Cyanocobalamin [c-lactam] inhibits vitamin B12 and causes cytotoxicity in HL60 cells: methionine protects cells completely.
The [c-lactam] derivative of cobalamin antagonizes vitamin B12 in vivo. Therefore, we investigated its effects in tissue culture to develop a model in which to study vitamin B12-deficient hemopoiesis. HL60 cells were cultured in medium containing either methionine or L-homocysteine thiolactone, and various concentrations of 5-methyltetrahydrofolate or pteroylglutamic acid. In medium with L-homocysteine thiolactone, 5-methyltetrahydrofolate, and dialyzed serum, cyanocobalamin [c-lactam] caused cell death, reversible by additional vitamin B12. Pteroylglutamic acid did not prevent this cytotoxic effect. Methionine completely protected cells against cyanocobalamin [c-lactam] for periods of up to 4 months of culture, irrespective of the folate source. Cyanocobalamin [c-lactam] reversibly impaired the incorporation of 5-[14CH3]-tetrahydrofolate and [1-(14)C] propionic acid by intact cells, consistent with inhibition of methionine synthase and methylmalonyl-CoA mutase. A substantial proportion of 5-[14CH3]-tetrahydrofolate uptake could not be suppressed by methionine and may, therefore, have occurred outside of the methionine synthase pathway. These findings are the first indication that cyanocobalamin [c-lactam] antagonizes vitamin B12 in vitro and causes cell death from methionine deficiency. The model should be valuable for investigating the biochemical pathology of vitamin B12-deficient hemopoiesis. The results suggest that methylfolate is not trapped when methionine synthase is inhibited in HL60 cells, but they do not disprove the methylfolate trap hypothesis as applied to normal blood cells.
Matthews
J H
JH
Department of Medicine, Queen's University, Kingston, Ontario, Canada.
D013766:5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase / Q000037:antagonists & inhibitors; D016923:Cell Death / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005492:Folic Acid / Q000494:pharmacology; D018922:HL-60 Cells / Q000187:drug effects*; D006710:Homocysteine / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D006879:Hydroxocobalamin / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D008715:Methionine / Q000172:deficiency / Q000494:pharmacology*; D008765:Methylmalonyl-CoA Mutase / Q000037:antagonists & inhibitors; D011422:Propionates / Q000378:metabolism; D013763:Tetrahydrofolates / Q000378:metabolism; D014805:Vitamin B 12 / Q000037:antagonists & inhibitors*
D004791:Enzyme Inhibitors; D011422:Propionates; D013763:Tetrahydrofolates; D006710:Homocysteine; D005492:Folic Acid; D008715:Methionine; C007957:homocysteine thiolactone; D013766:5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase; D008765:Methylmalonyl-CoA Mutase; C029658:propionic acid; D014805:Vitamin B 12; D006879:Hydroxocobalamin