Investigating Ikaros deletions in cohort of South African acute lymphoblastic leukemia patients
Date
2014-08-22
Authors
Moodley, Mishalan
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Abstract
INTRODUCTION: Despite best current therapy, acute lymphoblastic leukemia (ALL) still remains the most common cause of cancer-related death in children and young adults. Relapse is the main reason for treatment failure in ALL patients and occurs in 15-20% of these patients. Current risk stratification criteria have not been sufficient to predict relapse in ALL patients. The Philadelphia (Ph) chromosome is a chromosomal abnormality found in a subset of high risk ALL patients and is associated with a poor prognosis. Recent genome-wide studies have identified focal deletions of the Ikaros gene (IKZF1) in 70-80% of B-cell ALL patients that have the Philadelphia (Ph) chromosome. Subsequent studies have also found a strong correlation between IKZF1 deletions and ALL patients (Ph+ and Ph-) that relapse. IKZF1 is required for normal lymphoid development and loss of IKZF1 results in haploidinsufficiency or the overexpression of dominant negative IKZF1 isoforms, in particular Ik6 in high risk ALL patients. Most studies used DNA microarrays to detect IKZF1 deletions. Multiplex ligation probe dependent amplification (MLPA) is a low cost, rapid technique that can detect small DNA copy number changes of up to 50 targets in a single reaction and is not as technically challenging to analyse as arrays. MLPA has also been suggested to be used as an alternative to array based techniques in developing countries. METHODS: There were 31 ALL (paediatric and adult) patients that were tested using MLPA and 24 ALL patients tested using reverse transcriptase PCR (RT-PCR) to detect IKZF1 copy number changes and IKZF1 isoform expression pattern respectively. RT-PCR was validated with DNA sequencing and MLPA was validated with Fluorescent in situ hybridization (FISH). MLPA was also compared to cytogenetics in certain cases. RESULTS: MLPA detected 156 copy number changes (7.1 aberrations per sample) in 22 leukemic patients. IKZF1 deletions accounted for the majority of the aberrations (41%) and were detected in 53% of Ph+ ALL patients (n=15) by MLPA. IKZF1 deletions were detected at presentation and relapse in Ph+ and Ph- ALL patients. IKZF1 isoform Ik6 was detected in 70% of Ph+ and relapsed ALL patients after performing RT-PCR. IKZF1 deletions of exons 4-7 resulted in exclusive expression of Ik6. MLPA results were also correlated with certain aneuploidies detected
with cytogenetics. CONCLUSION: This study showed that IKZF1 deletions could have assisted with prognosis in certain ALL cases and thus, newly diagnosed ALL patients should be screened for IKZF1 deletions. MLPA proved to be a reliable, rapid and cost effective technique to detect small copy number changes in multiple genes and should be implemented as a diagnostic test to detect IKZF1 deletions.