To eliminate systemic toxicity, including the hypotension associated with human tumor necrosis factor alpha (TNF-alpha), we constructed mutant proteins (muteins) by mean of genetic engineering. A novel mutein, F4614, containing mutations of 5Thr-->Gly and 6Pro-->Asp, which resulted in the introduction of cell-adhesive Arg-Gly-Asp and 29Arg-->Val, had remarkably reduced hypotensive effects and lower lethality. We present evidence that the Arg-->Val mutation at position 29 is largely responsible for the reduced hypotensive effect. This effect of F4614 was thought to be closely correlated with its low inducibility of nitric oxide and prostaglandin E2 in vivo. In addition, the therapeutically effective dose of F4614 to MethA fibrosarcoma-transplanted mice was increased compared with that of TNF-alpha, indicating a wide therapeutic index. These results indicated that F4614 has several advantages as a systemic therapeutic drug in the treatment of cancer.