Villanueva-Meyer,5 Erin R. Bonner,6 Takahide Nejo,2 Rishi R. Lulla,7 Stewart Goldman,8 Anu Banerjee,2,3 Susan N. Chi,9 Nicholas S. Whipple,10 John R. Crawford,11 Karen Gauvain,12 Kellie J. Nazemi,13 Payal B. Watchmaker,2 Neil D. Almeida,14 Kaori Okada,2 Andres M. Salazar,15 Ryan D. Gilbert,2 Javad Nazarian,4,6 Annette M. Molinaro,2 Lisa H. Butterfield,16,17 Michael D. Prados,2,3 and Hideho Okada2,16,18 1Department of Neurology, 2Department of Neurosurgery and 3Department of Pediatrics, UCSF, San Francisco, California, USA. 4Children’s University Hospital Zurich, Switzerland. 5Department of Radiology and Biomedical Imaging, UCSF, San Francisco, California, USA. 6Children’s National Medical Center, Washington, DC, USA. 7Division of Pediatric Hematology/Oncology, Hasbro Children’s Hospital, Department of Pediatrics, The Warren Alpert Medical School of Brown University, Providence, Rhode Island, USA. 8Ann & Robert H. Lurie Children’s Hospital of Chicago, Chicago, Illinois, USA. 9Dana-Farber Cancer Institute, Boston, Massachusetts, USA. 10Division of Hematology/Oncology, Department of Pediatrics, University of Utah, Salt Lake City, Utah, USA. 11Department of Neurosciences and Pediatrics, UCSD and Rady Children’s Hospital, San Diego, California, USA. 12St. Louis Children’s Hospital, Washington University in St. Louis, St. Louis, Missouri, USA. 13Doernbecher Children’s Hospital, Oregon Health & Science University, Portland, Oregon, USA. 14The George Washington University School of Medicine and Health Sciences, The George Washington University, Washington, District of Columbia, USA. 15Oncovir Inc., Washington, DC, USA. 16Parker Institute for Cancer Immunotherapy, San Francisco, California, USA. 17Department of Microbiology and Immunology, UCSF, San Francisco, California, USA. 18Helen Diller Family Comprehensive Cancer Center, UCSF, San Francisco, California, USA. Authorship note: SM and JMT contributed equally to this work. Conflict of interest: HO is an inventor of a utility patent application titled “H3.3 CTL peptides and uses thereof” (attorney docket no: 81906-938904-220400 US, client reference no. SF15-163), which has been exclusively licensed to Tmunity Therapeutics Inc. Copyright: © 2020, American Society for Clinical Investigation. Submitted: May 29, 2020; Accepted: August 11, 2020; Published: October 26, 2020. Reference information: J Clin Invest. 2020;130(12):6325–6337. https://doi.org/10.1172/JCI140378. The Journal of Clinical Investigation CLINICAL MEDICINE 6326 jci.org Volume 130 Number 12 December 2020 40% male) in stratum B (Table 1). DMGs in stratum B included 7 thalamic tumors, 2 pontine-centered tumors that did not meet the imaging criteria for DIPGs, with extension into the cerebellum in both cases and the cervical spine in 1, and 1 spinal cord tumor located within the thoracic spine. The median time between tumor tissue collection and the start of radiation therapy (RT) was 17 days (range, 4–25 days) for patients in stratum A and 22 days (range, 2–49 days) for those in stratum B, except for 1 patient, who had a biopsy performed after RT completion. The median time from completion of RT to the first vaccine was 39 days (range, 27–71 days) for patients in stratum A and 39 days (range, 22–53 days) for patients in stratum B. Treatment and safety . The median number of vaccinations administered was 7 (IQR = 4) and 5 (IQR = 4.5) for strata A and B, respectively. Treatment was generally well tolerated with no grade 4 treatment-related adverse events (TRAEs). The most common TRAE was injection site reaction (Table 2). One patient from stra - tum A experienced a regimen-limiting toxicity (RLT), presenting with signs and symptoms of meningitis. This patient was treated according to institutional guidelines with antibiotics and was hos- pitalized until recovery. Blood and cerebrospinal fluid remained negative for meningitis, although the patient was started on antibi- otics before sample collection. After recovery from this event, the patient was taken off the study, as TRAEs could not be excluded. There were no other instances of meningitis.for cancers harboring tumor-specific neoantigens. We have iden- tified a 10 mer peptide spanning position 26–35 (H3.3K27M26–35) of the H3.3K27M protein as an HLA-A *02:01–restricted cytotoxic T lymphocyte (CTL) epitope (4). Although most missense mutation– derived neoantigens are unique to individual patients (5), a major- ity of patients with DMG and more than 70% of patients with diffuse intrinsic pontine glioma (DIPG) have the H3.3K27M muta - tion (6). The H3.3K27M-derived neoantigen epitope represents a valuable target, as it is not only tumor specific, but also K27M- mutant protein is present throughout all tumor nuclei in each of the 47 cases evaluated by immunohistochemistry (7), suggesting that this may be a truncal mutation and less likely to lead to antigen loss– mediated escape from the tumor. Within this multicenter study conducted through the Pacific Pediatric Neuro-Oncology Con- sortium (PNOC), we evaluated the safety, immunoreactivity, and efficacy of the synthetic H3.3K27M26–35 peptide in combination with the helper tetanus toxoid (TT) peptide and polyinosinic- polycytidylic acid-poly-I-lysine carboxymethylcellulose (poly-ICLC) adjuvant in patients with HLA-A *02:01+, H3.3K27M+ DMGs. Results Patient characteristics .