title: The combi-targeting concept: a novel 3,3-disubstituted nitrosourea with EGFR tyrosine kinase inhibitory properties.
abstract: OBJECTIVE
  To study the dual mechanism of action of FD137, a 3,3-disubstituted nitrosourea designed to block signaling mediated by the epidermal growth factor receptor (EGFR) on its own and to be hydrolyzed to an inhibitor of EGFR plus a DNA-damaging species.
  
  
  METHODS
  HPLC was used to determine the half-life (t(1/2)) of FD137 and to characterize its derived metabolite FD110. The dual mechanisms of DNA damaging and EGFR tyrosine kinase (TK) targeting were ascertained by the comet assay for DNA damage and by inmunodetection of phosphotyrosine in an ELISA and a whole-cell assay for EGFR-mediated signaling. The antiproliferative effects of the different drugs and their combinations were determined by the sulforhodamine B (SRB) assay.
  
  
  RESULTS
  In contrast to BCNU, FD137 significantly blocked EGF-induced EGFR autophosphorylation (IC(50) 4 micro M) in the human solid tumor cell line A431. DNA damage induced by FD137 could only be observed after 24 h exposure, but the level of DNA damage remained 3.6-fold lower than that induced by BCNU. This difference was rationalized by the 160-fold greater stability of FD137 when compared with BCNU in serum-containing medium. Further, degradation of FD137 was accompanied by the slow release of FD110, an extremely potent inhibitor of EGFR TK [IC(50) (EGFR autophosphorylation) <0.3 micro M]. The complex properties of FD137 translated into a 55-fold greater antiproliferative activity than BCNU against the EGFR-overexpressing A431 cells that coexpresses the O(6)-alkylguanine transferase (AGT). Depletion of AGT in these cells by the use of O(6)-benzylguanine (O(6)-BG) enhanced their sensitivity to BCNU by 8-fold, but only by 3-fold to FD137.
  
  
  CONCLUSIONS
  The results overall suggest that the superior antiproliferative activity of FD137 when compared with BCNU may be associated with its ability to behave as a combination of many species with different mechanisms of action. However, the enhancement of its potency by O(6)-BG suggests that its antiproliferative effect was at least partially mitigated by AGT and perhaps it may be largely dominated by its signal transduction inhibitory component.
authors.affiliation: Cancer Drug Research Laboratory, Department of Medicine, Division of Medical Oncology, McGill University Health Center/Royal Victoria Hospital, 687 Pine Avenue West, Montreal, Quebec, H3A 1A1, Canada.
authors.forename: Qiyu
authors.initials: Q
authors.lastname: Qiu
authors.forename: Fabienne
authors.initials: F
authors.lastname: Dudouit
authors.forename: Stephanie L
authors.initials: SL
authors.lastname: Matheson
authors.forename: Fouad
authors.initials: F
authors.lastname: Brahimi
authors.forename: Ranjita
authors.initials: R
authors.lastname: Banerjee
authors.forename: James P
authors.initials: JP
authors.lastname: McNamee
authors.forename: Bertrand J
authors.initials: BJ
authors.lastname: Jean-Claude
mesh_terms: D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D004249:DNA Damage; D066246:ErbB Receptors / Q000037:antagonists & inhibitors*; D006801:Humans; D009607:Nitrosourea Compounds / Q000494:pharmacology*; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000502:physiology; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases; D014407:Tumor Cells, Cultured
chemical_list: D018906:Antineoplastic Agents, Alkylating; C471268:FD137 nitrosourea; D009607:Nitrosourea Compounds; D019853:O(6)-Methylguanine-DNA Methyltransferase; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases