Abstract Background Primary central nervous system lymphoma (PCNSL) is a unique type of non-Hodgkin lymphoma restricted to the central nervous system. Most cases are diffuse large B-cell lymphomas, but prognosis is far less favorable compared to extracerebral cases. Many mutations found in PCNSL converge on the NF-κB signaling pathway, leading to downstream activation of B-cell receptor (BCR) and NF-κB pathway. The differences in mutation frequency when compared to non-CNS lymphoma and how mutational profiles affect treatment response are not well understood. We compared the genomic profile of PCNSL and non-CNS lymphoma, evaluated the association between specific molecular alterations and survival, and analyzed the association between various mutations and response to therapy. Materials and Methods A total of 60 patients with PCNSL, 8 with secondary CNS lymphoma, and 202 with non-CNS B-cell lymphomas underwent molecular profiling at Caris Life Sciences (Phoenix, AZ). Analyses included next-generation sequencing of DNA (592 Genes, NextSeq or WES, NovaSeq) and RNA (WTS, NovaSeq). X2/Fisher’s-exact U tests were used for comparison, and significance was determined as p-value adjusted for multiple comparison by the Benjamini-Hochberg method (q<0.05). Overall survival (OS) was calculated from the start of temozolomide (TMZ) or tissue collection to last contact using insurance claims data. Results When compared to non-CNS lymphomas, PCNSL tumors showed significantly higher mutation rates in MYD88 (70% vs. 7%), PIM-1 (58% vs. 7%), CD79B (42% vs. 3%) and higher rates of TMB-High (cutoff 10mut/MB; 28% vs. 10%) (q<0.01). In addition, mutations in CARD11 (12% vs. 4%) and IRF4 (8% vs. 1%) trended to be more prevalent in PCNSL (p<0.05). In contrast, mutations in KMT2D (35% vs. 16%), EZH2 (13% vs. 2%), CREBBP (22% vs. 7%), TNFRSF14 (10% vs. 0) and BCL2 (19% vs. 2%) trended to be more prevalent in non-CNS lymphomas compared to PCNSL (p<0.05). In patients with PCNSL, there was no difference in survival in those with or without MYD88, PIM1 or CD79B mutations. No between-group differences were observed in the small cohort of patients with secondary CNS lymphoma (n=8). In the subgroup in whom these data were available, 25% of tumors were MGMT methylated, 71% had a “high or intermediate” tumor mutational burden, 8% were MSI high, and 54% were PD-L1 expression positive when tested by immunohistochemistry. Patients with PCNSL had worse overall survival (OS) than non-CNS lymphomas (30 vs 81 months, p<0.001). Conclusion MYD88, PIM-1, and CD79B mutations are more frequent in PCNSL compared to non-CNS lymphomas, but none of these mutations affected OS. Only 25% of PCNSL patients are MGMT methylated, but a majority have expression of PD-L1 suggesting a benefit from PD-1-targeted therapy. Response and survival in patients with mutation-guided therapy will be presented.