Missense variants in CYP4B1 associated with increased risk of lung cancer among Chinese Han population

A systematic analysis of the Global Burden of Disease (GBD) study (2019) showed that the global burden of cancer is substantial and growing [1]. A study comparing the latest cancer profiles in China and the United States found that the most common cancer is lung cancer in China and breast cancer in the United States, with lung cancer being the leading cause of cancer deaths in both countries [2]. According to World Health Organization statistics, the lung cancer worldwide average annual mortality rate is very high, it has brought serious adverse to human health and the social economy [3]. Therefore, the urgency of lung cancer prevention and control should be widely recognized and paid great attention to. Understanding the etiology and risk factors of lung cancer is the key to developing scientific and effective prevention and control strategies for lung cancer.

Tobacco is considered to be one of the main reasons affecting the occurrence and development of lung cancer [4]. However, it is not the only reason. Many studies have found that the lung cancer risk of females among Chinese with low smoking rates is about the same as that of females with high smoking rates in Western European countries [5, 6]. Therefore, in addition to smoking, there are other factors that may affect the occurrence and development of lung cancer, such as individual genetic factors and lampblack [7‐9]. Researchers around the world have been studying the risk factors for lung cancer and ways to prevent it. Although there is no effective method to completely prevent lung cancer at present, it is recognized to reduce the risk of lung cancer by reducing exposure to risk factors and implementing precise and personalized prevention strategies. Therefore, it is particularly urgent to find new biomarkers and more accurate predictors of lung cancer to provide better diagnosis and prognosis for lung cancer patients [10].

SNP (single nucleotide polymorphism) is the most common single nucleotide variation among individuals and occurs in more than 1% of individual genomes in a population. Among the variations in the human genome, more than 90% belong to this kind of variation [11, 12]. As third-generation genetic markers, SNPs have the advantages of high density, representativeness, stability, etc. Therefore, they have been used in genetic analysis to locate quantitative trait loci (QTL) in lung cancer and other diseases, and a large number of related single nucleotide polymorphism loci have been identified [13, 14]. A large number of studies have reported that SNPs can change or regulate the function of genes, and play an important role in diseases including lung cancer. Such as, SNP rs35705950 is associated with familial interstitial pneumonia and idiopathic pulmonary fibrosis, and is involved in the pathogenesis of pulmonary fibrosis by regulating the expression of MUC5B in the lung [15]; SNP rs769236 may be involved in the occurrence of colon cancer, liver cancer and lung cancer by influencing the regulation of CCNA2 expression, and this SNP has been shown to be a valuable biomarker for evaluating individual patients' susceptibility to cancer [16]. SNP rs12587742 is involved in the occurrence of lung cancer by up-regulating the mRNA expression of DCAF4 and reducing its methylation status [17]. Although many genetic polymorphisms associated with lung cancer susceptibility have been identified, the molecular mechanisms involved in the development and development of lung cancer are still unclear. Screening SNPS related to the development of lung cancer in specific populations will lay a theoretical foundation for studying the molecular mechanism of lung cancer. And then further promote the individualized prevention and treatment of lung cancer.

Cytochrome P450 (CYP) family is a typical phase I drug metabolizing enzyme located in the inner mitochondrial membrane or the endoplasmic membrane of eukaryotic cells [18]. A variety of CYP proteins encoded by the human genome are responsible for the metabolism of many endogenous and exogenous compounds [19‐21]. Several previous studies have reported the potential of CYP polymorphisms in cancer treatment. CYPs gene polymorphisms such as CYP1A1, CYP1B1, CYP2E1, CYP2D6, and CYP3A4 have been reported may be play an important role in chemotherapy and survival in lung cancer patients [22]. In the CYP family, CYP4 enzymes are involved in the metabolism of fatty acids, which is associated with susceptibility to genetic diseases [18]. A recent study using the Cancer Genome Atlas (TCGA) project and gene Expression Synthesis (GEO) database showed that CYP4B1 is a potential therapeutic target for lung adenocarcinoma [23]. Fat is an important source of energy during tumorigenesis [24]. Studies have found that reprogramming of fatty acid metabolism plays an important role in the development of several cancers, including lung cancer [25]. Metabolic reprogramming is one of the hallmarks of tumor cells [26]. It is of great significance to understand the mechanism of metabolic reprogramming, which determines how we target metabolic reprogramming to treat cancer. The association between CYP4B1 genetic polymorphism and susceptibility to prostate cancer, bladder cancer or other cancers has been reported [18, 27, 28]. However, no studies have reported the association between CYP4B1 genetic polymorphism and lung cancer risk in the Chinese Han population. In conclusion, our study aimed to investigate the association between CYP4B1 single nucleotide polymorphism (SNP) and susceptibility to lung cancer. We also conducted stratified analysis according to the potential risk factors related to the development of lung cancer, such as age, sex, smoking/drinking, tumor staging, tumor metastasis, tumor type, and so on, so as the relationship between CYP4B1 SNP and these potential risk factors will be evaluated. This study will provide a data supplement for exploring novel biomarkers associated with susceptibility to lung cancer. Due to the potential role of CYP4B1 in fatty acid metabolism, the study will help to explore the molecular mechanism of the occurrence and development of lung cancer and then lay a theoretical foundation for targeted metabolic reprogramming to treat lung cancer.

There were 1339 participants in the case group (668) and the control group (671) enrolled in this study. The average ages of the case and control groups were 60.18 ± 9.91 years and 59.87 ± 9.30 years (Table 1). The number of males and females in the case group was 460 (68.9%) and 208 (31.1%), and in the control group were 457 (68.1%) and 214 (31.9%), respectively. The number of smoking and non-smoking participants in the case group was 351 (52.5%) and 317 (47.5%), and in the control group were 357 (53.2%) and 314 (46.8%), respectively. The number of drinking and non-drinking participants in the case group was 351 (52.5%) and 317 (47.5%), and in the control group were 361 (53.8%) and 310 (46.2%), respectively. The basic information about the participants can be found in Table 1.

We conducted a study on the association between three missense variants in CYP4B1 and LC susceptibility in 1339 participants. Combined with correlation analysis and FPRP results, three candidate missense variants in CYP4B1 (rs2297810, rs4646491, and rs2297809) were found to be associated with LC risk. The genotype GA of CYP4B1-rs2297810 was significantly associated with an increased risk of lung cancer in both overall and stratified analyses. Similarly, genotype CT of CYP4B1-rs4646491 and genotype CT of CYP4B1-rs2297809 are also associated with an increased risk of lung cancer. These indicate that CYP4B1-rsS2297810, -rs4646491, and -rs2297809 are potential genetic risk factors for lung cancer. To our knowledge, this study is the first to report that CYP4B1 genetic polymorphisms are associated with lung cancer susceptibility in the Chinese Han population.

The analysis showed that the presence of these three missense variant heterozygous genotypes will increase the risk of lung cancer whether participants were younger or older than 60 years. Genotype GA of CYP4B1-rs2297810 is a risk factor for both female and male lung cancer. CYP4B1-rs2297809 is associated with an increased risk of female lung cancer. Although no significant results of CYP4B1-rs2297809 associated with lung cancer were found in the male population, the overall trend was that the presence of the CT genotype of CYP4B1-rs2297809 will also increase the risk of male lung cancer (OR > 1). We also found that the three missense variants in CYP4B1 were significantly associated with an increased risk of lung cancer in non-smokers, while no positive results were found in smokers. Age, smoking, and gender differences have previously been reported as risk factors for lung cancer [34‐36]. Czerwinski M, et al have found that CYP4B1 is not induced by compounds present in cigarette smoke in lung cancer patients [37]. Combined with previous studies and the results of our study, we speculated that the three missense variants in CYP4B1 are risk factors for lung cancer in the Chinese Han population, and the above genetic risk factors related to CYP4B1 may not be affected by these potential risk factors.

In recent years, research on CYP4B1 in cancer has attracted special attention, which may be due to its different expression in patients with various cancers, including lung cancer, compared with normal individuals [23, 37, 38]. CYP4B1 was significantly downregulated in lung cancer patients, which was further confirmed by UALCAN online database analysis in this study. In addition, through the database, we also found that low expression of CYP4B1 was significantly associated with the prognosis of LUAD patients. Thus, the expression level of CYP4B1 is closely related to lung cancer, but its regulation mechanism in lung cancer is still not clear. Searching in the genotype-tissue expression database showed that CYP4B1 expression levels were different in lung tissues under different genotypes of these missense variants. And we found that these variants in CYP4B1 can cause changes in amino acid sequence when we used the dbSNP database to search the information related to candidate genetic variants. Changes in amino acid sequence can cause changes in protein structure, which is directly related to its function [39]. In addition, CYP4 is involved in fatty acid metabolism, and the reprogramming of fatty acid metabolism is very important for the occurrence of lung cancer [25]. Combined with previous studies and the results of our study, we speculate that these missense variants may cause changes in the amino acid sequence, which may cause changes in the protein structure of CYP4B1, thereby affecting the gene expression level, thereby affecting the fatty acid metabolism process, and ultimately affecting the LC risk. However, the above is only speculation, and further mechanism research is necessary to explore how these three candidate missense variants affect the susceptibility to LC by affecting the expression of CYP4B1 among the Chinese Han population. In any case, this study has laid a reliable theoretical foundation for the mechanism of CYP4B1 in the development of lung cancer. At the same time, it has provided new ideas for risk assessment and clinical individualized prevention and treatment of lung cancer among Chinese Han.

However, we must face the fact that this study has certain limitations. First, it is necessary to conduct a large sample size or confirmatory study in people with different genetic backgrounds, which will help ensure the reproducibility and reliability of the results of this study. Secondly, further design of functional validation tests will help to accurately understand the mechanism of three CYP4B1 gene polymorphisms in the occurrence and development of lung cancer. The above studies will further help us understand the potential molecular mechanism of the three CYP4B1 genetic loci in LC risk, which in turn will help to further understand the pathogenesis of LC. We believe this will be a very interesting research direction. In any case, this study is the first to explore the association of CYP4B1 gene polymorphism with lung cancer susceptibility in the Chinese Han population and has achieved positive results.