The central role of Sphingosine kinase 1 in the development of neuroendocrine prostate cancer (NEPC): A new targeted therapy of NEPC.

BACKGROUND: Neuroendocrine prostate cancer (NEPC) is often diagnosed as a sub-type from the castration-resistant prostate cancer (CRPC) recurred from the second generation of anti-androgen treatment and is a rapidly progressive fatal disease. The molecular mechanisms underlying the trans-differentiation from CRPC to NEPC are not fully characterized, which hampers the development of effective targeted therapy. METHODS: Bioinformatic analyses were conducted to determine the clinical correlation of sphingosine kinase 1 (SphK1) in CRPC progression. To investigate the transcriptional regulation SphK1 and neuroendocrine (NE) transcription factor genes, both chromosome immunoprecipitation and luciferase reporter gene assays were performed. To demonstrate the role of SphK1 in NEPC development, neurosphere assay was carried out along with several biomarkers determined by quantitative PCR and western blot. Furthermore, in vivo NEPC xenograft models and patient-derived xenograft (PDX) model were employed to determine the effect of SphK1 inhibitors and target validation. RESULTS: Significant prevalence of SphK1 in NEPC development is observed from clinical datasets. SphK1 is transcriptionally repressed by androgen receptor-RE1-silencing transcription factor (REST) complex. Furthermore, sphingosine 1-phosphate produced by SphK1 can modulate REST protein turnover via MAPK signaling pathway. Also, decreased REST protein levels enhance the expression of NE markers in CRPC, enabling the transition to NEPC. Finally, specific SphK1 inhibitors can effectively inhibit the growth of NEPC tumors and block the REST protein degradation in PDX. CONCLUSIONS: SphK1 plays a central role in NEPC development, which offers a new target for this lethal cancer using clinically approved SphK1 inhibitors.

Adenovirus-mediated Drosophila melanogaster deoxyribonucleoside kinase mutants combined with gemcitabine harbor a safe cancer treatment profile.

The purpose of this analysis was to investigate the enzyme activity and specificity of adenovirus-mediated Drosophila melanogaster deoxyribonucleoside kinase (Dm-dNK) mutants in combination with gemcitabine. Compared to herpes simplex type 1 thymidine kinases (HSV-TK) and other known dNKs, this Dm-dNK enzyme has a broader substrate specificity and a higher catalytic rate. We created the Dm-dNK mutants (dNKmut) by site-directed mutagenesis at the sites of 244E, 245S, 251S, and 252R, with the last 10 amino acids in the amino acid sequence randomly alternated. We subsequently evaluated the enzyme activity and substrate specificity. The engineered enzymes showed a relative increase in phosphorylation in the nucleoside analogs of gemcitabine (dFdC, 2',2'-difluoro-deoxycytidine) compared to the wild-type enzyme. The dNKmut enzymes were expressed in breast (Bcap37) and gastric (SGC-7901) cancer cell lines. In studying the sensitivity of the cell lines to dFdC, conditionally replicative adenovirus (CRAd) ZD55-dNKmut showed higher expression and enzymatic activity than the replication-defective adenovirus Ad-dNKmut in cancer cells, but with less cytotoxicity to cancer cells than that of Ad-dNKmut. Our data suggest that the triple phosphorylated dFdC catalyzed by dNKmut inhibited the replication of adenovirus with a simultaneous positive therapeutic effect on cancer cells. Therefore, concomitant use of the ZD55-dNKmut and dFdC could be a novel targeted strategy in suicide gene therapy with safe control of excessive virus replication.