scans, laboratory values) will be provided upon request. 12.3 Multicenter Guidelines NA 12.4 Collaborative Agreements Language NA STATISTICAL CONSIDER ATIONS 13.1 Study Design/Endpoints This is a pilot study to test the potential utility of PBR28 PET in measuring treatment related inflammation . We will measure if increased uptake of PBR28 in the tumor during treatment is associated with longer time to progression (i.e. suggesting that the response to therapy is inflammatory rather than tumor growth). 13.2 Sample Size, Accrual Rate and Study Duration Protocol A pilot study to evaluate PBR PET as an indicator of inflammation in brain tumor patients treated with chemoradiation or immunotherapy. Version Date: June 20, 2018 31 Total sample size is anticipated to be 40 patients: 15 patients in cohort A (metastatic brain tumor patients) and 15 in cohort C (GBM patients suspected of having pseudoprogression) and 10 in cohort B (GBM patients treated with immunothera py). Several factors influence the sample size for this study. This is a non- therapeutic study in a patient population which typically chooses to enroll in therapeutic trials if they are going to participate in a clinical trial at all so this trial will b e less appealing to patients . In addition, since PBR PET remains investigational it is a very expensive test so budgetary constraints limit accrual . In order to assess if PBR PET helps to assess tumor inflammation, we will quant itate PBR uptake in the tumor and correlate that with tumor progression as determined by the treating physician or tissue histology if obtained. The hypothesis is that if tumor PBR retention is high, this would be consistent with a strong response to immunotherapy and should be associated with prolonged progression free survival. If PBR retention is low, indicating an immunosuppressive, active tumor microenvironment, PFS should be short. We selected 9 months because by this time, we should know who has true progression since trea tment related inflammation has peaked by this time. The aim of this study is to demonstrate higher levels of PBR uptake in the non-progression group. Therefore the sample size calculation is based on two group means comparison and the test used is a one-sided two- sample equal variance t -test (with mean and SD estimates as cited below). Since there is no a priory indication for the breakdown between progression and non-progression, we assumed equal group sizes. With 15 patients in cohort A (metastatic brain tumor patients) and 15 in cohort C (GBM patients suspected of having pseudoprogression), the study will have at least 80% power to detect a 60% difference in PBR uptake between those progression free at 9 months and those not progression free at 9 months using a one- side hypothesis test with a 0.1 significance level. By 9 months, we should know who has true progression and who has pseudoprogression. Patients who die before nine months will be considered true tumor progression. Given this conservative approach we will have the power to detect a smaller difference in PBR uptake. For the patients who have baseline and mid treatment PET (cohort A), we will use the change in PBR uptake. There is limited data on PBR28 in brain tumors so this calculation assum es a baseline of 125 ml/cm 3 and equal variance of 100 ml/cm3 (based on literature repo rted rates from healthy brain) .9 PFS at 9 months was selected as most patients would have progressed at 9 months. All patients will also be followed for 3 years after study completion for progression and survival. Time to event analysis will be used to incorporate other potential prognostic factors such as age and functional status. Protocol A pilot study to evaluate PBR PET as an indicator of inflammation in brain tumor patients treated with chemoradiation or immunotherapy. Version Date: June 20, 2018 32 Given the small number of patients anticipated to accrue to cohort B (GBM patients treated with immunotherapy), th ese patients will be analyzed with descriptive statistics alone. This accrual is anticipated to be low because of the small number of GBM patients currently treated with immunotherapy. We anticipate enrolling a maximum of 10 patients in this cohort. Accrual Targets Ethnic Category Sex/Gender Females Males Total Hispanic or Latino 2 + 2 = 4 Not Hispanic or Latino 3 + 8 = 11 Ethnic Category: Total of all subjects 5 + 10 = 15 Racial Category American Indian or Alaskan Native 0 + 0 = 0 Asian 0 + 1 = 1 Black or African American 0 + 1 = 1 Native Hawaiian or other Pacific Islander 0 + 0 = 0 White 5 + 8 = 13 Racial Category: Total of all subjects 5 + 10 = 15 13.3 Stratification Factors NA 13.4 Interim Monitoring Plan NA 13.5 Analysis of Primary Endpoints As above Protocol A pilot study to evaluate PBR PET as an indicator of inflammation in brain tumor patients treated with chemoradiation or immunotherapy. Version Date: June 20, 2018 33 13.6 Analysis of Secondary Endpoints Given the small sample size, any secondary endpoint analysis will be exploratory and reported as descriptive. 13.7 Reporting and Exclusions 13.7.1 Evaluation of T oxicity Patients will be evaluable for toxicity related to PBR PETs within 3.5 hours of the PET scan. 13.7.2 Evaluation of the Primary Efficacy Endpoint NA PUBLICATION PLAN Dr. Gerstner will have sole responsibility for publishing results of this study. Protocol A pilot study to evaluate PBR PET as an indicator of inflammation in brain tumor patients treated with chemoradiation or immunotherapy. Version Date: June 20, 2018 34 REFERENCES 1.