Immunoglobulin Heavy Chain (IgH) Gene Rearrangement in Egyptian Precursor B-Acute Lymphoplastic Leukemia - Implication for Assessment of Minimal Residual Disease.
Authors: El-Beshir Dalia A. , Shammaa Sameh S. , Evans Paul P. , Abraham Nader G. , Abd-Elghaffar Hasan A. 
Published: October 2019

Abstract
The level of MRD determined with the use of PCR based techniques is the most reliable prognostic factor in therapy of ALL. In contrast to the western countries, molecular based techniques of MRD is still not included in routine assessment of ALL therapy effectiveness in Egypt. Therefore the aim of this study was early detection of MRD in precursor B-cell neoplasm to correlate it with the known prognostic factors, to assess the applicability gene scan(GS)for the detection of MRD together with assessment if there features indicating the existence of Egyptian specific characterization for IgH rearrangement. The study was conducted on 33 Egyptian patients(16 children and 17 adults)with precursor-B-ALL as confirmed by immunophenotyping. Bone marrow samples were taken on diagnosis and after completion of induction treatment. Additional 14 samples from 6 patients were collected from HMDS, Leeds General Infirmary, Leeds, UK to compare the results of gene scan and four color flow cytometry (FC) with ASO RQ PCR technique as a reference method used in the MRD detection. MRD detection was done for these cases using GS. Presentation samples DNA were amplified using Vh-Jh Biomed -2 multiplex primer sets. GS was done to the fluorescent PCR product to detect the distinct size of this product at presentation and the follow up samples. ASO RQ PCR was done for some cases as a reference method to compare it s results with FC and GS. All cases were in clinical remission after completion of induction treatment. When monitored for MRD, only 56.3% of children and 23.5% of adults found to be in true molecular remission. The MRD positivity in children in our study(62.5%)is higher than the Western studies using either FC, sequence-based techniques or heteroduplex technique. On the other hand, the MRD positivity in adult ALL(78.2%)in our study was comparable to the other Western studies. It is found that there is a very high significant concordance between the presence of MRD at post-induction and the occurrence of relapse in both childhood (p=0.000) and adult ALL (p=0.002). There was no significant correlation between the conventional prognostic criteria and the existence of MRD apart from percentage of peripheral blood at day 7 (p=0.009) in childhood ALL. GS results were compared with four color FC, ASO RQ PCR. Sensitivity of GS in detection of MRD in precursor B-ALL is 75% compared with ASO RQ PCR. There was complete VH-JH rearrangement in all patients. It is noticed that the most frequently used VH gene segments in our study belongs to VH3 family in both children (56.3%) and adult (64.7%). This is followed by VH6(25%), VH1(18.75%), and VH 4(12.5%)in children, and by VH1(29.41%), and VH2, and VH4 together covering 11.76% in adult group. The prevalence of VH3 family usage noticed in this study is in concordance to the most of published studies. However, the frequency of other VH other than VH3 segment usage varied from the Western studies. In conclusion, MRD monitoring in precursor-B-ALL patients is a powerful independent prognostic factor. A wide scale study on Egyptian precursor B-ALL patients to define the frequency of VH usage and the presence of other rearrangements that may characterize MRD kinetics and clonal evolution for stratification of treatment protocol.