL, vol. 144, 2011, pages 646 - 674
FLYGARE, J.A. ET AL., CHEM BIOL DRUGDES., vol. 81, 2013, pages 113 - 121
KASPERKIEWICZ, P. ET AL., PROC. NAT. ACAD. SCI., vol. 111, pages 2518 - 2523
ZERVOUDI E ET AL., BIOCHEM, vol. Y435, no. 2, 2011, pages 411 - 420
POREBA M ET AL., PLOS ONE, vol. 7, no. 2, 2012, pages e31938
MAHLA, RS ET AL., FRONTIERS IN IMMUNOLOGY, vol. 4, no. 248, 2013
KUMAR, H ET AL., INTL. REV. OF IMMUN., vol. 30, 2011, pages 16 - 34
SCHRODER K ET AL., CELL, vol. 140, no. 6, 2010, pages 821 - 832
"GenBank", Database accession no. AAK62677
ALTSCHUL ET AL., J. MOLECULAR BIOL., vol. 215, no. 3, 1990, pages 403 - 410
BEIGERT ET AL., PROC. NATL. ACAD. SCI. USA, vol. 106, no. 10, 2009, pages 3770 - 3775
JEFFERIS ET AL., MABS, vol. 1, no. 4, 2009, pages 332 - 338
SODING, BIOINFORMATICS, vol. 21, no. 7, 2005, pages 951 - 960
ALTSCHUL ET AL., NUCLEIC ACIDS RES., vol. 25, no. 17, 1997, pages 3389 - 3402
GUSFIELD: "Algorithms on Strings, Trees and Sequences", 1997, CAMBRIDGE UNIVERSITY PRESS
RUSSELL ET AL., J. MOLBIOL., vol. 244, 1994, pages 332 - 350
PAQUETTE, LEO A.: "Principles of Modern Heterocyclic Chemistry", 1968, W.A. BENJAMIN
"The Chemistry of Heterocyclic Compounds, A series of Monographs", 1950, JOHN WILEY & SONS
J. AM. CHEM. SOC., vol. 82, no. 5566, 1960
LIEBERMAN, PHARMACEUTICAL DOSAGE FORMS, vol. 1-3, 1992
LLOYD, THE ART, SCIENCE AND TECHNOLOGY OF PHARMACEUTICAL COMPOUNDING, 1999
PICKAR: "Dosage Calculations", 1999, MCGRAW-HILL, article "Goodman & Gilman's The Pharmacological Basis of Therapeutics"
COUSSENS ET AL., SCIENCE, vol. 230, 1985, pages 1132 - 9
SLAMON ET AL., SCIENCE, vol. 244, 1989, pages 707 - 12
SLAMON ET AL., NEW ENGL. J. MED., vol. 344, 2001, pages 783 - 792
YARDEN, ONCOGENE, vol. 19, 2000, pages 6102 - 14
YARDENSLIWKOWSKI., NAT REV MOL CELL BIOL, vol. 2, 2001, pages 127 - 37
SLIWKOWSKI, NAT STRUCT BIOL, vol. 10, 2003, pages 158 - 9
CHO ET AL., NATURE, vol. 421, 2003, pages 756 - 60
MALIK ET AL., PRO AM SOC CANCER RES, vol. 44, 2003, pages 176 - 7
ELLIS JALUZIO JP, JBIOL CHEM., vol. 270, no. 35, 1995, pages 20717 - 23
WANG B ET AL., J IMMUNOL., vol. 175, no. 7, 2005, pages 4274 - 4282
SOLOMON S ET AL., MOL CELL BIOL., vol. 27, no. 6, 2007, pages 2324 - 42
YANG, Z-S ET AL., ONCOLOGY LETTERS, vol. 18, 2019, pages 15 - 21
BLUMENTHAL, R. ET AL., CANCER IMMUNOLOGY IMMUNOTHERAPY, vol. 54, no. 4, 2005, pages 315 - 327
CARDILLO, T. ET AL., MOLECULAR CANCER THERAPEUTICS, vol. 17, no. 1, 2018, pages 150 - 160
LINNENBACH AJ ET AL., MOL CELL BIOL, vol. 13, no. 3, 1993, pages 1507 - 15
CALABRESE G ET AL., CYTOGENET CELL GENET, vol. 92, no. 1-2, 2001, pages 164 - 5
OHMACHI T ET AL., CLIN. CANCER RES., vol. 12, no. 10, 2006, pages 3057 - 3063
MUHLMANN G ET AL., J. CLIN. PATHOL., vol. 62, no. 2, 2009, pages 152 - 158
FONG D ET AL., BR. J. CANCER, vol. 99, no. 8, 2008, pages 1290 - 1295
FONG D ET AL., MOD. PATHOL., vol. 21, no. 2, 2008, pages 186 - 191
NING S ET AL., NEUROL. SCI., vol. 34, no. 10, 2013, pages 1745 - 1750
FAULK W P ET AL., PROC. NATL. ACAD. SCI., vol. 75, no. 4, 1978, pages 1947 - 1951
LIPINSKI M ET AL., PROC. NATL. ACAD. SCI., vol. 78, no. 8, 1981, pages 5147 - 5150
LINNENBACH A J ET AL., PROC. NATL. ACAD. SCI., vol. 86, no. 1, 1989, pages 27 - 31
FORNARO M ET AL., INT. J. CANCER, vol. 62, no. 5, 1995, pages 610 - 618
JUNUTULA ET AL., NATURE BIOTECH.,, vol. 26, no. 8, 2008, pages 925 - 932
DORNAN ET AL., BLOOD, vol. 114, no. 13, 2009, pages 2721 - 2729
JAVAID, N. ET AL., PHARMACEUTICS, vol. 11, no. 9, 2019, pages 441
RAMANJULU, J.M. ET AL., NATURE, vol. 564, 2018, pages 439 - 443
BARBER, G.N., NATURE REV IMMUNOL, vol. 15, 2015, pages 760 - 770
NEGRONI, A. ET AL., J. INFLAMM. RES., vol. 11, 2018, pages 49 - 60
COULOMBE, F. ET AL., J. EXP. MED., vol. 206, no. 8, 2009, pages 1709 - 1716
ELION, D.L. ET AL., ONCOTARGET, vol. 9, no. 48, 2018, pages 29007 - 29017
KOHLWAY, A., EMBO REP, vol. 14, 2013, pages 772 - 779
MANGAN, M. ET AL., NATURE REVIEWS DRUG DISCOVERY, vol. 17, 2018, pages 588 - 606
HERMANSON: "Bioconjugate Techniques", 2008, ACADEMIC PRESS
KHOT, A. ET AL., BIOANALYSIS, vol. 7, no. 13, 2015, pages 1633 - 1648
LI, F. ET AL., CANCER RES., vol. 76, 2016, pages 2710 - 2719
LYON, R. ET AL., METHODS IN ENZYM., vol. 502, 2012, pages 123 - 138
MCDONAGH ET AL., PROT. ENGR. DESIGN & SELECTION, vol. 19, no. 7, 2006, pages 299 - 307
HAMBLETT ET AL., CLIN. CANCER RES., vol. 10, 2004, pages 7063 - 7070
HAMBLETT, K.J. ET AL.: "Annual Meeting, March 27-31, 2004, Proceedings of the AACR", vol. 45, March 2004, AMERICAN ASSOCIATION FOR CANCER RESEARCH, article "Effect of drug loading on the pharmacology, pharmacokinetics, and toxicity of an anti-CD30 antibody-drug conjugate"
ALLEY, S.C. ET AL.: "2004 Annual Meeting, March 27-31, 2004, Proceedings of the AACR", vol. 45, March 2004, AMERICAN ASSOCIATION FOR CANCER RESEARCH, article "Controlling the location of drug attachment in antibody-drug conjugates"
CLAIMS: 1. An immunoconjugate comprising a cell-binding agent covalently attached to one or more immunostimulatory moieties by a linker comprising an elastase-substrate, peptide linker unit. 2. The immunoconjugate of claim 1 wherein the cell-binding agent is an antibody. 3. The immunoconjugate of claim 2 wherein the antibody is an antibody construct that has an antigen binding domain that binds PD-L1. 4. The immunoconjugate of claim 3 wherein the antibody is selected from the group consisting of atezolizumab, durvalumab, and avelumab, or a biosimilar or a biobetter thereof. 5. The immunoconjugate of claim 2 wherein the antibody is an antibody construct that has an antigen binding domain that binds HER2. 6. The immunoconjugate of claim 5 wherein the antibody is selected from the group consisting of trastuzumab and pertuzumab, or a biosimilar or a biobetter thereof. 7. The immunoconjugate of claim 2 wherein the antibody is an antibody construct that has an antigen binding domain that binds CEA. 8. The immunoconjugate of claim 7 wherein the antibody is labetuzumab, or a biosimilar or a biobetter thereof. 9. The immunoconjugate of claim 2 wherein the antibody is an antibody construct that has an antigen binding domain that binds Trop2. 10. The immunoconjugate of claim 7 wherein the antibody is sacituzumab, or a biosimilar or a biobetter thereof. 11. The immunoconjugate of any one of claims 1-10 wherein the one or more immunostimulatory moieties is a pattern-recognition receptor. 12. The immunoconjugate of any one of claims 1-11 wherein the one or more immunostimulatory moieties interact with or modulate a receptor selected from the group consisting of TLR, STING, NOD2, RIG-1, and NLRP3. 13. The immunoconjugate of any one of claims 2-12 having the Formula I: Ab−[L−Ims]p I or a pharmaceutically acceptable salt thereof, wherein: Ab is the antibody; L is the linker comprising an elastase-substrate, peptide linker unit; Ims is the immunostimulatory moiety; and p is an integer from 1 to 8. 14. The immunoconjugate of claim 13 wherein Ims is a Toll-like receptor (TLR) agonist. 15. The immunoconjugate of claim 13 wherein Ims is selected from formulas Ia-f: wherein R1, R2, R3, R4, and R5 are independently selected from the group consisting of H, C1-C12 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C12 carbocyclyl, C6-C20 aryl, C2-C9 heterocyclyl, and C1-C20 heteroaryl, where alkyl, alkenyl, alkynyl, carbocyclyl, aryl, heterocyclyl, and heteroaryl are independently and optionally substituted with one or more groups selected from: −(C1-C12 alkyldiyl)−N(R6)−*; −(C1-C12 alkyldiyl)−N(R6)2; −(C1-C12 alkyldiyl)−OR6; −(C3-C12 carbocyclyl); −(C3-C12 carbocyclyl)−*; −(C3-C12 carbocyclyl)−(C1-C12 alkyldiyl)−NR6−*; −(C3-C12 carbocyclyl)−(C1-C12 alkyldiyl)−N(R6)2; −(C3-C12 carbocyclyl)−NR5−C(=NR6a)NR6−*; −(C6-C20 aryl); −(C6-C20 aryldiyl)−*; −(C6-C20 aryldiyl)−N(R6)−*; −(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−N(R6)−*; −(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−(C2-C20 heterocyclyldiyl)−*; −(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−N(R6)2; −(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−NR6−C(=NR6a)N(R6)−*; −(C2-C20 heterocyclyl); −(C2-C20 heterocyclyl)−*; −(C2-C9 heterocyclyl)−(C1-C12 alkyldiyl)−NR6−*; −(C2-C9 heterocyclyl)−(C1-C12 alkyldiyl)−N(R6)2; −(C2-C9 heterocyclyl)−C(=O)−(C1-C12 alkyldiyl)−N(R6)−*; −(C2-C9 heterocyclyl)−NR5−C(=NR6a)NR6−*; −(C2-C9 heterocyclyl)−NR6−(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−N(R6)−*; −(C2-C9 heterocyclyl)−(C6-C20 aryldiyl)−*; −(C1-C20 heteroaryl); −(C1-C20 heteroaryldiyl)−*; −(C1-C20 heteroaryldiyl)−(C1-C12 alkyldiyl)−N(R6)−*; −(C1-C20 heteroaryldiyl)−(C1-C12 alkyldiyl)−N(R6)2; −(C1-C20 heteroaryldiyl)−NR6−C(=NR6a)N(R6)−*; −(C1-C20 heteroaryldiyl)−N(R6)C(=O)−(C1-C12 alkyldiyl)−N(R6)−*; −C(=O)−*; −C(=O)−(C1-C12 alkyldiyl)−N(R6)−*; −C(=O)−(C2-C20 heterocyclyldiyl)−*; −C(=O)N(R6)2; −C(=O)N(R6)−*; −C(=O)N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)R5; −C(=O)N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)N(R6)2; −C(=O)NR6−(C1-C12 alkyldiyl)−N(R6)CO2R6; −C(=O)NR6−(C1-C12 alkyldiyl)−N(R6)C(=NR6a)N(R6)2; −C(=O)NR6−(C1-C12 alkyldiyl)−NR6C(=NR6a)R6; −C(=O)NR6−(C1-C8 alkyldiyl)−NR6(C2-C5 heteroaryl); −C(=O)NR6−(C1-C20 heteroaryldiyl)−N(R6)−*; −C(=O)NR6−(C1-C20 heteroaryldiyl)−*; −C(=O)NR6−(C1-C20 heteroaryldiyl)−(C1-C12 alkyldiyl)−N(R6)2; −C(=O)NR6−(C1-C20 heteroaryldiyl)−(C2-C20 heterocyclyldiyl)−C(=O)NR6−(C1-C12 alkyldiyl)−NR6−*; −N(R6)2; −N(R6)−*; −N(R6)C(=O)R6; −N(R6)C(=O)−*; −N(R6)C(=O)N(R6)2; −N(R6)C(=O)N(R6)−*; −N(R6)CO2R6; −N(R6)CO2(R6)−*; −NR6C(=NR6a)N(R6)2; −NR6C(=NR6a)N(R6)−*; −NR6C(=NR6a)R6; −N(R6)C(=O)−(C1-C12 alkyldiyl)−N(R6)−*; −N(R6)−(C2-C5 heteroaryl); −N(R6)−S(=O)2−(C1-C12 alkyl); −O−(C1-C12 alkyl); −O−(C1-C12 alkyldiyl)−N(R6)2; −O−(C1-C12 alkyldiyl)−N(R6)−*; −OC(=O)N(R6)2; −OC(=O)N(R6)−*; −S(=O)2−(C2-C20 heterocyclyldiyl)−*; −S(=O)2−(C2-C20 heterocyclyldiyl)−(C1-C12 alkyldiyl)−N(R6)2; −S(=O)2−(C2-C20 heterocyclyldiyl)−(C1-C12 alkyldiyl)−NR6−*; and −S(=O)2−(C2-C20 heterocyclyldiyl)−(C1-C12 alkyldiyl)−OH; or R2 and R3 of formulas Ic-If together form a 5- or 6-membered heterocyclyl ring; X1, X2, X3, X4, and X5 are independently selected from the group consisting of a bond, C(=O), C(=O)N(R6), O, N(R6), S, S(O)2, and S(O)2N(R6); R6 is selected from the group consisting of H, C6-C20 aryl, C6-C20 aryldiyl, C1-C12 alkyl, and C1-C12 alkyldiyl, or two R6 groups together form a 5- or 6-membered heterocyclyl ring; R6a is selected from the group consisting of C6-C20 aryl and C1-C20 heteroaryl; where the asterisk * indicates the attachment site of L, and where one of R1, R2, R3, R4 and R5 is attached to L; and alkyl, alkyldiyl, alkenyl, alkenyldiyl, alkynyl, alkynyldiyl, aryl, aryldiyl, carbocyclyl, carbocyclyldiyl, heterocyclyl, heterocyclyldiyl, heteroaryl, and heteroaryldiyl are independently and optionally substituted with one or more groups independently selected from F, Cl, Br, I, − CN, −CH3, −CH2CH3, −CH=CH2, −C ≡CH, −C ≡CCH3, −CH2CH2CH3, −CH(CH3)2, − CH2CH(CH3)2, −CH2OH, −CH2OCH3, −CH2CH2OH, −C(CH3)2OH, −CH(OH)CH(CH3)2, − C(CH3)2CH2OH, −CH2CH2SO2CH3, −CH2OP(O)(OH)2, −CH2F, −CHF2, −CF3, −CH2CF3, − CH2CHF2, −CH(CH3)CN, −C(CH3)2CN, −CH2CN, −CH2NH2, −CH2NHSO2CH3, −CH2NHCH3, −CH2N(CH3)2, −CO2H, −COCH3, −CO2CH3, −CO2C(CH3)3, −COCH(OH)CH3, −CONH2, − CONHCH3, −CON(CH3)2, −C(CH3)2CONH2, −NH2, −NHCH3, −N(CH3)2, −NHCOCH3, − N(CH3)COCH3, −NHS(O)2CH3, −N(CH3)C(CH3)2CONH2, −N(CH3)CH2CH2S(O)2CH3, − NHC(=NH)H, −NHC(=NH)CH3, −NHC(=NH)NH2, −NHC(=O)NH2, −NO2, =O, −OH, −OCH3, −OCH2CH3, −OCH2CH2OCH3, −OCH2CH2OH, −OCH2CH2N(CH3)2, −O(CH2CH2O)n− (CH2)mCO2H, −O(CH2CH2O)nH, −OP(O)(OH)2, −S(O)2N(CH3)2, −SCH3, −S(O)2CH3, and − S(O)3H. 16. The immunoconjugate of claim 15 wherein one of R1, R2, R3, R4 and R5 is selected from the formulas: . 17. The immunoconjugate of claim 13 wherein Ims is a STING agonist. 18. The immunoconjugate of claim 13 wherein Ims has formula Ig: wherein Xa and Xb are independently selected from a five-membered heteroaryl; R1 is selected from the group consisting of F, Cl, Br, I, −CN, −OH, and −O−(C1-C6 alkyldiyl). R2a and R2b are independently selected from −C(=O)N(R5)2; R3 is selected from C1-C6 alkyldiyl, −(C1-C3 alkyldiyl)−O−(C1-C3 alkyldiyl)−, C2-C6 alkenyldiyl and C2-C6 alkynyldiyl, optionally substituted with one or more groups selected from F, Cl, −OH, −OCH3, −OCH2CH3, −OCH2CH2OCH3, −OCH2CH2OH, −OCH2CH2N(CH3)2; R4 is selected from the group consisting of: −(C1-C12 alkyldiyl)−N(R5)−*; −(C1-C12 alkyldiyl)−(C2-C20 heterocyclyldiyl)−*; −O−(C1-C12 alkyldiyl)−N(R5)−*; −O−(C1-C12 alkyldiyl)−(C2-C20 heterocyclyldiyl)−*; −O−(C1-C12 alkyldiyl)−(C2-C20 heterocyclyldiyl)−N(R5)−*; −OC(=O)N(R5)−*; −N(R5)−(C1-C12 alkyldiyl)−N(R5)−*; −N(R5)−(C1-C12 alkyldiyl)−(C2-C20 heterocyclyldiyl)−*; −C(=O)N(R5)−*; −(C2-C20 heterocyclyldiyl)−*; −S(=O)2−(C2-C20 heterocyclyldiyl)−*; −S(=O)2−(C2-C20 heterocyclyldiyl)−(C1-C12 alkyldiyl)−NR5−*; where the asterisk * indicates the attachment site of L; R5 is independently H or C1-C6 alkyl, or two R5 groups together form a 5- or 6- membered heterocyclyl ring; and alkyl, alkyldiyl, alkenyl, alkenyldiyl, alkynyl, alkynyldiyl, aryl, aryldiyl, carbocyclyl, carbocyclyldiyl, heterocyclyl, heterocyclyldiyl, heteroaryl, and heteroaryldiyl are independently and optionally substituted with one or more groups independently selected from F, Cl, Br, I, − CN, −CH3, −CH2CH3, −CH=CH2, −C ≡CH, −C ≡CCH3, −CH2CH2CH3, −CH(CH3)2, − CH2CH(CH3)2, −CH2OH, −CH2OCH3, −CH2CH2OH, −C(CH3)2OH, −CH(OH)CH(CH3)2, − C(CH3)2CH2OH, −CH2CH2SO2CH3, −CH2OP(O)(OH)2, −CH2F, −CHF2, −CF3, −CH2CF3, − CH2CHF2, −CH(CH3)CN, −C(CH3)2CN, −CH2CN, −CH2NH2, −CH2NHSO2CH3, −CH2NHCH3, −CH2N(CH3)2, −CO2H, −COCH3, −CO2CH3, −CO2C(CH3)3, −COCH(OH)CH3, −CONH2, − CONHCH3, −CON(CH3)2, −C(CH3)2CONH2, −NH2, −NHCH3, −N(CH3)2, −NHCOCH3, − N(CH3)COCH3, −NHS(O)2CH3, −N(CH3)C(CH3)2CONH2, −N(CH3)CH2CH2S(O)2CH3, − NHC(=NH)H, −NHC(=NH)CH3, −NHC(=NH)NH2, −NHC(=O)NH2, −NO2, =O, −OH, −OCH3, −OCH2CH3, −OCH2CH2OCH3, −OCH2CH2OH, −OCH2CH2N(CH3)2, −O(CH2CH2O)n− (CH2)mCO2H, −O(CH2CH2O)nH, −OCH2F, −OCHF2, −OCF3, −OP(O)(OH)2, −S(O)2N(CH3)2, − SCH3, −S(O)2CH3, and −S(O)3H. 19. The immunoconjugate of claim 18 wherein Xa and Xb are independently selected from the group consisting of imidazolyl, pyrazolyl, triazolyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxadiazolyl, oxazolyl, isothiazolyl, pyrrolyl, oxadiazolyl, and thiadiazolyl. 20. The immunoconjugate of claim 19 wherein Xa and Xb are each pyrazolyl, substituted with one or more C1-C12 alkyl groups. 21. The immunoconjugate of claim 18 wherein R1 is selected from the group consisting of −OCH3, −OCH2CH3, −OCH2CH2OCH3, −OCH2CH2OH, and −OCH2CH2N(CH3)2 22. The immunoconjugate of claim 21 wherein R1 is −OCH3. 23. The immunoconjugate of claim 18 wherein R1 is F. 24. The immunoconjugate of claim 18 wherein R2a and R2b are each −C(=O)NH2. 25. The immunoconjugate of claim 18 wherein R3 is selected from −CH2CH2−, − CH=CH−, and −C ≡C−. 26. The immunoconjugate of claim 18 wherein R3 is C2-C4 alkenyldiyl, substituted with one or more groups selected from F, −OH, and −OCH3. 27. The immunoconjugate of claim 18 wherein R4 is −O−(C1-C12 alkyldiyl)−(C2-C20 heterocyclyldiyl)−*. 28. The immunoconjugate of claim 27 wherein C1-C12 alkyldiyl is propyldiyl and C2- C20 heterocyclyldiyl is piperidiyl. 29. The immunoconjugate of claim 13 wherein L is selected from the group consisting of: −C(=O)−(PEG)−C(=O)−(EsPEP)−; −C(=O)−(PEG)−C(=O)−(EsPEP)−N(R6)−; −C(=O)−(PEG)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−; −C(=O)−(PEG)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)−(C2-C5 monoheterocyclyldiyl)−; −C(=O)−(PEG)−N(R6)−(PEG)−C(=O)−(EsPEP)−; −C(=O)−(PEG)−N+(R6)2−(PEG)−C(=O)−(EsPEP)−; −C(=O)−(PEG)−C(=O)−N(R6)CH(AA1)C(=O)−(PEG)−C(=O)−(EsPEP)−; −C(=O)−(C1-C12 alkyldiyl)−C(=O)−(EsPEP)−; −C(=O)−(C1-C12 alkyldiyl)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−; −C(=O)−(C1-C12 alkyldiyl)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−N(R6)−C(=O); −C(=O)−(C1-C12 alkyldiyl)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)− (C2-C5 monoheterocyclyldiyl)−; −succinimidyl−(CH2)m−C(=O)N(R6)−PEG−C(=O)−(EsPEP)−; −succinimidyl−(CH2)m−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−; −succinimidyl−(CH2)m−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)N(R6)C(=O)−; and −(succinimidyl)−(CH2)m−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)−(C2- C5 monoheterocyclyldiyl)−; PEG has the formula: −(CH2CH2O)n−(CH2)m−; m is an integer from 1 to 5, and n is an integer from 2 to 50; EsPEP is the elastase-substrate, peptide linker unit comprising 2 to 12 amino acid residues; and R6 is selected from the group consisting of H, C6-C20 aryl, C6-C20 aryldiyl, C1-C12 alkyl, and C1-C12 alkyldiyl, or two R6 groups together form a 5- or 6-membered heterocyclyl ring; alkyl, alkyldiyl, alkenyl, alkenyldiyl, alkynyl, alkynyldiyl, aryl, aryldiyl, carbocyclyl, carbocyclyldiyl, heterocyclyl, heterocyclyldiyl, heteroaryl, and heteroaryldiyl are independently and optionally substituted with one or more groups independently selected from F, Cl, Br, I, − CN, −CH3, −CH2CH3, −CH=CH2, −C ≡CH, −C ≡CCH3, −CH2CH2CH3, −CH(CH3)2, − CH2CH(CH3)2, −CH2OH, −CH2OCH3, −CH2CH2OH, −C(CH3)2OH, −CH(OH)CH(CH3)2, − C(CH3)2CH2OH, −CH2CH2SO2CH3, −CH2OP(O)(OH)2, −CH2F, −CHF2, −CF3, −CH2CF3, − CH2CHF2, −CH(CH3)CN, −C(CH3)2CN, −CH2CN, −CH2NH2, −CH2NHSO2CH3, −CH2NHCH3, −CH2N(CH3)2, −CO2H, −COCH3, −CO2CH3, −CO2C(CH3)3, −COCH(OH)CH3, −CONH2, − CONHCH3, −CON(CH3)2, −C(CH3)2CONH2, −NH2, −NHCH3, −N(CH3)2, −NHCOCH3, − N(CH3)COCH3, −NHS(O)2CH3, −N(CH3)C(CH3)2CONH2, −N(CH3)CH2CH2S(O)2CH3, − NHC(=NH)H, −NHC(=NH)CH3, −NHC(=NH)NH2, −NHC(=O)NH2, −NO2, =O, −OH, −OCH3, −OCH2CH3, −OCH2CH2OCH3, −OCH2CH2OH, −OCH2CH2N(CH3)2, −O(CH2CH2O)n− (CH2)mCO2H, −O(CH2CH2O)nH, −OP(O)(OH)2, −S(O)2N(CH3)2, −SCH3, −S(O)2CH3, and − S(O)3H. 30. The immunoconjugate of claim 29 wherein EsPEP is comprised of one or more unnatural amino acid residues. 31. The immunoconjugate of claim 29 wherein EsPEP is comprised of amino acid residues of amino acids selected from the group consisting of: 32. The immunoconjugate of claim 29 wherein EsPEP is selected from the group consisting of Ala-Pro-Val, Asn-Pro-Val, Ala-Ala-Val, Ala-Ala-Pro-Ala (SEQ ID NO: 639), Ala- Ala-Pro-Val (SEQ ID NO: 640), and Ala-Ala-Pro-Nva (SEQ ID NO: 641). 33. The immunoconjugate of claim 29 wherein EsPEP has the formula: where AA is independently selected from a natural or unnatural amino acid side chain, or one or more of AA, and an adjacent nitrogen atom form a 5-membered ring proline amino acid, and the wavy line indicates a point of attachment; Cyc is selected from C6-C20 aryldiyl and C1-C20 heteroaryldiyl, optionally substituted with one or more groups selected from F, Cl, NO2, −OH, −OCH3, and a glucuronic acid having the structure: R7 is selected from the group consisting of −CH(R8)O−, −CH2−, −CH2N(R8)−, and − CH(R8)O−C(=O)−, where R8 is selected from H, C1-C6 alkyl, C(=O)−C1-C6 alkyl, and − C(=O)N(R9)2, where R9 is independently selected from the group consisting of H, C1-C12 alkyl, and −(CH2CH2O)n−(CH2)m−OH, where m is an integer from 1 to 5, and n is an integer from 2 to 50, or two R9 groups together form a 5- or 6-membered heterocyclyl ring; y is an integer from 2 to 12; and z is 0 or 1. 34. The immunoconjugate of claim 33 wherein y is selected from 2, 3, and 4. 35. The immunoconjugate of claim 29 wherein EsPEP is a tripeptide having the formula: where AA1, AA2 and AA3 are independently selected from a natural or unnatural amino acid, and the wavy line indicates a point of attachment; Cyc is selected from C6-C20 aryldiyl and C1-C20 heteroaryldiyl, optionally substituted with one or more groups selected from F, Cl, NO2, −OH, −OCH3, and a glucuronic acid having the structure: R7 is selected from the group consisting of −CH(R8)O−, −CH2−, −CH2N(R8)−, and − CH(R8)O−C(=O)−, where R8 is selected from H, C1-C6 alkyl, C(=O)−C1-C6 alkyl, and − C(=O)N(R9)2, where R9 is independently selected from the group consisting of H, C1-C12 alkyl, and −(CH2CH2O)n−(CH2)m−OH, where m is an integer from 1 to 5, and n is an integer from 2 to 50, or two R9 groups together form a 5- or 6-membered heterocyclyl ring; and z is 0 or 1. 36. The immunoconjugate of claim 35 wherein AA1 is methyl, AA2 forms proline, and AA3 is isopropyl. 37. The immunoconjugate of claim 35 wherein EsPEP has the formula: . 38. The immunoconjugate of claim 37 wherein EsPEP is selected from the formulas: . 39. The immunoconjugate of claim 29 wherein L is: −C(=O)−(PEG)−C(=O)−(EsPEP)− . 40. The immunoconjugate of claim 39 wherein PEG is: −(CH2CH2O)25−(CH2)2− or −(CH2CH2O)10−(CH2)2− . 41. The immunoconjugate of claim 40 selected from the formulas: . 42. The immunoconjugate of claim 41 wherein Ims has the structure: and the wavy line indicates the site of attachment. 43. The immunoconjugate of claim 29 wherein EsPEP is a tetrapeptide having the formula: where AA1, AA2, AA3 and AA4 are independently selected from a natural or unnatural amino acid, and the wavy line indicates a point of attachment; Cyc is selected from C6-C20 aryldiyl and C1-C20 heteroaryldiyl, optionally substituted with one or more groups selected from F, Cl, NO2, −OH, −OCH3, and a glucuronic acid having the structure: R7 is selected from the group consisting of −CH(R8)O−, −CH2−, −CH2N(R8)−, and − CH(R8)O−C(=O)−, where R8 is selected from H, C1-C6 alkyl, C(=O)−C1-C6 alkyl, and − C(=O)N(R9)2, where R9 is independently selected from the group consisting of H, C1-C12 alkyl, and −(CH2CH2O)n−(CH2)m−OH, where m is an integer from 1 to 5, and n is an integer from 2 to 50, or two R9 groups together form a 5- or 6-membered heterocyclyl ring; and z is 0 or 1. 44. The immunoconjugate of claim 43 wherein AA1 is selected from the group consisting of Abu, Ala, and Val; AA2 is selected from the group consisting of Nle(O-Bzl), Oic and Pro; AA3 is selected from the group consisting of Ala and Met(O)2; and AA4 is selected from the group consisting of Oic, Arg(NO2), Bpa, and Nle(O-Bzl). 45. The immunoconjugate of claim 44 wherein EsPEP has the formula: . 46. The immunoconjugate of claim 45 wherein EsPEP has the formula: . 47. The immunoconjugate of claim 29 comprising a structure selected from IIe-h: where the wavy line indicates the attachment through L to the antibody. 48. The immunoconjugate of claim 29 comprising a structure selected from IIi-l: where the wavy line indicates the attachment through L to the antibody. 49. The immunoconjugate of claim 48 wherein R2 and R3 are each C1-C8 alkyl. 50. The immunoconjugate of claim 49 wherein R2 and R3 are each −CH2CH2CH3. 51. The immunoconjugate of claim 48 wherein X2 and X3 are each a bond, and R2 or R3 is −O−(C1-C12 alkyl). 52. The immunoconjugate of claim 51 wherein R2 or R3 is −OCH2CH3. 53. The immunoconjugate of any one of claims 2-12 wherein the elastase-substrate, peptide linker is cleaved by elastase. 54. An immunostimulant-elastase substrate, peptide linker compound selected from formulas IIa-f: wherein R1, R2, R3, R4, and R5 are independently selected from the group consisting of H, C1-C12 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C12 carbocyclyl, C6-C20 aryl, C2-C9 heterocyclyl, and C1-C20 heteroaryl, where alkyl, alkenyl, alkynyl, carbocyclyl, aryl, heterocyclyl, and heteroaryl are independently and optionally substituted with one or more groups selected from: −(C1-C12 alkyldiyl)−N(R6)−*; −(C1-C12 alkyldiyl)−N(R6)2; −(C1-C12 alkyldiyl)−OR6; −(C3-C12 carbocyclyl); −(C3-C12 carbocyclyl)−*; −(C3-C12 carbocyclyl)−(C1-C12 alkyldiyl)−NR6−*; −(C3-C12 carbocyclyl)−(C1-C12 alkyldiyl)−N(R6)2; −(C3-C12 carbocyclyl)−NR5−C(=NR6a)NR6−*; −(C6-C20 aryl); −(C6-C20 aryldiyl)−*; −(C6-C20 aryldiyl)−N(R6)−*; −(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−N(R6)−*; −(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−(C2-C20 heterocyclyldiyl)−*; −(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−N(R6)2; −(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−NR6−C(=NR6a)N(R6)−*; −(C2-C20 heterocyclyl); −(C2-C20 heterocyclyl)−*; −(C2-C9 heterocyclyl)−(C1-C12 alkyldiyl)−NR6−*; −(C2-C9 heterocyclyl)−(C1-C12 alkyldiyl)−N(R6)2; −(C2-C9 heterocyclyl)−C(=O)−(C1-C12 alkyldiyl)−N(R6)−*; −(C2-C9 heterocyclyl)−NR5−C(=NR6a)NR6−*; −(C2-C9 heterocyclyl)−NR6−(C6-C20 aryldiyl)−(C1-C12 alkyldiyl)−N(R6)−*; −(C2-C9 heterocyclyl)−(C6-C20 aryldiyl)−*; −(C1-C20 heteroaryl); −(C1-C20 heteroaryldiyl)−*; −(C1-C20 heteroaryldiyl)−(C1-C12 alkyldiyl)−N(R6)−*; −(C1-C20 heteroaryldiyl)−(C1-C12 alkyldiyl)−N(R6)2; −(C1-C20 heteroaryldiyl)−NR6−C(=NR6a)N(R6)−*; −(C1-C20 heteroaryldiyl)−N(R6)C(=O)−(C1-C12 alkyldiyl)−N(R6)−*; −C(=O)−*; −C(=O)−(C1-C12 alkyldiyl)−N(R6)−*; −C(=O)−(C2-C20 heterocyclyldiyl)−*; −C(=O)N(R6)2; −C(=O)N(R6)−*; −C(=O)N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)R5; −C(=O)N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)N(R6)2; −C(=O)NR6−(C1-C12 alkyldiyl)−N(R6)CO2R6; −C(=O)NR6−(C1-C12 alkyldiyl)−N(R6)C(=NR6a)N(R6)2; −C(=O)NR6−(C1-C12 alkyldiyl)−NR6C(=NR6a)R6; −C(=O)NR6−(C1-C8 alkyldiyl)−NR6(C2-C5 heteroaryl); −C(=O)NR6−(C1-C20 heteroaryldiyl)−N(R6)−*; −C(=O)NR6−(C1-C20 heteroaryldiyl)−*; −C(=O)NR6−(C1-C20 heteroaryldiyl)−(C1-C12 alkyldiyl)−N(R6)2; −C(=O)NR6−(C1-C20 heteroaryldiyl)−(C2-C20 heterocyclyldiyl)−C(=O)NR6−(C1-C12 alkyldiyl)−NR6−*; −N(R6)2; −N(R6)−*; −N(R6)C(=O)R6; −N(R6)C(=O)−*; −N(R6)C(=O)N(R6)2; −N(R6)C(=O)N(R6)−*; −N(R6)CO2R6; −N(R6)CO2(R6)−*; −NR6C(=NR6a)N(R6)2; −NR6C(=NR6a)N(R6)−*; −NR6C(=NR6a)R6; −N(R6)C(=O)−(C1-C12 alkyldiyl)−N(R6)−*; −N(R6)−(C2-C5 heteroaryl); −N(R6)−S(=O)2−(C1-C12 alkyl); −O−(C1-C12 alkyl); −O−(C1-C12 alkyldiyl)−N(R6)2; −O−(C1-C12 alkyldiyl)−N(R6)−*; −OC(=O)N(R6)2; −OC(=O)N(R6)−*; −S(=O)2−(C2-C20 heterocyclyldiyl)−*; −S(=O)2−(C2-C20 heterocyclyldiyl)−(C1-C12 alkyldiyl)−N(R6)2; −S(=O)2−(C2-C20 heterocyclyldiyl)−(C1-C12 alkyldiyl)−NR6−*; and −S(=O)2−(C2-C20 heterocyclyldiyl)−(C1-C12 alkyldiyl)−OH; or R2 and R3 of formulas Ic or Id together form a 5- or 6-membered heterocyclyl ring; X1, X2, X3, X4, and X5 are independently selected from the group consisting of a bond, C(=O), C(=O)N(R6), O, N(R6), S, S(O)2, and S(O)2N(R6); R6 is selected from the group consisting of H, C6-C20 aryl, C6-C20 aryldiyl, C1-C12 alkyl, and C1-C12 alkyldiyl, or two R6 groups together form a 5- or 6-membered heterocyclyl ring; R6a is selected from the group consisting of C6-C20 aryl and C1-C20 heteroaryl; where the asterisk * indicates the attachment site of L, and where one of R1, R2, R3, R4 and R5 is attached to L; L is selected from the group consisting of: Q−C(=O)−(PEG)−C(=O)−(EsPEP)−; Q−C(=O)−(PEG)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−; Q−C(=O)−(PEG)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)−(C2-C5 monoheterocyclyldiyl)−; Q−C(=O)−(PEG)−N(R6)−(PEG)−C(=O)−(EsPEP)−; Q−C(=O)−(PEG)−N+(R6)2−(PEG)−C(=O)−(EsPEP)−; Q−C(=O)−(PEG)−C(=O)−N(R6)CH(AA1)C(=O)−(PEG)−C(=O)−(EsPEP)−; Q−C(=O)−(C1-C12 alkyldiyl)−C(=O)−(EsPEP)−; Q−C(=O)−(C1-C12 alkyldiyl)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−; Q−C(=O)−(C1-C12 alkyldiyl)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−N(R6)−C(=O); Q−C(=O)−(C1-C12 alkyldiyl)−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)− (C2-C5 monoheterocyclyldiyl)−; Q−(CH2)m−C(=O)N(R6)−PEG−C(=O)−(EsPEP)−; Q−(CH2)m−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−; Q−(CH2)m−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)N(R6)C(=O)−; and Q−(CH2)m−C(=O)−(EsPEP)−N(R6)−(C1-C12 alkyldiyl)−N(R6)C(=O)−(C2-C5 monoheterocyclyldiyl)−; PEG has the formula: −(CH2CH2O)n−(CH2)m−; m is an integer from 1 to 5, and n is an integer from 2 to 50; EsPEP is an elastase-substrate, peptide linker unit comprising 2 to 12 amino acid residues; and Q is selected from the group consisting of N-hydroxysuccinimidyl, N- hydroxysulfosuccinimidyl, maleimide, and phenoxy substituted with one or more groups independently selected from F, Cl, NO2, and SO3-; alkyl, alkyldiyl, alkenyl, alkenyldiyl, alkynyl, alkynyldiyl, aryl, aryldiyl, carbocyclyl, carbocyclyldiyl, heterocyclyl, heterocyclyldiyl, heteroaryl, and heteroaryldiyl are independently and optionally substituted with one or more groups independently selected from F, Cl, Br, I, − CN, −CH3, −CH2CH3, −CH=CH2, −C ≡CH, −C ≡CCH3, −CH2CH2CH3, −CH(CH3)2, − CH2CH(CH3)2, −CH2OH, −CH2OCH3, −CH2CH2OH, −C(CH3)2OH, −CH(OH)CH(CH3)2, − C(CH3)2CH2OH, −CH2CH2SO2CH3, −CH2OP(O)(OH)2, −CH2F, −CHF2, −CF3, −CH2CF3, − CH2CHF2, −CH(CH3)CN, −C(CH3)2CN, −CH2CN, −CH2NH2, −CH2NHSO2CH3, −CH2NHCH3, −CH2N(CH3)2, −CO2H, −COCH3, −CO2CH3, −CO2C(CH3)3, −COCH(OH)CH3, −CONH2, − CONHCH3, −CON(CH3)2, −C(CH3)2CONH2, −NH2, −NHCH3, −N(CH3)2, −NHCOCH3, − N(CH3)COCH3, −NHS(O)2CH3, −N(CH3)C(CH3)2CONH2, −N(CH3)CH2CH2S(O)2CH3, − NHC(=NH)H, −NHC(=NH)CH3, −NHC(=NH)NH2, −NHC(=O)NH2, −NO2, =O, −OH, −OCH3, −OCH2CH3, −OCH2CH2OCH3, −OCH2CH2OH, −OCH2CH2N(CH3)2, −O(CH2CH2O)n− (CH2)mCO2H, −O(CH2CH2O)nH, −OP(O)(OH)2, −S(O)2N(CH3)2, −SCH3, −S(O)2CH3, and − S(O)3H. 55. The immunostimulant-elastase substrate, peptide linker compound of claim 54 wherein PEG has the formula: −(CH2CH2O)25−(CH2)2− or −(CH2CH2O)10−(CH2)2−. 56. The immunostimulant-elastase substrate, peptide linker compound of claim 54 wherein R2 and R3 are each C1-C8 alkyl. 57. The immunostimulant-elastase substrate, peptide linker compound of claim 56 wherein R2 and R3 are each −CH2CH2CH3. 58. The immunostimulant-elastase substrate, peptide linker compound of claim 54 wherein X2 and X3 are each a bond, and R2 or R3 is −O−(C1-C12 alkyl). 59. The immunostimulant-elastase substrate, peptide linker compound of claim 58 wherein R2 or R3 is −OCH2CH3. 60. The immunostimulant-elastase substrate, peptide linker compound of claim 54 is selected from the formulas: where TFP is 2,3,5,6-tetrafluorophenoxy. 61. The immunostimulant-elastase substrate, peptide linker compound of claim 54 comprising a structure selected from IIe-h: where the wavy line indicates the attachment through L to the antibody. 62. The immunostimulant-elastase substrate, peptide linker compound of claim 54 comprising a structure selected from IIi-l: where the wavy line indicates the attachment through L to the antibody. 63. An immunoconjugate prepared by conjugation of a cell-binding agent with an immunostimulant-elastase substrate, peptide linker compound of claim 54. 64. An immunoconjugate prepared by conjugation of a cell-binding agent with an immunostimulant-elastase substrate, peptide linker compound having the structure: . 65. The immunoconjugate of claims 63 or 64 wherein the cell-binding agent is an antibody. 66. A pharmaceutical composition comprising a therapeutically effective amount of an immunoconjugate according to any one of claims 1-53 and one or more pharmaceutically acceptable diluent, vehicle, carrier or excipient. 67. The pharmaceutical composition of claim 66 for use in therapy. 68. A method of treatment comprising administering a therapeutically-effective dose of the pharmaceutical composition of claim 66 to a patient with an immune-related disorder. 69. The method of claim 68 wherein the elastase-substrate, peptide linker of the immunoconjugate is cleaved by elastase. 70. An immunoconjugate of any one of claims 1 to 53 for use in therapy. 71. A method for treating cancer comprising administering a therapeutically effective amount of the pharmaceutical composition according to claim 66 to a patient in need thereof. 72. The method of claim 71, wherein the cancer is susceptible to a pro-inflammatory response induced by TLR7 and/or TLR8 agonism. 73. The method of claim 71, wherein the cancer is susceptible to a pro-inflammatory response induced by STING agonism. 74. The method of claim 71, wherein the cancer is a PD-L1-expressing cancer. 75. The method of claim 71, wherein the cancer is a HER2-expressing cancer. 76. The method of claim 71, wherein the cancer is a CEA-expressing cancer. 77. The method of claim 71, wherein the cancer is a Trop2-expressing cancer. 78. The method of any one of claims 72-77, wherein the cancer is selected from bladder cancer, urinary tract cancer, urothelial carcinoma, lung cancer, non-small cell lung cancer, Merkel cell carcinoma, colon cancer, colorectal cancer, gastric cancer, and breast cancer. 79. The method of claim 78, wherein the breast cancer is triple-negative breast cancer. 80. The method of claim 78, wherein the Merkel cell carcinoma cancer is metastatic Merkel cell carcinoma. 81. The method of claim 78, wherein the gastric cancer is HER2 overexpressing gastric cancer. 82. The method of claim 78, wherein the cancer is gastroesophageal junction adenocarcinoma. 83. Use of an immunoconjugate according to any one of claims 1-53 for treating cancer. |
wherein substituents X 1-4 and R 1-4 are described herein. An exemplary amino-benzazepine immunostimulatory TLR moiety has the structure: where the wavy line indicates the site of attachment to the elastase-substrate, peptide linker, L. ELASTASE-SUBSTRATE, PEPTIDE LINKERS The invention includes a linking unit, i.e. L or linker, between the cell-binding agent and the immunostimulatory moiety. The linker is a peptide radical based on a linear sequence of specific amino acid residues which can be selectively cleaved by a tumor-associated elastase enzyme or an enzyme with elastase-like activity. The peptide radical may be about two to about twelve amino acids. Cleavage of a bond within the elastase-substrate, peptide linker by elastase releases an active form of the immunostimulatory moiety. This leads to an increase in the tissue specificity of the conjugates according to the invention and thus to an additional decrease of toxicity of the conjugates according to the invention in other tissue types. The linker provides sufficient stability of the immunoconjugate in biological media, e.g. culture medium or serum and, at the same time, the desired intracellular action within tumor tissue as a result of its specific enzymatic or hydrolytic cleavability with release of the immunostimulatory moiety, i.e. “payload”. The enzymatic activity of elastase can catalyze cleavage of a covalent bond of the immunoconjugate under physiological conditions. The enzymatic activity being the expression product of cells associated with tumor tissue. The enzymatic activity on the cleavage site of the targeting peptide converts the immunoconjugate to an active immunostimulatory drug free of targeting peptide and linking group. The cleavage site may be specifically recognized by elastase. Elastase may catalyze the cleavage of a specific peptidic bond between the C-terminal amino acid residue of the specific peptide and the immunostimulatory moiety of the immunoconjugate. Specific cleavage of the immunoconjugates of the invention takes advantage of the presence of tumor infiltrating cells of the immune system and leukocyte-secreted enzymes, to promote the activation of an anticancer drug at the tumor site. In one embodiment, the elastase-substrate, peptide linker (EsPEP) has the formula: where AA is independently selected from a natural or unnatural amino acid side chain, or one or more of AA, and an adjacent nitrogen atom form a 5-membered ring proline amino acid, and the wavy line indicates a point of attachment; R 7 is selected from the group consisting of C 6 -C 20 aryldiyl and C 1 -C 20 heteroaryldiyl, substituted with a group selected from −CH 2 O−C(=O)−, −CH 2 O−, −CH 2 −, −CH 2 N(R 8 )−, and − CH(R 8 )O−C(=O)−, where R 8 is H or C 1 -C 6 alkyl; y is an integer from 2 to 12; and z is 0 or 1. In exemplary embodiments, EsPEP is a tripeptide and has the formulas: or EsPEP is a tetrapeptide and has the formulas: where AA1, AA2 and AA3 are independently selected from a natural or unnatural amino acid side chain, or one or more of AA 1 , AA 2 , AA 3 , AA 4 , and an adjacent nitrogen atom form a 5-membered ring proline amino acid, and the wavy line indicates a point of attachment. Cyc is selected from C 6 - C 20 aryldiyl and C 1 -C 20 heteroaryldiyl, optionally substituted with one or more groups selected from F, Cl, NO 2 , −OH, −OCH 3 , and a glucuronic acid having the structure: R 7 is selected from the group consisting of −CH(R 8 )O−, −CH 2 −, −CH 2 N(R 8 )−, and − CH(R 8 )O−C(=O)−, where R 8 is selected from H, C 1 -C 6 alkyl, C(=O)−C 1 -C 6 alkyl, and − C(=O)N(R 9 ) 2 , where R 9 is independently selected from the group consisting of H, C 1 -C 12 alkyl, and −(CH 2 CH 2 O)n−(CH 2 )m−OH, where m is an integer from 1 to 5, and n is an integer from 2 to 50, or two R 9 groups together form a 5- or 6-membered heterocyclyl ring; z is 0 or 1. In an exemplary embodiment, EsPEP is a tripeptide wherein AA 1 is methyl, AA 2 forms proline, and AA3 is isopropyl. In an exemplary embodiment, EsPEP is a tetrapeptide wherein AA 1 is selected from the group consisting of Abu, Ala, and Val; AA2 is selected from the group consisting of Nle(O-Bzl), Oic and Pro; AA3 is selected from the group consisting of Ala and Met(O) 2 ; and AA 4 is selected from the group consisting of Oic, Arg(NO 2 ), Bpa, and Nle(O-Bzl). In an exemplary embodiment, EsPEP is comprised of amino acid residues of amino acids selected from the group consisting of: Ala D-Ala Met(O 2 ) . In an exemplary embodiment, EsPEP is selected from the group consisting of Ala-Pro- Val, Asn-Pro-Val, Ala-Ala-Val, Ala-Ala-Pro-Ala (SEQ ID NO: 639), Ala-Ala-Pro-Val (SEQ ID NO: 640), and Ala-Ala-Pro-Nva (SEQ ID NO: 641). In an exemplary embodiment, EsPEP has the formula: . In an exemplary embodiment, EsPEP has the formula: . IMMUNOSTIMULANT-ELASTASE SUBSTRATE, PEPTIDE LINKER COMPOUNDS The immunoconjugates of the invention are prepared by conjugation of a cell-binding agent with an immunostimulant-elastase substrate, peptide linker compound. Immunostimulant- elastase substrate, peptide linker compounds comprise an immunostimulatory moiety covalently attached to a linker unit. The linker units comprise an elastase-substrate, peptide unit and functional groups and subunits which affect stability, permeability, solubility, and other pharmacokinetic, safety, and efficacy properties of the immunoconjugates. The linker unit includes a reactive functional group which reacts, i.e. conjugates, with a reactive functional group of the antibody. For example, a nucleophilic group such as a lysine side chain amino of the antibody reacts with an electrophilic reactive functional group of the immunostimulant- elastase substrate, peptide linker compound to form the immunoconjugate. A cysteine thiol of the cell-binding agent, e.g. an antibody, reacts with a maleimide or bromoacetamide group of the immunostimulant-elastase substrate, peptide linker compound to form the immunoconjugate. Electrophilic reactive functional groups suitable for the immunostimulant-elastase substrate, peptide linker compounds include, but are not limited to, N-hydroxysuccinimidyl (NHS) esters and N-hydroxysulfosuccinimidyl (sulfo-NHS) esters (amine reactive); carbodiimides (amine and carboxyl reactive); hydroxymethyl phosphines (amine reactive); maleimides (thiol reactive); halogenated acetamides such as N-iodoacetamides (thiol reactive); aryl azides (primary amine reactive); fluorinated aryl azides (reactive via carbon-hydrogen (C- H) insertion); pentafluorophenyl (PFP) esters (amine reactive); tetrafluorophenyl (TFP) esters (amine reactive); tetrafluorophenyl, sulfonate (sulfo-TFP) esters, imidoesters (amine reactive); isocyanates (hydroxyl reactive); vinyl sulfones (thiol, amine, and hydroxyl reactive); pyridyl disulfides (thiol reactive); and benzophenone derivatives (reactive via C-H bond insertion). Further reagents include, but are not limited, to those described in Hermanson, Bioconjugate Techniques 2 nd Edition, Academic Press, 2008. The invention provides solutions to the limitations and challenges to the design, preparation and use of immunoconjugates. Some linkers may be labile in the blood stream, thereby releasing unacceptable amounts of the adjuvant/drug prior to internalization in a target cell (Khot, A. et al (2015) Bioanalysis 7(13):1633–1648). Other linkers may provide stability in the bloodstream, but intracellular release effectiveness may be negatively impacted. Linkers that provide for desired intracellular release typically have poor stability in the bloodstream. Alternatively stated, bloodstream stability and intracellular release are typically inversely related. In addition, in standard conjugation processes, the amount of adjuvant/drug moiety loaded on the antibody, i.e. drug loading, the amount of aggregate that is formed in the conjugation reaction, and the yield of final purified conjugate that can be obtained are interrelated. For example, aggregate formation is generally positively correlated to the number of equivalents of adjuvant/drug moiety and derivatives thereof conjugated to the antibody. Under high drug loading, formed aggregates must be removed for therapeutic applications. As a result, drug loading-mediated aggregate formation decreases immunoconjugate yield and can render process scale-up difficult. Exemplary embodiments of immunostimulant-elastase substrate, peptide linker compounds are selected from formulas IIa-f:
wherein R 1 , R 2 , R 3 , R 4 , and R 5 are independently selected from the group consisting of H, C 1 -C 12 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C 3 -C 12 carbocyclyl, C 6 -C 20 aryl, C 2 -C 9 heterocyclyl, and C 1 -C 20 heteroaryl, where alkyl, alkenyl, alkynyl, carbocyclyl, aryl, heterocyclyl, and heteroaryl are independently and optionally substituted with one or more groups selected from: −(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 1 -C 12 alkyldiyl)−OR 6 ; −(C 3 -C 12 carbocyclyl); −(C 3 -C 12 carbocyclyl)−*; −(C 3 -C 12 carbocyclyl)−(C 1 -C 12 alkyldiyl)−NR 6 −*; −(C 3 -C 12 carbocyclyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 3 -C 12 carbocyclyl)−NR 5 −C(=NR 6a )NR 6 −*; −(C 6 -C 20 aryl); −(C 6 -C 20 aryldiyl)−*; −(C 6 -C 20 aryldiyl)−N(R 6 )−*; −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−(C 2 -C 20 heterocyclyldiyl)−*; −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−NR 6 −C(=NR 6a )N(R 6 )−*; −(C 2 -C 20 heterocyclyl); −(C 2 -C 20 heterocyclyl)−*; −(C 2 -C 9 heterocyclyl)−(C 1 -C 12 alkyldiyl)−NR 6 −*; −(C 2 -C 9 heterocyclyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 2 -C 9 heterocyclyl)−C(=O)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 2 -C 9 heterocyclyl)−NR 5 −C(=NR 6a )NR 6 −*; −(C 2 -C 9 heterocyclyl)−NR 6 −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 2 -C 9 heterocyclyl)−(C 6 -C 20 aryldiyl)−*; −(C 1 -C 20 heteroaryl); −(C 1 -C 20 heteroaryldiyl)−*; −(C 1 -C 20 heteroaryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 1 -C 20 heteroaryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 1 -C 20 heteroaryldiyl)−NR 6 −C(=NR 6a )N(R 6 )−*; −(C 1 -C 20 heteroaryldiyl)−N(R 6 )C(=O)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −C(=O)−*; −C(=O)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −C(=O)−(C 2 -C 20 heterocyclyldiyl)−*; −C(=O)N(R 6 ) 2 ; −C(=O)N(R 6 )−*; −C(=O)N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)R 5 ; −C(=O)N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)N(R 6 ) 2 ; −C(=O)NR 6 −(C 1 -C 12 alkyldiyl)−N(R 6 )CO 2 R 6 ; −C(=O)NR 6 −(C 1 -C 12 alkyldiyl)−N(R 6 )C(=NR 6a )N(R 6 ) 2 ; −C(=O)NR 6 −(C 1 -C 12 alkyldiyl)−NR 6 C(=NR 6a )R 6 ; −C(=O)NR 6 −(C 1 -C8 alkyldiyl)−NR 6 (C 2 -C5 heteroaryl); −C(=O)NR 6 −(C 1 -C 20 heteroaryldiyl)−N(R 6 )−*; −C(=O)NR 6 −(C 1 -C 20 heteroaryldiyl)−*; −C(=O)NR 6 −(C 1 -C 20 heteroaryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −C(=O)NR 6 −(C 1 -C 20 heteroaryldiyl)−(C 2 -C 20 heterocyclyldiyl)−C(=O)NR 6 −(C 1 -C 12 alkyldiyl)−NR 6 −*; −N(R 6 ) 2 ; −N(R 6 )−*; −N(R 6 )C(=O)R 6 ; −N(R 6 )C(=O)−*; −N(R 6 )C(=O)N(R 6 ) 2 ; −N(R 6 )C(=O)N(R 6 )−*; −N(R 6 )CO 2 R 6 ; −N(R 6 )CO 2 (R 6 )−*; −NR 6 C(=NR 6a )N(R 6 ) 2 ; −NR 6 C(=NR 6a )N(R 6 )−*; −NR 6 C(=NR 6a )R 6 ; −N(R 6 )C(=O)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −N(R 6 )−(C 2 -C 5 heteroaryl); −N(R 6 )−S(=O) 2 −(C 1 -C 12 alkyl); −O−(C 1 -C 12 alkyl); −O−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −O−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −OC(=O)N(R 6 ) 2 ; −OC(=O)N(R 6 )−*; −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−*; −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−(C 1 -C 12 alkyldiyl)−NR 6 −*; and −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−(C 1 -C 12 alkyldiyl)−OH; or R 2 and R 3 of formulas Ic or Id together form a 5- or 6-membered heterocyclyl ring; X 1 , X 2 , X 3 , X 4 , and X 5 are independently selected from the group consisting of a bond, C(=O), C(=O)N(R 6 ), O, N(R 6 ), S, S(O) 2 , and S(O) 2 N(R 6 ); R 6 is selected from the group consisting of H, C 6 -C 20 aryl, C 6 -C 20 aryldiyl, C 1 -C 12 alkyl, and C 1 -C 12 alkyldiyl, or two R 6 groups together form a 5- or 6-membered heterocyclyl ring; R 6a is selected from the group consisting of C 6 -C 20 aryl and C 1 -C 20 heteroaryl; where the asterisk * indicates the attachment site of L, and where one of R 1 , R 2 , R 3 , R 4 and R 5 is attached to L; L is selected from the group consisting of: Q−C(=O)−(PEG)−C(=O)−(EsPEP)−; Q−C(=O)−(PEG)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−; Q−C(=O)−(PEG)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)−(C 2 -C 5 monoheterocyclyldiyl)−; Q−C(=O)−(PEG)−N(R 6 )−(PEG)−C(=O)−(EsPEP)−; Q−C(=O)−(PEG)−N + (R 6 ) 2 −(PEG)−C(=O)−(EsPEP)−; Q−C(=O)−(PEG)−C(=O)−N(R 6 )CH(AA1)C(=O)−(PEG)−C(=O)−(EsPEP)−; Q−C(=O)−(C 1 -C 12 alkyldiyl)−C(=O)−(EsPEP)−; Q−C(=O)−(C 1 -C 12 alkyldiyl)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−; Q−C(=O)−(C 1 -C 12 alkyldiyl)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )−C(=O); Q−C(=O)−(C 1 -C 12 alkyldiyl)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)− (C 2 -C 5 monoheterocyclyldiyl)−; Q−(CH 2 ) m −C(=O)N(R 6 )−PEG−C(=O)−(EsPEP)−; Q−(CH 2 ) m −C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−; Q−(CH 2 ) m −C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)N(R 6 )C(=O)−; and Q−(CH 2 ) m −C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)−(C 2 -C 5 monoheterocyclyldiyl)−; PEG has the formula: −(CH 2 CH 2 O) n −(CH 2 ) m −; m is an integer from 1 to 5, and n is an integer from 2 to 50; EsPEP is an elastase-substrate, peptide linker unit comprising 2 to 12 amino acid residues; and Q is selected from the group consisting of N-hydroxysuccinimidyl, N- hydroxysulfosuccinimidyl, maleimide, and phenoxy substituted with one or more groups independently selected from F, Cl, NO 2 , and SO 3 -; alkyl, alkyldiyl, alkenyl, alkenyldiyl, alkynyl, alkynyldiyl, aryl, aryldiyl, carbocyclyl, carbocyclyldiyl, heterocyclyl, heterocyclyldiyl, heteroaryl, and heteroaryldiyl are independently and optionally substituted with one or more groups independently selected from F, Cl, Br, I, − CN, −CH 3 , −CH 2 CH 3 , −CH=CH 2 , −C ≡CH, −C ≡CCH 3 , −CH 2 CH 2 CH 3 , −CH(CH 3 ) 2 , − CH 2 CH(CH 3 ) 2 , −CH 2 OH, −CH 2 OCH 3 , −CH 2 CH 2 OH, −C(CH 3 ) 2 OH, −CH(OH)CH(CH 3 ) 2 , − C(CH 3 ) 2 CH 2 OH, −CH 2 CH 2 SO 2 CH 3 , −CH 2 OP(O)(OH) 2 , −CH 2 F, −CHF2, −CF3, −CH 2 CF3, − CH 2 CHF2, −CH(CH 3 )CN, −C(CH 3 ) 2 CN, −CH 2 CN, −CH 2 NH 2 , −CH 2 NHSO 2 CH 3 , −CH 2 NHCH 3 , −CH 2 N(CH 3 ) 2 , −CO 2 H, −COCH 3 , −CO 2 CH 3 , −CO 2 C(CH 3 )3, −COCH(OH)CH 3 , −CONH 2 , − CONHCH 3 , −CON(CH 3 ) 2 , −C(CH 3 ) 2 CONH 2 , −NH 2 , −NHCH 3 , −N(CH 3 ) 2 , −NHCOCH 3 , − N(CH 3 )COCH 3 , −NHS(O) 2 CH 3 , −N(CH 3 )C(CH 3 ) 2 CONH 2 , −N(CH 3 )CH 2 CH 2 S(O) 2 CH 3 , − NHC(=NH)H, −NHC(=NH)CH 3 , −NHC(=NH)NH 2 , −NHC(=O)NH 2 , −NO 2 , =O, −OH, −OCH 3 , −OCH 2 CH 3 , −OCH 2 CH 2 OCH 3 , −OCH 2 CH 2 OH, −OCH 2 CH 2 N(CH 3 ) 2 , −O(CH 2 CH 2 O)n− (CH 2 )mCO 2 H, −O(CH 2 CH 2 O)nH, −OP(O)(OH) 2 , −S(O) 2 N(CH 3 ) 2 , −SCH 3 , −S(O) 2 CH 3 , and − S(O)3H. An exemplary embodiment of an immunostimulant-elastase substrate, peptide linker compound has the formula: ; where TFP is 2,3,5,6-tetrafluorophenoxy. An exemplary embodiment of an immunostimulant-elastase substrate, peptide linker compound includes wherein PEG has the formula: −(CH 2 CH 2 O) 25 −(CH 2 ) 2 − or −(CH 2 CH 2 O) 10 − (CH 2 ) 2 −. Exemplary embodiments of immunostimulant-elastase substrate, peptide linker compounds comprise a structure selected from IIe-h:
where the wavy line indicates the attachment through L to the antibody. Exemplary embodiments of immunostimulant-elastase substrate, peptide linker compounds comprise a structure selected from IIi-l: where the wavy line indicates the attachment through L to the antibody. The invention includes all reasonable combinations, and permutations of the features, of the Formula II embodiments. An exemplary embodiment of the immunostimulant-elastase substrate, peptide linker compound of Formula II is selected from Tables 1a and 1b. Each compound was synthesized and purified by the methods in the Examples provided herein, characterized by mass spectrometry, and shown to have the mass indicated. When conjugated to an antibody, the compounds of Tables 1a and 1b demonstrate surprising and unexpected properties which may predict useful therapeutic activity to treat cancer and other disorders. Table 1a: Immunostimulant-elastase substrate, peptide linker compound of Formula II and intermediates thereof
V
Table 1b: STING agonist immunostimulant-elastase substrate, peptide linker compound IMMUNOCONJUGATES Exemplary embodiments of immunoconjugates comprise a cell-binding agent covalently attached to one or more immunostimulatory moieties by an elastase-substrate, peptide linker, having the Formula I: Targeting properties of the cell-binding agent may drive the immunoconjugate accumulation at a tumor site, where the proinflammatory stimuli result in the recruitment of tumor-infiltrating leukocytes, such as neutrophils. The activation of the latter promotes the release of elastase, which triggers the payload release in the tumor microenvironment. This mode of activation may possess potential therapeutic benefits, since the free payload would diffuse in the tumor mass, and act against a large variety of cells (e.g., antigen-negative cancer cells, endothelial and other cancer-associated host cells) leading to a localized damage (Li, F. et al (2016) Cancer Res.76:2710-2719). Lipophilic immunostimulatory moieties may be most suited for this strategy, as the membrane permeability may facilitate biological activity by the so-called “bystander effect.” The invention includes an immunoconjugates comprising a cell-binding agent covalently attached to one or more immunostimulatory moieties by an elastase-substrate, peptide linker. An exemplary embodiment of the immunoconjugate includes wherein the cell-binding agent is an antibody. The antibody may be an antibody construct that has an antigen binding domain that binds PD-L1. The antibody may be selected from the group consisting of atezolizumab, durvalumab, and avelumab, or a biosimilar or a biobetter thereof. The antibody may be an antibody construct that has an antigen binding domain that binds HER2. The antibody may be selected from the group consisting of trastuzumab and pertuzumab, or a biosimilar or a biobetter thereof. The antibody may be an antibody construct that has an antigen binding domain that binds CEA. The antibody may be labetuzumab, or a biosimilar or a biobetter thereof. An exemplary embodiment of the immunoconjugate includes wherein the one or more immunostimulatory moieties is a pattern-recognition receptor. An exemplary embodiment of the immunoconjugate includes wherein the one or more immunostimulatory moieties interact with or modulate a receptor selected from the group consisting of TLR, STING, NOD2, RIG-1, and NLRP3. An exemplary embodiment of the immunoconjugate has Formula I: I or a pharmaceutically acceptable salt thereof, wherein: Ab is the antibody; L is the linker comprising an elastase-substrate, peptide linker unit; Ims is the immunostimulatory moiety; and p is an integer from 1 to 8. An exemplary embodiment of the immunoconjugate includes wherein Ims is selected from formulas Ia-f: Ia; wherein R 1 , R 2 , R 3 , R 4 , and R 5 are independently selected from the group consisting of H, C 1 -C 12 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C 3 -C 12 carbocyclyl, C 6 -C 20 aryl, C 2 -C 9 heterocyclyl, and C 1 -C 20 heteroaryl, where alkyl, alkenyl, alkynyl, carbocyclyl, aryl, heterocyclyl, and heteroaryl are independently and optionally substituted with one or more groups selected from: −(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 1 -C 12 alkyldiyl)−OR 6 ; −(C 3 -C 12 carbocyclyl); −(C 3 -C 12 carbocyclyl)−*; −(C 3 -C 12 carbocyclyl)−(C 1 -C 12 alkyldiyl)−NR 6 −*; −(C 3 -C 12 carbocyclyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 3 -C 12 carbocyclyl)−NR 5 −C(=NR 6a )NR 6 −*; −(C 6 -C 20 aryl); −(C 6 -C 20 aryldiyl)−*; −(C 6 -C 20 aryldiyl)−N(R 6 )−*; −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−(C 2 -C 20 heterocyclyldiyl)−*; −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−NR 6 −C(=NR 6a )N(R 6 )−*; −(C 2 -C 20 heterocyclyl); −(C 2 -C 20 heterocyclyl)−*; −(C 2 -C 9 heterocyclyl)−(C 1 -C 12 alkyldiyl)−NR 6 −*; −(C 2 -C 9 heterocyclyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 2 -C 9 heterocyclyl)−C(=O)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 2 -C 9 heterocyclyl)−NR 5 −C(=NR 6a )NR 6 −*; −(C 2 -C 9 heterocyclyl)−NR 6 −(C 6 -C 20 aryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 2 -C 9 heterocyclyl)−(C 6 -C 20 aryldiyl)−*; −(C 1 -C 20 heteroaryl); −(C 1 -C 20 heteroaryldiyl)−*; −(C 1 -C 20 heteroaryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −(C 1 -C 20 heteroaryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −(C 1 -C 20 heteroaryldiyl)−NR 6 −C(=NR 6a )N(R 6 )−*; −(C 1 -C 20 heteroaryldiyl)−N(R 6 )C(=O)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −C(=O)−*; −C(=O)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −C(=O)−(C 2 -C 20 heterocyclyldiyl)−*; −C(=O)N(R 6 ) 2 ; −C(=O)N(R 6 )−*; −C(=O)N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)R 5 ; −C(=O)N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)N(R 6 ) 2 ; −C(=O)NR 6 −(C 1 -C 12 alkyldiyl)−N(R 6 )CO 2 R 6 ; −C(=O)NR 6 −(C 1 -C 12 alkyldiyl)−N(R 6 )C(=NR 6a )N(R 6 ) 2 ; −C(=O)NR 6 −(C 1 -C 12 alkyldiyl)−NR 6 C(=NR 6a )R 6 ; −C(=O)NR 6 −(C 1 -C 8 alkyldiyl)−NR 6 (C 2 -C 5 heteroaryl); −C(=O)NR 6 −(C 1 -C 20 heteroaryldiyl)−N(R 6 )−*; −C(=O)NR 6 −(C 1 -C 20 heteroaryldiyl)−*; −C(=O)NR 6 −(C 1 -C 20 heteroaryldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −C(=O)NR 6 −(C 1 -C 20 heteroaryldiyl)−(C 2 -C 20 heterocyclyldiyl)−C(=O)NR 6 −(C 1 -C 12 alkyldiyl)−NR 6 −*; −N(R 6 ) 2 ; −N(R 6 )−*; −N(R 6 )C(=O)R 6 ; −N(R 6 )C(=O)−*; −N(R 6 )C(=O)N(R 6 ) 2 ; −N(R 6 )C(=O)N(R 6 )−*; −N(R 6 )CO 2 R 6 ; −N(R 6 )CO 2 (R 6 )−*; −NR 6 C(=NR 6a )N(R 6 ) 2 ; −NR 6 C(=NR 6a )N(R 6 )−*; −NR 6 C(=NR 6a )R 6 ; −N(R 6 )C(=O)−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −N(R 6 )−(C 2 -C 5 heteroaryl); −N(R 6 )−S(=O) 2 −(C 1 -C 12 alkyl); −O−(C 1 -C 12 alkyl); −O−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −O−(C 1 -C 12 alkyldiyl)−N(R 6 )−*; −OC(=O)N(R 6 ) 2 ; −OC(=O)N(R 6 )−*; −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−*; −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−(C 1 -C 12 alkyldiyl)−N(R 6 ) 2 ; −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−(C 1 -C 12 alkyldiyl)−NR 6 −*; and −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−(C 1 -C 12 alkyldiyl)−OH; or R 2 and R 3 of formulas Ic or Id together form a 5- or 6-membered heterocyclyl ring; X 1 , X 2 , X 3 , X 4 , and X 5 are independently selected from the group consisting of a bond, C(=O), C(=O)N(R 6 ), O, N(R 6 ), S, S(O) 2 , and S(O) 2 N(R 6 ); R 6 is selected from the group consisting of H, C 6 -C 20 aryl, C 6 -C 20 aryldiyl, C 1 -C 12 alkyl, and C 1 -C 12 alkyldiyl, or two R 6 groups together form a 5- or 6-membered heterocyclyl ring; R 6a is selected from the group consisting of C 6 -C 20 aryl and C 1 -C 20 heteroaryl; where the asterisk * indicates the attachment site of L, and where one of R 1 , R 2 , R 3 , R 4 and R 5 is attached to L; and alkyl, alkyldiyl, alkenyl, alkenyldiyl, alkynyl, alkynyldiyl, aryl, aryldiyl, carbocyclyl, carbocyclyldiyl, heterocyclyl, heterocyclyldiyl, heteroaryl, and heteroaryldiyl are independently and optionally substituted with one or more groups independently selected from F, Cl, Br, I, − CN, −CH 3 , −CH 2 CH 3 , −CH=CH 2 , −C ≡CH, −C ≡CCH 3 , −CH 2 CH 2 CH 3 , −CH(CH 3 ) 2 , − CH 2 CH(CH 3 ) 2 , −CH 2 OH, −CH 2 OCH 3 , −CH 2 CH 2 OH, −C(CH 3 ) 2 OH, −CH(OH)CH(CH 3 ) 2 , − C(CH 3 ) 2 CH 2 OH, −CH 2 CH 2 SO 2 CH 3 , −CH 2 OP(O)(OH) 2 , −CH 2 F, −CHF 2 , −CF 3 , −CH 2 CF 3 , − CH 2 CHF 2 , −CH(CH 3 )CN, −C(CH 3 ) 2 CN, −CH 2 CN, −CH 2 NH 2 , −CH 2 NHSO 2 CH 3 , −CH 2 NHCH 3 , −CH 2 N(CH 3 ) 2 , −CO 2 H, −COCH 3 , −CO 2 CH 3 , −CO 2 C(CH 3 ) 3 , −COCH(OH)CH 3 , −CONH 2 , − CONHCH 3 , −CON(CH 3 ) 2 , −C(CH 3 ) 2 CONH 2 , −NH 2 , −NHCH 3 , −N(CH 3 ) 2 , −NHCOCH 3 , − N(CH 3 )COCH 3 , −NHS(O) 2 CH 3 , −N(CH 3 )C(CH 3 ) 2 CONH 2 , −N(CH 3 )CH 2 CH 2 S(O) 2 CH 3 , − NHC(=NH)H, −NHC(=NH)CH 3 , −NHC(=NH)NH 2 , −NHC(=O)NH 2 , −NO 2 , =O, −OH, −OCH 3 , −OCH 2 CH 3 , −OCH 2 CH 2 OCH 3 , −OCH 2 CH 2 OH, −OCH 2 CH 2 N(CH 3 ) 2 , −O(CH 2 CH 2 O)n− (CH 2 )mCO 2 H, −O(CH 2 CH 2 O)nH, −OP(O)(OH) 2 , −S(O) 2 N(CH 3 ) 2 , −SCH 3 , −S(O) 2 CH 3 , and − S(O)3H. An exemplary embodiment of the immunoconjugate includes wherein one of R 1 , R 2 , R 3 , R 4 and R 5 is selected from the formulas:
. An exemplary embodiment of the immunoconjugate includes wherein Ims has formula wherein X a and X b are independently selected from a five-membered heteroaryl; R 1 is selected from the group consisting of F, Cl, Br, I, −CN, −OH, and −O−(C 1 -C 6 alkyldiyl). R 2a and R 2b are independently selected from −C(=O)N(R 5 ) 2 ; R 3 is selected from C 1 -C 6 alkyldiyl, −(C 1 -C 3 alkyldiyl)−O−(C 1 -C 3 alkyldiyl)−, C 2 -C 6 alkenyldiyl and C 2 -C 6 alkynyldiyl, optionally substituted with one or more groups selected from F, Cl, −OH, −OCH 3 , −OCH 2 CH 3 , −OCH 2 CH 2 OCH 3 , −OCH 2 CH 2 OH, −OCH 2 CH 2 N(CH 3 ) 2 ; R 4 is selected from the group consisting of: −(C 1 -C 12 alkyldiyl)−N(R 5 )−*; −(C 1 -C 12 alkyldiyl)−(C 2 -C 20 heterocyclyldiyl)−*; −O−(C 1 -C 12 alkyldiyl)−N(R 5 )−*; −O−(C 1 -C 12 alkyldiyl)−(C 2 -C 20 heterocyclyldiyl)−*; −O−(C 1 -C 12 alkyldiyl)−(C 2 -C 20 heterocyclyldiyl)−N(R 5 )−*; −OC(=O)N(R 5 )−*; −N(R 5 )−(C 1 -C 12 alkyldiyl)−N(R 5 )−*; −N(R 5 )−(C 1 -C 12 alkyldiyl)−(C 2 -C 20 heterocyclyldiyl)−*; −C(=O)N(R 5 )−*; −(C 2 -C 20 heterocyclyldiyl)−*; −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−*; −S(=O) 2 −(C 2 -C 20 heterocyclyldiyl)−(C 1 -C 12 alkyldiyl)−NR 5 −*; where the asterisk * indicates the attachment site of L; R 5 is independently H or C 1 -C 6 alkyl, or two R 5 groups together form a 5- or 6- membered heterocyclyl ring; and alkyl, alkyldiyl, alkenyl, alkenyldiyl, alkynyl, alkynyldiyl, aryl, aryldiyl, carbocyclyl, carbocyclyldiyl, heterocyclyl, heterocyclyldiyl, heteroaryl, and heteroaryldiyl are independently and optionally substituted with one or more groups independently selected from F, Cl, Br, I, − CN, −CH 3 , −CH 2 CH 3 , −CH=CH 2 , −C ≡CH, −C ≡CCH 3 , −CH 2 CH 2 CH 3 , −CH(CH 3 ) 2 , − CH 2 CH(CH 3 ) 2 , −CH 2 OH, −CH 2 OCH 3 , −CH 2 CH 2 OH, −C(CH 3 ) 2 OH, −CH(OH)CH(CH 3 ) 2 , − C(CH 3 ) 2 CH 2 OH, −CH 2 CH 2 SO 2 CH 3 , −CH 2 OP(O)(OH) 2 , −CH 2 F, −CHF2, −CF3, −CH 2 CF3, − CH 2 CHF2, −CH(CH 3 )CN, −C(CH 3 ) 2 CN, −CH 2 CN, −CH 2 NH 2 , −CH 2 NHSO 2 CH 3 , −CH 2 NHCH 3 , −CH 2 N(CH 3 ) 2 , −CO 2 H, −COCH 3 , −CO 2 CH 3 , −CO 2 C(CH 3 )3, −COCH(OH)CH 3 , −CONH 2 , − CONHCH 3 , −CON(CH 3 ) 2 , −C(CH 3 ) 2 CONH 2 , −NH 2 , −NHCH 3 , −N(CH 3 ) 2 , −NHCOCH 3 , − N(CH 3 )COCH 3 , −NHS(O) 2 CH 3 , −N(CH 3 )C(CH 3 ) 2 CONH 2 , −N(CH 3 )CH 2 CH 2 S(O) 2 CH 3 , − NHC(=NH)H, −NHC(=NH)CH 3 , −NHC(=NH)NH 2 , −NHC(=O)NH 2 , −NO 2 , =O, −OH, −OCH 3 , −OCH 2 CH 3 , −OCH 2 CH 2 OCH 3 , −OCH 2 CH 2 OH, −OCH 2 CH 2 N(CH 3 ) 2 , −O(CH 2 CH 2 O) n − (CH 2 ) m CO 2 H, −O(CH 2 CH 2 O) n H, −OCH 2 F, −OCHF 2 , −OCF 3 , −OP(O)(OH) 2 , −S(O) 2 N(CH 3 ) 2 , − SCH 3 , −S(O) 2 CH 3 , and −S(O) 3 H. An exemplary embodiment of formula Ig includes wherein wherein X a and X b are independently selected from the group consisting of imidazolyl, pyrazolyl, triazolyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxadiazolyl, oxazolyl, isothiazolyl, pyrrolyl, oxadiazolyl, and thiadiazolyl. An exemplary embodiment of formula Ig includes wherein wherein X a and X b are each pyrazolyl, substituted with one or more C 1 -C 12 alkyl groups. An exemplary embodiment of formula Ig includes wherein wherein R 1 is selected from the group consisting of −OCH 3 , −OCH 2 CH 3 , −OCH 2 CH 2 OCH 3 , −OCH 2 CH 2 OH, and − OCH 2 CH 2 N(CH 3 ) 2 An exemplary embodiment of formula Ig includes wherein R 1 is −OCH 3 . An exemplary embodiment of formula Ig includes wherein R 1 is F. An exemplary embodiment of formula Ig includes wherein R 2a and R 2b are each − C(=O)NH 2 . An exemplary embodiment of formula Ig includes wherein R 3 is selected from − CH 2 CH 2 −, −CH=CH−, and −C ≡C−. An exemplary embodiment of formula Ig includes wherein R 3 is C 2 -C 4 alkenyldiyl, substituted with one or more groups selected from F, −OH, and −OCH 3 . An exemplary embodiment of formula Ig includes wherein R 4 is −O−(C 1 -C 12 alkyldiyl)− (C 2 -C 20 heterocyclyldiyl)−*. An exemplary embodiment of formula Ig includes wherein C 1 -C 12 alkyldiyl is propyldiyl and C 2 -C 20 heterocyclyldiyl is piperidiyl. An exemplary embodiment of the immunoconjugate includes wherein L is selected from the group consisting of: −C(=O)−(PEG)−C(=O)−(EsPEP)−; −C(=O)−(PEG)−C(=O)−(EsPEP)−N(R 6 )−; −C(=O)−(PEG)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−; −C(=O)−(PEG)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)−(C 2 -C 5 monoheterocyclyldiyl)−; −C(=O)−(PEG)−N(R 6 )−(PEG)−C(=O)−(EsPEP)−; −C(=O)−(PEG)−N + (R 6 ) 2 −(PEG)−C(=O)−(EsPEP)−; −C(=O)−(PEG)−C(=O)−N(R 6 )CH(AA 1 )C(=O)−(PEG)−C(=O)−(EsPEP)−; −C(=O)−(C 1 -C 12 alkyldiyl)−C(=O)−(EsPEP)−; −C(=O)−(C 1 -C 12 alkyldiyl)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−; −C(=O)−(C 1 -C 12 alkyldiyl)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )−C(=O); −C(=O)−(C 1 -C 12 alkyldiyl)−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)− (C 2 -C5 monoheterocyclyldiyl)−; −succinimidyl−(CH 2 )m−C(=O)N(R 6 )−PEG−C(=O)−(EsPEP)−; −succinimidyl−(CH 2 )m−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−; −succinimidyl−(CH 2 )m−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)N(R 6 )C(=O)−; and −(succinimidyl)−(CH 2 )m−C(=O)−(EsPEP)−N(R 6 )−(C 1 -C 12 alkyldiyl)−N(R 6 )C(=O)−(C 2 - C5 monoheterocyclyldiyl)−; PEG has the formula: −(CH 2 CH 2 O)n−(CH 2 )m−; m is an integer from 1 to 5, and n is an integer from 2 to 50; EsPEP is the elastase-substrate, peptide linker unit comprising 2 to 12 amino acid residues; and R 6 is selected from the group consisting of H, C 6 -C 20 aryl, C 6 -C 20 aryldiyl, C 1 -C 12 alkyl, and C 1 -C 12 alkyldiyl, or two R 6 groups together form a 5- or 6-membered heterocyclyl ring; alkyl, alkyldiyl, alkenyl, alkenyldiyl, alkynyl, alkynyldiyl, aryl, aryldiyl, carbocyclyl, carbocyclyldiyl, heterocyclyl, heterocyclyldiyl, heteroaryl, and heteroaryldiyl are independently and optionally substituted with one or more groups independently selected from F, Cl, Br, I, − CN, −CH 3 , −CH 2 CH 3 , −CH=CH 2 , −C ≡CH, −C ≡CCH 3 , −CH 2 CH 2 CH 3 , −CH(CH 3 ) 2 , − CH 2 CH(CH 3 ) 2 , −CH 2 OH, −CH 2 OCH 3 , −CH 2 CH 2 OH, −C(CH 3 ) 2 OH, −CH(OH)CH(CH 3 ) 2 , − C(CH 3 ) 2 CH 2 OH, −CH 2 CH 2 SO 2 CH 3 , −CH 2 OP(O)(OH) 2 , −CH 2 F, −CHF 2 , −CF 3 , −CH 2 CF 3 , − CH 2 CHF 2 , −CH(CH 3 )CN, −C(CH 3 ) 2 CN, −CH 2 CN, −CH 2 NH 2 , −CH 2 NHSO 2 CH 3 , −CH 2 NHCH 3 , −CH 2 N(CH 3 ) 2 , −CO 2 H, −COCH 3 , −CO 2 CH 3 , −CO 2 C(CH 3 ) 3 , −COCH(OH)CH 3 , −CONH 2 , − CONHCH 3 , −CON(CH 3 ) 2 , −C(CH 3 ) 2 CONH 2 , −NH 2 , −NHCH 3 , −N(CH 3 ) 2 , −NHCOCH 3 , − N(CH 3 )COCH 3 , −NHS(O) 2 CH 3 , −N(CH 3 )C(CH 3 ) 2 CONH 2 , −N(CH 3 )CH 2 CH 2 S(O) 2 CH 3 , − NHC(=NH)H, −NHC(=NH)CH 3 , −NHC(=NH)NH 2 , −NHC(=O)NH 2 , −NO 2 , =O, −OH, −OCH 3 , −OCH 2 CH 3 , −OCH 2 CH 2 OCH 3 , −OCH 2 CH 2 OH, −OCH 2 CH 2 N(CH 3 ) 2 , −O(CH 2 CH 2 O) n − (CH 2 )mCO 2 H, −O(CH 2 CH 2 O)nH, −OP(O)(OH) 2 , −S(O) 2 N(CH 3 ) 2 , −SCH 3 , −S(O) 2 CH 3 , and − S(O)3H. An exemplary embodiment of the immunoconjugate includes wherein EsPEP has the formula: where AA is independently selected from a natural or unnatural amino acid side chain, or one or more of AA, and an adjacent nitrogen atom form a 5-membered ring proline amino acid, and the wavy line indicates a point of attachment; Cyc is selected from C 6 -C 20 aryldiyl and C 1 -C 20 heteroaryldiyl, optionally substituted with one or more groups selected from F, Cl, NO 2 , −OH, −OCH 3 , and a glucuronic acid having the structure: R 7 is selected from the group consisting of −CH(R 8 )O−, −CH 2 −, −CH 2 N(R 8 )−, and − CH(R 8 )O−C(=O)−, where R 8 is selected from H, C 1 -C 6 alkyl, C(=O)−C 1 -C 6 alkyl, and − C(=O)N(R 9 ) 2 , where R 9 is independently selected from the group consisting of H, C 1 -C 12 alkyl, and −(CH 2 CH 2 O)n−(CH 2 )m−OH, where m is an integer from 1 to 5, and n is an integer from 2 to 50, or two R 9 groups together form a 5- or 6-membered heterocyclyl ring; y is an integer from 2 to 12; and z is 0 or 1. An exemplary embodiment of the immunoconjugate includes wherein y is selected from 2, 3, and 4. An exemplary embodiment of the immunoconjugate wherein EsPEP is a tripeptide having the formula: where AA 1 , AA 2 and AA 3 are independently selected from a natural or unnatural amino acid, and the wavy line indicates a point of attachment; Cyc is selected from C 6 -C 20 aryldiyl and C 1 -C 20 heteroaryldiyl, optionally substituted with one or more groups selected from F, Cl, NO 2 , −OH, −OCH 3 , and a glucuronic acid having the structure: R 7 is selected from the group consisting of −CH(R 8 )O−, −CH 2 −, −CH 2 N(R 8 )−, and − CH(R 8 )O−C(=O)−, where R 8 is selected from H, C 1 -C 6 alkyl, C(=O)−C 1 -C 6 alkyl, and − C(=O)N(R 9 ) 2 , where R 9 is independently selected from the group consisting of H, C 1 -C 12 alkyl, and −(CH 2 CH 2 O)n−(CH 2 )m−OH, where m is an integer from 1 to 5, and n is an integer from 2 to 50, or two R 9 groups together form a 5- or 6-membered heterocyclyl ring; and z is 0 or 1. An exemplary embodiment of the immunoconjugate includes wherein AA 1 is methyl, AA 2 forms proline, and AA 3 is isopropyl. An exemplary embodiment of the immunoconjugate includes wherein EsPEP has the formula: . An exemplary embodiment of the immunoconjugate includes wherein EsPEP is selected from the formulas: . An exemplary embodiment of the immunoconjugate includes wherein L is: −C(=O)−(PEG)−C(=O)−(EsPEP)− . An exemplary embodiment of the immunoconjugate includes wherein PEG is: −(CH 2 CH 2 O) 25 −(CH 2 ) 2 − . An exemplary embodiment of the immunoconjugate has the formula: . An exemplary embodiment of the immunoconjugate includes wherein Ims has formula IIc: IIc. Exemplary embodiments of the immunoconjugate includes wherein Ims have the structures: where the wavy line indicates the site of attachment to the linker. An exemplary embodiment of the immunoconjugate includes wherein EsPEP is a tetrapeptide having the formula: where AA 1 , AA 2 , AA 3 and AA 4 are independently selected from a natural or unnatural amino acid, and the wavy line indicates a point of attachment; Cyc is selected from C 6 -C 20 aryldiyl and C 1 -C 20 heteroaryldiyl, optionally substituted with one or more groups selected from F, Cl, NO 2 , −OH, −OCH 3 , and a glucuronic acid having the structure: ; R 7 is selected from the group consisting of −CH(R 8 )O−, −CH 2 −, −CH 2 N(R 8 )−, and − CH(R 8 )O−C(=O)−, where R 8 is selected from H, C 1 -C 6 alkyl, C(=O)−C 1 -C 6 alkyl, and − C(=O)N(R 9 ) 2 , where R 9 is independently selected from the group consisting of H, C 1 -C 12 alkyl, and −(CH 2 CH 2 O)n−(CH 2 )m−OH, where m is an integer from 1 to 5, and n is an integer from 2 to 50, or two R 9 groups together form a 5- or 6-membered heterocyclyl ring; and z is 0 or 1. An exemplary embodiment of the immunoconjugate includes wherein AA1 is selected from the group consisting of Abu, Ala, and Val; AA 2 is selected from the group consisting of Nle(O-Bzl), Oic and Pro; AA 3 is selected from the group consisting of Ala and Met(O) 2 ; and AA4 is selected from the group consisting of Oic, Arg(NO 2 ), Bpa, and Nle(O-Bzl). An exemplary embodiment of the immunoconjugate includes wherein EsPEP has the formula: . An exemplary embodiment of the immunoconjugate includes wherein EsPEP has the formula:
. An exemplary embodiment of the immunoconjugate includes a structure selected from IIe-h: ' where the wavy line indicates the attachment through L to the antibody. An exemplary embodiment of the immunoconjugate includes a structure selected from IIi-l: where the wavy line indicates the attachment through L to the antibody. An exemplary embodiment of the immunoconjugate includes wherein R 2 and R 3 are each C 1 -C 8 alkyl. An exemplary embodiment of the immunoconjugate includes wherein R 2 and R 3 are each −CH 2 CH 2 CH 3 . An exemplary embodiment of the immunoconjugate includes wherein X 2 and X 3 are each a bond, and R 2 or R 3 is −O−(C 1 -C 12 alkyl). An exemplary embodiment of the immunoconjugate includes wherein R 2 or R 3 is − OCH 2 CH 3 . An exemplary embodiment of the immunoconjugate includes wherein the elastase- substrate, peptide linker is cleaved by elastase. Exemplary embodiments of the immunoconjugate have the structures:
The invention includes all reasonable combinations, and permutations of the features, of the Formula I embodiments. Drug loading is represented by p, the number of immunostimulatory moieties per antibody in an immunoconjugate of Formula I. Drug (immunostimulant) loading may range from 1 to about 8 drug moieties (D) per antibody. Immunoconjugates of Formula I include mixtures or collections of antibodies conjugated with a range of drug moieties, from 1 to about 8. In some embodiments, the number of drug moieties that can be conjugated to an antibody is limited by the number of reactive or available amino acid side chain residues such as lysine and cysteine. In some embodiments, free cysteine residues are introduced into the antibody amino acid sequence by the methods described herein. In such aspects, p may be 1, 2, 3, 4, 5, 6, 7, or 8, and ranges thereof, such as from 1 to 8 or from 2 to 5. In any such aspect, p and n are equal (i.e., p = n = 1, 2, 3, 4, 5, 6, 7, or 8, or some range there between). Exemplary immunoconjugates of Formula I include, but are not limited to, antibodies that have 1, 2, 3, or 4 engineered cysteine amino acids (Lyon, R. et al. (2012) Methods in Enzym.502:123-138). In some embodiments, one or more free cysteine residues are already present in an antibody forming intrachain disulfide bonds, without the use of engineering, in which case the existing free cysteine residues may be used to conjugate the antibody to a drug. In some embodiments, an antibody is exposed to reducing conditions prior to conjugation of the antibody in order to generate one or more free cysteine residues. For some immunoconjugates, p may be limited by the number of attachment sites on the antibody. For example, where the attachment is a cysteine thiol, as in certain exemplary embodiments described herein, an antibody may have only one or a limited number of cysteine thiol groups, or may have only one or a limited number of sufficiently reactive thiol groups, to which the drug may be attached. In other embodiments, one or more lysine amino groups in the antibody may be available and reactive for conjugation with an immunostimulant-linker compound of Formula II. In certain embodiments, higher drug loading, e.g. p >5, may cause aggregation, insolubility, toxicity, or loss of cellular permeability of certain antibody-drug conjugates. In certain embodiments, the average drug loading for an immunoconjugate ranges from 1 to about 8; from about 2 to about 6; or from about 3 to about 5. In certain embodiments, an antibody is subjected to denaturing conditions to reveal reactive nucleophilic groups such as lysine or cysteine. The loading (drug/antibody ratio) of an immunoconjugate may be controlled in different ways, and for example, by: (i) limiting the molar excess of the immunostimulant-linker intermediate compound relative to antibody, (ii) limiting the conjugation reaction time or temperature, and (iii) partial or limiting reductive denaturing conditions for optimized antibody reactivity. Where more than one nucleophilic group of the antibody reacts with a drug, then the resulting product is a mixture of immunoconjugate compounds with a distribution of one or more drug moieties attached to an antibody. The average number of drugs per antibody may be calculated from the mixture by a dual ELISA antibody assay, which is specific for antibody and specific for the drug. Individual immunoconjugate molecules may be identified in the mixture by mass spectroscopy and separated by HPLC, e.g. hydrophobic interaction chromatography (see, e.g., McDonagh et al. (2006) Prot. Engr. Design & Selection 19(7):299-307; Hamblett et al. (2004) Clin. Cancer Res.10:7063-7070; Hamblett, K.J., et al. “Effect of drug loading on the pharmacology, pharmacokinetics, and toxicity of an anti-CD30 antibody-drug conjugate,” Abstract No.624, American Association for Cancer Research, 2004 Annual Meeting, March 27- 31, 2004, Proceedings of the AACR, Volume 45, March 2004; Alley, S.C., et al. “Controlling the location of drug attachment in antibody-drug conjugates,” Abstract No.627, American Association for Cancer Research, 2004 Annual Meeting, March 27-31, 2004, Proceedings of the AACR, Volume 45, March 2004). In certain embodiments, a homogeneous immunoconjugate with a single loading value may be isolated from the conjugation mixture by electrophoresis or chromatography. BIOLOGICAL ACTIVITY OF IMMUNOCONJUGATES Figure 9 shows a graph measuring potency as measured by TNFa production in a co- culture experiment with RAW 264.7 murine macrophage cells and HCC1954 HER2 expressing tumor cells. This experiment compared an elastase cleavable linker (Ala-Pro-Val) immunoconjugate ISAC-1, and a cathepsin B cleavable linker (Val-Cit) immunoconjugate ISAC-2. The antibody of ISAC-1 and ISAC-2 is anti-HER2 trastuzumab. The Val-Cit linker unit of ISAC-2 is a known cathepsin B substrate. Cells were cultured overnight at a 10:1 effector (macrophage) to target ( HCC1954 tumor cell) ratio, and mouse TNFa was measured by ELISA as a readout of a proinflammatory response. The data demonstrated that ISAC-1 has increased potency relative to the cathepsin B cleavable peptide (Val-Cit) ISAC-2. The RAW 264.7 murine macrophage cell line was cultured according to vendor protocols (Invivogen) and Example 203. PHARMACEUTICAL COMPOSITIONS OF IMMUNOCONJUGATES The invention provides a composition, e.g., a pharmaceutically or pharmacologically acceptable composition or formulation, comprising a plurality of immunoconjugates as described herein and optionally a carrier therefor, e.g., a pharmaceutically or pharmacologically acceptable carrier. The immunoconjugates can be the same or different in the composition, i.e., the composition can comprise immunoconjugates that have the same number of immunostimulants linked to the same positions on the antibody construct and/or immunoconjugates that have the same number of immunostimulants linked to different positions on the antibody construct, that have different numbers of immunostimulants linked to the same positions on the antibody construct, or that have different numbers of immunostimulants linked to different positions on the antibody construct. In an exemplary embodiment, a pharmaceutical composition comprises a therapeutically effective amount of the immunoconjugate and one or more pharmaceutically acceptable diluent, vehicle, carrier or excipient. In an exemplary embodiment, a composition comprising the immunoconjugate compounds comprises a mixture of the immunoconjugate compounds, wherein the average drug (immunostimulatory moieties) loading per antibody in the mixture of immunoconjugate compounds is about 2 to about 5. A composition of immunoconjugates of the invention can have an average adjuvant to antibody construct ratio (DAR) of about 0.4 to about 10. A skilled artisan will recognize that the number of immunostimulatory moieties conjugated to the antibody construct may vary from immunoconjugate to immunoconjugate in a composition comprising multiple immunoconjugates of the invention and thus the adjuvant to antibody construct (e.g., antibody) ratio can be measured as an average which may be referred to as the drug to antibody ratio (DAR). The adjuvant to antibody construct (e.g., antibody) ratio can be assessed by any suitable means, many of which are known in the art. The average number of adjuvant moieties per antibody (DAR) in preparations of immunoconjugates from conjugation reactions may be characterized by conventional means such as mass spectrometry, ELISA assay, and HPLC. The quantitative distribution of immunoconjugates in a composition in terms of p may also be determined. In some instances, separation, purification, and characterization of homogeneous immunoconjugates where p is a certain value from immunoconjugates with other drug loadings may be achieved by means such as reverse phase HPLC or electrophoresis. In some embodiments, the composition further comprises one or more pharmaceutically or pharmacologically acceptable excipients. For example, the immunoconjugates of the invention can be formulated for parenteral administration, such as IV administration or administration into a body cavity or lumen of an organ. Alternatively, the immunoconjugates can be injected intra-tumorally. Compositions for injection will commonly comprise a solution of the immunoconjugate dissolved in a pharmaceutically acceptable carrier. Among the acceptable vehicles and solvents that can be employed are water and an isotonic solution of one or more salts such as sodium chloride, e.g., Ringer's solution. In addition, sterile fixed oils can conventionally be employed as a solvent or suspending medium. For this purpose, any bland fixed oil can be employed, including synthetic monoglycerides or diglycerides. In addition, fatty acids such as oleic acid can likewise be used in the preparation of injectables. These compositions desirably are sterile and generally free of undesirable matter. These compositions can be sterilized by conventional, well known sterilization techniques. The compositions can contain pharmaceutically acceptable auxiliary substances as required to approximate physiological conditions such as pH adjusting and buffering agents, toxicity adjusting agents, e.g., sodium acetate, sodium chloride, potassium chloride, calcium chloride, sodium lactate and the like. The composition can contain any suitable concentration of the immunoconjugate. The concentration of the immunoconjugate in the composition can vary widely, and will be selected primarily based on fluid volumes, viscosities, body weight, and the like, in accordance with the particular mode of administration selected and the patient's needs. In certain embodiments, the concentration of an immunoconjugate in a solution formulation for injection will range from about 0.1% (w/w) to about 10% (w/w). METHOD OF TREATING CANCER WITH IMMUNOCONJUGATES The invention provides a method for treating cancer. The method includes administering a therapeutically effective amount of an immunoconjugate as described herein (e.g., as a composition as described herein) to a subject in need thereof, e.g., a subject that has cancer and is in need of treatment for the cancer. The method includes administering a therapeutically effective amount of an immunoconjugate (IC). In certain embodiments, the immunoconjugate compounds of the invention include those with immunostimulatory activity. The immunoconjugates of the invention selectively deliver an effective dose of an immunostimulatory drug to tumor tissue, whereby greater selectivity (i.e., a lower efficacious dose) may be achieved while increasing the therapeutic index (“therapeutic window”) relative to the unconjugated immunostimulatory drug. In an exemplary embodiment, the invention provides a pharmaceutical composition for use in therapy. The invention provides a method of treatment comprising administering a therapeutically-effective dose of the immunoconjugate to a patient with an immune-related disorder. The elastase-substrate, peptide linker of the immunoconjugate may be cleaved by elastase. It is contemplated that the immunoconjugate of the present invention may be used to treat various hyperproliferative diseases or disorders, e.g. characterized by the overexpression of a tumor antigen. Exemplary hyperproliferative disorders include benign or malignant solid tumors and hematological disorders such as leukemia and lymphoid malignancies. In another aspect, an immunoconjugate for use as a medicament is provided. In certain embodiments, the invention provides an immunoconjugate for use in a method of treating an individual comprising administering to the individual an effective amount of the immunoconjugate. In one such embodiment, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent, e.g., as described herein. In a further aspect, the invention provides for the use of an immunoconjugate in the manufacture or preparation of a medicament. In one embodiment, the medicament is for treatment of cancer, the method comprising administering to an individual having cancer an effective amount of the medicament. In one such embodiment, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent, e.g., as described herein. Carcinomas are malignancies that originate in the epithelial tissues. Epithelial cells cover the external surface of the body, line the internal cavities, and form the lining of glandular tissues. Examples of carcinomas include, but are not limited to, adenocarcinoma (cancer that begins in glandular (secretory) cells such as cancers of the breast, pancreas, lung, prostate, stomach, gastroesophageal junction, and colon) adrenocortical carcinoma; hepatocellular carcinoma; renal cell carcinoma; ovarian carcinoma; carcinoma in situ; ductal carcinoma; carcinoma of the breast; basal cell carcinoma; squamous cell carcinoma; transitional cell carcinoma; colon carcinoma; nasopharyngeal carcinoma; multilocular cystic renal cell carcinoma; oat cell carcinoma; large cell lung carcinoma; small cell lung carcinoma; non-small cell lung carcinoma; and the like. Carcinomas may be found in prostrate, pancreas, colon, brain (usually as secondary metastases), lung, breast, and skin. In some embodiments, methods for treating non-small cell lung carcinoma include administering an immunoconjugate containing an antibody construct that is capable of binding PD-L1 (e.g., atezolizumab, durvalumab, avelumab, biosimilars thereof, or biobetters thereof). In some embodiments, methods for treating breast cancer include administering an immunoconjugate containing an antibody construct that is capable of binding PD-L1 (e.g., atezolizumab, durvalumab, avelumab, biosimilars thereof, or biobetters thereof). In some embodiments, methods for treating triple-negative breast cancer include administering an immunoconjugate containing an antibody construct that is capable of binding PD-L1 (e.g., atezolizumab, durvalumab, avelumab, biosimilars thereof, or biobetters thereof). Soft tissue tumors are a highly diverse group of rare tumors that are derived from connective tissue. Examples of soft tissue tumors include, but are not limited to, alveolar soft part sarcoma; angiomatoid fibrous histiocytoma; chondromyoxid fibroma; skeletal chondrosarcoma; extraskeletal myxoid chondrosarcoma; clear cell sarcoma; desmoplastic small round-cell tumor; dermatofibrosarcoma protuberans; endometrial stromal tumor; Ewing’s sarcoma; fibromatosis (Desmoid); fibrosarcoma, infantile; gastrointestinal stromal tumor; bone giant cell tumor; tenosynovial giant cell tumor; inflammatory myofibroblastic tumor; uterine leiomyoma; leiomyosarcoma; lipoblastoma; typical lipoma; spindle cell or pleomorphic lipoma; atypical lipoma; chondroid lipoma; well-differentiated liposarcoma; myxoid/round cell liposarcoma; pleomorphic liposarcoma; myxoid malignant fibrous histiocytoma; high-grade malignant fibrous histiocytoma; myxofibrosarcoma; malignant peripheral nerve sheath tumor; mesothelioma; neuroblastoma; osteochondroma; osteosarcoma; primitive neuroectodermal tumor; alveolar rhabdomyosarcoma; embryonal rhabdomyosarcoma; benign or malignant schwannoma; synovial sarcoma; Evan’s tumor; nodular fasciitis; desmoid-type fibromatosis; solitary fibrous tumor; dermatofibrosarcoma protuberans (DFSP); angiosarcoma; epithelioid hemangioendothelioma; tenosynovial giant cell tumor (TGCT); pigmented villonodular synovitis (PVNS); fibrous dysplasia; myxofibrosarcoma; fibrosarcoma; synovial sarcoma; malignant peripheral nerve sheath tumor; neurofibroma; pleomorphic adenoma of soft tissue; and neoplasias derived from fibroblasts, myofibroblasts, histiocytes, vascular cells/endothelial cells, and nerve sheath cells. A sarcoma is a rare type of cancer that arises in cells of mesenchymal origin, e.g., in bone or in the soft tissues of the body, including cartilage, fat, muscle, blood vessels, fibrous tissue, or other connective or supportive tissue. Different types of sarcoma are based on where the cancer forms. For example, osteosarcoma forms in bone, liposarcoma forms in fat, and rhabdomyosarcoma forms in muscle. Examples of sarcomas include, but are not limited to, askin's tumor; sarcoma botryoides; chondrosarcoma; ewing's sarcoma; malignant hemangioendothelioma; malignant schwannoma; osteosarcoma; and soft tissue sarcomas (e.g., alveolar soft part sarcoma; angiosarcoma; cystosarcoma phyllodesdermatofibrosarcoma protuberans (DFSP); desmoid tumor; desmoplastic small round cell tumor; epithelioid sarcoma; extraskeletal chondrosarcoma; extraskeletal osteosarcoma; fibrosarcoma; gastrointestinal stromal tumor (GIST); hemangiopericytoma; hemangiosarcoma (more commonly referred to as “angiosarcoma”); kaposi’s sarcoma; leiomyosarcoma; liposarcoma; lymphangiosarcoma; malignant peripheral nerve sheath tumor (MPNST); neurofibrosarcoma; synovial sarcoma; and undifferentiated pleomorphic sarcoma). A teratoma is a type of germ cell tumor that may contain several different types of tissue (e.g., can include tissues derived from any and/or all of the three germ layers: endoderm, mesoderm, and ectoderm), including, for example, hair, muscle, and bone. Teratomas occur most often in the ovaries in women, the testicles in men, and the tailbone in children. Melanoma is a form of cancer that begins in melanocytes (cells that make the pigment melanin). Melanoma may begin in a mole (skin melanoma), but can also begin in other pigmented tissues, such as in the eye or in the intestines. Merkel cell carcinoma is a rare type of skin cancer that usually appears as a flesh-colored or bluish-red nodule on the face, head or neck. Merkel cell carcinoma is also called neuroendocrine carcinoma of the skin. In some embodiments, methods for treating Merkel cell carcinoma include administering an immunoconjugate containing an antibody construct that is capable of binding PD-L1 (e.g., atezolizumab, durvalumab, avelumab, biosimilars thereof, or biobetters thereof). In some embodiments, the Merkel cell carcinoma has metastasized when administration occurs. Leukemias are cancers that start in blood-forming tissue, such as the bone marrow, and cause large numbers of abnormal blood cells to be produced and enter the bloodstream. For example, leukemias can originate in bone marrow-derived cells that normally mature in the bloodstream. Leukemias are named for how quickly the disease develops and progresses (e.g., acute versus chronic) and for the type of white blood cell that is affected (e.g., myeloid versus lymphoid). Myeloid leukemias are also called myelogenous or myeloblastic leukemias. Lymphoid leukemias are also called lymphoblastic or lymphocytic leukemia. Lymphoid leukemia cells may collect in the lymph nodes, which can become swollen. Examples of leukemias include, but are not limited to, Acute myeloid leukemia (AML), Acute lymphoblastic leukemia (ALL), Chronic myeloid leukemia (CML), and Chronic lymphocytic leukemia (CLL). Lymphomas are cancers that begin in cells of the immune system. For example, lymphomas can originate in bone marrow-derived cells that normally mature in the lymphatic system. There are two basic categories of lymphomas. One category of lymphoma is Hodgkin lymphoma (HL), which is marked by the presence of a type of cell called the Reed-Sternberg cell. There are currently 6 recognized types of HL. Examples of Hodgkin lymphomas include nodular sclerosis classical Hodgkin lymphoma (CHL), mixed cellularity CHL, lymphocyte- depletion CHL, lymphocyte-rich CHL, and nodular lymphocyte predominant HL. The other category of lymphoma is non-Hodgkin lymphomas (NHL), which includes a large, diverse group of cancers of immune system cells. Non-Hodgkin lymphomas can be further divided into cancers that have an indolent (slow-growing) course and those that have an aggressive (fast-growing) course. There are currently 61 recognized types of NHL. Examples of non-Hodgkin lymphomas include, but are not limited to, AIDS-related Lymphomas, anaplastic large-cell lymphoma, angioimmunoblastic lymphoma, blastic NK-cell lymphoma, Burkitt’s lymphoma, Burkitt-like lymphoma (small non-cleaved cell lymphoma), chronic lymphocytic leukemia/small lymphocytic lymphoma, cutaneous T-Cell lymphoma, diffuse large B-Cell lymphoma, enteropathy-type T-Cell lymphoma, follicular lymphoma, hepatosplenic gamma- delta T-Cell lymphomas, T-Cell leukemias, lymphoblastic lymphoma, mantle cell lymphoma, marginal zone lymphoma, nasal T-Cell lymphoma, pediatric lymphoma, peripheral T-Cell lymphomas, primary central nervous system lymphoma, transformed lymphomas, treatment- related T-Cell lymphomas, and Waldenstrom's macroglobulinemia. Brain cancers include any cancer of the brain tissues. Examples of brain cancers include, but are not limited to, gliomas (e.g., glioblastomas, astrocytomas, oligodendrogliomas, ependymomas, and the like), meningiomas, pituitary adenomas, and vestibular schwannomas, primitive neuroectodermal tumors (medulloblastomas). Immunoconjugates of the invention can be used either alone or in combination with other agents in a therapy. For instance, an immunoconjugate may be co-administered with at least one additional therapeutic agent, such as a chemotherapeutic agent. Such combination therapies encompass combined administration (where two or more therapeutic agents are included in the same or separate formulations), and separate administration, in which case, administration of the immunoconjugate can occur prior to, simultaneously, and/or following, administration of the additional therapeutic agent and/or adjuvant. Immunoconjugates can also be used in combination with radiation therapy. The immunoconjugates of the invention (and any additional therapeutic agent) can be administered by any suitable means, including parenteral, intrapulmonary, and intranasal, and, if desired for local treatment, intralesional administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. Dosing can be by any suitable route, e.g. by injections, such as intravenous or subcutaneous injections, depending in part on whether the administration is brief or chronic. Various dosing schedules including but not limited to single or multiple administrations over various time-points, bolus administration, and pulse infusion are contemplated herein. Atezolizumab, durvalumab, avelumab, biosimilars thereof, and biobetters thereof are known to be useful in the treatment of cancer, particularly breast cancer, especially triple negative (test negative for estrogen receptors, progesterone receptors, and excess HER2 protein) breast cancer, bladder cancer, and Merkel cell carcinoma. The immunoconjugate described herein can be used to treat the same types of cancers as atezolizumab, durvalumab, avelumab, biosimilars thereof, and biobetters thereof, particularly breast cancer, especially triple negative (test negative for estrogen receptors, progesterone receptors, and excess HER2 protein) breast cancer, bladder cancer, and Merkel cell carcinoma. The immunoconjugate is administered to a subject in need thereof in any therapeutically effective amount using any suitable dosing regimen, such as the dosing regimens utilized for atezolizumab, durvalumab, avelumab, biosimilars thereof, and biobetters thereof. For example, the methods can include administering the immunoconjugate to provide a dose of from about 100 ng/kg to about 50 mg/kg to the subject. The immunoconjugate dose can range from about 5 mg/kg to about 50 mg/kg, from about 10 µg/kg to about 5 mg/kg, or from about 100 µg/kg to about 1 mg/kg. The immunoconjugate dose can be about 100, 200, 300, 400, or 500 µg/kg. The immunoconjugate dose can be about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 mg/kg. The immunoconjugate dose can also be outside of these ranges, depending on the particular conjugate as well as the type and severity of the cancer being treated. Frequency of administration can range from a single dose to multiple doses per week, or more frequently. In some embodiments, the immunoconjugate is administered from about once per month to about five times per week. In some embodiments, the immunoconjugate is administered once per week. In another aspect, the invention provides a method for preventing cancer. The method comprises administering a therapeutically effective amount of an immunoconjugate (e.g., as a composition as described above) to a subject. In certain embodiments, the subject is susceptible to a certain cancer to be prevented. For example, the methods can include administering the immunoconjugate to provide a dose of from about 100 ng/kg to about 50 mg/kg to the subject. The immunoconjugate dose can range from about 5 mg/kg to about 50 mg/kg, from about 10 µg/kg to about 5 mg/kg, or from about 100 µg/kg to about 1 mg/kg. The immunoconjugate dose can be about 100, 200, 300, 400, or 500 µg/kg. The immunoconjugate dose can be about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 mg/kg. The immunoconjugate dose can also be outside of these ranges, depending on the particular conjugate as well as the type and severity of the cancer being treated. Frequency of administration can range from a single dose to multiple doses per week, or more frequently. In some embodiments, the immunoconjugate is administered from about once per month to about five times per week. In some embodiments, the immunoconjugate is administered once per week. Some embodiments of the invention provide methods for treating cancer as described above, wherein the cancer is breast cancer. Breast cancer can originate from different areas in the breast, and a number of different types of breast cancer have been characterized. For example, the immunoconjugates of the invention can be used for treating ductal carcinoma in situ; invasive ductal carcinoma (e.g., tubular carcinoma; medullary carcinoma; mucinous carcinoma; papillary carcinoma; or cribriform carcinoma of the breast); lobular carcinoma in situ; invasive lobular carcinoma; inflammatory breast cancer; and other forms of breast cancer such as triple negative (test negative for estrogen receptors, progesterone receptors, and excess HER2 protein) breast cancer. In some embodiments, methods for treating breast cancer include administering an immunoconjugate containing an antibody construct that is capable of binding HER2 (e.g. trastuzumab, pertuzumab, biosimilars, or biobetters thereof ) and PD-L1 (e.g., atezolizumab, durvalumab, avelumab, biosimilars, or biobetters thereof). In some embodiments, methods for treating colon cancer lung cancer, renal cancer, pancreatic cancer, gastric cancer, and esophageal cancer include administering an immunoconjugate containing an antibody construct that is capable of binding CEA, or tumors over-expressing CEA (e.g. labetuzumab, biosimilars, or biobetters thereof). In some embodiments, the cancer is susceptible to a pro-inflammatory response induced by TLR7 and/or TLR8. EXAMPLES Preparation of immunostimulatory compounds and intermediates Example 1 Synthesis of 2,3,5,6-tetrafluorophenyl (R)-1-((S)-2-(((S)-1-((4-(((((1-((3- (2-amino-4-(dipropylcarbamoyl)-3H-benzo[b]azepin-8-yl)phenyl )sulfonyl)azetidin-3- yl)methyl)carbamoyl)oxy)methyl)phenyl)amino)-3-methyl-1-oxob utan-2- yl)carbamoyl)pyrrolidin-1-yl)-2-methyl-1,4-dioxo- 7,10,13,16,19,22,25,28,31,34,37,40,43,46,49,52,55,58,61,64,6 7,70,73,76,79-pentacosaoxa-3- azadooctacontan-82-oate, II-1 Preparation of O 2 -(2,5-dioxopyrrolidin-1-yl) O1-(9H-fluoren-9-ylmethyl) (2S)- pyrrolidine-1,2-dicarboxylate, II-1b
To a solution of (2S)-1-(9H-fluoren-9-ylmethoxycarbonyl)pyrrolidine-2-carboxy lic acid, II-1a (15 g, 44.5 mmol, 1.0 eq) in DCM (200 mL) was added 1-hydroxypyrrolidine-2,5-dione (5.12 g, 44.5 mmol, 1.0 eq) and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride, EDCI (10.2 g, 53.44 mmol, 1.2 eq). The mixture was stirred at 20 °C for 12 h and then washed by saturated aqueous solution of NaHCO 3 (70 mL x 3). The organic layer was dried over Na2SO4 and concentrated to give II-1b (17.5 g, 40.28 mmol, 90.60% yield) as a white solid. Preparation of (2S)-2-[[(2S)-1-(9H-fluoren-9-ylmethoxycarbonyl)pyrrolidine- 2- carbonyl]amino]-3-methyl-butanoic acid, II-1c To a solution of (2S)-2-amino-3-methyl-butanoic acid (4.95 g, 42.3 mmol, 1.05 eq) in THF (200 mL) was added NaHCO 3 (3.55 g, 42.3 mmol, 1.64 mL, 1.05 eq) in H 2 O (50 mL) and II-1b (17.5 g, 40.28 mmol, 1.0 eq) and it was stirred at 20°C for 12 h. The mixture was extracted with methyl, tert-butylether, MTBE (2 x 100 mL) (discarded). The pH of aqueous layer was adjusted to 5-6 with HCl (6 M) and extracted with EtOAc (3 x 200 ml). The combined organic layer was dried over Na 2 SO 4 and concentrated to give II-1c (15 g, 34.36 mmol, 85.31% yield) as white solid. 1 H NMR (MeOD, 400 MHz) δ7.80 (d, J = 7.2 Hz, 2H), 7.70-7.54 (m, 2H), 7.43-7.28 (m, 4H), 4.49-4.15 (m, 5H), 3.69-3.38 (m, 2H), 2.42-2.01 (m, 3H), 2.00-1.82 (m, 2H), 1.01-0.86 (m, 6H) Preparation of 9H-fluoren-9-ylmethyl(2S)-2-[[(1S)-1-[[4-(hydroxymethyl)phen yl] carbamoyl]-2-methyl-propyl]carbamoyl]pyrrolidine-1-carboxyla te, II-1d To a solution of II-1c (10 g, 22.9 mmol, 1.0 eq) and (4-aminophenyl)methanol (4.23 g, 34.4 mmol, 1.5 eq) in MeOH (80 mL) and DCM (80 mL) was added N-ethoxycarbonyl-2- ethoxy-1,2-dihydroquinoline, EEDQ (8.50 g, 34.36 mmol, 1.5 eq) and then stirred at 20 °C for 12 h. The mixture was concentrated in vacuum to give a residue and the residue was purified by flash silica gel chromatography (Silica Flash Column, Eluent of 0 to 40% Ethyl acetate/MeOH at 65 mL/min). The crude product was triturated with EtOAc at 20 o C for 20 min to give II-1d (13 g, 24.0 mmol, 52.38% yield) as yellow solid. 1 H NMR (MeOD, 400 MHz) δ 7.85-7.71 (m, 2H), 7.68-7.48 (m, 3H), 7.47-7.16 (m, 7H), 4.53 (d, J = 15.2 Hz, 2H), 4.49-4.41 (m, 1H), 4.40- 4.33 (m, 2H), 4.32-4.27 (m, 1H), 4.26-4.17 (m, 1H), 4.16-4.07 (m, 1H), 3.69-3.38 (m, 2H), 2.40- 2.05 (m, 2H), 1.99-1.82 (m, 2H), 1.08-0.88 (m, 6H) Preparation of (2S)-N-[(1S)-1-[[4-(hydroxymethyl)phenyl]carbamoyl]-2-methyl - propyl]pyrrolidine-2-carboxamide, II-1e To a solution of II-1d (13 g, 24.0 mmol, 1.0 eq) in DCM (130 mL) was added piperidine (10.22 g, 120 mmol, 11.85 mL, 5.0 eq) and then stirred at 20 °C for 2 h. The mixture was concentrated to give a residue and the residue was triturated with EtOAc at 20 °C for 20 min to give II-1e (8 g, crude) as white solid. Preparation of 9H-fluoren-9-ylmethylN-[(1R)-2-[(2S)-2-[[(1S)-1-[[4-(hydroxy methyl) phenyl]carbamoyl]-2-methyl-propyl]carbamoyl]pyrrolidin-1-yl] -1-methyl-2-oxo- ethyl]carbamate, II-1f To a solution of (2R)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propanoic acid (2.73 g, 8.77 mmol, 1.4 eq) in DCM (30 mL) was added 1-[Bis(dimethylamino)methylene]-1H-1,2,3- triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate; Hexafluorophosphate Azabenzotriazole Tetramethyl Uronium, HATU (2.50 g, 6.57 mmol, 1.05 eq), 4-methylmorpholine (1.90 g, 18.8 mmol, 2.07 mL, 3.0 eq) and II-1e (2 g, 6.26 mmol, 1.0 eq), and then stirred at 20°C for 2 h. The mixture was diluted with water (40 mL) and extracted with DCM (30 mL x 3). The organic layer was washed with brine (30 mL), dried over Na 2 SO 4 , filtered and concentrated. The residue was purified by flash silica gel chromatography (ISCO®; 3.5 g SepaFlash® Silica Flash Column, Eluent of 0~100% Ethyl acetate/Petroleum ether gradient at 45 mL/min) to give II-1f (1.8 g, 2.94 mmol, 46.92% yield) as white solid. 1 H NMR (MeOD, 400 MHz) δ 7.83-7.71 (m, 4H), 7.56-7.45 (m, 2H), 7.39-7.20 (m, 6H), 4.53-4.41 (m, 4H), 4.05-3.93 (m, 2H), 3.82-3.64 (m, 2H), 3.18-3.08 (m, 1H), 2.53-2.42 (m, 1H), 2.38-2.26 (m, 1H), 2.10-2.04 (m, 2H), 1.37 (d, J = 6.8 Hz, 3H), 1.06-0.96 (m, 6H) Preparation of [4-[[(2S)-2-[[(2S)-1-[(2R)-2-(9H-fluoren-9- ylmethoxycarbonylamino)propanoyl]pyrrolidine-2-carbonyl]amin o]-3-methyl- butanoyl]amino]phenyl]methyl (4-nitrophenyl) carbonate, II-1g To a solution of II-1f (1.8 g, 2.94 mmol, 1.0 eq) in DMF (15 mL) was added DIEA (569 mg, 4.41 mmol, 767 uL, 1.5 eq) and bis(4-nitrophenyl) carbonate (1.07 g, 3.53 mmol, 1.2 eq) and then stirred at 20°C for 12 h. The mixture was diluted with water (50 mL) and extracted with EtOAc (30 mL x 3). The organic layer was washed with brine (30 mL), dried over Na 2 SO 4 , filtered and concentrated. The residue was purified by flash silica gel chromatography (ISCO ® ; 4 g SepaFlash® Silica Flash Column, Eluent of 0 to 100% Ethyl acetate/Petroleum ether gradient at 45 mL/min) to give II-1g (1.1 g, 1.41 mmol, 48.14% yield) as light yellow solid. 1 H NMR (MeOD, 400 MHz) δ 8.57 (s, 1H), 8.19-8.10 (m, 2H), 8.00 (d, J = 8.4 Hz, 2H), 7.74-7.63 (m, 2H), 7.38-7.25 (m, 7H), 7.23-7.14 (m, 3H), 7.08 (d, J = 10.0 Hz, 1H), 5.18-5.16 (m, 2H), 4.77 (dd, J = 10.0, 4.0 Hz, 1H), 4.64 (dd, J = 8.4, 3.6 Hz, 1H), 4.44-4.32 (m, 1H), 4.10-4.03 (m, 1H), 3.98-3.82 (m, 2H), 3.68-3.55 (m, 2H), 2.79-2.67 (m, 1H), 2.41-2.21 (m, 2H), 2.17-2.06 (m, 2H), 1.46 (d, J = 7.2 Hz, 3H), 0.97 (d, J = 7.2 Hz, 6H) Preparation of II-1l Preparation of tert-butyl ((1-((3-bromophenyl)sulfonyl)azetidin-3-yl)methyl)carbamate, II-1h To a mixture of tert-butyl N-(azetidin-3-ylmethyl)carbamate (1.6 g, 8.59 mmol, 1.2 eq) in DCM (5 mL) was added triethylamine, TEA (1.45 g, 14.32 mmol, 1.99 mL, 2 eq) and 3- bromobenzenesulfonyl chloride (1.83 g, 7.16 mmol, 1.03 mL, 1 eq) at 0 °C. The mixture was stirred at 20°C for 1 hr. The mixture was diluted with water (50 mL) and extracted with DCM (25 ml x 3). The organic layer was washed with brine (25 mL), dried over Na2SO4, filtered and concentrated. The residue was purified by flash silica gel chromatography (ISCO ® ; 4 g SepaFlash® Silica Flash Column, Eluent of 0 to 100% Ethyl acetate/Petroleum ether gradient at 35 mL/min). Compound II-1h (2.5 g, 6.17 mmol, 86.16% yield) was obtained as white solid. 1 H NMR (CDCl3, 400 MHz) δ 7.99 (t, J = 4.0 Hz, 1H), 7.74-7.81 (m, 2H), 7.47 (t, J = 8.0 Hz, 1H), 4.61 (s, 1H), 3.86 (t, J = 8.0 Hz, 2H), 3.50-3.58 (m, 2H), 3.19 (t, J = 4.02H), 2.58-2.70 (m, 1H), 1.42 (s, 9H). Preparation of tert-butyl N-[[1-[3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan- 2- yl)phenyl]sulfonylazetidin-3-yl]methyl]carbamate, II-1i To a mixture of II-1h (1 g, 2.47 mmol, 1 eq) in dioxane (10 mL) was added 4,4,4',4',5,5,5',5'-octamethyl-2,2'-bi(1,3,2-dioxaborolane), Pin2B2 (939.80 mg, 3.70 mmol, 1.5 eq) and KOAc (484.29 mg, 4.93 mmol, 2 eq), [1,1′- bis(diphenylphosphino)ferrocene]dichloropalladium(II), Pd(dppf)Cl 2 (90.27 mg, 123.36 µmol, 0.05 eq) at 15°C under N2. The mixture was stirred at 110°C for 2 hrs. The product II-1i was not isolated and used into next step. Preparation of tert-butyl ((1-((3-(2-amino-4-(dipropylcarbamoyl)-3H-benzo[b]azepin-8- yl)phenyl)sulfonyl)azetidin-3-yl)methyl)carbamate, II-1k To a mixture of II-1i (1.12 g, 2.48 mmol, 1 eq) and 2-amino-8-bromo-N,N-dipropyl-3H- 1-benzazepine-4-carboxamide, II-1j (901.90 mg, 2.48 mmol, 1 eq) in dioxane (3 mL) was added K 2 CO 3 (684.35 mg, 4.95 mmol, 2 eq) and Pd(dppf)Cl 2 (90.58 mg, 123.79 µmol, 0.05 eq) at 15°C under N2. The mixture was stirred at 120 °C for 2 hrs. The mixture was filtered and concentrated. The residue was purified by flash silica gel chromatography (ISCO®; 2 g SepaFlash® Silica Flash Column, Eluent of 0 to 100% Ethyl acetate/Petroleum ether gradient @ 60 mL/min) to give II-1k (600 mg, 983.97 µmol, 39.74% yield, 100% purity) as yellow solid. 1 H NMR (MeOD-d4, 400 MHz) δ 7.99-8.10 (m, 2H), 7.74-7.86 (m, 2H), 7.36-7.52 (m, 3H), 6.89 (s, 1H), 3.83 (t, J = 8.0 Hz, 2H), 3.54 (t, J = 8.0 Hz, 2H), 3.34-3.48 (m, 6H), 3.02 (d, J = 8.0 Hz, 2H), 2.48-2.64 (m, 1H), 1.59-1.76 (m, 4H), 1.37 (s, 9H), 0.96-0.89 (m, 6H). LC/MS [M+H] 610.31 (calculated); LC/MS [M+H] 610.40 (observed). Preparation of 2-amino-8-[3-[3-(aminomethyl)azetidin-1-yl]sulfonylphenyl]-N ,N- dipropyl-3H-1-benzazepine-4-carboxamide, II-1l To a solution of II-1k (0.15 g, 245.99 µmol, 1 eq) in DCM (20 mL) was added TFA (56.10 mg, 491.98 µmol, 36.43 µL, 2 eq) at 25 °C and stirred for 1 hour. The mixture was concentrated in reduced pressure at 40 °C. The residue was purified by prep-HPLC (column: Nano-micro Kromasil C 1 8100 x 30mm 5um; mobile phase: [water (0.1%TFA)-ACN]; B%: 25%-50%, 10min) to give, II-1l (0.0546 g, 105.69 µmol, 42.97% yield, 98.66% purity) as a yellow solid. 1 H NMR (MeOD-d4, 400 MHz) δ 8.16-8.07 (m, 2H), 7.92 (d, J = 8.0 Hz, 1H), 7.83 (t, J = 7.6 Hz, 1H), 7.79-7.72 (m, 2H), 7.68 (d, J = 8.4 Hz, 1H), 7.09 (s, 1H), 3.96 (t, J = 8.4 Hz, 2H), 3.67-3.63 (m, 2H), 3.50-3.42 (m, 4H), 3.37 (s, 2H), 3.05 (d, J = 7.4 Hz, 2H), 2.78-2.65 (m, 1H), 1.75-1.66 (m, 4H), 1.08-0.82 (m, 6H). LC/MS [M+H] 510.25 (calculated); LC/MS [M+H] 510.10 (observed). Preparation of [4-[[(2S)-2-[[(2S)-1-[(2R)-2-(9H- fluoren-9- ylmethoxycarbonylamino)propanoyl]pyrrolidine-2-carbonyl]amin o]-3-methyl- butanoyl]amino]phenyl]methyl N-[[1-[3-[2-amino-4-(dipropylcarbamoyl)-3H-1-benzazepin-8- yl] phenyl]sulfonylazetidin-3-yl]methyl]carbamate, II-1m To a mixture of II-1l (100 mg, 196 umol, 1.0 eq) and II-1g (153 mg, 196 umol, 1.0 eq) in DMF (0.5 mL) was added DIEA (50.7 mg, 392 umol, 68.3 uL, 2.0 eq) at 25 °C under N2, and then stirred at 25 °C for 1 hour. The mixture was filtered and purified by prep-HPLC (column: Nano-micro Kromasil C 1 8100*30mm 8um; mobile phase: [water (0.1%TFA)-ACN]; B%: 35%-60%, 10 min) to afford II-1m (70 mg, 60.96 umol, 31.07% yield) as white solid. 1 H NMR (DMSO-d6, 400 MHz) δ12.16 (s, 1H), 10.10 (s, 1H), 9.82 (s, 2H), 9.55 (s, 1H), 9.23 (s, 2H), 8.33 (d, J = 8.4 Hz, 1H), 8.09-8.06 (m, 2H), 7.97 (s, 1H), 7.90-7.48 (m, 14H), 7.42-7.11 (m, 6H), 7.01 (s, 1H), 4.83-4.79 (m, 3H), 4.40-3.93 (m, 9H), 3.79-3.66 (m, 3H), 3.57-3.45 (m, 3H), 3.29 (s, 3H), 2.95-2.83 (m, 3H), 2.20-1.95 (m, 2H), 1.91-1.83 (m, 3H), 1.73-1.67 (m, 1H), 1.61-1.46 (m, 3H), 1.20 (d, J = 6.4 Hz, 3H), 1.05 (d, J = 6.4 Hz, 2H), 0.93-0.66 (m, 12H). LC/MS [M+H] 1148.5 (calculated); LC/MS [M+H] 1148.6 (observed). Preparation of (R)-1-((S)-2-(((S)-1-((4-(((((1-((3-(2-amino-4-(dipropylcarb amoyl)-3H- benzo[b]azepin-8-yl)phenyl)sulfonyl)azetidin-3- yl)methyl)carbamoyl)oxy)methyl)phenyl)amino)-3-methyl-1-oxob utan-2- yl)carbamoyl)pyrrolidin-1-yl)-2-methyl-1,4-dioxo- 7,10,13,16,19,22,25,28,31,34,37,40,43,46,49,52,55,58,61,64,6 7,70,73,76,79-pentacosaoxa-3- azadooctacontan-82-oic acid, II-1n Fmoc-amine, II-1m (0.014 g, 0.015 mmol, 1 eq.) was dissolved in 50% diethylamine in DCM and allowed to sit at room temperature. Upon complete deprotection, the solution was concentrated overnight. To the crude material was added a solution of PEG25-NHS, 79-((2,5- dioxopyrrolidin-1-yl)oxy)-79-oxo- 4,7,10,13,16,19,22,25,28,31,34,37,40,43,46,49,52,55,58,61,64 ,67,70,73,76- pentacosaoxanonaheptacontanoic acid (0.020 g, 0.015 mmol, 1 eq.) in DCM, followed by TEA (0.01 ml, 0.072 mmol, 3 eq.). The reaction mixture was concentrated and purified by HPLC to give II-1n (0.021 g, 0.01 mmol, 65%). LC/MS [M+2H] 1064.57 (calculated); LC/MS [M+2H] 1064.70 (observed). Preparation of II-1: Acid, II-1n (0.021 g, 0.0097 mmol, 1 eq.) and 2,3,5,6- tetrafluorophenol, TFP (0.0032 g, 0.019 mmol, 2 eq.) were dissolved in 1 ml dimethylformamide, DMF. Collidine (trimethylpyridine, 0.006 ml, 0.048 mmol, 5 eq.) was added, followed by EDC-HCl (0.0056 g, 0.029 mmol, 3 eq.). The reaction was stirred at room temperature and monitored by LCMS, then purified by HPLC to give tetrafluorophenol, TFP ester, II-1 (0.0097 g, 0.0043 mmol, 44%). LC/MS [M+2H] 1138.57 (calculated); LC/MS [M+2H] 1138.73 (observed). Example 2 Synthesis of 2,3,5,6-tetrafluorophenyl (R)-1-((S)-2-(((S)-1-((4-(((((1-((3- (2-amino-4-((3-((tert-butoxycarbonyl)amino)propyl)(propyl)ca rbamoyl)-3H-benzo[b]azepin-8- yl)phenyl)sulfonyl)azetidin-3-yl)methyl)carbamoyl)oxy)methyl )phenyl)amino)-3-methyl-1- oxobutan-2-yl)carbamoyl)pyrrolidin-1-yl)-2-methyl-1,4-dioxo- 7,10,13,16,19,22,25,28,31,34,37,40,43,46,49,52,55,58,61,64,6 7,70,73,76,79-pentacosaoxa-3- azadooctacontan-82-oate, II-2 Preparation of [4-[[(2S)-2-[[(2S)-1-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonyl amino) propanoyl]pyrrolidine-2-carbonyl]amino]-3-methyl-butanoyl]am ino]phenyl]methyl N-[[1-[3- [2-amino-4-[3-(tert-butoxycarbonylamino)propyl-propyl-carbam oyl]-3H-1-benzazepin-8- yl]phenyl]sulfonylazetidin-3-yl]methyl]carbamate, II-2b To a mixture of tert-butyl N-[3-[[2-amino-8-[3-[3-(aminomethyl)azetidin-1-yl] sulfonylphenyl]-3H-1-benzazepine-4-carbonyl]-propyl-amino]pr opyl]carbamate, II-2a (100 mg, 160 µmol, 1.0 eq) and [4-[[(2S)-2-[[(2S)-1-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonyl amino) propanoyl]pyrrolidine-2-carbonyl]amino]-3-methyl-butanoyl]am ino]phenyl]methyl (4- nitrophenyl) carbonate, II-1g (125 mg, 160 µmol, 1.0 eq) in DMF (0.5 mL) was added DIEA (41.4 mg, 320 umol, 55.7 uL, 2.0 eq) at 25 °C under N2, and then stirred at 25 °C for 0.5 hours. The mixture was filtered and purified by prep-HPLC (column: Nano-micro Kromasil C 1 8 100*30mm 8um;mobile phase: [water(0.1%TFA)-ACN];B%: 35%-60%,10min) to give II-2b (58 mg, 45.90 umol, 28.68% yield) as white solid. 1 H NMR (DMSO-d 6 , 400 MHz) δ12.14 (s, 2H), 10.09 (s, 1H), 9.82 (s, 1H), 9.55 (s, 1H), 9.20 (s, 2H), 8.32 (d, J = 8.8 Hz, 1H), 8.08 (s, 1H), 7.97 (s, 1H), 7.89-7.62 (m, 8H), 7.62-7.49 (m, 4H), 7.41-7.13 (m, 5H), 7.01 (s, 1H), 6.81 (s, 1H), 4.83-4.80 (m, 3H), 4.41-3.96 (m, 7H), 3.78-3.65 (m, 2H), 3.38-3.22 (m, 7H), 2.94-2.86 (m, 5H), 2.19-1.97 (m, 1H), 1.92-1.84 (m, 3H), 1.74-1.61 (m, 3H), 1.58-1.48 (m, 3H), 1.41-1.00 (m, 12H), 0.93-0.75 (m, 9H). LC/MS [M+H] 1263.6 (calculated); LC/MS [M+H] 1263.6 (observed). Preparation of (R)-1-((S)-2-(((S)-1-((4-(((((1-((3-(2-amino-4-((3-((tert- butoxycarbonyl)amino)propyl)(propyl)carbamoyl)-3H-benzo[b]az epin-8- yl)phenyl)sulfonyl)azetidin-3-yl)methyl)carbamoyl)oxy)methyl )phenyl)amino)-3-methyl-1- oxobutan-2-yl)carbamoyl)pyrrolidin-1-yl)-2-methyl-1,4-dioxo- 7,10,13,16,19,22,25,28,31,34,37,40,43,46,49,52,55,58,61,64,6 7,70,73,76,79-pentacosaoxa-3- azadooctacontan-82-oic acid, II-2c Fmoc-amine, II-2b (0.03 g, 0.024 mmol, 1 eq.) was dissolved in 50% diethylamine in DCM and allowed to sit at room temperature. Upon complete deprotection, the solution was concentrated overnight. To the crude material was added a solution of PEG25-NHS, 79-((2,5- dioxopyrrolidin-1-yl)oxy)-79-oxo- 4,7,10,13,16,19,22,25,28,31,34,37,40,43,46,49,52,55,58,61,64 ,67,70,73,76- pentacosaoxanonaheptacontanoic acid (0.031 g, 0.024 mmol, 1 eq.) in DCM, followed by TEA (0.01 ml, 0.072 mmol, 3 eq.). The reaction mixture was concentrated and purified by HPLC to give, II-2c (0.015 g, 0.0066 mmol, 28%). LC/MS [M+2H] 1122.10 (calculated); LC/MS [M+2H] 1122.40 (observed). Preparation of II-2: Acid II-2c (0.015 g, 0.0066 mmol, 1 eq.) and TFP (0.0022 g, 0.013 mmol, 2 eq.) were dissolved in 1 ml DMF. Collidine (0.004 ml, 0.033 mmol, 5 eq.) was added, followed by EDC-HCl; 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride; CAS 25952-53-8 (0.0038 g, 0.020 mmol, 3 eq.). The reaction was stirred at room temperature and monitored by LCMS, then purified by HPLC to give TFP ester, II-2 (0.009 g, 0.0038 mmol, 57%). LC/MS [M+2H] 1196.10 (calculated); LC/MS [M+2H] 1196.40 (observed). Example 3 Synthesis of 2,3,5,6-tetrafluorophenyl 34-((S)-2-(((S)-1-((4-(((((1-((3-(2- amino-4-((3-((tert-butoxycarbonyl)amino)propyl)(propyl)carba moyl)-3H-benzo[b]azepin-8- yl)phenyl)sulfonyl)azetidin-3-yl)methyl)carbamoyl)oxy)methyl )phenyl)amino)-3-methyl-1- oxobutan-2-yl)carbamoyl)pyrrolidin-1-yl)-34-oxo-4,7,10,13,16 ,19,22,25,28,31- decaoxatetratriacontanoate, II-3 Preparation of 9H-fluoren-9-ylmethyl(2S)-2-[[(1S)-2-methyl-1-[[4-[(4-nitrop henoxy) carbonyloxymethyl]phenyl]carbamoyl]propyl]carbamoyl]pyrrolid ine-1-carboxylate, II-3a To a mixture of 9H-fluoren-9-ylmethyl(2S)-2-[[(1S)-1-[[4-(hydroxymethyl) phenyl]carbamoyl]-2-methyl-propyl]carbamoyl]pyrrolidine-1-ca rboxylate, II-1d (1 g, 1.85 mmol, 1.0 eq) in DMF (20 mL) was added DIEA (358 mg, 2.77 mmol, 482 uL, 1.5 eq) and bis(4-nitrophenyl) carbonate (674 mg, 2.22 mmol, 1.2 eq), and then stirred at 25 °C for 16 hr. The reaction mixture was partitioned between EtOAc (50 mL) and water (50 mL). The organic phase was separated, washed with brine (20 mL), dried over Na2SO4, filtered and concentrated. The crude product was purified by column chromatography (SiO 2 , Petroleum ether/Ethyl acetate=1:0-0:1) to give II-3a (550 mg, 778 umol, 42.2% yield). Preparation of 9H-fluoren-9-ylmethyl (2S)-2-[[(1S)-1-[[4-[[1-[3-[2-amino-4-[3-(tert- butoxycarbonylamino)propyl-propyl-carbamoyl]-3H-1-benzazepin -8- yl]phenyl]sulfonylazetidin-3-yl]methylcarbamoyloxymethyl]phe nyl]carbamoyl]-2-methyl- propyl]carbamoyl]pyrrolidine-1-carboxylate, II-3b To a mixture of tert-butyl N-[3-[[2-amino-8-[3-[3-(aminomethyl)azetidin-1-yl] sulfonylphenyl]-3H-1-benzazepine-4-carbonyl]-propyl-amino]pr opyl]carbamate, II-2a (100 mg, 160 umol, 1.0 eq) and II-3a (113 mg, 160 umol, 1.0 eq) in DMF (0.5 mL) was added diisopropylethylamine, DIEA (41.4 mg, 320.1 umol, 55.7 uL, 2.0 eq) at 25 °C. The mixture was stirred at 25°C for 0.5 hours. Then it was filtered and purified by prep-HPLC (column: Nano- micro Kromasil C 1 8100*30mm 8um;mobile phase: [water (0.1%TFA)-ACN]; B%: 35%-65%, 10 min) to afford II-3b (70 mg, 58.70 umol, 36.68% yield) as white solid. 1 H NMR (DMSO-d 6 , 400 MHz) δ12.04 (s, 1H), 10.13-9.99 (m, 2H), 9.81 (s, 1H), 9.09 (s, 2H), 8.20 (d, J = 8.8 Hz, 1H), 8.09 (s, 1H), 7.98 (s, 1H), 7.89-7.70 (m, 5H), 7.68-7.60 (m, 2H), 7.51 (t, J = 8.8 Hz, 2H), 7.42-7.22 (m, 5H), 7.18 (t, J = 8.8 Hz, 2H), 7.02 (s, 1H), 6.81 (s, 1H), 4.82 (s, 2H), 4.56-4.53 (m, 1H), 4.36-4.14 (m, 4H), 4.08-3.92 (m, 2H), 3.74 (t, J = 8.0 Hz, 2H), 3.36-3.22 (m, 6H), 2.93- 2.89 (m, 5H), 2.24-2.20 (m, 1H), 2.03-1.73 (m, 6H), 1.72-1.61 (m, 2H), 1.56-1.52 (m, 2H), 1.41- 1.08 (m, 9H), 0.93-0.60 (m, 9H). LC/MS [M+H] 1192.6 (calculated); LC/MS [M+H] 1192.6 (observed). Preparation of 34-((S)-2-(((S)-1-((4-(((((1-((3-(2-amino-4-((3-((tert- butoxycarbonyl)amino)propyl)(propyl)carbamoyl)-3H-benzo[b]az epin-8- yl)phenyl)sulfonyl)azetidin-3-yl)methyl)carbamoyl)oxy)methyl )phenyl)amino)-3-methyl-1- oxobutan-2-yl)carbamoyl)pyrrolidin-1-yl)-34-oxo-4,7,10,13,16 ,19,22,25,28,31- decaoxatetratriacontanoic acid, II-3c Fmoc-amine, II-3b (0.037 g, 0.031 mmol, 1 eq.) was dissolved in 0.5 ml DMF. Diethylamine (0.1 ml) was added, and the reaction allowed to sit at room temperature for one hour. The reaction mixture was concentrated and the product triturated three times with diethyl ether. To the crude material was added a solution of PEG10-TFP, 34-oxo-34-(2,3,5,6- tetrafluorophenoxy)-4,7,10,13,16,19,22,25,28,31-decaoxatetra triacontanoic acid (0.0188 g, 0.027 mmol, 0.86 eq.) in acetonitrile, ACN, followed by triethylamine, TEA (0.022 ml, 0.16 mmol, 5 eq.) and HOAt (0.0017 g, 0.012 mmol, 0.4 eq.). Upon completion, the reaction mixture was diluted with water and the product purified by HPLC to give II-3c (0.016 g, 0.01 mmol, 33%). LC/MS [M+H] 1510.76 (calculated); LC/MS [M+H] 1511.14 (observed). Preparation of II-3: Acid II-3a (0.0156 g, 0.0103 mmol, 1 eq.) and TFP (0.004 g, 0.024 mmol, 2.3 eq.) were dissolved in 1 ml ACN. Collidine (0.0068 ml, 0.051 mmol, 5 eq.) was added, followed by EDC-HCl (0.0045 g, 0.023 mmol, 2.23 eq.). The reaction was stirred at room temperature and monitored by LCMS, then purified by HPLC to give TFP ester, II-3 (0.0097 g, 0.0058 mmol, 57%). LC/MS [M+H] 1658.56 (calculated); LC/MS [M+H] 1659.15 (observed). Example 4 Synthesis of [4-[[(2S)-2-[[(2S)-1-[(2R)-2-(9H-fluoren-9- ylmethoxycarbonylamino)propanoyl]pyrrolidine-2-carbonyl]amin o]-3-methyl- butanoyl]amino]phenyl]methyl 4-[[5-amino-7-(dipropylcarbamoyl)-6H-thieno[3,2-b]azepin-2- yl]methyl] piperazine-1-carboxylate, II-4
To a mixture of 5-amino-2-(piperazin-1-ylmethyl)-N,N-dipropyl-6H-thieno[3,2- b]azepine-7-carboxamide, II-4a (100 mg, 162 umol, 1.0 eq, 2 TFA) and [4-[[(2S)-2-[[(2S)-1- [(2R)-2- (9H-fluoren-9-ylmethoxycarbonylamino)propanoyl]pyrrolidine-2 -carbonyl]amino]-3- methyl-butanoyl]amino]phenyl]methyl (4-nitrophenyl) carbonate, II-1g (139 mg, 178 umol, 1.1 eq) in DMF (1 mL) was added DIEA (62.8 mg, 486 umol, 84.6 uL, 3.0 eq) at 20 °C under N2, and then stirred at 20 °C for 1 hour. The mixture was filtered and purified by prep-HPLC (column: Phenomenex Luna C 1 8150*30mm*5um; mobile phase: [water (0.1% TFA)-ACN]; B%: 35%-65%, 8 min) to afford II-4 (24.5 mg, 23.40 umol, 14.45% yield, 98.19% purity) as white solid. 1 H NMR (MeOD, 400 MHz) δ7.91 (d, J = 8.4 Hz, 2H), 7.75 (dd, J = 7.6, 3.6 Hz, 2H), 7.57 (d, J = 8.0 Hz, 2H), 7.43-7.31 (m, 6H), 7.30-7.20 (m, 3H), 7.12 (s, 1H), 5.03 (s, 2H), 4.58-4.50 (m, 2H), 4.46 (q, J = 6.8 Hz, 1H), 4.38 (s, 2H), 4.07-3.98 (m, 2H), 3.78-3.69 (m, 2H), 3.62-3.56 (m, 1H), 3.53-3.43 (m, 6H), 3.41 (s, 3H), 3.11-2.83 (m, 4H), 2.60-2.48 (m, 1H), 2.44- 2.31 (m, 1H), 2.17-2.03 (m, 3H), 1.75-1.63 (m, 4H), 1.42 (d, J = 6.8 Hz, 3H), 1.03-0.87 (m, 12H). LC/MS [M+H] 1028.5 (calculated); LC/MS [M+H] 1028.5 (observed). Example 5 Synthesis of [4-[[(2S)-2-[[(2S)-1-[(2R)-2-(9H-fluoren-9- ylmethoxycarbonylamino)propanoyl]pyrrolidine-2-carbonyl]amin o]-3-methyl- butanoyl]amino]phenyl]methyl N-[4-[[(1S)-1-(2-hydroxyethyl) pentyl]amino]quinazolin-2- yl]carbamate, II-5
Preparation of N4-[(1S)-1-[2-[tert-butyl(dimethyl)silyl]oxyethyl] pentyl]quinazoline-2,4- diamine, II-5b To a mixture of (3S)-3-[(2-aminoquinazolin-4-yl)amino]heptan-1-ol, II-5a (0.69 g, 2.51 mmol, 1.0 eq) in DCM (10 mL) was added imidazole (514 mg, 7.54 mmol, 3.0 eq) and tert- butyldimethylsilyl chloride, TBSCl (379 mg, 2.51 mmol, 1.0 eq) at 0 °C, and then stirred at 15°C for 10hr. The mixture was washed by water (10mL). The organic layer was dried over Na2SO4, filtered and concentrated. The residue was purified by column chromatography (SiO 2 , Petroleum ether/Ethyl acetate=1:0 to 0/1) to give II-5b (400 mg, 1.03 mmol, 40.93% yield) as yellow solid. 1 H NMR (DMSO-d6, 400 MHz) δ8.08 (d, J = 8.8 Hz, 1H), 7.49 (t, J = 7.2 Hz, 1H), 7.40 (d, J = 8.8 Hz, 1H), 7.23 (d, J = 8.0 Hz, 1H), 7.05 (t, J = 7.2 Hz, 1H), 5.92 (s, 2H), 3.73- 3.64 (m, 2H), 3.22-3.21 (m, 1H), 1.89-1.78 (m, 2H), 1.65-1.61 (m, 2H), 1.34-1.31 (m, 4H), 0.90- 0.86 (m, 12H), 0.00 (s, 6H). Preparation of [4-[[(2S)-2-[[(2S)-1-[(2R)-2-aminopropanoyl]pyrrolidine-2- carbonyl]amino]-3-methyl-butanoyl]amino]phenyl]methyl N-[4-[[(1S)-1-[2-[tert-butyl (dimethyl)silyl]oxyethyl]pentyl]amino] quinazolin-2-yl]carbamate, II-5c To a mixture of II-5b (115 mg, 296 umol, 1.0 eq) in THF (8 mL) was added LiHMDS (1 M, 2.96 mL, 10 eq) at -78 °C, and then stirred at this temperature for 0.5 hr, [4-[[(2S)-2-[[(2S)- 1-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propanoyl]pyr rolidine-2-carbonyl]amino]-3- methyl-butanoyl]amino]phenyl]methyl (4-nitrophenyl) carbonate, II-1g (230 mg, 296 umol, 1.0 eq) was added and it was stirred at -78 °C for 2 hr. The reaction was quenched by aq NH 4 Cl (15 mL) and extracted with EtOAc (10mL x 3), the organic layer was dried over Na 2 SO 4 and concentrated to give II-5c (0.3 g, crude) as yellow solid. Preparation of [4-[[(2S)-2-[[(2S)-1-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonyl amino) propanoyl]pyrrolidine-2-carbonyl]amino]-3-methyl-butanoyl]am ino]phenyl]methyl N-[4- [[(1S)-1-[2-[tert-butyl(dimethyl)silyl]oxyethyl]pentyl]amino ]quinazolin-2-yl]carbamate, II-5d To a mixture of II-5c (290 mg, 360 umol, 1.0 eq) in DCM (10 mL) was added imidazole (73.6 mg, 1.08 mmol, 3.0 eq) and Fmoc-Cl; 9H-fluoren-9-ylmethyl carbonochloridate (93.2 mg, 360 umol, 1.0 eq) at 0 °C, and then stirred at 15 °C for 10 hr. The reaction mixture was washed by water (5mL), and the organic layer was dried over Na2SO4, filtered and concentrated to give II-5d (0.37 g, 360 umol, 99% yield) as yellow oil. Preparation of II-5: To a mixture of II-5d (300 mg, 292 umol, 1.0 eq) in MeOH (5 mL) was added acetyl chloride (22.9 mg, 292 umol, 20.8 uL, 1.0 eq) at 0°C, and then stirred at 15 °C for 2hr. The mixture was concentrated to give a residue, the residue was purified by prep-HPLC (column: Phenomenex Synergi C 1 8150*25*10um; mobile phase: [water(0.1%TFA)-ACN]; B%: 40%-56%, 10 min) to obtain (30.4 mg, 28.66 umol, 9.82% yield, 96.84% purity, TFA) as white solid 1 H NMR (MeOD, 400 MHz) δ8.83 (s, 1H), 8.15 (d, J = 8.0 Hz, 1H), 7.91 (d, J = 8.4 Hz, 2H), 7.77-7.63 (m, 1H), 7.54-7.48 (m, 1H), 7.47-7.36 (m, 3H), 7.33-7.17 (m, 3H), 7.10-6.97 (m, 4H), 5.23-5.02 (m, 2H), 4.61-4.28 (m, 4H), 3.99-3.73 (m, 2H), 3.65-3.26 (m, 5H), 2.49-2.16 (m, 2H), 2.10-1.57 (m, 7H), 1.44-1.16 (m, 7H), 1.02-0.58 (m, 9H). LC/MS [M+H] 913.3 (calculated); LC/MS [M+H] 913.3 (observed). Example 6 Synthesis of [4-[[(2S)-2-[[(2S)-1-[(2R)-2-(9H-fluoren-9- ylmethoxycarbonylamino)propanoyl]pyrrolidine-2-carbonyl]amin o]-3-methyl- butanoyl]amino]phenyl]methyl 4-[[5-amino-7-[3-(3,3-dimethyl butanoylamino)propyl-propyl- carbamoyl]-6H-thieno[3,2-b]azepin-2-yl]methyl]piperazine-1-c arboxylate, II-6
To a solution of 5-amino-N-[3-(3,3-dimethylbutanoylamino)propyl]-2-(piperazin -1- ylmethyl)-N-propyl-6H-thieno[3,2-b]azepine-7-carboxamide, II-6a (100 mg, 174 umol, 1.0 eq, 2HCl) and DIEA (89.8 mg, 695 umol, 121 uL, 4.0 eq) in DMF (0.5 mL) was added [4-[[(2S)-2- [[(2S)-1-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propan oyl]pyrrolidine-2- carbonyl]amino]-3-methyl-butanoyl]amino]phenyl]methyl (4-nitrophenyl) carbonate, II-1g (133 mg, 170 umol, 1.0 eq). The mixture was stirred at 20 °C for 1 hr under N 2 atmosphere. The reaction mixture was filtered and purified by prep-HPLC (column: Phenomenex Synergi C 1 8 150*25*10um; mobile phase: [water (0.1%TFA)-ACN]; B%: 35%-45%, 7 min) to give II-6 (58.3 mg, 51.08 umol, 29.40% yield) as white solid. 1 H NMR (MeOD, 400 MHz) δ7.89 (d, J = 8.4 Hz, 2H), 7.71 (dd, J = 4.0, 7.6 Hz, 2H), 7.54 (d, J = 8.0 Hz, 1H), 7.38-7.28 (m, 5H), 7.27- 7.18 (m, 2H), 7.13 (d, J = 5.2 Hz, 2H), 4.99 (s, 2H), 4.56-4.48 (m, 2H), 4.43 (q, J = 7.2 Hz, 1H), 4.21-4.05 (m, 2H), 4.04-3.94 (m, 2H), 3.75-3.65 (m, 2H), 3.59-3.47 (m, 4H), 3.46-3.35 (m, 6H), 3.21-3.17 (m, 2H), 2.86-2.62 (m, 2H), 2.55-2.49 (m, 1H), 2.40-2.28 (m, 1H), 2.15-1.97 (m, 5H), 1.88-1.78 (m, 2H), 1.71-1.59 (m, 2H), 1.39 (d, J = 6.8 Hz, 3H), 1.01-0.97 (m, 9H), 0.92-0.86 (m, 3H). LC/MS [M+H] 1141.6 (calculated); LC/MS [M+H] 1141.5 (observed). Example 7 Synthesis of [4-[[(2S)-2-[[(2S)-1-[(2S)-2-[3-[2-[2-[2-[2-[2-[2-[2-[2-[2-[ 2- [[2-(2,5-dioxopyrrol-1- yl)acetyl]amino]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]et hoxy]ethoxy]ethoxy]ethoxy]pro panoylamino]propanoyl]pyrrolidine-2-carbonyl]amino]-3-methyl - butanoyl]amino]phenyl]methyl 4-[3-[[(2E)-6-carbamoyl-3-[(E)-4-[(2E)-5-carbamoyl-2-(2-ethy l- 5-methyl-pyrazole-3-carbonyl)imino-7-methoxy-3H-benzimidazol -1-yl]but-2-enyl]-2-(2-ethyl- 5-methyl-pyrazole-3-carbonyl)imino-1H-benzimidazol-4-yl]oxy] propyl]piperazine-1- carboxylate, II-7 V
Preparation of (S)-1-((9H-fluoren-9-yl)methyl) 2-(2,5-dioxopyrrolidin-1-yl) pyrrolidine- 1,2-dicarboxylate, 7b To a solution of (2S)-1-(9H-fluoren-9-ylmethoxycarbonyl)pyrrolidine-2-carboxy lic acid, 7a (15 g, 44.5 mmol, 1.0 eq) in DCM (200 mL) was added 1-hydroxypyrrolidine-2,5-dione (5.12 g, 44.5 mmol, 1.0 eq) and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, EDCI, EDAC, EDC, CAS Reg. No.25952-53-8 (10.2 g, 53.44 mmol, 1.2 eq), and it was stirred at 20°C for 12 h. The mixture was washed by saturated aqueous solution of NaHCO 3 (70 mL x 3). The organic layer was dried over Na 2 SO 4 and concentrated to give 7b (17.5 g, 40.28 mmol, 90.60% yield) as a white solid. Preparation of (2S)-2-[[(2S)-1-(9H-fluoren-9-ylmethoxycarbonyl)pyrrolidine- 2- carbonyl]amino]-3-methyl-butanoic acid, 7c To a solution of (2S)-2-amino-3-methyl-butanoic acid, L-valine (4.95 g, 42.3 mmol, 1.05 eq) in THF (200 mL) was added NaHCO 3 (3.55 g, 42.3 mmol, 1.64 mL, 1.05 eq) in H 2 O (50 mL) and 7b (17.5 g, 40.28 mmol, 1.0 eq) and it was stirred at 20°C for 12 h. The mixture was extracted with MTBE (2 x 100 mL) (discarded). The pH of aqueous layer was adjusted to 5-6 with HCl(6 M) and extracted with EtOAc (3 x 200 ml). The combined organic layer was dried over Na2SO4 and concentrated to give 7c (15 g, 34.36 mmol, 85.31% yield) as white solid. 1 H NMR (MeOD, 400 MHz) δ7.80 (d, J = 7.2 Hz, 2H), 7.70-7.54 (m, 2H), 7.43-7.28 (m, 4H), 4.49- 4.15 (m, 5H), 3.69-3.38 (m, 2H), 2.42-2.01 (m, 3H), 2.00-1.82 (m, 2H), 1.01-0.86 (m, 6H). Preparation of 9H-fluoren-9-ylmethyl(2S)-2-[[(1S)-1-[[4-(hydroxymethyl)phen yl] carbamoyl]-2-methyl-propyl]carbamoyl]pyrrolidine-1-carboxyla te, 7d To a solution 7c (10 g, 22.9 mmol, 1.0 eq) and (4-aminophenyl)methanol (4.23 g, 34.4 mmol, 1.5 eq) in MeOH (80 mL) and DCM (80 mL) was added 2-ethoxy-1-ethoxycarbonyl-1,2- dihydroquinoline, EEDQ, CAS Reg. No.16357-59-8 (8.50 g, 34.36 mmol, 1.5 eq) and then stirred at 20 °C for 12 h. The mixture was concentrated in vacuum to give a residue and the residue was purified by flash silica gel chromatography (Silica Flash Column, Eluent of 0-40% Ethyl acetate/MeOH @ 65 mL/min) to give 7d (13 g, 24.0 mmol, 52.38% yield) as yellow solid. 1 H NMR (MeOD, 400 MHz) δ7.85-7.71 (m, 2H), 7.68-7.48 (m, 3H), 7.47-7.16 (m, 7H), 4.53 (d, J = 15.2 Hz, 2H), 4.49-4.41 (m, 1H), 4.40-4.33 (m, 2H), 4.32-4.27 (m, 1H), 4.26-4.17 (m, 1H), 4.16-4.07 (m, 1H), 3.69-3.38 (m, 2H), 2.40-2.05 (m, 2H), 1.99-1.82 (m, 2H), 1.08-0.88 (m, 6H). Preparation of (2S)-N-[(1S)-1-[[4-(hydroxymethyl)phenyl]carbamoyl]-2-methyl - propyl]pyrrolidine-2-carboxamide, 7e To a solution of 7d (13 g, 24.0 mmol, 1.0 eq) in DCM (130 mL) was added piperidine (10.22 g, 120 mmol, 11.85 mL, 5.0 eq) and then stirred at 20°C for 2 h. The mixture was concentrated to give a residue and the residue was triturated with EtOAc at 20°C for 20 min to give 7e (8 g, crude) as white solid. Preparation of 9H-fluoren-9-ylmethyl N-[(1S)-2-[(2S)-2-[[(1S)-1-[[4-(hydroxymethyl) phenyl]carbamoyl]-2-methyl-propyl]carbamoyl]pyrrolidin-1-yl] -1-methyl-2-oxo- ethyl]carbamate, 7f To a solution of (2S)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propanoic acid (3.95 g, 12.68 mmol, 1.5 eq) in DMF (30 mL) was added 1-bBis(dimethylamino)methylene]-1H-1,2,3- triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate, Hexafluorophosphate Azabenzotriazole Tetramethyl Uronium, HATU, CAS Reg. No.148893-10-1 (4.82 g, 12.68 mmol, 1.5 eq) and DIPEA (3.28 g, 25.36 mmol, 4.42 mL, 3 eq) at 0°C. After addition, the mixture was stirred at this temperature for 5min, and then 7e (2.7 g, 8.45 mmol, 1 eq) was added at 0 °C and then the resulting mixture was stirred at 0°C for 25 min. The reaction mixture was quenched by addition of H 2 O (150 mL) and then extracted with EtOAc (70 mL x 3). The combined organic layers were washed with brine (50 mL x 3), dried over Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by column chromatography (SiO 2 , Petroleum ether:Ethyl acetate = 1:0 to 0:1) and then (SiO 2 , EtOAc:MeOH = 1:0 to 10:1) to give 7f (2.94 g, 4.80 mmol, 56.76% yield) as an off-white solid. 1 H NMR (MeOD-d4, 400MHz) δ7.79 (d, J = 7.6 Hz, 2H), 7.66 (t, J = 6.4 Hz, 2H), 7.54 (d, J = 8.4 Hz, 2H), 7.39 (t, J = 7.2 Hz, 2H), 7.35-7.26 (m, 4H), 4.59-4.51 (m, 3H), 4.50-4.40 (m, 1H), 4.39-4.30 (m, 2H), 4.29-4.18 (m, 2H), 3.83-3.71 (m, 1H), 3.68-3.63 (m, 1H), 2.31-2.09 (m, 2H), 2.07-1.91 (m, 3H), 1.36 (dd, J = 6.4, 9.6 Hz, 4H), 1.03 (dd, J = 4.0, 6.8 Hz, 6H). Preparation of [4-[[(2S)-2-[[(2S)-1-[(2S)-2-(9H-fluoren-9-ylmethoxycarbonyl amino) propanoyl]pyrrolidine-2-carbonyl]amino]-3-methyl-butanoyl]am ino]phenyl]methyl (4- nitrophenyl) carbonate, 7g To a solution of 9H-fluoren-9-ylmethyl N-[(1S)-2-[(2S)-2-[[(1S)-1-[[4- (hydroxymethyl)phenyl]carbamoyl]-2-methyl-propyl]carbamoyl]p yrrolidin-1-yl]-1-methyl-2- oxo-ethyl]carbamate (2.4 g, 3.92 mmol, 1 eq) in DMF(20 mL) was added bis(4-nitrophenyl) carbonate (2.38 g, 7.83 mmol, 2 eq)and DIPEA (1.01 g, 7.83 mmol, 1.36 mL, 2 eq) and then stirred at 20°C for 1 h. The reaction mixture was quenched by addition of H 2 O (100 mL) at 0°C, and then extracted with EtOAc (80 mL x 3). The combined organic layers were washed with brine (50 mL x 3), dried over Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by column chromatography (SiO 2 , Petroleum ether:Ethyl acetate = 1:0 to 0:1) to give 7g (2.7 g, 3.47 mmol, 88.62% yield) as a white solid. 1 H NMR (CDCl 3, 400MHz) δ 8.32-8.22 (m, 3H), 7.78 (d, J = 7.6 Hz, 2H), 7.67 (br d, J = 8.4 Hz, 2H), 7.60 (br d, J = 7.6 Hz, 2H), 7.46-7.29 (m, 8H), 7.18 (br d, J = 8.4 Hz, 1H), 5.60 (br d, J = 7.6 Hz, 1H), 5.25 (s, 2H), 4.70-4.66 (m, 1H), 4.62-4.52 (m, 1H), 4.46-4.28 (m, 3H), 4.28-4.19 (m, 1H), 3.81-3.68 (m, 1H), 3.62-3.58 (m, 1H), 2.48-2.29 (m, 2H), 2.19-1.98 (m, 3H), 1.41 (d, J = 7.2 Hz, 3H), 1.10-0.94 (m, 6H). LC/MS [M+H] 778.3 (calculated); LC/MS [M+H] 778.2 (observed). Preparation of tert-butyl 3-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[[2-(2,5-dioxopyrrol-1-yl) acetyl]amino]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethox y]ethoxy]ethoxy]ethoxy]propa noate, 7i To a solution of tert-butyl 3-[2-[2-[2-[2-[2-[2-[2-[2-[2-(2- aminoethoxy)ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy ]ethoxy]ethoxy]propanoate, 7h (11.3 g, 19.3 mmol, 1 eq), 2-(2,5-dioxopyrrol-1-yl)acetic acid (3 g, 19.3 mmol, 1 eq) and DIPEA (10.0 g, 77.4 mmol, 13.5 mL, 4 eq) in DCM (100 mL) was added HATU (8.09 g, 21.3 mmol, 1.1 eq) at 0°C and then the mixture was stirred at 0 °C for 30 min. The reaction mixture was concentrated under reduced pressure. The residue was purified by prep-HPLC (TFA condition; column: Phenomenex luna c18250mm*100mm*10um;mobile phase: [water(0.1%TFA)-ACN];B%: 25%-55%,25min) to give 7i (4.5 g, 6.23 mmol, 32.2% yield) as a yellow oil. 1 H NMR (CDCl 3 , 400 MHz) δ6.88-6.80 (m, 1H), 6.78 (s, 2H), 4.22 (s, 2H), 3.77- 3.54 (m, 40H), 3.47 (q, J = 5.2 Hz, 2H), 2.51 (t, J = 6.4 Hz, 2H), 1.46 (s, 9H). Preparation of 3-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[[2-(2,5-dioxopyrrol-1-yl) acetyl]amino]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethox y]ethoxy]ethoxy]ethoxy]propa noic acid, 7j To a solution of 7i (4.5 g, 6.23 mmol, 1 eq) in CH 3 CN (25 mL) and H 2 O (25 mL) was added TFA (5.68 g, 49.8 mmol, 3.69 mL, 8 eq) and then stirred at 80°C for 1 h. The reaction mixture was concentrated under reduced pressure to remove CH 3 CN. The residue was extracted with MTBE (10 mL x 3) and discarded. The water phase was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (TFA condition; column: Phenomenex Luna c18250mm x 100mm x 10um;mobile phase: [water(0.1%TFA)-ACN];B%: 0%-25%,24min) to give 7j (1.6 g, 2.40 mmol, 38.6% yield) as a light yellow oil. 1 H NMR (CDCl3, 400 MHz) δ6.95 (br s, 1H), 6.78 (s, 2H), 4.22 (s, 2H), 3.78 (t, J = 6.4 Hz, 2H), 3.70- 3.63 (m, 36H), 3.60-3.54 (m, 2H), 3.46 (q, J = 5.2 Hz, 2H), 2.61 (t, J = 6.0 Hz, 2H). LCMS (ESI): mass calcd. for C 42 H 43 N 5 O 10 667.3, m/z found 667.2 [M+H] + . Preparation of [4-[[(2S)-2-[[(2S)-1-[(2S)-2-aminopropanoyl]pyrrolidine-2- carbonyl]amino]-3-methyl-butanoyl]amino]phenyl]methyl 4-[3-[[(2E)-6-carbamoyl-3-[(E)- 4- [(2E)-5-carbamoyl-2-(2-ethyl-5-methyl-pyrazole-3-carbonyl)im ino-7-methoxy-3H- benzimidazol-1-yl]but-2-enyl]-2-(2-ethyl-5-methyl-pyrazole-3 -carbonyl)imino-1H- benzimidazol-4-yl]oxy]propyl]piperazine-1-carboxylate, 7l To a solution of (2E)-1-[(E)-4-[(2E)-5-carbamoyl-2-(2-ethyl-5-methyl-pyrazole -3 - carbonyl)imino-7-(3-piperazin-1-ylpropoxy)-3H-benzimidazol-1 -yl]but-2-enyl]-2-(2-ethyl-5- methyl-pyrazole-3-carbonyl)imino-7-methoxy-3H-benzimidazole- 5-carboxamide, 7k (150 mg, 151 umol, 1 eq, 4HCl) and 7g (129 mg, 166 umol, 1.1 eq) in DMF (3.00 mL) was added DIPEA (97.0 mg, 754 umol, 131 uL, 5 eq) at 20°C, the mixture was stirred at this temperature for 2 h, and then piperidine (39.0 mg, 452.35 umol, 45.0 uL, 3 eq) was added. The mixture was stirred at 20 °C for another 2 h. The mixture was filtered and the residue was purified by prep-HPLC (column: Phenomenex Luna 80*30mm*3um;mobile phase: [water(0.1%TFA)-ACN];B%: 20%- 40%,8min) to give 7l (90 mg, 60.26 umol, 39.97% yield, 2TFA) as light yellow solid. 1 H NMR (MeOD, 400 MHz) δ7.67-7.53 (m, 4H), 7.37 (d, J = 8.4 Hz, 2H), 7.25 (dd, J = 1.2, 14.0 Hz, 2H), 6.60 (d, J = 7.6 Hz, 2H), 5.85-5.68 (m, 2H), 5.14 (s, 2H), 5.00 (br s, 5H), 4.64-4.54 (m, 4H), 4.33-4.18 (m, 2H), 3.95 (br t, J = 6.0 Hz, 2H), 3.74-3.58 (m, 5H), 3.25-3.13 (m, 4H), 2.21 (s, 3H), 2.19 (s, 3H), 2.16-1.92 (m, 6H), 1.52 (d, J = 7.2 Hz, 3H), 1.40-1.29 (m, 6H), 1.08-0.98 (m, 6H). Preparation of 7 To a solution 7l (50 mg, 33.5 umol, 1 eq, 2TFA) and 7j (22.0 mg, 33.5 umol, 1 eq) in DMF (1.00 mL) was added Et3N (7.00 mg, 66.9 umol, 9.00 uL, 2 eq) and 1-Propanephosphonic anhydride, T3P, CAS Reg. No.68957-94-8 (32.0 mg, 50.2 umol, 30.0 uL, 50% purity, 1.5 eq), and then stirred at 20°C for 2 h. The mixture was filtered and the residue was purified by prep- HPLC (column: Phenomenex Luna 80*30mm*3um;mobile phase: [water(0.1%TFA)- ACN];B%: 15%-40%,8min) to give 7 (16 mg, 7.47 umol, 22.31% yield, 2TFA) as white solid. 1 H NMR (MeOD, 400 MHz) δ7.68-7.53 (m, 4H), 7.37 (d, J = 8.4 Hz, 2H), 7.30-7.20 (m, 2H), 6.88 (s, 2H), 6.60 (d, J = 12 Hz, 2H), 5.78 (br s, 2H), 5.14 (s, 2H), 4.99 (br s, 4H), 4.68-4.47 (m, 8H), 4.29-4.25 (m, 1H), 4.16 (s, 3H), 3.96 (br t, J = 5.6 Hz, 2H), 3.80 (td, J = 6.8, 9.2 Hz, 2H), 3.74-3.71 (m, 2H), 3.70 (s, 4H), 3.67-3.56 (m, 36H), 3.53 (br t, J = 5.6 Hz, 2H), 3.48 (td, J = 1.6, 3.2 Hz, 1H), 3.36 (br t, J = 5.2 Hz, 2H), 3.23-3.13 (m, 4H), 2.54-2.43 (m, 2H), 2.20 (d, J = 10.4 Hz, 6H), 2.18-2.10 (m, 2H), 2.06-1.94 (m, 5H), 1.40-1.28 (m, 9H), 1.02 (dd, J = 5.2, 6.4 Hz, 6H). LCMS (ESI): mass calcd. for C42H43N5O101913.9, m/z found 1914.0 [M+H] + . Example 201 Preparation of Immunoconjugates (IC) In an exemplary procedure, an antibody is buffer exchanged into a conjugation buffer containing 100 mM boric acid, 50 mM sodium chloride, 1 mM ethylenediaminetetraacetic acid at pH 8.3, using G-25 SEPHADEX TM desalting columns (Sigma-Aldrich, St. Louis, MO). The eluates are then each adjusted to 6 mg/ml using the buffer and then sterile filtered. The antibody at 6 mg/ml is pre-warmed to 30 °C and rapidly mixed with 2-20 (e.g., 7-10) molar equivalents of immunostimulant-elastase substrate, peptide linker compound. The reaction is allowed to proceed for 16 hours at 30 °C and Immunoconjugate A is separated from reactants by running over two successive G-25 desalting columns equilibrated in phosphate buffered saline (PBS) at pH 7.2 to provide the Immunoconjugate (IC) of Table 3. Adjuvant-antibody ratio (DAR) is determined by liquid chromatography mass spectrometry analysis using a C4 reverse phase column on an ACQUITY TM UPLC H-class (Waters Corporation, Milford, Massachusetts) connected to a XEVO TM G2-XS TOF mass spectrometer (Waters Corporation). For conjugation, the antibody may dissolved in a physiological buffer system known in the art that will not adversely impact the stability or antigen-binding specificity of the antibody. Phosphate buffered saline may be used. The immunostimulant-elastase substrate, peptide linker compound is dissolved in a solvent system comprising at least one polar aprotic solvent as described elsewhere herein. In some such aspects, immunostimulant-elastase substrate, peptide linker intermediate is dissolved to a concentration of about 5 mM, 10 mM, about 20 mM, about 30 mM, about 40 mM or about 50 mM, and ranges thereof such as from about 50 mM to about 50mM or from about 10 mM to about 30 mM in pH 8 Tris buffer (e.g., 50 mM Tris). In some aspects, the immunostimulant-elastase substrate, peptide linker intermediate is dissolved in DMSO or acetonitrile, or in DMSO. In the conjugation reaction, an equivalent excess of immunostimulant-elastase substrate, peptide linker intermediate solution is diluted and combined with chilled antibody solution (e.g. from about 1 °C to about 10 °C). The immunostimulant-elastase substrate, peptide linker intermediate solution may suitably be diluted with at least one polar aprotic solvent and at least one polar protic solvent, examples of which include water, methanol, ethanol, n-propanol, and acetic acid. In some particular aspects the thienoazepine-linker intermediate is dissolved in DMSO and diluted with acetonitrile and water prior to admixture with the antibody solution. The molar equivalents of immunostimulant- elastase substrate, peptide linker intermediate to antibody may be about 1.5:1, about 3:1, about 5:1, about 10:1 about 15:1 or about 20:1, and ranges thereof, such as from about 1.5:1 to about 20:1 from about 1.5:1 to about 15:1, from about 1.5:1 to about 10:1, from about 3:1 to about 15:1, from about 3:1 to about 10:1, from about 5:1 to about 15:1 or from about 5:1 to about 10:1. The reaction may suitably be monitored for completion by methods known in the art, such as LC-MS, and the reaction is typically complete in from about 1 hour to about 24 hours. After the reaction is complete, a reagent may be added to the reaction mixture to quench the reaction and/or cap unreacted antibody thiol groups. An example of a suitable capping reagent is ethylmaleimide. Following conjugation, the immunoconjugates may be purified and separated from unconjugated reactants and/or conjugate aggregates by purification methods known in the art such as, for example and not limited to, size exclusion chromatography, hydrophobic interaction chromatography, ion exchange chromatography, chromatofocusing, µLtrafiltration, centrifugal µLtrafiltration, and combinations thereof. For instance, purification may be preceded by diluting the immunoconjugate, such in 20 mM sodium succinate, pH 5. The diluted solution is applied to a cation exchange column followed by washing with, e.g., at least 10 column volumes of 20 mM sodium succinate, pH 5. The conjugate may be suitably eluted with a buffer such as PBS. Example 202 HEK Reporter Assay HEK293 reporter cells expressing human TLR7 or human TLR8 were purchased from Invivogen and vendor protocols were followed for cellular propagation and experimentation. Briefly, cells were grown to 80-85% confluence at 5% CO 2 in DMEM supplemented with 10% FBS, Zeocin, and Blasticidin. Cells were then seeded in 96-well flat plates at 4x10 4 cells/well with substrate containing HEK detection medium and immunostimulatory molecules. Activity of immunostimulatory compounds are measured using a plate reader at 620-655 nm wavelength. Example 203 Assessment of Immunoconjugate Activity In Vitro The RAW 264.7 murine macrophage cell line was cultured according to vendor protocols (Invivogen). RAW 264.7 cells were co-cultured with the human HER2-expressing tumor cell line, HCC1954, and then stimulated with a range of concentrations of trastuzumab- based ISACs, ISAC 1 and ISAC 2 (Figure 9). Stimulated cells were cultured for 18 hours and then assessed for cellular activation based on production of proinflammatory cytokine TNFa. Cell-free supernatant was collected and analyzed by ELISA. This example shows that Immunoconjugates of the invention are effective at eliciting myeloid activation, and therefore are useful for the treatment of cancer. Isolation of Human Antigen Presenting Cells: Human myeloid antigen presenting cells (APCs) were negatively selected from human peripheral blood obtained from healthy blood donors (Stanford Blood Center, Palo Alto, California) by density gradient centrifugation using a ROSETTESEP TM Human Monocyte Enrichment Cocktail (Stem Cell Technologies, Vancouver, Canada) containing monoclonal antibodies against CD14, CD16, CD40, CD86, CD123, and HLA-DR. Immature APCs were subsequently purified to >90% purity via negative selection using an EASYSEP TM Human Monocyte Enrichment Kit (Stem Cell Technologies) without CD16 depletion containing monoclonal antibodies against CD14, CD16, CD40, CD86, CD123, and HLA-DR. Myeloid APC Activation Assay: 2 x 10 5 APCs were incubated in 96-well plates (Corning, Corning, NY) containing Gibco Iscove’s modified Dulbecco’s medium, IMDM (Lonza) supplemented with 10% FBS, 100 U/mL penicillin, 100 µg/mL (micrograms per milliliter) streptomycin, 2 mM L-glutamine, sodium pyruvate, non-essential amino acids, and where indicated, various concentrations of unconjugated (naked) PD-L1 or HER2 antibodies and immunoconjugates of the invention (as prepared according to the Example above). Trastuzumab and avelumab were used as the antibody constructs. Cell-free supernatants were analyzed after 18 hours via ELISA to measure TNF ^ secretion as a readout of a proinflammatory response. Myeloid cell types for use in screening assays: Conventional myeloid cell types include Monocytes, M-CSF MΦ (M0), GM/IL4 DC, ex vivo cDC for TLR7/8 stimulation assays in a Cytokine read out to exclude inactive agonists. Polarized myeloid cells include Monocytes differentiated toward immunosuppressive state such as M2a MΦ (IL4/IL13), M2c MΦ (IL10/TGFb) and tumor-educated monocytes (TEM) – differentiation w/ tumor-conditioned media (786.O, MDA-MB-231, HCC1954) GM/IL6 “MDSC” in a cytokine read out to exclude agonists with limited activity across assays. Tumor-associated myeloid cells include Myeloid cells present in dissociated tumor cell suspensions (Discovery Life Sciences) in an assay for discovery of agonists. Other useful cell lines for screening may include murine cell lines such as the RAW 264.7 described above, bone marrow derived monocytes, bone marrow derived dendritic cells or macrophages, splenic dendritic cells, TAMs (tumor associated macrophages), and myeloid- derived suppressor cells (MDSCs) in the murine setting. All references, including publications, patent applications, and patents, cited herein are hereby incorporated by reference to the same extent as if each reference were individually and specifically indicated to be incorporated by reference and were set forth in its entirety herein.