Acute myeloid leukemia with 4q12 rearrangements does not involve pdgfra

Abstract Background:Chromosome 4q12 rearrangements are recurrent molecular alterations in hematologic malignancies, and the most common oncogenic driver involved is PDGFRA. PDGFRA rearrangements serve as defining molecular markers for Myeloid/Lymphoid Neoplasms with Eosinophilia and PDGFRA rearrangement (MLNE-PDGFRA), a distinct entity characterized by exquisite sensitivity to tyrosine kinase inhibitors (TKIs) such as imatinib. Fluorescence in situ hybridization (FISH) is widely used for detecting PDGFRA rearrangements, its reliance on known gene targets and breakpoints can limit diagnostic accuracy. In recent years, RNA sequencing (RNA-seq) has emerged as a critical tool for comprehensive characterization of gene fusions in hematologic malignancies. This study applies RNA-seq to resolve cryptic gene fusions within 4q12 rearrangements misinterpreted by FISH and defines the transcriptomic profile of PDGFRA-independent 4q12 aberrations in AML. Methods:RNA-seq was employed for transcriptomic profiling of bone marrow samples from 9 AML patients, comprising 2 cases previously diagnosed with PDGFRA rearrangement by FISH and 7 AML with NPM1 mutation serving as controls. A heat map was utilized to visualize RNA expression profiles, a volcano plot was employed to identify differentially expressed genes (DEGs), and the KEGG Orthology-Based Annotation System (KOBAS) online database was used for KEGG pathway enrichment analysis. Meanwhile, the literature on the 4q12 rearrangement not involving PDGFRA, as identified by molecular or FISH analyses, was reviewed. Results: We reports 2 cases of AML initially suspected to have PDGFRA rearrangement based on FISH analysis, which was later identified as harboring CDK6::CHIC2 (novel fusion) and ETV6::CHIC2 fusions by RNA-Seq. The two fusion genes was confirmed through PCR and Sanger sequencing. In both cases, RNA-seq mapped the 4q12 breakpoints to CHIC2 intron 1 (28–43 kb upstream of the CHIC2 probe's 3ʹ end), a region physically covered by the CHIC2 FISH probe but near its detection limit. This positional breakpoints generated a suboptimal hybridization signal, resulting in misinterpretation of CHIC2 disruption as PDGFRA rearrangement due to observed displacement of the PDGFRA probe to derivative chromosome. Based on literature review and two cases reported in this study, 14 cases with 4q12 rearrangements revealed a 64.3% (9/14) false-positive rate for PDGFRA rearrangement by FISH in rearrangements lacking PDGFRA involvement. Discordant results occurred in 9 cases where FISH falsely indicated PDGFRA rearrangement, while molecular testing identified rearrangements involving adjacent genes (GSX2, CHIC2, SCFD2). Transcriptome analysis revealed a distinct transcriptional profile between 2 CHIC2-rearranged AML cases compared to NPM1-mutated AML controls. A total of 492 DEGs were identified between the case and controls. KEGG pathway enrichment analysis revealed significant enrichment of upregulated genes in hematopoietic cell lineage, cytokine-cytokine receptor interactions, and JAK-STAT signaling pathways (FDR < 0.05). Conclusion: Collectively, this study establishes RNA-seq as a superior method for detecting cryptic genetic rearrangements within the 4q12 locus compared to FISH. The distinct transcriptional signature in CHIC2-rearranged AML suggests dysregulation of hematopoietic cell lineage, cytokine-cytokine receptor interactions and JAK-STAT signaling as potential contributors to disease pathogenesis, indicating that dysregulated nodes within these pathways may be potential therapeutic targeting.