Hydroxyureamethyl-Acylfulvene for Treating Brain Cancer or CNS Cancer TECHNICAL FIELD

[0001] This application relates to the field of chemistry and oncology. More particularly, this application relates to methods for treating brain cancer using hydroxyureamethyl-acylfulvene.

BACKGROUND

[0002] Glioblastoma multiforme (GB, GBM or glioblastoma), with a median survival rate of around 15 months, ranks amongst the most aggressive of human cancers. The current standard of care for GB consists of de-bulking surgery followed by combined treatments with fractionated ionizing radiation (IR) and the DNA alkylating agent temozolomide (TMZ). The effectiveness of standard therapy with TMZ is limited because sensitivity of GBM to TMZ is dependent upon the methylation status of the DNA repair enzyme 06 Methyl Guanine Methyl Transferase (MGMT).

[0003] Glioblastoma is the most common and aggressive malignancy. Despite advances in therapy, improvement in overall survival has been limited. Patients with GB almost uniformly experience relapse and have a median survival time of only 15 to 20 months despite aggressive treatment with surgery, radiation, and chemotherapy. Surgical removal of the entire tumor is very difficult and impossible in most cases. Glioblastoma have fmger-like tentacles that extend from the main tumor mass into surrounding normal brain tissue. Often surgical excision is limited by the balance between tumor removal and risks to cognitive function, or indeed immediate patient survival. GB can grow or metastasize to other areas in the brain.

[0004] Another reason GB is difficult to treat is that many drugs cannot efficiently cross the “blood-brain barrier” to enter the brain to act on the tumor. Glioblastomas comprise various populations of cells, some of which respond to treatment and others which do not. These other cells linger and spread through the brain, resulting in little long-term success. These and other factors make GB a pernicious and difficult tumor to characterize and treat.

[0005] Accordingly, there is always a need for an improved method and process for treating GB and other brain cancer. DEFINITIONS

[0006] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in art to which the subject matter herein belongs. As used herein, the following definitions are supplied in order to facilitate the understanding of the present invention.

[0007] Amino acid sequence aligned with the amino acid sequence set out in SEQ ID NO: X (when referring to a variant polypeptide) means that the variant amino acid sequence and the amino acid sequence set out in SEQ ID NO: X are aligned by a suitable method which allows comparison of the sequences with each other and identifications of the positions in the amino acid sequence of the variant wherein either the same amino acid is present (identical position), or another amino acid is present (substitution), or one or more extra amino acids are present (insertion or extension) or no amino acid is present (deletion or truncation) if compared with the amino acid sequence set out in SEQ ID NO: X.

[0008] The term “antibody” as used herein includes intact molecules as well as molecules comprising or consisting of fragments thereof, such as, for example, Fab, F(ab')2, Fv and scFv, as well as engineered variants including diabodies, triabodies, mini-bodies and single-domain antibodies which are capable of binding an epitopic determinant. Thus, antibodies may exist as intact immunoglobulins, or as modifications in a variety of forms.

[0009] The terms “patient,” “subject,” “individual,” and “host” refer to either a human or a non-human animal suffering from or suspected of suffering from a disease or disorder associated with aberrant biological or cell growth activity.

[0010] [0024] The term “biomarker” refers to any molecule, such as a gene, gene transcript (for example mRNA), peptide or protein or fragment thereof produced by a subject which is useful in differentiating subjects to predict the responsiveness of patients to treatments including disodium 2,2'-dithio-bis-ethane sulfonate or its analogs. A biomarker that is differentially present (i.e., increased or decreased) in a biological sample from a subject or a group of subjects has a first phenotype (e.g., having a disease) as compared to a biological sample from a subject or group of subjects having a second phenotype (e.g., not having the disease). A biomarker may be differentially present at any level, but is generally present at a level that is increased by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, by at least 100%, by at least 110%, by at least 120%, by at least 130%, by at least 140%, by at least 150%, or more; or is generally present at a level that is decreased by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, or by 100% (i.e., absent). A biomarker is preferably differentially present at a level that is statistically significant (e.g., a p- value less than 0.05 and/or a q-value of less than 0.10 as determined using either Welch's T- test or Wilcoxon's rank-sum Test).

[0011] The term “glioblastoma multiforme” or “GB” or “GBM” refers to the most common and aggressive type of primary brain tumor in humans. GB tumors are characterized by the presence of small areas of necrotizing tissue that is surrounded by anaplastic cells (pseudopabsading necrosis). This characteristic, as well as the presence of hyperplastic blood vessels, differentiates the tumor from Grade 3 astrocytomas, which do not have these features. In some cases, the term “glioblastoma multiforme” or “glioblastoma” “or malignant glioma” are used interchangeably herein and refer to a brain tumor that arises from astrocytes.

[0012] The terms “treat” and “treating” such a disease or disorder refers to ameliorating at least one symptom of the disease or disorder. These terms, when used in connection with a condition such as a cancer, refer to one or more of: impeding growth of the cancer, causing the cancer to shrink by weight or volume, extending the expected survival time of the patient, inhibiting tumor growth, reducing tumor mass, reducing size or number of metastatic lesions, inhibiting the development of new metastatic lesions, prolonging survival, prolonging progression-free survival, prolonging time to progression, and/or enhancing quality of life. The terms “treatment” or “treating” glioblastoma multiforme can include arresting the development or reversing the symptom or symptoms of glioblastoma and/or an improvement in clinical outcome of the patient suffering from glioblastoma or recurrent glioblastoma. Example of improvements in clinical outcome include longer survival time, reduction in tumor size, non growth in tumor size, and/or lack of exacerbation in neurological symptoms. Non-limiting examples of neurological symptoms include double vision, vomiting, loss of appetite, changes in mood and personality, changes in ability to think and learn, seizures, speech difficulty, and cognitive impairment.

[0013] The term “preventing” when used in relation to a condition or disease such as cancer, refers to a reduction in the frequency of, or delay in the onset of, symptoms of the condition or disease. Thus, prevention of cancer includes, for example, reducing the number of detectable cancerous growths in a population of patients receiving a prophylactic treatment relative to an untreated control population, and/or delaying the appearance of detectable cancerous growths in a treated population versus an untreated control population, e.g., by a statistically and/or clinically significant amount.

[0014] The terms “expression level” and “level of expression,” as used herein, refer to the amount of a gene product in a cell, tissue, biological sample, organism, or patient, e.g., amounts of DNA, RNA (e.g. messenger RNA (mRNA)), or proteins corresponding to a given gene. [0015] The term “pharmaceutically acceptable” means that, which is useful in preparing a pharmaceutical composition that is generally safe, non-toxic, and neither biologically nor otherwise undesirable and includes that which is acceptable for veterinary as well as human pharmaceutical use.

[0016] “Pharmaceutically acceptable salt” refers to a salt which is acceptable for administration to a patient, such as a mammal (e.g., salts having acceptable mammalian safety for a given dosage regime). Such salts can be derived from pharmaceutically acceptable inorganic or organic bases and from pharmaceutically-acceptable inorganic or organic acids, depending on the particular substituents found on the compounds described herein. When compounds of the present disclosure contain relatively acidic functionalities, base addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired base, either neat or in a suitable inert solvent. Salts derived from pharmaceutically acceptable inorganic bases include aluminum, ammonium, calcium, copper, ferric, ferrous, lithium, magnesium, manganic, manganous, potassium, sodium, zinc and the like. Salts derived from pharmaceutically acceptable organic bases include salts of primary, secondary, tertiary and quaternary amines, including substituted amines, cyclic amines, naturally-occurring amines and the like, such as arginine, betaine, caffeine, choline, N,N'- dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydrabamine, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine, N,N'-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, meglumine (N-methyl-glucamine) and the like. When compounds of the present disclosure contain relatively basic functionalities, acid addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired acid, either neat or in a suitable inert solvent. Salts derived from pharmaceutically acceptable acids include acetic, trifluoroacetic, propionic, ascorbic, benzenesulfonic, benzoic, camphosulfonic, citric, ethanesulfonic, fumaric, glycolic, gluconic, glucoronic, glutamic, hippuric, hydrobromic, hydrochloric, isethionic, lactic, lactobionic, maleic, malic, mandelic, methanesulfonic, mucic, naphthalenesulfonic, nicotinic, nitric, pamoic, pantothenic, phosphoric, succinic, sulfuric, hydroiodic, carbonic, tartaric, p- toluenesulfonic, pyruvic, aspartic, benzoic, anthranilic, mesylic, salicylic, p-hydroxybenzoic, phenylacetic, embonic (pamoic), ethanesulfonic, benzenesulfonic, 2-hydroxyethanesulfonic, sulfanibc, stearic, cyclohexylaminosulfonic, algenic, hydroxybutyric, galactaric and galacturonic acid and the like.

[0017] The term “therapeutic effect” refers to a beneficial local or systemic effect in animals, particularly mammals, and more particularly humans, caused by administration of a compound or composition of the invention. The phrase “therapeutically-effective amount” means that amount of a compound or composition of the invention that is effective to treat a disease or condition caused by aberrant biological activity at a reasonable benefit/risk ratio. In some embodiments, the therapeutically effective amount of hydroxyureamethyl-acylfulvene or a pharmaceutically acceptable salt thereof is selected from the group consisting of 0.5 mg/day, 1 mg/day, 2.5 mg/day, 5 mg/day, 10 mg/day, 20 mg/day, 30 mg/day, 60 mg/day, 90 mg/day, 120 mg/day, 150 mg/day, 180 mg/day, 210 mg/day, 240 mg/day, 270 mg/day, 300 mg/day, 360 mg/day, 400 mg/day, 440 mg/day, 480 mg/day, 520 mg/day 580 mg/day, 600 mg/day, 620 mg/day, 640 mg/day, 680 mg/day, and 720 mg/day.

[0018] The term “healthy individual” shall be taken to mean an individual who is known not to suffer from cancer (e.g., brain cancer), such knowledge being derived from clinical data on the individual, including, but not limited to, a different diagnostic assay to that described herein.

[0019] A “reference level” means a level of the compound of the present invention or additional biomarker(s) that is indicative of a particular disease state, phenotype, or lack thereof, as well as combinations of disease states, phenotypes, or lack thereof.

[0020] A “reference sample” refers to a sample containing reference level of a biomarker. For example, a reference sample can be obtained from a subject that does not have a particular disease, disease state or phenotype, such as cancer or acute injury.

[0021] The therapeutically effective amount of such substance will vary depending upon the subject and disease condition being treated, the weight and age of the subject, the severity of the disease condition, the manner of administration and the like, which can readily be determined by one of skill in the art.

SUMMARY

[0022] One aspect of this application includes a method of therapeutically treating brain cancer comprising administering an effective amount of hydroxyureamethyl-acylfulvene to a subject in need thereof. In one embodiment, the brain cancer is a glioblastoma multiforme. [0023] Another aspect of this application includes a method for treating brain cancer using an effective amount of hydroxyureamethyl-acylfulvene together with an effective amount of one or more of an additional therapeutic agent selected from the group consisting of temozolomide, bevacizumab, everolimus, carmustine, lomustine, procarbazine, vincristine, irinotecan, cisplatin, carboplatin, methotrexate, etoposide, vinblastine, bleomycin, actinomycin, cyclophosphamide, and ifosfamide.

[0024] Another aspect of this application includes a method for treating brain cancer using an effective amount of hydroxyureamethyl-acylfulvene together with radiation therapy. The radiation therapy can be selected from whole-brain irradiation, fractionated radiotherapy, radio surgery, and a combination thereof.

[0025] Another aspect of this application includes a method in which the patient is human or animal.

BRIEF DESCRIPTION OF THE DRAWINGS

[0026] FIG. 1 shows hydroxyureamethyl-acylfulvene exhibits nanomolar potency in GB cell lines;

[0027] FIG. 2 shows hydroxyureamethyl-acylfulvene exhibits nanomolar potency in additional GB cell lines;

[0028] FIG. 3 shows the analysis of the modulation of the blood-brain barrier by hydroxyureamethyl-acylfulvene;

[0029] FIG. 4 shows the impact on cell viability in the presence of hydroxyureamethyl- acylfulvene;

[0030] FIG. 5 shows the profile of hydroxyureamethyl-acylfulvene activity in vivo in the U87 subcutaneous (Sc) xenograft tumor model of GBM;

[0031] FIG. 6A shows results from mice implanted with GBM neurospheres (Ml 123) treated with a vehicle as a control;

[0032] FIG. 6B shows results from mice implanted with GBM neurospheres (Ml 123) treated with 4, every other day, intravenous doses of 4 mg/kg hydroxyureamethyl-acylfulvene.

[0033] FIG. 7 shows images of physical observations of mice implanted with GBM neurospheres (Ml 123) treated with vehicle control or hydroxyureamethyl-acylfulvene.

SEQUENCES

[0034] SEQ ID NO: 1— amino acid sequence of EGFR,

[0035] SEQ ID NO:2— amino acid sequence of NF1,

[0036] SEQ ID NO:3— amino acid sequence of IDH1, [0037] SEQ ID NO:4--amino acid sequence of LABI, [0038] SEQ ID NO:5~amino acid sequence of IGDH, [0039] SEQ ID NO: 6-amino acid sequence of ANXA2, [0040] SEQ ID NO:7-amino acid sequence of S100A11, [0041] SEQ ID NO:8-amino acid sequence of CTSB, [0042] SEQ ID NO:9-amino acid sequence of TP53, and [0043] SEQ ID NO: 10-amino acid sequence of MGMT

DETAILED DESCRIPTION

[0044] HydroxyUreaMethyl Acylfulvene or hydroxyureamelhyl-acylfulvene (currentiy, termed as LP-184 by Lantern Pharma, Inc.) is a semisynthetic or synthetic antitumor agent derived from the mushroom toxin illudin S. The structure of each isomer is shown below.

[0045] Specific embodiments relate to methods of treating brain cancer, the methods including the administration of an effective amount of hydroxyureamethyl-acylfulvene to a subject in need thereof. In some embodiments, the brain cancer may be from a metastatic brain tumor, glioblastoma multiforme, and a combination thereof. In specific examples, the brain tumor is a glioblastoma multiforme. In one example, hydroxyureamethyl-acylfulvene can be administered as a monotherapy.

[0046] One embodiment includes co-administering hydroxyureamethyl-acylfulvene and an additional therapeutic agent in separate compositions or the same composition. Thus, some embodiments include a first pharmaceutical composition comprising: (a) a safe and therapeutically effective amount of hydroxyureamethyl-acylfulvene or pharmaceutically acceptable salts thereof and (b) a pharmaceutically acceptable carrier, diluent, excipient or combination thereof; and a second pharmaceutical composition comprising: (1) a safe and therapeutically effective amount of an additional therapeutic agent and (2) a pharmaceutically

7

SUBSTITUTE SHEET (RULE 26) acceptable carrier, diluent, excipient or combination thereof. Some embodiments include a pharmaceutical composition comprising: (a) a safe and therapeutically effective amount of an additional therapeutic agent; and (b) a pharmaceutically acceptable carrier, diluent, excipient or combination thereof. In some embodiments, the method described herein can further include subjecting the subject to a radiation therapy. In some embodiments, the radiation therapy can be a whole-brain irradiation, fractionated radiotherapy, and radiosurgery.

[0047] Another embodiment includes a method of inhibiting proliferation of a brain tumor cell, tire method including contacting the brain tumor cell with hydroxyureamethyl-acylfulvene. In some embodiments, the contacting comprises administering an effective amount of hydroxyureamethyl-acylfulvene to a subject having the brain tumor cell. In some embodiments, the brain tumor is a glioblastoma multiforme. In some embodiments, the method can be used to treat primary Central Nervous System (CNS) tumors. CNS cancer is a type of cancer that forms in the central nervous system. It includes brain stem glioma, craniopharyngioma, medulloblastoma and meningioma. CNS tumors start in the normal cells of the brain and spinal cord called “neurons” and “glia.” Tumors that start from neurons include medulloblastoma and primitive neuroectodermal tumors (PNETs). Tumors that start from glia include glioma, astrocytoma, oligodendroglioma, and ependymoma. The tumor's specific name often reflects the CNS tumor's tissue of origin.

[0048] Another embodiment includes a method of therapeutically treating brain cancer or CNS tumors in a subject including obtaining a tumor sample from the subject; measuring the expression of MGMT+ or MGMT- in the tumor sample; and administering an effective amount of hydroxyureamethyl-acylfulvene or a pharmaceutically acceptable salt thereof to a subject in need thereof when MGMT+ measured to be greater than a reference, wherein the reference is the level of MGMT in a healthy person. In one example, the characterization further comprises a molecular subtype classification as a Classical, Mesenchymal, Proneural, or Neural glioblastoma.

[0049] Protein sequences of certain proteins identified herein include EGFR(SEQ ID NO: 1), NF1 (SEQ ID NO: 2), IDH1 (SEQ ID NO: 3), LAMB1 (SEQ ID NO: 4), UGDH (SEQ ID NO: 5), ANXA2 (SEQ ID NO: 6), S100A11 (SEQ ID NO: 7), CTSB (SEQ ID NO: 8), TP53 (SEQ ID NO: 9), and MGMT (SEQ ID NO: 10).

[0050] Some embodiments relate to a method of inducing apoptosis in a brain tumor cell, the method including contacting the brain tumor cell with hydroxyureamethyl-acylfulvene. In some embodiments, the contacting comprises administering an effective amount of hydroxyureamethyl-acylfulvene to a subject having the brain tumor cell, In some embodiments, the brain tumor is a glioblastoma multiforme.

8 [0051] Some embodiments include a treatment for GB that includes (I) conducting a physical exam of the patient; (2) using a magnetic resonance imaging (“MRI”) to confirm the presence of a tumor in the patient’s brain; (3) conducting a brain biopsy to obtain genetic information about the cancer, which includes molecular subtype and genetic markers; and (4) prescribing effective treatment of hydroxyureamethyl-acyifulvene to a subject having GB.

[0052] The administration period can be a multi-week treatment cycle as long as the tumor remains under control and the regimen is clinically tolerated. In some embodiments, a single dosage of hydroxyureamethyl-acyifulvene or other therapeutic agent can be administered once a week, and preferably once on each of day 1 and day 8 of a three- week (21 -day) treatment cycle. In some embodiments, a single dosage of hydroxyureamethyl-acyifulvene or other therapeutic agent can be administered once a week, twice a week, three times per week, four times per week, five times per week, six times per week, or daily during a one-week, two-week, three-week, four-week, or five-week treatment cycle. The administration can be on the same or different day of each week in the treatment cycle.

[0053] Another embodiment includes a method for treating or determining the sensitivity of brain cancer (e.g., Glioblastoma or CNS cancer) to a hydroxyureamethyl-acyifulvene treatment by assessing the level of MGMT expression. A level of MGMT greater than that in a healthy individual indicates that LP-184 can be effective.

[0054] A method of treating brain cancer or CNS cancer that includes detecting, in a human subject, the presence of certain genetic information; administering hydroxyureamethyl- acyifulvene or a pharmaceutically acceptable salt thereof, to the subject, if the human subject overexpress or under expresses certain markers. The marker can be substantially similar to EGFR (SEQ ID NO: 1), NF1 (SEQ ID NO: 2), IDH1 (SEQ ID NO: 3), LAMB1 (SEQ ID NO: 4), UGDH (SEQ ID NO: 5), ANXA2 (SEQ ID NO: 6), S100A11 (SEQ ID NO: 7), CTSB (SEQ ID NO: 8), TP53 (SEQ ID NO: 9), and MGMT (SEQ ID NO: 10).

[0055] Another embodiment includes a method for treating or determining the sensitivity of brain cancer (e.g., Glioblastoma) to a Hydroxyureamethyl-acyifulvene treatment by assessing the level of MGMT expression and/or at least one gene selected from the group consisting of: Laminin Subunit Beta 1 (LAMB1), UDP-Glucose 6-Dehydrogenase (UGDH), Annexin A2 (ANXA2), S100 Calcium Binding Protein All (S100A11), and Cathepsin B (CTSB). The nucleotide sequence and/or amino acid sequences are well known and published in public database (e.g., Genebank) and available papers.

[0056] Another embodiment includes a method of treating brain cancer in a subject, comprising: (a) obtaining or having obtained an expression level in a sample from a subject for a plurality of targets, wherein the plurality of targets comprises the group consisting of MGMT (SEQ ID NO: 10), LAMB1 (SEQ ID NO: 4), UGDH(SEQ ID NO: 5), ANXA2(SEQ ID NO: 6), S100A11(SEQ ID NO: 7), and/or CTSB(SEQ ID NO: 8); (b) determining that the subject is sensitive to a treatment with a Hydroxyureamethyl-acylfulvene; and (c) administering a cancer treatment including a Hydroxyureamethyl-acylfulvene.

[0057] Another embodiment includes a method of treating brain cancer in a subject, comprising: (a) obtaining or having obtained an expression level in a sample from a subject for a plurality of targets, wherein the plurality of targets comprises the group consisting of EGFR, NF1, and/or PDGFRA/IDHI; (b) determining that the subject is sensitive to a treatment with a Hydroxyureamethyl-acylfulvene: and (c) administering a cancer treatment including a Hydroxyureamethyl-acylfulvene.

[0058] Another embodiment includes the detection the genetic information including high- level EGFR amplification, deletions in NF i gen, high expression of MET gene, alternations of PDFRA gene, point mutations in IDH1, TP53 mutation, expression of neuron markers by astrocytes, mutations in oncogenes, MGMT methylation, or a combination thereof.

[0059] Hydroxyureamethyl-acylfulvene for use in accordance with the present invention can be mainly administered by parenteral administration, specifically including subcutaneous administration, intramuscular administration, intravenous administration, transcutaneous administration, intrahecal administration, epidural administration, intra joint administration and local administration, or may also be administered in various dosage forms, for example by oral administration if possible.

[0060] The injections for parenteral administration include for example sterile, aqueous or non- aqueous solutions, suspensions and emulsions. The aqueous solutions and suspensions include for example distilled water for injections and physiological saline. The non-aqueous solutions and suspensions include for example propylene glycol, polyethylene glycol, vegetable oils such as olive oil, alcohols such as ethanol, and Polysorbate 80 (under trade name). Such composition may contain auxiliary agents such as preservatives, moistening agents, emulsifying agents, dispersing agents, stabilizers (for example, lactose) and dissolution auxiliary agents (for example, meglumine). These are sterilized by filtering through bacteria-retaining filters, blending sterilizing agents, or irradiation. Alternatively, these may be produced once into a sterile solid composition and then dissolved or suspended in sterile water or sterile solvents for injections, prior to use.

[0061] In some embodiments, the brain tumor can be selected from metastatic brain tumor, anaplastic astrocytoma, glioblastoma multiforme, oligodendroglioma, ependymomas, meningioma, mixed glioma, and a combination thereof. In some embodiments, the brain tumor is a glioblastoma multiforme. In some embodiments, the brain tumor is a metastatic brain tumor.

[0062] In some embodiments, the brain tumor can be selected from Anaplastic astrocytoma, Central neurocytoma, Choroid plexus carcinoma, Choroid plexus papilloma, Choroid plexus tumor, Dysembryoplastic neuroepithelial tumor, Ependymal tumor, Fibrillary astrocytoma, Giant-cell glioblastoma, Glioblastoma multiforme, Gliomatosis cerebri, Gliosarcoma, Hemangiopericytoma, Medulloblastoma, Medulloepithelioma, Meningeal carcinomatosis, Neuroblastoma, Neurocytoma, Oligoastrocytoma, Oligodendroglioma, Optic nerve sheath meningioma, Pediatric ependymoma, Pilocytic astrocytoma, Pinealoblastoma, Pineocytoma, Pleomorphic anaplastic neuroblastoma, Pleomorphic xanthoastrocytoma, Primary central nervous system lymphoma, Sphenoid wing meningioma, Subependymal giant cell astrocytoma, Subependymoma, central nervous system myeloma, and Trilateral retinoblastoma.

[0063] The liquid composition for oral administration includes for example pharmaceutically acceptable emulsions, liquids, suspensions, syrups and elixirs and contains inert diluents for general use, for example, distilled water and ethanol. The composition may contain auxiliary agents such as moistening agents and suspending agents, sweetening agents, flavoring agents, aromatic agents and preservatives, other than the inert diluents.

[0064] The active compound may also be administered intravenously or intrapentoneally by infusion or injection. Solutions of the active compound can be prepared in water, optionally mixed with a nontoxic surfactant. Dispersions can also be prepared in glycerol, liquid polyethylene glycols, triacetin, and mixtures thereof and in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.

[0065] It should also be understood that a specific dosage and treatment regimen for any particular patient will depend upon a variety of factors, including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, rate of excretion, drug combination, and the judgment of the treating physician and the severity of the particular disease being treated. The amount of a compound of the present invention in the composition will also depend upon the particular compound in the composition.

EXAMPLES

Example 1

[0066] Table 1 shows that LP-184/hydroxyureamethyl-acylfulvene shows enhanced sensitivity in tumors that express MGMT. In addition to MGMT expression, other GB-specific genes also show multi-omic parameter correlations with the possibility that LP-184/hydroxyureamethyl-acylfulvene will show sensitivity to GB.

[0067] Table 1 shows genes that can be used to show sensitivity of GB to hydroxyureamethyl- acylfulvene.

TABLE 1

[0068] Table 2 shows genes downregulated in GB and associated with TMZ resistance that are negatively correlated with LP-184 sensitivity, and thus there can be a benefit of LP-184 in such tumors.

TABLE 2

[0069] Table 3 shows genes whose upregulation are known to promote GB. Tumors with elevated expression of such genes (compared to those in healthy people) can be more sensitive to hydroxyureamethyl-acylfulvene.

TABLE 3

Example 2

[0070] FIG. 1 shows that hydroxy ureamethyl-acylfulvene exhibits nanomolar potency in GB cell lines. The cytotoxicity of LP-184 in terms of IC50 in the 4 cell lines is in the range of 45 to 230 nM.

[0071] The results from a tool for evaluating chemical ADMET (absorption, distribution, metabolism, excretion - toxicity) properties indicates that LP-184 crosses the blood brain barrier. NCI60 data generated drug sensitivity - gene expression correlation analysis of Hydroxyureamethyl-acylfulvene and Mitozolomide, a TMZ precursor molecule, showed that there are no overlapping genes that likely mediate resistance to either drug. Hydroxyureamethyl-acylfulvene can serve as a potential monotherapy or as a combination therapy for GB tumors.

Example 3

[0072] Hydroxyureamethyl-acylfulvene was tested and showed activity in an expanded spectrum of in vitro and in vivo brain cancer models that represent a variety of molecular and clinical subtypes of glioblastomas. Hydroxyureamethyl-acylfulvene was shown to cross the blood-brain barrier in an in vitro model system.

[0073] FIG. 2 shows that Hydroxyureamethyl-acylfulvene exhibits nanomolar potency in various GB cell lines beyond those shown in FIG. 1. The cytotoxicity of hydroxyureamethyl- acylfulvene in terms of IC50 in a set of 6 cell lines was in the range of 46 to 2620 nM.

[0074] FIG. 3 shows the analysis of the modulation of the blood-brain barrier due to compound interaction. Hydroxyureamethyl-acylfulvene had insignificant impact on cell viability and blood-brain barrier integrity was not compromised. To evaluate the in vivo therapeutic activity of Hydroxyureamethyl-acylfulvene in mice bearing glioblastoma U87-SC cells (without any drug treatment in vitro) were inoculated subcutaneously.

[0075] By way of background, the physically, metabolically and immunologically privileged status of the brain presents diagnostic and therapeutic limitations along with treatment-related neurotoxicity issues. A critical property of any drug for GBM would be its ability to penetrate the blood-brain barrier (BBB). Independent in vitro studies assessing the penetration kinetics of Hydroxyureamethyl-acylfulvene showed Hydroxyureamethyl-acylfulvene passing across the blood-brain barrier. In the highly representative Neuromics in vitro 3D model that closely recapitulates the human blood-brain barrier, transport properties of the blood-brain barrier are reflected due to the formation of tight junctions, higher expression of specific carriers, or great cell viability. This 3D in vitro model of the blood-brain barrier created by co-culturing brain endothelial cells with pericytes and astrocytes layered in an insert, improves endothelial cell polarization and enhances the formation of tight junctions, provides better endothelial cell-to- cell contact that is important for barrier development, and prevents the dilution of secreted neurotrophic factors. These conditions collectively lead to the development of an in vitro model that can truly mimic the blood-brain barrier. This assay leverages the novel 3D blood- brain barrier model that allows studying both compound transport across the barrier as well as the effect of compounds on the structure and function of the blood-brain barrier. Along with LP-184 and TMZ, the current standard of care chemotherapeutic for GBM, we also compared the behavior of known positive and negative control agents Antipyrine and Cyclosporin A respectively as benchmarks. LP-184 is as effective as TMZ in penetrating the blood-brain barrier. Apparent blood brain barrier permeability measured for TMZ was 1.72 * 10-4 cm/s at 30 minutes and for LP-184 was 1.53 * 10-4 cm/s at 30 minutes, as illustrated in the following graph.

[0076] FIG. 4 shows the impact on cell viability after treatment with hydroxyureamethyl- acylfulvene.

[0077] FIG. 5 shows the profile of hydroxyureamethyl-acylfulvene activity in vivo in the U87- Sc xenograft tumor model of GB. The mice were treated with 4, every other day, intravenous doses of Hydroxyureamethyl-acylfulvene at 4 mg/kg in the U87-SC model resulted in complete tumor regression with no measurable tumors 12 days post the last dosing in all 5 mice treated. [0078] FIG. 6A and FIG. 6B shows that mice (Ml 123) treated with 4, every other day, intravenous doses of hydroxyureamethyl-acylfulvene at 4 mg/kg resulted in complete tumor regression. FIG. 6A shows results from mice implanted with GBM neurospheres (Ml 123) treated with a vehicle (control). FIG. 6B shows results from mice implanted with GBM neurospheres (Ml 123) treated with 4 every other day intravenous doses of 4 mg/kg hydroxyureamethyl-acylfulvene. Three of four mice showed no measurable tumor after 12 days of the last dose.

[0079] FIG. 7 shows images of physical observations of mice implanted with GBM neurospheres (Ml 123) treated with vehicle control or hydroxyureamethyl-acylfulvene. The tumor volume physically displayed clear tumor growth inhibition in Hydroxyureamethyl- acylfulvene treated mice relative to vehicle-treated control mice. This example shows in vivo efficacy of Hydroxyureamethyl-acylfulvene in subcutaneous tumor models of GB. Protein sequences of EGFR(SEQ ID NO: 1), NF1 (SEQ ID NO: 2), IDH1 (SEQ ID NO: 3), LAMB1 (SEQ ID NO: 4), UGDH (SEQ ID NO: 5), ANXA2 (SEQ ID NO: 6), S100A11 (SEQ ID NO: 7), CTSB (SEQ ID NO: 8), TP53 (SEQ ID NO: 9), and MGMT (SEQ ID NO: 10) from Uniprot database:

SEQUENCE ID NO: 1 (SEQ ID NO: 1)

EGFR/Homo sapiens/Human

MRPSGTAGAALLALLAALCPASRALEEKKVCQGTSNKLTQLGTFEDHFLSLQRMFN

NCEVVLGNLEITYVQRNYDLSFLKTIQEVAGYVLIALNTVERIPLENLQIIRGNMYY E

NSYALAVLSNYDANKTGLKELPMRNLQEILHGAVRFSNNPALCNVESIQWRDIVSSD

FLSNMSMDFQNHLGSCQKCDPSCPNGSCWGAGEENCQKLTKIICAQQCSGRCRGKS

PSDCCHNQCAAGCTGPRESDCLVCRKFRDEATCKDTCPPLMLYNPTTYQMDVNPEG

KY SF GATCVKKCPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRKV CN

GIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVAFRGDSFTHTPPLDPQELDIL KTV

KEITGFLLIQAWPENRTDLHAFENLEIIRGRTKQHGQFSLAVVSLNITSLGLRSLKE ISD

GDVIISGNKNLCYANTINWKKLFGTSGQKTKIISNRGENSCKATGQVCHALCSPEGC

WGPEPRDCVSCRNVSRGRECVDKCNLLEGEPREFVENSECIQCHPECLPQAMNITCT

GRGPDNCIQCAHYIDGPHCVKTCPAGVMGENNTLVWKYADAGHVCHLCHPNCTYG

CTGPGLEGCPTNGPKIPSIATGMVGALLLLLVVALGIGLFMRRRHIVRKRTLRRLLQ E

RELVEPLTPSGEAPNQALLRILKETEFKKIKVLGSGAFGTVYKGLWIPEGEKVKIPV AI

KELREATSPKANKEILDEAYVMASVDNPHVCRLLGICLTSTVQLITQLMPFGCLLDY

VREHKDNIGSQYLLNWCVQIAKGMNYLEDRRLVHRDLAARNVLVKTPQHVKITDF

GLAKLLGAEEKEYHAEGGKVPIKWMALESILHRIYTHQSDVWSYGVTVWELMTFGS

KPYDGIPASEISSILEKGERLPQPPICTIDVYMIMVKCWMIDADSRPKFRELIIEFS KMA

RDPQRYLVIQGDERMHLPSPTDSNFYRALMDEEDMDDVVDADEYLIPQQGFFSSPST

SRTPLLSSLSATSNNSTVACIDRNGLQSCPIKEDSFLQRYSSDPTGALTEDSIDDTF LPV

PEYIN Q S VPKRP AGS V QNP V YHN QPLNP AP S RDPHY QDPHST AV GNPEYLNTV QPTC

VNSTFDSPAHWAQKGSHQISLDNPDYQQDFFPKEAKPNGIFKGSTAENAEYLRVAPQ

SSEFIGA

SEQUENCE ID NO: 2 (SEQ ID NO: 2)

NF 1/Homo sapiens/Human

MAAHRPVEWVQAVVSRFDEQLPIKTGQQNTHTKVSTEHNKECLINISKYKFSLVISG

LTT

ILKNVNNMRIFGEAAEKNLYLSQLIILDTLEKCLAGQPKDTMRLDETMLVKQLLPEI C

HF

LHTCREGNQHAAELRNSASGVLFSLSCNNFNAVFSRISTRLQELTVCSEDNVDVHDI E

LL

QYINVDCAKLKRLLKETAFKFKALKKVAQLAVINSLEKAFWNWVENYPDEFTKLYQ

IPQTDMAECAEKLFDLVDGFAESTKRKAAVWPLQIILLILCPEIIQDISKDVVDENN M

NKKLFLDSLRKALAGHGGSRQLTESAAIACVKLCKASTYINWEDNSVIFLLVQSMVV

DLKNLLFNPSKPFSRGSQPADVDLMIDCLVSCFRISPHNNQHFKICLAQNSPSTFHY V

L VNSLHRIITNS ALD WWPKID AV Y CHS VELRNMF GETLHKAV Q GC GAHP AIRM AP S

LTFKEKVTSLKFKEKPTDLETRSYKYLLLSMVKLIHADPKLLLCNPRKQGPETQGST

AELITGL V QL VP Q SHMPEI AQE AME ALL VLHQLD SIDL WNPD AP VETF WEI S S QMLF

YICKKLTSHQMLS STEILKWLREILICRNKFLLKNKQ ADRS S CHFLLF Y GV GCDIPS SG

NTSQMSMDHEELLRTPGASLRKGKGNSSMDSAAGCSGTPPICRQAQTKLEVALYMF

LWNPDTEAVLVAMSCFRHLCEEADIRCGVDEV SVHNLLPNYNTFMEFASV SNMMST

GRAALQKRVMALLRRIEHPTAGNTEAWEDTHAKWEQATKLILNYPKAKMEDGQAA ESLHKTIVKRRMSHVSGGGSIDLSDTDSLQEWINMTGFLCALGGVCLQQRSNSGLAT

YSPPMGPV SERKGSMISVMSSEGNADTPV SKFMDRLLSLMVCNHEKV GLQIRTNVK

DLVGLELSPALYPMLFNKLKNTISKFFDSQGQVLLTDTNTQFVEQTIAIMKNLLDNH T

EGSSEHLGQASIETMMLNLVRYVRVLGNMVHAIQIKTKLCQLVEVMMARRDDLSFC

QEMKFRNKMVEYLTDWVMGTSNQAADDDVKCLTRDLDQASMEAVVSLLAGLPLQ

PEEGDGVELMEAKSQLFLKYFTLFMNLLNDCSEVEDESAQTGGRKRGMSRRLASLR

HCTVLAMSNLLNANVDSGLMHSIGLGYHKDLQTRATFMEVLTKILQQGTEFDTLAE

TVLADRFERLVELVTMMGDQGELPIAMALANVVPCSQWDELARVLVTLFDSRHLL

Y QLL WNMF S KEVEL AD SMQTLFRGN S L ASKIMTFCFKV Y GATYLQKLLDPLLRIVIT

SSDWQHVSFEVDPTRLEPSESLEENQRNLLQMTEKFFHAIISSSSEFPPQLRSVCHC LY

QATCHSLLNKATVKEKKENKKSVVSQRFPQNSIGAVGSAMFLRFINPAIVSPYEAGI L

DKKPPPRIERGLKLMSKILQSIANHVLFTKEEHMRPFNDFVKSNFDAARRFFLDIAS D

CPTSDAVNHSLSFISDGNVLALHRLLWNNQEKIGQYLSSNRDHKAVGRRPFDKMAT

LLAYLGPPEHKPVADTHWSSLNLTSSKFEEFMTRHQVHEKEEFKALKTLSIFYQAGT

SKAGNPIF YYV ARRFKT GQIN GDLLI YHVLLTLKP YY AKP YEI V VDLTHT GP SNRFKT

DFLSKWFVVFPGFAYDNVSAVYIYNCNSWVREYTKYHERLLTGLKGSKRLVFIDCP

GKLAEHIEHEQQKLPAATLALEEDLKVFHNALKLAHKDTKVSIKVGSTAVQVTSAER

TKVLGQSVFLNDIYYASEIEEICLVDENQFTLTIANQGTPLTFMHQECEAIVQSIIH IRT

RWELSQPDSIPQHTKIRPKDVPGTLLNIALLNLGSSDPSLRSAAYNLLCALTCTFNL KI

EGQLLETSGLCIPANNTLFIVSISKTLAANEPHLTLEFLEECISGFSKSSIELKHLC LEYM

TPWLSNLVRFCKHNDDAKRQRVTAILDKLITMTINEKQMYPSIQAKIWGSLGQITDL

LDVVLDSFIKTSATGGLGSIKAEVMADTAVALASGNVKLVSSKVIGRMCKIIDKTCL S

PTPTLEQHLMWDDIAILARYMLMLSFNNSLDVAAHLPYLFHVVTFLVATGPLSLRAS

THGLVINIIHSLCTCSQLHFSEETKQVLRLSLTEFSLPKFYLLFGISKVKSAAVIAF RSS

YRDRSFSPGSYERETFALTSLETVTEALLEIMEACMRDIPTCKWLDQWTELAQRFAF

QYNPSLQPRALVVFGCISKRVSHGQIKQIIRILSKALESCLKGPDTYNSQVLIEATV IAL

TKLQPLLNKDSPLHKALFWVAVAVLQLDEVNLYSAGTALLEQNLHTLDSLRIFNDK

SPEEVFM AIRNPLEWHCKQMDHFV GLNFN SNFNF ALV GHLLKGYRHP SP AIV ARTV

RILHTLLTLVNKHRNCDKFEVNTQSVAYLAALLTVSEEVRSRCSLKHRKSLLLTDIS

MENVPMDTYPIHHGDPSYRTLKETQPWSSPKGSEGYLAATYPTVGQTSPRARKSMS

LDMGQPSQANTKKLLGTRKSFDHLISDTKAPKRQEMESGITTPPKMRRVAETDYEM

ETQRISSSQQHPHLRKV SV SESNVLLDEEVLTDPKIQALLLTVLATLVKYTTDEFDQRI

LYEYLAEASVVFPKVFPVVHNLLDSKINTLLSLCQDPNLLNPIHGIVQSVVYHEESP P

QYQTSYLQSFGFNGLWRFAGPFSKQTQIPDYAELIVKFLDALIDTYLPGIDEETSEE SL

LTPTSPYPPALQSQLSITANLNLSNSMTSLATSQHSPGIDKENVELSPTTGHCNSGR TR

HGSASQVQKQRSAGSFKRNSIKKIV

SEQUENCE ID NO: 3 (SEQ ID NO: 3)

IDHl/Homo sapiens/Human

MSKKISGGSVVEMQGDEMTRIIWELIKEKLIFPYVELDLHSYDLGIENRDATNDQVT KDAAEAIKKHNVGVKCATITPDEKRVEEFKLKQMWKSPNGTIRNILGGTVFREAIIC KNIPRLV SGWVKPIIIGRHAY GDQYRATDFVVPGPGKVEITYTPSDGTQKVTYLVHN FEEGGGVAMGMYNQDKSIEDFAHSSFQMALSKGWPLYLSTKNTILKKYDGRFKDIF QEIYDKQYKSQFEAQKIWYEHRLIDDMVAQAMKSEGGFIWACKNYDGDVQSDSVA QGY GSLGMMTSVLV CPDGKTVEAEAAHGTVTRHYRMY QKGQETSTNPIASIFAWT RGLAHRAKLDNNKELAFFAN ALEEV SIETIEAGFMTKDLAACIKGLPNV QRSDYLNT FEFMDKLGENLKIKLAQAKL

SEQUENCE ID NO: 4 (SEQ ID NO: 4)

LAMBl/Homo sapiens/Human MGLLQLLAFSFLALCRARVRAQEPEFSYGCAEGSCYPATGDLLIGRAQKLSVTSTCG LHKPEP Y CIV SHLQEDKKCFICN S QDP YHETLNPD SHLIENV VTTF APNRLKI WW Q SE NGVENVTIQLDLEAEFHFTHLIMTFKTFRPAAMLIERSSDFGKTWGVYRYFAYDCEA SFPGISTGPMKKVDDIICDSRYSDIEPSTEGEVIFRALDPAFKIEDPYSPRIQNLLKITN L RIKFVKL

HTLGDNLLDSRMEIREKYYYAVYDMVVRGNCFCYGHASECAPVDGFNEEVEGMVH

GHCMCRHNTKGLNCELCMDFYHDLPWRPAEGRNSNACKKCNCNEHSISCHFDMAV

YLATGNVSGGVCDDCQHNTMGRNCEQCKPFYYQHPERDIRDPNFCERCTCDPAGSQ

NEGICDSYTDFSTGLIAGQCRCKLNVEGEHCDVCKEGFYDLSSEDPFGCKSCACNPL

GTIPGGNPCDSETGHCYCKRLVTGQHCDQCLPEHWGLSNDLDGCRPCDCDLGGALN

NSCFAESGQCSCRPHMIGRQCNEVEPGYYFATLDHYLYEAEEANLGPGVSIVERQYI

QDRIPSWTGAGFVRVPEGAYLEFFIDNIPYSMEYDILIRYEPQLPDHWEKAVITVQR P

GRIPTSSRCGNTIPDDDNQVVSLSPGSRYVVLPRPVCFEKGTNYTVRLELPQYTSSD S

DVESPYTLIDSLVLMPYCKSLDIFTVGGSGDGVVTNSAWETFQRYRCLENSRSVVKT

PMTDVCRNIIFSISALLHQTGLACECDPQGSLSSVCDPNGGQCQCRPNVVGRTCNRC

APGTF GF GP SGCKPCECHLQGS VN AFCNPVTGQCHCF QGVY ARQCDRCLPGHW GFP

SCQPCQCNGHADDCDPVTGECLNCQDYTMGHNCERCLAGYYGDPIIGSGDHCRPCP

CPDGPDSGRQFARSCYQDPVTLQLACVCDPGYIGSRCDDCASGYFGNPSEVGGSCQP

CQCHNNIDTTDPEACDKETGRCLKCLYHTEGEHCQFCRFGYYGDALQQDCRKCVC

NYLGTVQEHCNGSDCQCDKATGQCLCLPNVIGQNCDRCAPNTWQLASGTGCDPCN

CNAAHSFGPSCNEFTGQCQCMPGFGGRTCSECQELFWGDPDVECRACDCDPRGIETP

QCDQSTGQCVCVEGVEGPRCDKCTRGYSGVFPDCTPCHQCFALWDVIIAELTNRTH

RFLEKAKALKISGVIGPYRETVDSVERKVSEIKDILAQSPAAEPLKNIGNLFEEAEK LI

KDVTEMMAQVEVKLSDTTSQSNSTAKELDSLQTEAESLDNTVKELAEQLEFIKNSDI

RGALDSITKYFQMSLEAEERVNASTTEPNSTVEQSALMRDRVEDVMMERESQFKEK

QEEQARLLDELAGKLQSLDLSAAAEMTCGTPPGASCSETECGGPNCRTDEGERKCG

GPGCGGLVTVAHNAWQKAMDLDQDVLSALAEVEQLSKMVSEAKLRADEAKQSAE

DILLKTNATKEKMDKSNEELRNLIKQIRNFLTQDSADLDSIEAVANEVLKMEMPSTP

QQLQNLTEDIRERVESLSQVEVILQHSAADIARAEMLLEEAKRASKSATDVKVTADM

VKEALEEAEKAQVAAEKAIKQADEDIQGTQNLLTSIESETAASEETLFNASQRISEL ER

NVEELKRKAAQN S GE AEYIEKV V YTVKQ S AED VKKTLDGELDEKYKKVENLI AKKT

EESADARRKAEMLQNEAKTLLAQANSKLQLLKDLERKYEDNQRYLEDKAQELARL

EGEVRSLLKDISQKVAVYSTCL

SEQUENCE ID NO: 5 (SEQ ID NO: 5)

UGDH/Homo sapiens/Human

MFEIKKICCIGAGYVGGPTCSVIAHMCPEIRVTVVDVNESRINAWNSPTLPIYEPGL KE

V

VESCRGKNLFFSTNIDDAIKEADLVFIS VNTPTKTY GMGKGRAADLKYIEAC ARRIV Q NS

NGYKIVTEKSTVPVRAAESIRRIFDANTKPNLNLQVLSNPEFLAEGTAIKDLKNPDR V

LI

GGDETPEGQRAVQALCAVYEHWVPREKILTTNTWSSELSKLAANAFLAQRISSINSI S

AL

CEATGADVEEVATAIGMDQRIGNKFLKASVGFGGSCFQKDVLNLVYLCEALNLPEV ARYWQQVIDMNDYQRRRFASRIIDSLFNTVTDKKIAILGFAFKKDTGDTRESSSIYISK YLMDEGAHLHIYDPKVPREQIVVDLSHPGV SEDDQV SRLVTISKDPYEACDGAHAV VICTEWDMFKELDYERIHKKMLKPAFIFDGRRVLDGLHNELQTIGFQIETIGKKVSSK RIP Y AP S GEIPKF S LQDPPNKKPKV

SEQUENCE ID NO: 6 (SEQ ID NO: 6) ANXA2/Homo sapiens/Human

MSTVHEILCKLSLEGDHSTPPSAYGSVKAYTNFDAERDALNIETAIKTKGVDEVTIV N

IL

TNRSNAQRQDIAFAYQRRTKKELASALKSALSGHLETVILGLLKTPAQYDASELKAS MKGLGTDEDSLIEIICSRTNQELQEINRVYKEMYKTDLEKDIISDTSGDFRKLMVALA KGRRAEDGSVIDYELIDQDARDLYDAGVKRKGTDVPKWISIMTERSVPHLQKVFDR YKSYSPYDMLESIRKEVKGDLENAFLNLV QCIQNKPLYFADRLYDSMKGKGTRDKV LIRIMV SRSEVDMLKIRSEFKRKY GKSLYYYIQQDTKGDY QKALLYLCGGDD

SEQUENCE ID NO: 7 (SEQ ID NO: 7)

S100A11/Homo sapiens/Human

MAKISSPTETERCIESLIAVFQKYAGKDGYNYTLSKTEFLSFMNTELAAFTKNQKDP G

VL

DRMMKKLDTNSDGQLDFSEFLNLIGGLAMACHDSFLKAVPSQKRT

SEQUENCE ID NO: 8 (SEQ ID NO: 8)

CTSB/Homo sapiens/Human

MWQLWASLCCLLVLANARSRPSFHPLSDELVNYVNKRNTTWQAGHNFYNVDMSY

LKRLCGTFLGGPKPPQRVMFTEDLKLPASFDAREQWPQCPTIKEIRDQGSCGSCWAF

GAVEAISDRICIHTNAHVSVEVSAEDLLTCCGSMCGDGCNGGYPAEAWNFWTRKGL

VSGGLYESHVGCRPYSIPPCEHHVNGSRPPCTGEGDTPKCSKICEPGYSPTYKQDKH Y

GYN S YS V SNSEKDIM AEIYKN GP VEGAF S V Y SDFLL YKS GV Y QHVT GEMMGGH AIR

ILGWGVENGTPYWLVANSWNTDWGDNGFFKILRGQDHCGIESEVVAGIPRTDQYW

EKI

SEQUENCE ID NO: 9 (SEQ ID NO: 9)

TP53/Homo sapiens/Human

MEEPQSDPSVEPPLSQETFSDLWKLLPENNVLSPLPSQAMDDLMLSPDDIEQWFTED P

GP

DEAPRMPEAAPPVAPAPAAPTPAAPAPAPSWPLSSSVPSQKTYQGSYGFRLGFLHSG

TAKS VT CTY SP ALNKMFCQL AKT CP V QL WVD STPPP GTRVRAM AI YKQ S QHMTEV

VRRCPHHERCSDSDGLAPPQHLIRVEGNLRVEYLDDRNTFRHSVVVPYEPPEVGSDC

TTIHYNYMCNSSCMGGMNRRPILTIITLEDSSGNLLGRNSFEVRVCACPGRDRRTEE E

NLRKKGEPHHELPPGSTKRALPNNTSSSPQPKKKPLDGEYFTLQIRGRERFEMFREL N

EALELKDAQAGKEPGGSRAHSSHLKSKKGQSTSRHKKLMFKTEGPDSD

SEQUENCE ID NO: 10 (SEQ ID NO: 10)

MGMT/Homo sapiens/Human

MDKDCEMKRTTLDSPLGKLELSGCEQGLHEIKLLGKGTSAADAVEVPAPAAVLGGP EPLMQCTAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLKVVKFGEVIS Y QQL AAL AGNPKAARAV GGAMRGNP VPILIPCHRV V CSS GAV GNY S GGL AVKEWL LAHEGHRLGKPGLGGSSGLAGAWLKGAGATSGSPPAGRN