Abstract 4861: INCA33890 increases CD8+ T-cell effector function compared with pembrolizumab as assessed by single-cell RNA sequencing in human PD-1xTGFβR2 knock-in mouse model

Abstract INCA33890, a novel PD-1xTGFβR2 bispecific Biclonics® antibody, is designed to selectively inhibit TGFβR2 on PD-1+ cells, leveraging a higher binding affinity for PD-1 over TGFβR2. In preclinical studies using the MC38 colorectal tumor model in human PD-1xTGFβR2 Knock-in mice, INCA33890 had a superior antitumor efficacy compared with pembrolizumab, anti-TGFβR2 or isotype control. This study aimed to uncover the differential mechanism of tumor regression by applying single-cell TCR sequencing (scTCRseq) coupled with single-cell RNA sequencing (scRNAseq) on T cells derived from treated MC38 tumor-bearing mice. Intratumoral T cells were enriched and profiled using 10X Genomics scTCRseq and scRNAseq from MC38 tumor-bearing mice treated with INCA33890, pembrolizumab, anti-TGFβR2, or control antibodies. Results revealed that CD8+ T cells in INCA33890-treated mice exhibited a robust increase in RNA transcripts associated with cytotoxicity, proliferation, and T-cell exhaustion compared with pembrolizumab or control antibody treatments, signifying a potent effector response within the tumor microenvironment. Sub-grouping of CD8+ T cells into 13 distinct clusters based on marker expression and gene enrichment analysis demonstrated a notable expansion of proliferating, activated, and cytotoxic CD8+ T cell populations following INCA33890 treatment—an outcome not observed with pembrolizumab or control treatments. INCA33890 treatment also reduced the proportion of intratumoral-naïve CD8+ T cells, transforming their gene expression profile toward an effector-like phenotype. In intratumoral CD4+ T cells, treatment with INCA33890 resulted in signatures of enhanced activation, proliferation, and exhaustion compared with pembrolizumab and control treatments, suggesting broad engagement of the adaptive immune response. scTCRseq demonstrated a marked increase in the clonality of intratumoral T cells, further underscoring the impactful immune modulation induced by INCA033890. In summary, bispecific Biclonics® targeting of TGFβR2 and PD-1 with INCA33890 promotes the expansion and activation of effector CD8+ T-cell populations within the tumor microenvironment, driving enhanced antitumor activity in vivo. Citation Format: Michelle Kinder, Jin Lu, Justine Carl, Lifeng Zhang, Paul Collier, Paul Waeltz, Rodrigo Hess, Natalia Pardo, Maria Terradas, Liang-Chuan Wang, Cynthia Timmers. INCA33890 increases CD8 T-cell effector function compared with pembrolizumab as assessed by single-cell RNA sequencing in human PD-1xTGFβR2 knock-in mouse model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4861.