title: Potentiation of BCNU antitumor efficacy by 9-substituted O6-benzylguanines. Effect of metabolism.
abstract: OBJECTIVE
  O6-Benzylguanine (BG), an O6-methylguanine-DNA methyltransferase (MGMT) inactivator, potentiates the efficacy of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and of other DNA chloroethylating and methylating anticancer drugs and is currently undergoing clinical trials. O6-Benzyl-2'-deoxyguanosine (dBG), a less effective MGMT inactivator than BG in vitro, is at least as effective as BG in combination with BCNU against tumor xenografts in athymic mice. In order to identify the mechanism of dBG activation in in vivo systems we tested the metabolism, ability to inactivate MGMT, and efficacy to potentiate BCNU in vivo of two additional 9-substituted derivatives of BG, namely O6-benzyl-9-cyanomethylguanine (CMBG) and O6-benzylguanosine (BGS).
  
  
  METHODS
  Metabolism and disposition of these drugs was examined in athymic mice and Sprague-Dawley rats. MGMT suppression was determined in human medulloblastoma (Daoy) tumor xenografts in athymic mice following treatment with BGS, dBG, and CMBG and was compared with the loss of resistance to BCNU as determined by tumor growth delays.
  
  
  RESULTS
  Growth delays at 25 mg/m2 BCNU and 133 mg/m2 BG or equimolar doses of CMBG, BGS or dBG were 23.0, 2.5, 21.3 days. and 30.4 days, respectively. The above differences did not correlate with the ED50S of 0.2, 13, 11 microM, and 2 microM determined for the above compounds, respectively, in cell free extracts. Differences in the efficacies of the 9-substituted compounds did correlate, however, with the extent of their metabolic conversion to BG. The maximum concentrations of BG in blood achieved after the administration of equimolar (250 micromol/kg) doses of CMBG, BGS and dBG were 10, 30 microM, and 55 microM, respectively. Although such levels were lower than those achieved in circulation by administration of an equimolar amount of BG, BG levels persisted longer following treatments with BGS or dBG than after treatment with BG itself. Formation of BG was required for continuous and prolonged (> 16 h) suppression of MGMT activity to non-detectable levels (< 5 fmol/mg protein).
  
  
  CONCLUSIONS
  Metabolism of BGS and dBG to BG explains the unexpected high efficacy of these compounds in potentiating the antitumor activity of BCNU in the athymic mouse model. The faster and more effective suppression of tumor MGMT by dBG and its greater efficacy, as compared with BGS, also correlates with a more rapid accumulation of BG in blood after dBG than after BGS administration, which results in faster and complete suppression of MGMT in Daov xenografts. Thus, metabolism of dBG and BGS to BG appears to be the determining factor for continuous and prolonged suppression of MGMT activity, and that near complete suppression of such activity during and following BCNU administration is required for the higher efficacy of treatments. Similarly, the failure of CMBG to suppress tumor MGMT to the same extent as BGS, in spite of their similar ED50 values, could be attributed to the metabolism of this compound mainly by pathways other than conversion to BG.
authors.affiliation: Department of Neurosurgery, University of Texas Southwestern Medical Center, Dallas 75235-8855, USA.
authors.forename: D M
authors.initials: DM
authors.lastname: Kokkinakis
authors.forename: R C
authors.initials: RC
authors.lastname: Moschel
authors.forename: A E
authors.initials: AE
authors.lastname: Pegg
authors.forename: S C
authors.initials: SC
authors.lastname: Schold
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001921:Brain / Q000378:metabolism; D002330:Carmustine / Q000494:pharmacology*; D004357:Drug Synergism; D006147:Guanine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000037:antagonists & inhibitors; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D000970:Antineoplastic Agents; C064976:O(6)-benzylguanine; D006147:Guanine; D019853:O(6)-Methylguanine-DNA Methyltransferase; D002330:Carmustine