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ORIGINAL ARTICLE
Year : 2016  |  Volume : 1  |  Issue : 3  |  Page : 31-37

Real-time quantitative polymerase chain reaction detection of minimal residual disease in acute lymphoblastic leukemia: a single-center experience


1 Department of Clinical Pathology, Assiut University Hospital, Assiut, Egypt
2 Department of Clinical Pathology, South Egypt Cancer Institute, Assiut, Egypt

Correspondence Address:
Eman M Nagiub Abdelsalam
Department of Clinical Pathology, Assiut University Hospital, Assiut
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/2357-0121.199351

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Background Monitoring of minimal residual disease (MRD) has become a frontline clinical practice in the treatment of virtually all childhood acute lymphoblastic leukemia (ALL) cases and in many cases of adult patients with ALL. The MRD diagnostics has proven to be the strongest prognostic factor allowing for risk group assignment into different treatment arms. The MRD techniques need to be sensitive (≤10–4), which means, the ability to detect one malignant cell among 10 000 normal cells; broadly applicable; accurate; reliable; fast; and affordable. Aim The objective of this study is to evaluate the analysis of immunoglobulin heavy chain (IGH) or T-cell receptor (TCR) gene rearrangements as targets for MRD assessment in ALL, allowing early detection of relapsed cases, compare with the results of morphological evaluation of the same cases and to risk stratify patients with ALL according to the MRD assessment as a prognostic marker independent and superior to other conventional risk factors. Patients and methods Overall, 30 patients (15 males and 15 females) with age ranged from 1 to 25 years old were included in this study. Patients were subjected to full medical history, clinical examination, laboratory examinations such as complete blood count, bone marrow aspirate smear examination, cytochemistry, and immunophenotyping. The molecular studies done by real-time PCR were performed using consensus primers and allele-specific primers for (IGH) or (TCR) gene rearrangements as targets to detect MRD. The ALL cases were assessed by real-time quantitative PCR at the time of diagnosis and at the end of induction chemotherapy, and comparative cycle threshold (Ct) relative quantification method was used for quantitative gene expression. Results The results showed discrepancy between the morphologic examination for ALL assessment depending on the 5% blast index as an indicator of remission or nonremission; morphological nonremissions (>5% blast) were detected in eight (26.7%) cases whereas 22 (73.3%) cases showed morphological remissions (<5% blast). Real-time quantitative PCR for IGH/TCR gene rearrangements was done for 30 patients. Of the 22 cases that showed morphological remission, there were 18 (81.8%) cases which were MRD positive (i.e. not in molecular remission) and four (18.2%) cases which were MRD negative (i.e., in molecular remission). Patients who showed nonremission morphologically (right cases) were confirmed to be MRD positive. None of the patient's clinical variables such as age, sex, total leukocyte count, and ALL immunophenotype have been identified as predictors of MRD risk. Conclusion Approximately 81% of ALL cases previously diagnosed as being in the remission state depending on the morphologic assessment only tend to harbor MRD as evident by positive MRD assessment using real-time PCR. The molecular assessment of MRD allows early detection of relapse with chance of intervention and tailoring of treatment according to the patients' need. It was found that there was no relation between MRD results and risk stratification in ALL according to age and white blood cell at diagnosis; this confirms that detection of MRD of leukemic cells can be considered a superior prognostic marker of relapse, independent from conventional prognostic factors. The risk classification in ALL based on MRD results could be used to guide the final treatment strategy and predict early relapse; this should allow prediction of response and relapse while patients are still in the state of clinical remission and morphological remission too.


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