Novel mutations in NCF4 gene confer non-classic chronic granulomatous disease with disseminated histoplasmosis in a Colombian child

Carlos Andrés Arango-Franco, Alejandro Nieto-Patlán, Marcela Moncada-Vélez, Jesús Armando Álvarez, Carmen Oleaga-Quinta, Caroline Deswarte, Juan Fernando Alzate, Felipe Cabarcas, Carlos Garcés, Julio César Orrego, Susana Pamela Mejía, Luz Elena Cano, Jean-Laurent Casanova, Jacinta Bustamante, José Luis Franco, Andrés Augusto Arias


Background: Chronic granulomatous disease (CGD) is a primary immunodeficiency characterized by susceptibility to early-onset life-threatening bacterial and fungal infections as well as dysregulated chronic inflammation. CGD results from mutations in one of the components of the phagocyte NADPH oxidase, a multimeric complex that consists of two membrane-bound components (gp91phox and p22phox) and 3 cytoplasmic subunits (p40phox, p47phox y p67phox) that function to induce reactive O2 species (ROS) in phagocytic cells to induce microbial killing. To date, only a single patient with granulomatous colitis and compound heterozygous mutations in NCF4 encoding p40phox has been reported as a genetic subgroup of CGD.

Method: We performed whole exome-sequencing in a patient with early-onset systemic histoplasmosis. Functional testing to Investigate phagocyte NADPH oxidase included dihydrorhodamine oxidation assay as well as amplex red and luminol. Protein expression was assessed by FACS and immunoblotting.

Results: We found a missense homozygous variation in NCF4 within the phox homology (PX) domain, predicted to be damaging by polyphen and SIFT2 with a CADD score of 35. RT-PCR and immunoblotting demonstrated decreased p40phox protein expression protein both in neutrophils and EBV-transformed B cells from the patient, but not from controls. In addition, intracellular (IC) ROS production was significantly impaired after physiological stimulation with fMLP, Histoplasma capsulatum and Candida albicans on neutrophils and EBV-B, but not with phorbol 12-myristate 13-acetate (PMA).

Conclusions: We report a novel homozygous mutation in NCF4 selectively impairing IC ROS production in a Colombian child. Remarkably, systemic histoplasmosis has not been previously reported in association with classical CGD, therefore our results expand the spectrum of genetic and infectious diseases underlying CGD in humans.

Palabras clave

NADPH; Whole exome sequencing; NCF4; Chronic granulomatous diseases

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