Preparation and characterization of prostate cell lines for functional cloning studies to identify regulators of apoptosis

Abstract

Because apoptotic evasion is a central feature of prostate cancer, there is an urgent need for increased understanding of the key regulatory molecules that control the life/death decision of prostate cells. Functional expression cloning permits the isolation of genes that control the rate-limiting steps of cell death and offers a possible solution to this problem. This technique requires the availability of prostate cells that meet several stringent requirements. Therefore, the main objective was to obtain prostate cell clones that undergo cell death with minimal survival of spontaneously resistant cells and that can be infected at a high efficiency with viral vectors. Initial characterization of 5 prostate cell lines with a range of apoptotic inducers revealed cell line - dependent and treatment-dependent effects. In general, the colony-forming ability of non-tumorigenic PNT2C2 cells showed the highest sensitivity to most chemical agents and ultraviolet (UV) irradiation, whereas the metastases-derived cell lines, LNCaP and PC-3, showed resistance to UV and etoposide, respectively. Clones of PNT2C2, 22Rv1, and PC-3 were produced, which displayed heterogeneous responses to UV irradiation. Further characterization of UV-sensitive clones revealed at least 1 clone per cell line with high sensitivity (mean clonogenic survival ≤0.02% control cells) to 3 or more apoptotic inducers. These clones could be infected at a high efficiency (>90%) with a lentiviral vector. In conclusion, we have isolated clones of nontumorigenic prostate cells (PNT2C2), androgen-sensitive prostate cancer cells (22Rv1), and androgen-independent, metastatic prostate cancer cells (PC-3), which are suitable as host cells for functional cloning studies to address cell death control mechanisms in the prostate during cancer progression. Copyright © American Society of Andrology.