Protease Activated Masked Interferon to Treat Malignancy

PROJECT SUMMARY ABSTRACT IFNand IFNtype Iand IFNtype IIare potent mediators of anti cancer immunity having both direct antiproliferative effects against many cancers as well as a multitude of anti tumor immunotherapeutic effectsHoweverwhile IFN has efficacy against multiple human cancers its clinical utility to date has been limited by the inability to achieve effective concentrations of IFN at tumor sites without causing systemic toxicitySeveral groups including ours have successfully approached this problem by using the tumor targeting ability of monoclonal antibodies to carry IFNs directly to tumor sitesThese approaches have shown great promise and anti CDIFNis in clinical trials by ImmunGene and anti CDIFNis being developed by Qwixel Therapeutics under a license from UCLAAlthough using the antibody binding specificity to target tumorassociated antigens delivers a greater percentage of the IFN to the site of the tumor than is achieved when IFN is injected by itselfthe attached interferon still is recognized and bound by interferon receptors expressed throughout the body that are not tumor associatedThis binding can result both in less IFN reaching the tumor and in unwanted off target toxicityThe goal of the present studies will be to produce anti IFN fusion proteins in which the IFN will only bind IFN receptors once it reaches the tumorTo achieve that goal we will attach a mask to the IFNwhich will inhibit its activityThis mask will be separated from the IFN by a peptide linker that is a site for proteolytic cleavageA hallmark of cancer is increased proteolytic activity in tumor tissue and the surrounding microenvironmentWe therefore hypothesize that we can achieve improved tumor targeting of IFN activity by producing it as a prodrug that will be activated only in the tumor microenvironment following removal of the mask by the tumor associated proteasesPreliminary studies have identified a potential mask and cleavable peptide linker for IFNWe will now use tumor cell lines and extracts to confirm that the mask inhibits IFNactivitythat it can be removed within the tumor microenvionment and that the unmasked protein is now active both in vitro and in xenograft modelsHumanized NSG mice will be used to determine if the masked interferon fusion protein elicits a decreased inflammatory response compared to unmasked IFNSimilar approaches will be used to produce a masked IFNusing either a peptide or scFv maskThe resulting masked IFNwill be characterized as will be done for IFNWith the successful completion of these experimentsQwixel will have novel IFN fusion proteins that are even more effective in delivering large amounts of IFN to tumor sites in the absence of systemic toxicity PROJECT NARRATIVE Although fusion proteins in which IFN is genetically fused to an antibody that recognizes a tumor associated antigen have shown great clinical promisethe attached interferon still is recognized and bound by interferon receptors expressed throughout the body that are not tumor associatedThe goal of the present studies will be to produce antibody IFN fusion pro drugs in which the IFN will only bind IFN receptors once it reaches the tumorsIt is hypothesized that these pro drug fusion proteins will have enhanced PK and increased therapeutic window making them more effective cancer therapeutics