Gene Expression Profiling of MLL-PTD in AML Identifies Differentially Regulated Members of the HOX Gene Family.

Partial tandem duplications (PTD) of the mixed lineage leukaemia (MLL) gene occur in approximately 7% of acute myeloid leukaemia (AML) patients. MLL-PTD has a frequency of approximately 10% in patients with normal karyotype and is reported to be associated with an adverse prognosis. Recent papers have reported that MLL-PTD may be useful as a marker for minimal residual disease (MRD) in AML patients with no other markers available due to normal cytogenetics. Diagnostic RNA from 133 AML patients was analysed for the presence of the MLL-PTD, twelve samples were positive and three of the twelve were also positive for FLT3 ITD. The MLL-PTD gene dosage was verified using quantitative polymerase chain reaction (QPCR). In order to identify the molecular disruption caused by MLL PTD mutations, fifty-four of the 133 patients were profiled using the Affymetrix U133A chip containing 22,283 probe sets. Four of the 54 samples analysed were MLL-PTD positive and three of these contained FLT3 mutations. Groups of genes were identified that had significance to MLL and showed more than 2-fold change in regulation in the MLL-PTD compared with wild type. Up-regulated genes found common to the two groups include HOXA9, HOX2B, HOX2C, HOX1 and CREBBP/EP 300. Whilst significant genes down-regulated more than 2-fold in MLL-PTD include MONDOA, a transcriptional activator. To identify whether MLL-PTD and FLT3 mutations used a common pathway, a comparison was made between genes with significance to either MLL-PTD or FLT3 mutations and no common genes were found. Hierarchical cluster analysis on the most significant genes, using Genespring software, created 4 main groups and distinguishes non-leukaemic samples, AML patients with MLL-PTD and those with a wild type MLL gene. This gene signature showed that MLL- PTD is a unique entity and could be identified amongst other AML subtypes. Moreover, over expression of Hox genes plays a role in MLL PTD positive AMLs and may provide an insight into the mechanism of the MLL-PTD with potential clinical implications.