title: Molecular dynamics simulation and free energy landscape methods in probing L215H, L217R and L225M βI-tubulin mutations causing paclitaxel resistance in cancer cells.
abstract: Drug resistance poses a threatening challenge for mankind, as the development of resistance to already well-established drugs causes serious therapeutic problems. Resistance to paclitaxel (Ptxl), a complex diterpenoid working as microtubule stabilizer, is one such issue in cancer treatment. Microtubule stabilizer drugs, stabilises microtubules upon binding to β-tubulin subunit of tubulin heterodimer thus causing mitotic arrest leading to death of cancer cell. Leucine point mutations viz. L215H, L217R, and L225M were reported for Ptxl resistance in various cancers. In the current study, molecular mechanism of these resistance causing mutations was explored using molecular docking, molecular dynamics (MD) simulation, binding energy estimation (MMPBSA), free energy decomposition, principle component analysis (PCA) and free energy landscape (FEL) methods. A total of five systems including unbound βI-tubulin (Apo), docked wild+Ptxl, L215H+Ptxl, L217R+Ptxl and L225M+Ptxl were prepared, and 50 ns MD simulation was performed for each system. Binding energy estimation indicated that leucine mutation reduces the binding affinity of Ptxl in mutant types (MTs) as compared to wild type (WT). Further, in contrast to WT Ptxl interactions with the M-loop (PHE270-VAL286), S6-S7 loop and H9-H10 were significantly altered in MTs. Results showed that in MTs, Ptxl had weak interaction with M-loop residues, while having strong affinity with S6-S7 loop and H6-H7 loop. Moreover, PCA and FEL analysis revealed that M-loop flexible region (THR274-LEU284) was strongly bound with Ptxl in WT preventing its flexible movement and the causing factor for microtubule stabilization. In MTs due to poor interaction with Ptxl, M-loop flexible region retains its flexibility, therefore unable to stabilize microtubule. This study will give an insight into the importance of M-loop flexible region interaction with Ptxl for microtubule stabilization. In addition, it clearly provides the molecular basis of Ptxl resistance mechanism in leucine MTs. This work will help in developing novel microtubule stabilizers molecules active against MTs.
authors.affiliation: Biotechnology Division, CSIR-Central Institute of Medicinal and Aromatic Plants (CSIR-CIMAP), Kukrail Picnic Spot Road, P.O. CIMAP, Lucknow 226015, India.
authors.forename: Shubhandra
authors.initials: S
authors.lastname: Tripathi
authors.affiliation: Biotechnology Division, CSIR-Central Institute of Medicinal and Aromatic Plants (CSIR-CIMAP), Kukrail Picnic Spot Road, P.O. CIMAP, Lucknow 226015, India.
authors.forename: Gaurava
authors.initials: G
authors.lastname: Srivastava
authors.affiliation: Biotechnology Division, CSIR-Central Institute of Medicinal and Aromatic Plants (CSIR-CIMAP), Kukrail Picnic Spot Road, P.O. CIMAP, Lucknow 226015, India. Electronic address: ashoksharma@cimap.res.in.
authors.forename: Ashok
authors.initials: A
authors.lastname: Sharma
mesh_terms: D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D019008:Drug Resistance, Neoplasm; D006801:Humans; D007930:Leucine / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008870:Microtubules / Q000737:chemistry / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D009369:Neoplasms / Q000188:drug therapy / Q000235:genetics; D017239:Paclitaxel / Q000494:pharmacology*; D017354:Point Mutation; D013816:Thermodynamics; D014404:Tubulin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D050257:Tubulin Modulators / Q000494:pharmacology*
chemical_list: D000972:Antineoplastic Agents, Phytogenic; C578986:TUBB1 protein, human; D014404:Tubulin; D050257:Tubulin Modulators; D007930:Leucine; D017239:Paclitaxel
keywords: Drug resistance; Free energy landscape; Microtubule; Molecular dynamics simulation; Paclitaxel; βI-tubulin