13. Diagnostic utility of FISH testing for Ph-like B-ALL: A 5-year institutional experience

Ph-like B-ALL accounts for 15-30% of B-ALL in children, adolescents, and adults and is associated with high rates of treatment failure. Current clinical trials are assessing the efficacy of adding tyrosine kinase inhibitors (TKIs) to chemotherapy in these patients. However, real-time diagnosis of these patients is challenging given the genetic heterogeneity of this disease and the often cytogenetically cryptic nature of Ph-like B-ALL alterations. The ACMG standards recommend a tiered approach for B-ALL FISH testing (PMID: 38349293). This study aims to assess the diagnostic utility of Ph-like B-ALL FISH testing in pediatric/young adult and adult B-ALL patients in our institution over a period of 5-years. Between March 2020 and March 2025, 157 newly diagnosed B-ALL patients (M:F ratio 1.38:1) were studied at the UAB Cytogenetics Lab using the Ph-like B-ALL FISH panel, including 117 pediatric/young adult and 40 adult patients. The Ph-like B-ALL FISH panel includes break-apart probes (BAPs) for ABL1, ABL2, CRLF2, JAK2, and PDGFRB . The testing scheme for pediatric/young adult patients includes chromosome analysis and both the standard B-ALL and Ph-like B-ALL FISH panels performed simultaneously, whereas in adult patients it includes chromosome analysis and the standard B-ALL FISH panel, and reflexing to the Ph-like B-ALL FISH panel in patients lacking a diagnostic abnormality using the standard B-ALL panel. A total of 27 patients (27/157, 17.2%) demonstrated evidence for a rearrangement of one of the genes included in the Ph-like B-ALL FISH panel, including 19 patients (19/157, 12.1%) with CRLF2 rearrangement, 4 patients (4/157, 2.5%) with JAK2 rearrangement, 2 patients (2/157, 1.3%) with ABL1 rearrangement (other than BCR::ABL1 fusion), and 2 patients (2/157, 1.3%) with PDGFRB rearrangement. Our CRLF2 FISH testing scheme was used to further characterize the 19 patients with CRLF2 rearrangement, which included additional testing using the IGH::CRLF2 dual-fusion probe and P2RY8 BAP. A total of 12 patients (12/19, 63.2%) had the IGH::CRLF2 fusion due to the t(X/Y;14) and 5 patients (5/19, 26.3%) had the P2RY8::CRLF2 fusion due to the PAR1 deletion, whereas 2 patients (2/19, 10.5%) had both rearrangements which were shown to involve the same CRLF2 allele. Based on available evidence, the co-presence of both rearrangements is thought to function like the IGH::CRLF2 fusion only. Our CRLF2 FISH testing scheme will be presented. Finally, a notable M:F ratio (2.8:1) bias was noted in the CRLF2 -rearranged group. This study illustrates the diagnostic utility of the Ph-like B-ALL FISH testing. Although FISH analysis often cannot identify the 5’ fusion gene partner in ABL -class and JAK2 rearrangements, it can provide the first clinical suspicion for Ph-like B-ALL and allocate patients efficiently who require further molecular characterization. This could facilitate earlier therapeutic intervention with JAK or ABL/PDGFR inhibitor addition in induction chemotherapy.