One of the most recent advances in the treatment of HER2⁺ breast cancer is the development of the antibody-drug conjugate, T-DM1. T-DM1 has proven clinical benefits for patients with advanced and/or metastatic breast cancer who have progressed on prior HER2-targeted therapies. However, T-DM1 resistance ultimately occurs and represents a major obstacle in the effective treatment of this disease. Because anti-apoptotic BCL-2 family proteins can affect the threshold for induction of apoptosis and thus limit the effectiveness of the chemotherapeutic payload, we examined whether inhibition of BCL-2/X_L would enhance the efficacy of T-DM1 in five HER2-expressing patient-derived breast cancer xenograft models. Inhibition of BCL-2/X_L via navitoclax/ABT-263 significantly enhanced the cytotoxicity of T-DM1 in two of three models derived from advanced and treatment-exposed metastatic breast tumors. No additive effects of combined treatment were observed in the third metastatic tumor model, which was highly sensitive to T-DM1, as well as a primary treatment-exposed tumor, which was refractory to T-DM1. A fifth model, derived from a treatment naïve primary breast tumor, was sensitive to T-DM1 but markedly benefited from combination treatment. Notably, both PDXs that were highly responsive to the combination therapy expressed low HER2 protein levels and lacked ERBB2 amplification, suggesting that BCL-2/X_L inhibition can enhance sensitivity of tumors with low HER2 expression. Toxicities associated with combined treatments were significantly ameliorated with intermittent ABT-263 dosing. Taken together, these studies provide evidence that T-DM1 cytotoxicity could be significantly enhanced via BCL-2/X_L blockade and support clinical investigation of this combination beyond ERBB2-amplified and/or HER2-overexpressed tumors.

The breast is a dynamic organ whose response to physiological and pathophysiological conditions alters its disease susceptibility, yet the specific effects of these clinical variables on cell state remain poorly annotated. We present a unified, high-resolution breast atlas by integrating single-cell RNA-seq, mass cytometry, and cyclic immunofluorescence, encompassing a myriad of states. We define cell subtypes within the alveolar, hormone-sensing, and basal epithelial lineages, delineating associations of several subtypes with cancer risk factors, including age, parity, and BRCA2 germline mutation. Of particular interest is a subset of alveolar cells termed basal-luminal (BL) cells, which exhibit poor transcriptional lineage fidelity, accumulate with age, and carry a gene signature associated with basal-like breast cancer. We further utilize a medium-depletion approach to identify molecular factors regulating cell-subtype proportion in organoids. Together, these data are a rich resource to elucidate diverse mammary cell states.

PURPOSE:The Adjuvant Paclitaxel and Trastuzumab trial was designed to address treatment of patients with small human epidermal growth factor receptor 2 (HER2)-positive breast cancer. The primary analysis of the Adjuvant Paclitaxel and Trastuzumab trial demonstrated a 3-year disease-free survival (DFS) of 98.7%. In this planned secondary analysis, we report longer-term outcomes and exploratory results to characterize the biology of small HER2-positive tumors and genetic factors that may predispose to paclitaxel-induced peripheral neuropathy (TIPN). PATIENTS AND METHODS:In this phase II study, patients with HER2-positive breast cancer with tumors 3 cm or smaller and negative nodes received paclitaxel (80 mg/m2) with trastuzumab for 12 weeks, followed by trastuzumab for 9 months. The primary end point was DFS. Recurrence-free interval (RFI), breast cancer-specific survival, and overall survival (OS) were also analyzed. In an exploratory analysis, intrinsic subtyping by PAM50 (Prosigna) and calculation of the risk of recurrence score were performed on the nCounter analysis system on archival tissue. Genotyping was performed to investigate TIPN. RESULTS:A total of 410 patients were enrolled from October 2007 to September 2010. After a median follow-up of 6.5 years, there were 23 DFS events. The 7-year DFS was 93% (95% CI, 90.4 to 96.2) with four (1.0%) distant recurrences, 7-year OS was 95% (95% CI, 92.4 to 97.7), and 7-year RFI was 97.5% (95% CI, 95.9 to 99.1). PAM50 analyses (n = 278) showed that most tumors were HER2-enriched (66%), followed by luminal B (14%), luminal A (13%), and basal-like (8%). Genotyping (n = 230) identified one single-nucleotide polymorphism, rs3012437, associated with an increased risk of TIPN in patients with grade 2 or greater TIPN (10.4%). CONCLUSION:With longer follow-up, adjuvant paclitaxel and trastuzumab is associated with excellent long-term outcomes. Distribution of PAM50 intrinsic subtypes in small HER2-positive tumors is similar to that previously reported for larger tumors.

Myoepithelial cells play key roles in normal mammary gland development and in limiting pre-invasive to invasive breast tumor progression, yet their differentiation and perturbation in ductal carcinoma in situ (DCIS) are poorly understood. Here, we investigated myoepithelial cells in normal breast tissues of BRCA1 and BRCA2 germline mutation carriers and in non-carrier controls, and in sporadic DCIS. We found that in the normal breast of non-carriers, myoepithelial cells frequently co-express the p63 and TCF7 transcription factors and that p63 and TCF7 show overlapping chromatin peaks associated with differentiated myoepithelium-specific genes. In contrast, in normal breast tissues of BRCA1 mutation carriers the frequency of p63⁺TCF7⁺ myoepithelial cells is significantly decreased and p63 and TCF7 chromatin peaks do not overlap. These myoepithelial perturbations in normal breast tissues of BRCA1 germline mutation carriers may play a role in their higher risk of breast cancer. The fraction of p63⁺TCF7⁺ myoepithelial cells is also significantly decreased in DCIS, which may be associated with invasive progression.