Identification of breast cancer cell subtypes sensitive to ATG4B inhibition

Autophagy, a lysosome-mediated intracellular degradation and recycling pathway, plays multiple context-dependent roles in tumorigenesis and treatment resistance. Encouraging results from various preclinical studies have led to the initiation of numerous clinical trials with the intention of targeting autophagy in various cancers. Accumulating knowledge of the particular mechanisms and players involved in different steps of autophagy regulation led to the ongoing discovery of small molecule inhibitors designed to disrupt this highly orchestrated process. However, the development of validated autophagy-related biomarkers, essential for rational selection of patients entering clinical trials involving autophagy inhibitors, is lagging behind. One possible source of biomarkers for this purpose is the autophagy machinery itself. In this review, we address the recent trends, challenges and advances in the assessment of the biomarker potential of clinically relevant autophagy proteins in human cancers.

Multidisciplinary approaches are increasingly being used to elucidate the role of autophagy in health and disease and to harness it for therapeutic purposes. The broad range of topics included in the program of the Vancouver Autophagy Symposium (VAS) 2013 illustrated this multidisciplinarity: structural biology of Atg proteins, mechanisms of selective autophagy, in silico drug design targeting ATG proteins, strategies for drug screening, autophagy-metabolism interplay, and therapeutic approaches to modulate autophagy. VAS 2013 took place at the British Columbia Cancer Research Centre, and was hosted by the CIHR Team in Investigating Autophagy Proteins as Molecular Targets for Cancer Treatment. The program was designed as a day of research exchanges, featuring two invited keynote speakers, internationally recognized for their groundbreaking contributions in autophagy, Dr Ana Maria Cuervo (Albert Einstein College of Medicine, Bronx, NY) and Dr Jayanta Debnath (University of California, San Francisco). By bringing together international and local experts in cell biology, drug discovery, and clinical translation, the symposium facilitated rich interdisciplinary discussions focused on multiple forms of autophagy and their regulation and modulation in the context of cancer.

Autophagy is a lysosomal catabolic pathway responsible for the degradation of cytoplasmic constituents. Autophagy is primarily a survival pathway for recycling cellular material in times of nutrient starvation, and in response to hypoxia, endoplasmic reticulum stress, and other stresses, regulated through the mammalian target of rapamycin pathway. The proteasomal pathway is responsible for degradation of proteins, whereas autophagy can degrade cytoplasmic material in bulk, including whole organelles such as mitochondria (mitophagy), bacteria (xenophagy), or lipids (lipophagy). Although signs of autophagy can be present during cell death, it remains controversial whether autophagy can execute cell death in vivo. Here, we will introduce protocols for detecting autophagy in mammalian primary cells by using western blots, immunofluorescence, immunohistochemistry, flow cytometry, and imaging flow cytometry.

E23275 Background: During the past decade, a tremendous amount of effort has been devoted to better understanding the dynamic role of autophagy, a lysosome-mediated degradation and recycling process, in different cancers. However, little progress has been made in applying this knowledge clinically for diagnostic and therapeutic purposes. In this study, we addressed one of the outstanding challenges in the autophagy field: determining whether certain cancer subtypes are sensitive to specific autophagy-related inhibition strategies, with the longer term aim of better patient selection for relevant treatments. Methods: We examined a tissue microarray from 330 breast cancer patients to determine the prognostic value of two autophagy-related markers, LC3B and ATG4B, in all major breast cancer subtypes. Results: We found that punctate LC3B positivity significantly correlated with the triple-negative subtype and was prognostic of poor outcome (disease-specific survival) across all breast cancer subtypes, and especially in triple-negative breast cancer. We also found that ATG4B expression was significantly associated with the HER2-positive subtype. We validated this novel correlation in vitro and established a functional link between ATG4B and HER2. HER2-positive breast cancer cells demonstrated high sensitivity to ATG4B inhibition under stress, and combination of trastuzumab with ATG4B inhibition resulted in a greater reduction in cell viability than trastuzumab treatment alone. Conclusions: Together these results demonstrate an association of ATG4B with HER2-positive breast cancers and indicate that this subtype might be a good candidate for ATG4B inhibition.