Systematic cysteine scanning identifies a druggable pocket in oncogenic KRAS

  • Cell Chem Biol. 2026 Feb 19;33(2):241-255.e8. doi: 10.1016/j.chembiol.2026.01.007.
Laurens M van Tienen  1 ,  Shadwa Bayoumi  2 ,  Khaja Muneeruddin  3 ,  Nancy Leymarie  3 ,  Alexandra Popa  4 ,  Mrinal Shekhar  3 ,  Madeline Mueller  3 ,  Ruitong Li  2 ,  Krzysztof M Zak  4 ,  Harsha Chilukuri  3 ,  David J P Kornfilt  3 ,  Thomas C Atack  3 ,  Devishi Kesar  3 ,  Yuemin Bian  3 ,  Katharin L Shaw  5 ,  Zuzana Jandova  4 ,  Philipp Trollmann  4 ,  Leonhard Geist  4 ,  Peggy Stolt-Bergner  4 ,  Klaus Rumpel  4 ,  Dirk Kessler  4 ,  William R Sellers  6
Affiliations
  • 1. The Broad Institute of MIT and Harvard, Cambridge, MA, USA; Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA; Harvard Medical School, Boston, MA, USA. Electronic address: [email protected].
  • 2. The Broad Institute of MIT and Harvard, Cambridge, MA, USA; Harvard Medical School, Boston, MA, USA.
  • 3. The Broad Institute of MIT and Harvard, Cambridge, MA, USA.
  • 4. Boehringer Ingelheim RCV GmbH & Co KG, Vienna, Austria.
  • 5. The Broad Institute of MIT and Harvard, Cambridge, MA, USA; Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA; Harvard Medical School, Boston, MA, USA.
  • 6. The Broad Institute of MIT and Harvard, Cambridge, MA, USA; Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA; Harvard Medical School, Boston, MA, USA. Electronic address: [email protected].
PMID: 41679298 DOI: 10.1016/j.chembiol.2026.01.007
Abstract

The discovery of druggable pockets within proteins that lack traditional active sites remains a significant challenge in the development of therapeutics. To address this, we developed Cysteine Mapping of Accessible Pockets (CysMAP), a method for identifying druggable pockets in proteins. CysMAP employs systematic pooled cysteine (Cys)-variant libraries screened against diverse covalent compound libraries by intact LC-MS. We applied CysMAP to 189 KRAS(G12D) variants, purifying KRAS Cys-variants and screening them against 47 covalent compounds, quantifying accessibility, and reactivity across KRAS(G12D). We discovered previously unidentified ligand-bound states of Cys-variants surrounding the KRAS switch-II pocket. Structural studies of the D92C variant in complex with the compound BI-1830 uncovered a distinct novel binding pocket, highlighting the inherent plasticity of the region between switch-II and α3, that can accommodate diverse chemical entities in various conformations. This method holds significant potential for advancing drug discovery efforts against elusive targets such as oncogenic Ras mutants.

Keywords
KRAS; covalent compounds; cysteine scanning; intact mass spectrometry; screening.
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