Locoregional Recurrence Is Associated With Genomic Down-Regulation of the Immune Response in Women with Locally Invasive Breast Cancer

Abstract

Sustained locoregional control of breast cancer is a significant issue in patients with high risk disease. We recently reported the results of a phase I clinical trial describing acute and late toxicity of PARP1 inhibition (PARPi) with concurrent veliparib and RT. The purpose of this study was to determine predictive biomarkers of acute (during treatment) and late (1, 2, and 3 year) toxicity in patients treated with PARPi and RT from that trial. We hypothesized that protein and gene expression changes in skin epithelial cells after concurrent PARPi and RT might be used as predictive biomarkers of toxicity. Materials/Methods: Acute toxicity was any dose-limiting toxicity defined in the protocol; late toxicity was defined as any CTCAE v4 Grade 3 event, regardless of attribution. Skin punch biopsies from the irradiated field were taken from patients at baseline (pretreatment TZ0) and 6 hrs after first fraction of RT alone (TZ1). Veliparib was then started and a 3rd punch taken 6 hrs after the second fraction of RT (TZ2). RNA was isolated from skin epithelial cells and gene expression assessed using Affymetrix Human Gene ST 2.1 arrays. GSEA and MSigDB software was used for analysis. Significant difference was predetermined as a log2 fold change of+/-1.5 and FDR adjusted p-value <0.05. Protein changes (pgH2AX, PAR, Ki67) were detected using IHC and quantitated using Aperio Digital Path software. Results: Severe acute and late toxicity rates with combined therapy were 15% (5/33) and 21% (7/33), respectively. Acute toxicity was not a predictor of late toxicity. Indeed, the two were almost mutually exclusive. Gene expression analysis identified 31 genes whose expression was significantly different 6 hrs after RT and 54 genes differed after combined treatment, including genes associated with DNA damage repair (ATM, MDM2, XPC) and proliferation (Ki67, TP53TG1). 67 genes were associated with acute toxicity, including overrepresentation of miRNAs associated with gene repression. Additionally, 63 genes were associated with late toxicity and were associated with metabolism, inflammation, and DNA damage response. There was no overlap in biomarkers of acute and late toxicity. Both pgH2AX and PAR protein levels increased and Ki67 decreased significantly (more than 2 fold for all proteins) with RT and combined therapy (T1 and T2) but were not significantly associated with acute or late toxicity at any time point. Conclusion: This demonstrates the feasibility of using skin punch biopsy in the irradiated field for biomarker analysis and identification of a number of putative biomarkers of early and late radiation toxicity following combined PARPi and RT treatment. Additionally, for the first time in patient skin punch samples we report (1) gene expression changes in acute responding genes shortly after RT and (2) decreased proliferation after combined treatment. Future trials will validate the utility of these biomarkers for predicting toxicity and will seek to identify biomarkers of treatment efficacy.