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Haploinsufficiency and Intractable Epilepsy Rescue Increasing Endogenous Gene Promoter Efficiency

Periodic Reporting for period 1 - PromoTeRapy (Haploinsufficiency and Intractable Epilepsy Rescue Increasing Endogenous Gene PromoterEfficiency)

Reporting period: 2016-02-01 to 2018-01-31

The possibility to regulate gene expression for gene therapy without adding new extra genes potentially can revolutionise this field. But at the moment there are not scientific evidences to prove that this is feasible.
PromoTerapy demonstrated that is possible to treat acquired or genetic epilepsies modifying gene expression. It has been the first time that was possible to increase the activity of genes without inserting new extra genes in the DNA. We used this system to restore normal level of a gene which loss of function cause a severe childhood epilepsy with autistic features (Dravet syndrome) but also to increase a protective gene against acquired intractable epilepsy induced by hippocampal damage.
This project is important for the society because is the first proof of principle that severe diseases can be cured with new gene editing techniques. Diseases that till now had no treatments available.
During PromoTerapy we developed a new gene therapy approach to treat both acquired or genetic epilepsies. We used CRISPRon, a modified version of the gene editing tool not able to cut the DNA but to increase promoter activity, to increase the expression of a potassium channel in an hippocampal model of epilepsy and to increase a sodium channel which down regulation is responsible of a severe childhood epilepsy (Dravet syndrome). Initially we design the gene therapy approach testing with bioinformatics and molecular biology the efficiency in cell line. Then this approach, in parallel for sodium and potassium channel, was tested functionally with electrophysiology in rodent neurons. This validation produced also the development off a new technique, activity clamp, used to understand the action of an anti-epileptic drug working on sodium channels.This work has been published as a side story to the main project. Once we proved that we were be able to increase the expression of endogenous genes in neurons we moved to study the effect of this potential treatment for epilepsy. The sodium channel approach was used in vitro and in vivo in an animal model of Dravet syndrome in collaboration with Vania Broccoli in Milan. The potassium channel approach was used in vivo in the hippocampus in a model of intractable temporal lobe epilepsy.
The results of PromoTerapy have been presented at national and international scientific conferences, and 2 papers will be published in peer-reviewed scientific journals.
Gene therapy can potentially cure d Epilepsy and other neurological disorders but some limitations are still present. One big limitation is the size of the extra-gene we would like to deliver to treat the disease. At the moment this limitation is due to the maximal size that a viral vector can carry. PromoTerapy overcome this limitation, acting directly on the genes present in the genome and not rely on extra-gene to add. For this reason the size of the gene therapy tool will be always the same changing only the targeted gene promoter. This is a huge step forward for cure neurological disease.
Recent years have seen enormous advances in viral vectors as delivering shuttles to overexpress transgenes of interest in vivo. In the last year 3 novel gene-therapies were approved by the FDA. This highlights the importance of this research area in the development of novel strategies for intractable diseases. With PromoTerapy we were able to be part of this development and propose a novel strategy to overcome an important limitation of current gene therapy.
PromoTerapy Strategy