If no, this is a negative dechallenge. 81 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 (Note: This criterion is not applicable if: (1) the AE resulted in death or permanent disability; (2) the AE resolved/improved despite continuation of the Sponsor’s product; or (3) the trial is a single-dose drug trial); or (4) Sponsor’s product(s) is/are only used one time.) 8.4.2.9 Rechallenge Was the subject re-exposed to Merck product in this study? If yes, did the AE recur or worsen? If yes, this is a positive rechallenge. If no, this is a negative rechallenge. (Note: This criterion is not applicable if: (1) the initial AE resulted in death or permanent disability, or (2) the trial is a single-dose drug trial); or (3) Sponsor’s product(s) is/are used only one time). NOTE: IF A RECHALLENGE IS PLANNED FOR AN ADVERSE EVENT WHICH WAS SERIOUS AND WHICH MAY HAVE BEEN CAUSED BY MERCK PRODUCT, OR IF REEXPOSURE TO MERCK PRODUCT POSES ADDITIONAL POTENTIAL SIGNIFICANT RISK TO THE SUBJECT, THEN THE RECHALLENGE MUST BE APPROVED IN ADVANCE BY THE SPONSOR AS PER DOSE MODIFICATION GUIDELINES IN THE PROTOCOL. 8.4.2.10 Consistency with Trial Treatment Profile Is the clinical/pathological presentation of the AE consistent with previous knowledge regarding Merck product or drug class pharmacology or toxicology? The assessment of relationship will be reported on the case report forms /worksheets by an investigator who is a qualified physician according to his/her best clinical judgment, including consideration of the above elements. Record one of the following: Use the following scale of criteria as guidance (not all criteria must be present to be indicative of Merck product relationship). 1.Yes, there is a reasonable possibility of Merck product relationship. There is evidence of exposure to Merck product. The temporal sequence of the AE onset relative to the administration of Merck product is reasonable. The AE is more likely explained by Merck product than by another cause. 2.No, there is not a reasonable possibility of Merck product relationship. Subject did not receive the Merck product OR temporal sequence of the AE onset relative to administration of Merck product is not reasonable OR the AE is more likely explained by another cause than the Merck product. (Also entered for a subject with overdose without an associated AE.) 8.4.3 Events of Clinical Interest These selected non-serious and serious adverse events are also known as Events of Clinical Interest (ECI) and must be reported within 24 hours to Merck. For reporting of ECI refer to section 8.3. For the time period beginning when the consent form is signed until treatment initiation, any ECI, or follow up to an ECI, that occurs to any subject must be reported within 24 hours to the 82 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 Sponsor if it causes the subject to be excluded from the trial, or is the result of a protocol- specified intervention, including but not limited to washout or discontinuation of usual therapy, diet, placebo treatment or a procedure. For the time period beginning at treatment initiation through 90 days following cessation of treatment, or 30 days following cessation of treatment if the subject initiates new anticancer therapy, whichever is earlier, any ECI, or follow up to an ECI, whether or not related to study treatment, must be reported within 24 hours to the Sponsor (NCI). Merck Global Safety facsimile number: +1-215-993-1220 (Sponsor [NCI] will contact Merck) 8.4.4 Reporting Serious Adverse Events to Merck and Agenus Upon receipt, Sponsor (NCI) will forward SAE reports and any other relevant safety information to the Merck Global Safety facsimile number: +1-215-993-1220 and to Agenus Pharmacovigilance via email: adverse.events@agenusbio.com or fax: 1-781-674-4261. A copy of all 15 Day Reports and Annual Progress Reports will be submitted to Merck & Co., Inc. (Attn: Worldwide Product Safety; FAX 215 993-1220) and to Agenus Pharmacovigilance (email: adverse.events@agenusbio.com or fax: 1-781-674-4261) at the time of submission to FDA. For the time period beginning at treatment initiation through 90 days following cessation of treatment, or 30 days following cessation of treatment if the subject initiates new anticancer therapy, whichever is earlier, any ECI, or follow up to an ECI, whether or not related to study treatment, must be reported within 24 hours to the Sponsor (NCI). Sites will submit all reportable events to the Sponsor (NCI), and NCI will submit to Agenus and Merck within 2 working days. The DSMB Sponsor will monitor unblinded aggregated efficacy endpoint events and safety data to ensure the safety of the subjects in the trial. Any suspected endpoint which upon review is not progression of the cancer under study will be forwarded to Merck Global Safety and Agenus Pharmacovigilance as a SAE within 2 working days of determination that the event is not progression of the cancer under study. For reporting pregnancy: The designated Sponsor representative will work with the investigator to ensure that all relevant information is provided to Merck and Agenus within 1 to 5 calendar days for SAEs and within 30 days for all other pregnancies. The same timelines apply when outcome information is available. 8.5 REPORTING PREGNANCY All required pregnancy reports/follow-up to OSRO will be submitted to: OSROSafety@mail.nih.gov and to the CCR PI and study coordinator. Forms and instructions can be found here: https://ccrod.cancer.gov/confluence/display/CCRCRO/Forms+and+Instructions. 8.5.1 Maternal exposure If a participant becomes pregnant during the course of the study, the study treatment should be discontinued immediately, and the pregnancy reported to the Sponsor no later than 24 hours of 83 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 when the Investigator becomes aware of it. The Investigator should notify the Sponsor no later than 24 hours of when the outcome of the Pregnancy becomes known, Pregnancy itself is not regarded as an SAE. However, congenital abnormalities or birth defects and spontaneous miscarriages that meet serious criteria (section 8.3) should be reported as SAEs. The outcome of all pregnancies should be followed up and documented. 8.5.2 Paternal exposure Male participants should refrain from fathering a child or donating sperm during the study and for 120 days after the last dose of pembrolizumab. Pregnancy of the participant’s partner is not considered to be an AE. The outcome of all pregnancies occurring from the date of the first dose until 120 days after the last dose should, if possible, be followed up and documented. Pregnant partners may be offered the opportunity to participate in an institutional pregnancy registry protocol (e.g., the NIH IRP pregnancy registry study) to provide data about the outcome of the pregnancy for safety reporting purposes. 8.6 REGULATORY R EPORTING FOR STUDIES C ONDUCTED UNDER CCR SPONSORED IND Following notification from the investigator, CCR, the IND sponsor, will report any suspected adverse reaction that is both serious and unexpected in expediated manner to the FDA in accordance to 21 CFR 312. 32. CCR will report an AE as a suspected adverse reaction only if there is evidence to suggest a causal relationship between the study product and the adverse event. CCR will notify FDA and all participating investigators (i.e., all investigators to whom the sponsor is providing drug under its INDs or under any investigator’s IND) in an IND safety report of potential serious risks from clinical trials or any other source, as soon as possible, in accordance to 21 CFR Part 312.32. All serious events will be reported to the FDA at least annually in a summary format. 9 CLINICAL MONITORING Clinical site monitoring is conducted to ensure: that the rights of the participants are protected; that the study is implemented per the approved protocol, Good Clinical Practice and standard operating procedures; and, the quality and integrity of study data and data collection methods are maintained. Monitoring for this study will be performed by NCI CCR Office of Sponsor and Regulatory Oversight (OSRO) Sponsor and Regulatory Oversight Support (SROS) Services contractor. Clinical site monitoring activities will be based on OSRO standards, FDA Guidance E6(R2) Good Clinical Practice: Integrated Addendum to ICH E6(R1) March 2018, and applicable regulatory requirements. Details of clinical site monitoring will be documented in a Clinical Monitoring Plan (CMP) developed by OSRO. CMPs will be protocol-specific, risk-based and tailored to address human subject protections and integrity of the study data. OSRO will determine the intensity and frequency of monitoring based on several factors, including study type, phase, risk, complexity, 84 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 expected enrollment rate, and any unique attributes of the study and the site. The Sponsor will conduct a periodic review of the CMP to confirm the plan's continued appropriateness. A change to the protocol, significant or pervasive non-compliance with GCP, or the protocol may trigger CMP updates. OSRO SROS Monitoring visits and related activities will be conducted throughout the life cycle of each protocol. The first activity is before the study starts to conduct a Site Assessment Visit (SAV) (as warranted), followed by a Site Initiation Visit (SIV), Interim Monitoring Visit(s) (IMVs), and a study Close-Out Visit (COV). Some monitoring activities may be performed remotely, while others will occur at the study site(s). Monitoring visit reports will describe visit activities, observations, and associated action items or follow-up required for resolution of any issues, discrepancies, or deviations. Monitoring reports will be distributed to the study PI, NCI CCR QA, CCR Protocol Support Office, coordinating center (if applicable), and the Sponsor regulatory file. The site Monitor will inform the study team of any deviations observed during monitoring visits. If unresolved, the Monitor will request that the site Staff enter the deviations in the CCR Protocol Deviation Tracking System (PDTS) for deviation reporting to the Sponsor and as applicable per institutional and IRB guidance. 10 STATISTICAL CONSIDERATIONS This is a phase II blinded, randomized trial in newly diagnosed GBM. Evaluable patients must complete 1 cycle of treatment post RT. A Bayesian two-stage design will be used to monitor the efficacy of two experimental arms independently based on the OS at 8 months (OS8) using the following Bayesian rule (Thall and Simon, 1994) [46]: Stop the experimental arm for futility if Pr (OS8 < 0.75 | data ) > 0.6. That is, if the observed data indicate that there is greater than 60% chance that the OS8 is smaller than 0.75, we stop the experimental arm. We choose 0.75 as the threshold because that is the estimate of OS8 for the standard of care (RT+TMZ). In order to speed up the trial, we use the OS8, rather than the OS at 12 months (i.e., the primary endpoint), for the purpose of interim monitoring. Assuming vague Beta prior Beta (0.25, 0.25) for the OS8, the two-stage Bayesian design can be described as follows: Enroll 22 patients for each of the experimental arms, if 16 or more patients survive at 8 months, we continue to enroll the remaining 23 patients; otherwise stop that experimental arm. At the interim analysis, if some patients are followed less than 8 months, we will calculate Pr (OS8 < 0.75 | data) based on the methodology of Cai, Liu and Yuan (2014) by imputing the OS8 for these patients. The following table shows the operating characteristics of the Bayesian phase II design: True OS8 0.6 0.7 0.8 0.9 85 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 Stopping probability 0.845 0.507 0.135 0.004 Average sample size 25.6 33.3 41.9 44.9 Similar Bayesian method will also be used to monitor the rate of DLT. We monitor two experimental arms independently-patients that are randomized. Specifically, for each of the arms, we will stop enrolling patients and inspect the safety data if the Pr(DLT > 25% | data) > 0.9. That is, we will stop enrolling patients if the data indicate that there is more than 90% chance that the true DLT rate is higher than 25%. This decision rule gives the following stopping rule, assuming a Beta(0.1, 0.4) prior distribution for DLT rate, Stop enrolling pts if [# of pts with DLT] / [# of pts evaluated] ≥ 2/3, 4/6, 5/10 or 8/20. Below are the operating characteristics of the toxicity monitoring rule. True toxicity rate 0.1 0.2 0.3 0.4 0.5 Early stop probability0.028 0.136 0.372 0.692 0.906 Average sample size27.3 25.0 20.6 14.9 9.8 In the case that one arm is discontinued due to futility or safety, enrollment would shift to the remaining arm as the primary objective of this trial is not to compare two arms. 10.1 SAMPLE SIZE AND ACCRUAL Based on previous experience of newly diagnosed (ND) unmethylated GBM patients treated with Pembrolizumab, the estimated one-year OS for the RT+TMZ+Pembrolizumab arm is 70%, and the estimated one-year OS for the RT+TMZ+Pembrolizumab +HSPPC-96 arm is 90%. Given one-sided significance level of 0.10, the sample size of 45 patients per arm provides 80% power to detect the difference between two experimental arms. Treatment accrual for the randomization arms will be 90 patients (45 patients per arm). Considering vaccine production failures for patients when HSPPC-96 is not produced, accrual in the ancillary cohort is up to 18 patients. Therefore, the randomized treatment accrual ceiling will be 108 patients. Assuming a screen failure rate of 65%, the total accrual ceiling will be 310 patients. Approximately 8 potentially eligible patients are seen per month, and it is anticipated that at least 1-2 per month will be accrued per site. NOTE: DSMB recommended in May 2021 to terminate study early due to slow accrual. Study is closed to enrolment. Patients will be unblinded but will be kept on treatment until completion of TMZ, Pembro, and/or HSPPC-96 and followed for overall survival. Placebo will be discontinued. 86 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 10.2 DATA ANALYSES PLANS Progression-free and overall survival will be determined using a Kaplan-Meier survival curve. Median PFS and OS will be determined from this curve. A one-sided lower 90% confidence limit for median PFS and OS will be determined by the method of Brookmeyer and Crowley [47]. The Chi-square test will be used to compare the one-year OS between two experimental arms. In addition, the weighted log rank test will also be used to compare the OS and PFS between two experimental arms. 10.3 SECONDARY OBJECTIVES & ENDPOINTS PFS-6 (percentage of subjects alive and progression-free at 6 months post-registration): determined from the value of the Kaplan-Meier curve at 6 months. Response rate: determined as described by RANO Table 2. Rates and 95% confidence intervals will be determined using exact binomial probability distributions. Overall survival: Kaplan-Meier curves will be used to analyze overall survival. OS-6, 12 and 24 (overall survival at 6, 12, and 24 months post-registration, respectively): determined from the level of the Kaplan-Meier curves at these time points. Safety of Pembrolizumab in GBM: Adverse events will be described descriptively by giving frequencies of each event by type, severity, frequency, timing, and attribution. To evaluate the occurrence of symptoms and correlate to disease progression and tolerance to treatment using the MD Anderson Symptom Inventory-Brain Tumor Module (MDASI-BT). 11 COLLABORATIVE AGREEMENTS 11.1 AGREEMENT TYPE There is a BTTC consortia agreement in place between all of the participating institutions listed in the NIH IRB system. In addition, there are CRADA agreements in place with Merck (03128), and Agenus (03127) who are supporting this study. Transfers that are associated with correlative studies conducted under an approved protocol: Investigators in the NIH intramural program may participate in multi-site clinical trials (either as a site or as the coordinating center) under which human materials are transferred from the intramural program to another site for correlative studies that are part of the approved protocol. In such a situation, the protocol clearly documents the tests conducted under the correlative studies, and each institution participating in the clinical study is bound by the terms of their Protocol and their obligations under the statutes and regulations. In addition, intramural protocols are cleared by the IC Clinical Director. In such a situation, use of an HM-MTA is not necessary for these transfers. 11.2 MULTI-INSTITUTIONAL GUIDELINES 11.2.1 Participating Institution Site Initiation Before an institution may begin participating in a BTTC protocol, they must complete the following steps: 87 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 Submit all required regulatory documents to the BTTC Coordinating Center Participate in a site initiation visit, webcast, or conference call Receive training regarding study specific CRF’s and/or databases After these requirements have been fulfilled, the participating institution will receive by fax, e- mail, or hard copy, a Site Activation Memo. Once the Site Activation Memo has been received, the participating institution may begin to register patients to the protocol. 11.2.2 IRB Approvals The lead PI will provide the NIH Intramural IRB with a copy of the participating institution’s initial approval and approved yearly continuing review. Registration will be halted at any participating institution in which a current continuing approval is not on file at the NIH Intramural IRB. 12 HUMAN SUBJECTS PROTECTIONS 12.1 RATIONALE F OR S UBJECT SELECTION This study was designed to include women and minorities, but was not designed to measure differences of intervention effects. Males and females will be recruited with no preference to gender. No exclusion to this study will be based on race. Minorities will actively be recruited to participate. High grade gliomas occur in patients of all races and although there is a slight predominance in men, this is a disease that is also common in women. The molecular targets of pembrolizumab and HSPPC-96 within the tumor are not known to be different among patients based on gender or race; hence this study will be open to all adults. 12.2 PARTICIPATION OF C HILDREN Individuals under the age of 18 will not be eligible to participate in this study because they are unlikely to have glioblastoma, and because of unknown toxicities of the study agents in the pediatric patient. Furthermore, the targets of pembrolizumab and HSPPC-96 are not as prevalent in pediatric malignant gliomas and therefore, the efficacy of this regimen will be initially determined in the adult population before consideration of its use in pediatrics. 12.3 PARTICIPATION OF S UBJECTS UNABLE TO GIVE C ONSENT (NCI) Adults unable to give consent are excluded from enrolling in the protocol. However, re-consent may be necessary and there is a possibility, though unlikely, that subjects could become decisionally impaired. In circumstances where the subject becomes decisionally impaired due to disease progression, the subject will be taken off-treatment per protocol, but may continue in long term follow-up. In long term follow-up, the risks associated with study related interventions are no more than minimal. However, because in other circumstances, there is a prospect of direct benefit from research participation, all subjects ≥ age 18 will be offered the opportunity to fill in their wishes for research and care, and assign a substitute decision maker on the “NIH Advance Directive for Health Care and Medical Research Participation” form so that another person can make decisions about their medical care in the event that they become incapacitated or cognitively 88 Abbreviated Title: GBM Pembro HSPPC Version Date: 03/26/2024 impaired during the course of the study. Note: The PI or AI will contact the NIH Ability to Consent Assessment Team (ACAT) for evaluation to assess ongoing capacity of the subjects and to identify an LAR, as needed. Please see section 12.6.1 for consent procedure. 12.4 PARTICIPATION OF A DULTS UNABLE TO GIVE C ONSENT (BTTC) Each institution will follow their own procedure for re-consenting subjects unable to re-consent and will notify the Coordinating Center at time of continuing review of these occurrences. 12.5 EVALUATION OF BENEFITS AND RISKS/DISCOMFORTS 12.5.1 Risks 12.5.1.1 Study Agents The primary risk to patients participating in this research study is from the toxicity of pembrolizumab or HSPPC-96 or all drugs as described in section 14 and the consent document. Pembrolizumab and HSPPC-96 are investigational agents in the treatment of gliomas. The protocol provides for detailed and careful monitoring of all patients to assess for toxicity.