Abstract 1341: Inhibition of cellular protein prenylation enhances persistence and efficacy of CAR-T therapy.

Abstract Significance: Chimeric antigen receptor (CAR) T cell therapy has demonstrated great success in treating patients with hematological malignancy. However, its broader application has been constrained by the limited persistence of CAR-T cells in vivo. Here we show that inhibition of protein prenylation in human CAR-T cells can promote their persistence and tumor clearance by enhancing memory-like phenotype and reducing exhaustion. Results: By reanalyzing a human pan-cancer single-cell RNA sequencing dataset (316 patients with 21 cancer types), we discovered that genes in the mevalonate-prenylation pathway were markedly elevated in exhausted T cells but strikingly low in central memory T cells. We also found low expression level of PGGT1B in tumor-infiltrating CD8+ T cells indicated a favorable outcome to ICI therapy in individuals with melanoma using a cohort with 29 patients. Importantly, using a click-chemistry method combined, we demonstrated the over-all level of protein prenylation significantly increases in exhausted CAR-T cells. To determine whether inhibition of protein prenylation could reduce exhaustion of CAR-T cells, we treated CAR-T cells with GGTI to selectively impair the function of the geranylgeranyl-transferase type 1 (encoded by PGGT1B). Following GGTI treatment, we observed accumulation of CD45RO+ CD62L+ central-memory like cells. Moreover, GGTI treatment resulted in better tumor clearance function of CAR-T cells after multiple rounds of tumor challenge. We also transfused leukemia bearing mice with GGTI pre-treated or control CAR-T cells and found GGTI treated CAR-T cells significantly delayed leukemia relapse and extended survival of recipients. We also observed similar efficacy using osteosarcoma model. All these effects were recapitulated in shRNA mediated PGGT1B knock-down CAR-T cells. Transcriptome profiling in GGTI treated CAR-T cells showed upregulated genes associated with memory T cell differentiation (SELL, IL7R and FOXO1) and downregulated exhaustion genes (TOX, HAVCR2, ENTPD1 and LAG3). To find out down-stream effectors that affected by prenylation in CAR-T cells, we performed prenylated-protein mass spectrometry and discovered 8 proteins (RAB18, RAB8B, RAB1B, LMNA, LMNB, PPIA, RAC1 and RAP1A) that have significantly reduced prenylation levels in GGTI treated CAR-T cells. Mechanistical studies revealed that proximal TCR signaling is attenuated by PGGT1B-mediated prenylation of these protein effectors, which helps to prevent high tonic signaling induced exhaustion of CAR-T cells. Conclusion: Our findings indicate that PGGT1B-mediated protein prenylation constitutes a pivotal therapeutic target, which can be leveraged to enhance the effectiveness of T-cell-based cancer immunotherapies through reducing exhaustion and enriching memory-like T cells to provide long-term protection. Citation Format: Kai Liu, Kexin Wang, Leijin Yu, Gang Xiao, He Huang, . Inhibition of cellular protein prenylation enhances persistence and efficacy of CAR-T therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1341.