Mechanisms of CRLF2 Rearrangement with IGH and P2RY8 in B-Cell Acute Lymphoblastic Leukemia.

Precursor B-cell acute lymphoblastic leukemia (B-ALL) is the most common malignancy in children. While the great majority of these children achieve long-term remission, the disease exacts a high mortality in adults, and children with high-risk features. Fusion oncogenes such as BCR-ABL, MLL-AF4, E2A-PBX and TEL-AML1 are present in 60% of B-ALL cases. Cloning of these rearrangements has provided significant insight into the mechanisms involved in their formation. The gene alterations that affect the remaining 40% are poorly understood. We and others have recently identified CRLF2, a subunit of the thymic stromal lymphopoietin receptor, as a novel oncoprotein in B-ALL. Approximately 15% of adult and high-risk pediatric B-ALL that lack other characteristic gene rearrangements overexpress CRLF2, while leukemias with these rearrangements do not. In cases with CRLF2 overexpression, CRLF2 appears to be the driver of STAT activation, either alone or in combination with gain-of-function mutations in Janus Kinases (JAKs). CRLF2 overexpression results from rearrangements involving the CRLF2 locus, which is located in the pseudoautosomal regions on chromosomes X and Y. These rearrangements can either involve translocation with the immunoglobulin heavy chain (IGH) locus (t(X;14)(p22;q32) or t(Y;14)(p11;q32)) or interstitial deletion (del(X)(p22.33p22.33) or del(Y)(p11.32p11.32)). We sought to define the mechanisms of cleavage and repair that mediate these rearrangements.

For the translocations, we performed a series of polymerase chain reaction (PCR) assays to amplify junctions between IGHJ segments on chr.14 and the region upstream (i.e., centromeric) of CRLF2 on chr.X/Y. For the deletions, we used single nucleotide polymorphism (SNP) arrays and quantitative PCR to define the extent of deletions and then amplified junctions by PCR.

We successfully amplified IGHJ/CRLF2 translocation junctions from six B-ALL with CRLF2 overexpression. Junctions involved chr.X/Y sequence between 8-16kb upstream of the CRLF2 translation start site. Five of 6 clustered near putative V(D)J recombinase recognition signal sequences (RSS). Additional evidence for involvement of the V(D)J recombinase was identified in all cases, including the presence of nontemplated nucleotides. In contrast, deletions resulted in juxtaposition of the full length CRLF2 coding sequence to P2RY8. A similar event involving t(X;12) that resulted in SOX5 translocation to P2RY8 has been described in a single case of splenic follicular lymphoma. P2RY8 is a member of the purine nucleotide G-protein coupled receptor gene family and is highly expressed in lymphocytes.

CRLF2 rearrangements result in overexpression through juxtaposition to alternate transcriptional control elements. While translocations appear to be mediated by aberrant V(D)J recombination, deletions likely involve an alternate sequence-dependent mechanism that targets downstream of the P2RY8 promoter.

No relevant conflicts of interest to declare.