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Higher order genes interaction in DNA repair and cytokine genes polymorphism and risk to lung cancer in North Indians

1 Department of Biochemistry, University of Allahabad, Allahabad, Uttar Pradesh, India
2 Department of Zoology, University of Lucknow, Lucknow, Uttar Pradesh, India
3 Department of Radiation Oncology, Kamala Nehru Memorial Hospital, Allahabad, Uttar Pradesh, India
4 Narayana Translational Research Centre, Narayana Medical College, Nellore, Andhra Pradesh, India
5 Department of Biological Sciences, Cleveland State University, Cleveland, Ohio, United States of America

Date of Submission14-Jan-2020
Date of Decision14-Jul-2020
Date of Acceptance12-Aug-2020
Date of Web Publication23-Oct-2021

Correspondence Address:
Munish Kumar,
Assistant Professor, Department of Biochemistry, University of Allahabad, Allahabad - 211 002, Uttar Pradesh
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jcrt.JCRT_51_20

 > Abstract 

Context: Lung cancer pathological process involves cumulative effects exerted by gene polymorphism(s), epigenetic modifications, and alterations in DNA repair machinery. Further, DNA damage due to oxidative stress, chronic inflammation, and the interplay between genetic and environmental factors is also an etiologic milieu of this malignant disease.
Aims: The present study aims to assess the prognostic value of DNA repair, cytokines, and GST gene polymorphism in lung cancer patients who had not received any neoadjuvant therapy.
Materials and Methods: In this case–control study, 127 cases and 120 controls were enrolled. DNA from the blood samples of both patients and controls was used to genotype XRCC1Arg399Gln, XPDLys751Gln, and interleukin-1 (IL-1β) genes by polymerase chain reaction (PCR)-restriction fragment length polymorphism method, whereas multiplex PCR was performed to genotype GSTT1 and GSTM1.
Results: Binary logistic regression analysis showed that XRCC1Arg399Gln-mutant genotype (Gln/Gln, odds ratio [OR] = 4.6, 95% confidence interval [CI]: 2.2–9.6) and GSTT1 null (OR = 2.7, 95% CI: 1.6–4.5) were linked to cancer susceptibility. Generalized multidimensional reduction analysis of higher order gene–gene interaction using cross-validation testing (CVT) accuracy showed that GSTT1 (CVT 0.62, P = 0.001), XPD751 and IL- (CVT 0.6, P = 0.001), and XRCC1399, XPD751, and interleukin-1 receptor antagonists (IL-1RN) (CVT 0.98, P = 0.001) were single-, two-, and three-factor best model predicted, respectively, for lung cancer risk. Classification and regression tree analysis results showed that terminal nodes which contain XRCC1399-mutant genotype (AA) had increased the risk to lung cancer.
Conclusion: The present study demonstrated that XRCC1399 (Gln/Gln), GSTT1, and IL-1RN allele I, I/II served as the risk genotypes. These genes could serve as the biomarkers to predict lung cancer risk.

Keywords: Classification and regression tree analysis, cytokine, generalized multifactor dimensionality reduction, genetic polymorphism, GSTM1, GSTT1, lung cancer, XPD, XRCC1

How to cite this URL:
Ritambhara, Kumar R, Gupta MK, Gautam P, Tiwari S, Vijayraghavalu S, Shukla GC, Kumar M. Higher order genes interaction in DNA repair and cytokine genes polymorphism and risk to lung cancer in North Indians. J Can Res Ther [Epub ahead of print] [cited 2022 Aug 16]. Available from: https://www.cancerjournal.net/preprintarticle.asp?id=329055

 > Introduction Top

Lung cancer is the third most common malignancy reported across the world. In the present scenario, lung cancer (18.4%) accounts for the highest mortality rate among all cancer types and the 5-year prevalence is 56.6% higher than other cancers.[1] Genetic makeups in an individual have the unique polymorphic trait of detoxification enzymes and DNA repair genes, and their functions are modulated by environmental exposures. Phase II detoxification enzymes include GSTT1 and GSTM1 involved in the activation of carcinogens. Deletion polymorphism of GSTT1 and GSTM1 is associated with lung cancer risk.[2],[3] DNA repair mechanism is one of the most versatile systems of the human cells to combat new mutations, environmental exposures, and carcinogenic compounds. Earlier studies had shown the interplay between the following genes ERCC2/XPD involved in the nucleotide excision repair (NER) pathway and XRCC1 in the base excision repair (BER) pathway to increase the cancer risk.[4]

The XRCC1 functions as a scaffold protein coordinated with DNA ligase III and DNA polymerase β to target damaged sites. XRCC1 protein is involved with several BER proteins to prevent DNA strand break and prevent deficiency in DNA repair. XRCC1Arg399Gln found to be strongly correlated to increased DNA damage, leading to reduced activity of XRCC1 scaffold protein.[5] Single-nucleotide polymorphisms (>10%) in XRCC1 codon 399 lead to amino acid changes and thereby result in compromised DNA repair activity.[6]

The XPD gene coexisting with XRCC1 on chromosome (Chr) 19q13.2–q13.3[7] functions as helicase unwinds DNA in 5/–3/direction. Hence, mutations in this gene downregulate its helicase activity and affect the basal transcription level and NER pathway.[8],[9],[10]

Interleukin-1 (IL-1) cytokine gene family consists of interleukin-1beta (IL-1β) and interleukin-1 receptor antagonists (IL-1RN), which are parts of the innate immune system and are involved in inflammatory responses. Both IL-1β and IL-1RN are located on Chr 2 within 430-kb gene segment. IL-1β is pro-inflammatory and is produced by monocytes and tissue macrophage.[11] Various other cytokines are also found in lung cancer tumor microenvironment, but cellular and molecular functions are not well established.[12] Cytokine gene polymorphism of IL-1β511C/T showed an increased risk in lung cancer as well as chronic obstructive pulmonary disease.[13],[14] IL-1RN is an anti-inflammatory cytokine that has 86 bp variable number tandem repeats (VNTR) polymorphism at intron 2 region located on Chr 2q14.2. The IL-1RN is an antagonist of IL-1β and competitively binds to the same membrane receptor of IL-1β.[15],[16] Inflammatory cytokine genes showed a strong association with lung cancer risk; it was reported that circulating free cytokine levels were increased in Non-Small Cell Lung Cancer (NSCLC) patients.[17] The prime focus of the present study is to assess the prognostic value and risk associated to phase II enzymes, DNA repair genes, and cytokine gene polymorphism in patients who had not undergone any neoadjuvant therapy. There are many studies reported on the status of these enzymes on patients who underwent this chemo-radiotherapy but only a few reports at the prognostic level. DNA repair genes XRCC1 rs25487 (BER pathway), XPD rs13181 (NER pathway) and cytokine genes IL-1β, IL-1RN were selected because either they lie in the coding region or promoter region. Both the DNA repair genes and cytokine genes co-exist on the same Chr, which might be responsible for increased susceptibility to lung cancer risk.

 > Materials and Methods Top

Study design and subjects

The study population consists of 247 participants, including 120 healthy controls and 127 lung cancer patients. This study was performed after ethical approval from the population resource and research center, Allahabad Institute Ethical Committee (IERB Reference: 18/9.39). The study was conducted from February 2017 to July 2019. The protocol followed for this study was according to the Declaration of Helsinki provisions. All the participants recruited in this study were given informed consent for withdrawing blood samples. A detailed questionnaire was prepared in both local language and English for all the study subjects, including information of occupational history, residence, dietary habits, smoking, and history of malignancy. All the lung cancer patients recruited in the present case–control study had not undergone any prior neoadjuvant therapy. The inclusion criteria for lung cancer patients were North Indian ethnicity, age range between 18 and 75 years, histopathologically confirmed biopsy of lung cancer, and lung cancer patients who have more than 10 pack-year of smoking. Inclusion criteria for healthy controls were age-, sex-, and ethnicity-matched individuals without any reported cancer history. The exclusion criteria for both the cases and controls were comorbid conditions such as other diseases, double metastasis, and age more than 75 years.

Genomic DNA extraction

Genomic DNA was extracted from 2 ml of the peripheral blood lymphocytes of lung cancer patients and controls using Qiagen DNA blood mini isolation kit (Germany) according to the manufacturers' protocol and its concentration quantified on a NanoDrop spectrophotometer (Agilent Technologies, Santa Clara, USA).


The genotyping was performed for XRCC1 codon 399 genotyping was performed according to earlier method[5][Figure 1]. The XPD codon 751 genotyping was done byPCR-fragment length polymorphism (PCR-RFLP) as described as earlier method[18] [Figure 2]. Genotyping of IL-1β-511C/T and IL-1RN polymorphisms performed according to earlier method[19] [Figure 3]. IL-1RN 86bp repeats in Intron 2 was genotyped using polymerase chain reaction (PCR). The amplified product of IL-1RN was electrophoresed [Figure 4] stained with EtBr and visualized by Geldoc System (Biorad System, Canada). The multiplex PCR method was used to assess GSTM1 and GSTT1 polymorphism, according to earlier method[2] [Figure 5]. The 2X PCR master mix and primer sequences were added together to make the PCR reaction volume [Table 1].
Figure 1: Polymerase chain reaction restriction fragment length polymorphism analysis of XRCC1399Gln. Lane M showing 100 bp DNA ladder; lane 1 and 4 showing Arg/Gln; and lane 2, 3, 5, and 6 showing Arg/Arg genotype

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Figure 2: Polymerase chain reaction-restriction fragment length polymorphism analysis of XPD751Gln. Lane M showing 100 bp DNAladder; lane 2, 4, and 6 showing Lys/Lys; lane 3 and 5 Lys/Gln;andlane 7 showing Gln/Gln genotype

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Figure 3: Polymerase chain reaction-restriction fragment length polymorphism analysis of IL-1β-511C/T. Lane M showing 100 bp DNA ladder; lane 1, 3, 5, and 7 showing TT genotype; lane 2, 4, and 6 showing CC genotype

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Figure 4: Representative 2% gel electrophoresis of 86 bp variable number tandem repeats in intron 2IL-1RN showing lane M 100 bp DNA ladder; lane 1, 3 and 5 alleles I; lane 2 and 6 I/II allele; and lane 4 II allele

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Figure 5: 2% gel electrophoresis multiplex polymerase chain reaction of GSTT1 and GSTM1 lane M 100 bp DNA ladder; lane 2,3 and 5 showing both, GSTT1 and GSTM1 genotypes; lane 6 showed both null genotypes; lane 4 and 7 showing deletion of GSTT1

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Table 1: Characteristic features of selected gene polymorphism of DNA repair, cytokine gene, and xenobiotic-metabolizing genes and primer sequences

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Statistical analysis

The minor allele frequency (MAF) of XRCC1Arg399Gln, XPDLys751Gln, IL-1β511C/T, IL-1RN, GSTT1, and GSTM1 considered for the study using QUANTO 1.1 (Open source software) for sample size analysis. The present case–control study (127 lung cancer cases and 120 controls) was adequate to give 80% power. The Chi-square test was performed to estimate the difference between lung cancer cases and control with distribution to gender and smoking status including smokers and nonsmokers. The Chi-square test was also performed to analyze the Hardy–Weinberg equilibrium (HWE) in the control group. Binary logistic regression was performed to assess the odds ratio (OR) and 95% confidence interval (CI), considering wild-type homozygote as a reference group. SPSS 16.0 Software Packages (SPSS, Chicago, IL, USA) was used for the test assessment. Based on the observed genotype frequencies, we performed haplotype analysis by using the SHEsis analysis platform.[20] To further evaluate gene–gene and gene–environment interaction, generalized multidimensional reduction (GMDR) analysis was performed.[21] GMDR method based on the generalized linear model is suitable to assess both dichotomous and quantitative phenotype with adjustment to covariates. The characterization of the best interaction model was based on maximum cross-validation testing (CVT) accuracy and cross-validation consistency (CVC). The P-value of 0.05 is considered as statistically significant.

 > Results Top

Demographic variables in lung cancer cases and controls

The mean age differences of lung cancer patients (58 ± 11) were higher than the controls (53 ± 11). The frequency of smokers in lung cancer patients (91%) was higher than controls (30%) and statistically significant. The percentage distribution of histology in lung cancer patients was squamous cell carcinoma (SQCC, 24), adenocarcinoma cell carcinoma (ADCC, 62), and large cell carcinoma (13) [Table 2].
Table 2: Baseline demographic variables of lung cancer patients and controls

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Risk estimation of xenobiotic-metabolizing enzymes, DNA repair gene, and cytokine gene polymorphism

The XRCC1 codon 399 gene showed association to lung cancer susceptibility (OR = 4.6; 95% CI: 2.2–9.6) which was statistically significant, P < 0.001. The genotype Lys/Gln showed a protective effect in lung cancer patients (OR = 0.42; 95% CI: 0.24–0.73) with statistical significance value (P = 0.002). The genotype combination of Lys/Gln+Gln/Gln also showed protective effect (OR = 0.57; 95% CI: 0.34–0.95) with P = 0.03 [Table 3]. The 86 bp VNTR of IL-1RN genotypic distributions of allele II showed no exposure to lung cancer risk and found statistically significant (OR = 0.42; 95% CI: 0.21–0.84, P = 0.01) [Table 3]. The null genotype of GSTT1 showed two times increased risk toward lung cancer (OR = 2.6; 95% CI: 1.5–4.4, P < 0.001) [Table 3]. GSTM1 showed no association to lung cancer risk (OR = 1.3; 95% CI: 0.7–2.1, P = 0.280) [Table 3]. The allele frequency estimation showed XRCC1399 codon significantly associated with lung cancer risk (OR = 2.3; 95% CI: 1.6–1.3, P < 0.001). 86 bp VNTR polymorphism in the intron 2 IL-1RN allele frequency showed significant association to lung cancer (P = 0.004) [Table 4]. The MAF of the following genes XRCC1399, XPD751, GSTT1, and GSTM1 was higher in lung cancer patients oncomparison to healthy controls recruited, in contrast thegene - IL-1β-511C/T was higher in control population thanpatients in this study [Table 5]. XRCC1Arg399Gln (P < 0.001), XPDLys751Gln (P = 0.001), and IL-1RN (P = 0.02) were in HWE [Table 5].
Table 3: Genotypic frequencies of XRCC1399Gln, XPD751Gln, IL-1β-511C/T, IL-1RN, GSTT1, and GSTM1 in cases and controls

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Table 4: Allelic frequencies of XRCC1399Gln, XPD751Gln, IL-1β-511C/T, IL-1RN, GSTT1, and GSTM1 in cases and controls

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Table 5: Minor allele frequencies in cases and control and two-sided Chi-square test for genotypic distribution and allele frequencies

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 > Haplotype analysis Top

The four-haplotype combination constructed for XRCC1399 codon and XPD751 codon showed that A-A (Gln-Gln) combination of XRCC1399Gln and XPD751Gln (OR = 3.5; 95% CI: 2.1–5.7) has 3.5 times increased risk associated to lung cancer. The haplotype combination of XRCC1399Gln and XPD751Gln G-A (Arg-Gln) showed reduced risk (OR = 0.4; 95% CI: 0.3–0.6). In cytokine gene polymorphism, IL-1β-511-C/T and IL-1RN showed two-haplotype combination in which T-I (IL-1β-511-C/T-IL-1RN) was significantly associated to reduced risk (OR = 0.6; 95% CI: 0.4–0.8) for lung cancer. I/II haplotype combination (IL-1β-511C/T-IL-1RN) showed significant association (OR = 1.5; 95% CI: 1.0–2.2) to lung cancer [Table 6].
Table 6: Haplotype frequencies of XRCC1399Gln, XPD751Gln, IL-1β-511C/T, and IL-1RN and pairwise linkage disequilibrium Haploview (SHEsis, ver. online)

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Higher order gene–gene and gene–environment interaction models by generalized multidimensional reduction analysis

The best interaction models for gene–gene interaction analysis were performed with the help of GMDR (GMDR software Beta 0.9, Free open source interaction analysis tool). GMDR analysis constructed the best model for one, two, and three factors. The three-factor model – XRCC1Arg399Gln, XPDLys751Gln, and IL-1RN [Figure 6]c was found the best model with the association to lung cancer risk with a testing accuracy of 0.87 (P = 0.001) [Table 7]. The best two-factor model XPDLys751Gln and IL-1β-511-C/T [Figure 6]b with overall testing accuracy of 0.63 was significantly associated with lung cancer (P = 0.001) [Table 7]. The single-factor model of GSTT1 [Figure 6]a showed the highest CVC (CVT = 0.62, CVC = 10/10, P = 0.001) [Table 7]. The distribution of null genotypes of GSTT1 lung cancer cases was lower comparatively to controls in single-factor model of GSTT1. The best interaction model for SQCC histology of lung cancer was predicted as a single-factor XPD751 [Figure 7]a and two-factor model XPD751/IL-1β [Figure 7]b. The single-factor model XPD codon 751 served as the best prediction model for SQCC with the highest CVC (CVT = 0.51, CVC = 10/10, P = 0.001) [Table 7]. The best two-factor model constructed of XPD751, IL-1β with testing accuracy (CVT = 0.70, CVC = 9/10, P = 0.01) showed significant association to SQCC [Table 7].
Figure 6: Generalized multidimensional reduction analysis showing best prediction model with association to lung cancer risk, high-risk cells are represented by dark shading and low-risk cell are represented by light shading. (a) Single-factor model of GSTT1 for lung cancer risk. (b) Two factor model of XPD751Gln and IL-1β to increased risk of lung cancer. (c) Three-factor model of XRCC1399Gln, XPD751Gln, and IL-1RN with risk to lung cancer

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Figure 7: Generalized multidimensional reduction analysis showing interaction model with an association to lung cancer histology of squamous cell carcinoma. (a) Single-factor model of XPDLys751Gln with the association to the risk of squamous cell carcinoma. (b) Two factor model of XPD751Gln and IL-1β-511C/T with increased risk of lung cancer in squamous cell carcinoma

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Table 7: Gene–gene and gene–environment interaction model by generalized multidimensional reduction analysis

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Classification and regression tree analysis

The tree construction based on the classification and regression tree (CART) model was performed on CART software (version 6.0, Salford Systems, San Diego, California, USA) using gene–gene and gene–environment factors [Figure 8]a and [Figure 8]b. The first root node 1 split was XRCC1399, genotype (AA) (OR = 3.26; 95% CI: 1.11–9.59, P = 0.03) showed clearly that it is strongly associated withlung cancer risk. Node 3 showed significant risk to lung cancer consisting of XPD751 (AA)/IL-1RN (I/II) XRCC1399 (GG, AA) (OR = 4.34; 95% CI: 1.35–13.92, P = 0.01). Terminal node 2 comprised of XPD751 (AA, AC)/IL-1RN (II) (OR = 12.13; 95% CI: 3.13–46.94, P < 0.001) stated its association to lung cancer [Table 8]. The tree construction based on smoking showed that IL-1RN (I, I/II) in node 1 showed strong risk (OR = 4.76; 95% CI: 1.51–15.03, P = 0.008) in lung cancer patients [Figure 8]b and [Table 8].
Figure 8: Classification and regression tree analysis displaying gene–gene and gene–environment risk factor associated with lung cancer. The blue boxes are the nodes which can be further divided into other nodes. The red boxes are terminal nodes which cannot divide further. (a) Gene–gene interaction, (b) gene–environment interaction

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Table 8: Risk estimates in lung cancer patients based on classification and regression tree analysis

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 > Discussion Top

In the present case–control study, we assess whether the DNA repair gene (XRCC1Arg399Gln, XPDLys751Gln), cytokines (IL-1β, IL-1RN), and GST (GSTM1, GSTT1) gene polymorphism associated to lung cancer risk among North Indian population. XRCC1 prominently involved in the BER pathway, Gln/Gln-mutant variant of XRCC1399Gln, was associated with higher sister chromatid exchange and DNA damage.[5] There is various polymorphism reported for XRCC1 in which XRCC1399Gln showed that genotypes Gln/Gln and Arg/Gln having increased risk to lung cancer and served as a biomarker in the prognosis of lung cancer.[22] XRCC1 repair efficiency was found to be affected in lung cancer patients carrying XRCC1 Arg/Gln and Arg/Gln+Gln/Gln.[23] In lung cancer patients of Northeast India, mutant genotype (Gln/Gln) of XRCC1Arg399Gln and XPDLys751Gln showed association (OR = 3.3, 95% CI: 1.2–8.5), and in our study performed in North Indian population, it was reported increased risk of lung cancer with XRCC1399 codon Gln/Gln whereas protective effect of Lys/Gln+Gln/Gln toward lung cancer.[24] Similar to our findings; an earlier study in East Chinese Han population, reported that the people with Gln/Gln genotype of XRCC1399 shown two-fold increased lung cancer risk.[25] DNA repair studies done in NSCLC tissues revealed increased expression of XRCC1 gene than in healthy lung tissues.[26] In an European case control study also reported an association of XRCC1399 codon Gln/Gln genotype with increased risk for squamous cell carcinoma (SQCC) of the lungs.[27] Similarly, a study done in South Indian lung cancer patients also shown strong association of XRCC1399 Gln/Gln genotype to the lung cancer risk.[22] These studies strengthens our findings that impaired activity of XRCC1399Gln results more mutant. A meta-analysis report suggested that combined Arg/Gln+ Gln/Gln variant of codon 399 XRCC1 has risk associated to lung cancer[28], however we found risk associated to Gln/Gln genotype in North Indian Population, results from different research group also shown increased lung cancer susceptibility in North Indian Population with Gln/Gln genotype.[29] In a study, it was shown that haplotypes of XRCC1399 codon polymorphism sequence GCC (OR = 1.6, 95% CI: 1.1–2.2) had increased risk toward lung cancer, whereas we found that gene–gene interaction of XRCC1399Gln and XPD751Gln sequence Gln-Gln showed threefold increased risk to lung cancer.[30] In a recent study, it was reported that XRCC1399 Gln/Gln genotype (OR = 1.8, 95% CI = 1.3–2.5) has a strong association with lung cancer risk, similar to our findings. They reported that XPD751 Gln/Gln associated (OR = 1.4, 95% CI: 1.1–1.7) with lung cancer risk, whereas we found that Lys/Gln genotype and Lys/Gln + Gln/Gln combination showed protective effects toward lung cancer risk. They also performed GMDR analysis and showed that XRCC1Arg399Gln and XPDLys751Gln was the best two-factor model (CVC = 10/10, CVT = 71.28), whereas in our study, we found XRCC1Arg399Gln, XPDLys751Gln, and IL-1RN was the best three-factor model.[31] Another recent study on lung cancer patients using multifactor dimensionality reduction methods with DNA repair genes NBS1 (rs1805794)–XRCC1 (rs25487)–hOGG1 (rs1052133)–XPG (rs17655) (P = 0.0001); similarly, the present study finds the best prediction model with XRCC1, XPD, and IL-1RN (P = 0.001).[32]

In various cancers, XPDLys751Gln polymorphism leads to amino acid substitution of Lys to Gln found strongly associated to esophageal cancer,[33] hepatocellular carcinoma,[34] and ovarian cancer.[35] There were studies reported which lack association of XPDLys751Gln evidenced in colorectal cancer[36] and bladder cancer,[37] which was in agreement to our study. In contrary to our research, researchers reported increased susceptibility of XPD751Gln in Asians to lung cancer, whereas we discovered its protective effect toward lung cancer.[38] In meta-analysis study, XPDLys751Gln associated with increased risk of lung cancer in Caucasians with an odds ratio of 1.15. Our finding on the basis of GMDR analysis suggested that SQCC histology in lung cancer cases XPD751 codon serves as the best predicted single-factor model. In the present study, we also found XRCC1399Gln to be associated with increased cancer risk, whereas this study found no relation in Asians.[39] The study reported in early-stage NSCLC patients that XPD codon 751 found increased DNA adducts levels (204.9%, 95% CI: 0.8–822.2, P = 0.059) in SQCC subjects of lung cancer patients and we also found that XPD751Gln was a best-predicted model for SQCC subjects by GMDR method.[40] The tumor microenvironment of lung cancer has increased number of pro-inflammatory cytokine-like IL-1β and produces an immune response toward cancer cell transformation. There are factors such as tobacco smoking and air pollution, which accelerated lung carcinogenesis because of chronic inflammation. IL-1RN 86 bp VNTR polymorphism in intron 2 was reported to have three protein-binding sites and related to immune response and cancer risk.[41] Researchers showed differences in immune cell expression in lung ADCC and SQCC. They demonstrated ADCC with low immune cell expression and SQCC with higher immune cell expression; similarly, we also found with GMDR analysis that XPD751Gln and IL-1β-511-C/T were the best-predicted models in SQCC. This puts strength to our study that cytokine gene polymorphism involved in lung carcinogenesis.[42] The allele 2 of IL-1RN was found to be with the decreased association (OR = 0.68, 95% CI: 0.52–0.89) to lung cancer risk in Chinese population which was in favor to our study conducted in the North Indian population; we also found the decreased risk of IL-1RN allele 2 in lung cancer cases. IL-1RN also served as the best prediction in lung cancer risk in combination with XRCC1399Gln and XPD751Gln.[41] Recent study showed that IL-1β-511C/T has shown decreased association (OR = 0.74, 95% CI: 0.58–0.94) to lung cancer, and in our case–control study, we did not find any association to IL-1β-511C/T lung cancer risk.[43] In contrary to our finding, it was shown that IL-1β-511C/T gene associated with lung cancer (P < 0.05), whereas we found no significant exposure of IL-1β-511C/T with an association to lung cancer susceptibility.[44] Researchers found no significant differences of IL-1β in both lung cancer cases and controls (OR = 0.72, 95% CI: 0.41–1.28, P = 0.3) which was in accordance with our findings.[45]

We found a significant association with GSTT1 null genotype (OR = 2.6, 95% CI: 2.2–9.6, P < 0.001) and also predicted as best predicted single factor model by GMDR analysis which was similar to other study (OR = 1.8, 95% CI: 1.2–2.8, P = 0.002) lung cancer patients.[46] A study reported in Northeast India showed no significant association to GSTM1 (Ptrend = 0.13) and reduced risk to GSTT1 (OR = 0.3, 95% CI: 0.1–0.7, P = 0.005) in lung cancer patients. As per our study, we have reported a significant association with GSTT1 and no association to GSTM1.[47]

In recent study, it showed that protein expression of GSTT1 was twofold higher in lung cancer patients in the absence of GSTM1 genes comparatively to those patients having GSTM1 genes (P = 0.018). Similarly, in our study, we found twofold (OR = 2.6, 95% CI: 1.5–4.4, P < 0.001) increased risk toward lung cancer patients having deletion of GSTT1.[48] Researchers in meta-analysis found no effect of deletion of GSTM1 and GSTT1 genes as well as survival in lung cancer patients, whereas the present study found increased risk toward lung cancer.[49]

The limitation of our study is that few genes were taken for the study because lung cancer involved many genetic factors associated with various genes. Further studies with larger sample size provide a brief insight into these findings.

 > Conclusions Top

In conclusion, our results demonstrated a higher order of gene–gene interaction of phase II metabolizing enzymes, DNA repair genes, and cytokine gene. The loci investigated in the present study for XRCC1399Gln, XPD751Gln, IL-1β, and IL-1RN are located on same Chr, which provides detailed insight to risk associated with lung cancer. Till date, no research reported on cytokine gene polymorphism in North Indian population. Cytokine gene showed the protective effect as IL-1RN is an anti-inflammatory cytokine in lung cancer susceptibility.


This study was supported by a grant from the Department of Science and Technology and University Grants Commission (UGC). We also thank both the cancer patients and controls enrolled in the study. Ritambhara is thankful for her UGC fellowship.

Financial support and sponsorship

This study was supported by a grant from the Department of Science and Technology and University Grants Commission.

Conflicts of interest

There are no conflicts of interest.

 > References Top

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  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8]

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6], [Table 7], [Table 8]


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