Effect of the antileukemic agent L-asparaginase on thyroxine-binding globulin and albumin synthesis in cultured human hepatoma (HEP G2) cells.

L-Asparaginase (ASNase), a drug widely used in the treatment of acute lymphoblastic leukemia, has been reported to decrease serum T4-binding globulin (TBG) levels, while results of serum albumin determinations were conflicting. This effect in vivo has been attributed to depressed liver protein synthesis, but this hypothesis has not been proved. To investigate this problem, human hepatoma (Hep G2) cells were continuously labeled for 4 h with 100 microCi/ml [35S]methionine in the absence or presence of graded amounts of ASNase (from 0.1 nM to 0.1 mM). Media and cell lysates were collected, immunoprecipitated with antialbumin or anti-TBG serum and protein A, and submitted to sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Gels were sliced, and the radioactivity was counted in a beta-counter. A dose-dependent inhibition of TBG and albumin biosynthesis (as well as of total protein synthesis) was demonstrable, but TBG appeared to be more sensitive to the action of the drug. In fact, TBG biosynthesis was reduced by 8% with 0.1 nM ASNase, while an effect on albumin was observed only at 1 nM ASNase; 50% inhibition was obtained with 30 nM ASNase in the case of TBG and with 800 nM in the case of albumin. At the highest concentration (0.1 mM), TBG biosynthesis was reduced by 94%, and albumin biosynthesis by 75%. ASNase also proved to have a time-dependent effect, as assessed by the measurement of radioimmunoassayable TBG in the media from Hep G2 cells grown in the presence of 10 nM ASNase for 1-4 days. The TBG concentration was progressively reduced, by 40% after 1 day to 85% after 4 days. In pulse-chase experiments, a reduction of total (intracellular plus secreted) immunoprecipitable TBG and, to a lesser extent, albumin was observed, suggesting that the drug also affected the catabolism of newly synthesized proteins. These results provide the first in vitro evidence that ASNase actually inhibits TBG biosynthesis. This effect is not specific for TBG, but this protein appears to be more susceptible than albumin to ASNase action. This can explain why in patients treated with ASNase for leukemia, a decrease in serum TBG concentrations has not always been associated with a reduction in serum albumin levels.