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Pharmacological perturbation of CDK9 using selective CDK9 inhibition or degradation

Abstract

Cyclin-dependent kinase 9 (CDK9), an important regulator of transcriptional elongation, is a promising target for cancer therapy, particularly for cancers driven by transcriptional dysregulation. We characterized NVP-2, a selective ATP-competitive CDK9 inhibitor, and THAL-SNS-032, a selective CDK9 degrader consisting of a CDK-binding SNS-032 ligand linked to a thalidomide derivative that binds the E3 ubiquitin ligase Cereblon (CRBN). To our surprise, THAL-SNS-032 induced rapid degradation of CDK9 without affecting the levels of other SNS-032 targets. Moreover, the transcriptional changes elicited by THAL-SNS-032 were more like those caused by NVP-2 than those induced by SNS-032. Notably, compound washout did not significantly reduce levels of THAL-SNS-032-induced apoptosis, suggesting that CDK9 degradation had prolonged cytotoxic effects compared with CDK9 inhibition. Thus, our findings suggest that thalidomide conjugation represents a promising strategy for converting multi-targeted inhibitors into selective degraders and reveal that kinase degradation can induce distinct pharmacological effects compared with inhibition.

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Figure 1: THAL-SNS-032 selectively induces degradation of CDK9.
Figure 2: NVP-2 selectively inhibits CDK9.
Figure 3: THAL-SNS-032 exhibits CRBN-dependent anti-proliferative and pro-apoptotic effects.
Figure 4: THAL-SNS-032 exhibits transcriptional effects consistent with a selective CDK9 inhibitor.
Figure 5: THAL-SNS-032 diminishes elongating polymerase II.

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Acknowledgements

We thank D. Buckley, E. Wang, J. Paulk, and members of the Gray and Bradner laboratories for helpful discussions. This work was supported by the US National Institutes of Health (CA179483-03 to N.K. and N.S.G., and T32GM007753 to Z.Z.), the Koch Institute and Dana-Farber/Harvard Cancer Center Bridge Grant (N.K., N.S.G., R.A.Y., C.M.O.),

Author information

Authors and Affiliations

Authors

Contributions

N.S.G., C.M.O., N.K. and T.Z. conceived the project. N.S.G., Y.L., B.J. and T.Z. conceived and directed chemistry effort. Chemical synthesis and small-molecule structure determination was performed by T.Z., Y.L. and Z.Z. R.A.Y., N.S.G., C.M.O., M.A.E., G.E.W. and N.K. conceived genomics effort. N.K. and C.M.O. designed and executed cellular biological experimental research with input from N.S.G. and R.A.Y. W.H. and M.B. designed and executed the proteomics efforts. E.S.F. designed and executed statistical analysis of proteomics. T.N. and J.G. designed and executed the Kinativ efforts. M.A.E. designed and performed genomics data analyses. C.M.O., N.K. and N.S.G. wrote the manuscript. All of the authors edited the manuscript.

Corresponding author

Correspondence to Nathanael S Gray.

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Competing interests

N.S.G. is a Scientific Founder and member of the Scientific Advisory Board of C4 Therapeutics, Syros Pharmaceuticals and Petra Pharmaceuticals and is the inventor on intellectual property licensed to these entities. J.E.B. is a Scientific Founder of Syros Pharmaceuticals, SHAPE Pharmaceuticals, Acetylon Pharmaceuticals, Tensha Therapeutics (now Roche) and C4 Therapeutics and is the inventor on intellectual property licensed to these entities. J.E.B. is now an executive and shareholder in Novartis AG. R.A.Y. is a Scientific Founder of Syros Pharmaceuticals and Marauder Therapeutics.

Supplementary information

Supplementary Text and Figures

Supplementary Results, Supplementary Tables 1–2, Supplementary Figures 1–9 (PDF 2413 kb)

Life Sciences Reporting Summary (PDF 158 kb)

Supplementary Note 1

Chemical Synthesis of THAL-SNS-032, NVP-2, THAL-NVP-2-02-099, THAL-NVP-2- 3 03-069, HAL-NVP-2-03-099, THAL-NVP-2-03-084, THAL-NVP-2-03-105. (PDF 668 kb)

Supplementary Dataset 1

Lysate Kinativ of SNS-032, THAL-SNS-032, and NVP-2 (XLSX 39 kb)

Supplementary Dataset 2

Proteomics of THAL-SNS-032 treatment (XLSX 328 kb)

Supplementary Dataset 3

Selectivity of NVP-2 by Ambit profiling (XLSX 23 kb)

Supplementary Dataset 4

Live cell Kinativ of SNS-032 and THAL-SNS-032 (XLSX 25 kb)

Supplementary Dataset 5

Live cell Kinativ of NVP-2 (XLSX 29 kb)

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Olson, C., Jiang, B., Erb, M. et al. Pharmacological perturbation of CDK9 using selective CDK9 inhibition or degradation. Nat Chem Biol 14, 163–170 (2018). https://doi.org/10.1038/nchembio.2538

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