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Lestaurtinib inhibits histone phosphorylation and androgen-dependent gene expression in prostate cancer cells.

ABSTRACT: BACKGROUND: Epigenetics is defined as heritable changes in gene expression that are not based on changes in the DNA sequence. Posttranslational modification of histone proteins is a major mechanism of epigenetic regulation. The kinase PRK1 (protein kinase C related kinase 1, also known as PKN1) phosphorylates histone H3 at threonine 11 and is involved in the regulation of androgen receptor signalling. Thus, it has been identified as a novel drug target but little is known about PRK1 inhibitors and consequences of its inhibition. METHODOLOGY/PRINCIPAL FINDING: Using a focused library screening approach, we identified the clinical candidate lestaurtinib (also known as CEP-701) as a new inhibitor of PRK1. Based on a generated 3D model of the PRK1 kinase using the homolog PKC-theta (protein kinase c theta) protein as a template, the key interaction of lestaurtinib with PRK1 was analyzed by means of molecular docking studies. Furthermore, the effects on histone H3 threonine phosphorylation and androgen-dependent gene expression was evaluated in prostate cancer cells. CONCLUSIONS/SIGNIFICANCE: Lestaurtinib inhibits PRK1 very potently in vitro and in vivo. Applied to cell culture it inhibits histone H3 threonine phosphorylation and androgen-dependent gene expression, a feature that has not been known yet. Thus our findings have implication both for understanding of the clinical activity of lestaurtinib as well as for future PRK1 inhibitors.

SUBMITTER: Kohler J

PROVIDER: S-EPMC3332061 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Lestaurtinib inhibits histone phosphorylation and androgen-dependent gene expression in prostate cancer cells.

Köhler Jens J Erlenkamp German G Eberlin Adrien A Rumpf Tobias T Slynko Inna I Metzger Eric E Schüle Roland R Sippl Wolfgang W Jung Manfred M

PloS one 20120420 4

<h4>Background</h4>Epigenetics is defined as heritable changes in gene expression that are not based on changes in the DNA sequence. Posttranslational modification of histone proteins is a major mechanism of epigenetic regulation. The kinase PRK1 (protein kinase C related kinase 1, also known as PKN1) phosphorylates histone H3 at threonine 11 and is involved in the regulation of androgen receptor signalling. Thus, it has been identified as a novel drug target but little is known about PRK1 inhib ...[more]

PMID: 22532837

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Project description:BackgroundClooxygenase-2 (COX-2) expression is overexpressed in human prostate cancer, and aberrant methylation of the COX-2 promoter has also been elucidated. However, how the methylation of CpG islands at COX-2 regulates its expression in prostate cancer is still unclear. We will determine the methylated 5' CpG island of the COX-2 gene and its role in the expression of COX-2 in prostate androgen-dependent and androgen-independent cancer cells, LNCaP and DU145.MethodsWe used western blotting and quantitative reverse transcription polymerase chain reaction (qRT-PCR) to confirm the COX-2 expression in prostate cancer cell lines, including LNCaP (androgen-dependent) and DU145 (androgen-independent) cells. To investigate whether the COX-2 expression was regulated by the methylation status of the 5' CpG island, we treated LNCaP and DU145 cells with the DNA methylation inhibitor, 5-aza-2'-deoxycytidine, and determined COX-2 expression in the treated/untreated cells by western blotting and qRT-PCR. Subsequently, bisulfite sequencing was performed to study the methylation sites in the treated/untreated cells. The effects of 5-aza-2'-deoxycytidine to cell proliferation, cell migration and cell cycle process in DU145 and LNCaP cells were determined using Cell Counting Kit-8 (CCK-8) assay, transwell assay and flow cytometry, respectively.ResultsThe results revealed that the expression of COX-2 in androgen-dependent LNCaP cells was 5.44-fold (in protein level) and 2.46-fold (in mRNA level) higher than that in androgen-independent DU145 cells. After 5-aza-2'-deoxycytidine treatment, COX-2 expression in DU145 cells was elevated significantly, but no change was found in LNCaP cells. The A and C regions of the COX-2 CpG island exhibited reduced methylation along with that an increased expression of COX-2 was noted in DU145 cell after 5-aza-2'-deoxycytidine treatment. Also, the treatment with 5-aza-2'-deoxycytidine inhibited cell proliferation, cell migration and influenced the cell cycle progression in both DU145 and LNCaP cells.ConclusionsOur results reveal that androgen receptor (AR)-dependent/independent prostate cancer cell lines exhibit different regulation of methylation in COX-2 that regulate its expression. Additionally, 5-aza-2'-deoxycytidine treatment of DU145 and LNCaP cells inhibits their ability of tumor progression.

Project description:BackgroundWith almost 30,000 deaths per year, prostate cancer is the second-leading cause of cancer-related death in men. Androgen Deprivation Therapy (ADT) has been the corner stone of prostate cancer treatment for decades. However, despite an initial response of prostate cancer to ADT, this eventually fails and the tumors recur, resulting in Castration Resistant Prostate Cancer (CRPC). Triptolide, a diterpene triepoxide, has been tested for its anti-tumor properties in a number of cancers for over a decade. Owing to its poor solubility in aqueous medium, its clinical application had been limited. To circumvent this problem, we have synthesized a water-soluble pro-drug of triptolide, Minnelide, that is currently being evaluated in a Phase 1 clinical trial against gastrointestinal tumors. In the current study, we assessed the therapeutic potential of Minnelide and its active compound triptolide against androgen dependent prostate cancer both in vitro as well as in vivo.MethodsCell viability was measured by a MTT based assay after treating prostate cancer cells with multiple doses of triptolide. Apoptotic cell death was measured using a caspase 3/7 activity. Androgen Receptor (AR) promoter-binding activity was evaluated by using luciferase reporter assay. For evaluating the effect in vivo, 22Rv1 cells were implanted subcutaneously in animals, following which, treatment was started with 0.21 mg/kg Minnelide.ResultsOur study showed that treatment with triptolide induced apoptotic cell death in CRPC cells. Triptolide treatment inhibited AR transcriptional activity and decreased the expression of AR and its splice variants both at the mRNA and the protein level. Our studies show that triptolide inhibits nuclear translocation of Sp1, resulting in its decreased transcriptional activity leading to downregulation of AR and its splice variants in prostate cancer cells. In vivo, Minnelide (0.21 mg/kg) regressed subcutaneous tumors derived from CRPC 22RV1 at our study endpoint. Our animal studies further confirmed that Minnelide was more efficacious than the standard of care therapies, Docetaxel and Enzalutamide.ConclusionOur study indicates that Minnelide is very effective as a therapeutic option against CRPC at a dose that is currently tolerated by patients in the ongoing clinical trials. Prostate 77: 584-596, 2017. © 2017 Wiley Periodicals, Inc.

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Lestaurtinib inhibits histone phosphorylation and androgen-dependent gene expression in prostate cancer cells.

Publications

Lestaurtinib inhibits histone phosphorylation and androgen-dependent gene expression in prostate cancer cells.

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