title: A DC-81-indole conjugate agent suppresses melanoma A375 cell migration partially via interrupting VEGF production and stromal cell-derived factor-1α-mediated signaling.
abstract: Pyrrolo[2,1-c][1,4]benzodiazepine (PBD) chemicals are antitumor antibiotics inhibiting nucleic acid synthesis. An indole carboxylate-PBD hybrid with six-carbon spacer structure (IN6CPBD) has been previously demonstrated to induce melanoma cell apoptosis and reduce metastasis in mouse lungs. This study aimed at investigating the efficacy of the other hybrid compound with four-carbon spacer (IN4CPBD) and elucidating its anti-metastatic mechanism. Human melanoma A375 cells with IN4CPBD treatment underwent cytotoxicity and apoptosis-associated assays. Transwell migration assay, Western blotting, and ELISA were used for mechanistic study. IN4CPBD exhibited potent melanoma cytotoxicity through interrupting G1/S cell cycle progression, increasing DNA fragmentation and hypodipoidic DNA contents, and reducing mitochondrial membrane potential. Caspase activity elevation suggested that both intrinsic and extrinsic pathways were involved in IN4CPBD-induced melanoma apoptosis. IN4CPBD up-regulated p53 and p21, thereby concomitantly derailing the equilibrium between Bcl-2 and Bax levels. Transwell migration assay demonstrated that stromal cell-derived factor-1α (SDF-1α) stimulated A375 cell motility, while kinase inhibitors treatment confirmed that Rho/ROCK, Akt, ERK1/2, and p38 MAPK pathways were involved in SDF-1α-enhanced melanoma migration. IN4CPBD not only abolished the SDF-1α-enhanced chemotactic motility but also suppressed constitutive MMP-9 and VEGF expression. Mechanistically, IN4CPBD down-regulated Akt, ERK1/2, and p38 MAPK total proteins and MYPT1 phosphorylation. In conclusion, beyond the fact that IN4CPBD induces melanoma cell apoptosis at cytotoxic dose, the interruption in the VEGF expression and the SDF-1α-related signaling at cytostatic dose may partially constitute the rationale for its in vivo anti-metastatic potency.
authors.affiliation: Graduate Institute of Pharmacy, Faculty of Pharmacy, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: Ming-Chu
authors.initials: MC
authors.lastname: Hsieh
authors.forename: Wan-Ping
authors.initials: WP
authors.lastname: Hu
authors.forename: Hsin-Su
authors.initials: HS
authors.lastname: Yu
authors.forename: Wen-Chuan
authors.initials: WC
authors.lastname: Wu
authors.forename: Long-Sen
authors.initials: LS
authors.lastname: Chang
authors.forename: Ying-Hsien
authors.initials: YH
authors.lastname: Kao
authors.forename: Jeh-Jeng
authors.initials: JJ
authors.lastname: Wang
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D001569:Benzodiazepines / Q000494:pharmacology*; D015153:Blotting, Western; D020169:Caspases / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects*; D054377:Chemokine CXCL12 / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D008545:Melanoma / Q000188:drug therapy* / Q000473:pathology; D009943:Organophosphorus Compounds / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011758:Pyrroles / Q000494:pharmacology* / Q000627:therapeutic use; D015398:Signal Transduction / Q000187:drug effects; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis
chemical_list: C474072:(11aS)-8-hydroxy-7-methoxy-1,2,3,11a-tetrahydro-5H-pyrrolo(2,1-c)(1,4)benzodiazepin-5-one; D000970:Antineoplastic Agents; D054377:Chemokine CXCL12; D007211:Indoles; D009943:Organophosphorus Compounds; D047428:Protein Kinase Inhibitors; D011758:Pyrroles; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D001569:Benzodiazepines; D020169:Caspases