Abstract Brain cancer is the most common cause of cancer death in patients age 0-19. Diffuse intrinsic pontine glioma (DIPG) has a median overall survival (OS) of approximately 8 months. High grade glioma (HGG) is similarly devastating, with a median OS of 16 months, and recurrent medulloblastoma (MB) has an OS of approximately 12 months. We and others have found that cytomegalovirus (CMV) antigens are present in MB and HGG/DIPG but not in healthy brain tissue. Human CMV nucleic acids and protein epitopes in found in 70- 100% of HGG/DIPG and 92% of medulloblastoma. Given the prevalence of CMV antigen expression in HGG/DIPG and medulloblastoma, targeting CMV antigens through peptide vaccination represents a novel treatment to selectively kill tumor while leaving normal tissue unharmed. Our group has developed a peptide vaccine (PEP-CMV) directed to the CMV antigen, pp65, in children and young adults that is being evaluated in a Phase I clinical trial (NCT03299309). Our preliminary results in 17 patients have shown that there have been no ≥ 3 Grade toxicities and that vaccines elicited immune responses in 75% of patients. In this multiply recurrent cohort of patients, six of 11patients who received at least three vaccines demonstrated stable disease or partial responses on MRI. Given this data, our hypothesis is that treatment of children with a newly diagnosed HGG and DIPG or recurrent MB using the PEP-CMV vaccine will generate a robust immune response resulting in improved PFS and OS compared to historical controls. Our Primary Objectives are: 1) to determine the objective response rate to PEP-CMV in patients with recurrent MB, 2) to determine the progression-free survival at 4 months in patients with recurrent MB, 3) to determine 1 year OS in patients with newly-diagnosed DIPG, and 4) to determine 1 year PFS in patients with newly diagnosed HGG. Our Secondary Objectives are: 1) to further define treatment related toxicities of this regimen and 2) to determine 1 year PFS in patients with recurrent MB and 2 year OS in HGG. Our Exploratory objectives are 1) to describe and quantify the immune response to PEP-CMV and 2) to estimate the vascularity and permeability of tumors in response to PEP-CMV using perfusion MRI to better differentiate tumor progression from pseudoprogression. To attain the overall objectives, the following specific aims will be pursued. Aim 1: Conduct a multi-institutional trial to assess the PFS and OS of children and young adults with newly diagnosed HGG, newly diagnosed DIPG, and recurrent MB who receive PEP-CMV. Aim 2: Determine the immunogenicity and radiographic responses of patients who receive PEP-CMV. This study is impactful because it has the potential to cause a paradigm shift in the way malignant pediatric brain tumors are treated, using a novel immunotherapeutic vaccine approach in lieu of toxic radiation and chemotherapy.