Nutrition based gene regulation system applicable for gene therapy in human. – NUTRITHERAGENE

1- Optimization of the existing construct: we have tested several regulatory sequences to improve the inducibility
2- Transgene induction and Mapping of the GCN2/eIF2a/ATF4 pathway in tissues: We have subjected transgenic mice expressing the luciferase gene under the control of AARE to different diets lacking a single essential amino acid. Then, luciferase expression has been studied in several tissues
3- Proof of concept: genes of interest have been tested according their relevance in the fields of cancer and/or neurodegenerative diseases (in collaboration with specialized laboratories)

We have developed a promoter containing several copies of AARE and a diet that can rapidly induce a blood decrease of one EAA, which dramatically activates the AARE-dependent transcription. This system provides a cogent mean to regulate transgene in the context of gene therapy. The characteristics and benefits of the “nutrition-based” regulatory system are: (1) precise control of the induction period, (2) very low basal expression of the transgene and a strongly induced expression level, (3) control of transgene expression in several tissues including the brain, and (4) absence of toxicity.

This novel nutrition-based regulatory system stands as a physiological approach with the ability to resolve one of the major remaining hurdles in human gene therapy It is presently the only available system that is easily applicable to humans and thus represents a major step forward in bringing gene therapy to clinical fruition with the prospect of having a global impact on the entire gene therapy field.

2 articles , 1 patent

Nutrition based gene regulation system applicable for gene therapy in human

Gene therapy is a treatment that consists in replacing defective genes with healthy ones, delivering therapeutic genes such as trophic factors to prevent cell deaths, modulating the expression of endogenous genes/network of genes using shRNA/miRNA. Several barriers need to be overcome before this type of therapy becomes a widely accepted treatment for a broad group of medical diseases. Besides the concerns of the safety of available vector systems, the major limitation in the use of gene therapy is the requirement to exogenously modulate the expression of transgenes in term of time course and expression level.

We have developed a regulatory system, which meet all requirements expressed above. Our work on nutrition led us to delineate the transcriptional mechanisms that underline the control of gene expression by nutritional limitation in Essential Amino Acid (EAA). More particularly, we have shown that AAE limitation activates a signaling pathway that include the protein kinase GCN2, the translation initiation factor eIF2a and the transcription factor ATF4 which binds the AARE to turn-on transcription. We have developed a promoter containing several copies of AARE and a diet that can rapidly induce a blood decrease of one EAA, which dramatically activates the AARE-dependent transcription. This system provides a cogent mean to regulate transgene in the context of gene therapy. It is novel and overcome the remaining hurdle in the development of gene therapy for numerous diseases. The main characteristics and benefits of this system are: 1) a precise control of the induction period 2) a low basal expression level of the transgene expression and a good induced-expression level. 3) the possibility to control transgene expression in brain (the blood brain barrier amplifies the decrease of the limiting AA) and other tissues (eyes, pancreas, liver…) 4) No toxicity: A single meal deficient in one EAA has absolutely no toxicity. If treatment must be renewed periodically, it will be possible to alternate starvation of several EAA.

This system has been patented and this application aims to bring this technology to clinical fruition. This project results from the merging of two disciplines: nutrition and gene therapy through the collaboration between two groups that are leader in these two fields and that have collaborated fruitfully in the recent years. The main tasks of this project are straightforward and can be summarized as follow: The first work package consists in optimizing and improving the gene expression system. In task1 we will design new vectors for an optimal induction of transgene expression. Then in task2 we will optimize the nutritional conditions leading to the transgene induction. The final goal is to generate several vectors and several inducing-diets with different properties in term of expression level in both condition non-induced and induced. Thus, it will be possible to adapt both the vector and the AA-deficient diet to specific needs. Finally, task3 will aim to evaluate the signal/noise ratio of the transgene expression in certain tissues in pathophysiological conditions. In the second work package, we will exemplify the use of the “nutrition-based” gene expression system by proposing two applications. The first one concerns a clinical application and consists in the inhibition of food intake by controlled leptin expression in the arcuate nucleus of the hypothalamus. The second one relates to experimental studies in mice and consists in the development of a cell type specific AARE-regulated transgene system.

Briefly, the objective of the present application consists in optimizing the “nutritional-based” gene expression system we have developed in order to make the advance from proof of concept to proof of practice for clinical applications.