Targeted Gene Regions

Genes interrogated, including relevant transcripts and exons, are listed in alphabetical order.

Clinical Indications

This assay is intended for patients with known or suspected hematologic neoplasms including myelodysplastic syndromes, myeloproliferative neoplasms, acute myeloid leukemia, acute lymphoblastic leukemia, and selected mature lymphoid leukemias.

Interpretation

All variants are classified using Association for Molecular Pathology guidelines for interpretation of somatic variants in cancer.^[8]

Detailed interpretations are provided for each variant, and an overall interpretation of the entire mutational profile summarizes the case findings.

Reported variants include those of strong or potential clinical significance as well as variants of unclear clinical significance.

Known benign polymorphisms are not reported.

Methodology

Nucleic acid extracted from the specimen is subjected to nested multiplex PCR-based target enrichment.

Coding and non-coding regions of targeted genes are amplified and sequenced on an Illumina instrument (San Diego, CA) with paired-end, 150×2 cycle reads.

A customized bioinformatic analytical pipeline is used to map reads to the reference human genome (Genomic Build GRCh37/hg19).

Limitations

This test does not detect structural variants or copy number changes and does not distinguish between variants that are inherited versus acquired.

During internal validation, this test delivered an average of >500X coverage and >98% of targeted regions showed over 100X coverage. The test demonstrated 95.2% sensitivity and 99.9% specificity in identifying single nucleotide variants, small insertions and deletions (indels) (≤10bp) of >5% variant allele fraction (VAF). For the identification of large indels (>10bp) at >5% VAF the test demonstrated 87.5% sensitivity and 99.9% specificity.

Due to the limitations of next-generation sequencing technology, some large insertions may not be detected.

References

1. Papaemmanuil E, Gerstung M, Malcovati L, et al. Clinical and biological implications of driver mutations in myelodysplastic syndromes. Blood. 2013 Nov 21; 122(22):3616-27.

2. Bejar R, Stevenson K, Abdel-Wahab O, et al. Clinical effect of point mutations in myelodysplastic syndromes. N Engl J Med. 2011 Jun 30;364(26):2496-506.

3. Döhner H, Weisdorf DJ, Bloomfield CD. Acute Myeloid Leukemia. N Engl J Med. 2015 Sep 17;373(12):1136-52.

4. Nazha A, Zarzour A, Al-Issa K, et al. The complexity of interpreting genomic data in patients with acute myeloid leukemia. Blood Cancer J. 2016 Dec 16;6(12):e510.

5. Arber DA, Orazi A, Hasserjian R, et al. The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia. Blood. 2016 May 19;127(20):2391-405.

6. “NCCN Guidelines for Treatment of Cancer by Site,” National Comprehensive Cancer Network: https://www.nccn.org/professionals/physician_gls/default.aspx#site.

7. McClure RF, Ewalt MD, Crow J, et al. Clinical Significance of DNA Variants in Chronic Myeloid Neoplasms: A Report of the Association for Molecular Pathology. J Mol Diagn. 2018 Nov;20(6):717-737. DOI: 10.1016/j.jmoldx.2018.07.002. Epub 2018 Aug 20.

8. Li MM, Datto M, Duncavage EJ, et al. Standards and Guidelines for the Interpretation and Reporting of Sequence Variants in Cancer: A Joint Consensus Recommendation of the Association for Molecular Pathology, American Society of Clinical Oncology, and College of American Pathologists. J Mol Diagn. 2017 Jan;19(1):4-23.