Genomic correlates of clinical CAR-T cell activity

Chimeric antigen receptor (CAR)-T cell therapies demonstrate potent anti-tumor efficacy in hematologic malignancies, yet clinical outcomes remain unpredictable due to the bespoke nature of the treatment, which is manufactured from each patient’s own T-cells. While germline variants are known to influence response to immune checkpoint inhibitors ^{1, 2} , their role in CAR-T cell therapy is unknown. Here, we pair whole-genome germline sequencing of lymphoma patients from the ZUMA-1 ³ and ZUMA-7 ⁴ clinical trials of axicabtagene ciloleucel, along with correlative biomarkers and functional assays, to ascertain the impact of germline variants on CAR-T cell behavior. Hypothesizing shared mechanisms of the most common toxicities of CAR-T cells, namely cytokine release syndrome (CRS) with hemophagocytic lymphohistiocytosis (HLH)—a hyperinflammatory syndrome driven by T cell overactivation ⁵ , we first looked within 17 canonical HLH-associated genes ⁶ , and identified putative deleterious STXBP2 variants in 15% of ZUMA-1 patients with toxicity, which were absent in control subjects who did not experience high grade toxicity. Subjects with these variants had elevated baseline IFN-γ and inflammatory cytokines, findings that were recapitulated in engineered STXBP2 -deficient and STXBP2 -variant-expressing primary CAR-T cells derived from healthy donors. However, STXBP2 variant enrichment was absent in ZUMA-7 for this toxicity phenotype, possibly reflecting differences in underlying disease burden and evolving clinical management between the trials. A more expansive genome-wide analysis revealed ADAMTSL3 (a negative regulator of TGFβ ⁷ ) as the only gene nominally enriched for putative deleterious variants in both ZUMA-1 and ZUMA-7 among control subjects, suggesting a protective effect. Finally, we focused on associations between germline variants and CAR-T cell expansion after infusion, a more objective and granular continuous variable that is strongly associated with clinical response across most CAR-T products ⁸ . We found a strong association between PTPN22 , a known negative regulator of T-cell activation ^9–12 and an autoimmune risk gene ^{13, 14} , variant status and CAR-T cell expansion in both ZUMA-1 and ZUMA-7, with the patients having the highest level of CAR-T expansion across clinical trials harboring variants in the gene. Together, these findings demonstrate the first clear association between germline variants and the clinical behavior of engineered immune cell therapies, which has implications for cellular therapy design, monitoring, testing, clinical trial design, and patient care.