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Correction of Respiratory Burst Activity in X-Linked Chronic Granulomatous Cells to Therapeutically Relevant Levels after Gene Transfer into Bone Marrow CD34 + Cells

1998; Mary Ann Liebert, Inc.; Volume: 9; Issue: 11 Linguagem: Inglês

10.1089/hum.1998.9.11-1561

1557-7422

Sven Becker, Sandra Wasser, Martin Hauses, Johann Peter Hossle, Marion Ott, Mary C. Dinauer, Arnold Ganser, Dieter Hoelzer, Reinhard Seger, Manuel Grez,

Calcium signaling and nucleotide metabolism

Chronic granulomatous disease (CGD) is a disorder of the lymphohematopoietic system, whereby phagocytes of affected patients are unable to kill microorganisms. CGD is caused by a functional defect in the phagocytic nicotinamide adenine dinucleotide phosphatase (NADPH) oxidase (phox) enzyme complex, leading to a lack of microbicidal metabolites. As a therapeutic approach toward the predominant X-linked form of CGD, we have developed a bicistronic retroviral vector containing the coding sequences of gp91-phox and a cytoplasmically truncated version of the human low-affinity receptor for nerve growth factor (ΔLNGFR). Full reconstitution of superoxide-generating activity was achieved with this vector in a gp91-phox-deficient cell line. Using an optimized gene transfer protocol, up to 85% of the CD34+ cells obtained from the bone marrow of X-CGD patients were transduced. CD15+ cells differentiated in vitro from transduced X-CGD CD34+ cells showed correction of NADPH oxidase activity to 45–52% of normal levels whereas ΔLNGFR expression was found in 40–67% of the CD15+ cells. Moreover, immunoblots prepared from extracts of transduced CD15+ cells revealed gp91-phox protein levels similar to those found in neutrophils derived from normal CD34+ cells. Taking into consideration that superoxide production in only 5 to 10% of wild-type neutrophils is sufficient to protect X-CGD heterozygotes from serious infections, the results achieved in this study shows that for X-CGD patients a curative approach based on the genetic modification of hematopoietic stem/progenitor cells is feasible. In this study we describe reconstitution of respiratory burst activity in X-CGD cells after gene transfer into CD34+ cells of an optimized retroviral vector containing the gp91-phox gene and a cytoplasmically truncated version of the human low-affinity receptor for nerve growth factor (ΔLNGFR). NADPH oxidase activity in neutrophils derived from transduced X-CGD CD34+ cells was reconstituted to therapeutically relevant levels (45–52%) in three of three samples.