Case Report: Hypomorphic Function and Somatic Reversion in DOCK8 Deficiency in One Patient With Two Novel Variants and Sclerosing Cholangitis.

DOCK8 deficiency is a combined immunodeficiency due to biallelic variants in dedicator of cytokinesis 8 (DOCK8) gene. The disease has a wide clinical spectrum encompassing recurrent infections (candidiasis, viral and bacterial infections), virally driven malignancies and immune dysregulatory features, including autoimmune (cytopenia and vasculitis) as well as allergic disorders (eczema, asthma, and food allergy). Hypomorphic function and somatic reversion of DOCK8 has been reported to result in incomplete phenotype without IgE overproduction. Here we describe a case of DOCK8 deficiency in a 8-year-old Caucasian girl. The patient's disease was initially classified as autoimmune thrombocytopenia, which then evolved toward a combined immunodeficiency phenotype with recurrent infections, persistent EBV infection and lymphoproliferation. Two novel variants (one deletion and one premature stop codon) were characterized, resulting in markedly reduced, but not absent, DOCK8 expression. Somatic reversion of the DOCK8 deletion was identified in T cells. Hypomorphic function and somatic reversion were associated with restricted T cell repertoire, decreased STAT5 phosphorylation and impaired immune synapse functioning in T cells. Although the patient presented with incomplete phenotype (absence of markedly increase IgE and eosinophil count), sclerosing cholangitis was incidentally detected, thus indicating that hypomorphic function and somatic reversion of DOCK8 may delay disease progression but do not necessarily prevent from severe complications.

Generation of 3 clones of induced pluripotent stem cells (iPSCs) from a patient affected by Autosomal Recessive Osteopetrosis due to mutations in TCIRG1 gene.

Autosomal recessive osteopetrosis (ARO) is a rare inherited disorder leading to increased bone density with impairment in bone resorption. Among the genes responsible for ARO, the TCIRG1 gene, coding for the a3 subunit of the osteoclast proton pump, is mutated in more than 50% of the cases, increasing the importance of TCIRG1-iPSCs as disease model. We generated 3 iPSC clones derived from Peripheral Blood Mononuclear Cells (PBMCs) of a patient carrying the heterozygous mutations p.Y512X and c.2236 + 1G > A. A Sendai virus-based vector was used and the iPSCs were characterized for genetic identity to parental cells, genomic integrity, pluripotency, and differentiation ability.

Generation of induced pluripotent stem cells (iPSCs) from patient with Cri du Chat Syndrome.

The Cri du Chat Syndrome (CdCS) is a genetic disease resulting from variable size deletion occurring on the short arm of chromosome 5. The main clinical features are a high-pitched monochromatic cry, microcephaly, severe psychomotor and mental retardation with characteristics of autism spectrum disorders such as hand flapping, obsessive attachments to objects, twirling objects, repetitive movements, and rocking. We reprogrammed to pluripotency peripheral blood mononuclear cells derived from a patient carrying large deletion on the short arm of chromosome 5, using a commercially available non-integrating expression system. The iPSCs expressed pluripotency markers and differentiated in the three embryonic germ layers.

Generation of three isogenic induced Pluripotent Stem Cell lines (iPSCs) from fibroblasts of a patient with Aicardi Goutières Syndrome carrying a c.2471G>A dominant mutation in IFIH1 gene.

Aicardi-Goutières syndrome (AGS) is an early-onset monogenic encephalopathy characterized by intracranial calcification, leukodystrophy and cerebrospinal fluid lymphocytosis. To date, seven genes have been related to AGS. Among these, IFIH1 encodes for MDA5, a cytosolic double-stranded RNA receptor, and is responsible for AGS type 7. We generated three isogenic iPSC clones, using a Sendai virus-based vector, starting from fibroblasts of a patient carrying a dominant mutation in IFIH1. All lines were characterized for genomic integrity, genetic uniqueness, pluripotency, and differentiation capability. Our clones might offer a good model to investigate AGS7 pathophysiological mechanism and to discover new biomarkers for this condition treatment.

Establishment of three iPSC lines from fibroblasts of a patient with Aicardi Goutières syndrome mutated in RNaseH2B.

We report the generation of three isogenic iPSC clones (UNIBSi007-A, UNIBSi007-B, and UNIBSi007-C) obtained from fibroblasts of a patient with Aicardi Goutières Syndrome (AGS) carrying a homozygous mutation in RNaseH2B. Cells were transduced using a Sendai virus based system, delivering the human OCT4, SOX2, c-MYC and KLF4 transcription factors. The resulting transgene-free iPSC lines retained the disease-causing DNA mutation, showed normal karyotype, expressed pluripotent markers and could differentiate in vitro toward cells of the three embryonic germ layers.

Generation of three iPSC lines from fibroblasts of a patient with Aicardi Goutières Syndrome mutated in TREX1.

Fibroblasts from a patient with Aicardi Goutières Syndrome (AGS) carrying a compound heterozygous mutation in TREX1, were reprogrammed into induced pluripotent stem cells (iPSCs) to establish isogenic clonal stem cell lines: UNIBSi006-A, UNIBSi006-B, and UNIBSi006-C. Cells were transduced using the episomal Sendai viral vectors, containing human OCT4, SOX2, c-MYC and KLF4 transcription factors. The transgene-free iPSC lines showed normal karyotype, expressed pluripotent markers and displayed in vitro differentiation potential toward cells of the three embryonic germ layers.

Generation of 3 clones of induced pluripotent stem cells (iPSCs) from a patient affected by Crohn's disease.

Crohn's disease is a debilitating and incurable chronic inflammatory bowel disease, affecting millions of individuals worldwide, with an increasing frequency. Surgery must be applicable in half of the cases often with a disabling course, and pharmacological treatments may have adverse complications. We generated three isogenic clones of iPSCs from peripheral blood mononuclear cells (PBMCs) of a patient with Crohn's Disease under pharmacological treatment without adverse effects. Sendai virus based vector was used and the iPSCs were characterized for genetic uniqueness, genomic integrity, pluripotency, and differentiation ability. These iPSCs will be a powerful tool to develop tailored therapies.