The Extended Clinical Phenotype of 26 Patients with Chronic Mucocutaneous Candidiasis due to Gain-of-Function Mutations in STAT1

Abstract

Purpose: Gain-of-function (GOF) mutations in the signal transducer and activator of transcription 1 (STAT1) result in unbalanced STAT signaling and cause immune dysregulation and immunodeficiency. The latter is often characterized by the susceptibility to recurrent Candida infections, resulting in the clinical picture of chronic mucocutaneous candidiasis (CMC). This study aims to assess the frequency of GOF STAT1 mutations in a large international cohort of CMC patients.

Methods: STAT1 was sequenced in genomic DNA from 57 CMC patients and 35 healthy family members. The functional relevance of nine different STAT1 variants was shown by flow cytometric analysis of STAT1 phosphorylation in patients' peripheral blood cells (PBMC) after stimulation with interferon (IFN)-α, IFN-γ or interleukin-27 respectively. Extended clinical data sets were collected and summarized for 26 patients.

Results: Heterozygous mutations within STAT1 were identified in 35 of 57 CMC patients (61%). Out of 39 familial cases from 11 families, 26 patients (67%) from 9 families and out of 18 sporadic cases, 9 patients (50%) were shown to have heterozygous mutations within STAT1. Thirteen distinct STAT1 mutations are reported in this paper. Eight of these mutations are known to cause CMC (p.M202V, p.A267V, p.R274W, p.R274Q, p.T385M, p.K388E, p.N397D, and p.F404Y). However, five STAT1 variants (p.F172L, p.Y287D, p.P293S, p.T385K and p.S466R) have not been reported before in CMC patients.

Conclusion: STAT1 mutations are frequently observed in patients suffering from CMC. Thus, sequence analysis of STAT1 in CMC patients is advised. Measurement of IFN- or IL-induced STAT1 phosphorylation in PBMC provides a fast and reliable diagnostic tool and should be carried out in addition to genetic testing.

Keywords: CMC; Chronic mucocutaneous candidiasis; GOF; PID; STAT1; gain-of-function; phosphorylation; primary immunodeficiency; signal transducer and activator of transcription 1.

Fig. 1

Flow chart of STAT1 mutation detection results. The cohort consists of 92 individuals, including 18 sporadic and 39 familial patients, and 35 healthy family members

Fig. 2

GOF STAT1 mutations. Linear representation of the human STAT1 alpha isoform with GOF mutations associated with mycoses. Coding exons are numbered with roman numerals. Regions corresponding to the N-terminal segment (NTS), coiled-coil domain (CCD), DNA binding domain (DBD), linker domain (L), SH2 domain (SH2), tail segment domain (TS) and transactivator domain (TA) are indicated by rectangles. Mutations colored in blue affect the coiled-coil domain and colored in red affect the DNA-binding domain. GOF mutations in the upper part of the Figure have been published previously [–, –45]. GOF mutations listed in the lower part were found in the cohort under study. Each dot represents one patient

Fig. 3

a Increased STAT1 phosphorylation in patients’ monocytes after IFN-α and IFN-γ treatment. PBMC from patients (solid line) and healthy controls (dashed lines) were stimulated for 15 min with IFN-α (500 U/ml) or IFN-γ (100 ng/ml). Cells were fixed and permeabilized prior to staining with anti-CD14 and anti-phospho-STAT1 (pY701) antibodies. The gate was set on CD14+ monocytes. b Increased STAT1 phosphorylation in patients’ CD4+ T cells following IFN-α and IL-27 stimulation. PBMC were stimulated with either IFN-α (10,000 U/ml) or IL-27 (200 ng/ml) for 7.5, 15 and 30 min. Cells were then treated as described in (a) and phospho-STAT1 levels were evaluated gating on CD4+ T cells (Red – unstimulated patient cells; blue – stimulated healthy control cells; orange – stimulated patient cells)