The effect of menstrual cycling on genomic predictive biomarkers in premenopausal breast cancer

Abstract

Emerging in the clinic are new assays that use gene expression profiling to predict breast cancer response to treatment. A leading genomic assay is Oncotype DX, which quantifies a panel of 21 genes to calculate a Recurrence Score. The Oncotype DX 21-gene Recurrence Score is predictive of adjuvant chemotherapy benefit for women with early-stage, hormone receptor (HR)-positive, HER2-negative breast cancer and is recommended in clinical guidelines for guiding adjuvant chemotherapy treatment decisions. However, genomic assays such as Oncotype DX were largely developed in postmenopausal women and it remains unclear whether they are suitable for use in premenopausal women, where fluctuations in estrogen and progesterone during the menstrual cycle could affect genomic biomarker expression. The studies described in this thesis aimed to determine how menstrual cycling affects the Oncotype DX 21-gene signature using paired breast cancer samples and mouse models of breast cancer. To investigate how Oncotype DX Recurrence Scores fluctuate in HR-positive breast cancer, paired formalin-fixed paraffin-embedded invasive breast cancer samples were collected approximately 2 weeks apart from women <50 years old, and compared to women >50 years old and women with HR-negative disease. The Oncotype DX 21-gene signature was assessed through quantitative RT-PCR and 21-gene Recurrence Scores were calculated using the Oncotype DX Recurrence Score algorithm. Increased discordance in Recurrence Scores was observed between paired breast cancer samples collected from younger women (3.2±2.5; mean±stdev) compared to older women (2.0±1.7; p=0.04) and women with HR-negative disease (2.3±1.0; p=0.02). Linear regression analysis revealed that for every one-year decrease in age, discordance in Recurrence Scores increased by 0.07 units (p=0.0035). In young women, discordances were driven by variable expression of Proliferation- and HER2-group genes. To determine whether the ovarian cycle contributes to the increased variability in Recurrence Scores observed in younger women, HR-positive mammary tumours were dissected from naturally cycling MMTV-PyMT mice at either the estrus or diestrus phase of the ovarian cycle. Tumours collected at diestrus show significant differences in expression of 6 Oncotype DX signature genes (Ki67, Ccnb1, Esr1, Erbb2, Grb7, Bag1; p≤0.05) and a significant increase in Recurrence Score (21.8±2.4; mean±SEM) compared to tumours dissected at estrus (15.5±1.9; p=0.03). Clustering analysis revealed a subgroup of tumours collected at diestrus characterised by increased expression of Proliferation- (p<0.001) and Invasion-group (p=0.01) genes, and increased Recurrence Score (p=0.01). These tumours also exhibited higher expression of Estrogen-group genes (p=0.005) suggesting increased sensitivity to hormonal fluctuations during the ovarian cycle.