التقييم الجزيئي لنسخ جين المقاومة الدوائية MDR1 في بعض المرضى العراقيين البالغين المصابين بسرطان ابيضاض الدم الحاد == Molecullar Assessment of Multidrug Resistance Gene (MDR1) Transcript In Some Adult Iraqi Patients With Acute Leukemia
Author name:
كفاح جبار شاكر اليعقوبي
Supervisor name:
عبد الحسين الفيصل
General topic:
Biology
Specific topic:
Biotechnologies
Degree:
Doctorate
University:
University of Baghdad - Institute Of Genetic Engineering And Biotechnology - Department Of Biotechnology
Language:
English
University location:
Baghdad
First pages:
24T2952 - p.pdf
Abstract:
The present study aims to shed light on the follow up of acute leukemic (AL) patients at initial diagnosis and after treatment to assess the response and early relapse through evaluating the gene expression level of one of the major multidrug resistance genes which is the multidrug resistance 1 (MDR1) to investigate the possible association between level of MDR1 gene expression and the clinical outcomes and this may be considered as a potential marker for response to chemotherapy of acute leukemic patients. Furthermore, the current study correlates between the MDR1 gene phenotype and MDR1 genotype in three important coding regions (C1236T, G2677T/A, and C3435T considering the potential influence of altering MDR1 activity and its effect on therapy outcome as well as susceptibility to develop cancer.White blood cells (WBCs) isolated from 106 blood sample of acute leukemic patients were provided by Iraqi hospitals in Medical City. These samples were distributed as follows : 46 newly diagnosed patients with acute leukemia who had not received chemotherapy and follow - up 25 after 1st induction, 17 after 2nd induction and 8 at consolidation, with 10 blood samples of healthy voluntaries. Two comparative groups were taken included 46 sample of peripheral blood (PB) and 26 sample of bone marrow biopsy (BMB) in paraffin blocks to validate the level of gene expression compare to WBCs. For genotyping analysis, 56 of blood sample were taken to study genetic variation of MDR1 gene polymorphism. The samples preservation with TRIzol was done. Samples subjected to total RNA and DNA extraction, then molecular study by using reverse transcription, Quantitative Real Time - polymerase chain reaction (qRT - PCR) and direct sequencing, at Molecular Oncology Unit in Guy´s Hospital - Kings College / London.The study reached at the following results : 1 - The results showed age groups (20 - 39 years) were associated with acute myeloid leukemia (AML), while (13 - 19 years) with acute lymphoblastic leukemia (ALL).2 - The level of MDR1 gene expression showed high significant differences with WBCs compared to PB and BMB.3 - The clinical outcomes indicated that the rate of complete response (CR) of newly diagnosed acute leukemic patients was 19(41%), while 27(58.7%) was non - responder (NR).4 - Statistical analysis showed significant differences with NR at initial diagnosis in acute myeloid leukemia, while appeared after 1st induction in lymphoid type.5 - The results of positivity MDR1 gene expression were 10(21.7%) out of 46 newly diagnosed in acute leukemia, while 36(78.3%) were MDR1 - negative depend on (1.1±0.03) cutoff value.6 - The positivity MDR1 gene expression appeared mainly in non - responders patients at initial diagnosis, and with early relapse patients, after complete remission, in consolidation.7 - The MDR1 mRNA expression showed significant differences with high level in NR compared to CR patients at initial diagnosis. During treatment follow up the increased level of MDR1 gene expression in CR patients and appeared non - significantly with NR.8 - The results of MDR1 C1236T genotype and allele frequency showed that 1236CC wild type genotype and C allele were significantly frequent in healthy control. While CT heterozygous genotype frequency was highly significant in AML and no significant difference in allele frequency. ALL showed non - significant difference in genotype and allele frequency of MDR1 C1236T.9 - Odds ratio and 95% confidence interval (ORs and 95%CI) analysis showed no evidence associated with risk factor in MDR1 C1236T ALL carriers. While risk factor observed in AML with MDR1 1236CT carriers.10 - The results of MDR1 phenotype - genotype association indicate that MDR1 1236CC wild type was significantly high expression among healthy and it was aprotective genotype. While the MDR1 1236CT showed significant differences with high level of MDR1 gene expression in AML patients. Whereas ALL revealed significant differences in high level of MDR1 gene expression with MDR1 1236TT genotype. Both CT and TT were affected genotypes.11 - The results of MDR1 G2677T genotype and allele frequency indicated that 2677GA genotype significantly appeared with low frequency in healthy control with no significant difference in allele frequency. Both ALL and AML showed high significant frequency in 2677GT genotype. G allele frequency was showed significant differences in AML while non - significant in with ALL.12 - Odds ratio and 95% confidence interval (ORs and 95%CI) analysis showed the MDR1 2677GT genotype was associated with risk factor to developing ALL and AML. Whereas the GG appeared associated with AML only.13 - MDR1 phenotype - genotype association, indicate that MDR1 2677GA genotype was significantly high expression in healthy individual. While AML patients showed significant differences with high level of MDR1gene expression in 2677GT genotype. ALL showed significant differences with high level of MDR1 gene expression in MDR1 2677TT genotype.14 - The results of MDR1 C3435T genotype and allele frequency showed significant difference in genotype and allele frequency with heterozygous CT in both control and AML patients and mutant T allele. Whereas non - significant genotype and allele frequency with ALL.15 - Odds ratio and 95% confidence interval (ORs and 95%CI) analysis showed that the MDR1 3435CC genotype carriers associated with risk to developing ALL. While no risk factor associate with MDR1 C3435T variants to develop AML.16 - MDR1 phenotype - genotype association, indicate that the wild type 3435CC genotype was significantly high expression in healthy control. The MDR1 3453CT genotype showed high significance with high level of MDR1 gene expression inAML. While ALL showed significantly high level of MDR1 gene expression in 3435TT genotype.17 - The results of MDR1 genotype - phenotype association showed similar impact of MDR1C1236T, G2677T/A and C3435T genotypes in AML clinical outcomes. The MDR1 CT/GT/TT genotypes were associated in NR AML with high level of expression at presentation, compared to significant low level in CC/GG genotype. In contrast, CR patients were observed non - significant with MDR1 gene expression at presentation and significant with low MDR1CC/GG genotypes in post treatment. In regards to ALL patients the MDR1 TT genotype showed significant differences with high level of MDR1 gene expression in NR and CR ALL at presentation and significant only with NR at post treatment. So there was no clear evidence between MDR1 genotypes and clinical outcome with ALL.18 - The haplotype results showed that the three MDR1 C1236T, G2677T/A and C3435T genotype were linkage disequilibrium significantly with heterozygous haplotype B (CT - GT - CT) compared to A(CGC) and C(TTT). Also B haplotype appeared significantly with high level of MDR1 gene expression compared to A and C. According to the clinical outcome, haplotype B was observed significant differences in NR AML patients while other haplotypes were non - significant
