HSP90AA1 Knockdown Enhances Doxorubicin Sensitivity by Promoting Immunogenic Cell Death-Related Signaling and Immune-Related Tumor Microenvironment Remodeling in Breast Cancer

Background: Doxorubicin (DOX) is widely used in breast cancer treatment; however, therapeutic resistance and an immunosuppressive tumor microenvironment remain major obstacles to effective therapy. Immunogenic cell death (ICD) contributes to chemotherapy-induced antitumor immunity, but the molecular regulators linking DOX response, ICD-related signaling, and immune infiltration in breast cancer remain incompletely understood. This study aimed to identify and experimentally validate HSP90AA1 as a potential regulator of DOX response and ICD-associated immune remodeling in breast cancer. Methods: ICD-related genes were analyzed using public breast cancer transcriptomic datasets to construct prognostic models and evaluate their associations with survival, immune infiltration, immune checkpoint expression, and HLA-related molecules. HSP90AA1 was selected as a candidate gene for further validation. HSP90AA1 expression was examined in breast cancer tissues and cell lines. HSP90AA1 knockdown was performed in breast cancer cells, followed by DOX treatment. Western blotting was used to detect apoptosis-related proteins, ICD-related markers, and immune-associated molecules, including cleaved-caspase-3, BAX, BCL-2, CALR, HMGB1, p-eIF2α/eIF2α, and PD-L1. In vivo, a 4T1 syngeneic breast cancer mouse model was established to assess the effects of HSP90AA1 knockdown combined with DOX on tumor growth and tumor immune-related protein expression. Results: Bioinformatic analyses showed that ICD-related molecular patterns were associated with prognosis and immune microenvironment features in breast cancer. HSP90AA1 was upregulated in breast cancer and associated with unfavorable survival, supporting its potential relevance as an ICD-related prognostic candidate. In vitro, HSP90AA1 knockdown enhanced DOX-induced apoptotic responses, as indicated by increased cleaved-caspase-3 and BAX expression and decreased BCL-2 expression. HSP90AA1 knockdown also strengthened DOX-induced ICD-related molecular changes, including increased CALR, HMGB1, and p-eIF2α/eIF2α levels, accompanied by altered PD-L1 expression. In vivo, Hsp90aa1 knockdown combined with DOX resulted in a greater reduction in terminal tumor burden than DOX treatment alone. Tumor tissues from the combination group showed increased CALR, HMGB1, CD8A, and GZMB expression, together with altered PD-L1 expression. Tumor tissues from the combination group showed increased CALR, HMGB1, CD8A, and GZMB expression, together with altered PD-L1 expression, indicating ICD-related and immune-associated molecular changes. Conclusions: This study identifies HSP90AA1 as a potential ICD-related regulator of DOX response in breast cancer. HSP90AA1 knockdown may enhance the response to DOX by promoting apoptotic responses and ICD-related molecular changes, together with immune-related tumor microenvironment remodeling. These findings provide experimental evidence supporting HSP90AA1 as a potential therapeutic target for improving chemotherapy response in breast cancer.