Rapid Molecular Profiling of Myeloproliferative Neoplasms with Next-Generation Sequencing

Myeloproliferative neoplasms (MPNs) are a heterogeneous group of blood disorders characterized by the abnormal proliferation of one or more of the terminal myeloid cell lines in the peripheral blood. These disorders include chronic myeloid leukemia (CML), polycythemia vera (PV), primary myelofibrosis (PMF), and essential thrombocythemia (ET), among other subtypes.1 Accurate and comprehensive molecular profiling is crucial for the effective diagnosis, prognosis, and treatment of MPNs.

 

Traditionally, sequential single gene testing (SGT) using methodologies, such as Sanger sequencing or PCR, have been utilized to identify driver mutations in JAK2, MPL, and CALR. Next-generation sequencing (NGS) is then utilized to further investigate triple negative MPNs, assess clonality, and understand prognosis and therapy selection (figure 1).

Figure 1. Single gene testing with reflex to traditional NGS testing.

However, this approach may require significant resources and time, often taking weeks to provide in-depth genomic profiling for MPN, potentially delaying patient management. Advancements in NGS have enabled laboratories to simultaneously analyze multiple biomarkers at initial workup, leading to more comprehensive, timely, and potentially cost-effective genomic profiling.

Advantages of NGS at initial MPN workup

Using NGS, specifically a rapid panel, at the initial work-up offers several advantages over sequential SGT, including:

 

  • Delivers in-depth genomic profiling: NGS can simultaneously detect all clinically significant mutations implicated in MPN. WHO and NCCN guidelines recommend investigating driver mutations in JAK2, MPL, and CALR, which are critical to MPN diagnosis. These guidelines also suggest examining other MPN-related genes such as ASXL1, IDH1, IDH2, SF3B1, SRSF2, and TET2, among others, which can provide crucial information about prognosis, clonal evolution, and aid in therapy selection. 2,3

  • Identifies rare and novel variants: NGS may identify rare and novel variants in driver genes (e.g., JAK2) that may be overlooked by sequential SGT.4

  • Clarifies MDS/MPN neoplasms: NGS enables the classification and diagnosis of myelodysplastic/myeloproliferative (MDS/MPN) neoplasms, a rare myeloid neoplasm exhibiting both dysplastic and proliferative features.5

 

By using a rapid NGS panel at the initial workup stage, healthcare providers can conduct a comprehensive analysis of multiple genes in a single test. This approach provides a more complete picture of the genetic landscape of MPN, enabling the identification of mutations implicated in diagnosis/prognosis and aiding timely therapy selection.3,6  (figure 2)

 

Figure 2. A rapid NGS panel at initial workup.

Timely molecular profiling

SGT often involves sequential testing, where each genetic mutation is analyzed separately. This process can be time-consuming and may delay the appropriate diagnosis and therapy initiation. With rapid NGS, healthcare providers can obtain genetic profiling faster, enabling more timely and informed clinical decision-making.

Cost-effective

While the initial cost of NGS is higher compared to single analyte testing, it offers potential long-term cost savings through its efficiency and comprehensiveness. SGT can become expensive when multiple tests are required to identify different mutations. As the number of targets increase, the feasibility of efficiently performing multiple testing diminishes7.

 

By using NGS at initial workup, multiple genes are consolidated into a single assay, reducing the need for sequential testing and minimizing the overall cost of additional resources. Moreover, the comprehensive information obtained from NGS may help guide more effective treatment decisions, potentially reducing the costs associated with delaying patient care and adverse outcomes.

Pathologist perspective: utilizing NGS for MPN diagnosis and risk assessment

During a recent conference, Dr. Yi Ding of Geisinger Medical Laboratories described how her laboratory overcame the MPN testing challenge. Before they brought NGS in-house, MPN genetic testing took 4 to 6 weeks in their lab. Patient follow-ups were scheduled as far out as three months after the initial visit. By bringing a multi-gene NGS panel in-house, they helped reduce the time to diagnosis.

 

Takeaway: Multi-gene NGS panels that include genes and mutations recommended by National Comprehensive Cancer Network (NCCN) guidelines and deliver timely results offer unprecedented opportunities to streamline diagnosis and enhance disease management for improved outcomes and quality of life.

Yi Ding, MD, PhD

Systems & Core Laboratory Director of Molecular Diagnostics

Geisinger Medical Laboratories

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References

  1. Thapa B, Fazal S, Parsi M, Rogers HJ. Myeloproliferative Neoplasms. 2023 Aug 8. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2025 Jan–. PMID: 30285359.
  2. Khoury, J.D., Solary, E., Abla, O. et al. The 5th edition of the World Health Organization Classification of Haematolymphoid Tumours: Myeloid and Histiocytic/Dendritic Neoplasms. Leukemia 36, 1703–1719 (2022). https://doi.org/10.1038/s41375-022-01613-1
  3. National Comprehensive Cancer Network® (NCCN®), Myeloproliferative neoplasms 2024. https://www.nccn.org/login?ReturnURL=https://www.nccn.org/professionals/physician_gls/pdf/mpn.pdf
  4. Zhang J, Shen K, Xiao M, Huang J, Wang J, Wang Y, Hong Z. Case report: Application of targeted NGS for the detection of non-canonical driver variants in MPN. Front Genet. 2023 Jun 16;14:1198834. doi: 10.3389/fgene.2023.1198834. PMID: 37396034; PMCID: PMC10313112.
  5. Arber et al. International Consensus Classification of Myeloid Neoplasms and Acute Leukemias: integrating morphologic, clinical, and genomic data. https://doi.org/10.1182/blood.2022015850
  6. Guglielmelli P et al. MIPSS70: Mutation-Enhanced International Prognostic Score System for Transplantation-Age Patients with Primary Myelofibrosis. J Clin Oncol. 2018 Feb 1;36(4):310-318. doi: 10.1200/JCO.2017.76.4886. Epub 2017 Dec 9. PMID: 29226763.
  7. Kuo FC, Mar BG, Lindsley RC, Linderman NI. The relative utilities of genome-wide, gene-panel, and individual gene sequencing in clinical practice. Blood. 2017;130(4):433-439.

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