High-Throughput Genetic Testing for Thrombotic Microangiopathies and C3 Glomerulopathies

Abstract

The thrombotic microangiopathies (TMAs) and C3 glomerulopathies (C3Gs) include a spectrum of rare diseases such as atypical hemolytic uremic syndrome, thrombotic thrombocytopenic purpura, C3GN, and dense deposit disease, which share phenotypic similarities and underlying genetic commonalities. Variants in several genes contribute to the pathogenesis of these diseases, and identification of these variants may inform the diagnosis and treatment of affected patients. We have developed and validated a comprehensive genetic panel that screens all exons of all genes implicated in TMA and C3G. The closely integrated pipeline implemented includes targeted genomic enrichment, massively parallel sequencing, bioinformatic analysis, and a multidisciplinary conference to analyze identified variants in the context of each patient's specific phenotype. Herein, we present our 1-year experience with this panel, during which time we studied 193 patients. We identified 17 novel and 74 rare variants, which we classified as pathogenic (11), likely pathogenic (12), and of uncertain significance (68). Compared with controls, patients with C3G had a higher frequency of rare and novel variants in C3 convertase (C3 and CFB) and complement regulator (CFH, CFI, CFHR5, and CD46) genes (P,0.05). In contrast, patients with TMA had an increase in rare and novel variants only in complement regulator genes (P,0.01), a distinction consistent with differing sites of complement dysregulation in these two diseases. In summary, we were able to provide a positive genetic diagnosis in 43% and 41% of patients carrying the clinical diagnosis of C3G and TMA, respectively.

Combined study of ADAMTS13 and complement genes in the diagnosis of thrombotic microangiopathies using next‐generation sequencing

“…Nevertheless, this NGS study ended with 17 (50%) genetically unresolved aHUS patients, as described in others aHUS studies . Therefore, the expectation that the whole‐exome sequencing approaches could identify other genes involved in aHUS is high; however, these studies still remain under investigation . Conversely, NGS‐targeted gene panels are being introduced into clinical practice providing substantial benefits for definitive diagnoses in hematological diseases …”

C4 Nephritic Factors in C3 Glomerulopathy: A Case Series

“…All biopsies were reviewed in a multi-disciplinary forum as described. 4 Each patient provided informed consent, which was approved by the Institutional Review Board of Carver College of Medicine at the University of Iowa.…”

“…4, 13 As a source of genomic DNA, the buffy coat was isolated and DNA was extracted using the Gentra Puregene Kit (Qiagen Inc., Valencia, CA). Serum was prepared from whole blood allowed to clot in plain (untreated) tubes for 45min and then centrifuged for 10min at 1000 g .…”

Complement component C3 – The “Swiss Army Knife” of innate immunity and host defense

“…Although the molecular consequences of this polymorphism are still being explored, the C3F variant appears to be associated with an unfavorable outcome in several diseases, including IgA nephropathy, partial lipodystrophy, and systemic vasculitis . Meanwhile, numerous distinct C3 variants have been correlated with certain pathologic conditions, particularly with age‐related macular degeneration (AMD), aHUS, and C3G . Complement dysregulation is a common feature in these diseases, producing a situation featuring chronic inflammation and tissue damage .…”