title: A synthetic polyvalent ligand for α5β1 integrin activates components of the urothelial carcinoma cell response to bacillus Calmette-Guérin.
abstract: OBJECTIVE
  Prior study has shown that bacillus Calmette-Guérin binds to and cross-links α5β1 integrins present on the surface of urothelial carcinoma cells. Antibody mediated cross-linking of α5β1 integrins can reproduce signal transduction, gene transactivation and phenotypic changes, similar to those observed in response to bacillus Calmette-Guérin. We evaluated the effect of a synthetic polyvalent ligand for α5β1 on these elements of the tumor response to bacillus Calmette-Guérin.
  
  
  METHODS
  The consensus α5β1 integrin binding tripeptide RGD was linked to a MAP8 backbone to result in an octavalent construct targeting α5β1 integrin. RGD-MAP8 was used to determine its effect on signaling pathway activation (nuclear factor-κB, NRF2 and CEBP), gene expression (p21, interleukin-6 and 8, CXCL1, CXCL2 and CCL20) and cytotoxicity (trypan blue exclusion and HMGB1 release) in human urothelial carcinoma cells. Results were compared to those of treatment with bacillus Calmette-Guérin or the missense peptide GRD-MAP8.
  
  
  RESULTS
  The RDG-MAP8 construct significantly increased nuclear factor-κB signaling and p21 expression relative to controls. Compared to bacillus Calmette-Guérin treatment, only p21 expression was comparable for cells treated with RGD-MAP8, averaging 70% of bacillus Calmette-Guérin induced expression. RGD-MAP8 failed to have a significant effect on CEBP or NRF2 activation, gene expression or cell viability.
  
  
  CONCLUSIONS
  Intracellular signaling, gene transactivation and phenotypic changes in response to RGD-MAP8 were qualitatively and quantitatively different than those observed in response to bacillus Calmette-Guérin. Results suggest that while α5β1 integrin cross-linking contributes to the bacillus Calmette-Guérin response, it alone is insufficient to duplicate the full spectrum of bacillus Calmette-Guérin induced changes in urothelial carcinoma cell biology.
authors.affiliation: Department of Urology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
authors.forename: Fanghong
authors.initials: F
authors.lastname: Chen
authors.forename: Guangjian
authors.initials: G
authors.lastname: Zhang
authors.forename: Yanli
authors.initials: Y
authors.lastname: Cao
authors.forename: Bassam
authors.initials: B
authors.lastname: Wakim
authors.forename: William A
authors.initials: WA
authors.lastname: See
mesh_terms: D000276:Adjuvants, Immunologic / Q000494:pharmacology; D001500:BCG Vaccine / Q000494:pharmacology*; D002295:Carcinoma, Transitional Cell / Q000188:drug therapy / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D004273:DNA, Neoplasm / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D039081:Integrin alpha5beta1 / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D001749:Urinary Bladder Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000378:metabolism
chemical_list: D000276:Adjuvants, Immunologic; D001500:BCG Vaccine; D004273:DNA, Neoplasm; D039081:Integrin alpha5beta1