Supporting data for “Androgen receptor modulates chemotherapy response in breast cancer”

Breast cancer ranks as the second most diagnosed cancer globally and the most common among women. Chemotherapies, encompassing both systemic and targeted approaches, are critical for managing the disease, yet drug resistance often compromises treatment efficacy. The androgen receptor (AR), a widely expressed hormonal receptor in breast cancer, has an unclear role in disease progression. Emerging evidence suggests AR may contribute to drug resistance in various cancers, including breast cancer; however, the underlying mechanisms remain underexplored.

This study investigated the role of AR in chemotherapy response using two AR-positive breast cancer cell lines. We assessed responses to multiple chemotherapeutic agents—palbociclib, docetaxel, doxorubicin, paclitaxel, and cisplatin. In vitro and in vivo results revealed that AR activation reduced cell sensitivity to palbociclib and doxorubicin, indicating that AR modulated responses to these drugs. For palbociclib, a CDK4/6 inhibitor, AR activation induced RB1 polyubiquitination and degradation, diminishing RB1 levels—a key downstream target of CDK4/6—and thus impairing palbociclib efficacy. Chromatin immunoprecipitation (ChIP) sequencing identified 1,209 AR-regulated genes, with gene ontology (GO) analysis highlighting significant enrichment of ubiquitin-related genes. qPCR validation pinpointed TRIM37, an AR-regulated E3 ligase, as a mediator of RB1 degradation. TRIM37 overexpression decreased palbociclib sensitivity, while its knockdown enhanced sensitivity and attenuated the effect of AR on RB1 levels and drug response, establishing TRIM37 as a novel E3 ligase that bound RB1 via its MATH and C-terminal domains.

For doxorubicin, a DNA-damaging agent, AR activation mitigated induced DNA damage, suggesting a role in DNA repair. GO analysis of AR-regulated genes from ChIP sequencing implicated DNA damage-binding genes in AR-mediated doxorubicin response. qPCR validation identified CRY2 as an AR-regulated gene reducing DNA damage. CRY2 overexpression lowered doxorubicin sensitivity, whereas its knockdown increased sensitivity and diminished the influence of AR on drug response. Mechanistically, CRY2 enhanced non-homologous end joining (NHEJ) repair—rather than homologous recombination (HR), by binding to NHEJ proteins (DNA-PKcs, Artemis, Ku80, Lig4, and XLF). Interactions of CRY2 to DNA-PKcs, Artemis, Ku80, and Lig4 strengthened post-doxorubicin treatment. Inhibition of NHEJ with the DNA-PKcs inhibitor KU-57788 reversed the protective effect of CRY2, confirming AR modulates doxorubicin response via CRY2-driven NHEJ activation.

In conclusion, this study elucidates AR-mediated resistance mechanisms to palbociclib and doxorubicin in breast cancer through TRIM37 and CRY2, respectively. These findings highlight AR as a potential therapeutic target and underscore the need for further clinical validation to optimize treatment strategies for AR-positive breast cancer patients.

These are the supporting data for the PhD thesis of “Androgen receptor modulates chemotherapy response in breast cancer”. The data included western blot, luciferase assay, cell viability assay, qPCR, ChIP qPCR, cell cycle assay, CoIP assay, Ubiquitination assay, IF staining, animal study, DNA repair assay including HR and NHEJ activities, and TUNEL assay. The procedures of the assays were described in the thesis.