title: Biochemical modulation of iododeoxyuridine by N6-[4-(morpholinosulfonyl)benzyl]-N6-methyl-2,6-diaminobenz[cd]indole glucuronate (AG-331) leading to enhanced cytotoxicity.
abstract: Inhibition of thymidylate synthase (TS) may increase incorporation of thymidine analogues into DNA, leading to increased inhibition of colony formation in tumor cells. We have reported previously that TS inhibition by N-(5-[N-(3,4-dihydro-2-methyl-4-oxoquinazolin-6,-ylmethyl)-N -methylamino]-2 - thenoyl)-L-glutamic acid (ICI D1694 or Tomudex), a folate-based TS inhibitor, increases the cytotoxicity of iododeoxyuridine (IdUrd), a thymidine analogue, in MGH-U1 human bladder and HCT-8 human colon cancer cells. N6-[4-(Morpholinosulfonyl)benzyl]-N6-methyl-2,6-diaminobenz[ cd]-indole glucuronate (AG-331) differs from ICI D1694 in that it is a de novo designed lipophilic TS inhibitor, it does not require a specific carrier for cellular uptake, and it does not undergo intracellular polyglutamation. Exposure of MGH-U1 cells to 5 microM AG-331 for 24 hr decreased clonogenic survival by 30%, but almost completely inhibited TS activity. IdUrd is a cytotoxic thymidine analogue, with IC50 and IC90 values after 24-hr exposures in MGH-U1 cells of 13 and 81 microM, respectively. The combination of IdUrd and AG-331 resulted in an enhanced antitumor effect, as compared with the effect of either agent alone. The cytotoxic IC50 of IdUrd decreased from 13 to 1.5 microM, and the IC90 decreased from 81 to 5 microM with the addition of 5 microM AG-331. Biochemical studies of the combination revealed that pretreating MGH-U1 cells with 5 microM AG-331 increased IdUrd incorporation into cellular DNA by 3.8-fold. This increased incorporation was associated with a greater proportion of DNA single-strand breaks than observed with either agent alone, and the combination of 5 microM AG-331 plus IdUrd produced up to a 2.5-fold increase in DNA single-strand breaks as compared with IdUrd alone. The effects of AG-331, IdUrd, and the combination of IdUrd and AG-331 on the colony-forming ability of normal human bone marrow CFU-GM cells was determined as a measure of myelosuppression. The combination of IdUrd and AG-331, at the same concentrations as those used in the MGH-U1 cells, produced a wider therapeutic index relative to that of IdUrd alone, and the therapeutic index for the combination was 6.5, as compared with 4.0 for IdUrd plus ICI D1694 in previous studies from this laboratory. These observations suggest that the combination of IdUrd and AG-331 may enhance antitumor effects with minimal myelosuppression in vivo.
authors.affiliation: Division of Experimental Therapeutics, Ontario Cancer Institute, Toronto, Canada.
authors.forename: J
authors.initials: J
authors.lastname: Pressacco
authors.forename: B
authors.initials: B
authors.lastname: Mitrovski
authors.forename: D W
authors.initials: DW
authors.lastname: Hedley
authors.forename: R
authors.initials: R
authors.lastname: Tsang
authors.forename: C
authors.initials: C
authors.lastname: Erlichman
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D001853:Bone Marrow / Q000187:drug effects; D001854:Bone Marrow Cells; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D004357:Drug Synergism; D006801:Humans; D007065:Idoxuridine / Q000378:metabolism / Q000494:pharmacology*; D007211:Indoles / Q000494:pharmacology*; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors*; D014407:Tumor Cells, Cultured
chemical_list: D000964:Antimetabolites, Antineoplastic; D007211:Indoles; C087748:AG 331; D004247:DNA; D013940:Thymidylate Synthase; D007065:Idoxuridine