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Purpose: The aim of this study was to determine the clinical and molecular characteristics of 2,079 patients who underwent hereditary cancer multigene panel testing. Methods: Panels included comprehensive analysis of 14–22 cancer susceptibility genes ( BRCA1 and BRCA2 not included), depending on the panel ordered (BreastNext, OvaNext, ColoNext, or CancerNext). Next-generation sequencing and deletion/duplication analyses were performed for all genes except EPCAM (deletion/duplication analysis only). Clinical histories of ColoNext patients harboring mutations in genes with well-established diagnostic criteria were assessed to determine whether diagnostic/testing criteria were met. Results: Positive rates were defined as the proportion of patients with a pathogenic mutation/likely pathogenic variant(s) and were as follows: 7.4% for BreastNext, 7.2% for OvaNext, 9.2% for ColoNext, and 9.6% for CancerNext. Inconclusive results were found in 19.8% of BreastNext, 25.6% of OvaNext, 15.1% of ColoNext, and 23.5% of CancerNext tests. Based on information submitted by clinicians, 30% of ColoNext patients with mutations in genes with well-established diagnostic criteria did not meet corresponding criteria. Conclusion: Our data point to an important role for targeted multigene panels in diagnosing hereditary cancer predisposition, particularly for patients with clinical histories spanning several possible diagnoses and for patients with suspicious clinical histories not meeting diagnostic criteria for a specific hereditary cancer syndrome. Genet Med 16 11, 830–837.

Background Panel‐based targeted exome sequencing was used to analyze the genetic and clinical findings of targeted genes in a cohort of northeast Chinese with retinitis pigmentosa. Methods A total of 87 subjects, comprising 23 probands and their family members (total patients: 32) with confirmed retinitis pigmentosa were recruited in the study. Panel‐based targeted exome sequencing was used to sequence the patients and family members, all subjects with retinitis pigmentosa underwent a complete ophthalmologic examination. Results Of the 23 probands, the clinical manifestations include night blindness, narrowing of vision, secondary cataracts, choroidal atrophy, color blindness, and high myopia, the average age of onset of night blindness is 12.9 ± 14 (range, 0–65; median, 8). Posterior subcapsular opacities is the most common forms of secondary cataracts (nine cases, 39.1%), and peripheral choroidal atrophy is the most common form of secondary choroidal atrophy (12 cases, 52.2%). Of these probands with complication peripheral choroidal atrophy, there were eight probands (66.7%, 8/12) caused by the pathogenic variation in USH2A gene. A total of 17 genes and 45 variants were detected in 23 probands. Among these genes, the commonest genes were USH2A (40%; 18/45), RP1 (15.6%; 7/45), and EYS (8.9%; 4/45), and the top three genes account for 56.5% (13/23) of diagnostic probands. Among these variants, comprising 22 (48.9%) pathogenic variants, 14 (31%) likely pathogenic variants, and nine (20%) uncertain clinical significance variants, and 22 variants was discovered first time. Most of the mutations associated with RP were missense (53.3%, 24/45), and the remaining mutation types include frameshift (35.6%, 16/45), nonsense (6.7%, 3/45), and spliceSite (4.4%, 2/45). Among the probands with mutations detected, compound heterozygous forms was detected in 13 (56.5%, 13/23) probands, and digenic inheritance (DI) forms was detected in five (21.7%, 5/23) probands. Conclusion Panel‐based targeted exome sequencing revealed 23 novel mutations, recognized different combinations forms of variants, and extended the mutational spectrum of retinitis pigmentosa and depicted common variants in northeast China. We applied panel‐based targeted exome sequencing to explore the pathogenic variation spectrum and genetic characteristics of retinitis pigmentosa populations in northeast China, and have a deeper understanding of the relationship between clinical manifestations and genotypes.

Inherited retinal dystrophies (IRDs) are a group of clinically and genetically heterogeneous diseases involving more than 280 genes and no less than 20 different clinical phenotypes. In this study, our aims were to identify the disease-causing gene variants of 319 Chinese patients with IRD, and compare the pros and cons of targeted panel sequencing and whole exome sequencing (WES). Patients were assigned for analysis with a hereditary eye disease enrichment panel (HEDEP) or WES examination based on time of recruitment. This HEDEP was able to capture 441 hereditary eye disease genes, which included 291 genes related to IRD. As RPGR ORF15 was difficult to capture, all samples were subjected to Sanger sequencing for this region. Among the 163 disease-causing variants identified in this study, 73 had been previously reported, and the other 90 were novel. Genes most commonly implicated in different inheritances of IRDs in this cohort were presented. HEDEP and WES achieved diagnostic yield with 41.2% and 33.0%, respectively. In addition, nine patients were found to carry pathogenic mutations in the RPGR ORF15 region with Sanger sequencing. Our study demonstrates that HEDEP can be used as a first-tier test for patients with IRDs.

Hyper-IgE syndromes and chronic mucocutaneous candidiasis constitute rare primary immunodeficiency syndromes with an overlapping clinical phenotype. In recent years, a growing number of underlying genetic defects have been identified. To characterize the underlying genetic defects in a large international cohort of 275 patients, of whom 211 had been clinically diagnosed with hyper-IgE syndrome and 64 with chronic mucocutaneous candidiasis, targeted panel sequencing was performed, relying on Agilent HaloPlex and Illumina MiSeq technologies. The targeted panel sequencing approach allowed us to identify 87 (32 novel and 55 previously described) mutations in 78 patients, which generated a diagnostic success rate of 28.4%. Specifically, mutations in DOCK8 (26 patients), STAT3 (21), STAT1 (15), CARD9 (6), AIRE (3), IL17RA (2), SPINK5 (3), ZNF341 (2), CARMIL2/RLTPR (1), IL12RB1 (1), and WAS (1) have been detected. The most common clinical findings in this cohort were elevated IgE (81.5%), eczema (71.7%), and eosinophilia (62.9%). Regarding infections, 54.7% of patients had a history of radiologically proven pneumonia, and 28.3% have had other serious infections. History of fungal infection was noted in 53% of cases and skin abscesses in 52.9%. Skeletal or dental abnormalities were observed in 46.2% of patients with a characteristic face being the most commonly reported feature (23.1%), followed by retained primary teeth in 18.9% of patients. Targeted panel sequencing provides a cost-effective first-line genetic screening method which allows for the identification of mutations also in patients with atypical clinical presentations and should be routinely implemented in referral centers.

Unintended genetic modifications that occur during the differentiation and proliferation of human induced pluripotent stem cells (hiPSCs) can lead to tumorigenicity. This is a crucial concern in the development of stem cell-based therapies to ensure the safety and efficacy of the final product. Moreover, conventional genetic stability testing methods are limited by low sensitivity, which is an issue that remains unsolved. In this study, we assessed the genetic stability of hiPSCs and hiPSC-derived cardiomyocytes using various testing methods, including karyotyping, CytoScanHD chip analysis, whole-exome sequencing, and targeted sequencing. Two specific genetic mutations in KMT2C and BCOR were selected from the 17 gene variants identified by whole-exome and targeted sequencing methods, which were validated using droplet digital PCR. The applicability of this approach to stem cell-based therapeutic products was further demonstrated with associated validation according to the International Council for Harmonisation (ICH) guidelines, including specificity, precision, robustness, and limit of detection. Our droplet digital PCR results showed high sensitivity and accuracy for quantitatively detecting gene mutations, whereas conventional qPCR could not avoid false positives. In conclusion, droplet digital PCR is a highly sensitive and precise method for assessing the expression of mutations with tumorigenic potential for the development of stem cell-based therapeutics.

Background Age‐related cataracts (ARC) is the most common blinding eye disease worldwide, and its incidence tend to become younger. However, the relationship between genetic factors and mechanisms is not fully understood. The aim of the study was to further clarify the relationship between ARC and genetic mechanisms in East Asian populations and to elucidate the pathogenesis. Methods The study collected 191 sporadic cataracts and 208 healthy people from the eastern provinces of China, with an average age of about 60 years. All participants were subjected to a comprehensive ophthalmic clinical examination and peripheral blood samples were collected and their genomic DNA was extracted. Mutations were screened among 792 candidate genes to enhance understanding of the disease through targeted capture and high‐throughput sequencing. Results We identified novel candidate susceptibility gene, which may serve as a potential susceptibility factor leading to an increase in the incidence of age‐related cataracts. Three novel loci are associated with age‐related cataracts significant significance: rs129882 in DBH (p = 5.27E‐07, odds ratio = 3.9), rs1800280 in DMD (p = 2.85E‐06, odds ratio = 1.4) and rs2871776 in ATP13A2 (p = 4.18E‐05, odds ratio = 0.04). Gene–gene interaction analysis revealed that the most significant interactions between genes include the interaction between DBH and TUB (rs17847537 in TUB, rs129882 in DBH, p‐value = 2.12E‐14), and the interaction between DBH and DMD (rs1800280 in DMD, rs129882 in DBH, p‐value = 2.12E‐14). Pathway analysis shows that the most significant processes are concentrated in response to light stimulation (adjusted p‐Value = 5.56E‐03), response to radiation (adjusted P‐Value = 5.56E‐03), abiotic stimulus (adjusted p‐Value = 5.56E‐03). eQTL analysis shows that DBH rs129882 could regulate the expression of DBH mRNA in various tissues including retina. Conclusion Our study indicates rs129882 and rs1800280 loci are associated with age‐related cataracts, which enlarge the gene map of age‐related cataracts. We identified novel candidate susceptibility gene, which may serve as a potential susceptibility factor leading to an increase in the incidence of age‐related cataracts. Our study indicates rs129882 and rs1800280 loci are associated with age‐related cataracts, which enlarge the gene map of age‐related cataracts.

Background Breast cancer is one of the leading causes of cancer-related deaths in women, and there is a demand in developing an Asian-based genetic profiling database for breast cancer in improving the treatment response. This study aimed to determine molecular alternations and identify potential therapeutic targets by analyzing the genetic profiling from a cohort of Taiwanese breast cancers using a commercialized next-generation sequencing (NGS) targeted panel. Methods The study population comprised a broad spectrum of breast cancer patients in Taiwan, including Group 1: planned to receive first-line surgery and followed by adjuvant therapy, or early relapse within three years, Group 2: planned to receive first-line neoadjuvant therapy and followed by surgery, and Group 3: de novo stage IV, or stage IV with recurrence beyond three years. Molecular profiles were determined using Thermo Fisher™ Oncomine™ Comprehensive Assay version 3 (TMO comprehensive assay) from Formalin-Fixed Paraffin-Embedded (FFPE) tissues. Level of actionability was evaluated with the ESMO Scale of clinical actionability of molecular targets (ESCAT). Results A total of 380 TMO comprehensive assays were conducted on 372 patients, and we presented targeted sequencing analyses of Tier I: alteration-drug match associated with improved outcome in clinical trials including ERBB2 amplification, BRCA1/2 germline mutation, PIK3CA mutation, and NTRK translocation, and Tier II: antitumor activity associated with the matched alteration-drug but lack of prospective outcome data including PTEN loss, ESR1 mutation, AKT1 mutation, and ERBB2 mutation, and Tier III: matched drug-alteration that led to clinical benefit in another tumor type including MDM2 amplification, and ERBB3 mutation. Among them, 249 (66%) showed at least one actionable alternation based on the ESCAT criteria. The most frequent impacted genes (all variants combined within each sample) were PIK3CA (38%), followed by ERBB2 (23%), ESR1 (10%), AKT1 (6%), and BRCA2 (5%), and the remaining rare variants (less than 5% of assayed cohort) were BRCA1 (3%), MDM2 (2.2%), and ERBB3 (1.1%). Conclusion Targeted sequencing of actionable genes is believed to provide clinical applicability and substantial benefits for Taiwanese breast cancer patients. A valid scale of clinical actionability should be adopted for precision medicine practice under multidisciplinary molecular tumor board.

Background Panel‐based targeted exome sequencing was applied to identify the pathogenic variants and genetic characteristics of retinitis pigmentosa (RP) in two Chinese families, and to gain a deeper understanding of the relationship between clinical manifestations and genotypes. Methods A total of 17 subjects, comprising two probands (total patients: four subjects) and their family member, were recruited in this study. All subjects underwent comprehensive ophthalmic examinations and clinical evaluations, and the complete history and medical records were collected according to the standard procedures. All participants were screened using the multigene panel test (Target_Eye_792_V2 chip), and Sanger sequencing was used to confirm the candidate variants. Results Among these two families, a total of three novel mutations in the EYS gene were identified in patients, including a homozygous frameshift mutation c.9252_9253insT detected in two patients in one family, and the compound heterozygous splicesite mutation c.5644+2T>C and frameshift mutation c.1920_1923delTGAG detected in two patients in the another family. All patients in both families had early onset of night blindness and poor visual acuity, and with typical posterior capsule opacification. The mutation co‐segregated within all recruited individuals. In addition, one patient with compound heterozygous mutations was found to have typical blue‐blindness symptoms and detected a previously reported disease‐causing mutation c.235G>A in OPN1SW gene, which caused blue blindness manifestations and was first discovered in patient combined with RP causative genes. Conclusions Panel‐based targeted exome sequencing was used to identify three novel variants of RP causative gene, and we also detected a known pathogenic variants of blue‐blindness causative genes in two patients. Our finding will provide a powerful basis for genetic counseling and enhance our current understanding of the genetics factors for RP families. We applied panel‐based targeted exon sequencing to explore the pathogenic variation spectrum and genetic characteristics of retinitis pigmentosa populations in two Chinese families, and have a deeper understanding of the relationship between clinical manifestations and genotypes.

Next-generation sequencing (NGS), comprising targeted panels (TP), exome sequencing (ES), and genome sequencing (GS) became robust clinical tools for diagnosing hereditary ataxia (HA). Determining their diagnostic yield (DY) is crucial for optimal clinical decision-making. We conducted a comprehensive systematic literature review on the DY of NGS tests for HA. We searched PubMed and Embase databases for relevant studies between 2016 and 2022 and manually examined reference lists of relevant reviews. Eligible studies described the DY of NGS tests in patients with ataxia as a significant feature. Data from 33 eligible studies showed a median DY of 43% (IQR = 9.5–100%). The median DY for TP and ES was 46% and 41.9%, respectively. Higher DY was associated with specific phenotype selection, such as episodic ataxia at 68.35% and early and late onset of ataxia at 46.4% and 54.4%. Parental consanguinity had a DY of 52.4% ( p = 0.009), and the presumed autosomal recessive (AR) inheritance pattern showed 62.5%. There was a difference between the median DY of studies that performed targeted sequencing (tandem repeat expansion, TRE) screening and those that did not ( p = 0.047). A weak inverse correlation was found between DY and the extent of previous genetic investigation (rho = − 0.323; p = 0.065). The most common genes were CACNA1A and SACS . DY was higher for presumed AR inheritance pattern, positive family history, and parental consanguinity. ES appears more advantageous due to the inclusion of rare genes that might be excluded in TP.

Timely and accurate diagnosis of rare genetic disorders is critical, as it enables improved patient management and prognosis. In a resource-constrained environment such as the South African State healthcare system, the challenge is to design appropriate and cost-effective assays that will enable accurate genetic diagnostic services in patients of African ancestry across a broad disease spectrum. Next-generation sequencing (NGS) has transformed testing approaches for many Mendelian disorders, but this technology is still relatively new in our setting and requires cost-effective ways to implement. As a proof of concept, we describe a feasible diagnostic strategy for genetic disorders frequently seen in our genetics clinics (RASopathies, Cornelia de Lange syndrome, Treacher Collins syndrome, and CHARGE syndrome). The custom-designed targeted NGS gene panel enabled concurrent variant screening for these disorders. Samples were batched during sequencing and analyzed selectively based on the clinical phenotype. The strategy employed in the current study was cost-effective, with sequencing and analysis done at USD849.68 per sample and achieving an overall detection rate of 54.5%. The strategy employed is cost-effective as it allows batching of samples from patients with different diseases in a single run, an approach that can be utilized with rare and less frequently ordered molecular diagnostic tests. The subsequent selective analysis pipeline allowed for timeous reporting back of patients results. This is feasible with a reasonable yield and can be employed for the molecular diagnosis of a wide range of rare monogenic disorders in a resource-constrained environment.