peloruside-a and Carcinoma

Microtubules (MTs) play an essential role in mitosis; hence they are identified as potential targets to design novel anti-mitotic agents. MT's are composed of α/β-tubulin isotypes that are associated with differential drug-resistant effects against MT-targeting agents. Peloruside-A (PLA) is a potent anti-mitotic agent showing excellent activity against taxol-resistant carcinoma. PLA alters MT dynamics by binding to the 'non-taxoid' site of β-tubulin. The abundance of βII and βIII tubulin isotypes in human ovarian carcinoma affects the efficacy of PLA. Nevertheless, the mechanism of PLA resistance due to βII and βIII tubulin isotype is not well understood. Therefore, we investigated the interactions of PLA with αβIIa, αβIIb, and αβIII tubulin isotypes which are predominantly expressed in the human ovarian carcinoma, using a molecular modeling approach. A sequence analysis study shows that the βIII isotype has seven residue variations at the 'non-taxoid' site compared to the βIIa and βIIb isotypes. Molecular docking and molecular dynamics simulation revealed that residue variation at the 'non-taxoid' site of βIII isotype affect PLA binding. Furthermore, binding energy calculations showed that αβIIa has the highest binding towards PLA, whereas αβIIb and αβIII isotypes shows weaker associations with PLA. Our computational study provides valuable structural and energetic information to increase understanding into the origin of PLA resistance in human ovarian carcinoma and could be helpful to develop potential PLA analogs against specific β-tubulin isotypes expressed in cancer cells.Communicated by Ramaswamy H. Sarma.

Topics: Antibiotics, Antineoplastic; Bridged Bicyclo Compounds, Heterocyclic; Carcinoma; Drug Resistance, Neoplasm; Female; Humans; Lactones; Microtubules; Molecular Docking Simulation; Ovarian Neoplasms; Protein Binding; Tubulin

Acquired β-tubulin alterations in human ovarian carcinoma 1A9 cells were previously shown to confer resistance to the microtubule stabilizing agents peloruside A (PLA) and laulimalide (LAU). We examined the proteome of resistant cells to see what other protein changes occurred as a result of the acquired drug resistance.. Two-dimensional differential in-gel electrophoresis was performed to explore differentially expressed proteins in the resistant 1A9-R1 (R1) and 1A9-L4 (L4) cells. The proteins on the gels were identified by MALDI-TOF MS, and altered protein abundance was confirmed by Western blotting and immunocytochemistry. Vimentin expression was restored in vimentin-deficient L4 cells by transfecting a full-length human vimentin cDNA, and sensitivity to PLA and LAU were tested using an MTT cell proliferation assay.. Proteomic analysis identified several proteins that were significantly altered in the resistant cells relative to the parental 1A9 cells. Using Western blotting and immunocytochemistry, a decreased vimentin abundance in the L4 cells was validated. Vimentin levels were unchanged in PLA-resistant R1 cells and paclitaxel/epothilone-resistant derivatives of 1A9 cells. Vimentin cDNA transfection into L4 cells partially restored PLA and LAU sensitivity.. Downregulation of vimentin contributes to the resistance of 1A9 cells to the microtubule stabilizing agents, PLA and LAU.

Topics: Antineoplastic Agents; Bridged Bicyclo Compounds, Heterocyclic; Carcinoma; Cell Line, Tumor; DNA, Complementary; Down-Regulation; Drug Resistance, Neoplasm; Female; Gene Expression Regulation, Neoplastic; Humans; Lactones; Macrolides; Microtubules; Ovarian Neoplasms; Proteomics; Transfection; Vimentin