Abstract 5807: Differential T cell response to anti-PD1 in breast cancer Sub-types is driven by activity of intra-tumoral immune cells

Abstract Immune checkpoint inhibition (ICI) holds great promise for triple-negative breast cancer (TNBC), while it shows limited response hormone receptor positive (ER/PR+) breast cancer (CaBr). We employed the Farcast CaBr TruTumor histoculture platform, that preserves the native tumor microenvironment (TME), to study the role of tumor resident immune cell types in determining T cell activation levels on ICI treatment, in the two CaBr sub-types. Freshly resected tumor tissue samples along with matched blood were collected from consented patients. Tumor explants were generated and distributed into arms and cultured for 72 h. Media was replenished every 24 h and the supernatant was stored. The response to stimulation with anti-CD3 (100 ng/mL) + Interleukin-2 (IL-2, 100 IU/mL) and treatment with Nivolumab (132 µg/mL) was evaluated using cytokine release and flow cytometry based immune profiling. CaBr (n=115) samples had lower immune component than head and neck squamous cell carcinoma (n=113) (p<0.0001) but similar to ovarian (n=21) and renal cell (n=52) cancer prior to treatment with ICI. Amongst the two sub-types, immune component in TNBC (n=43) was higher than ER/PR+ subtypes (n=46) (p<0.05). TNBC (n=10) showed a greater proportion of lymphocyte (58.46%) compared to myeloid (39.21%) compartment. This bias was not observed in ER/PR+ samples (n=12). Anti-CD3+IL2 stimulation showed similar response between the two CaBr sub-types. Upon treatment with Nivolumab, 2 out of 4 TNBC samples exhibited a response phenotype, with >1.6-fold increase in CD8+GzmB+ cells and interferon-gamma (IFN-γ) release. In contrast, only 1 out of 4 ER/PR+ CaBr samples showed a modest increase of 1.3 fold for CD8+GzmB+ cells with no detectable IFN-γ release. To understand the basis for the differential response in the two sub-types, we studied in detail, one ER/PR+ (H1) and two TNBC (T1 and T2) samples, displaying varying levels of response. T2 and H1 did not show a Nivolumab response phenotype, whereas T1 demonstrated a strong T-cell reinvigoration and tumor cytotoxicity. Interestingly, all three samples showed anti-CD3+IL2 stimulation driven T cell response. tSNE analysis of CTLs in the control arm showed two distinct sub-populations of exhausted CTLs (CD8+PD1+). Population1 (Pop1) was Granzyme B-positive, while population2 (Pop2) was not. Upon anti-CD3 stimulation, Pop1 showed further increase of activation whereas Pop2 did not, indicative of Pop2 being an irreversibly exhausted T cell population. T2 notably had lower Pop1 and Pop2 CTLs, along with highest proportion of monocytes across all three samples pointing towards an immunosuppressive TME. H1 mainly contained over-exhausted Pop2 and negligible Pop1 CTLs. Interplay between different TME immune sub-types, thus influence response to Nivolumab in CaBr. This is effectively captured by the TruTumor platform. Citation Format: Mouniss MM, Biswajit Das, Kowshik Jaganathan, Syamkumar V, Moumita Nath, Chandan Bhowal, Dharanidharan M, Saikrishna S, Abdul Haseeb, Pallavi R, Kubera Chandran, Rajashekar M, Oliyarasi M, Méhul Kapur, Jayaprakash C, Venkatesh T, Ganesh MS, Amritha Prabha, Prakash BV, Ravi Krishnappa, Upendra K, Ritu Malhotra, Govindaraj K, Pavithira ., Mohit Malhotra, Nandini Pal Basak, Satish Sankaran. Differential T cell response to anti-PD1 in breast cancer Sub-types is driven by activity of intra-tumoral immune cells [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 5807.