A founder truncating variant in GDF1 causes autosomal-recessive right isomerism and associated congenital heart defects in multiplex Arab kindreds.

The genetic basis of congenital heart malformations associated with disruption of left-right (L-R) asymmetry is broad and heterogenous, with variants in over 25 genes implicated thus far. Of these, deleterious variants in the Growth/Differentiation Factor 1 (GDF1) gene have been shown to cause heterotaxy with varied complex heart malformations of left-right patterning, in 23 individuals reported to date, either in monoallelic or biallelic state. We report three unrelated individuals exhibiting right isomerism with congenital heart defects, each originating from a consanguineous kindred of Arab-Muslim descent. Using whole exome sequencing, a shared novel homozygous truncating c.608G > A (p.W203*) variant in the GDF1 gene was revealed as the molecular basis of their disease. Subsequently, targeted sequencing of this variant showed full segregation with the disease in these families, with a total of over 15 reportedly affected individuals, enabling genetic counseling, prenatal diagnosis, and planning of future pregnancies. Our findings further confirm the association of biallelic GDF1 variants, heterotaxy and congenital heart defects of left-right patterning, and expand the previously described phenotypic spectrum and mutational profile. Moreover, we suggest targeted screening for the p.W203* variant in relevant clinical circumstances.

SurviveAI: Long Term Survival Prediction of Cancer Patients Based on Somatic RNA-Seq Expression.

Motivation: Prediction of cancer outcome is a major challenge in oncology and is essential for treatment planning. Repositories such as The Cancer Genome Atlas (TCGA) contain vast amounts of data for many types of cancers. Our goal was to create reliable prediction models using TCGA data and validate them using an external dataset. Results: For 16 TCGA cancer type cohorts we have optimized a Random Forest prediction model using parameter grid search followed by a backward feature elimination loop for dimensions reduction. For each feature that was removed, the model was retrained and the area under the curve of the receiver operating characteristic (AUC-ROC) was calculated using test data. Five prediction models gave AUC-ROC bigger than 80%. We used Clinical Proteomic Tumor Analysis Consortium v3 (CPTAC3) data for validation. The most enriched pathways for the top models were those involved in basic functions related to tumorigenesis and organ development. Enrichment for 2 prediction models of the TCGA-KIRP cohort was explored, one with 42 genes (AUC-ROC = 0.86) the other is composed of 300 genes (AUC-ROC = 0.85). The most enriched networks for both models share only 5 network nodes: DMBT1, IL11, HOXB6, TRIB3, PIM1. These genes play a significant role in renal cancer and might be used for prognosis prediction and as candidate therapeutic targets. Availability And Implementation: The prediction models were created and tested using Python SciKit-Learn package. They are freely accessible via a friendly web interface we called surviveAI at https://tinyurl.com/surviveai.

Transcriptomic profiling and genomic mutational analysis of Human coronavirus (HCoV)-229E -infected human cells.

Human coronaviruses (HCoVs) cause mild to severe respiratory infection. Most of the common cold illnesses are caused by one of four HCoVs, namely HCoV-229E, HCoV-NL63, HCoV-HKU1 and HCoV-OC43. Several studies have applied global transcriptomic methods to understand host responses to HCoV infection, with most studies focusing on the pandemic severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome CoV (MERS-CoV) and the newly emerging SARS-CoV-2. In this study, Next Generation Sequencing was used to gain new insights into cellular transcriptomic changes elicited by alphacoronavirus HCoV-229E. HCoV-229E-infected MRC-5 cells showed marked downregulation of superpathway of cholesterol biosynthesis and eIF2 signaling pathways. Moreover, upregulation of cyclins, cell cycle control of chromosomal replication, and the role of BRCA1 in DNA damage response, alongside downregulation of the cell cycle G1/S checkpoint, suggest that HCoV-229E may favors S phase for viral infection. Intriguingly, a significant portion of key factors of cell innate immunity, interferon-stimulated genes (ISGs) and other transcripts of early antiviral response genes were downregulated early in HCoV-229E infection. On the other hand, early upregulation of the antiviral response factor Apolipoprotein B mRNA editing enzyme catalytic subunit 3B (APOBEC3B) was observed. APOBEC3B cytidine deaminase signature (C-to-T) was previously observed in genomic analysis of SARS-CoV-2 but not HCoV-229E. Higher levels of C-to-T mutations were found in countries with high mortality rates caused by SARS-CoV-2. APOBEC activity could be a marker for new emerging CoVs. This study will enhance our understanding of commonly circulating HCoVs and hopefully provide critical information about still-emerging coronaviruses.

A single center experience with publicly funded clinical exome sequencing for neurodevelopmental disorders or multiple congenital anomalies.

Exome sequencing (ES) is an important diagnostic tool for individuals with neurodevelopmental disorders (NDD) and/or multiple congenital anomalies (MCA). However, the cost of ES limits the test's accessibility for many patients. We evaluated the yield of publicly funded clinical ES, performed at a tertiary center in Israel, over a 3-year period (2018-2020). Probands presented with (1) moderate-to-profound global developmental delay (GDD)/intellectual disability (ID); or (2) mild GDD/ID with epilepsy or congenital anomaly; and/or (3) MCA. Subjects with normal chromosomal microarray analysis who met inclusion criteria were included, totaling 280 consecutive cases. Trio ES (proband and parents) was the default option. In 252 cases (90.0%), indication of NDD was noted. Most probands were males (62.9%), and their mean age at ES submission was 9.3 years (range 1 month to 51 years). Molecular diagnosis was reached in 109 probands (38.9%), mainly due to de novo variants (91/109, 83.5%). Disease-causing variants were identified in 92 genes, 15 of which were implicated in more than a single case. Male sex, families with multiple-affected members and premature birth were significantly associated with lower ES yield (p < 0.05). Other factors, including MCA and coexistence of epilepsy, autism spectrum disorder, microcephaly or abnormal brain magnetic resonance imaging findings, were not associated with the yield. To conclude, our findings support the utility of clinical ES in a real-world setting, as part of a publicly funded genetic workup for individuals with GDD/ID and/or MCA.

BRPF1-associated intellectual disability, ptosis, and facial dysmorphism in a multiplex family.

BACKGROUND: Over 500 epigenetic regulators have been identified throughout the human genome. Of these, approximately 30 chromatin modifiers have been implicated thus far in human disease. Recently, variants in BRPF1, encoding a chromatin reader, have been associated with a previously unrecognized autosomal dominant syndrome manifesting with intellectual disability (ID), hypotonia, dysmorphic facial features, ptosis, and/or blepharophimosis in 22 individuals. PATIENTS AND METHODS: We report a multiply affected nonconsanguineous family of mixed Jewish descent who presented due to ID in three male siblings. Molecular analysis of the family was pursued using whole exome sequencing (WES) and subsequent Sanger sequencing. RESULTS: Whole exome sequencing analysis brought to the identification of a novel heterozygous truncating mutation (c.556C>T, p.Q186*) in the BRPF1 gene in the affected siblings and their mother. The four affected individuals showed varying degrees of intellectual disability, distinct facial features including downslanted palpebral fissures, ptosis, and/or blepharophimosis. Their clinical characteristics are discussed in the context of previously reported patients with the BRPF1-related phenotype. CONCLUSION: The reported family contributes to the current knowledge regarding this unique and newly recognized genetic disorder, and further implicates the role of BRPF1 in human brain development.

Rare homozygosity in amyotrophic lateral sclerosis suggests the contribution of recessive variants to disease genetics.

OBJECTIVE: to determine the occurrence of homozygous rare, in-silico damaging variants in a genetically relatively homogenous group of amyotrophic lateral sclerosis (ALS) patients. METHODS: Whole-exome-sequencing of 43 ALS patients of North-Africa Jewish origin was performed. Data were filtered to identify very rare homozygous recessive in-silico damaging variants, in genes annotated to ALS-associated cellular pathways. RESULTS: We identified a rare missense homozygous variant, p.Arg663Cys in MFN2, predicted to be damaging, in a patient with an early age at disease onset (36 years) and fast progression. An additional ALS patient carried the mutation and together established its association to ALS (p = .01). Additional homozygous variants were identified, including the risk allele p.Arg261His in NEK1, as well as variants in genes known to be associated with other neurodegenerative diseases, such as HTT (Huntington's disease), ATM (Ataxia-Telangiectasia), and ZFYVE26 (SPG15), and variants in genes previously reported as upregulated (LZTS3) or downregulated (ARMC4, CFAP54, and MTHFSD) in ALS patients. Altogether, 13 patients (30%) carried at least one homozygous rare in-silico damaging variant, of them 10 carried either another rare homozygous variant and/or a variant in a known ALS gene, which is categorized as pathogenic, likely-pathogenic or variant of uncertain significance. CONCLUSIONS: Our results suggest the contribution of recessive alleles to ALS and the possibility of burden of mutations, emphasizing the complexity of ALS genetics.

Reduced Function and Diversity of T Cell Repertoire and Distinct Clinical Course in Patients With IL7RA Mutation.

The alpha subunit of IL-7 receptor (IL7R7α) is critical for the differentiation of T cells, specifically for the development and maintenance of γδT cells. Mutations in IL7RA are associated with Severe Combined Immunodeficiency (SCID). Infants with IL7RA deficiency can be identified through newborn screening program. We aimed at defining the immunological and genetic parameters that are directly affected by the IL7RA mutation on the immune system of five unrelated patients which were identified by our newborn screening program for SCID. The patients were found to have a novel identical homozygote mutation in IL7RA (n.c.120 C>G; p.F40L). Both surface expression of IL7Rα and functionality of IL-7 signaling were impaired in patients compared to controls. Structural modeling demonstrated instability of the protein structure due to the mutation. Lastly the TRG immune repertoire of the patients showed reduced diversity, increased clonality and differential CDR3 characteristics. Interestingly, the patients displayed significant different clinical outcome with two displaying severe clinical picture of immunodeficiency and three had spontaneous recovery. Our data supports that the presented IL7RA mutation affects the IL-7 signaling and shaping of the TRG repertoire, reinforcing the role of IL7RA in the immune system, while non-genetic factors may exist that attribute to the ultimate clinical presentation and disease progression.

High frequency of C9orf72 hexanucleotide repeat expansion in amyotrophic lateral sclerosis patients from two founder populations sharing the same risk haplotype.

We characterized the C9orf72 hexanucleotide repeat expansion (RE) mutation in amyotrophic lateral sclerosis (ALS) patients of 2 distinct origins, Ashkenazi and North Africa Jews (AJ, NAJ), its frequency, and genotype-phenotype correlations. In AJ, 80% of familial ALS (fALS) and 11% of sporadic ALS carried the RE, a total of 12.9% of all AJ-ALS compared to 0.3% in AJ controls (odds ratio [OR] = 44.3, p < 0.0001). In NAJ, 10% of fALS and 9% of sporadic ALS carried the RE, a total of 9.1% of all NAJ-ALS compared to 1% in controls (OR = 9.9, p = 0.0006). We identified a risk haplotype shared among all ALS patients, although an association with age at disease onset, fALS, and dementia were observed only in AJ. Variations were identified downstream the repeats. The risk haplotype and these polymorphisms were at high frequencies in alleles with 8 repeats or more, suggesting sequence instability. The different genotype-phenotype correlations and OR, together with the large range in age at onset, suggest that other modifiers and risk factors may affect penetrance and phenotype in ALS.

OPTN 691_692insAG is a founder mutation causing recessive ALS and increased risk in heterozygotes.

OBJECTIVE: To detect genetic variants underlying familial and sporadic amyotrophic lateral sclerosis (ALS). METHODS: We analyzed 2 founder Jewish populations of Moroccan and Ashkenazi origins and ethnic matched controls. Exome sequencing of 2 sisters with ALS from Morocco was followed by genotyping the identified causative null mutation in 379 unrelated patients with ALS and 1,000 controls. The shared risk haplotype was characterized using whole-genome single nucleotide polymorphism array. RESULTS: We identified 5 unrelated patients with ALS homozygous for the null 691_692insAG mutation in the optineurin gene (OPTN), accounting for 5.8% of ALS of Moroccan origin and 0.3% of Ashkenazi. We also identified a high frequency of heterozygous carriers among patients with ALS, 8.7% and 2.9%, respectively, compared to 0.75% and 1.0% in controls. The risk of carriers for ALS was significantly increased, with odds ratio of 13.46 and 2.97 in Moroccan and Ashkenazi Jews, respectively. We determined that 691_692insAG is a founder mutation in the tested populations with a minimal risk haplotype of 58.5 Kb, encompassing the entire OPTN gene. CONCLUSIONS: Our data show that OPTN 691_692insAG mutation is a founder mutation in Moroccan and Ashkenazi Jews. This mutation causes autosomal recessive ALS and significantly increases the risk to develop the disease in heterozygous carriers, suggesting both a recessive mode of inheritance and a dominant with incomplete penetrance. These data emphasize the important role of OPTN in ALS pathogenesis, and demonstrate the complex genetics of ALS, as the same mutation leads to different phenotypes and appears in 2 patterns of inheritance.

Cellularized biosynthetic microhydrogel polymers for intravascular liver tissue regeneration therapy.

INTRODUCTION: The liver is the natural microenvironment for hepatocytes transplantation but unfortunately engraftment efficiency is low. Cell-laden microhydrogels made of fibrinogen attached to poly(ethylene glycol) (PEG)-diacrylate side chains, were used as a cell carrier, for intravascular transplantation. This approach may reduce shear stress and immediate immunological pressure after intravascular transplantation and provide biomatrix for environmental support. AIMS: In vitro assessment of HuH-7 viability and function after polymerization within PEGylated fibrinogen-hydrogel. In vivo assessment of intraportal transplantation of cell-laden microhydrogels with rat adult parenchymal cells. METHODS: (1) In vitro assessment of HuH-7 cell viability and function, after cell-laden hydrogel (hydrogel volume 30 µL) fabrication, by propidium iodide (PI)/fluorescein diacetate (FDA), and MTT assays, albumin concentration and CYP1A activity. (2) Fabrication of cell-laden microhydrogels and their intraportal transplantion. Engraftment efficiency in vivo was evaluated by real-time qPCR of Y chromosome (SRY gene) and histology. RESULTS: The viability of cells in hydrogels in culture was comparable to viability of not embedded cells during the first 48 h. However, the viability of cells in hydrogels was reduced after 72 h compared with not embedded cells. Activity of CYP1A in hydrogel was comparable to that of not embedded cells (4.33±1 pmole/µg DNA/4 h vs. 5.13±1 pmole/µg DNA/4 h, respectively). Albumin concentration increased at day 3 in hydrogels to 1.4±0.6 µg/10(4)/24 h and was greater to that of free cells, 0.3±0.1 µg/10(4)/24 h. Cell-laden microhydrogels at a size of 150-150-600 µm (6×10(6) cells/rat) showed better engraftment efficiency at 21 days post-transplantation, compared with isolated cell transplantation (54.6%±5% vs. 1.8%±1.2%, p<0.001). CONCLUSIONS: The in vitro HuH-7 viability and function after polymerization in PEGylated fibrinogen hydrogel was comparable to cells without the hydrogel. Long-term survival and engraftment efficiency of intravascular transplanted adult hepatocytes is much better in within cell-laden microhydrogels compared with isolated cells. The overall efficiency of the procedure needs to be improved.