## 4 Discussion
However, the results of the evaluation of the correlation between promoter methylation and gene/protein expression were not always in accordance. The MEXPRESS data showed a weak negative correlation between gene expression and the methylation of CpG dinucleotides in the region corresponding to the proximal promoter sequence (near 87,600 K; GenBank; area marked in red in Figures 11 and 12). Methylated samples from patients with bronchioloalveolar carcinoma showed no presence of P-gp [50]. On the contrary, a study by [PERSON] and colleagues reported that the expression of the \(ABCB1\) gene was enhanced at both the mRNA and protein levels in hypermethylated lung cancer tissue samples [37]. These contradictory results reflect the fact that promoter methylation is only one of several mechanisms affecting gene/protein expression; thus, the level of gene methylation is often not directly related to the amounts of transcripts or protein. Furthermore, thorough the analysis in the group of lung cancer patients, a slight change in promoter methylation in the course of both LUAD and LUSC was found when taking into account clinicopathological factors. The methylation patterns of the gene for P-gp seem to be reciprocal between LUSC and LUAD in a number of categories. In lung adenocarcinoma, the level of methylation increased with the age of the patient, while a higher stage of the disease was associated with a reduction in the promoter methylation in squamous-cell carcinomas. This may indicate a quite different significance of \(ABCB1\) promoter methylation and/or the strong influence of individual/environmental factors on the course of cancer in these two histological types of NSCLC. To the best of our knowledge, the presented results are not supported by data from other lung cancer studies. So far, findings have suggested that \(ABCB1\) promoter methylation cannot be considered a predictive or prognostic biomarker in lung cancer [48; 50]. Data from the TCGA database indicated that promoter methylation of this gene has the potential to become a predicator of patient prognosis, which is consistent with observations for breast carcinomas [53] and other types of solid tumors [52]. In lung squamous-cell carcinoma, the correlation of \(ABCB1\) promoter methylation with the histopathological stage that was observed in this analysis is a novelty of the present report. For LUAD, no similar association could be attributed to the cancer staging. The present study is not without limitations. The first is the simple model of statistical analysis. This model, which is available in the bioinformatics tools that were used does not include a multi-factor adjustment. Moreover, only bioinformatics techniques were used to analyze DNA methylation in the \(ABCB1\) promoter, which is the second limitation of our research. Nevertheless, the expression of this gene was evaluated through an in vitro study and was previously published [54]. In this preliminary study, there were no statistically significant differences between the level of \(ABCB1\) and the clinicopathological features of the disease, with the exception of histological malignancy. The associations may not be noticeable due to the small number of patients that were included in the experimental analysis. In summary, the physiologically low levels of \(ABCB1\) expression in lung tissue are significantly downregulated in cancer cells. Thus, a close link between the reduction in efflux pump function and the transformation of normal lung cells to malignant cells was confirmed. In addition, high \(ABCB1\) expression can also be used as an independent prognostic factor for lung cancer patients. Finally, it is equally important that the main methylation promoter is one of the causes of decreased gene expression in lung cancer cells. Taken together, these data establish the \(ABCB1\) transporter gene as an important prognostic factor that could alter disease progression and contribute to the survival of cancer patients. Conceptualization, A.M.J. and E.B. ; Methodology, A.M.J. and E.B.