KEY SYNTHETIC ANALOGS OF BLEOMYCIN A(2) THAT DIRECTLY ADDRESS THE EFFECT AND ROLE OF THE DISACCHARIDE - DEMANNOSYLBLEOMYCIN A(2) AND ALPHA-D-MANNOPYRANOSYLDEGLYCOBLEOMYCIN A(2)

Two key synthetic analogs of bleomycin A(2) (1) are described. The synthesis and evaluation of demannosylbleomycin A(2) (3) lacking the terminal 2-O-(3-O-carbamoyl)-alpha-D-mannopyranoside including the putative carbamoyl sixth metal ligand and the monosaccharide 4 in which an alpha-D-mannopyranose has been substituted for the authentic alpha-L-gulopyranose with the inversion of the single C5 center of the monosaccharide are detailed. Both agents were prepared through diastereoselective O-glycosidation of N-alpha-CBZ-N-pi-trityl-beta-hydroxy-L-histidine methyl ester (11) with the diphenyl phosphate of 3,4,6-tri-O-acetyl-2-O-methyl-beta-L-gulopyranose (10) and 2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranose (24), respectively, followed by sequential tetrapeptide S and pyrimidoblamic acid couplings. The former proceeded with clean inversion of the glycosyl C1 stereochemistry to provide 12 (greater than or equal to 20:1 alpha:beta, 72%) while the latter proceeded with net retention of the glycosyl C1 stereochemistry to provide 26. Both the low reactivity of the glycosyl acceptor 11 which favors formation of the more stable alpha-anomer and mechanistic features characteristic of a glycosyl phosphate donor which favor reaction with inversion of the stereochemistry at the reacting anomeric center account for the clean generation of 12 while the mannosylpyranose C2 acetate neighboring group participation in the glycosidation reaction of 24 accounts for the clean generation of 26. The observation that the DNA cleavage efficiency, selectivity (5'-GC, 5'-GT > 5'-GA), and the double strand versus single strand DNA cleavage ratio of demannosylbleomycin A(2) (3) are similar or nearly indistinguishable from bleomycin A(2) (1) indicate that the terminal 2-O-(3-O-carbamoyl)-alpha-D-mannopyranoside inclusive of the carbamoyl group may have little impact on the DNA cleavage capabilities of the natural agent. In contrast, the substantially diminished DNA cleavage efficiency and ratio of double strand to single strand cleavage events of 4 relative to bleomycin A(2) (1), demannosylbleomycin A(2) (3), or even deglycobleomycin A(2) (2) indicate that the first carbohydrate of the disaccharide may greatly influence the DNA cleavage capabilities of the natural agent. Importantly, the studies indicate that it is the alpha-L-gulopyranoside segment, not the alpha-D-mannopyranoside segment inclusive of the carbamoyl group, of the disaccharide that permits the full potentiation of the DNA cleavage properties of bleomycin A(2).