Two analogs of
N-acetylmannosamine, 2-acetamido-1,3,4,6-tetra-O-acetyl-2-deoxy-D-mannopyranose (Ac4-NAcMan) and the 2-trifluoroacetamido derivative (AC4F3-NAcMan), were synthesized as potential inhibitors of the formation of
sialic acid-containing
glycoconjugates and were examined for their ability to modify the incorporation of N-[3H]acetylamannosamine into cellular
glycoconjugates of Friend murine
erythroleukemia cells.
Ac4F3-NAcMan and
Ac4-NAcMan inhibited cellular replication in
suspension culture at concentrations of 0.02 and 0.08 mM, respectively. The cytotoxicity of
Ac4-NAcMan was relatively reversible, whereas that produced by Ac4-F3-NAcMan was not, as judged by measurement of the cloning efficiencies of cells exposed to these agents. The analogs inhibited incorporation of N-[3H]acetylmannosamine into
ethanol-soluble and -insoluble materials. Separation of
ethanol-soluble metabolites by HPLC demonstrated that
Ac4F3-NAcMan caused accumulation of radioactivity from N-[3H]acetylmannosamine in
CMP-N-acetylneuraminic acid (
CMP-NeuNAc) equal to the decrease in macromolecular-bound 3H caused by this agent. In contrast, similar exposure to
Ac4-NAcMan produced a large increase in the amount of radioactivity in
ethanol-soluble
N-acetylneuraminic acid while decreasing the amount of label from N-[3H]acetylmannosamine in cellular
CMP-NeuNAc, suggesting that the analogs differ in their biochemical sites of action. Treatment of cells with either analog increased the amount of
neuraminidase-hydrolyzable
sialic acid-like material on the cell surface; this appeared to be due to the incorporation of the analogs into cellular
glycoconjugates, since incubation of cells with 3H-labeled analogs resulted in the appearance of radioactivity in cellular
ethanol-insoluble and
neuraminidase-hydrolyzable material. Incubation of cells with
Ac4-NAcMan labeled with 14C in the 4-O-acetyl group further demonstrated that incorporation occurred with approx. 50% retention of this substituent. Thus, both the amount and the nature of the surface
sialic acid constituents of treated cells were altered, suggesting that these or similar analogs could potentially be used to modify cellular membrane function.