Abstract 1036: Development of a comprehensive RNA-based NGS panel for detecting tyrosine kinase fusion genes associated with primary hypereosinophilia

Abstract

Abstract Hypereosinophilia (HE) is a medical condition diagnosed when the count of eosinophils in the blood is greater than 1.5 × 109 per liter. Primary HE is a clonal stem cell disorder manifesting as myeloid and/or lymphoid malignancies and is typically driven by aberrant fusion genes involving a tyrosine kinase (TK) gene (e.g., PDGFRA, PDGFRB, FGFR1, JAK2, FLT3, ABL1). The precise identification of the underlying molecular genetic abnormalities is crucial for timely diagnosing and managing primary HE. There has been no HE-specific assay that can concomitantly detect all known or novel TK fusion genes. We've developed an RNA-based panel for detecting TK fusion genes in HE. This panel, developed using 5' RACE technology, allows for the identification of fusion transcripts in total RNA extracted from whole blood sample. It covers 48 known TK fusion genes and also has the capability of detecting novel gene fusions. We balanced and optimized the panel using an in-house fusion RNA mix containing 11 fusion transcripts. We additionally validated the panel using samples from five patients with hematopoietic neoplasms and two fusion-positive cell lines to ensure its accuracy. The Limit of Detection (LoD) was established by analyzing the in-house RNA mix and normal whole blood RNA. It was found that the minimum RNA input required to detect all 11 fusion transcripts was 3540 copies in the presence of a background of 10 ng normal whole blood RNA. The panel successfully identified two fusion transcripts using 10 ng of total RNA obtained from cancer cell lines. The panel was then used to test five patient RNA samples extracted from whole blood. We successfully confirmed the presence of BCR-ABL1 fusion in 3 patient samples and ETV6-PDGFRB fusion in 1 patient sample, respectively. In summary, we developed HE-specific NGS panel for detecting all known and novel TK fusion genes and demonstrated its analytical accuracy and clinical effectiveness. The next phase of our study is to further validate the tests on a large patient cohort. Citation Format: Bongyong Lee, Andrew Chern, Aiguo Zhang, Chuanyi M. Lu, Michael Y. Sha. Development of a comprehensive RNA-based NGS panel for detecting tyrosine kinase fusion genes associated with primary hypereosinophilia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1036.