Molecular Docking and Molecular Dynamics Studies Reveal Secretory Proteins as Novel Targets of Temozolomide in Glioblastoma Multiforme.
Cancer Genetics and Therapeutics
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Primary Categories:
- Cancer
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Secondary Categories:
- Cancer
Introduction:
Glioblastoma multiforme (GBM) is the most prevalent malignant primary brain tumor, with an incidence of 3.2 per 100,000, predominantly affecting men and occurring at a median age of 64. GBM can be primary (arising spontaneously) or secondary (progressing from a lower-grade tumor). Standard treatment involves surgery, followed by Temozolomide (TMZ) and radiation. Limitations in diagnostic imaging and tissue biopsies have driven interest in non-invasive liquid biopsies, which detect circulating tumor biomarkers in blood and CSF. This study highlights brain-enriched secretory proteins, such as CNTN2, NPTX1, OPCML, and SLIT1, involved in GBM pathways. CNTN2 and NPTX1 enhance tumor proliferation via pathways like PI3K/AKT, while downregulated proteins (e.g., OPCML) show potential diagnostic and therapeutic value. Molecular docking and simulations were employed to investigate the binding of these proteins with TMZ, highlighting their significance in GBM pathogenesis and potential as therapeutic targets.
Methods:
The 3D structure of Temozolomide (CID:5394) was obtained from PubChem in SDF format and converted to MOL2 using Open Babel. Protein structures were chosen based on their FASTA sequences retrieved from UniProt, analyzed via PDB BLAST for high identity matches. Structures with over 97% identity, such as contactin 2 (PDB ID: 2OM5) and SERPINI1 (PDB ID: 3F5N), were downloaded from the RCSB Protein Data Bank and cleaned of duplicates and nonstandard residues using BIOVIA Discovery Studio.
For proteins lacking PDB structures or with lower identity, Phyre2 modeling was used to generate 3D structures, selecting top-confidence models. Pocket identification was performed using DoGSiteScorer to find high drug score pockets, guiding grid coordinates for docking.
Molecular docking was conducted with AutoDock 4.2, applying polar hydrogen and Kollman charges. Twenty runs were executed, and the best binding poses were visualized with PyMol and BIOVIA Discovery Studio.
MD simulations for protein–TMZ complexes were analyzed using CABS-flex (10 ns) for RMSF and structural flexibility, while iMODS provided insight into stability, molecular motion, deformability, B-factors, eigenvalues, variance, and covariance mapping. This confirmed the stability and dynamics of TMZ-protein interactions, supporting their therapeutic potential in GBM.
Results:
Molecular docking was employed to predict interactions between Temozolomide (TMZ) and nine GBM-related secretory proteins. SLIT1 showed the highest binding affinity with TMZ, despite lacking hydrogen bonds, due to interactions enhancing binding energy. GDF1 followed with high affinity, supported by hydrogen bonds. Both proteins were stable in docked complexes, highlighting their potential as novel TMZ targets. NPTX1 and SERPINI1 also demonstrated notable binding, with NPTX1 showing greater overall interaction. CREG2 showed non-hydrogen binding interactions, while OPCML and LGI1 had moderate binding with significant hydrogen bonds, warranting further exploration.
CNTN2 and LY6H, although having lower TMZ affinity, formed numerous hydrogen bonds, indicating stable interactions. RMSF analysis using CABS-flex revealed significant flexibility in SERPINI1, SLIT1, GDF1, and LY6H. iMODS analysis confirmed structural flexibility, with all proteins showing peaks of deformability and low eigenvalues, indicating ease of structural adaptation. B-factor analysis supported the flexibility and mobility of TMZ-bound proteins. Variance and covariance analyses revealed reasonable stability, with specific correlations in complex movements.
These findings emphasize the potential of SLIT1, GDF1, NPTX1, and other proteins as promising targets for TMZ therapy in GBM, reinforcing drug-target interactions that could enhance therapeutic strategies.
Conclusion:
Molecular docking and dynamics simulation studies provide a better understanding of the intermolecular-level interactions of TMZ with the circulating/secretory proteins involved in GBM pathogenesis. The present findings provide substantial evidence that these proteins are potential targets of TMZ, highlighting a novel aspect of TMZ’s therapeutic potential. However, extensive in vivo and in vitro studies are warranted to decipher the exact molecular mechanism and mode of action of TMZ by targeting these potential secretory proteins involved in various cellular and molecular pathways associated with GBM.
Glioblastoma multiforme (GBM) is the most prevalent malignant primary brain tumor, with an incidence of 3.2 per 100,000, predominantly affecting men and occurring at a median age of 64. GBM can be primary (arising spontaneously) or secondary (progressing from a lower-grade tumor). Standard treatment involves surgery, followed by Temozolomide (TMZ) and radiation. Limitations in diagnostic imaging and tissue biopsies have driven interest in non-invasive liquid biopsies, which detect circulating tumor biomarkers in blood and CSF. This study highlights brain-enriched secretory proteins, such as CNTN2, NPTX1, OPCML, and SLIT1, involved in GBM pathways. CNTN2 and NPTX1 enhance tumor proliferation via pathways like PI3K/AKT, while downregulated proteins (e.g., OPCML) show potential diagnostic and therapeutic value. Molecular docking and simulations were employed to investigate the binding of these proteins with TMZ, highlighting their significance in GBM pathogenesis and potential as therapeutic targets.
Methods:
The 3D structure of Temozolomide (CID:5394) was obtained from PubChem in SDF format and converted to MOL2 using Open Babel. Protein structures were chosen based on their FASTA sequences retrieved from UniProt, analyzed via PDB BLAST for high identity matches. Structures with over 97% identity, such as contactin 2 (PDB ID: 2OM5) and SERPINI1 (PDB ID: 3F5N), were downloaded from the RCSB Protein Data Bank and cleaned of duplicates and nonstandard residues using BIOVIA Discovery Studio.
For proteins lacking PDB structures or with lower identity, Phyre2 modeling was used to generate 3D structures, selecting top-confidence models. Pocket identification was performed using DoGSiteScorer to find high drug score pockets, guiding grid coordinates for docking.
Molecular docking was conducted with AutoDock 4.2, applying polar hydrogen and Kollman charges. Twenty runs were executed, and the best binding poses were visualized with PyMol and BIOVIA Discovery Studio.
MD simulations for protein–TMZ complexes were analyzed using CABS-flex (10 ns) for RMSF and structural flexibility, while iMODS provided insight into stability, molecular motion, deformability, B-factors, eigenvalues, variance, and covariance mapping. This confirmed the stability and dynamics of TMZ-protein interactions, supporting their therapeutic potential in GBM.
Results:
Molecular docking was employed to predict interactions between Temozolomide (TMZ) and nine GBM-related secretory proteins. SLIT1 showed the highest binding affinity with TMZ, despite lacking hydrogen bonds, due to interactions enhancing binding energy. GDF1 followed with high affinity, supported by hydrogen bonds. Both proteins were stable in docked complexes, highlighting their potential as novel TMZ targets. NPTX1 and SERPINI1 also demonstrated notable binding, with NPTX1 showing greater overall interaction. CREG2 showed non-hydrogen binding interactions, while OPCML and LGI1 had moderate binding with significant hydrogen bonds, warranting further exploration.
CNTN2 and LY6H, although having lower TMZ affinity, formed numerous hydrogen bonds, indicating stable interactions. RMSF analysis using CABS-flex revealed significant flexibility in SERPINI1, SLIT1, GDF1, and LY6H. iMODS analysis confirmed structural flexibility, with all proteins showing peaks of deformability and low eigenvalues, indicating ease of structural adaptation. B-factor analysis supported the flexibility and mobility of TMZ-bound proteins. Variance and covariance analyses revealed reasonable stability, with specific correlations in complex movements.
These findings emphasize the potential of SLIT1, GDF1, NPTX1, and other proteins as promising targets for TMZ therapy in GBM, reinforcing drug-target interactions that could enhance therapeutic strategies.
Conclusion:
Molecular docking and dynamics simulation studies provide a better understanding of the intermolecular-level interactions of TMZ with the circulating/secretory proteins involved in GBM pathogenesis. The present findings provide substantial evidence that these proteins are potential targets of TMZ, highlighting a novel aspect of TMZ’s therapeutic potential. However, extensive in vivo and in vitro studies are warranted to decipher the exact molecular mechanism and mode of action of TMZ by targeting these potential secretory proteins involved in various cellular and molecular pathways associated with GBM.