Identifying Cancers Impacted by CDK8/19

CDK8 and CDK19 Mediator kinases are transcriptional co-regulators implicated in several types of cancer. Small-molecule CDK8/19 inhibitors have recently entered or are entering clinical trials, starting with breast cancer and acute myeloid leukemia (AML). To identify other cancers where these novel drugs may provide benefit, we queried genomic and transcriptomic databases for potential impact of CDK8, CDK19, or their binding partner CCNC. sgRNA analysis of a panel of tumor cell lines showed that most tumor types represented in the panel, except for some central nervous system tumors, were not dependent on these genes. In contrast, analysis of clinical samples for alterations in these genes revealed a high frequency of gene amplification in two highly aggressive subtypes of prostate cancer and in some cancers of the GI tract, breast, bladder, and sarcomas. Analysis of survival correlations identified a group of cancers where CDK8 expression correlated with shorter survival (notably breast, prostate, cervical cancers, and esophageal adenocarcinoma). In some cancers (AML, melanoma, ovarian, and others), such correlations were limited to samples with a below-median tumor mutation burden. These results suggest that Mediator kinases are especially important in cancers that are driven primarily by transcriptional rather than mutational changes and warrant an investigation of their role in additional cancer types

The Inhibition of CDK8/19 Mediator Kinases Prevents the Development of Resistance to EGFR-Targeting Drugs

Drug resistance is the main obstacle to achieving cures with both conventional and targeted anticancer drugs. The emergence of acquired drug resistance is initially mediated by non-genetic transcriptional changes, which occur at a much higher frequency than mutations and may involve population-scale transcriptomic adaptation. CDK8/19 kinases, through association with transcriptional Mediator complex, regulate transcriptional reprogramming by co-operating with different signal-responsive transcription factors. Here we tested if CDK8/19 inhibition could prevent adaptation to drugs acting on epidermal growth factor receptor (EGFR/ERBB1/HER1). The development of resistance was analyzed following long-term exposure of BT474 and SKBR3 breast cancer cells to EGFR-targeting small molecules (gefitinib, erlotinib) and of SW48 colon cancer cells to an anti-EGFR monoclonal antibody cetuximab. In all cases, treatment of small cell populations (~10 cells) with a single dose of the drug initially led to growth inhibition that was followed by the resumption of proliferation and development of drug resistance in the adapted populations. However, this adaptation was always prevented by the addition of selective CDK8/19 inhibitors, even though such inhibitors alone had only moderate or no effect on cell growth. These results indicate that combining EGFR-targeting drugs with CDK8/19 inhibitors may delay or prevent the development of tumor resistance to therapy

Structural insight into substrate and inhibitor discrimination by human P-glycoprotein

ISSN:0036-8075ISSN:1095-920

Structure of a zosuquidar and UIC2-bound human-mouse chimeric ABCB1

The multidrug transporter ABCB1 (P-glycoprotein) is an ATP-binding cassette transporter that has a key role in protecting tissues from toxic insult and contributes to multidrug extrusion from cancer cells. Here, we report the near-atomic resolution cryo-EM structure of nucleotide-free ABCB1 trapped by an engineered disulfide cross-link between the nucleotide-binding domains (NBDs) and bound to the antigen-binding fragment of the human-specific inhibitory antibody UIC2 and to the third-generation ABCB1 inhibitor zosuquidar. Our structure reveals the transporter in an occluded conformation with a central, enclosed, inhibitor-binding pocket lined by residues from all transmembrane (TM) helices of ABCB1. The pocket spans almost the entire width of the lipid membrane and is occupied exclusively by two closely interacting zosuquidar molecules. The external, conformational epitope facilitating UIC2 binding is also visualized, providing a basis for its inhibition of substrate efflux. Additional cryo-EM structures suggest concerted movement of TM helices from both halves of the transporters associated with closing the NBD gap, as well as zosuquidar binding. Our results define distinct recognition interfaces of ABCB1 inhibitory agents, which may be exploited for therapeutic purposes.ISSN:0027-8424ISSN:1091-649

The Inhibition of CDK8/19 Mediator Kinases Prevents the Development of Resistance to EGFR-Targeting Drugs

Drug resistance is the main obstacle to achieving cures with both conventional and targeted anticancer drugs. The emergence of acquired drug resistance is initially mediated by non-genetic transcriptional changes, which occur at a much higher frequency than mutations and may involve population-scale transcriptomic adaptation. CDK8/19 kinases, through association with transcriptional Mediator complex, regulate transcriptional reprogramming by co-operating with different signal-responsive transcription factors. Here we tested if CDK8/19 inhibition could prevent adaptation to drugs acting on epidermal growth factor receptor (EGFR/ERBB1/HER1). The development of resistance was analyzed following long-term exposure of BT474 and SKBR3 breast cancer cells to EGFR-targeting small molecules (gefitinib, erlotinib) and of SW48 colon cancer cells to an anti-EGFR monoclonal antibody cetuximab. In all cases, treatment of small cell populations (~105 cells) with a single dose of the drug initially led to growth inhibition that was followed by the resumption of proliferation and development of drug resistance in the adapted populations. However, this adaptation was always prevented by the addition of selective CDK8/19 inhibitors, even though such inhibitors alone had only moderate or no effect on cell growth. These results indicate that combining EGFR-targeting drugs with CDK8/19 inhibitors may delay or prevent the development of tumor resistance to therapy

Structure of a zosuquidar and UIC2-bound human-mouse chimeric ABCB1

The multidrug transporter ABCB1 (P-glycoprotein) is an ATP-binding cassette transporter that has a key role in protecting tissues from toxic insult and contributes to multidrug extrusion from cancer cells. Here, we report the near-atomic resolution cryo-EM structure of nucleotide-free ABCB1 trapped by an engineered disulfide cross-link between the nucleotide-binding domains (NBDs) and bound to the antigen-binding fragment of the human-specific inhibitory antibody UIC2 and to the third-generation ABCB1 inhibitor zosuquidar. Our structure reveals the transporter in an occluded conformation with a central, enclosed, inhibitor-binding pocket lined by residues from all transmembrane (TM) helices of ABCB1. The pocket spans almost the entire width of the lipid membrane and is occupied exclusively by two closely interacting zosuquidar molecules. The external, conformational epitope facilitating UIC2 binding is also visualized, providing a basis for its inhibition of substrate efflux. Additional cryo-EM structures suggest concerted movement of TM helices from both halves of the transporters associated with closing the NBD gap, as well as zosuquidar binding. Our results define distinct recognition interfaces of ABCB1 inhibitory agents, which may be exploited for therapeutic purposes

Inhibition of Cyclin-Dependent Kinases 8/19 Restricts Bacterial and Virus-Induced Inflammatory Responses in Monocytes

Hyperactivation of the immune system remains a dramatic, life-threatening complication of viral and bacterial infections, particularly during pneumonia. Therapeutic approaches to counteract local and systemic outbreaks of cytokine storm and to prevent tissue damage remain limited. Cyclin-dependent kinases 8 and 19 (CDK8/19) potentiate transcriptional responses to the altered microenvironment, but CDK8/19 potential in immunoregulation is not fully understood. In the present study, we investigated how a selective CDK8/19 inhibitor, Senexin B, impacts the immunogenic profiles of monocytic cells stimulated using influenza virus H1N1 or bacterial lipopolysaccharides. Senexin B was able to prevent the induction of gene expression of proinflammatory cytokines in THP1 and U937 cell lines and in human peripheral blood-derived mononuclear cells. Moreover, Senexin B substantially reduced functional manifestations of inflammation, including clustering and chemokine-dependent migration of THP1 monocytes and human pulmonary fibroblasts (HPF)