Proteomic and transcriptomic profiling identifies mediators of anchorage-independent growth and roles of inhibitor of differentiation proteins in invasive lobular carcinoma

Nilgun Tasdemir,Nancy E Davidson,Adrian V Lee,Steffi Oesterreich,Ashuvinee Elangovan,Tian Du,Kevin M Levine,Laura Savariau,Kai Ding,Emily A Bossart

doi:10.1038/s41598-020-68141-9

Abstract

Invasive lobular carcinoma (ILC) is a histological subtype of breast cancer with distinct molecular and clinical features from the more common subtype invasive ductal carcinoma (IDC). ILC cells exhibit anchorage-independent growth in ultra-low attachment (ULA) suspension cultures, which is largely attributed to the loss of E-cadherin. In addition to anoikis resistance, herein we show that human ILC cell lines exhibit enhanced cell proliferation in ULA cultures as compared to IDC cells. Proteomic comparison of ILC and IDC cell lines identified induction of PI3K/Akt and p90-RSK pathways specifically in ULA culture in ILC cells. Further transcriptional profiling uncovered unique upregulation of the inhibitors of differentiation family transcription factors ID1 and ID3 in ILC ULA culture, the knockdown of which diminished the anchorage-independent growth of ILC cell lines through cell cycle arrest. We find that ID1 and ID3 expression is higher in human ILC tumors as compared to IDC, correlated with worse prognosis uniquely in patients with ILC and associated with upregulation of angiogenesis and matrisome-related genes. Altogether, our comprehensive study of anchorage independence in human ILC cell lines provides mechanistic insights and clinical implications for metastatic dissemination of ILC and implicates ID1 and ID3 as novel drivers and therapeutic targets for lobular breast cancer.

Highlights

Invasive lobular carcinoma (ILC) is a histological subtype of breast cancer with distinct molecular and clinical features from the more common subtype invasive ductal carcinoma (IDC)
We have previously shown that the estrogen receptor alpha (ER)-positive human ILC cell lines MDA-MB-134 (MM134), SUM44PE (SUM44), MDA-MB-330 (MM330) and BCK4 can grow efficiently in ultra-low attachment (ULA) suspension culture, as compared to the limited growth of the ER-positive human IDC cell lines MCF7 and T47D, and the ER-negative human IDC cell line MDA-MB-231 (MM231) under the same conditions relative to their 2D g rowth[25]
We showed that the ER-negative, HER2-positive human breast cancer cell line SKBR332 exhibits limited ULA growth (Supplementary Fig. S1c) and that seeding the ILC and IDC cell lines at lower or higher starting numbers yields similar results (Supplementary Fig. S1d,e)

Summary

Introduction

Invasive lobular carcinoma (ILC) is a histological subtype of breast cancer with distinct molecular and clinical features from the more common subtype invasive ductal carcinoma (IDC). ILC tumors exhibit single-file growth invading the surrounding stroma in a diffuse, linear pattern[2] This unusual feature is largely attributed to the hallmark genetic loss of CDH1, which encodes the adherens junction protein E-cadherin[3,4,5]. Loss of other adherens junction proteins such as p120catenin (p120) drive the survival of mouse ILC cell lines and primary metastatic human ILC cells through ROCK1-mediated anoikis resistance[22,23,24]. To complement these findings, we need additional studies of anchorage-independence in ER-positive human ILC cell lines assessing the potential roles of anoikis resistance and cell proliferation

Methods

Results

Conclusion