HUMAN GENE THERAPY 11:1647–1656 (August 10, 2000) Mary Ann Liebert, Inc. Biolistic-Mediated Interleukin 4 Gene Transfer Prevents the Onset of Type 1 Diabetes MARK J. CAMERON, 1,2 CRAIG A. STRATHDEE, 2,3 KEVIN D. HOLMES, 2,3 GUILLERMO A. ARREAZA, 1,2 GREGORY A. DEKABAN, 2,3 and TERRY L. DELOVITCH 1,2,4 ABSTRACT We tested the efficacy of biolistic-mediated gene transfer as a noninvasive therapy for type 1 diabetes (T1D) in nonobese diabetic (NOD) mice by expression of murine interleukin 4 (mIL-4) cDNA. Epidermal delivery of 2 m g of DNA yielded transient detection of serum mIL-4, using a conventional cDNA expression vector. A vector stabilized by incorporation of the Epstein–Barr virus (EBV) EBNA1/ oriP episomal maintenance repli- con produced higher levels of serum mIL-4 that persisted for 12 days after inoculation. Although biolistic in- oculation of either vector reduced insulitis and prevented diabetes, the protracted mIL-4 expression afforded by the EBV vector resulted in Th2-type responses in the periphery and pancreas and more significant pro- tection from the onset of diabetes. Our studies demonstrate the efficacy of biolistic gene delivery of stabilized cytokine expression as a viable therapeutic approach to prevent the onset of T1D. 1647 OVERVIEW SUMMARY We have previously shown that systemic injection of multi- ple low doses of recombinant mIL-4 can prevent T1D in NOD mice. To provide a more efficient and advanced means of reducing pancreas inflammation, we adopted a biolistic- mediated gene transfer approach to sustaining mIL-4 lev- els in NOD mice. In this article, we tested the therapeutic potential of two mIL-4 cDNA-expressing vectors. We found that only three biolistic inoculations with the conventional pWRG-IL-4 vector or the novel pHERO-IL-4 vector reduce insulitis and the incidence of diabetes in NOD mice with no adverse effects. pHERO-IL-4 elicited peripheral and intra- pancreatic Th2 cell responses, and proved more effective as a therapeutic agent than pWRG-IL-4. This observation was dependent on the EBV EBNA1/oriP episomal maintenance replicon of pHERO-IL-4, which prolongs transgene ex- pression. Thus, biolistic gene delivery of mIL-4 represents a clinically viable approach to the therapy of T1D. INTRODUCTION T YPE 1 DIABETES (T1D) is an organ-specific autoimmune dis- ease. Although not inherited, susceptibility to the disease is genetically transmitted and is under multigenic control. To- gether, type 1 and type 2 diabetes afflict about 5% of Western populations. T1D affects bout 1 in 6000 North Americans and is most prominent among young children, with the peak age of onset at about 11–12 years of age. Patients with T1D develop a lifelong dependence on insulin for survival; however, insulin therapy does not prevent the eventual complications of T1D. An- other immune-specific type of therapy may be more efficacious. Nonobese diabetic (NOD) mice spontaneously develop a form of T1D that shares many features of the human disease (Delovitch and Singh, 1997). T1D in NOD mice occurs as a re- sult of a progressive CD4 1 helper T type 1 (Th1) cell-medi- ated destruction of insulin-producing b cells in the absence of regulatory CD4 1 Th2 cell function (Shehadeh et al., 1993; Ra- binovitch et al ., 1995; Arreaza et al., 1997; Cameron et al ., 1 Autoimmunity/Diabetes Group, John P. Robarts Research Institute, London, Ontario N6G 2V4, Canada. 2 Department of Microbiology and Immunology, University of Western Ontario, London, Ontario N6A 5C1, Canada. 3 Gene Therapy and Molecular Virology Group, John P. Robarts Research Institute, London, Ontario N6G 2V4, Canada. 4 Department of Medicine, University of Western Ontario, London, Ontario N6A 5C1, Canada.