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Efficient targeted degradation via reversible and irreversible covalent PROTACs

preprint
revised on 15.03.2020 and posted on 17.03.2020 by Ronen Gabizon, Amit Shraga, Paul Gehrtz, Ella Livnah, Yamit Shorer, Neta Gurwicz, Liat Avram, Tamar Unger, Hila Aharoni, Shira Albeck, Alexander Brandis, Ziv Shulman, Ben- Zion Katz, Yair Herishanu, Nir London

PROteolysis Targeting Chimeras (PROTACs) represent an exciting inhibitory modality with many advantages, including sub-stoichiometric degradation of targets. Their scope, though, is still limited to-date by the requirement for a sufficiently potent target binder. A solution that proved useful in tackling challenging targets is the use of electrophiles to allow irreversible binding to the target. However, such binding will negate the catalytic nature of PROTACs. Reversible covalent PROTACs potentially offer the best of both worlds. They possess the potency and selectivity associated with the formation of the covalent bond, while being able to dissociate and regenerate once the protein target is degraded. Using Bruton’s tyrosine kinase (BTK) as a clinically relevant model system, we show efficient covalent degradation by non-covalent, irreversible covalent and reversible covalent PROTACs, with <10 nM DC50’s and >85% degradation. Our data suggests that part of the degradation by our irreversible covalent PROTACs is driven by reversible binding prior to covalent bond formation, while the reversible covalent PROTACs drive degradation primarily by covalent engagement. The PROTACs showed enhanced inhibition of B cell activation compared to Ibrutinib, and exhibit potent degradation of BTK in patients-derived primary chronic lymphocytic leukemia cells. The most potent reversible covalent PROTAC, RC-3, exhibited enhanced selectivity towards BTK compared to non-covalent and irreversible covalent PROTACs. These compounds may pave the way for the design of covalent PROTACs for a wide variety of challenging targets.

Funding

S.A. is the incumbent of Prof. David Casson Research Fellowship, N.L. is the incumbent of the Alan and Laraine Fischer Career Development Chair; N.L. would like to acknowledge funding from the Israel Science Foundation (grant No. 2462/19), The Rising Tide Foundation, The Israel Cancer Research Fund, the Israeli Ministry of Science. Technology (grant No. 3-14763), and the Moross integrated cancer center. N.L. is also supported by the Helen and Martin Kimmel Center for Molecular Design, Joel and Mady Dukler Fund for Cancer Research, the Estate of Emile Mimran and Virgin JustGiving and the George Schwartzman Fund. R.G. was supported by the state of Israel, ministry of Aliyah, Center for Integration in Science. Y.H. is supported by grants from the Israeli Science Foundation (1707/19), Israeli Cancer Association and Sackler Faculity of Medicine, Tel-Aviv University.

History

Email Address of Submitting Author

nir.london@weizmann.ac.il

Institution

The Weizmann Institute of Science

Country

Israel

ORCID For Submitting Author

https://orcid.org/0000-0003-2687-0699

Declaration of Conflict of Interest

no conflict of interest

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