NOTCH signaling is activated in and contributes to resistance in enzalutamide-resistant prostate cancer cells

Elia Farah,Chaohao Li,Lijun Cheng,Yifan Kong,Nadia A Lanman,Pete Pascuzzi,Gabrielle Renee Lorenz,Yanquan Zhang,Nihal Ahmad,Lang Li,Tim Ratliff,Xiaoqi Liu

doi:10.1074/jbc.ra118.006983

Abstract

Prostate cancer is the second leading cause of cancer death among men in the United States. The androgen receptor (AR) antagonist enzalutamide is a Food and Drug Administration-approved drug for treatment of patients with late-stage prostate cancer and is currently under clinical study for early-stage prostate cancer treatment. After a short positive response period, tumors will develop drug resistance. In this study using RNA-Seq and bioinformatics analyses, we observed that NOTCH signaling is a deregulated pathway in enzalutamide-resistant cells. NOTCH2 and c-MYC gene expression positively correlated with AR expression in samples from patient with hormone refractory disease in which AR expression levels correspond to those typically observed in enzalutamide resistance. Cleaved NOTCH1, HES1 (Hes family BHLH transcription factor 1), and c-MYC protein expression levels are elevated in two enzalutamide-resistant cell lines, MR49F and C4-2R, indicating NOTCH signaling activation. Moreover, inhibition of the overexpressed ADAM metallopeptidase domain 10 (ADAM10) in the resistant cells induces an exclusive reduction in cleaved NOTCH1 expression. Furthermore, exposure of enzalutamide-resistant cells to both PF-03084014 and enzalutamide increased cell death, decreased colony formation ability, and resensitized cells to enzalutamide. Knockdown of NOTCH1 in C4-2R increased enzalutamide sensitivity by decreasing cell proliferation and increasing cleaved PARP expression. In a 22RV1 xenograft model, PF-03084014 and enzalutamide decreased tumor growth through reducing cell proliferation and increasing apoptosis. These results indicate that NOTCH1 signaling may contribute to enzalutamide resistance in prostate cancer, and inhibition of NOTCH signaling can resensitize resistant cells to enzalutamide.

Highlights

Prostate cancer is the second leading cause of cancer death among men in the United States
These cell lines were developed by extensive treatment with high concentrations of enzalutamide in a tissue culture setting or in an in vivo xenograft mouse model to serve as a model to study enzalutamide resistance [5, 6]
To compare enzalutamide-sensitive and enzalutamide-resistant cell lines, RNA was isolated from LNCaP, C4-2, MR49F, and C4-2R after a 4-h treatment with enzalutamide

Summary

Introduction

Prostate cancer is the second leading cause of cancer death among men in the United States. In a 22RV1 xenograft model, PF-03084014 and enzalutamide decreased tumor growth through reducing cell proliferation and increasing apoptosis These results indicate that NOTCH1 signaling may contribute to enzalutamide resistance in prostate cancer, and inhibition of NOTCH signaling can resensitize resistant cells to enzalutamide. Despite the high survival rate of prostate cancer patients, most develop resistance to therapies, leading to a more aggressive form of the disease and death. We hypothesized that genes and pathways outside of the AR signaling pathway contribute to enzalutamide resistance in prostate cancer cells. Our results show that the Notch signaling pathway is linked to enzalutamide resistance Abrogation of this pathway in vitro and in vivo restores sensitivity of prostate cancer cells to enzalutamide. The data discussed in this publication have been deposited in NCBI’s Gene Expression Omnibus and are accessible through GEO Series accession number GSE123379

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