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Nelfinavir and nelfinavir analogs block site-2 protease cleavage to inhibit castration-resistant prostate cancer.

Guan M, Su L, Yuan YC, Li H, Chow WA - Sci Rep (2015)

Bottom Line: Nelfinavir and its analogs are more potent inhibitors of S2P cleavage activity than 1,10-phenanthroline, a metalloprotease-specific inhibitor.Further, cluster analysis of gene expression from treated DU145 and PC3 cell lines demonstrate a close similarity of nelfinavir, its analogs, and 1,10-phenanthroline.This leads to accumulation of precursor SREBP-1 and ATF6, and development of insufficient reserves of their transcriptionally-active forms.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Pharmacology, Beckman Research Institute of the City of Hope, Duarte, CA, USA.

ABSTRACT
Nelfinavir and its analogs inhibit proliferation and induce apoptosis of castration-resistant prostate cancer through inhibition of site-2 protease (S2P) activity, which leads to suppression of regulated intramembrane proteolysis. Western blotting in nelfinavir and its analog treated cells confirms accumulation of precursor SREBP-1 and ATF6. Nelfinavir and its analogs inhibit human homolog M. jannaschii S2P cleavage of an artificial protein substrate CED-9 in an in vitro proteolysis assay in a dose-dependent manner. Nelfinavir and its analogs are more potent inhibitors of S2P cleavage activity than 1,10-phenanthroline, a metalloprotease-specific inhibitor. Further, cluster analysis of gene expression from treated DU145 and PC3 cell lines demonstrate a close similarity of nelfinavir, its analogs, and 1,10-phenanthroline. These results show nelfinavir and its analogs inhibit castration-resistant prostate cancer proliferation by blocking regulated intramembrane proteolysis through suppression of S2P cleavage activity. This leads to accumulation of precursor SREBP-1 and ATF6, and development of insufficient reserves of their transcriptionally-active forms. The present results validate S2P and regulated intramembrane proteolysis as novel therapeutic targets for castration-resistant prostate cancer therapeutics. A clinical trial of nelfinavir or its analogs should be developed for castration-resistant prostate cancer.

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Nelfinavir analogs inhibit proliferation and induce apoptosis in CRPC.(A) Nelfinavir analogs inhibit CRPC proliferation. DIMSCAN assay was performed in nelfinavir- or analog #6, 7, 8, 31, 39-treated DU145 and PC-3 cells. 1000 cells were seeded in a 96-well plate for overnight incubation followed by treatment with DMSO, nelfinavir or its analogs (10 μM) for 3 days. Each experiment was performed in triplicate. Results are presented as mean ± SD. *p < 0.01, +p < 0.05 compared to DMSO control. (B) Nelfinavir analogs induce CRPC apoptosis. DU145 and PC-3 cells were treated with DMSO or 10 μM of nelfinavir or analog #6, 7, 8, 31, 39. After 24 hr, cells were harvested and stained with Annexin V-FITC for detection of apoptosis. Image represents three repeated experiments.
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f2: Nelfinavir analogs inhibit proliferation and induce apoptosis in CRPC.(A) Nelfinavir analogs inhibit CRPC proliferation. DIMSCAN assay was performed in nelfinavir- or analog #6, 7, 8, 31, 39-treated DU145 and PC-3 cells. 1000 cells were seeded in a 96-well plate for overnight incubation followed by treatment with DMSO, nelfinavir or its analogs (10 μM) for 3 days. Each experiment was performed in triplicate. Results are presented as mean ± SD. *p < 0.01, +p < 0.05 compared to DMSO control. (B) Nelfinavir analogs induce CRPC apoptosis. DU145 and PC-3 cells were treated with DMSO or 10 μM of nelfinavir or analog #6, 7, 8, 31, 39. After 24 hr, cells were harvested and stained with Annexin V-FITC for detection of apoptosis. Image represents three repeated experiments.

Mentions: DU145 and PC-3 cells were treated with 10 μM NFV and the available analogs for 72 hr prior to assaying for proliferation using the DIMSCAN system. As shown in Figure 2A, proliferation was significantly reduced in analogs #6, #7 and #8 treated CRPC cells comparing to nelfinavir treated cells. Only moderate effects were observed in CRPC cells treated with analogs #31 and #39 compared to nelfinavir. Results for analogs less potent than nelfinavir are not shown. Moreover, increased apoptosis was detected in CRPC cells by FACS detection of FITC-annexin V treated with analogs #6, #7 and #8 compared to nelfinavir-treated CRPC cells (Figure 2B). Both the proliferation and apoptosis assays suggest analogs #6, #7 and #8 are more potent than nelfinavir at equimolar concentrations.


Nelfinavir and nelfinavir analogs block site-2 protease cleavage to inhibit castration-resistant prostate cancer.

Guan M, Su L, Yuan YC, Li H, Chow WA - Sci Rep (2015)

Nelfinavir analogs inhibit proliferation and induce apoptosis in CRPC.(A) Nelfinavir analogs inhibit CRPC proliferation. DIMSCAN assay was performed in nelfinavir- or analog #6, 7, 8, 31, 39-treated DU145 and PC-3 cells. 1000 cells were seeded in a 96-well plate for overnight incubation followed by treatment with DMSO, nelfinavir or its analogs (10 μM) for 3 days. Each experiment was performed in triplicate. Results are presented as mean ± SD. *p < 0.01, +p < 0.05 compared to DMSO control. (B) Nelfinavir analogs induce CRPC apoptosis. DU145 and PC-3 cells were treated with DMSO or 10 μM of nelfinavir or analog #6, 7, 8, 31, 39. After 24 hr, cells were harvested and stained with Annexin V-FITC for detection of apoptosis. Image represents three repeated experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4816264&req=5

f2: Nelfinavir analogs inhibit proliferation and induce apoptosis in CRPC.(A) Nelfinavir analogs inhibit CRPC proliferation. DIMSCAN assay was performed in nelfinavir- or analog #6, 7, 8, 31, 39-treated DU145 and PC-3 cells. 1000 cells were seeded in a 96-well plate for overnight incubation followed by treatment with DMSO, nelfinavir or its analogs (10 μM) for 3 days. Each experiment was performed in triplicate. Results are presented as mean ± SD. *p < 0.01, +p < 0.05 compared to DMSO control. (B) Nelfinavir analogs induce CRPC apoptosis. DU145 and PC-3 cells were treated with DMSO or 10 μM of nelfinavir or analog #6, 7, 8, 31, 39. After 24 hr, cells were harvested and stained with Annexin V-FITC for detection of apoptosis. Image represents three repeated experiments.
Mentions: DU145 and PC-3 cells were treated with 10 μM NFV and the available analogs for 72 hr prior to assaying for proliferation using the DIMSCAN system. As shown in Figure 2A, proliferation was significantly reduced in analogs #6, #7 and #8 treated CRPC cells comparing to nelfinavir treated cells. Only moderate effects were observed in CRPC cells treated with analogs #31 and #39 compared to nelfinavir. Results for analogs less potent than nelfinavir are not shown. Moreover, increased apoptosis was detected in CRPC cells by FACS detection of FITC-annexin V treated with analogs #6, #7 and #8 compared to nelfinavir-treated CRPC cells (Figure 2B). Both the proliferation and apoptosis assays suggest analogs #6, #7 and #8 are more potent than nelfinavir at equimolar concentrations.

Bottom Line: Nelfinavir and its analogs are more potent inhibitors of S2P cleavage activity than 1,10-phenanthroline, a metalloprotease-specific inhibitor.Further, cluster analysis of gene expression from treated DU145 and PC3 cell lines demonstrate a close similarity of nelfinavir, its analogs, and 1,10-phenanthroline.This leads to accumulation of precursor SREBP-1 and ATF6, and development of insufficient reserves of their transcriptionally-active forms.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Pharmacology, Beckman Research Institute of the City of Hope, Duarte, CA, USA.

ABSTRACT
Nelfinavir and its analogs inhibit proliferation and induce apoptosis of castration-resistant prostate cancer through inhibition of site-2 protease (S2P) activity, which leads to suppression of regulated intramembrane proteolysis. Western blotting in nelfinavir and its analog treated cells confirms accumulation of precursor SREBP-1 and ATF6. Nelfinavir and its analogs inhibit human homolog M. jannaschii S2P cleavage of an artificial protein substrate CED-9 in an in vitro proteolysis assay in a dose-dependent manner. Nelfinavir and its analogs are more potent inhibitors of S2P cleavage activity than 1,10-phenanthroline, a metalloprotease-specific inhibitor. Further, cluster analysis of gene expression from treated DU145 and PC3 cell lines demonstrate a close similarity of nelfinavir, its analogs, and 1,10-phenanthroline. These results show nelfinavir and its analogs inhibit castration-resistant prostate cancer proliferation by blocking regulated intramembrane proteolysis through suppression of S2P cleavage activity. This leads to accumulation of precursor SREBP-1 and ATF6, and development of insufficient reserves of their transcriptionally-active forms. The present results validate S2P and regulated intramembrane proteolysis as novel therapeutic targets for castration-resistant prostate cancer therapeutics. A clinical trial of nelfinavir or its analogs should be developed for castration-resistant prostate cancer.

Show MeSH
Related in: MedlinePlus