Fulcrum Occupancy–Leverage Perturbation Strategy Enables Rapid Discovery of Potent CDK2–Cyclin A2 Interaction Inhibitors

Traditional strategies for developing small-molecule inhibitors of protein-protein interactions (PPIs) are time-consuming and often yield low success rates due to the flat and dynamic interfaces of PPIs. To enable the rapid design of highly potent PPI inhibitors, we proposed a novel strategy named "Fulcrum Occupancy-Leverage Perturbation (FOLP)". In this strategy, high-affinity fragments serve as the "Fulcrum" by binding to the orthosteric pocket, while suitable moieties extend into allosteric sites near the PPI interface as "Leverage" to modulate the protein-protein interaction. As a proof of concept, the potent CDK2-Cyclin A2 PPI inhibitor LC-K2CAin-3, which fits the "FOLP" paradigm, was discovered with an IC<sub>50</sub> of 32.1 nM for inhibiting the interaction. Molecular dynamics simulations and cryptic pocket identification were employed, revealing the activation loop (A-loop) of CDK2 was flexible and targetable. X-ray crystallography and hydrogen deuterium exchange mass spectrometry (HDX-MS) analysis showed that LC-K2CAin-3 indeed bound to and stabilized the A-loop. LC-K2CAin-3 effectively inhibited the CDK2-Cyclin A2 interaction in CDK2 highly expressed melanoma cells, leading to cell cycle arrest and apoptosis and inhibition of CDK2 mediated signaling. In conclusion, the "FOLP" strategy offers a novel approach for PPI inhibitor discovery and could accelerate the development of PPI inhibitors.

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Publication Details

Journal
Angewandte Chemie International Edition
Published
2025-08-19
DOI
https://doi.org/10.1002/anie.202513542
Primary Topic
Chronic Lymphocytic Leukemia Research
Type
article
Field-Weighted Citation Impact
0.00

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article

Fulcrum Occupancy–Leverage Perturbation Strategy Enables Rapid Discovery of Potent CDK2–Cyclin A2 Interaction Inhibitors

Cheng Luo, Shuaishuai Chi, Mingchen Wang, Junyi Qiu et al.
Angewandte Chemie International Edition
Chronic Lymphocytic Leukemia Research
article

Fulcrum Occupancy–Leverage Perturbation Strategy Enables Rapid Discovery of Potent CDK2–Cyclin A2 Interaction Inhibitors

Cheng Luo, Shuaishuai Chi, Mingchen Wang, Junyi Qiu, Yaxi Yang, Hongbo Han, Bing Zhou, Jiacheng Li, Pan Xu, Mingyu Wang, Liping Liu, Chao Chen, Yi Chen, Yu-Lin Yang, Shijie Chen, Wei Fan, Tingting Lin, Ge Sun, Jie Zheng, Kaixian Chen, Ruyu Yan, Szu‐Chieh Fu
article en

Abstract

Traditional strategies for developing small-molecule inhibitors of protein-protein interactions (PPIs) are time-consuming and often yield low success rates due to the flat and dynamic interfaces of PPIs. To enable the rapid design of highly potent PPI inhibitors, we proposed a novel strategy named "Fulcrum Occupancy-Leverage Perturbation (FOLP)". In this strategy, high-affinity fragments serve as the "Fulcrum" by binding to the orthosteric pocket, while suitable moieties extend into allosteric sites near the PPI interface as "Leverage" to modulate the protein-protein interaction. As a proof of concept, the potent CDK2-Cyclin A2 PPI inhibitor LC-K2CAin-3, which fits the "FOLP" paradigm, was discovered with an IC<sub>50</sub> of 32.1 nM for inhibiting the interaction. Molecular dynamics simulations and cryptic pocket identification were employed, revealing the activation loop (A-loop) of CDK2 was flexible and targetable. X-ray crystallography and hydrogen deuterium exchange mass spectrometry (HDX-MS) analysis showed that LC-K2CAin-3 indeed bound to and stabilized the A-loop. LC-K2CAin-3 effectively inhibited the CDK2-Cyclin A2 interaction in CDK2 highly expressed melanoma cells, leading to cell cycle arrest and apoptosis and inhibition of CDK2 mediated signaling. In conclusion, the "FOLP" strategy offers a novel approach for PPI inhibitor discovery and could accelerate the development of PPI inhibitors.

Angewandte Chemie International EditionVol. 64(41)
Zhejiang Chinese Medical University (CN), Guiyang Medical University (CN), Chinese Academy of Sciences (CN), Shanghai Institute of Materia Medica (CN), Zhejiang Academy of Medical Sciences (CN), Drug Discovery Laboratory (Norway) (NO), Institute for Advanced Study (DE), University of Chinese Academy of Sciences (CN), East China Normal University (CN), Zhejiang University (CN)
Shanghai Municipal Health Commission, National Natural Science Foundation of China, Science and Technology Commission of Shanghai Municipality, National Key Research and Development Program of China, Program of Shanghai Academic Research Leader
Openalex Percentile: Top 71%
Chronic Lymphocytic Leukemia Research
DOISourceOpenAlex
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