Abstract PS12-04: HDACi combined with anthracycline elicits interactions between MHC-II+ triple-negative breast cancer and CD69+CD4+Trm orchestrating synergistic immunotherapy

Abstract Background: Early TNBC (eTNBC) lacks biomarkers for predicting the benefits of immune checkpoint blockade (ICB). Therefore, identifying indicators for the high-benefit patients and finding synergistic target for low-benefit populations are crucial. Tumor-associated MHC-II (tsMHC-II) are linked with the benefits of ICB in eTNBC, though the mechanisms remain to be explored. This study aimed to research the interaction mechanisms and translational targets between tsMHC-II and CD4+ T cell subgroups by NeoTennis clinical trial cohorts. Method: RNA-seq, mIHC, and spatial analysis were used to quantify MHC expression in the tumor and spatial relations within the TME. In vivo murine cohorts combined with CyTOF were utilized to elucidate the mechanisms of tsMHC-II and CD4+ Trm cells in chemo-ICB. Co-culture of cell line, organoid, and T cell were used to investigate the regulatory factors of tsMHC-II. TMA and sc-RNA seq were used to explore the clinical significance, molecular characteristics of MHC-II+ tumor. Results: The NeoTennis trial included eTNBC patients in a two-phase clinical trial involving anthracycline induction followed by sequential nab-paclitaxel and PD-1 inhibitor (Toripalimab). Analysis of baseline and anthracycline-induced biopsy samples using mIHC (Pan-CK, HLA-A, HLA-DR, CD4, CD8, CD69, Foxp3, T-bet) allowed for the clustering of patients based on MHC-I and MHC-II expression in the tumor epithelium, resulting in four subtypes. The ROC predictive model based on the outcomes and MHC subtypes was constructed to find that the MHC-I+ MHC-II+ subtype and tsMHC-II had the highest AUC scores (0.811 and 0.848, respectively), whereas the CPS was only 0.419. Meanwhile, tsMHC-II expression in the pCR group was significantly higher than in the Non-pCR group at baseline while MHC-I showed no difference. Spatial analysis revealed that CD69+CD4+ resident-memory T cells (Trm) had the strongest spatial association with MHC-II+ tumors, with this relationship being most prevalent in the pCR group. Dynamic analysis of changes before and after anthracycline showed that the proportion of MHC-II+ tumors and CD4+ Trm cells increased in the pCR group after treatment. In vivo models combined with CyTOF revealed that there was a significant increase in CD4+ Trm cells, Th1 cells, and iNOS+ macrophages in the tumor after anthracycline. After knocking out the MHC-II molecules in tumor cells, the anthracycline-induced increase in CD4+ Trm cells and Th1 cells was reduced, and the tumors grew faster than the control group. Conversely, when CD4+ T cells were depleted following doxorubicin-treated, the efficacy of sequential anti-PD1 was diminished. The above findings showed that anthracyclines could upregulate tsMHC-II, leading to the production of CD4+ Trm. Analysis of TMA from 300 cases in FDSCC and sc-RNA seq of TNBC confirmed that MHC-II+ TNBC was associated with an extremely good prognosis (HR=0.28, 95%CI=0.12-0.64; p=0.003), while 22.4% of TNBC patients had high tsMHC-II. Moreover, the tsMHC-II+ tumor cells exhibited upregulated IFN-gamma response. RNA-seq showed that patients resistant to anthracycline-induced MHC-II upregulation had highly active HDAC. In vitro, HDAC inhibitors (Entinostat) could upregulate MHC-II in TNBC cell line and PDOs, thereby promoting increased contact with CD4+ T cells. In vivo, combining anthracyclines with Entinostat increased the infiltration of CD4+ Trm cells, Th1 cells, and CD8+ T cells, and inhibited tumor proliferation, after which, sequential anti-PD1 therapy further inhibited tumor growth for a longer time. Conclusion eTNBC patients with high tsMHC-II expression benefited from neoadjuvant anthracycline with sequential anti-PD1 ICB. Anthracyclines could promote the upregulation of tsMHC-II in tumor cells and induce the production of CD4+ Trm. For anthracyclines-resistant patients, combining neoadjuvant anthracycline with HDACi could significantly enhance the effectiveness of ICB. Citation Format: Zehao Wang, Xue Jingyan, Xiu Bingqiu, Wu Jiong. HDACi combined with anthracycline elicits interactions between MHC-II+ triple-negative breast cancer and CD69+CD4+Trm orchestrating synergistic immunotherapy [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr PS12-04.