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Immortalization of human primary prostate epithelial cells via CRISPR inactivation of the CDKN2A locus and expression of telomerase
Prostate Cancer Prostatic Dis. 2021 Mar;24(1) :233-243
PMID: 32873916 PMCID: PMC7917161 URL: https://www.ncbi.nlm.nih.gov/pubmed/32873916
AbstractBACKGROUND: Immortalization of primary prostate epithelial cells (PrEC) with just hTERT expression is particularly inefficient in the absence of DNA tumor viral proteins or p16(INK4A) 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.
Notes1476-5608 Zhao, Ziran Fowle, Holly Valentine, Henkel Liu, Zemin Tan, Yinfei Pei, Jianming Badal, Simone Orcid: 0000-0001-5214-3896 Testa, Joseph R Graña, Xavier Orcid: 0000-0001-7134-0473 R03 CA216134-01/U.S. Department of Health & Human Services | NIH | National Cancer Institute (NCI)/ P30 CA006927/CA/NCI NIH HHS/United States U54 CA22170/U.S. Department of Health & Human Services | NIH | National Cancer Institute (NCI)/ R01 GM117437/GM/NIGMS NIH HHS/United States U54 CA221704/CA/NCI NIH HHS/United States Journal Article England Prostate Cancer Prostatic Dis. 2020 Sep 1. doi: 10.1038/s41391-020-00274-4.