IMMUNOREGULATORY EFFECTS OF THE INTERFERONS

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The interferons mediate a wide variety of biologic functions, including the ability to induce differentiation, inhibit normal and tumor cell growth, and reverse the action of several oncogenes; we hypothesize that they function as negative growth regulators and interact with T cells, B cells and monocytes as immunoregulatory lymphokines. They have demonstrated significant clinical activity in hematologic neoplasms and our own data imply a direct, antiproliferative mechanism of action mediated via specific cell surface receptors, possibly as a second message growth inhibitor leading to oncogene modulation. Nevertheless, they are not widely used as anti-cancer agents in spite of the presence of specific receptors for interferon on a variety of tumor cell lines and freshly explanted tumors. To understand further their molecular biology and their possible role in the lymphokine network, we now propose to prepare specific anti-idiotype antibodies to type I and II interferon receptors, to purify the human receptor for alpha-2 interferon from the Burkitt lymphoma cell line Daudi, and to explore the role of alpha and gamma interferon and receptor expression as negative growth regulators in interferon sensitive and resistant human cell lines and tumor cells. Recombinant alpha and gamma interferon will be radiolabeled and used to assess specific receptor binding, turnover and subcellular localization. We will prepare monoclonal antibodies in mice and polyclonal antibodies in rabbits specific for the type I and II receptor using, alternatively, purified receptor proteins and/or the production of anti-idiotypes to monoclonal antibodies already prepared in our laboratory that bind to receptor-specific domains on the ligands and block antiviral and anti-proliferative function. Purification of the receptor will be accomplished using affinity chromatography on interferonsepharose, on anti-receptor antibody sepharose, on wheat germ sepharose, and with HPLC. The amino acid sequence of the purified receptor and of isolated tryptic peptides will be determined and used to construct several oligonucleotide probes that will be used to screen a genomic human DNA library contained in lambda phage. Alternatively, the anti-receptor antibodies or oligonucleotide probes can be used to screen a cDNA expression library in E. coli. The receptor gene will be isolated, cloned, and its nucleotide and flanking sequences determined as well as its chromosome localization. Appropriate gene fragments will be used to analyze the DNA and RNA in normal and cancerous human tissues to determine the mechanism of susceptibility and resistance to interferon's action and its effects on specific oncogenes. These studies will clarify the biology of the interferon system and may permit its clinical use in combination with other lymphokines or possibly identify alternative therapeutic strategies in the treatment of human malignancy.