Abstract 3718: Deciphering the role of TREM2+ tumor-associated macrophages as mediators of immunoresistance in micro-satellite stable colorectal metastases
Authors: Mirza Muhammad Bilal , Cottam Matthew Alexander , Raoof Mustafa , Idrees Kamran 
Published: March 2024

Introduction: Most Colorectal Cancer (CRC)-related deaths are caused by metastatic disease, and more than 95% of CRC metastases (mCRC) are microsatellite stable (MSS) which respond poorly to Immune checkpoint inhibitors (ICI). Tumor-associated macrophages (TAMs) have been recognized as major drivers of immunoresistance in primary CRC; however, their role in mCRC remains poorly understood. We analyzed primary CRC and major metastatic sites (Liver, Peritoneum, Lungs) using single-cell RNA-sequencing (scRNAseq) to elucidate the TAM driven mechanisms of immunoresistance in the mCRC tumor microenvironment (TME) and used a novel TAM directed targeting strategy to improve responses to ICI in a preclinical orthotopic model of MSS-mCRC.

Methods: Using scRNA-seq, we analyzed 22 samples from 14 patients with mCRC. Target antigens identified were validated using flow cytometry and immunofluorescence (IF). The antitumor activity of anti-TREM2 and anti-PD1 mABs was evaluated in-vivo using CT26 CRC cell line in BALB-C mice, assessing tumor immune response via flow cytometry.

Results: We characterized the TME in MSS-mCRC and identified 17 clusters of malignant epithelial, immune, and stromal cells. Our sub-clustering analysis revealed 6 unique sub-populations of TAMs which were differentially distributed across liver, lung, and peritoneal metastases (PM).Two subsets of TAMs, 5 and 6, were enriched in PM and malignant ascites. TAMs 5/6 expressed canonical lipid-associated genes; TREM2, APOE, and PLTP. TREM2+ TAMs were also enriched in immunosuppressive and pro-angiogenic pathways, and expressed genes known to drive EMT and ECM remodeling in the TME. Thus, TREM2 was identified as a potential therapeutic target. We validated our transcriptomic findings using flow cytometry and IF analysis of MSS-mCRC showing an abundance of TREM2+ TAMs in PM. TREM2+ TAMs co-expressed CD163, known marker for M2-TAMs and stained positive for BODIPY. In an aggressive PM mouse model, treatment with anti-PD1, anti-TREM2, or combination anti-PD1/anti-TREM2 was insufficient to reduce tumor burden. However, a trend towards a reduction in total intratumoral F4/80+CD11b+ TAMs was observed in the combination treatment compared to other groups. All three treatments trended towards increased CD8+ intratumoral T-cell infiltration compared to control mice.

Conclusion: Our study provides new insights into the TME of MSS-CRC metastases at a single-cell level. Our findings indicate that TREM2+ TAMs contribute to the immunosuppressive milieu of MSS-CRC metastases. Targeting TREM2 in combination with anti-PD1 resulted in a modulation of TME via reduction of TAMs paralleled by an increased infiltration of CD8+ T cells. This reprogramming of TME suggests that inhibiting TREM2 might augment response to ICI in MSS-CRC metastases, traditionally resistant to ICI.

Citation Format: Muhammad Bilal Mirza, Matthew Alexander Cottam, Mustafa Raoof, Kamran Idrees. Deciphering the role of TREM2+ tumor-associated macrophages as mediators of immunoresistance in micro-satellite stable colorectal metastases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3718.