Method for optimizing the in vitro production of clinical grade induced pluripotent human stem cells.

摘要

A method of optimizing the in vitro production of human induced pluripotent stem (iPS) cells from hematopoietic cells, characterized by being performed in accordance with clinical grade good manufacturing practices and comprising the following steps: a separation step of a population of hematopoietic mononuclear cells by centrifugation of peripheral blood isolated from a patient; a first step of amplification of said hematopoietic mononuclear cell population by culturing said cell population on a first growth medium which promotes its expansion; a step for monitoring the rate of expansion of said population of expanded hematopoietic cells; a step of collecting said population of expanded hematopoietic cells carried out at the expansion peak; a transfection step of said collection of expanded hematopoietic cells through the introduction of reprogramming factors comprising one or more plasmids selected from pCLXE-hOCT3 / 4-shp53-F, pCXLE-hSK, pCXLE-hUL, MOS, MMK, MBX, pEB- C5 and pEB-Tg; a second step of amplification of said transfected cells; a step of plating on at least one matrix for the engraftment of said amplified transfected cells; a step in the production of human pluripotent stem cells induced by direct culture of said cells plated on a specific medium for human stem cells.