Selective CDK9 Inhibition by Natural Compound Toyocamycin in Cancer Cells

Abstract

Aberrant transcription in cancer cells involves the silencing of tumor suppressor genes (TSGs) and activation of oncogenes. Transcriptomic changes are associated with epigenomic alterations such as DNA-hypermethylation, histone deacetylation, and chromatin condensation in promoter regions of silenced TSGs. To discover novel drugs that trigger TSG reactivation in cancer cells, we used a GFP-reporter system whose expression is silenced by promoter DNA hypermethylation and histone deacetylation. After screening a natural product drug library, we identified that toyocamycin, an adenosine-analog, induces potent GFP reactivation and loss of clonogenicity in human colon cancer cells. Connectivity-mapping analysis revealed that toyocamycin produces a pharmacological signature mimicking cyclin-dependent kinase (CDK) inhibitors. RNA-sequencing revealed that the toyocamycin transcriptomic signature resembles that of a specific CDK9 inhibitor (HH1). Specific inhibition of RNA Pol II phosphorylation level and kinase assays confirmed that toyocamycin specifically inhibits CDK9 (IC50 = 79 nM) with a greater efficacy than other CDKs (IC50 values between 0.67 and 15 µM). Molecular docking showed that toyocamycin efficiently binds the CDK9 catalytic site in a conformation that differs from other CDKs, explained by the binding contribution of specific amino acids within the catalytic pocket and protein backbone. Altogether, we demonstrated that toyocamycin exhibits specific CDK9 inhibition in cancer cells, highlighting its potential for cancer chemotherapy.

Recent Discovery and Development of Inhibitors that Target CDK9 and Their Therapeutic Indications

“…Furthermore, numerous natural products serve as potential candidates for CDK9 inhibitors, as inspiration for synthetic endeavors. The natural product toyocamycin ( 99 ) itself functions as an effective selective inhibitor with an IC 50 of 79 nM . Other natural or semisynthetic chemical structures, such as oroxylin A ( 100 ) and bufalin ( 101 ), also demonstrate promising pharmacological effects (Figure ).…”

“…The natural product toyocamycin (99) itself functions as an effective selective inhibitor with an IC 50 of 79 nM. 153 Other natural or semisynthetic chemical structures, such as oroxylin A (100) and bufalin (101), also demonstrate promising pharmacological effects (Figure 17). 4 In summary, natural products play a crucial role in inspiring the development of inhibitors.…”

Multifunctional nanomedicines-enabled chemodynamic-synergized multimodal tumor therapy via Fenton and Fenton-like reactions

“…Recently, chemotherapy drugs acting on endoplasmic reticulum stress (ERS) may be ideal for combination with CDT to achieve enhanced combined antitumor therapy 55 . Toyocamycin (Toy), as a promising anti-cancer drug, can prevent IRE1α splicing X-box binding protein 1 (XBP1u) to effectively inhibit the survival of cancer cells 60 . In the latest study, a dendritic polymer-drug conjugate using tannic acid (TA) chelated Fe 3+ was used for MR imaging-guided tumor CDT/chemotherapy by ERS amplification 61 .…”

Epigenetic Mechanisms Underlying Melanoma Resistance to Immune and Targeted Therapies

“…Interestingly, DNMT inhibitors sensitized a pre-clinical melanoma model to CTLA-4 immunotherapy [ 107 ] by de-repressing endogenous retroviruses (ERVs) and activating an IFN response. Additionally, cyclin-dependent kinase 9 (CDK9) inhibition with toyocamycin activated the SWI/SNF catalytic subunit BRG1 and synergized with DNMT inhibition to de-repress endogenous retroviral elements and interferon signaling, sensitizing an in vivo ovarian cancer model to PD-1 immunotherapy [ 108 , 109 ]. Here we examine recent studies advancing our understanding of epigenetic regulation of ERVs and the emerging role of viral mimicry in overcoming melanoma immunotherapy resistance.…”