Abstract P1-09-22: Mechanistic Insight into DAXX-Modulated Sensitivity to Chemotherapy in Triple Negative Breast Cancer

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Abstract Background: Mechanisms of response and/or resistance to chemotherapy, immunotherapy, or PARP inhibitors in triple negative breast cancer (TNBC) are not well understood. Death Domain-Associated Protein 6 (DAXX) is a multifunctional protein that promotes cell death, represses gene transcription, and maintains a heterochromatin state. We showed previously that DAXX is a potent inhibitor of breast cancer stem cells and resistance to endocrine therapy in estrogen receptor positive (ER+) breast cancer (Peiffer et al., Cancer Res. 2018). In this study, we explored the role of DAXX in TNBC and sensitivity to chemotherapy and olaparib in vitro and in vivo. We also investigated mechanisms by which DAXX modulated growth and response to chemotherapy in TNBC cell lines using RNA sequencing. Methods: Proliferation and cell cycle analysis were performed for three TNBC cell lines (MDA-MB-231, BT549, and MDA-MB-468) that expressed DAXX or were depleted for DAXX using siRNA. Sensitivity to carboplatin, paclitaxel, and doxorubicin was determined by measuring proliferation. PARP-1 activity was assessed by detecting global protein PARylation levels using Western blotting. Olaparib was used to assess the role of PARP-1 activation in DAXX-depleted TNBC cell lines. Xenografts studies in vivo measured tumor growth of MDA-MB-231 DAXX-expressing or DAXX-depleted tumord treated with vehicle, paclitaxel, olaparib, or the combination. Overall survival of mice was assessed using Kaplan-Meier and a Mantel-Cox Log Rank test. RNA-sequencing was performed on RNA from two TNBC cell lines (MDA-MB-231 and MDa-MB-468) expressing DAXX or depleted for DAXX. The KMplotter tool interrogated recurrence free survival of patients with high DAXX versus low DAXX RNA expressing TNBC. Results: Depletion of DAXX increased cell proliferation of three TNBC cell lines by promoting cell cycle progression through the S-phase compared to DAXX-expressing cell lines. DAXX-depleted TNBC cell lines were 2-5 fold more sensitive to carboplatin and paclitaxel, but not to doxorubicin as measured by IC50 calculations. Depletion of DAXX increased PARP-1 activity and modestly increased sensitivity to olaparib. In vivo, DAXX-depleted TNBC tumors grew at a faster rate than DAXX-expressing tumors. Although DAXX expression did not change the anti-tumor efficacy of paclitaxel or olaparib in vivo, overall survival of mice was 100% up to 40 days when TNBC tumors had low DAXX compared to only 25% of mice surviving with TNBC expressing high DAXX and treated with paclitaxel. Mechanistically, RNA-sequencing followed by Gene Ontology pathway analysis showed that DAXX-depleted TNBC cells have higher expression of genes that regulate cell cycle progression and lower expression of genes that regulate the unfolded protein response. The KMPlotter tool revealed that patients (N=370) with TNBC have better recurrence free survival (hazard ratio = 1.79, P=0.0032) if their tumors have lower DAXX RNA levels. Conclusions: These results suggest that DAXX is a critical growth regulator and potential predictor of response to carboplatin or paclitaxel in TNBC. The mechanism by which DAXX modulates sensitivity to chemotherapy and drug resistance could be through regulation of cell cycle genes. Future studies will focus on elucidating the mechanism by which DAXX regulates these enriched genes and pathways and if DAXX is a potential predictive biomarker for recurrence free survival. Citation Format: Debra Wyatt, Michelle Fernandez, Ava Gureghian, Kathy S. Albain, Clodia Osipo. Mechanistic Insight into DAXX-Modulated Sensitivity to Chemotherapy in Triple Negative Breast Cancer [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 P1-09-22.

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