Therapeutic Effects of Islets Isolated From Genetically Engineered Pigs Transgenic for hCD46 and hCD39 in Immune-Deficient Mice.

Abstract

Background: Xenotransplantation using neonatal porcine islets is being considered as an alternative to allotransplantation as treatment for type 1 diabetes mellitus due to a more stable supply of islets. To overcome severe immunological responses, genetically engineered (GE) pigs have been developed with islets that may be quiescent to the human immune system. In this study, we investigated therapeutic effects of islets isolated from two different types of neonatal GE pigs. Group 1 GE pigs (GT KO+hCD46), the epitope alpha1, 3-galactose (αGal) was absent (GT KO), and transgenic for a human complement regulatory protein (hCD46). Group 2 GE pigs (GT KO+hCD46+hCD39) had the same genetic modification as Group 1 GE pigs but also were transgenic for a human anticoagulant gene (hCD39). Methods: Pancreases were harvested from Group 1 GE pigs aged 5 days, and from Group 2 GE pigs aged 2, 6 and 7 days. Islets were isolated with collagenase processing and followed by 7-day culture purification. Morphological and cellular composition of the islets were determined by immunohistochemical analysis and 2000 islet equivalents were transplanted under the kidney capsule of Sterptozotocin-induced diabetic immune-deficient mice to assess their function in vivo. Results: Morphological evaluation of the islets from Group 1 and Group 2 GE pigs showed no difference in the insulin and glucagon-positive cells. In addition, neonatal islets from both groups had no detectable αGal-positive cells. Immunoflorescence staining of pancreas sections taken from pigs cloned from the same exact cells showed strong CD46 and CD39 expression. The in vivo study showed that six of the 10 mice that received islets from Group 1 GE pigs and two of the four mouse recipients of islets from 6 day-old Group 2 GE pigs reached normal glycemia after 54.0 and 55.5 days of transplantation respectively. The mouse recipients of islets from 2 and 7 day-old Group 2 GE pigs remained diabetic for 113 days post-transplantation. Conclusions: In this study, islets from both types of GE pigs showed therapeutic effect on chemically induced diabetic immune-deficient mice. Furthermore, it remains to be determined whether islets from GT KO+hCD46 ± hCD39 pigs will be less susceptible to rejection mediated by human immune cells. DISCLOSURES:Bianchi, J.: Employee, Revivicor/United Therapeutics. Ayares, D.: Employee, Revivicor/United Therapeutics.