Homozygous deletion 8p21 in prostate cancer

Research background and aimsThe aim of this research project was to investigate the effect of the chromosome 8p21 homozygous deletion of genes BNIP3L and PPP2R2A on the phenotype of the prostate cancer cell model called VCaP. The chromosome 8p21 deletion has been known for a long time, but the contribution of these deletion genes has not been verified in prostate cancer before. VCaP is known to be difficult to transfect; hence the first aim was to find a functional transfection method. MethodsDifferent transient transfection methods were studied to optimize the VCaP transfection protocol and to be able to transfect the deletion genes back into VCaP. Accordingly, plasmid reconstruction was prepared for virus transduction and the usability of two cell proliferation analysis methods were compared in VCaP. ResultsLiposome-mediated transfection methods provided only modest results, not sufficient transfection efficiency for subsequent functional analysis. VCaP cells showed atypical morphology and severely suffered after transfection in many cases. Reverse transfection with polyethylenimine (PEI25) was found to be cytotoxic with all studied variables. Viral transduction preparations including vector ligation were inefficient, showing no positive clones of the correct vector size. Thus, proceeding to more efficient viral infection was not possible. ConclusionsProstate cancer cell model VCaP was shown to be difficult to transfect. Adherent cells suffered in all studied transfection protocols. The best alternative for transfecting VCaP was shown to be a commercial liposome-mediated forward transfection reagent. VCaP cell line needs thorough optimization for each experiment and further studies to address the function of chromosome 8p21 deletion genes. Asiasanat:prostate cance

Homozygous deletion 8p21 in prostate cancer

Research background and aimsThe aim of this research project was to investigate the effect of the chromosome 8p21 homozygous deletion of genes BNIP3L and PPP2R2A on the phenotype of the prostate cancer cell model called VCaP. The chromosome 8p21 deletion has been known for a long time, but the contribution of these deletion genes has not been verified in prostate cancer before. VCaP is known to be difficult to transfect; hence the first aim was to find a functional transfection method. MethodsDifferent transient transfection methods were studied to optimize the VCaP transfection protocol and to be able to transfect the deletion genes back into VCaP. Accordingly, plasmid reconstruction was prepared for virus transduction and the usability of two cell proliferation analysis methods were compared in VCaP. ResultsLiposome-mediated transfection methods provided only modest results, not sufficient transfection efficiency for subsequent functional analysis. VCaP cells showed atypical morphology and severely suffered after transfection in many cases. Reverse transfection with polyethylenimine (PEI25) was found to be cytotoxic with all studied variables. Viral transduction preparations including vector ligation were inefficient, showing no positive clones of the correct vector size. Thus, proceeding to more efficient viral infection was not possible. ConclusionsProstate cancer cell model VCaP was shown to be difficult to transfect. Adherent cells suffered in all studied transfection protocols. The best alternative for transfecting VCaP was shown to be a commercial liposome-mediated forward transfection reagent. VCaP cell line needs thorough optimization for each experiment and further studies to address the function of chromosome 8p21 deletion genes. Asiasanat:prostate cance

CIP2A is a candidate therapeutic target in clinically challenging prostate cancer cell populations

Residual androgen receptor (AR)-signaling and presence of cancer stem-like cells (SCs) are the two emerging paradigms for clinically challenging castration-resistant prostate cancer (CRPC). Therefore, identification of AR-target proteins that are also overexpressed in the cancer SC population would be an attractive therapeutic approach. Our analysis of over three hundred clinical samples and patient-derived prostate epithelial cultures (PPECs), revealed Cancerous inhibitor of protein phosphatase 2A (CIP2A) as one such target. CIP2A is significantly overexpressed in both hormone-naïve prostate cancer (HN-PC) and CRPC patients . CIP2A is also overexpressed, by 3- and 30-fold, in HN-PC and CRPC SCs respectively. In vivo binding of the AR to the intronic region of CIP2A and its functionality in the AR-moderate and AR-high expressing LNCaP cell-model systems is also demonstrated. Further, we show that AR positively regulates CIP2A expression, both at the mRNA and protein level. Finally, CIP2A depletion reduced cell viability and colony forming efficiency of AR-independent PPECs as well as AR-responsive LNCaP cells, in which anchorage-independent growth is also impaired. These findings identify CIP2A as a common denominator for AR-signaling and cancer SC functionality, highlighting its potential therapeutic significance in the most clinically challenging prostate pathology: castration-resistant prostate cancer