Introduction Of Whole-exome Sequencing Research Articles

Whole exome sequencing revealed a novel dystrophin-related protein-2 (DRP2) deletion in an Iranian family with symptoms of polyneuropathy.

Objective(s):Charcot-Marie Tooth disease (CMT) is one of the main inherited causes of motor and sensory neuropathies with variable expressivity and age-of onset. Although more than 70 genes have been identified for CMT, more studies are needed to discover other genes involved in CMT. Introduction of whole exome sequencing (WES) to capture all the exons may help to find these genes.Materials and Methods:Here, we tried to find the genetic cause of the neuropathy in two Iranian brothers using WES. Blood sample was collected from probands and their family members to extract the genomic DNA. The extracted DNA from one of the affected case was subjected for WES. The variant calls were filtered to reveal the pathogenic variant. Presence of the candidate mutation was confirmed using Sanger sequencing. The pathogenic potential of the variant was examined using in silico software. Using ClustalW multiple alignment, the presence of variant in conserved domain of protein was investigated. The parent and another affected boy were also checked for presence of the variant using PCR-sequencing. Results:The obtained data presented a novel TTC del mutation in CDS 738 of dystrophin related protein 2 (DRP2) gene, which was validated by sequencing. The variant was located in a conserved domain of DRP2 protein and predicted as pathogenic. Two affected boys were hemizygous for the mutation and received the mutation from mother. Conclusion:Here, we provided the evidence for the contribution of DRP2 in CMT. Also, the symptoms shed light on molecular aspect of this genetically heterogeneous disease.

Infectious Disease Risk in Dialysis Patients: A Transdisciplinary Approach.

Purpose of review:Infections are a major contributor to morbidity and mortality in end-stage renal disease (ESRD) patients. A better understanding of the interplay between infectious processes and ESRD may eventually lead to the development of targeted treatment strategies aimed at lowering overall disease morbidity and mortality. Monogenic causes are a major contributor to the development of adult chronic kidney disease (CKD). Recent studies identified a genetic cause in 10% to 20% of adults with CKD. With the introduction of whole-exome sequencing (WES) into clinical mainstay, this proportion is expected to increase in the future. Once patients develop CKD/ESRD due to a genetic cause, secondary changes, such as a compromised immune status, affect overall disease progression and clinical outcomes. Stratification according to genotype may enable us to study its effects on secondary disease outcomes, such as infectious risk. Moreover, this knowledge will enable us to better understand the molecular interplay between primary disease and secondary disease outcomes.Sources of information:We conducted a literature review using search engines such as PubMed, PubMed central, and Medline, as well as cumulative knowledge from our respective areas of expertise.Methods:This is a transdisciplinary perspective on infectious complications in ESRD due to monogenic causes, such as autosomal dominant polycystic kidney disease (ADPKD), combining expertise in genomics and immunology.Key findings:In ADPKD, infection is a frequent complication manifesting primarily as lower urinary tract infection and less frequently as renal infection. Infectious episodes may be a direct consequence of a specific underlying structural abnormality, for example the characteristic cysts, among others. However, evidence suggests that infectious disease risk is also increased in ESRD due to secondary not-well-understood disease mechanisms. These disease mechanisms may vary depending on the underlying nature of the primary disease. While the infectious disease risk is well documented in ADPKD, there are currently insufficient data on the risk in other monogenic causes of ESRD. WES in combination with novel technologies, such as RNA sequencing and single-cell RNA sequencing, can provide insight into the molecular mechanisms of disease progression in different monogenic causes of CKD/ESRD and may lead to the development of novel risk-stratification profiles in the future.Limitations:This is not a systematic review of the literature and the proposed perspective is tainted by the authors’ point of view on the topic.Implications:WES in combination with novel technologies such as RNA sequencing may enable us to fully unravel underlying disease mechanisms and secondary disease outcomes in monogenic causes of CKD and better characterize individual risk profiles. This understanding will hopefully facilitate the development of novel targeted therapies.

ASCIA‐P74: AUSTRALIAN AUTOINFLAMMATORY DISEASES REGISTRY (AADRY): A NATIONAL APPROACH TO THE GENETIC AND IMMUNOLOGICAL EVALUATION OF PATIENTS WITH SUSPECTED AUTOINFLAMMATORY DISEASE

Traditionally defined as periodic fever syndromes lacking markers of autoimmunity such as high titre self-reactive T cells and auto-antibodies, the spectrum of monogenic autoinflammatory diseases has broadened substantially in recent years. The clinical heterogeneity of newly described conditions such as STING associated vasculopathy with onset in infancy (SAVI) and Deficiency in ADA2 (DADA2) has forced reconsideration of traditional designations, with the common link between these disorders being inappropriate inflammation and cytokine dysregulation. These new diagnoses can be attributed to the introduction of whole exome sequencing (WES) into the diagnostic algorithm of patients with suspected, but currently genetically undefined, autoinflammatory diseases. We believe it's important to take a national approach to the diagnosis of autoinflammatory disorders given their rarity and introduce the establishment of AADRY, an Australian Autoinflammatory Diseases RegistrY. AADRY proposes to collect epidemiological and clinical data on patients with suspected and confirmed autoinflammatory diseases, and will perform whole exome sequencing in patients who are yet to be genetically characterised. We also aim to perform in vitro testing and harness CRISPR-Cas9 gene editing technology to determine the functional significance of novel variants and their potential contribution to clinical phenotype. We describe bioinformatic analyses and other results from the first 12 AADRY cases analysed, including one trio and one family of five.

Cancer patients’ preferences for return of somatic and germline whole-exome sequencing results: Data from the CANSEQ study.

1535 Background: The introduction of whole-exome sequencing (WES) into routine cancer care presents novel challenges for patient education and informed consent. In light of the recent American Coll...

NOTCH1, SF3B1 and BIRC3 Mutations in Chronic Lymphocytic Leukemia (CLL) Patients Requiring First-LINE Treatment: Correlation with Biological Parameters and Response to Treatment

AbstractAbstract 1784 Introduction:The introduction of whole exome sequencing has allowed to unravel novel molecular lesions in CLL. NOTCH1, SF3B1 and BIRC3 mutations are detected, according to the phases of disease, in 4–12%, 5–17% and 4–24% of patients, respectively. In retrospective studies, their presence has been shown to correlate with overall survival (OS) and treatment-free interval shortening.Aims: To define the incidence, correlation with known prognostic factors and clinical impact of NOTCH1, SF3B1 and BIRC3 mutations in CLL patients undergoing first-line treatment. Methods:We evaluated 162 CLL patients enrolled in the GIMEMA LLC0405 protocol (n=80) for patients aged <60 yrs and in the ML21445 protocol (n=82) for elderly patients (aged >65 yrs or 60–65 if not eligible for fludarabine-based programs). In the GIMEMA LLC0405 protocol, patients were stratified into low and high-risk: patients with del17p or with del11q plus an unmutated IGHV status and/or CD38 positivity and/or ZAP70 positivity were considered as high-risk (HR) and underwent Fludarabine plus Campath, followed by stem cell transplantation procedures, whereas low-risk patients received Fludarabine and Cyclophosphamide.The MLL21445 protocol consisted of 8 cycles of Chlorambucil and 6 of Rituximab induction treatment.NOTCH1 (exon 34), SF3B1 (exons 14 and 15) and BIRC3 (exons 2–9, including splicing sites) were screened by Sanger sequencing on either genomic DNA (gDNA) or whole genome amplified DNA (WGA) collected at the time of treatment. These studies were not part of the clinical protocols. Results:NOTCH1 mutations were detected at the time of treatment in 18 cases (22%) enrolled in the LLC0405 study. There was a significant association with high-risk stratification (p=0.036), namely with an IGHV unmutated status (p=0.0035), CD38 (p=0.03), +12 (p=0.034) and, partly, ZAP-70 expression (p=0.059). While the overall response rate (ORR) did not differ between NOTCH1 mutated vs wild-type (WT) cases (82% vs 77%, respectively), the complete response (CR) rate was significantly lower in NOTCH1 mutated patients (43% for WT vs 17% for NOTCH1 mutated cases; p=0.05). So far, no significant difference between mutated and WT patients has emerged in terms of OS and progression-free survival (PFS); this may be contributed by the fact that most NOTCH1 mutated cases were HR and were therefore treated more aggressively. SF3B1 mutations were recorded in 9 cases (11%); no significant associations were found with known biological parameters and, so far, with the ORR and CR rate. A single case harbored a BIRC3 mutation; this patient had an IGHV unmutated status, no FISH abnormalities and a concomitant SF3B1 mutation.In the ML21445 cohort, NOTCH1 mutations were found in 12 cases (15%), were associated with an unmutated IGHV status (p=0.047) and ZAP-70 expression (p=0.007), and did not impact on the ORR and CR rate. SF3B1 mutations were found in 11 cases (13%); no significant associations were found with known biological parameters and the ORR rate. Of interest, only 1/11 SF3B1 mutated patients achieved a CR. BIRC3 mutations were recorded in 3 patients (3.6%); of these, 2 were IGHV mutated, 1 had no cytogenetic abnormalities and 1 carried a del11q, while the third patient was IGHV unmutated status and had no cytogenetic abnormalities. No NOTCH1 and/or SF3B1 mutations were detected.Overall, NOTCH1, SF3B1 and BIRC3 mutations were largely mutually exclusive among each other and with TP53 lesions in the whole cohort. Conclusions:This study confirms the association of NOTCH1 mutations with unfavorable biologic markers and +12, while the presence of SF3B1 mutations was not coupled to poor prognostic markers in CLL patients requiring first-line treatment. Furthermore, it suggests that NOTCH1 mutations impact on the CR rate of young patients receiving Fluda-based regimens, while SF3B1 appears to impact on the CR rate of elderly patients treated with Chlorambucil and Rituximab. Given the small numbers of patients harboring BIRC3, it is at present difficult to draw any conclusion on the clinical impact of this mutation in the cohort of patients hereby analyzed. Disclosures:No relevant conflicts of interest to declare.