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As controversy exists regarding the prognostic significance of genomic rearrangements of CRLF2 in pediatric B-precursor acute lymphoblastic leukemia (ALL) classified as standard/intermediate-risk (SR) or high-risk (HR), we assessed the prognostic significance of CRLF2 mRNA expression, CRLF2 genomic lesions (IGH@-CRLF2, P2RY8-CRLF2, CRLF2 F232C), deletion/mutation in genes frequently associated with high CRLF2 expression (IKZF1, JAK, IL7R), and minimal residual disease (MRD) in 1061 pediatric ALL patients (499 HR and 562 SR) on COG Trials P9905/P9906. Whereas very high CRLF2 expression was found in 17.5% of cases, only 51.4% of high CRLF2 expressors had CRLF2 genomic lesions. The mechanism underlying elevated CRLF2 expression in cases lacking known genomic lesions remains to be determined. All CRLF2 genomic lesions and virtually all JAK mutations were found in high CRLF2 expressors, whereas IKZF1 deletions/mutations were distributed across the full cohort. In multivariate analyses, NCI risk group, MRD, high CRLF2 expression, and IKZF1 lesions were associated with relapse-free survival. Within HR ALL, only MRD and CRLF2 expression predicted a poorer relapse-free survival; no difference was seen between cases with or without CRLF2 genomic lesions. Thus, high CRLF2 expression is associated with a very poor outcome in high-risk, but not standard-risk, ALL. This study is registered at www.clinicaltrials.gov as NCT00005596 and NCT00005603.

Analysis of 15 cases of T-cell acute lymphoblastic leukemia with spectral karyotyping (SKY), which can identify all chromosomes simultaneously, clarified the chromosome rearrangements in 3 cases and confirmed them in 11 others; no abnormal cells were identified in 1 case, which had only 10% abnormal cells. Five of the latter cases had a normal karyotype. Thus, the use of SKY substantially improves the precision of karyotype analysis of malignant cells, which in turn leads to a more accurate assessment of the genotypic abnormalities in those cells.

Using fluorescence in situ hybridization (FISH) and probes located on 12p12.1 to 13.3, we studied the breakpoints of 23 patients who had various hematologic malignant diseases and who had 12p13-balanced translocations (21 patients), inversion (1 patient), or insertion (1 patient). Among them, 14 patients had breakpoints within YAC964c10, which contains the TEL (ETV6 ) gene and in 12 of these with balanced translocations or insertion, the FISH results suggested that TEL was involved. Two of the 14 patients, patients no. 13 and 14, had breakpoints in YAC 964C10 that were centromeric to TEL but telomeric to KIP1. In the other 9 patients whose breakpoints did not fall within the YAC, the breakpoints were found telomeric to the YAC in at least three different locations on distal 12p. These results indicated that TEL was involved in only half (12 of 23) of the patients with balanced 12p13 rearrangements and that there probably were several other breakpoint cluster regions on 12p13, suggesting that genes other than TEL were involved in these rearrangements.

The involvement of 11q23-balanced translocations in acute leukemia after treatment with drugs that inhibit the function of DNA topoisomerase II (topo II) is being recognized with increasing frequency. We and others have shown that the gene at 11q23 that is involved in all of these treatment-related leukemias is MLL (also called ALL1, Htrx, and HRX). In general, the translocations in these leukemias are the same as those occurring in de novo leukemia [eg, t(9;11), t(11;19), and t(4;11)], with the treatment-related leukemias accounting for no more than 5% to 10% of any particular translocation type. We have cloned the t(11;16)(q23;p13.3) and have shown that it involves MLL and CBP (CREB binding protein). The CBP gene was recently identified as a partner gene in the t(8;16) that occurs in acute myelomonocytic leukemia (AML-M4) de novo and rarely in treatment-related acute myeloid leukemia. We have studied eight t(11;16) patients, all of whom had prior therapy with drugs targetting topo II with fluorescence in situ hybridization (FISH) using a probe for MLL and a cosmid contig covering the CBP gene. Both probes were split in all eight patients and the two derivative (der) chromosomes were each labeled with both probes. Use of an approximately 100-kb PAC located at the breakpoint of chromosome 16 from one patient revealed some variability in the breakpoint because it was on the der(16) in three patients, on the der(11) in another, and split in four others. We assume that the critical fusion gene is 5'MLL/3'CBP. Our series of patients is unusual because three of them presented with a myelodysplastic syndrome (MDS) most similar to chronic myelomonocytic leukemia (CMMoL) and one other had dyserythropoiesis; MDS is rarely seen in 11q23 translocations either de novo or with t-AML. Using FISH and these same probes to analyze the lineage of bone marrow cells from one patient with CMMoL, we showed that all the mature monocytes contained the fusion genes as did some of the granulocytes and erythroblasts; none of the lymphocytes contained the fusion gene. The function of MLL is not well understood, but many domains could target the MLL protein to particular chromatin complexes. CBP is an adapter protein that is involved in regulating transcription. It is also involved in histone acetylation, which is thought to contribute to an increased level of gene expression. The fusion gene could alter the CBP protein such that it is constitutively active; alternatively, it could modify the chromatin-association functions of MLL.