Inhibition of Cyclin-Dependent Kinase 4 (Cdk4) by Fascaplysin, a Marine Natural Product

Soni, Rajeev; Muller, Lionel; Furet, Pascal; Schoepfer, Joseph; Stephan, Christine; Zumstein-Mecker, Sabine; Fretz, Heinz; Chaudhuri, Bhabatosh

doi:10.1006/bbrc.2000.3349

Cited by 160 publications

(121 citation statements)

References 46 publications

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“…Figure 5c demonstrates that BRCA1 630-669 was efficiently phosphorylated by cyclin D1/cdk complexes in vitro, whereas BRCA1 112-141 showed minimal level of phosphorylation (compare lanes 1 and 2). To confirm further the specificity of this phosphorylation, the wild-type BRCA1 peptide was utilized in an in vitro kinase assay, using cyclin D1 immunoprecipitate as well as two other control kinases ( Cyclin D1/cdk4 phosphorylates BRCA1 in vivo To demonstrate that cyclin D1/cdk4 complexes phosphorylate BRCA1 in vivo, a cdk4 inhibitor, Fascaplysin, was utilized (Soni et al, 2000). It has been demonstrated in vitro to have an IC 50 of 0.35 mM against cyclin D1/cdk4.…”

Section: Cyclin D1 and Brca1 Colocalizementioning

confidence: 99%

Functional consequences of cyclin D1/BRCA1 interaction in breast cancer cells

et al. 2007

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The inheritance of one defective BRCA1 or BRCA2 allele predisposes an individual to developing breast and ovarian cancers. BRCA1 is a multifunctional tumor suppressor protein, which through interaction with a vast array of proteins has implications in processes such as cell cycle, transcription, DNA damage response and chromatin remodeling. Conversely, the oncogene, cyclin D1 is overexpressed in about 35% of all breast cancer cases. In this study, we provide detailed analyses on the phosphorylation state of BRCA1 by cyclin D1/cdk4 complexes. In particular, we have identified Ser 632 of BRCA1 as a cyclin D1/cdk4 phosphorylation site in vitro. Using chromatin immunoprecipitation assays, we observed that the inhibition of cyclin D1/cdk4 activity resulted in increased BRCA1 DNA binding at particular promoters in vivo. In addition, we identified multiple novel genes that are bound by BRCA1 in vivo. Collectively, these results indicate that cyclin D1/cdk4-mediated phosphorylation of BRCA1 inhibits the ability of BRCA1 to be recruited to particular promoters in vivo. Therefore, cyclin D1/Cdk4 phosphorylation of BRCA1 could provide a mechanism to interfere with the DNA-dependent activities of BRCA1.

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Section: Cyclin D1 and Brca1 Colocalizementioning

confidence: 99%

Functional consequences of cyclin D1/BRCA1 interaction in breast cancer cells

et al. 2007

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“…To further demonstrate that inhibition of CycD-CDK activity is not sufficient to induce all these changes in the absence of CDK2 inhibition, we determined the effects of a small chemical inhibitor specific for CDK4(6)-CycD1, fascaplysine. 24 This molecule appears to interfere with CycD-CDK activity without causing CKI redistribution. Thus, we anticipated that in the presence of fascaplysine CDK2-associated activity would remain unaltered, mimicking p16 actions on NP-18 cells.…”

Section: Effects Of Chemical Inhibition Of Cdk4 In Np-9 and Np-18 Cellsmentioning

confidence: 99%

The fate of pancreatic tumor cell lines following p16 overexpression depends on the modulation of CDK2 activity

et al. 2004

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Restitution of lost tumor-suppressor activities may be a promising strategy to target specifically cancer cells. However, the action of ectopically expressed tumor-suppressor genes depends on genetic background of tumoral cells. Ectopic expression of p16 INK4a induces either cell cycle arrest or apoptosis in different pancreatic cancer cell lines. We examined the molecular mechanisms mediating these two different cellular responses to p16 overexpression. Ectopic expression of p16 leads to G1 arrest in NP-9 cells by redistributing p21/p27 CKIs and inhibiting cyclin-dependent kinase CDK2 activity. In contrast, in NP-18 cells cyclin E (CycE)/ CDK2 activity is significantly higher and is not downregulated by p16-mediated redistribution of p21/p27. Moreover, inhibition of CDK4 activity with fascaplysine, which does not affect CycE/ CDK2 activity, reduces pocket protein phosphorylation in both cell lines, but fails to induce growth arrest. Like overexpression of p16, fascaplysine induces apoptosis in NP-18 cells, suggesting that inhibition of D-type cyclin/CDK activity in cells with high levels of CycE/CDK2 activity activates an apoptotic pathway. Inhibition of CycE/CDK2 activity via ectopic expression of p21 in NP-18 cells overexpressing p16 induces growth arrest and prevents p16-mediated apoptosis. Accordingly, silencing of p21 expression by using small interfering RNA switches the fate of p16-expressing NP-9 cells from cell cycle arrest to apoptosis. Our data suggest that, after CDK4/6 inactivation, the fate of pancreatic tumor cells depends on the ability to modulate CDK2 activity.

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“…So far, numerous specific CDK inhibitors have been identified on the basis of their ability to inhibit CDK1, CDK2 or CDK4: the purines olomoucine (Vesely et al, 1994), roscovitine de Azevedo et al, 1997); purvalanols (Gray et al, 1998;Chang et al, 1999;Villerbu et al, 2002), CVT-313 (Brooks et al, 1997), C2-alkylynated purines (Legraverend et al, 2000), H717 (Dreyer et al, 2001) and NU2058 (Arris et al, 2000), piperidine-substituted purines (Shum et al, 2001), toyocamycin (Park et al, 1996), flavopiridol (Losiewicz et al, 1994), indirubins (Hoessel et al, 1999;Leclerc et al, 2001), paullones Zaharevitz et al, 1999;Leost et al, 2000), g-butyrolactone (Kitagawa et al, 1993), hymenialdisine , indenopyrazoles (Nugiel et al, 2001), the pyrimidines NY6027 (Arris et al, 2000) and CGP60474 (Zimmermann, 1995), pyridopyrimidines (Barvian et al, 2000), the aminopyrimidine PNU 112455A (Clare et al, 2001), oxindoles (Kent et al, 1999;Davis et al, 2001), PD0183812 (Fry et al, 2001), cinnamaldehydes (Jeong et al, 2000), quinazolines (Shewchuk et al, 2000;Sielecki et al, 2001), fasclaplysin (Soni et al, 2000(Soni et al, , 2001, SU9516 (Lane et al, 2001) and benzocarbazoles (Carini et al, 2001). Despite their chemical diversity, these inhibitors all act by competing with ATP at the ATP-binding site of the kinase.…”

Section: Introductionmentioning

confidence: 99%

p42/p44 MAPKs are intracellular targets of the CDK inhibitor purvalanol

et al. 2002

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Chemical inhibitors of cyclin-dependent kinases (CDKs) have a great therapeutic potential against various proliferative and neurodegenerative disorders. Intensive screening of a combinatorial chemistry library of 2,6,9-trisubstituted purines has led to the identification of purvalanol, one of the most potent and selective CDK inhibitors to date. In preliminary studies, this compound demonstrates definite anti-mitotic properties, consistent with its nanomolar range efficiency towards purified CDK1 and CDK2. However, the actual intracellular targets of purvalanol remain to be identified, and a method for the determination of its in vivo selectivity was developed. In this technique, cell extracts were screened for purvalanolinteracting proteins by affinity chromatography on immobilized inhibitor. In addition to CDK1, p42/p44 MAPK were found to be two major purvalanol-interacting proteins in five different mammalian cell lines (CCL39, PC12, HBL100, MCF-7 and Jurkat cells), suggesting the generality of the purvalanol/p42/p44 MAPK interaction. The Chinese hamster lung fibroblast cell line CCL39 was used as a model to investigate the anti-proliferative properties of purvalanol. The compound inhibited cell growth with a GI 50 value of 2.5 mM and induced a G2/M block when added to exponentially growing cells. It did not appear to trigger massive activation of caspase. We next tested whether CDKs and p42/p44 MAPK were actually targeted by the compound in vivo. p42/p44 MAPK activity was visualized using an Elk -Gal4 luciferase reporter system and CDK1 activity was detected by the phosphonucleolin level. When cells were treated with purvalanol, p42/p44 MAPK and CDK1 activities were inhibited in a dose-dependent manner. Furthermore, purvalanol inhibited the nuclear accumulation of p42/p44 MAPK, an event dependent on the catalytic activity of these kinases. We conclude that the anti-proliferative properties of purvalanol are mediated by inhibition of both p42/p44 MAPK and CDKs. These observations highlight the potency of moderate selectivity compounds and encourage the search for new therapeutics which simultaneously target distinct but relevant pathways of cell proliferation.

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Inhibition of Cyclin-Dependent Kinase 4 (Cdk4) by Fascaplysin, a Marine Natural Product

Cited by 160 publications

References 46 publications

Functional consequences of cyclin D1/BRCA1 interaction in breast cancer cells

Functional consequences of cyclin D1/BRCA1 interaction in breast cancer cells

The fate of pancreatic tumor cell lines following p16 overexpression depends on the modulation of CDK2 activity

p42/p44 MAPKs are intracellular targets of the CDK inhibitor purvalanol

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