Peptide Targets for Glioblastoma Against Novel Cytomegalovirus Antigens (PERFORMANCE)

Annias Immunotherapeutics, Inc., National Institutes of Health (NIH), National Cancer Institute (NCI)

Newly diagnosed glioblastoma (GBM) patients with complete or partial surgical resection who were CMV seropositive patients were eligible to enroll on this trial. Patients were enrolled following standard of care chemoradiation and prior to initiation of post-radiation cycles of temozolomide (TMZ) provided they met all eligibility criteria. All eligible patients received a tetanus-diphtheria (Td) vaccination. Patients enrolled on study were randomized to receive either standard TMZ or dose-intensified TMZ (excluding the safety cohort who only received standard TMZ). All patients received a pre-conditioning injection of tetanus on day 22 of the first post-radiation cycle of TMZ. The following day, patients received the first of 3 intradermal (i.d.) injections of the study drug cytomegalovirus peptide (PEP-CMV), which contained either a combination of Component A and Component B or Component A only depending upon when they enrolled on study. Vaccines #2 and #3 will be given at 2 week intervals. Patients who were O[6]-methylguanine-DNA methyltransferase (MGMT) unmethylated received one adjuvant cycle of the TMZ regimen according to their assigned TMZ arm. Patients who were MGMT methylated or whose methylation status was inconclusive continue with up to 12 cycles of TMZ. After the completion of a patient's last TMZ cycle, vaccines continued every 4-6 weeks for a maximum number of 20 vaccines (unless tumor progression occurred). The study ended prematurely due to lack of funds. The preliminary results suggest that the vaccine may be capable of generating an immune response.

Patients were enrolled following radiation therapy and prior to initiation of post-radiation therapy cycles of adjuvant TMZ provided they met all eligibility criteria. After signing main consent, patients had immune monitoring blood work collected and received a Tetanus-diphtheria booster vaccination with 0.5 mL of Td (tetanus, diphtheria toxoid, adsorbed). After meeting all eligibility criteria, patients were randomized to receive either standard TMZ (150-200 mg/m^2/day on days 1-5 of each 28-day cycle) with vaccination on Day 23 (-1 day, + 2 days) of each TMZ cycle or to receive dose-intensified TMZ (75-100 mg/m^2/day on days 1-21 of each 28-day cycle) with vaccination on day 23 (-1 day, +2 days) of each TMZ cycle (excluding patients enrolled in the safety cohort who only received standard TMZ). Patients began their initial cycle of adjuvant TMZ as soon as possible following randomization, if applicable. For Arm 1, the adjuvant TMZ cycle(s) will be given as described above. If a patient had an MGMT unmethylated tumor, they discontinued TMZ after the 1st cycle. All patients received a tetanus pre-conditioning injection in the right groin on day 22 (+1 day) of cycle 1 of adjuvant TMZ. On the following day, patients receive their study vaccine (either a combination of Component A and Component B or Component A only depending upon when they enrolled on study). Vaccines #2 and #3 were given at 2 week intervals (+ 3 days), which will resulted in a ~35-day delay before starting TMZ cycle 2. MGMT unmethylated patients did not receive subsequent cycles of TMZ, but continued to receive vaccines approximately every 4 (+2) weeks. Originally, patients received the vaccine as follows: 500 µg of PEP-CMV Component A mixed with Montanide Incomplete Freund's Adjuvant (ISA)-51 intradermally administered in the right groin and 2 hours later, 500 µg of PEP-CMV Component B mixed in 150 µg of Granulocyte Macrophage Colony Stimulating Factor (GM-CSF) intradermally administered in the left groin. A safety cohort was added in 2017 following hypersensitivity reactions in which patient received different schedules of vaccine Components A and B to determine the source of the hypersensitivity reactions. Following this safety cohort, the randomized study was allowed to reinitiate with changes to vaccine administration procedure. Subsequent revisions were made to the study in the latter part of 2018, due to a continuation of hypersensitivity reactions, and Component B was removed from the study. Patients then received the vaccine as follows: 500 µg of PEP-CMV Component A mixed with Montanide ISA-51 administered intradermally with half in the right groin and half in the left groin. Patients were imaged with contrast-enhanced magnetic resonance imaging (MRI) within 2 weeks (+3 days) after vaccine 3 and then approximately every 8 weeks. RANO criteria was used for assessment of pseudo-progression, and patients demonstrating definitive progression were removed from study. Blood for immune monitoring was obtained at several time points. The study ended prematurely due to lack of funds.

All patients receive Tetanus-Diphtheria booster vaccine at time of enrollment. Cycles of standard TMZ (150-200 mg/m^2/day on days 1-5 of each 28 day cycle) with PEP-CMV vaccination on Day 23 (-1 day, + 2 days) of each TMZ cycle and tetanus pre-conditioning the day before the first vaccine.

All patients receive Tetanus-Diphtheria booster vaccine at time of enrollment. Cycles of dose-intensified TMZ (75-100 mg/m^2/day on days 1-21 of each 28 day cycle) with PEP-CMV vaccination on day 23 (-1 day, + 2 days) of each TMZ cycle and tetanus pre-conditioning the day before the first vaccine.

All patients receive Tetanus-Diphtheria booster vaccine at time of enrollment. Cycles of standard TMZ (150-200 mg/m^2/day on days 1-5 of each 28 day cycle) with PEP-CMV vaccination on Day 23 (-1 day, + 2 days) of each TMZ cycle and tetanus pre-conditioning the day before the first vaccine. Vaccine components A and B were administered separately, with a delay between them, to determine if it was an individual component or a combination of the components that resulted in adverse reactions.

All patients receive Tetanus-Diphtheria booster vaccine at time of enrollment. Cycles of standard TMZ (150-200 mg/m^2/day on days 1-5 of each 28 day cycle) with PEP-CMV vaccination on Day 23 (-1 day, + 2 days) of each TMZ cycle and tetanus pre-conditioning the day before the first vaccine.

All patients receive Tetanus-Diphtheria booster vaccine at time of enrollment. Cycles of dose-intensified TMZ (75-100 mg/m^2/day on days 1-21 of each 28 day cycle) with PEP-CMV vaccination on day 23 (-1 day, + 2 days) of each TMZ cycle and tetanus pre-conditioning the day before the first vaccine.

Chemotherapy agent FDA approved to treat newly-diagnosed glioblastoma given as cycles of 150-200 mg/m^2/day on days 1-5 of each 28 day cycle

Chemotherapy agent FDA approved to treat newly-diagnosed glioblastoma given as cycles of dose-intensified TMZ (75-100 mg/m2/day on days 1-21 of each 28 day cycle)

500 µg of PEP-CMV Component A (a synthetic long peptide) mixed with Montanide ISA-51 intradermally administered

At time of enrollment, after signing consent and undergoing immune monitoring blood work, patients will receive Tetanus-diphtheria booster vaccination with 0.5 mL of Td (tetanus, diphtheria toxoid, absorbed)

All patients will receive a tetanus pre-conditioning injection in the right groin intradermally on day 22 (±1 day) of the first post-radiation cycle of TMZ

500 µg of PEP-CMV Component B (a neutralizing antibody epitope from human CMV glycoprotein B conjugated to Keyhole Limpet Hemocyanin) mixed in 150 µg of GM-CSF intradermally administered

To assess the safety of PEP-CMV vaccination in combination with adjuvant TMZ, the percentage of patients with unacceptable toxicity will be estimated within each arm. All patients who received any PEP-CMV vaccine will be included in these analyses.

Immunologic Response as Measured by Peak Number of T Cells That Secrete IFNγ by ELISPOT in Response to Component A of PEP-CMV

The primary analysis will focus on patients who have follow-up immunologic monitoring after the 3rd vaccination with component A alone and before initiation of the second TMZ/vaccine cycle. Such a patient is considered "evaluable" for the immunologic response primary analyses. The Wilcoxon rank sum test will compare treatment groups with regard to the median peak number of T cells that secrete IFNγ by ELISPOT in response to component A of PEP-CMV. Analyses will include only those patients who have an assessment of immune response after receiving 3 vaccinations.

To determine if tumors are CMV antigen negative by immunohistochemical analysis for the presence of the antigen pp65 at the time of disease progression/recurrence

Inclusion Criteria: Age ≥ 18 years. Histopathologically proven newly-diagnosed primary glioblastoma with complete or partial surgical resection. Biopsy not acceptable. Patients must be cytomegalovirus (CMV) seropositive. The tumor must be supratentorial. Karnofsky performance status of ≥ 70. Stable or decreasing steroid dose (≤ 4 mg/day) at time of post-radiation treatment (XRT) adjuvant TMZ initiation. If patients are decreasing steroid use, once they are at 2 mg/day, they may be supplemented with physiologic replacement hydrocortisone therapy (20-30 mg/day in divided doses), at the discretion of the treating oncologist. Hematology: absolute neutrophil count (ANC) ≥ 1500 cells/µL, Platelet count ≥ 100,000 cells/µL, Hemoglobin ≥ 9.0 g/dl Chemistry: ALT/AST ≤ 3.0 times the upper limit of normal (ULN), Total bilirubin ≤ 1.5 mg x ULN (Exception: Patient has known Gilbert's Syndrome or patient has suspected Gilbert's Syndrome, for which additional lab testing of direct and/or indirect bilirubin supports this diagnosis. In these instances, a total bilirubin of ≤ 3.0 x ULN is acceptable.) Exclusion Criteria: Radiographic or cytologic evidence of leptomeningeal or multifocal disease at any time prior to study entry. Prior conventional antitumor therapy, other than steroids, RT or TMZ therapy given for glioblastoma. Pregnant or need to breast feed during the study period. Not adhering to pregnancy prevention recommendations. Active infection requiring intravenous antibiotics or an unexplained febrile (> 101.5 F) illness. Immunosuppressive disease or human immunodeficiency virus infection. Patients with unstable or severe intercurrent medical conditions such as severe heart or lung disease. Allergic or unable to tolerate TMZ for any reason. Any patient that successfully completed at least 5 weeks of Temodar during standard of care XRT/TMZ and whose blood counts meet the eligibility requirements (inclusion #7) within 4 weeks post XRT/TMZ is eligible. Patients with previous inguinal lymph node dissection, radiosurgery, brachytherapy, or radiolabeled monoclonal antibodies. Prior allogeneic solid organ transplant. Currently receiving or ever received immunosuppressive therapy for an autoimmune disorder or an organ transplant.