# Identification of Novel Markers of Prostate Cancer Progression, Potentially Modulated by Vitamin D
## 4 Discussion  
In this paper, we undertook an analysis to identify the key genes involved in PCa. We analyzed three PCa microarray datasets (GSE70770, GSE62872 and GSE6919), merged all the genes significantly up-regulated and subsequently, intersected the common genes identified (n = 276) with the transcriptome of genes down-regulated in the _LNCaP cell line_ treated with Vitamin D, in order to identify the common key genes involved in PCa and potentially modulated by the action of Vitamin D. The analysis yielded 43 genes involved in PCa significantly modulated by Vitamin D. These genes were involved in different molecular processes, among which the most interesting were _organogenesis, morphogenesis and cell migration_, all mechanisms involved in the genesis and in the tumor progression. Noteworthy, our analysis showed that six genes (PRSS8, SOX4, SMYD2, MCCC2, CCNG2 and CD2 AP) were significantly modulated in both PCa biopsies and in the _LNCaP cell line_ treated with Vitamin D. A literature investigation showed that all these genes were involved in different tumor progressions. Furthermore, a Z-score transformation of all datasets allowed us to group all the healthy prostate samples (n = 251) and PCa (n = 548), thus significantly increasing the court of subjects included in the study. A Spearman's \(\varrho\) correlation analysis showed that five genes out of six (PRSS8, SMYD2, MCCC2, CCNG2 and CD2 AP), were positively and statistically correlated with the gene expression levels of KLK3 and with the tumor %. Contrary note, SOX4 expression levels were positively correlated only with the tumor %, but not with KLK3 gene expression levels. Furthermore, the analysis of the data deriving from GSE107438, the treatment of LNCaPP cell line with siCYP27B1 and siVDR, showed that a downregulation of KLK3 expression levels depends strictly on the CYP27B1 gene transcription. More mildly, the PRSS8 and SMYD2 genes expression also depends on the transcription of the CYP27B1 gene. Co-silencing of the CYP27B1 and VDR genes was determinant in the transcription of the CCNG2, PRSS8, SMYD2 and SOX4 genes. Our results have been in part confirmed by the immunostaining obtained by the Human Atlas Project portal. The immunohistochemistry analysis showed that _five_ genes (PRSS8, SOX4, MCCC2, CCNG2 and CD2 AP) out of six appeared more expressed in PCa biopsies compared to healthy prostate tissue. No difference was observed for SYMD2. Prostasin (PRSS8) is a serine protease, mainly expressed in normal prostate epithelia, the prostate gland and in seminal fluid [37]. This protein is implicated in epithelial sodium channel regulation and essential for terminal epithelial differentiation [38]. Nowadays, the role played by this molecule in cancer is unclear. It was shown that it may be related to tumor promoter mechanisms [39] as well as tumor suppression [40; 41; 42]. In several studies it has been shown that PRSS8 plays a role as a tumor suppressor in various malignancies, such as in hepatocellular carcinoma [43]. In 2016 [PERSON] et al. showed that abundant amounts of secreted prostasin found in the sera of early stage ovarian cancer (OVC) can potentially be used as a minimally invasive screening biomarker for early stage OVC [44]. As regarding the role of PRSS8 and PCa, there is no large bibliography. Noteworthy, in 2003, [PERSON] and colleagues demonstrated that in 54 patients (Japanese), the PRSS8 mRNA expression levels, measured by northern blot analysis, were not correlated with the clinical stage of human PCa. Furthermore, almost in all cases of metastatic and hormone-refractory cancers, these demonstrated a down-regulation of _prostasin_ expression. The authors at the end of their manuscript suggested that _prostasin_ cannot be regarded as a prognostic indicator for human PCa, although it may be a useful marker for tumor differentiation [45]. In light of this, our results are totally contrasting with the results obtained by [PERSON] in 2003.