Phenotypic correction of dwarfism mediated by different growth parameters after plasmid DNA administration in an animal model of isolated growth hormone deficiency

Author(s)

Eliza Higuti

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Instituto de Pesquisas Energéticas e Nucleares (IPEN/BT)
• Defense date: 2016-01-22

Examining board members

Cibele Nunes Peroni; Paolo Bartolini; Claudia Regina Cecchi; Alexander Augusto de Lima Jorge; Poliana Cristina de Melo Martins

• Advisor: Cibele Nunes Peroni

Abstract

The human growth hormone deficiency (GHD) is the most common deficiency related to pituitary hormones. The current therapy is based on daily injections of recombinant human growth hormone (r-hGH). This therapy, however, presents some disadvantages, as the need for frequent injections of r-hGH during a long life time, depending on the deficiency severity and the high cost of this hormone, due to the expensive purification processes. An alternative to the standard treatment should be to avoid these inconveniences via a sustainable hormone release, acting for a long time and providing normal and sustainable levels of insulin-like growth factor-I (IGF-I). A possible alternative is in vivo gene therapy, based on the administration of plasmid DNA in several organs/tissues, followed by electroporation. This methodology is considered very promising and has been the target of many different studies for several types of systemic deficiencies. In the present work several administrations of a plasmid containing the human growth hormone gene were carried out, in the exposed quadriceps or non-exposed tibialis cranialis muscle, followed by electroporation, using immunodeficient dwarf mice 40-80 days old. The goal was to obtain a phenotypic correction of dwarfism, through the evaluation of different growth parameters. The administration of this plasmid, in the tibialis cranialis muscle of 40 day old mice, was able to provide a normalization of mIGF-I levels, when compared to non GHD mice. Furthermore, catch-up increases of longitudinal growth parameters of 36-77% were obtained. Aiming a high efficiency on GH expression, parental plasmids were constructed and from these DNA minicircles were generated with CMV and Ubiquitin C promoter and hGH or mGH cDNA sequences. These DNA minicircles were transfected into HEK 293 cells and were even 2 times moren efficient than conventional plasmids with CMV promoter. This data are very promising and pave the way for more efficient assays utilizing this type of gene therapy protocol for GHD, aiming at a normalization of all growth parameters. (AU)

• FAPESP's process: 11/21708-6 - Complete phenotypic correction of dwarfism by injection of plasmid DNA in an animal model of isolated growth hormone deficiency
• Grantee: Eliza Higuti Sousa
• Support Opportunities: Scholarships in Brazil - Doctorate