Development of a Targeted NGS Panel for Hematologic Malignancies According to Who/ICC 2022 Guidelines

Introduction: Since 2020, we have successfully developed a targeted blood cancer NGS panel and have been using it efficiently in clinical settings. However, it needed to be upgraded due to recent new blood cancer diagnosis and classification guidelines by the World Health Organization and the International Collaboration for Cancer Classification (ICC) in 2022. Therefore, in this study, an updated targeted NGS panel that covers the updated guidelines was evaluated using clinical samples.

Methods: To adhere to the recent guidelines, the currently used targeted NGS panel (KBB DNA/RNAseq NGS Leukemia PHB; KBlueBio Inc., Hwasun, Korea) was reviewed. Its analytical performance was validated using standard NA12878 and clinical samples. The clinical samples consisted of 19 genomic DNA and 20 total RNA samples extracted from the bone marrow or peripheral blood of patients with hematologic malignancies, including acute myeloid leukemia, acute lymphocytic leukemia, myelodysplastic syndrome/myeloproliferative neoplasm, multiple myeloma, and lymphoma.

Results: The updated DNA panel comprises 125 genes including 6 new genes ( BCORL1, BLM, GNB1, PRPF8, SAMD9 and SAMD9L). The updated RNA panel has 116 genes including 31 new genes ( FLT3, CBFA2T3, RPN1, BCL11B, ZNF362, HLF, NUTM1, UBRF, CDX2, ZEB2, CDK6, ZMYM2, AFDN, ELL, NSD1, HOXA9, KDM5A, ZBTB16, GLIS2, PRDM16, NPM1, KAT6A, MNX1, TAF15, MLLT1, P2RY8, HNRNPUL1, ECM1, ENAM, JCHAIN (IGJ) and MDFIC). The accuracy, repeatability, reproducibility, sensitivity, and detection limit of the updated NGS panels were evaluated using standard materials, and the results met the predefined criteria. A comparison of the updated DNA panel with the old panel using 19 clinical samples revealed an overall concordance rate of 100% between the two panels for all mutations (95% confidence interval (CI): 99.72-100.00%). Similarly, the updated RNA panel also met the predefined criteria. The overall concordance rate between the updated RNA panel and the old panel was 99.72% in 20 clinical samples (95% CI: 99.00-99.72%).

Conclusions: The old targeted DNA/RNA NGS panel was successfully updated according to the 2022 WHO and ICC guidelines, and can be used to accurately and efficiently detect the genetic variants of blood cancers.