Discovery of a potent ROR1 inhibitor using μs-scale MD simulations, wt-metadynamics, and absolute binding free energy calculations

IF 2.5 4区化学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY

Journal of Molecular Modeling Pub Date : 2025-09-25 DOI:10.1007/s00894-025-06508-3

Shradheya R. R. Gupta, Rashmi Rameshwari, Indrakant Kumar Singh

{"title":"Discovery of a potent ROR1 inhibitor using μs-scale MD simulations, wt-metadynamics, and absolute binding free energy calculations","authors":"Shradheya R. R. Gupta, Rashmi Rameshwari, Indrakant Kumar Singh","doi":"10.1007/s00894-025-06508-3","DOIUrl":null,"url":null,"abstract":"<div><h3>Context</h3><p>Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is a cancer-associated pseudokinase with low expression in normal adult tissues but elevated levels in various malignancies, making it a promising therapeutic target. Among ~ 4 million compounds, CHEMBL3926946 emerged as the most promising candidate, demonstrating a persistent binding pose and a well-defined free energy basin. Well-tempered metadynamics (wt-MetaD) revealed a deep minimum of 26.00 ± 2.44 kcal/mol, indicating a highly stable interaction. CHEMBL3926946 exhibited a favourable Absolute Binding Free Energy Perturbation (ABFEP) of − 16.52 ± 0.37 kcal/mol, significantly outperforming the inhibitor Ponatinib (− 8.67 ± 0.94 kcal/mol), supported by persistent interactions with GLU523 and LEU479. This study highlights CHEMBL3926946 as a robust lead for ROR1-targeted cancer therapy and emphasizes the utility of combining wt-MetaD and ABFEP for reliable hit prioritization.</p><h3>Methods</h3><p>We employed a multilayered in silico pipeline integrating high-throughput virtual screening, long-timescale molecular dynamics (MD), wt-MetaD, and ABFEP. Ligands and protein were prepared using the OPLS2005 force field, and all stages up to wt-MetaD were conducted in Maestro (v12.8.117) using the same force field. A library of ~ 4 million compounds yielded 137 candidates, Further shortlisted via MD. 7 high-confidence molecules underwent 5 independent MD replicates with randomized seeds to ensure statistical robustness. The top 3 compounds were validated by 1 μs (1000 ns) simulations to assess long-term conformational stability and wt-MetaD to reveal deep minimum. ABFEP calculations were performed using the CGenFF force field in NAMD 3.0. We benchmarked ABFEP protocol against experimentally validated ligands, successfully reproducing experimental binding free energies (ΔG), confirming the protocol’s predictive reliability.</p><h3>Graphical Abstract</h3><div><figure><div><div><picture><source><img></source></picture></div></div></figure></div></div>","PeriodicalId":651,"journal":{"name":"Journal of Molecular Modeling","volume":"31 10","pages":""},"PeriodicalIF":2.5000,"publicationDate":"2025-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Molecular Modeling","FirstCategoryId":"92","ListUrlMain":"https://link.springer.com/article/10.1007/s00894-025-06508-3","RegionNum":4,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}

引用次数: 0

Abstract

Context

Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is a cancer-associated pseudokinase with low expression in normal adult tissues but elevated levels in various malignancies, making it a promising therapeutic target. Among ~ 4 million compounds, CHEMBL3926946 emerged as the most promising candidate, demonstrating a persistent binding pose and a well-defined free energy basin. Well-tempered metadynamics (wt-MetaD) revealed a deep minimum of 26.00 ± 2.44 kcal/mol, indicating a highly stable interaction. CHEMBL3926946 exhibited a favourable Absolute Binding Free Energy Perturbation (ABFEP) of − 16.52 ± 0.37 kcal/mol, significantly outperforming the inhibitor Ponatinib (− 8.67 ± 0.94 kcal/mol), supported by persistent interactions with GLU523 and LEU479. This study highlights CHEMBL3926946 as a robust lead for ROR1-targeted cancer therapy and emphasizes the utility of combining wt-MetaD and ABFEP for reliable hit prioritization.

Methods

We employed a multilayered in silico pipeline integrating high-throughput virtual screening, long-timescale molecular dynamics (MD), wt-MetaD, and ABFEP. Ligands and protein were prepared using the OPLS2005 force field, and all stages up to wt-MetaD were conducted in Maestro (v12.8.117) using the same force field. A library of ~ 4 million compounds yielded 137 candidates, Further shortlisted via MD. 7 high-confidence molecules underwent 5 independent MD replicates with randomized seeds to ensure statistical robustness. The top 3 compounds were validated by 1 μs (1000 ns) simulations to assess long-term conformational stability and wt-MetaD to reveal deep minimum. ABFEP calculations were performed using the CGenFF force field in NAMD 3.0. We benchmarked ABFEP protocol against experimentally validated ligands, successfully reproducing experimental binding free energies (ΔG), confirming the protocol’s predictive reliability.

Graphical Abstract

查看原文本刊更多论文

利用μs尺度MD模拟、wt-元动力学和绝对结合自由能计算，发现了一种有效的ROR1抑制剂。

背景：受体酪氨酸激酶样孤儿受体1 （ROR1）是一种与癌症相关的假激酶，在正常成人组织中低表达，但在各种恶性肿瘤中表达升高，使其成为一个有希望的治疗靶点。在约400万种化合物中，CHEMBL3926946表现出持久的结合姿态和明确的自由能盆地，成为最有希望的候选化合物。温调元动力学（wt-MetaD）的深度最小值为26.00±2.44 kcal/mol，表明相互作用高度稳定。CHEMBL3926946表现出良好的绝对结合自由能扰动（ABFEP），为- 16.52±0.37 kcal/mol，显著优于抑制剂Ponatinib(- 8.67±0.94 kcal/mol)，并与GLU523和LEU479持续相互作用。这项研究强调了CHEMBL3926946作为ror1靶向癌症治疗的强有力先导，并强调了wt-MetaD和ABFEP联合用于可靠的命中优先级排序的实用性。方法：采用集成高通量虚拟筛选、长时间分子动力学（MD）、wt-MetaD和ABFEP的多层硅管道。使用OPLS2005力场制备配体和蛋白，在Maestro （v12.8.117）中使用相同的力场进行直至wt-MetaD的所有阶段。一个包含约400万个化合物的文库产生了137个候选化合物，通过MD进一步入围。7个高可信度分子进行了5次独立的MD重复，随机种子，以确保统计稳健性。通过1 μs （1000 ns）模拟验证了前3个化合物的长期构象稳定性，并通过wt-MetaD验证了深层最小值。在NAMD 3.0中使用CGenFF力场进行ABFEP计算。我们将ABFEP方案与实验验证的配体进行了基准测试，成功地再现了实验结合自由能（ΔG），证实了该方案的预测可靠性。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

求助全文

约1分钟内获得全文求助全文

来源期刊

Journal of Molecular Modeling 化学-化学综合

CiteScore

3.50

自引率

4.50%

发文量

362

审稿时长

2.9 months

期刊介绍： The Journal of Molecular Modeling focuses on "hardcore" modeling, publishing high-quality research and reports. Founded in 1995 as a purely electronic journal, it has adapted its format to include a full-color print edition, and adjusted its aims and scope fit the fast-changing field of molecular modeling, with a particular focus on three-dimensional modeling. Today, the journal covers all aspects of molecular modeling including life science modeling; materials modeling; new methods; and computational chemistry. Topics include computer-aided molecular design; rational drug design, de novo ligand design, receptor modeling and docking; cheminformatics, data analysis, visualization and mining; computational medicinal chemistry; homology modeling; simulation of peptides, DNA and other biopolymers; quantitative structure-activity relationships (QSAR) and ADME-modeling; modeling of biological reaction mechanisms; and combined experimental and computational studies in which calculations play a major role.