Immortalization of human primary prostate epithelial cells via CRISPR inactivation of the CDKN2A locus and expression of telomerase

Ziran Zhao, Holly Fowle, Henkel Valentine, Zemin Liu, Yinfei Tan, Jianming Pei, Simone Badal, Joseph R. Testa, Xavier Grana

Research output: Contribution to journalArticlepeer-review

10 Scopus citations

Abstract

Background: Immortalization of primary prostate epithelial cells (PrEC) with just hTERT expression is particularly inefficient in the absence of DNA tumor viral proteins or p16INK4A knockdown. Materials and methods: Here, we describe the establishment of immortalized normal prostate epithelial cell line models using CRISPR technology to inactivate the CDKN2A locus concomitantly with ectopic expression of an hTERT transgene. Results: Using this approach, we have obtained immortal cell clones that exhibit fundamental characteristics of normal cells, including diploid genomes, near normal karyotypes, normal p53 and pRB cell responses, the ability to form non-invasive spheroids, and a non-transformed phenotype. Based on marker expression, these clones are of basal cell origin. Conclusions: Use of this approach resulted in the immortalization of independent clones of PrEC that retained normal characteristics, were stable, and non-transformed. Thus, this approach could be used for the immortalization of normal primary prostate cells. This technique could also be useful for establishing cell lines from prostate tumor tissues of different tumor grades and/or from patients of diverse ethnicities to generate cell line models that facilitate the study of the molecular basis of disease disparity.

Original languageEnglish
Pages (from-to)233-243
Number of pages11
JournalProstate Cancer and Prostatic Diseases
Volume24
Issue number1
DOIs
StatePublished - Mar 2021

Keywords

  • Cell Line, Tumor
  • Clustered Regularly Interspaced Short Palindromic Repeats
  • Cyclin-Dependent Kinase Inhibitor p16/biosynthesis
  • Gene Expression Regulation, Neoplastic
  • Humans
  • Male
  • Prostatic Neoplasms/genetics
  • RNA, Neoplasm/genetics
  • Telomerase/biosynthesis

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