Abstract P3-11-02: CST1 Interaction with RAB1B Modulates Tamoxifen resistance of Breast Cancer by Regulating Autophagy

Abstract

Abstract Background: Breast cancer is a prominent cause of cancer-related death among women worldwide. Approximately,70% of breast cancers (BC) are estrogen and progestogen receptor-positive (HR+) at diagnosis. Adjuvant endocrine therapy is a pivotal component of treatment for this type of patient. Tamoxifen (TAM), a widely recommended drug in endocrine therapy, is confronted with a high risk of recurrence after 5 years of treatment among breast cancer patients, which shows big clinical demands. Therefore, it is urgent to find new targets for tamoxifen resistance therapy. Recently, our team found that enhanced expression of Cystain SN (CST1) was detected in tissues that had undergone tamoxifen treatment appearing to recurrence and metastasis in ER-positive breast cancer patients. However, the molecular mechanisms of modulating tamoxifen resistance remain unknown. Therefore, our team suspected that there might be a correlation between the abnormal expression of CST1 and tamoxifen resistance. To investigate whether CST1 is involved in tamoxifen resistance and its molecular mechanism, we conducted further research. It is critical to elucidate the molecular mechanisms of tamoxifen resistance and to explore a novel effective therapeutic target. Method: Enhanced expression of CST1 was confirmed by immunohistochemistry in breast cancer tissue samples and by quantitative real-time PCR in Tamoxifen-resistant cancer cell lines MCF-7R/T47DR. To further investigate the molecular function of CST1, we knockdown its expression in MCF-7R/T47DR cells by transfecting with the lentiviral particles of shCST1 and corresponding negative control (Shanghai Genechem Co., Ltd.) and overexpressed CST1 in MCF-7/T47D cells using ove-GST-CST1, ove-HA-RAB1B plasmids. The effects of CST1 on cell viability and proliferation in breast cancer cells were detected by cell counting kit-8 and colony formation assays, respectively. The interaction between CST1 and RAB1B was validated by Co-immunoprecipitation and Western blot. Autophagosomes were found using transmission electron microscopy (TEM) along with Western blotting to detect the transformation of LC3-I to LC3-II protein (i.e., the LC3-II/LC3-I ratio) and P62 protein levels. Results: We found that aberrant expression of Cystatin SN (CST1) was identified in ER-positive breast cancer (BC) cells (Figure 1) and which is crucial in contributing to Tamoxifen resistance (Figure 2). The direct promotion of autophagy by RAPA can faciliate cells to endocrine resistance. And knockdown of CST1 not only suppressed autophagy in Tamoxifen-resistant cells but also attenuated the resistance, which determined the correlation of CST1 with autophagy in BC (Figure 3). Rab1b (Ras-related protein Rab-1B), a membrane protein essential for autophagosome formation, was verified to have interaction with CST1 through Coimmunoprecipitation (CO-IP) experiments (Figure 4). Meanwhile, RAB1B expression was also reduced by CST1 knockdown in the resistant cells. The interaction between CST1 and RAB1B induces autophagy which decreases Tamoxifen’s therapeutic efficacy. Furthermore, rescue experiments suggested that CST1 knockdown–induced sensitization in the efficacy of TAM Therapy and decreased autophagy could be restored via RAB1B overexpression.Conclusions: These results indicate that CST1 interacts with RAB1B to facilitate BC resistant to TAM through regulation of autophagy and CST1 might be a potential therapeutic target to overcome endocrine therapy resistance. Figure 1. Enhanced expression of CST1 in Tamoxifen-resistance breast cancer Figure 1. Enhanced expression of CST1 in Tamoxifen-resistance breast cancer Figure 2. CST1 expression is positively correlated with autophagy in ER-positive breast cancer Figure 2. CST1 expression is positively correlated with autophagy in ER-positive breast cancer Figure 3. CST1 promotes cells resistant to tamoxifen therapy by activating auophagy Figure 3. CST1 promotes cells resistant to tamoxifen therapy by activating auophagy Figure 4. CST1 interacts with RAB1B in cell autophagy Figure 4. CST1 interacts with RAB1B in cell autophagy Citation Format: li liu. CST1 Interaction with RAB1B Modulates Tamoxifen resistance of Breast Cancer by Regulating Autophagy [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-11-02.