High-throughput sequencing aids in diagnosis of bleeding, platelet disorders

ATLANTA — The use of high-throughput sequencing approaches successfully identified blood and platelet disorder genes among patients with inherited, undiagnosed platelet or coagulation defects, according to results presented during a plenary session at the ASH Annual Meeting and Exposition.

Analyses identified 23 novel blood and platelet disorder genes, suggesting these approaches can aid in genetic diagnosis to prevent future bleeding, the research showed.

Laboratory investigations often fail to establish reasoning for bleeding symptoms or to identify individuals at significant risk for future bleeding. In addition, patients with congenital thrombocytopenia cannot be differentiated from acquired disorders, rendering investigation of platelet function illogical.

Heterogeneity of phenotypes associated with established blood and platelet disorders further complicates diagnosis.

Therefore, Claire Lentaigne, BSc, MB, MRCP, from the National Institute for Health Research Imperial Biomedical Research Centre for hematology of Imperial College London, and colleagues developed high-throughput sequencing approaches to identify patients with undiagnosed bleeding and platelet disorders using the ThromboGenomics 79 gene panel high-throughput sequencing test and by whole-genome sequencing.

“Our aim has been twofold; firstly, to identify novel genes causing rare platelet disorders and, secondly, to try to harness this technology to bring it to the frontline of patient care and provide precision diagnostics to patients,” Lentaigne said during her presentation.

Researchers recruited individuals for blood samples based on three categories: individuals suspected to have a defect in one of the 79 blood and platelet disorder genes known to harbor variants liable for disorder (group 1; n = 1,321); individuals being prepared for elective surgery with self-reported bleeding symptoms (group 2; n = 212); and individuals with a suspected inherited blood platelet disorder not caused by a variant in one of the known blood and platelet disorder genes (group 3; n = 1,916).

“All these patients with platelet disorders recruited by different conditions from around the world have created an enormous amount of heterogenous data. We have assigned human phenotype ontology to make sense of the data,” Lentaigne said.

Human phenotype ontology, or HPO, is a library of more than 11,000 terms which can describe any phenotype in any organ system in the body and brain in great detail, according to Lentaigne.

“Essentially this ... applies to each patient a set of standardized computer terms,” Lentaigne said.

This process grouped similar patients together and applied specific analyses to them.

Researchers used the ThromboGenomics 79 gene panel high-throughput sequencing test to sequence DNA samples from groups 1 and group 2, and they used whole-genome sequencing for group 3 as part of the first 35,000 samples for the 100,000 Genomes Project.

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A multidisciplinary team reviewed variants by where pathogenicity was assigned to variants using Association for Clinical Genetic Science criteria, according to Lentaigne.

Researchers assessed a mean of 6.6 variants for each case.

In group 1, pathogenic variants were found among 54% of cases (n = 713), 40% of which were novel and likely pathogenic. Also, nearly 20% of variants were of unknown significance.

In group 2, researchers detected pathogenic variants among 2.8% of cases (n = 2). In group 3, pathogenic variants were identified among 12% of cases (n = 106).

Novel blood and platelet disorder genes identified through whole-genome sequencing analysis included NBEAL2, RBM8A, SRC, DIAPH1, TPM4, ABCC4 and KDSR.

In more than 300 cases with nonsyndromic thrombocytopenia, researchers detected clearly pathogenic or likely pathogenic variants among approximately 30% of cases. Of 131 cases with a significant bleeding disorder but no laboratory platelet or coagulation abnormality causal, researchers detected variants in five.

“Our results, and the observations by others, are compatible with the notion that the genetic architecture of unresolved bleeding and platelet disorder cases is extremely diverse,” the researchers wrote. “Continuing efforts such as the 100,000 Genomes Project with sharing and transparency of data across projects will enable better interpretation of variants and the discovery of novel genes.” – by Melinda Stevens

Reference:

Lentaigne C, et al. Abstract 5. Presented at: ASH Annual Meeting and Exposition; Dec. 9-12, 2017; Atlanta.

Disclosures: The authors report no relevant financial disclosures.