Genetic Polymorphisms of Glutathione- S-Transferase and N-Acetyltransferase-2 among Sudanese Patients with Acute Lymphoblastic Leukemia

Abstract

Leukaemia represents the second type of the ten top most common cancers in Sudan, and acute lymphoblastic leukaemia (ALL) is the first malignancy of childhood cancer in Sudan. This study aimed to study the genetic plymrphism of N-acetyltransferase-2 (NAT2) {TagG590A (rs1799930), Dde A803 (rs1208), BamH1 G857A (rs1801279) and Kpn1C481T (rs1799929)} and glutathione S-transferases (GSTT1, M1 and P1). And to investigate the haematological picture and to determine the frequency of ALL phenotypes. Has been associated with acute lymphocytic leukemia (ALL) in the different regions of the Sudan. The blood samples are collected from 300, people of whom 150 were patients arriving to Khartoum Oncology Hospital (KOH) and the remaining 150 were healthy matched volunteered controls. Were genotyped for GSTs and NAT2 in the study from 2015 to 2018. Sequence technique to PCR product for 55 (NAT2/547bp) patients and controls samples and also 11(GSTP1) patients samples by Sanger Sequencing (BGI TECH SOLU TIONS (HONGKONG), CO. China). Bioinformatics has done using various publicly available soft ware's (Sift, PolyPhen-2 I Mutant 3.0, and Chimera)the effect of each SNP was predicted. The mean age of all subjects was 13.6 years. Their sex (63.8%) males and female (36.2%). The incidence of ALL among the adolescent group (7-18 years of age) was slightly higher (41.2%) than that registered among children group (1-6 years) (32.2%) and elderly group (>18 years of age) (22.6%). The majority of ALL patients showed ‘B’ immune- phenotype (75.5%). Afro-Asiatic linguistic ethnic was the most popular group (72%). According to residence, the high frequencies were reported in Khartoum (22%), Kordofan (17%), AL-Jazira and Darfur (~12% each). A significant association between affected tribes and disease frequency at states (P=0.000) was reported. The mean of the complete blood count parameters of ALL patients after treatment (Hb=10±1g/dl, RBC=4±1x106/ml, HCT34±1%, PLT 269± 125x109/L) was signific- antly lower (p =0.00) than that in controls (Hb= 12±1g/dl, RBC=5± 5.2 106 / ml, HCT36±3%, PLT 353±8x109/L) and the other parameters (WBC=19±51x103/ml, MCV=84±12fl, MCH=28±2pg, Blast= 8±21%) was significantly higher (p=0.00) than that in control (WBC=7.6 ±2x103/ml, MCV =81±3fl, MCH=27±1pg, Blast=0.00%). Regarding genetic analysis, the risk of ALL in patients with GSTT1 null genotype was not significant (OR=1.23, 95% CI=0.74 -2.0, P-0.44). The GSTM1 null genotype was significantly higher in the ALL group (79, 60.3 %) compared to controls (52, 39.78%) and increased the risk to ALL by about two folds (OR= 1.72, 95% CI=1.1-2.7, p=0.03). The frequency of GSTP1 genotypes was also significantly different between the two groups (P=0.000). AG, GG and AG+GG genotypes were found to increase the risk of ALL by more than three times (P<0.000) compared to the AA genotype. Allele frequencies were also significantly different (P= 0.001, OR= 1.79, CI= 1.247-2.58). In bioinformatics results for GSTP1, (Ile/Val 105; rs1695) SNP was deleterious and probably damaging by using SIFT and Polphen-2 software, respectively. The protein of GSTP1 changed from isoleucine to valine at codon 105 by the Chimera prediction software. The NAT2-Tag G590A (rs1799930) genotype was significantly different between the two groups (p=0.01). The AA and AA+AG genotypes were found to increase the risk of ALL by about two folds (P=0.01, 0.03, respectively) compared to the GG genotype. Allele frequencies were also significantly different (P=0.00) between ALL cases and controls. The Dde A803G (rs1208) genotype was significantly different between the two groups (p=0.02). GG genotype was found to increase the risk of ALL by more than two times (p=0.02) compared to the AA genotype. Allele frequencies were also significantly different (OR=1.16, 95% CI=0.77-1.74, P=0.01).The BamH1 G857A (rs1801279) genotype was significantly different between the two groups (p=0.00). The AA and AA+AG genotypes were found to increase the risk of ALL by about three times (P=0.01), compared to the GG genotype. Allele frequencies were also significantly different (P=0.00, OR=1.9, 95% CI= 1.3-2.84). Regarding Kpn1 C481T (rs1799929) genotype and allele frequencies, they were insignificant different between the two groups. The additive model exposed that SNPs with a significant association with ALL were Tag G590A (rs1799930) and BamH1 G857A (rs1801279) (OR=1.5, p=0.01). Sequence analysis (Multiple Alignment) shows changes of nucleotide from C to T at Kpn1 C481T (rs1799929) SNP and from G to A at Tag G590A (rs17 99930) and BamH1G857A (rs1801279). In bioinformatics analysis, the Tag G590A (rs179 9930) and BamH1 G857A(rs1801279)ware predicted as deleterious, probably damaging and decrease protein stability using the Sift, Poly- Phen-2 and I Mutant 3.0 soft wares respectively. Project HOPE showed that the three-dimensional structure of proteins was changed by homology modelling server from arginine (R) to glutamine (Q) at position 197 (rs1799930) and 64 (rs1 80 1279). Regarding the two other SNPs, they have no deleterious effect with the same software. The study concludes that, ALL among Sudanese children shows male predominance, with significant different males in haematological parameters than unaffected children. The more affected age group with ALL is 7-18 years, and B-phenotype represented high-frequency rate. This study indicates that GST (M1 and P1) and NAT2 (Tag (G590A), Dde (A803G) and BamH1 (G857A) mutant genotype exhibit significant association with the risk of developing ALL among Sudanese patients. The Western and Central Sudan recorded the highest rates of ALL. Most of ALL patients are from the Afro-Asiatic ethnic group.