Discovery of Novel CRK12 Inhibitors for the Treatment of Human African Trypanosomiasis: An Integrated Computational and Experimental Approach
<b>Background:</b> Human African trypanosomiasis (HAT), caused by <i>Trypanosoma brucei</i>, is a neglected tropical disease with limited treatments, highlighting the pressing need for new drugs. Cell division cycle-2-related kinase 12 (CRK12), a pivotal protein involved in t...
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2025-05-01
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| author | Qin Li Jiayi Luo Chenggong Fu Wenqingqing Kang Lingling Wang Henry Tong Zhaorong Lun Qianqian Zhang Dehua Lai Huanxiang Liu |
| author_facet | Qin Li Jiayi Luo Chenggong Fu Wenqingqing Kang Lingling Wang Henry Tong Zhaorong Lun Qianqian Zhang Dehua Lai Huanxiang Liu |
| author_sort | Qin Li |
| collection | DOAJ |
| description | <b>Background:</b> Human African trypanosomiasis (HAT), caused by <i>Trypanosoma brucei</i>, is a neglected tropical disease with limited treatments, highlighting the pressing need for new drugs. Cell division cycle-2-related kinase 12 (CRK12), a pivotal protein involved in the cell cycle regulation of <i>T. brucei</i>, has emerged as a promising therapeutic target for HAT, yet effective CRK12 inhibitors remain lacking. <b>Methods:</b> An integrated strategy combining computational modeling, virtual screening, molecular dynamics (MD) simulations, and experimental validation was adopted to discover potential inhibitors against CRK12. By using the predicted and refined 3D structure of CRK12 from AlphaFold2 and MD simulation, over 1.5 million compounds were screened based on multiple-scale molecular docking, and 26 compounds were selected for evaluation of biological activity based on anti-<i>T. brucei</i> bioassays. Dose–response curves were generated for the most potent inhibitors, and the interaction mechanism between the top four compounds and CRK12 was explored by MD simulations and MM/GBSA binding free energy analysis. <b>Results:</b> Of the 26 compounds, six compounds demonstrated sub-micromolar to low-micromolar IC<sub>50</sub> values (0.85–3.50 µM). The top four hits, F733-0072, F733-0407, L368-0556, and L439-0038, exhibited IC<sub>50</sub> values of 1.11, 1.97, 0.85, and 1.66 µM, respectively. Binding free energy and energy decomposition analyses identified ILE335, VAL343, PHE430, ALA433, and LEU482 as hotspot residues for compound binding. Hydrogen bonding analysis demonstrated that these compounds can form stable hydrogen bonds with the hinge residue ALA433, ensuring their stable binding within the active site. <b>Conclusions:</b> This study establishes a robust and cost-effective pipeline for CRK12 inhibitor discovery, identifying several novel inhibitors demonstrating promising anti-HAT activity. The newly discovered scaffolds exhibit structural diversity distinct from known CRK12 inhibitors, providing valuable lead compounds for anti-trypanosomal drug development. |
| format | Article |
| id | doaj-art-5b6aca9dac9145bca28bbf74fdb14c99 |
| institution | OA Journals |
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| publishDate | 2025-05-01 |
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| spelling | doaj-art-5b6aca9dac9145bca28bbf74fdb14c992025-08-20T02:21:57ZengMDPI AGPharmaceuticals1424-82472025-05-0118677810.3390/ph18060778Discovery of Novel CRK12 Inhibitors for the Treatment of Human African Trypanosomiasis: An Integrated Computational and Experimental ApproachQin Li0Jiayi Luo1Chenggong Fu2Wenqingqing Kang3Lingling Wang4Henry Tong5Zhaorong Lun6Qianqian Zhang7Dehua Lai8Huanxiang Liu9Faculty of Applied Sciences, Macao Polytechnic University, Macao SAR, ChinaMOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol and Guangdong Provincial Key Laboratory of Aquatic Economic Animals, School of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, ChinaFaculty of Applied Sciences, Macao Polytechnic University, Macao SAR, ChinaFaculty of Applied Sciences, Macao Polytechnic University, Macao SAR, ChinaFaculty of Applied Sciences, Macao Polytechnic University, Macao SAR, ChinaFaculty of Applied Sciences, Macao Polytechnic University, Macao SAR, ChinaMOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol and Guangdong Provincial Key Laboratory of Aquatic Economic Animals, School of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, ChinaFaculty of Applied Sciences, Macao Polytechnic University, Macao SAR, ChinaMOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol and Guangdong Provincial Key Laboratory of Aquatic Economic Animals, School of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, ChinaFaculty of Applied Sciences, Macao Polytechnic University, Macao SAR, China<b>Background:</b> Human African trypanosomiasis (HAT), caused by <i>Trypanosoma brucei</i>, is a neglected tropical disease with limited treatments, highlighting the pressing need for new drugs. Cell division cycle-2-related kinase 12 (CRK12), a pivotal protein involved in the cell cycle regulation of <i>T. brucei</i>, has emerged as a promising therapeutic target for HAT, yet effective CRK12 inhibitors remain lacking. <b>Methods:</b> An integrated strategy combining computational modeling, virtual screening, molecular dynamics (MD) simulations, and experimental validation was adopted to discover potential inhibitors against CRK12. By using the predicted and refined 3D structure of CRK12 from AlphaFold2 and MD simulation, over 1.5 million compounds were screened based on multiple-scale molecular docking, and 26 compounds were selected for evaluation of biological activity based on anti-<i>T. brucei</i> bioassays. Dose–response curves were generated for the most potent inhibitors, and the interaction mechanism between the top four compounds and CRK12 was explored by MD simulations and MM/GBSA binding free energy analysis. <b>Results:</b> Of the 26 compounds, six compounds demonstrated sub-micromolar to low-micromolar IC<sub>50</sub> values (0.85–3.50 µM). The top four hits, F733-0072, F733-0407, L368-0556, and L439-0038, exhibited IC<sub>50</sub> values of 1.11, 1.97, 0.85, and 1.66 µM, respectively. Binding free energy and energy decomposition analyses identified ILE335, VAL343, PHE430, ALA433, and LEU482 as hotspot residues for compound binding. Hydrogen bonding analysis demonstrated that these compounds can form stable hydrogen bonds with the hinge residue ALA433, ensuring their stable binding within the active site. <b>Conclusions:</b> This study establishes a robust and cost-effective pipeline for CRK12 inhibitor discovery, identifying several novel inhibitors demonstrating promising anti-HAT activity. The newly discovered scaffolds exhibit structural diversity distinct from known CRK12 inhibitors, providing valuable lead compounds for anti-trypanosomal drug development.https://www.mdpi.com/1424-8247/18/6/778CRK12human African trypanosomiasis (HAT)molecular dockingvirtual screeningneglected tropical diseasedrug discovery |
| spellingShingle | Qin Li Jiayi Luo Chenggong Fu Wenqingqing Kang Lingling Wang Henry Tong Zhaorong Lun Qianqian Zhang Dehua Lai Huanxiang Liu Discovery of Novel CRK12 Inhibitors for the Treatment of Human African Trypanosomiasis: An Integrated Computational and Experimental Approach Pharmaceuticals CRK12 human African trypanosomiasis (HAT) molecular docking virtual screening neglected tropical disease drug discovery |
| title | Discovery of Novel CRK12 Inhibitors for the Treatment of Human African Trypanosomiasis: An Integrated Computational and Experimental Approach |
| title_full | Discovery of Novel CRK12 Inhibitors for the Treatment of Human African Trypanosomiasis: An Integrated Computational and Experimental Approach |
| title_fullStr | Discovery of Novel CRK12 Inhibitors for the Treatment of Human African Trypanosomiasis: An Integrated Computational and Experimental Approach |
| title_full_unstemmed | Discovery of Novel CRK12 Inhibitors for the Treatment of Human African Trypanosomiasis: An Integrated Computational and Experimental Approach |
| title_short | Discovery of Novel CRK12 Inhibitors for the Treatment of Human African Trypanosomiasis: An Integrated Computational and Experimental Approach |
| title_sort | discovery of novel crk12 inhibitors for the treatment of human african trypanosomiasis an integrated computational and experimental approach |
| topic | CRK12 human African trypanosomiasis (HAT) molecular docking virtual screening neglected tropical disease drug discovery |
| url | https://www.mdpi.com/1424-8247/18/6/778 |
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