Refer to the commercially labeled bottles for expiration dating. Temozolomide is rapidly and completely absorbed after oral administration; peak plasma concentrations occur in 1 hour. Food reduces the rate and extent of temozolomide absorption. Mean peak plasma concentration and AUC decreased by 32% and 9%, respectively, and Tmax increased 2-fold (from 1.1 to 2.25 hours) when temozolomide was administered after a modified high-fat breakfast. Temozolomide is rapidly eliminated with a mean elimination half-life of 1.8 hours and exhibits linear kinetics over the therapeutic dosing range. Temozolomide has a mean apparent volume of distribution of 0.4 L/kg (%CV=13%). It is weakly bound to human plasma proteins; the mean percent bound of drug- related total radioactivity is 15%. 14.2.8 Metabolism and Elimination Temozolomide is spontaneously hydrolyzed at physiologic pH to the active species, 3-methyl- (triazen-1-yl)imidazole-4-carboxamide (MTIC) and to temozolomide acid metabolite. MTIC is further hydrolyzed to 5-amino-imidazole-4-carboxamide (AIC), which is known to be an intermediate in purine and nucleic acid biosynthesis and to methylhydrazine, which is believed to be the active alkylating species. Cytochrome P450 enzymes play only a minor role in the metabolism of temozolomide and MTIC. 14.2.9 Mechanism of Action An oral cytotoxic alkylating agent is the lead agent in a new class of compounds known as imidazotetrazines. Cytotoxic agents are designed to prevent the replication of cells that divide rapidly, including those in tumors. 14.2.10 Special Populations 14.2.10.1 Creatinine Clearance: Caution should be exercised when temozolomide is administered to patients with severe renal impairment. Temozolomide has not been studied in patients on dialysis. 14.2.10.2 Hepatically Impaired Patients: In a pharmacokinetic study, the pharmacokinetics of temozolomide in patients with mild to moderate hepatic impairment (Child’s-Pugh Class I-II) were similar to those observed in patients with normal hepatic function. Caution should be exercised when temozolomide is administered to patients with severe hepatic impairment. 14.2.10.3 Gender: Population pharmacokinetic analysis indicates that women have an approximately 5% lower clearance (adjusted for body surface area) for temozolomide than men. Women have higher incidences of Grade 4 neutropenia and thrombocytopenia in the first cycle of therapy than men. 99 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 14.2.10.4 Age: Population pharmacokinetic analysis indicates that age (range 19-78 years) has no influence on the pharmacokinetics of temozolomide. 14.2.11 Drug-Drug Interactions In a multiple dose study, administration of temozolomide with ranitidine did not change the Cmax or AUC values for temozolomide or MTIC. Population analysis indicates that administration of valproic acid decreases the clearance of temozolomide by about 5%. The clinical implication of this effect is not known. Population analysis failed to demonstrate any influence of co-administered dexamethasone, prochlorperazine, phenytoin, carbamazepine, ondansetron, H2-receptor antagonists, or phenobarbital on the clearance of orally administered temozolomide. 14.2.12 Adverse Events: Hematologic: Thrombocytopenia, leukopenia, myelodysplastic syndrome Gastrointestinal: Nausea, vomiting, anorexia Hepatic: Elevated liver enzymes (reversible) Skin: Rash Neurologic: Convulsions, weakness on one side of the body, abnormal coordination, paralysis Other: Constipation, diarrhea, stomatitis, fatigue, decreased performance status, headache 14.2.13 Special Handling and Precautions Capsules should not be opened or damaged. Rigorous precautions should be taken to avoid capsule contents having contact with skin or mucous membranes. Capsule contents may be irritating to skin and eyes. Mutagenic and prolonged exposure may cause serious health effects (outside of prescribed dosage in this trial). Temozolomide is potentially mutagenic and should be handled with appropriate precautions by both staff and patients. Capsules should not be opened. If capsules are accidentally opened or damaged, rigorous precautions should be taken with the capsule contents to avoid inhalation or contact with the skin or mucous membranes. Procedures for proper handling and disposal of anticancer drugs should be considered. 14.2.14 Contraindications Temozolomide is contraindicated in patients who have a history of a hypersensitivity reaction to any of its components or to DTIC. 14.2.15 Pregnancy Category D Temozolomide may cause fetal harm when administered to a pregnant woman. There are no adequate and well-controlled studies in pregnant women. If this drug is used during pregnancy, or if the patient becomes pregnant while taking this drug, the patient should be apprised of the potential hazard to the fetus. Women of childbearing potential should be advised to avoid becoming pregnant during therapy with temozolomide. 100 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 14.2.16 Fertility Treatment of a man with temozolomide may increase the risk of birth defects if he causes a woman to become pregnant while he is taking temozolomide. Men treated with temozolomide may have difficulty causing a woman to become pregnant after their treatment is completed. Men receiving temozolomide should be directed to use effective contraception while they are being treated. There is insufficient data to know what the risk of subsequent problems with fertility will be. Similarly, women who are treated with temozolomide may have difficulty becoming pregnant in the future and may at be at increased risk of having children with birth defects. There is insufficient evidence to determine what the risk of these complications will be. 14.3 HEAT SHOCK PROTEIN-PEPTIDE COMPLEX-96 VACCINE (HSPPC-96) HSPPC-96 is an autologous tumor-derived vaccine under clinical investigation for the treatment of a variety of cancer types. It is composed of the 96-kDa heat shock protein gp96 in complex with autologous tumor-derived peptides. The gp96 in HSPPC-96 is a highly conserved, abundant, nonpolymorphic stress protein found in every cell type of the body. The gp96 isolated from normal or tumor tissues is found in complex with a diverse repertoire of peptides, which are specific to the tissue of origin. Published studies in mouse tumor models have shown that HSPPC-96 confers protective immunity only to the tumor from which it is derived and not to antigenically distinct tumors. The specific immunogenicity of the HSP preparations can be attributed to the unique repertoire of antigenic peptides that exists in different cancers. When injected into the host, HSPPC-96 interacts with antigen presenting cells (APCs) such as macrophages, dendritic cells or Langerhans cells, which take up HSPPCs via HSP receptors, including CD91. Once internalized by APCs, the peptides chaperoned by the HSPs are transferred to major histocompatibility complex (MHC) class I and II molecules in intracellular compartments and subsequently expressed at the cell surface. The cells then migrate to lymph nodes, where the complexes are processed so that the peptides chaperoned by the HSPs are re- presented to naive T cells. The T cells recognize the peptides and, as a result, are stimulated by them. Vaccination with HSPPCs thus elicits both a CD8+ and CD4+ T-cell response targeting potentially all relevant tumor antigens. The interaction of HSPs with receptors on APCs also leads to activation of various components of innate immunity, including cytokine and chemokine release by macrophage and dendritic cells, maturation of dendritic cells and activation of natural killer cells, as shown in both murine and human systems. Tumor immunity is largely mediated by T cells and the ability of HSPPC-96 to stimulate both T-cell arms to recognize a large variety of tumor antigens, coupled with the ability to activate innate immune responses, uniquely positions the product among other cancer vaccine strategies. 14.3.1 Molecular Formula The molecular formula of HSPPC-96 is unknown since the antigenic peptide repertoire isolated from tumor cells is patient specific. 14.3.2 Peptide Component The peptide component of HSPPC-96 is not characterized on a patient specific basis due to the limited quantity of product available and the technical challenges in performing this type of peptide analysis. 101 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 14.3.3 Appearance HSPPC-96 is supplied in a single-use vial as a clear, colorless solution. Each vial contains 25 μg of HSPPC-96 in a solution of 9% sucrose-potassium phosphate for intradermal (ID) injection. The total volume of each vial of HSPPC-96 is 0.47 mL. The total volume that should be administered is approximately 0.4 mL. 14.3.4 Availability Blinded vaccine (HSPPC-96 or placebo) will be prepared by Agenus Inc. and shipped directly to sites within approximately 4-6 weeks of Agenus’ receipt of tumor tissue for randomized patients. The number of vaccine treatments will be based on the amount of vaccine generated. Although HSPPC-96 vaccine will be prepared for patients on the placebo arm, it will not be shipped to sites. Vaccine produced for patients randomized to placebo will be used for quality evaluation and will not be available for clinical use. 14.3.5 HSPPC-96 Vaccine Label Each vial of vaccine vial is labeled with the subject/patient study number, subject/patient initials, and subject/patient date of birth (DOB). 14.3.6 Storage and Stability Following successful production of HSPPC-96 vaccine at Agenus Inc. and randomization, vials of blinded vaccine are shipped overnight on dry ice and must be stored at -80 °C ± 20 °C until administration to the patient. NOTE: Once HSPPC-96 is drawn up into the syringe, the vaccine is stable for up to 2 hours at ambient temperature if immediate administration is not possible. Please refer to the Investigator’s Brochure for further details. Any unused or expired vaccine remaining after a patient has completed vaccine treatment should be returned to Agenus or destroyed at the site upon approval from Agenus. Destruction records are to be forwarded to Agenus. The shipping kit used to send vaccine from Agenus to the site may be stored until the patient has completed vaccine therapy and utilized for return of any unused or expired vaccine. See Appendix 16.6 for further detail. 14.3.7 Preparation Withdraw the vial from the freezer and roll gently between two fingers until it is completely thawed (5-10 minutes). Withdraw approximately 0.4 mL, using a low hold up TB or insulin syringe. 14.3.8 Administration The route of administration is intradermal injection. Inject into 1 site or into 2 adjacent sites (0.2 mL each) a few centimeters apart. Appropriate sites for vaccination include the anterior deltoid regions or subclavicular region bilaterally. Do not administer HSPPC-96/placebo to any side where lymph nodes have been removed or damaged. Rotate the injection sites so that injections are not repeated at the same site at 2 consecutive administrations, and utilize all potential sites for the patient before repeating 102 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 injections at a previously used injection site. Syringes with slip-tip detachable needles or luer hubs with greater than 0.1 mL dead space should not be utilized. 14.3.9 Toxicities Local injection-site reactions can occur and are usually of minimal extent and mild severity. If a local injection-site reaction requires medical attention, the investigator can provide local therapy as needed. Occasional systemic toxicities have been reported that have been mild in severity. The most frequently reported events include injection- site reactions of erythema and induration, fatigue, nausea, pyrexia, headache, back pain, arthralgia, constipation, and asthenia. 14.4 PLACEBO 14.4.1 Supply Placebo will be prepared by Agenus Inc. and shipped directly to sites within approximately 4-6 weeks of Agenus’ receipt of tumor tissue for randomized patients. The number of placebo treatments will be based on the amount of vaccine generated. 14.4.2 Product Description Each dose of placebo is supplied in a single-use vial as a clear, colorless solution. Each vial contains a solution of 9% sucrose-potassium phosphate for intradermal (ID) injection. The total volume in each vial of placebo is 0.47 mL. The total volume that should be administered is approximately 0.4 mL. 14.4.3 Placebo Label Each vial of vaccine vial is labeled with the subject/patient study number, subject/patient initials, and subject/patient date of birth (DOB). 14.4.4 Storage Vials of blinded vaccine are shipped overnight on dry ice and must be stored at–-80 °C ± 20 °C until administration to the patient. NOTE: Once the placebo is drawn up into the syringe, it is stable for up to 2 hours at ambient temperature if immediate administration is not possible. 14.4.5 Administration Withdraw the vial from the freezer and roll gently between two fingers until it is completely thawed (5-10 minutes). Withdraw approximately 0.4 mL, using a low hold up TB or insulin syringe. 14.4.6 Dose The route of administration is intradermal injection. Inject into 1 site or into 2 adjacent sites (0.2 mL each) a few centimeters apart. Appropriate sites for vaccination include the anterior deltoid regions or subclavicular region bilaterally. 103 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 Do not administer placebo to areas distal to lymph node basins that have been resected or in areas just distal to a surgical scar. Rotate the injection sites so that injections are not repeated at the same site at 2 consecutive administrations, and utilize all potential sites for the patient before repeating injections at a previously used injection site. Syringes with slip-tip detachable needles or luer hubs with greater than 0.1 mL dead space should not be utilized. 14.5 ORDERING S TUDY AGENT 14.5.1 Heat Shock Protein-Peptide Complex-96 Following vaccine manufacture and completion of quality testing, Agenus will send the site the Vaccine Production Notification form indicating whether manufacture of vaccine was successful. Upon randomization, Agenus will then send the appropriate blinded vaccine to the site. Agenus Contact Information for Tissue Procurement and Kit Ordering: supply&logistics@agenusbio.com or fax: 781-674-4220. See Appendix 16.3 and Appendix 16.4 for more details.