Homozygous variant in TKFC abolishing triokinase activities is associated with isolated immunodeficiency.

BACKGROUND: Triokinase and FMN cyclase (TKFC) is a bifunctional enzyme involved in fructose metabolism. Triokinase catalyses the phosphorylation of fructose-derived glyceraldehyde (GA) and exogenous dihydroxyacetone (DHA), while FMN cyclase generates cyclic FMN. TKFC regulates the antiviral immune response by interacting with IFIH1 (MDA5). Previously reported pathogenic variants in TKFC are associated with either a multisystemic disease or isolated hypotrichosis with loose anagen hairs. METHODS: Whole-exome sequencing identified a homozygous novel variant in TKFC (c.1624G>A; p.Gly542Arg) in an individual with a complex primary immunodeficiency disorder. The variant was characterised using enzymatic assays and yeast studies of mutant recombinant proteins. RESULTS: The individual presented with chronic active Epstein-Barr virus disease and multiple bacterial and viral infections. Clinical investigations revealed hypogammaglobulinaemia, near absent natural killer cells and decreased memory B cells. Enzymatic assays showed that this variant displayed defective DHA and GA kinase activity while maintaining FMN cyclase activity. An allogenic bone marrow transplantation corrected the patient's immunodeficiency. CONCLUSION: Our report suggests that TKFC may have a role in the immunological system. The pathological features associated with this variant are possibly linked with DHA/GA kinase inactivation through a yet an unknown mechanism. This report thus adds a possible new pathway of immunometabolism to explore further.

Generation of functional human T cell development in NOD/SCID/IL2rγnull humanized mice without using fetal tissue: Application as a model of HIV infection and persistence.

Humanization of mice with functional T cells currently relies on co-implantation of hematopoietic stem cells from fetal liver and autologous fetal thymic tissue (so-called BLT mouse model). Here, we show that NOD/SCID/IL2rγnull mice humanized with cord blood- derived CD34+ cells and implanted with allogeneic pediatric thymic tissues excised during cardiac surgeries (CCST) represent an alternative to BLT mice. CCST mice displayed a strong immune reconstitution, with functional T cells originating from CD34+ progenitor cells. They were equally susceptible to mucosal or intraperitoneal HIV infection and had significantly higher HIV-specific T cell responses. Antiretroviral therapy (ART) robustly suppressed viremia and reduced the frequencies of cells carrying integrated HIV DNA. As in BLT mice, we observed a complete viral rebound following ART interruption, suggesting the presence of HIV reservoirs. In conclusion, CCST mice represent a practical alternative to BLT mice, broadening the use of humanized mice for research.

PIGG variant pathogenicity assessment reveals characteristic features within 19 families.

PURPOSE: Phosphatidylinositol Glycan Anchor Biosynthesis, class G (PIGG) is an ethanolamine phosphate transferase catalyzing the modification of glycosylphosphatidylinositol (GPI). GPI serves as an anchor on the cell membrane for surface proteins called GPI-anchored proteins (GPI-APs). Pathogenic variants in genes involved in the biosynthesis of GPI cause inherited GPI deficiency (IGD), which still needs to be further characterized. METHODS: We describe 22 individuals from 19 unrelated families with biallelic variants in PIGG. We analyzed GPI-AP surface levels on granulocytes and fibroblasts for three and two individuals, respectively. We demonstrated enzymatic activity defects for PIGG variants in vitro in a PIGG/PIGO double knockout system. RESULTS: Phenotypic analysis of reported individuals reveals shared PIGG deficiency-associated features. All tested GPI-APs were unchanged on granulocytes whereas CD73 level in fibroblasts was decreased. In addition to classic IGD symptoms such as hypotonia, intellectual disability/developmental delay (ID/DD), and seizures, individuals with PIGG variants of null or severely decreased activity showed cerebellar atrophy, various neurological manifestations, and mitochondrial dysfunction, a feature increasingly recognized in IGDs. Individuals with mildly decreased activity showed autism spectrum disorder. CONCLUSION: This in vitro system is a useful method to validate the pathogenicity of variants in PIGG and to study PIGG physiological functions.

PIGH deficiency can be associated with severe neurodevelopmental and skeletal manifestations.

Phosphatidylinositol Glycan Anchor Biosynthesis class H (PIGH) is an essential player in the glycosylphosphatidylinositol (GPI) synthesis, an anchor for numerous cell membrane-bound proteins. PIGH deficiency is a newly described and rare disorder associated with developmental delay, seizures and behavioral difficulties. Herein, we report three new unrelated families with two different bi-allelic PIGH variants, including one new variant p.(Arg163Trp) which seems associated with a more severe phenotype. The common clinical features in all affected individuals are developmental delay/intellectual disability and hypotonia. Variable clinical features include seizures, autism spectrum disorder, apraxia, severe language delay, dysarthria, feeding difficulties, facial dysmorphisms, microcephaly, strabismus, and musculoskeletal anomalies. The two siblings homozygous for the p.(Arg163Trp) variant have severe symptoms including profound psychomotor retardation, intractable seizures, multiple bone fractures, scoliosis, loss of independent ambulation, and delayed myelination on brain MRI. Serum iron levels were significantly elevated in one individual. All tested individuals with PIGH deficiency had normal alkaline phosphatase and CD16, a GPI-anchored protein (GPI-AP), was found to be decreased by 60% on granulocytes from one individual. This study expands the PIGH deficiency phenotype range toward the severe end of the spectrum with the identification of a novel pathogenic variant.

Efficient and Robust NK-Cell Transduction With Baboon Envelope Pseudotyped Lentivector.

NK-cell resistance to transduction is a major technical hurdle for developing NK-cell immunotherapy. By using Baboon envelope pseudotyped lentiviral vectors (BaEV-LVs) encoding eGFP, we obtained a transduction rate of 23.0 ± 6.6% (mean ± SD) in freshly-isolated human NK-cells (FI-NK) and 83.4 ± 10.1% (mean ± SD) in NK-cells obtained from the NK-cell Activation and Expansion System (NKAES), with a sustained transgene expression for at least 21 days. BaEV-LVs outperformed Vesicular Stomatitis Virus type-G (VSV-G)-, RD114- and Measles Virus (MV)- pseudotyped LVs (p < 0.0001). mRNA expression of both BaEV receptors, ASCT1 and ASCT2, was detected in FI-NK and NKAES, with higher expression in NKAES. Transduction with BaEV-LVs encoding for CAR-CD22 resulted in robust CAR-expression on 38.3 ± 23.8% (mean ± SD) of NKAES cells, leading to specific killing of NK-resistant pre-B-ALL-RS4;11 cell line. Using a larger vector encoding a dual CD19/CD22-CAR, we were able to transduce and re-expand dual-CAR-expressing NKAES, even with lower viral titer. These dual-CAR-NK efficiently killed both CD19KO- and CD22KO-RS4;11 cells. Our results suggest that BaEV-LVs may efficiently enable NK-cell biological studies and translation of NK-cell-based immunotherapy to the clinic.

Polychlorinated biphenyl 126 exposure in rats alters skeletal muscle mitochondrial function.

In the past few years, polychlorinated biphenyls (PCBs), a class of environmental pollutants, have been associated with metabolism dysregulation. Muscle is one of the key regulators of metabolism because of its mass and its important role in terms of glucose consumption and glucose storage. It has been shown that muscle alterations, such as oxidative stress and mitochondrial dysfunction, contribute significantly to the development of metabolic diseases. No study has yet investigated the toxicological effect of PCBs on muscle mitochondrial function and oxidative stress in vivo. The aim of this study was to assess the effect of PCB126 in vivo exposure (single dose of 1.05 µmol/kg) on muscle mitochondrial function and oxidative stress in rats. PCB126-treated rats showed a marked increase in Cyp1a1 mRNA levels in skeletal muscles in association with a 40% reduction in state 3 oxygen consumption rate measured with complex I substrates in permeabilized muscle fibers. Furthermore, PCB126 exposure altered the expression of some enzymes involved in ROS detoxification such as catalase and glutaredoxin 2. Our results highlight for the first time a toxic effect of coplanar PCBs on skeletal muscle mitochondrial function and oxidative stress. This suggests that acute PCB exposure, by affecting muscle metabolism, could contribute to the development of metabolic disorders. Studies are needed to determine if lower-level but longer-term PCB exposure exhibits the same effect.