Terapia génica para la diabetes tipo I basada en la administración intramuscular de AAV1 Insulina-Glucoquinasa

Abstract

Diabetes is a complex metabolic disease for which there is currently no cure and is associated with severe secondary complications, caused largely by poor glycaemic control. Achievement of normoglycemia with exogenous insulin treatment requires the use of high doses of hormone, which increases the risk of life-threatening hypoglycemic episodes. We previously demonstrated that a single intramuscular administration of two AAV serotype 1 (AAV1) vectors expressing insulin and glucokinase produce 4 years of disease correction in diabetic dogs. The first part of this study focused on the evaluation of efficacy and safety after 8 years of treatment. It is demonstrated how these dogs maintain glycemic control without the need of exogenous insulin for a period of up to 8 years after a single administration of the therapeutic vectors. Long term metabolic normalization was demonstrated by the multi-annual quantification of serum levels of glycosylated proteins (fructosamine), triglycerides and cholesterol and the correct response of the treated animals to the oral glucose tolerance test. The persistence of the viral genomes and the expression and activity of the therapeutic transgenes were confirmed in multiple samples of the treated muscles. In addition, no signs of pathology were observed in the histopathological analysis of the same muscles. In the second part of the study, dual AAV1 vectors were generated that encoded together insulin and glucokinase. The generation of a dual vector will allow to increase the therapeutic efficacy since all the genetically modified cells will express the two components of the "glucose sensor", while a reduced viral dose with minimize potential toxicity, and simplify both the production and regulatory processes required for future clinical application. The design of the modular system allowed for the generation and evaluation of conformations and combinations of coding and regulatory sequences. These were evaluated in vitro to demonstrate that the dual vectors with the two units in opposite conformation mediated a greater expression of both transgenes. The data in the studies performed in healthy and diabetic mice administered with the dual vectors confirmed that efficacy was superior to that obtained in the mice treated with the individual vectors. Therefore, the data obtained in both parts of this work demonstrate the long-term functionality and safety of intramuscular administration of AAV1 vectors encoding insulin and glucokinase to counteract diabetes while providing the basis for future clinical translation of a this type of gene therapy strategy, using a single dual AAV1 vector expressing both genes.

Date of Award	24 Nov 2017
Original language	Spanish
Supervisor	Maria Fatima Bosch Tubert (Director) & Miguel García Martínez (Director)