KRas4B-PDE6d complex stabilization by small molecules obtained by virtual screening affects Ras signaling in pancreatic cancer 06 Biological Sciences 0601 Biochemistry and Cell Biology
Por:
Casique-Aguirre D., Briseño-Díaz P., García-Gutiérrez P., La Rosa C.H.G.-D., Quintero-Barceinas R.S., Rojo-Domínguez A., Vergara I., Medina L.A., Correa-Basurto J., Bello M., Hernández-Rivas R., Del Rociothompson-Bonilla M., Vargas M.
Publicada:
1 ene 2018
Resumen:
Background: The GTPase KRas4B has been utilized as a principal target in the development of anticancer drugs. PDE6d transports KRas4B to the plasma membrane, where it is released to activate various signaling pathways required for the initiation and maintenance of cancer. Therefore, identifying new small molecules that prevent activation of this GTPase by stabilizing the KRas4B-PDE6d molecular complex is a practical strategy to fight against cancer. Methods: The crystal structure of the KRas4B-PDE6d heterodimer was employed to locate possible specific binding sites at the protein-protein interface region. Virtual screening of Enamine-database compounds was performed on the located potential binding sites to identify ligands able to simultaneously bind to the KRas4B-PDE6d heterodimer. A molecular dynamics approach was used to estimate the binding free-energy of the complex. Cell viability and apoptosis were measured by flow cytometry. G-LISA was used to measure Ras inactivation. Western blot was used to measure AKT and ERK activation. MIA PaCa-2 cells implanted subcutaneously into nude mice were treated with D14 or C22 and tumor volumes were recorded. Results: According to the binding affinity estimation, D14 and C22 stabilized the protein-protein interaction in the KRas4B-PDE6d complex based on in vitro evaluation of the 38 compounds showing antineoplastic activity against pancreatic MIA PaCa-2 cancer cells. In this work, we further investigated the antineoplastic cellular properties of two of them, termed D14 and C22, which reduced the viability in the human pancreatic cancer cells lines MIA PaCa-2, PanC-1 and BxPC-3, but not in the normal pancreatic cell line hTERT-HPNE. Compounds D14 and C22 induced cellular death via apoptosis. D14 and C22 significantly decreased Ras-GTP activity by 33% in MIA PaCa-2 cells. Moreover, D14 decreased AKT phosphorylation by 70% and ERK phosphorylation by 51%, while compound C22 reduced AKT phosphorylation by 60% and ERK phosphorylation by 36%. In addition, compounds C22 and D14 significantly reduced tumor growth by 88.6 and 65.9%, respectively, in a mouse xenograft model. Conclusions: We identified two promising compounds, D14 and C22, that might be useful as therapeutic drugs for pancreatic ductal adenocarcinoma treatment. © 2018 The Author(s).
Filiaciones:
Departamento de Biomedicina Molecular, Centro de Investigación y de Estudios Avanzados Del Instituto Politécnico Nacional, Av. I.P.N, México City, 2508, Mexico
Departamento de Química, Universidad Autónoma Metropolitana, Unidad Iztapalapa, México City, Mexico
Departamento de Ciencias Naturales, Universidad Autónoma Metropolitana, Unidad Cuajimalpa, México City, Mexico
Instituto Nacional de Cancerología, Unidad de Investigación Biomédica en Cáncer, México City, Mexico
Laboratorio de Modelado Molecular y Diseño, Fármacos de la Escuela Superior de Medicina, Instituto Politécnico Nacional, México City, Mexico
Investigación Biomédica y Trasnacional, Laboratorio de Medicina Genómica, Hospital 1 de Octubre, ISSSTE, México City, Mexico
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