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Discovery and mechanistic characterization of A-485, a potent p300/CBP catalytic inhibitor

ABSTRACT: The dynamic and reversible acetylation of proteins catalyzed by histone acetyltransferases (HATs) and histone deacetylases (HDACs) was discovered more than 2 decades ago and the enzymatic function of these enzymes are established as a major epigenetic regulatory mechanism of gene transcription. Thus, these epigenetic modifiers are involved in multiple diseases and represent attractive targets for therapeutic intervention. While HDAC inhibitors have been developed and approved by the FDA to treat certain cancers, progress on the development of drug-like HAT inhibitors has lagged. The HAT paralogs p300 and CBP (here called p300/CBP) are key transcriptional co-activators that are essential for a multitude of cellular processes and also implicated in human pathological conditions, including cancer. Current p300/CBP HAT domain inhibitors including natural products and bisubstrate analogs such as Lys-CoA either lack potency and selectivity or suffer from poor cellular permeability. C646 is widely utilized as a tool to inhibit p300/CBP HAT activity, but its off-target activity and reactivity may limit its cellular specificity. Here, we describe A-485 as a potent, selective and drug-like p300/CBP catalytic inhibitor. We show the first high resolution (1.95Å) co-crystal structure of a pharmacologically active small molecule (A-485) bound to the catalytic active site of p300 HAT domain and demonstrate that A-485 is an acetyl-CoA competitive inhibitor of p300/CBP. A-485 selectively inhibited proliferation across lineage-specific tumor types, including several hematological malignancies and androgen receptor-positive prostate cancer. A-485 robustly inhibited the androgen receptor transcriptional program in both androgen sensitive and castrate resistant prostate cancer and inhibited tumor growth in a castration resistant xenograft model. These results demonstrate the feasibility of selectively drugging the catalytic activity of histone acetyltransferases, provide the framework for delineating the enzymatic functions of HATs, and pave the way for the development of novel therapeutics targeting HAT activity.

ORGANISM(S): Homo sapiens

PROVIDER: GSE94580 | GEO | 2017/09/27

SECONDARY ACCESSION(S): PRJNA371497

REPOSITORIES: GEO

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Project description:EP300 and CBP are paralogous, multidomain histone acetyltransferases (HATs) that regulate gene expression by binding to and acetylating diverse proteins. With widespread effects on gene regulation, these enzymes are attractive targets for therapeutic development. Discovery efforts for chemical inhibitors typically target protein domains that are amenable to probe binding. In some cases, probes have been identified for more than one domain within a single protein, including high potency inhibitors of both the catalytic HAT domain and bromodomain (BRD) that mediate the enzymatic and protein binding (“reader”) activities of EP300/CBP. In cancer cell systems, combination inhibition of both domains is more toxic than single inhibitor alone, indicating that inhibition of individual domain functions may elicit differential effects. Thus, we hypothesized that determining the relative effects of domain-specific inhibition of EP300/CBP on tumor cell growth may elucidate exceptional-responding tumor types to one inhibitor or the other. This would permit a broader therapeutic index by enabling reduced dosing of domain-targeted inhibitors, while maintaining enhanced inhibition of cancer cell growth. Here, using high-throughput cell line-based screening, we demonstrate that targeting the HAT or BRD domains of EP300/CBP using the chemical probes A485 and CCS1477 has differential effects in select tumor types. Group 3 medulloblastoma (G3MB) cells were especially sensitive to EP300/CBP BRD inhibition, as compared with HAT inhibition. Structurally, these effects are mediated by the difluorophenyl group in the catalytic core of CCS1477. Mechanistically, the effects of bromodomain and HAT-domain specific inhibition are distinct, with bromodomain inhibition causing rapid early disruption of genetic dependency networks that are required for G3MB growth. These studies provide a domain-specific structural foundation for drug discovery efforts targeting EP300/CBP and identify a selective role for the EP300/CBP bromodomain in maintaining genetic dependency networks in G3MB cells.

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Discovery and mechanistic characterization of A-485, a potent p300/CBP catalytic inhibitor

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