Abstract A081: Chromobox 2 (CBX2) drives an immunosuppressive tumor microenvironment in high grade serous carcinoma (HGSC)

Abstract

Abstract Introduction: Epigenetic reader and component of Polycomb Repressor Complex 1 (PRC1), Chromobox 2 (CBX2) is expressed in greater than 75% of high-grade serous carcinoma (HGSC) tumors. Increased CBX2 expression conveys a poorer disease-free and overall survival. CBX2 knockdown leads to increased chemotherapy sensitivity and attenuates stem-like properties, while CBX2 overexpression is sufficient to promote chemoresistance. In an HGSC immune competent murine model, knockdown of CBX2 decreased tumor progression. We sought to explore the impact of modulation of CBX2 on the tumor immune microenvironment (TIME), understanding that TIME plays a critical role in disease progression and development of therapy resistance. Methods: We performed an exploration of existing data sets, including The Cancer Genome Atlas (TCGA), TIMERv2, and Carcinoma EcoTyper. We used HGSC cell lines (OVCAR4 and PEO1) with knockdown of CBX2 and RNA-sequencing. KEGG pathway analysis performed. A syngeneic ID8 mouse model was completed with shCBX2 compared to shControl. Nanostring Immuno-Oncology Panel and Multispectral Immunohistochemistry (mIHC) were completed to evaluate the TIME. Cell lines with genetic CBX2 modulation were utilized to create spheroids to recapitulate HGSC growth and confocal microscopy was employed to evaluate macrophage recruitment. Monocyte recruitment and macrophage polarization were evaluated using flow cytometry. Results: We found that existing data sets demonstrate a shift in TIME dependent on up or downregulation of CBX2. For instance, high CBX2 is associated with decreased CD8+ T cells (14.69% vs. 4.89%, p=0.0032) and increased undifferentiated macrophages (14.77 vs. 29.93%, p=0.006). Also, CBX2 is a defining feature of the Cell State Type 6 Carcinoma EcoType, which is associated with poor prognosis. RNA sequencing identified an association between CBX2 and CCL2, a cytokine known to recruit immunosuppressive macrophages. Pathway analysis of differentially expressed genes demonstrates an enrichment of several immune signatures. CBX2 knockdown in the ID8 syngeneic mouse model led to decreased tumor progression compared to control (11.50 mg. vs 51.63 mg, p=0.0001). NanoString analysis of the specimens suggest a shift in immune cell composition, with an increase in macrophages. mIHC defined the spatial relationships in this composition and confirmed increase in macrophage infiltration (28.64% vs. 34.60%, p=0.0231). Confocal microscopy and co-culture experiments to explore monocyte recruitment, found that both knockdown and overexpression of CBX2 lead to increased recruitment and infiltration of macrophages. Flow cytometry of macrophages cultured with CBX2 overexpressing cells showed an increase in M2-like (CD206+) macrophages and decreased phagocytosis activity. Conclusion: Increased CBX2 expression leads to increased recruitment and polarization of M2-like macrophages leading to an immunosuppressive TIME. Targeting of CBX2 may serve to modulate the TIME to enhance the efficacy of immune therapies. Citation Format: Lindsay W. Brubaker, Ritsuko Iwanaga, Tomomi M Yamamoto, Elizabeth R. Woodruff, Nicole Marjon, Siddhartha Mitra, Benjamin G. Bitler. Chromobox 2 (CBX2) drives an immunosuppressive tumor microenvironment in high grade serous carcinoma (HGSC) [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr A081.