CONSTRUCTION OF A NONINTEGRATIVE PLURIPOTENCY GENE VECTOR FOR THE INDUCTION OF NEURAL PROGENITORS

MARIANNE LEHMANN, MICAELA LÓPEZ-LEÓN, SANTIAGO HAASE, MELISA FRAGOMENO, VÍCTOR ROMANOWSKI RODOLFO G. GOYA

Buenos Aires

Congreso; 2nd Congress of the Federation of Latin-American and Caribbean Societies for Neuroscience (FALAN); 2016

Federation of Latin-American and Caribbean Societies for Neuroscience

CONSTRUCTION OF A NONINTEGRATIVE PLURIPOTENCY GENE VECTOR FOR THE INDUCTION OF NEURAL PROGENITORS Marianne Lehmanna, Micaela López-Leóna, Santiago Haaseb, Melisa Fragomenoa, Víctor Romanowskib Rodolfo G. Goyaa aINIBIOLP-Histology B -Pathology B, School of Medicine, National University of La Plata (UNLP) bInstitute for Biotechnology and Molecular Biology, (IBBM), UNLP Keywords: cell reprogramming- nonintegrative ? Yamanaka genes ? STEMCCA- direct reprogramming- neural progenitorsABSTRACTThe absence of efficacious therapies for neurodegenerative diseases has increased the interest in regenerative medicine, particularly after the advent of cell reprogramming technologies, which have ushered in the era of personalized cell therapeutics. The use retroviral vectors for cell reprogramming, conveys the risk of insertional mutagenesis and the consequent emergence of tumors in the treated individuals. Currently, efforts are being made to develop nonintegrative gene delivery systems, thus avoiding the risk of insertional mutagenesis.We want to reprogram skin fibroblasts into neural progenitors by setting up a novel nonintegrative approach, termed direct cell reprogramming. We have already constructed a regulatable plasmid system for the simultaneous expression of the pluripotency genes sox2, c-myc, klf4 y oct4 and the reporter gene for green fluorescent protein (hGFP). The plasmid harbors a regulatable bidirectional cassette which expresses hGFP and a tandem (termed STEMCCA cassette, donated by G. Mostoslavsky) harboring the four Yamanaka pluripotency genes. Both the hGFP gene and the pluripotency gene tandem are under the control of a single regulatable (Tet-Off) bidirectional promoter which can be reversibly inhibited by the antibiotic doxycycline. We are currently characterizing the plasmid and setting up a magnetofection technique in order to optimize the delivery of this vector to primary cultures of mouse fibroblasts and the 3T3 fibroblast cell line.