|Ahead of print
Genetic polymorphism of hOGG1 ser326cys and its association with breast cancer in Jammu and Kashmir
Ashna Nagpal1, Sonali Verma1, Ruchi Shah1, Ghulam R Bhat1, Amrita Bhat1, Divya Bakshi1, Bhanu Sharma1, Sandeep Kaul2, Rakesh Kumar1
1 School of Biotechnology, Shri Mata Vaishno Devi University, Katra, Jammu and Kashmir, India
2 Department of Oncology, Shri Mata Vaishno Devi Superspeciality Narayana Hospital, Katra, Jammu and Kashmir, India
|Date of Submission||16-Oct-2018|
|Date of Decision||01-Nov-2018|
|Date of Acceptance||06-Feb-2019|
|Date of Web Publication||09-Mar-2020|
School of Biotechnology, Shri Mata Vaishno Devi University, Katra, Jammu and Kashmir
Source of Support: None, Conflict of Interest: None
Background: 8-Oxo-7, 8-dihydro-2'-deoxyguanosine (8-oxodG) is a potent DNA damage marker that leads to cellular oxidative stress. It is a DNA-repair enzyme that participates in “8-oxodG” DNA adducts removal. Previous studies show weak associations of rs1052133 (hOGG1) in breast cancer patients of Northern India. We performed this study to explore the variant rs1052133 (hOGG1) with breast in the population of Jammu and Kashmir (J and K).
Method: A polymerase chain reaction–restriction fragment length polymorphism -based single-nucleotide polymorphism (SNP) genotypic study was carried out in peripheral blood samples of 165 breast cancer patients and 200 healthy controls, using specific primers. Sanger sequencing verified the results.
Results: hOGG1-Ser326Cys polymorphism occurred frequently in cases as compared with controls. Data were evaluated by SPSS V.13 software, following Hardy–Weinberg equilibrium (P = 0.002 at OR 2.57; 95% CI [1.68–3.93]), which showed that the SNP rs1052133 had a significant association with increased risk of breast cancer.
Conclusion: Overall, the results of this analysis show that the hOGG1-Ser326Cys polymorphism may be associated with an increased risk for breast cancer in the J and K population.
Keywords: 8-oxodG, Base excision repair, DNA repair, human 8-oxoguanine DNA glycosylase, oxidative stress, reactive oxygen species
|How to cite this URL:|
Nagpal A, Verma S, Shah R, Bhat GR, Bhat A, Bakshi D, Sharma B, Kaul S, Kumar R. Genetic polymorphism of hOGG1 ser326cys and its association with breast cancer in Jammu and Kashmir. Indian J Cancer [Epub ahead of print] [cited 2020 Apr 1]. Available from: http://www.indianjcancer.com/preprintarticle.asp?id=280299
| » Introduction|| |
Breast cancer is one of the leading causes of death in women throughout the world. Various genome-wide association studies' analyses found that rs1052133 of the hOGG1 gene is associated with human cancers., The hOGG1 gene plays an important role in the base excision repair (BER) pathway. The main role of the hOGG1 gene in the BER pathway is to recognize the lesion 8-hydroxy-2'-deoxyguanine (8-OH-dG) and its excision. It acts as a scaffold for other DNA repair genes like XRCC1, APE1, Pol β, and LIG3. Environmental exposure and dietary habits influence the function of these genes that increases the breast cancer risk. A similar case–control study in colorectal carcinoma found no association in the Kashmiri population. Saudi and Polish studies have found hOGG1-Ser326Cys to be strongly associated with breast cancer in their respective populations. In India, a few studies have shown its association with breast cancer.,,, We designed this study to explore the variant rs1052133 (hOGG1) with breast cancer in the population of the Jammu and Kashmir (J and K), India.
| » Materials and Methods|| |
The study recruited 165 breast carcinoma patients and 200 healthy ethnicity-matched and age-matched women. Clinical details are given in [Table 1]. 2-3 ml of blood was collected from breast patients and normal healthy females (age and sex matched) of Jammu and Kashmir region and controls were collected by random sampling and stored in Ethylenediamine tetraacetic acid (EDTA) vials at –20°C until the time of DNA isolation. The genomic DNA was extracted using the manufacturer protocol of DNA Isolation Kit (Qiagen Kit). DNA concentration and quality were checked using a Nano-Drop Bio spectrophotometer (Eppendorf India Limited, Ambattur, Chennai) and agarose gel electrophoresis (Bio-Rad and Gel Doc EZ image-Bio-Rad House, Gurugram, Haryana), respectively.
The hOGG1 Ser326Cys polymorphism was assessed by polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) using primers from a previous study. The reaction mixture of PCR having 25 μL (total PCR reaction mix is 25μL) volume contains 0.5 μL Forward and Reverse Primer, 2.0 μL (50 ng/μL) of genomic DNA, 9.5 μL dH2O, and 12.5 μL master mix (MgCl2, dNTPs, Taq Polymerase) (Sigma Aldrich Srivastava Market, Hauz Khas, Delhi). For the PCR amplification process in a thermal cycler (Eppendorf India, Ambattur, Chennai), each step consisted of a 5-min denaturation at 94°C followed by 35 cycles of 94°C for 30 s, annealing at 58°C for 30 s and extension at 72°C for 1 min, and 72°C for 7 min. We used RFLP method to identify the Ser326Cys variant because the C-to-G transversion creates a new Fnu4HI (NEBL) restriction site (Thermo Fisher Scientific Pvt. Ltd, Hiranandani Business Park, Mumbai, Maharashtra). Fragments were separated on a 2% agarose gel and results were validated by Sanger Sequencing (Ms. Agri Genome Pvt. Ltd, Cochin and Kerala, India). The chromatograms were analyzed by ChromasPro V.2.6.4.
| » Results|| |
The 200 bp PCR product is digested by Sat1 (Fnu4HI) creating two 100 bp fragments for the 326Cys variant and is undigested for the 326Ser allele at exon 7. The chromatograms showed Ser/Cys heterozygote (C/G), Ser/Ser homozygote (C/C), and Cys/Cys homozygote (G/G) [Figure 1]. SNP polymorphism at Ser326Cys (rs1052133) at exon7 of chr3p25 positioned at codon326 in the hOGG1 gene had a significant association with breast cancer. The analysis was done in 200 healthy controls and 165 breast cancer patients using SPSS V.13 software, following Hardy–Weinberg equilibrium (HWE). Genotypic distribution, risk allele frequency, risk estimation, and association details are summarized in [Table 1]. The observed odds ratio of risk allele 'G' is 1.24 (0.71–2.16) and HWE = 0.893 for cases and 0.446 in controls, that predisposes them to a greater risk of having breast cancer, because of oxidative DNA damage. The covariates taken for the analysis included age and BMI.
|Figure 1: Representative chromatogram and gel picture showing Ser326Cys SNP (rs1052133), C to G transversion|
Click here to view
| » Discussion|| |
We found an association of rs1052133 of the hOGG1 gene with breast cancer in the J and K population and it has highlighted the role of the gene in a predisposition towards breast cancer in the population. The hOGG1 Ser326Cys was selected because of its critical role in genome integrity. It was reported that hOGG1 Ser326 Cys has deleterious effect on different cancers world wide. In Asian, Caucasian, and Chinese populations, this SNP showed a strong association with cancers like ovarian, lung, prostate, gastric etc., In Polish women, it showed a strong association with breast cancer and is even used as a biomarker in breast cancer patients. Pakistani women showed a strong association of this SNP with breast cancer. The variant rs1052133 under study also showed a strong association with breast cancer in the Indian population, whereas the study in the ethnic Kashmiri population of India in colorectal cancer had no association of this SNP. No such study has been carried in breast cancer patients of the J and K region.
Alanazi et al. found protein structural destabilization of OGG1 mutant 326cys on simulation. Mutant residues are more hydrophobic and have deleterious effects on the function of the hOGG1 protein by affecting the enzymatic activities of DNA glycosylases, lyase, and hydrolase. Hill and Evans et al. conducted in vivo functional studies using deletion mutants and confirmed the DNA-binding and enzymatic activity of hOGG1. Simonelli et al. found the lower enzymatic activity and DNA damage and repair potential of purified Cys326Cys mutant protein in an in vitro base excision assay. In this case–control study, Ser326Cys SNP was found to be significantly associated with the increased risk of breast cancer. The current findings suggested that allele G of SNP rs1052133 act as a risk factor in females of the J and K region. This is the first report from the J and K population, which shows strong association with breast cancer. In the future, it may act as a biomarker for breast cancer screening programmes.
The authors thank the DST and Govt. of Jammu and Kashmir for funding this project. Funding for this study was sanctioned by DST-SERB, GOI with the sanctioned number: YSS/2014/000659.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
The present study was approved by the Institutional Ethics Review Board [IERB] of Shri Mata Vaishno Devi University with approved serial no. SMVDU/IERB/14/28.
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