Germline ATG2B/GSKIP-containing 14q32 duplication predisposes to early clonal hematopoiesis leading to myeloid neoplasms.

The germline predisposition associated with the autosomal dominant inheritance of the 14q32 duplication implicating ATG2B/GSKIP genes is characterized by a wide clinical spectrum of myeloid neoplasms. We analyzed 12 asymptomatic carriers and 52 patients aged 18-74 years from six families, by targeted sequencing of 41 genes commonly mutated in myeloid malignancies. We found that 75% of healthy carriers displayed early clonal hematopoiesis mainly driven by TET2 mutations. Molecular landscapes of patients revealed two distinct routes of clonal expansion and leukemogenesis. The first route is characterized by the clonal dominance of myeloproliferative neoplasms (MPN)-driver events associated with TET2 mutations in half of cases and mutations affecting splicing and/or the RAS pathway in one-third of cases, leading to the early development of MPN, mostly essential thrombocythemia, with a high risk of transformation (50% after 10 years). The second route is distinguished by the absence of MPN-driver mutations and leads to AML without prior MPN. These patients mostly harbored a genomic landscape specific to acute myeloid leukemia secondary to myelodysplastic syndrome. An unexpected result was the total absence of DNMT3A mutations in this cohort. Our results suggest that the germline duplication constitutively mimics hematopoiesis aging by favoring TET2 clonal hematopoiesis.

Genetic screening of male patients with primary hypogammaglobulinemia can guide diagnosis and clinical management.

The precise diagnosis of an immunodeficiency is sometimes difficult to assess, especially due to the large spectrum of phenotypic variation reported among patients. Common variable immunodeficiency disorders (CVID) do not have, for a large part, an identified genetic cause. The identification of a causal genetic mutation is important to confirm, or in some cases correct, the diagnosis. We screened >150 male patients with hypogammaglobulinemia for mutations in three genes involved in pediatric X-linked primary immunoglobulin deficiency: CD40LG, SH2D1A and BTK. The SH2D1A screening allowed to reclassify two individuals with an initial CVID presentation as XLP after mutations identification. All these mutations were associated with a lack of protein expression. In addition, 4 patients with a primary diagnosis of CVID and one with a primary IgG subclass deficiency were requalified as XLA after identifying BTK mutations. Interestingly, two out of these 5 patients carried a damaging coding BTK mutation associated with a lower, but detectable, BTK expression in monocytes, suggesting that a dysfunctional protein explains the disease phenotype in these patients. In conclusion, our results advocate to include SH2D1A and BTK in newly developed targeted NGS genetic testing, to contribute to providing the most appropriate medical treatment and genetic counselling.

Regulated in development and DNA damage responses 1 (REDD1) links stress with IL-1ß-mediated familial Mediterranean fever attack through autophagy-driven neutrophil extracellular traps.

BACKGROUND: Familial Mediterranean fever (FMF) is an IL-1ß-dependent autoinflammatory disease caused by mutations of Mediterranean fever (MEFV) encoding pyrin and characterized by inflammatory attacks induced by physical or psychological stress. OBJECTIVE: We investigated the underlying mechanism that links stress-induced inflammatory attacks with neutrophil activation and release of IL-1ß-bearing neutrophil extracellular traps (NETs) in patients with FMF. METHODS: RNA sequencing was performed in peripheral neutrophils from 3 patients with FMF isolated both during attacks and remission, 8 patients in remission, and 8 healthy subjects. NET formation and proteins were analyzed by using confocal immunofluorescence microscopy, immunoblotting, myeloperoxidase-DNA complex ELISA, and flow cytometry. Samples from patients with Still's disease and bacterial infections were used also. RESULTS: The stress-related protein regulated in development and DNA damage responses 1 (REDD1) is significantly overexpressed during FMF attacks. Neutrophils from patients with FMF during remission are resistant to autophagy-mediated NET release, which can be overcome through REDD1 induction. Stress-related mediators (eg, epinephrine) decrease this threshold, leading to autophagy-driven NET release, whereas the synchronous inflammatory environment of FMF attack leads to intracellular production of IL-1ß and its release through NETs. REDD1 in autolysosomes colocalizes with pyrin and nucleotide-binding domain, leucine-rich repeat/pyrin domain-containing 3. Mutated pyrin prohibits this colocalization, leading to higher IL-1ß levels on NETs. CONCLUSIONS: This study provides a link between stress and initiation of inflammatory attacks in patients with FMF. REDD1 emerges as a regulator of neutrophil function upstream to pyrin, is involved in NET release and regulation of IL-1ß, and might constitute an important piece in the IL-1ß-mediated inflammation puzzle.

FAS promoter polymorphisms correlate with activity grade in hepatitis C patients.

OBJECTIVE: Hepatocytes are susceptible to FAS-mediated apoptosis. The impact of polymorphisms in the FAS gene on histopathological features of HCV infection was therefore investigated. DESIGN/METHODS: Three single-nucleotide polymorphisms in the FAS promoter were assessed in 190 patients with chronic hepatitis C. Associations between FAS haplotypes and fibrosis stage and activity grade were tested by univariate and multivariate analyses. RESULTS: While there was no correlation between FAS promoter genotype and fibrosis stage, patients carrying the GCA haplotype (P=0.03, Fisher's exact test) and those homozygous for the GTG haplotype (P = 0.06) tended to have lower activity scores. Logistic regression showed that these associations were independent of patient age, sex and alcohol consumption. In a logistic regression model incorporating only male gender (odds ratio 2.1, 95% confidence interval 1.1-4.1 P = 0.04), the presence of the GCA haplotype (OR 0.31 95% CI 0.13-0.78 P = 0.01), and GTG homozygosity (OR 0.26 95% CI 0.08-0.83 P = 0.02), all three factors were independently correlated with activity grade. Furthermore, the GTG haplotype appeared to have lower promoter activity than the wild type GTA haplotype in a hepatocellular carcinoma cell line. CONCLUSIONS: Genetic polymorphism in the FAS gene may account for some of the histopathological variability in chronic hepatitis C.

Deleterious genetic influence of CX3CR1 genotypes on HIV-1 disease progression.

We previously reported that patients homozygous for a specific mutation (M280) in the chemokine receptor CX3CR1 progressed to AIDS more rapidly than those with other genotypes. This deleterious effect would predict that a cohort of prevalent patients would be depleted in M280 carriers, because these patients would have disappeared before recruitment. This hypothesis is confirmed in this new study based on the French SEROCO cohort showing that patients homozygous for the M280 allele were rare among the seroprevalent group. These results may explain the conflicting results published on the impact of CX3CR1 polymorphism in seroconverters.

Cloning and functional characterization of the human fractalkine receptor promoter regions.

We have previously shown that reduced expression of the fractalkine receptor, CX3CR1, is correlated with rapid HIV disease progression and with reduced susceptibility to acute coronary events. In order to elucidate the mechanisms underlying transcriptional regulation of CX3CR1 expression, we structurally and functionally characterized the CX3CR1 gene. It consists of four exons and three introns spanning over 18 kb. Three transcripts are produced by splicing the three untranslated exons with exon 4, which contains the complete open reading frame. The transcript predominantly found in leucocytes corresponds to the splicing of exon 2 with exon 4. Transcripts corresponding to splicing of exons 1 and 4 are less abundant in leucocytes and splicing of exons 3 and 4 are rare longer transcripts. A constitutive promoter activity was found in the regions extending upstream from untranslated exons 1 and 2. Interestingly, exons 1 and 2 enhanced the activity of their respective promoters in a cell-specific manner. These data show that the CX3CR1 gene is controlled by three distinct promoter regions, which are regulated by their respective untranslated exons and that lead to the transcription of three mature messengers. This highly complex regulation may allow versatile and precise expression of CX3CR1 in various cell types.