Abstract P3-06-03: Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer
Authors: M Spears, M Braunstein, L Liao, C Yao, N Lyttle, N Lobo, KJ Taylor, PM Krzyzanowski, I Kalatskaya, R Marcellus, L Stein, P Boutros, CJ Twelves, JMS Bartlett
Journal: Cancer Research (2016)

Abstract Background: Meta-analyses performed by the Early Breast Cancer Trialists Collaborative Group demonstrated a significant increase in disease free and overall survival through the addition of anthracyclines to polychemotherapy. Anthracyclines have, however, significant toxicities including cardiotoxicity and leukaemia. It is, therefore, imperative to identify those patients who will benefit from adjuvant anthracycline treatment; other patients could then be spared unnecessary toxicities and be considered for alternative adjuvant therapy. Several markers that may predict anthracycline benefit have been explored in patient cohorts (HER2, TOP2A, Ch17CEP and TIMP1) with limited success. Methods: To identify markers that are clinically-relevant, we generated MDA-MB-231, MCF7, SKBR3 and ZR-75-1 breast cancer cell lines sensitive and resistant to epirubicin to identify pathways contributing to anthracycline resistance. A complementary approach including gene expression analyses to identify molecular pathways involved in resistance, and small-molecule inhibitors to reverse resistance were performed. RNA was extracted from patients in the BR9601 adjuvant trial evaluating the addition of epirubicin (E) to CMF and analysed through Nanostring technology. Log-rank analyses explored the predictive values of the signatures on distant relapse-free survival (DRFS). Cox-regression models tested independent predictive value on DRFS in the presence of treatment, age, tumour size, nodal status, ER status and grade, and treatment by marker interactions. Results: Gene expression analysis identified upregulaton of a histone gene module in all four cell lines which was validated by qRT-PCR. Histone deacetylase small-molecule inhibitors reversed resistance and were cytotoxic for epirubicin-resistant cell lines, with IC50's ranging from 0.1-3.69µM, confirming that histone pathways are associated with epirubicin resistance. Gene expression analysis of the 18-gene histone module in the BR9601 clinical cohort revealed that patients whose tumour had low expression had an increased DRFS (HR: 0.35, 95%CI 0.17-0.73, p=0.005) when treated with E-CMF compared with patients treated with CMF alone. Conversely, there was no apparent benefit of E-CMF vs CMF in patients with high histone module expression (HR: 0.96, 95%CI 0.58-1.59, p=0.87). After multivariate analysis and adjustment for HER2 status, nodal status, age, grade and ER status, the treatment by marker interaction was 0.35 (95%CI 0.13-0.96, p=0.042) for DRFS. Conclusion: Histone gene expression was an independent predictor of anthracycline benefit in terms of DRFS. In vitro data demonstrated that resistance could be reversed with histone deacetylase small-molecule inhibitors. The histone signature identified could be a potential theranostic candidate for patients with early breast cancer. Citation Format: Spears M, Braunstein M, Liao L, Yao C, Lyttle N, Lobo N, Taylor KJ, Krzyzanowski PM, Kalatskaya I, Marcellus R, Stein L, Boutros P, Twelves CJ, Bartlett JMS. Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P3-06-03.