الفهرس 
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Abstract
Acute lymphoblastic leukemia (ALL) is a malignant lymphoid cells proliferation at early stage of differentiation, it is primarily a disease of children under six years of age ; approximately 80-85% are of precursor B-cell phenotype. Diagnosis of ALL includes peripheral blood and bone marrow examination and cytochemistry together with immunophenotyping by flowcytometry, cytogenetics & molecular techniques.
With improvements in diagnosis and treatment, overall cure rate for children with ALL reached 85% in the developed world. Multiple recent studies indicate that genetic polymorphisms play an influential role in childhood ALL susceptibility, treatment response and prognosis.
The purine analog mercaptopurine is a key medication for the successful treatment of childhood ALL, Inosine triphosphate pyrophosphatase (ITPA) is one of several enzymes whose job is to cleanse the nucleotide pool. It catalyze the pyrophosphohydrolysis of 6-thioITP and-methylthioITP and prevents the accumulation of such potentially toxic compounds. Therefore, the end result is that the ITPase enzyme acts to reduce the amount of active forms of the 6-MP drug present in human cells.
Genotyping methods can reliably detect the major and rare mutant alleles at human ITPA locus, in particular when genetic polymorphism is highly likely to provide an explanation for ITPA deficiency in individuals. characterization of ITPA deficiency by genotyping for the most common inactivating single-nucleotide polymorphisms can prospectively identify patients at higher risk of mercaptopurine toxicity which can subsequently influence the outcome of the patients.
This study was done to determine ITPA polymorphisms 94C>A (rs1127354) and IVS2+21A>C (rs7270101) with RQ-PCR using TaqMan SNP genotyping assay and carried out on 80 ALL pediatric patients admitted to Alexandria University Children’s Hospital and assess their relations with the development of 6MP drug toxicity and outcome.
All the patients were subjected to full history taking, complete clinical examination & laboratory investigations including: CBC, ALT, AST, bone marrow aspiration and immunophenotyping by flowcytometry, then DNA were extracted by (QIA amp Genomic blood DNA purification Mini kit QIAGEN, Cat.no 51104 Germany) extraction kit from peripheral blood samples and quantified using Nano DROP 2000 Spectrophotometer. The purified DNA genotyped with RQ-PCR system using TaqMan SNP genotyping assay (Applied biosystems -Life Technologies, USA Cat.no 4351379,) on Rotor gene Q.
The results of the present study of IVS2+21AC ITPA gene SNP (rs7270101) were 68.8% of wild-type homozygous AA genotype, 27.5% of variant type heterozygous AC genotype, 3.8% of variant type homozygous CC genotype while the results of 94CA ITPA gene SNP (rs1127354) were 97.5% of wild-type homozygous CC genotype, 2.5% of variant type heterozygous AC genotype, 0% of variant type homozygous AA genotype.
There was no statistically significant difference between ITPA genotypes and 6-MP toxicity and overall survival while relapse free survival was lower in ITPA 94CA (rs1127354) variant type than wild type.
CONCLUSIONS
In conclusion, our study provides suggestive evidence that:
1. All ITPA SNPs genotype frequencies were normally equilibrated with Hardy-Weinberg equation.
2. The homozygous wild type ITPA rs7270101 AA genotype and homozygous wild type ITPA rs1127354 CC genotype were the most frequent genotypes.
3. The heterozygous variant type ITPA rs7270101 AC genotype and heterozygous variant type ITPA rs1127354 AC were the most prevalent mutant genotypes in this study.
4. ITPA polymorphisms 94C>A (rs1127354) was associated with lower relapse free survival.
5. No relation was detected between the different studied ITPA SNPs and 6-MP toxicity and overall survival.