## 4 Discussion  
This study assessed the prognostic values of _KRAS_ and _HRAS_ gene expression in NSCLC, compared to values in a public database, and evaluated the impact of different concentrations of deltarasin, FTS and Kobe065 (RAS-dependent signaling inhibitors) on the viability of the NSCLC A549 cell line, as well as its _KRAS_ and _HRAS_ genes expression levels. Initially, bioinformatic analysis was used to determine the differential expression patterns of _KRAS_ and _HRAS_ genes in all types of human cancers and normal tissues. Our findings provide the first indication of changes in the expression of _KRAS_ and _HRAS_ mRNA in NSCLC cells following stimulation by different RAS inhibitors. Reanalysis with the use of TNMplot's publicly available data found that _KRAS_ and _HRAS_ genes are widely overexpressed in different types of human cancers. Thus far, four main independent routes downstream of _KRAS_ have been described, viz. the PI3K/AKT, RAF/MEK/ERK/MAPK, RALGEF/TBK1 and RAF1-MAPK pathways; these influence tumorigenesis through the stimulation of cell proliferation, differentiation, survival and cell-cycle regulation. These pathways might be excessively activated in accordance with mutations in _KRAS_ and other RAS family members (i.e., _HRAS_, _NRAS_) as well as different expression of _KRAS_ isoforms. Although many studies have examined the role of RAS family genes in oncogenesis, there is still a need to recognize factors leading to the activation of KRAS-dependent pathways, particularly with regard to therapeutic applications based on direct or indirect inhibition of _KRAS_[26]. A comparison of _KRAS_ and _HRAS_ gene expression in normal and tumor tissues of NSCLC patients in the TNMplot database showed that the _KRAS_ gene was significantly overexpressed in LUAD and LUSC samples. _HRAS_ mRNA expression was significantly higher in LUSC than in normal tissue, but was downregulated in the LUAD samples. All of the tested _p_-values were \(<\)0.05. Moreover, analyses performed on the basis of the UALCAN database confirmed the above-mentioned results except for _HRAS_ mRNA expression in LUAD, where it was slightly higher than in normal non-cancerous tissues. The observed disparity between _HRAS_ mRNA expression in LUAD samples and in normal tissue could be due to the inequality in the amount of normal samples obtained from non-cancerous persons (n = 486 vs. n = 59, respectively). Both _KRAS_ and _HRAS_ are highly expressed in NSCLC tissues compared to the normal lung tissues at the mRNA level, which could suggest their possible function in NSCLC development and progression. There is some evidence that _KRAS_ mutations are involved in the forming of a tumor microenvironment (TME). Overstimulation of KRAS-dependent signaling might lead to increased secretion of interleukin-6 (IL-6), thus inducing inflammation and the initiation and progression of tumors [27]. In the next step, the relative expression levels of _KRAS_ and _HRAS_ genes in both tumor and normal samples in various NSCLC subgroups were related to clinicopathological features. Our findings indicate some correlation between _KRAS_ and _HRAS_ expression and cancer stage, smoking habits, age, and sex, as well as the nodal metastasis status of lung cancer patients. _KRAS_ and _HRAS_ overexpression was strongly associated with smoking status, which corroborates data from the literature that smoking is the major cause of lung cancer [28]. [PERSON] et al. showed that expression of K-RAS4B protein in adenocarcinoma subtype of NSCLC was associated with age; however, they did not report any associations between the mRNA expression of _KRAS_ isoforms and clinicopathological parameters [29]. Significant associations have been found between higher _HRAS_ mRNA expression in blood and smoking status. Interestingly, _HRAS_ overexpression was noticed in tumor tissues in LUAD, but in blood in LUSC. In addition, relative _KRAS_ expression level in tissue was only significantly higher in patients older than 67 years [30].