Quantitative Proteomic Profiling Identifies a Potential Novel Chaperone Marker in Resistant Breast Cancer.

Karen M Gallegos,Wensheng Zhang,Shawn D Llopis,Rashidra R Walker,Kun Zhang,Syreeta L Tilghman,A Michael Davidson,Jankiben R Patel

doi:10.3389/fonc.2021.540134

Abstract

Development of aromatase inhibitor resistant breast cancer among postmenopausal women continues to be a major clinical obstacle. Previously, our group demonstrated that as breast cancer cells transition from hormone-dependent to hormone-independent, they are associated with increased growth factor signaling, enhanced cellular motility, and the epithelial to mesenchymal transition (EMT). Given the complexity of cancer stem cells (CSC) and their implications on endocrine resistance and EMT, we sought to understand their contribution towards the development of aromatase inhibitor resistant breast cancer. Cells cultured three dimensionally as mammospheres are enriched for CSCs and more accurately recapitulates tumors in vivo. Therefore, a global proteomic analysis was conducted using letrozole resistant breast cancer cells (LTLT-Ca) mammospheres and compared to their adherent counterparts. Results demonstrated over 1000 proteins with quantitative abundance ratios were identified. Among the quantified proteins, 359 were significantly altered (p < 0.05), where 173 were upregulated and 186 downregulated (p < 0.05, fold change >1.20). Notably, midasin, a chaperone protein required for maturation and nuclear export of the pre-60S ribosome was increased 35-fold. Protein expression analyses confirmed midasin is ubiquitously expressed in normal tissue but is overexpressed in lobular and ductal breast carcinoma tissue as well as ER+ and ER- breast cancer cell lines. Functional enrichment analyses indicated that 19 gene ontology terms and one KEGG pathway were over-represented by the down-regulated proteins and both were associated with protein synthesis. Increased midasin was strongly correlated with decreased relapse free survival in hormone independent breast cancer. For the first time, we characterized the global proteomic signature of CSC-enriched letrozole-resistant cells associated with protein synthesis, which may implicate a role for midasin in endocrine resistance.

Highlights

Aromatase inhibitors (AI), e.g., letrozole, are the first-line treatment for estrogen receptor positive (ER+) breast cancer in post-menopausal women
To examine whether increased midasin expression was exclusive to the LTLT-Ca mammospheres, we investigated midasin expression across various cell lines including letrozole-sensitive and letrozole-resistant cell lines
When the 3D mammospheres were compared to their 2D culture counterparts, midasin expression was significantly increased in the letrozole resistant T47D breast cancer cells (T47DaromLR) (Figure S1A, B) as well as the MDA-MB-231 triple negative breast cancer cells (Figure S1C, D)

Summary

Introduction

Aromatase inhibitors (AI), e.g., letrozole, are the first-line treatment for estrogen receptor positive (ER+) breast cancer in post-menopausal women. The mortality rates for ER+ breast cancer in the US have declined due to successful endocrine therapy (AI and selective estrogen receptor modulators, SERMS), the development of resistance is still a critical lingering problem. In order to develop improved therapies for highly metastatic breast tumors, it is crucial to uncover the molecular underpinnings that drive resistance and proliferation. An exacerbating factor is the emergence of a small sub-population of breast cancer stem cells (CSC) in AI-resistant breast cancer [8]. This is a rate-limiting factor as these pluripotent cells are de novo resistant to radiation and chemotherapy. To gain an insight of those molecular mechanisms, we identified a chaperone, midasin, as a potential marker for AI-resistant cancer in CSC-enriched mammospheres

Methods

Results

Conclusion