ANTITUMOR-ACTIVITY OF TUMOR-NECROSIS-FACTOR-ALPHA CONJUGATED WITH DIVINYL ETHER AND MALEIC-ANHYDRIDE COPOLYMER ON SOLID TUMORS IN MICE

The purpose of this study is to further explore the usefulness of conj ugation with functional polymeric modifiers for clinical application o f bioactive proteins and to increase the therapeutic efficacy of tumor necrosis factor alpha (TNF-alpha) by conjugation in vivo. We synthesi zed TNF-alpha conjugated with the copolymer of divinyl ether and malei c anhydride (DIVEMA), which has intrinsic antitumor activity as a synt hetic biological response modifier. The synthesis of DIVEMA-TNF-alpha could be controlled by the addition of 2,3-dimethylmaleic anhydride (D MMAn), which binds to or separates from amino groups when the pH is ch anged. The specific activity of DIVEMA-TNF-alpha (+) synthesized with DMNAn was hardly decreased in vitro, However, DIVEMA-TNF-alpha (-), wh ich is conjugated without blocking by DMMAn, had a markedly diminished specific activity, DIVEMA-TNF-alpha (+) caused a dramatic hemorrhagic necrotic effect on the tumor when compared to native TNF-alpha 24 h a fter i.v. injection into mice bearing Sarcoma-180 solid tumors. In add ition, DIVEMA-TNF-alpha (+) at a dose of only 100 Japan reference unit s per mouse revealed a dramatic antitumor effect that is approximately 100 times greater than native TNF-alpha and that could induce complet e regression in all five mice bearing Meth-A solid tumors without any apparent side effects. Because neither DIVEMA alone nor a mixture of T NF-alpha and DIVEMA caused antitumor activity with i.v. administration , the increase in antitumor potency of TNF-alpha may be caused by the covalent conjugation with DIVEMA. DIVEMA-TNF-alpha at low dose reveale d dramatic antitumor potency, Because TNF-alpha injected in vivo is ex tremely low-dose, concentration of intrinsic TNF-alpha in vivo is not influenced. Therefore, the cytokine network in vivo is not destroyed. These results suggest that DIVEMA is a useful polymeric modifier for c onjugation of TNF-alpha to increase its antitumor activity.