DIFFERENTIATION INDUCTION BY A TUMOR-NECROSIS-FACTOR MUTANT-471 IN HUMAN MYELOGENOUS LEUKEMIC-CELLS VIA TUMOR-NECROSIS-FACTOR RECEPTOR-P55

The present study examined differentiation-inducing activity by variou s tumor-necrosis-factor(TNF) mutants against the human leukemic cell l ines HL-60 and U-937, Mutant TNF 471, from which 7 N-terminal amino ac ids of native TNF were deleted and pro(8), Ser(9) and Asp(10) were rep laced by Arg, Lys and Arg, possessed the highest activity among the TN F mutants, and its activity was 120-fold that of native TNF, The vario us biological activities of TNF were signaled through 2 distinct recep tors, p55 and p75, Although cytotoxicity was reported to involve mainl y p55, this differentiation inducing activity was not well understood, The fact that the affinity of TNF 471 was higher to p55 and lower to p75 than that of native TNF by a binding competition assay suggested t hat the differentiation-inducing activity was also signaled through p5 5. To verify this hypothesis, the human myelogenous leukemic cell line , KC-1, which scarcely expresses either receptor and does not differen tiate with TNF, was transduced with the p55 or p75 gene. Subsequently p55 transfectants manifested a greater ability to differentiate; howev er, p75 transfectants did not differ from parental cells or from mock- transfectants. Further, the differentiation of p55 transfectants induc ed by TNF was reduced by the inhibitor of protein-kinase-C (PKC), stau rosporine. These results indicate that the differentiation-inducing ac tivity was signaled through the TNF receptor, p55, via PKC and that th e excellent ability of TNF 471 to induce differentiation was related t o its high affinity for p55. (C) 1998 Wiley-Liss, Inc.