A Novel CDK9 Inhibitor Shows Potent Antitumor Efficacy in Preclinical Hematologic Tumor Models

Yin, Tinggui; Lallena, Marı́a José; Kreklau, Emiko L.; Fales, Kevin R.; Carballares, Santiago; Torrres, Raquel; Wishart, Graham N.; Ajamie, Rose T.; Cronier, Damien M.; Iversen, Phillip W.; Meier, Timothy I.; Foreman, Robert T.; Zeckner, Douglas J.; Sissons, Sean E.; Halstead, Bart W.; Lin, Aimee Bence; Donoho, Gregory P.; Qian, Yue‐Wei; Li, Shuyu; Wu, Song; Aggarwal, Amit; Ye, Xiang S.; Starling, James J.; Gaynor, Richard B.; Dios, Alfonso De; Du, Jinyan

doi:10.1158/1535-7163.mct-13-0849

Cited by 84 publications

(67 citation statements)

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“…Although FLVP inhibits CDK9 preferentially [45], inhibition of additional kinases by FLVP [52–54] could reduce pS187-H1.4 levels and bias the identification of CDK9 as a major kinase for H1.4-S187. We investigated this possibility using THZ1, a recently developed inhibitor that exhibits extraordinary selectivity for CDK7 due to a novel mechanism of inhibition that involves covalent binding to a cysteine residue outside of the kinase domain that is unique to CDK7 [55].…”

Section: Resultsmentioning

confidence: 99%

Site-specific regulation of histone H1 phosphorylation in pluripotent cell differentiation

Liao

Mizzen

2017

Epigenetics & Chromatin

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BackgroundStructural variation among histone H1 variants confers distinct modes of chromatin binding that are important for differential regulation of chromatin condensation, gene expression and other processes. Changes in the expression and genomic distributions of H1 variants during cell differentiation appear to contribute to phenotypic differences between cell types, but few details are known about the roles of individual H1 variants and the significance of their disparate capacities for phosphorylation. In this study, we investigated the dynamics of interphase phosphorylation at specific sites in individual H1 variants during the differentiation of pluripotent NT2 and mouse embryonic stem cells and characterized the kinases involved in regulating specific H1 variant phosphorylations in NT2 and HeLa cells.ResultsHere, we show that the global levels of phosphorylation at H1.5-Ser18 (pS18-H1.5), H1.2/H1.5-Ser173 (pS173-H1.2/5) and H1.4-Ser187 (pS187-H1.4) are regulated differentially during pluripotent cell differentiation. Enrichment of pS187-H1.4 near the transcription start site of pluripotency factor genes in pluripotent cells is markedly reduced upon differentiation, whereas pS187-H1.4 levels at housekeeping genes are largely unaltered. Selective inhibition of CDK7 or CDK9 rapidly diminishes pS187-H1.4 levels globally and its enrichment at housekeeping genes, and similar responses were observed following depletion of CDK9. These data suggest that H1.4-S187 is a bona fide substrate for CDK9, a notion that is further supported by the significant colocalization of CDK9 and pS187-H1.4 to gene promoters in reciprocal re-ChIP analyses. Moreover, treating cells with actinomycin D to inhibit transcription and trigger the release of active CDK9/P-TEFb from 7SK snRNA complexes induces the accumulation of pS187-H1.4 at promoters and gene bodies. Notably, the levels of pS187-H1.4 enrichment after actinomycin D treatment or cell differentiation reflect the extent of CDK9 recruitment at the same loci. Remarkably, the global levels of H1.5-S18 and H1.2/H1.5-S173 phosphorylation are not affected by these transcription inhibitor treatments, and selective inhibition of CDK2 does not affect the global levels of phosphorylation at H1.4-S187 or H1.5-S18.ConclusionsOur data provide strong evidence that H1 variant interphase phosphorylation is dynamically regulated in a site-specific and gene-specific fashion during pluripotent cell differentiation, and that enrichment of pS187-H1.4 at genes is positively related to their transcription. H1.4-S187 is likely to be a direct target of CDK9 during interphase, suggesting the possibility that this particular phosphorylation may contribute to the release of paused RNA pol II. In contrast, the other H1 variant phosphorylations we investigated appear to be mediated by distinct kinases and further analyses are needed to determine their functional significance.Electronic supplementary materialThe online version of this article (doi:10.1186/s13072-017-0135-3) contains supplementary material,...

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Section: Resultsmentioning

confidence: 99%

Site-specific regulation of histone H1 phosphorylation in pluripotent cell differentiation

Liao

Mizzen

2017

Epigenetics & Chromatin

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“…Vehicle alone or LY2606368 was administered BIDx3, followed by 4 days of rest and repeated for an additional two cycles. Tumor size and body weight were recorded biweekly and compared between vehicleand drug-treated groups (23).…”

Section: In Vivo Biochemistry and Tumor Growth Inhibitionmentioning

confidence: 99%

LY2606368 Causes Replication Catastrophe and Antitumor Effects through CHK1-Dependent Mechanisms

King

Diaz

McNeely

et al. 2015

Molecular Cancer Therapeutics

151

175

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CHK1 is a multifunctional protein kinase integral to both the cellular response to DNA damage and control of the number of active replication forks. CHK1 inhibitors are currently under investigation as chemopotentiating agents due to CHK1's role in establishing DNA damage checkpoints in the cell cycle. Here, we describe the characterization of a novel CHK1 inhibitor, LY2606368, which as a single agent causes double-stranded DNA breakage while simultaneously removing the protection of the DNA damage checkpoints. The action of LY2606368 is dependent upon inhibition of CHK1 and the corresponding increase in CDC25A activation of CDK2, which increases the number of replication forks while reducing their stability. Treatment of cells with LY2606368 results in the rapid appearance of TUNEL and pH2AX-positive double-stranded DNA breaks in the S-phase cell population. Loss of the CHK1-dependent DNA damage checkpoints permits cells with damaged DNA to proceed into early mitosis and die. The majority of treated mitotic nuclei consist of extensively fragmented chromosomes. Inhibition of apoptosis by the caspase inhibitor Z-VAD-FMK had no effect on chromosome fragmentation, indicating that LY2606368 causes replication catastrophe. Changes in the ratio of RPA2 to phosphorylated H2AX following LY2606368 treatment further support replication catastrophe as the mechanism of DNA damage. LY2606368 shows similar activity in xenograft tumor models, which results in significant tumor growth inhibition. LY2606368 is a potent representative of a novel class of drugs for the treatment of cancer that acts through replication catastrophe.

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“…Inhibition of CDK9 leads to reduced phosphorylation of Ser2 on RPB1, which results in the reduction of Mcl-1 levels. 19–21 …”

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confidence: 99%

Chemically induced degradation of CDK9 by a proteolysis targeting chimera (PROTAC)

Robb¹,

Contreras²,

Kour³

et al. 2017

Chem. Commun.

178

141

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Cyclin-dependent kinase 9 (CDK9), a member of the cyclin-dependent protein kinase (CDK) family, is involved in transcriptional elongation of several target genes. CDK9 is ubiquitously expressed and has been shown to contribute to a variety of malignancies such as pancreatic, prostate and breast cancers. Here we report the development of a heterobifunctional small molecule proteolysis targeting chimera (PROTAC) capable of cereblon (CRBN) mediated proteasomal degradation of CDK9. In HCT116 cells, it selectively degrades CDK9 while sparing other CDK family members. This is the first example of a PROTAC that selectively degrades CDK9.

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A Novel CDK9 Inhibitor Shows Potent Antitumor Efficacy in Preclinical Hematologic Tumor Models

Cited by 84 publications

References 47 publications

Site-specific regulation of histone H1 phosphorylation in pluripotent cell differentiation

Site-specific regulation of histone H1 phosphorylation in pluripotent cell differentiation

LY2606368 Causes Replication Catastrophe and Antitumor Effects through CHK1-Dependent Mechanisms

Chemically induced degradation of CDK9 by a proteolysis targeting chimera (PROTAC)

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