Customer Publications in October 2021

         October 28, 2021

The customer-published articles this October cite the range of Golden Helix’s VarSeq annotation range and capability. The following publications feature everything from annotating an Italian nobleman mummy, assisting in identifying mutations in primary congenital and juvenile glaucoma, new mutations associated with muscular dystrophinopathy, and germline variants associated with head and neck cancer. In each of these cases, VarSeq was utilized for annotating and calling variants allowing for clear and efficient workflow in the respective pipelines.

Whole-exome sequencing of the mummified remains of Cangrande della Scala (1291–1329 CE) indicates the first known case of late-onset Pompe disease

Mummified remains of relevant historical figures are nowadays an important source of information to retrace data concerning their private life and health, especially when historical archives are not available. Next-generation-sequencing was proved to be a valuable tool to unravel the characteristics of these individuals through their genetic heritage. Using the strictest criteria currently available for the validation of ancient DNA sequences, whole-genome and whole-exome sequencing were generated from the mummy remains of an Italian nobleman died almost 700 years ago, Cangrande della Scala. While its genome sequencing could not yield sufficient coverage for in depth investigation, exome sequencing could overcome the limitations of this approach to achieve significantly high coverage on coding regions, thus allowing to perform the first extensive exome analysis of a mummy genome. Similar to a standard “clinical exome analysis” conducted on modern DNA, an in-depth variant annotation, high-quality filtering and interpretation was performed, leading to the identification of a genotype associated with late-onset Pompe disease (glycogen storage disease type II). This genetic diagnosis was concordant with the limited clinical history available for Cangrande della Scala, who likely represents the earliest known case of this autosomal recessive metabolic disorder.

Iadarola, B., Lavezzari, D., Modi, A. et al. Whole-exome sequencing of the mummified remains of Cangrande della Scala (1291–1329 CE) indicates the first known case of late-onset Pompe disease. Sci Rep 11, 21070 (2021).

Exome sequencing identifies procollagen‑lysine 2‑oxoglutarate 5‑dioxygenase 2 mutations in primary congenital and juvenile glaucoma

To report the association of procollagen-lysine 2-oxoglutarate 5-dioxygenase 2 (PLOD2) mutations with bilateral primary congenital glaucoma (PCG) in monozygotic twins and with nondominant juvenile-onset primary open-angle glaucoma (JOAG). We utilized family-based whole-exome sequencing to detect disease-causing mutations in a pair of monozygotic twins with de-novo PCG and compared its existence in 50 nonfamilial cases of JOAG and 30 healthy controls. To validate the identified mutations, direct Sanger sequencing was performed. For further evaluation of gene expression in the ocular tissues, we performed whole-mount in situ hybridization in zebrafish embryos.We identified a novel missense mutation (c.1925A>G, p.Tyr642Cys) in the PLOD2 gene in the monozygotic twin pair with PCG and another missense mutation (c.1880G>A, p.Arg627Gln) in one JOAG patient. Both mutations identified were heterozygous. Neither the parents of the twins nor the parents of the JOAG patient harbored the mutation and it was probably a de-novo change. The zebrafish in situ hybridization revealed expression of the PLOD2 gene during embryogenesis of the eye. We observed an association of PLOD2 mutations with PCG and with nonfamilial JOAG. This new gene needs to be further investigated for its role in pathways associated with glaucoma pathogenesis.

Gupta, Viney,; Somarajan, Bindu I; Kaur, Gagandeep1; Gupta, Shikha; Singh, Renu; Pradhan, Dibyabhaba2; Singh, Harpreet2; Kaur, Punit3; Sharma, Anshul5; Chawla, Bindia4; Pahuja, Anisha4; Ramachandran, Rajesh4; Sharma, Arundhati5 Exome sequencing identifies procollagen-lysine 2-oxoglutarate 5-dioxygenase 2 mutations in primary congenital and juvenile glaucoma, Indian Journal of Ophthalmology: October 2021 – Volume 69 – Issue 10 – p 2710-2716 doi: 10.4103/ijo.IJO_1750_21

Genetic Profile of the Dystrophin Gene Reveals New Mutations in Colombian Patients Affected with Muscular Dystrophinopathy

Duchenne and Becker muscular dystrophies (DMD/BMD) are the most common human dystrophinopathies with recessive X-linked inheritance. Dystrophin gene deletions and duplications are the most common mutations, followed by point mutations. The aim of this study is to characterize the mutational profile of the dystrophin gene in Colombian patients with DMD/BMD. Mutational profiling was determined in 69 affected patients using Sanger sequencing, next-generation sequencing (NGS) and/or multiplex ligation dependent-probes amplification (MLPA). Genetic variants were classified according to molecular consequence and new variants were determined through database and literature analysis. Mutational profile in affected patients revealed that large deletions/duplications analyzed by MLPA accounted for 72.5% of all genetic variations. By using Sanger sequencing or NGS, we identified point mutations in 15.9% and small deletions in 11.6% of the patients. New mutations were found, most of them were point mutations or small deletions (10.1%). Our results described the genetic profile of the dystrophin gene in Colombian patients with DMD and contribute to efforts to identify molecular variants in Latin American populations. For our population, 18.8% of cases could be treated with FDA or MDA approved molecular therapies based on specific mutations. These data contribute to the establishment of appropriate genetic counseling and potential treatment.

Triana-Fonseca P, Parada-Márquez JF, Silva-Aldana CT, Zambrano-Arenas D, Arias-Gomez LL, Morales-Fonseca N, Medina-Méndez E, Restrepo CM, Silgado-Guzmán DF, Fonseca-Mendoza DJ. Genetic Profile of the Dystrophin Gene Reveals New Mutations in Colombian Patients Affected with Muscular Dystrophinopathy. Appl Clin Genet. 2021;14:399-408

Germline variants in DNA repair genes are associated with young-onset head and neck cancer

The genetic predisposition to head and neck carcinomas (HNSCC) and how the known risk factors (papillomavirus infection, alcohol, and tobacco consumption) contribute to the early-onset disease are barely explored. Although HNSCC at early onset is rare, its frequency is increasing in recent years. Germline and somatic variants were assessed to build a comprehensive genetic influence pattern in HNSCC predisposition and patient outcome. Whole-exome sequencing was performed in 45 oral and oropharynx carcinomas paired with normal samples of young adults (≤49 years). We found FANCG, CDKN2A, and TPP germline variants previously associated with HNSCC risk. At least one germline variant in DNA repair pathway genes was detected in 67% of cases. Germline and somatic variants (including copy number variations) in FAT1 gene were identified in 9 patients (20%) and 12 tumors (30%), respectively. Somatic variants were found in HNSCC associated genes, such as TP53, CDKN2A, and PIK3CA. To date, 55 of 521 cases from the large cohort of TCGA presented < 49 years old. A comparison between the somatic alterations of TCGA-HNSCC at early onset and our dataset revealed strong similarities. Protein-protein interaction analysis between somatic and germline altered genes revealed a central role of TP53. Altogether, germline alterations in DNA repair genes potentially contribute to an increased risk of developing HNSCC at early-onset, while FAT1 could impact the prognosis.

Sarah Santiloni Cury, Priscila Mayrink de Miranda, Fabio Albuquerque Marchi, Luisa Matos do Canto, Thiago Celestino Chulam, Annabeth Høgh Petersen, Mads M. Aagaard, Clóvis Antonio Lopes Pinto, Luiz Paulo Kowalski, Silvia Regina Rogatto, Germline variants in DNA repair genes are associated with young-onset head and neck cancer, Oral Oncology, Volume 122, 2021, 105545, ISSN 1368-8375,

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