Evaluation of multiplex ligation‐dependent probe amplification as a method for the detection of copy number abnormalities in B‐cell precursor acute lymphoblastic leukemia

Abstract

Recent genomic studies have shown that copy number abnormalities (CNA) of genes involved in lymphoid differentiation and cell cycle control are common in B-cell precursor acute lymphoblastic leukemia (BCP-ALL). We have evaluated Multiplex Ligation-dependent Probe Amplification (MLPA) on 43 BCP-ALL patients for the detection of the most common deletions among these genes and compared the results to those obtained by fluorescence in situ hybridization (FISH) and genomic quantitative PCR (qPCR). There was good correlation between methods for CDKN2A/B, IKZF1, and PAX5 deletions in the majority of cases and MLPA confirmed the presence of deletions within the PAR1 region in two of three cases identified by FISH. Small intragenic aberrations detected by MLPA, which were below the resolution of FISH for CDKN2A/B (n = 7), IKZF1 (n = 3), and PAX5 (n = 3) were confirmed by qPCR. MLPA and qPCR were unable to detect populations present at a low level (<20%) by FISH. In addition, although MLPA identified the presence of a deletion, it was unable to discern the presence of mixed cell populations which had been identified by FISH: CDKN2A/B (n = 3), IKZF1 (n = 1), PAX5 (n = 2), and PAR1 deletion (n = 1). Nevertheless, this study has demonstrated that MLPA is a robust technique for the reliable detection of CNA involving multiple targets in a single test and thus is ideal for rapid high throughput testing of large cohorts with a view to establishing incidence and prognostic significance.

The clinical characteristics, therapy and outcome of 85 adults with acute lymphoblastic leukemia and t(4;11)(q21;q23)/MLL-AFF1 prospectively treated in the UKALLXII/ECOG2993 trial

“…27 The dual presence of myeloid antigens CD15 (S) and CD65 (S) is, however, a feature of all CD10 NEG Blineage cases and not restricted to those with t(4;11). 20,28 Contrary to earlier reports, 12 we found CD33 and CD13 to be expressed by significantly fewer lymphoblasts in t(4;11)-positive ALL than in other Philadelphia-negative BCP-ALL. We expanded on the characteristic stem cell antigen signature of t(4;11)-positive ALL, namely CD133 high/CD34 low expression, previously proposed in pediatric ALL, 29 by showing overexpression also of CD135 and CD105, while CD123 was present though non-discriminating.…”

Prognostic significance of copy number alterations in adolescent and adult patients with precursor B acute lymphoblastic leukemia enrolled in PETHEMA protocols

“…15 Finally, because MLPA is a technique with limited sensitivity, we selected samples with a blast burden 30%. 31,32 With this approach, we assumed that, for instance, approximately 3% of WT IKZF1 samples may have had subclonal populations with Ikaros deletions not detected by MLPA in contrast to what could have been found with polymerase chain reaction. 32 The results of the current study could help to refine the risk stratification of adolescent and adult ALL patients according to their genetic features.…”

MEN1 intragenic deletions may represent the most prevalent somatic event in sporadic primary hyperparathyroidism

“…Altogether, these data may support the discontinuity of MEN1 intragenic deletions found in our work. Although MLPA analysis represents a powerful tool in genetic analysis being used in numerous studies (45,46), the confirmation of the observed intragenic deletions would require the subcloning of PCR products in plasmids and further sequencing. These experiments could not be performed due to scarcity of tumour tissue and large size of deletion, making it difficult to obtain a reliable PCR product from FFPE.…”