2 research outputs found
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Expanding the Scope of Electrophiles Capable of Targeting K‑Ras Oncogenes
There is growing interest in reversible
and irreversible covalent
inhibitors that target noncatalytic amino acids in target proteins.
With a goal of targeting oncogenic K-Ras variants (e.g., G12D) by
expanding the types of amino acids that can be targeted by covalent
inhibitors, we survey a set of electrophiles for their ability to
label carboxylates. We functionalized an optimized ligand for the
K-Ras switch II pocket with a set of electrophiles previously reported
to react with carboxylates and characterized the ability of these
compounds to react with model nucleophiles and oncogenic K-Ras proteins.
Here, we report that aziridines and stabilized diazo groups preferentially
react with free carboxylates over thiols. Although we did not identify
a warhead that potently labels K-Ras G12D, we were able to study the
interactions of many electrophiles with K-Ras, as most of the electrophiles
rapidly label K-Ras G12C. We characterized the resulting complexes
by crystallography, hydrogen/deuterium exchange, and differential
scanning fluorimetry. Our results both demonstrate the ability of
a noncatalytic cysteine to react with a diverse set of electrophiles
and emphasize the importance of proper spatial arrangements between
a covalent inhibitor and its intended nucleophile. We hope that these
results can expand the range of electrophiles and nucleophiles of
use in covalent protein modulation
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Novel K‑Ras G12C Switch-II Covalent Binders Destabilize Ras and Accelerate Nucleotide Exchange
The success of targeted covalent
inhibitors in the global pharmaceutical
industry has led to a resurgence of covalent drug discovery. However,
covalent inhibitor design for flexible binding sites remains a difficult
task due to a lack of methodological development. Here, we compared
covalent docking to empirical electrophile screening against the highly
dynamic target K-Ras<sup>G12C</sup>. While the overall hit rate of
both methods was comparable, we were able to rapidly progress a docking
hit to a potent irreversible covalent binder that modifies the inactive,
GDP-bound state of K-Ras<sup>G12C</sup>. Hydrogen–deuterium
exchange mass spectrometry was used to probe the protein dynamics
of compound binding to the switch-II pocket and subsequent destabilization
of the nucleotide-binding region. SOS-mediated nucleotide exchange
assays showed that, contrary to prior switch-II pocket inhibitors,
these new compounds appear to accelerate nucleotide exchange. This
study highlights the efficiency of covalent docking as a tool for
the discovery of chemically novel hits against challenging targets