Enzymatic activity of free‐prostate‐specific antigen (f‐PSA) is not required for some of its physiological activities

Abstract

BACKGROUND Prostate specific antigen (PSA) is a well known biomarker for early diagnosis and management of prostate cancer. Furthermore, PSA has been documented to have anti-angiogenic and anti-tumorigenic activities in both in vitro and in vivo studies. However, little is known about the molecular mechanism(s) involved in regulation of these processes, in particular the role of the serine-protease enzymatic activity of PSA. METHODS Enzymatic activity of PSA isolated directly from seminal plasma was inhibited specifically (>95%) by incubation with zinc2+. Human umbilical vein endothelial cells (HUVEC) were utilized to compare/contrast the physiological effects of enzymatically active versus inactive PSA. RESULTS Equimolar concentrations of enzymatically active PSA and PSA enzymatically inactivated by incubation with Zn2+ had similar physiological effects on HUVEC, including inhibiting the gene expression of pro-angiogenic growth factors, like VEGF and bFGF, and up-regulation of expression of the anti-angiogenic growth factor IFN-γ; suppression of mRNA expression for markers of blood vessel development, like FAK, FLT, KDR, TWIST-1; P-38; inhibition of endothelial tube formation in the in vitro Matrigel Tube Formation Assay; and inhibition of endothelial cell invasion and migration properties. DISCUSSION Our data provides compelling evidence that the transcriptional regulatory and the anti-angiogenic activities of human PSA are independent of the innate enzymatic activity

Anti-angiogenic activity of PSA-derived peptides

“…Our evidence that the anti-angiogenic activity of PSA was maintained in f-PSA protein in which the enzymatic active site was inactivated by incubation with Zn 2þ [15] is in contrast to reported data where the anti-angiogenic activity was not preserved in enzymatically inactive internally cleaved PSAisoforms [27]. This study suggests that rather than through enzymatic activity, the anti-tumoricidal activity of PSA may be mediated through the interaction of f-PSA with a cell surface protein(s) on prostate cancer cells and/or prostate endothelial cells.…”

“…The loss of anti‐angiogenic activity of enzymatically active PSA when exposed to anti‐PSA antibody may be due to loss of accessibility of the “active site” on PSA‐antibody complex required for its anti‐angiogenic activity. In our previous studies, we have shown that enzymatically “active” PSA and enzymatically “inactive” PSA, where enzymatic function was inhibited by Zinc, have comparable anti‐angiogenic activity , suggesting that enzymatic function of PSA is not required for its anti‐angiogenic activity. In addition, recombinant PSA mutated to ablate enzymatic activity for chromogenic PSA‐specific substrate has been reported to have significant anti‐angiogenic activity .…”

“…In addition, both enzymatically active and enzymatically inactive f‐PSA comparably modulated gene expression in human umbilical vein endothelial cells (HUVEC) of multiple growth factors, including suppression of expression of bFGF and VEGF, and up‐regulation of IFN, and of proteins that have a direct role in blood vessel development, including FAK, FLT, KDR, TWIST‐1, P‐38, and Cathepsin‐D. Furthermore, maintenance of PSA enzymatic activity was not required for inhibition of endothelial cell tube formation in the in vitro Matrigel Tube Formation Assay or for inhibition of endothelial cell invasion and migration by HUVEC cells . Therefore, our data suggests strongly that PSA has a significant physiological role in prostate tissue, potentially affecting both prostate epithelial cells/prostate cancer cells and prostate endothelial cells, which is independent of the serine protease enzymatic activity of PSA.…”

“…Consequently, the anti-tumorigenic activity of PSA, as for the degradation of semenogelins, was proposed to be related to the serine protease enzymatic activity of PSA in that denatured PSA did not demonstrate biological activity in these assays [13]. However, our group demonstrated in CaP cells (PC-3M) that both purified, enzymatically active PSA (free-PSA: f-PSA) and enzymatically inactive PSA, where enzymatic activity was inactivated by incubation with Zinc 2þ [14], equivalently down-regulated expression of pro-angiogenic factors/cancer-related genes/proteins, including VEGF, EphA2, Bcl2, Pim-1 oncogene, CYR61 and uPA, and up-regulated expression of anti-angiogenic genes/proteins, including interferon (IFN) and IFN-related genes [15]. In addition, both enzymatically active and enzymatically inactive f-PSA comparably modulated gene expression in human umbilical vein endothelial cells (HUVEC) of multiple growth factors, including suppression of expression of bFGF and VEGF, and up-regulation of IFN, and of proteins that have a direct role in blood vessel development, including FAK, FLT, KDR, TWIST-1, P-38, and Cathepsin-D.…”

Detection of Prostate Cancer Metastasis by Whole Body Magnetic Resonance Imaging Combined with Bone Scintigraphy and PSA Levels

“…The high serum PSA level in PCa patients has also been revealed to signal bone metastasis and PCa recurrence among patients who have received localized disease treatments, yet our study was confined for not exploring this [43][44][45]. Besides, scholars have provided a notion that detecting elements relative to dynamic PSA changes, including baseline PSA, PSA nadir, extent of PSA declining, TTPN and PSA doubling time, was able to improve the diagnostic accuracy of PCa [46]. Meanwhile, the PSA-relevant factors were also associated with PCa prognosis (Table 3).…”