Cancer stem cells (CSCs), characterized by self-renewal and unlimited proliferation, lead to therapeutic resistance in lung cancer. In this study, we aimed to investigate the expressions of stem cell-related genes in lung adenocarcinoma (LUAD). The stemness index based on mRNA expression (mRNAsi) was utilized to analyze LUAD cases in the Cancer Genome Atlas (TCGA). First, mRNAsi was analyzed with differential expressions, survival analysis, clinical stages, and gender in LUADs. Then, the weighted gene co-expression network analysis was performed to discover modules of stemness and key genes. The interplay among the key genes was explored at the transcription and protein levels. The enrichment analysis was performed to annotate the function and pathways of the key genes. The expression levels of key genes were validated in a pan-cancer scale. The pathological stage associated gene expression level and survival probability were also validated. The Gene Expression Omnibus (GEO) database was additionally used for validation. The mRNAsi was significantly upregulated in cancer cases. In general, the mRNAsi score increases according to clinical stages and differs in gender significantly. Lower mRNAsi groups had a better overall survival in major LUADs, within five years. The distinguished modules and key genes were selected according to the correlations to the mRNAsi. Thirteen key genes (CCNB1, BUB1, BUB1B, CDC20, PLK1, TTK, CDC45, ESPL1, CCNA2, MCM6, ORC1, MCM2, and CHEK1) were enriched from the cell cycle Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, relating to cell proliferation Gene Ontology (GO) terms, as well. Eight of the thirteen genes have been reported to be associated with the CSC characteristics. However, all of them have been previously ignored in LUADs. Their expression increased according to the pathological stages of LUAD, and these genes were clearly upregulated in pan-cancers. In the GEO database, only the tumor necrosis factor receptor associated factor-interacting protein (TRAIP) from the blue module was matched with the stemness microarray data. These key genes were found to have strong correlations as a whole, and could be used as therapeutic targets in the treatment of LUAD, by inhibiting the stemness features.

S-palmitoylation, catalyzed by a family of 23 zinc finger Asp-His-His-Cys (DHHC) domain-containing (ZDHHC) protein acyltransferases localized on the cell membrane. However, stemness genes modulated by ZDHHCs in lung adenocarcinoma (LUAD) remain to be defined. Previously, we have constructed a network of cancer stem cell genes, including INCENP, based on mRNA stemness indices (mRNAsi) of LUAD. INCENP has the function of a chromosomal passenger complex locating to centromeres, which is performed by the conserved region of its N-terminal domain. INCENP protein with a deletion of the first non-conserved 26 amino acid sequence failed to target centromeres. However, the exact function of the deleted sequence has not been elucidated. To identify novel cancer stem cell-relevant palmitoylated proteins and responsible ZDHHC enzymes in LUAD, we analyzed multi-omics data obtained from the database of The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), Clinical Proteomic Tumor Analysis Consortium (CPTAC), and the Human Protein Atlas (HPA). ZDHHC5 is distinguished from the ZDHHC family for being up-regulated in mRNA and protein levels and associated with malignant prognosis. ZDHHC5 was positively associated with INCENP, and the correlation score increased with LUAD stages. CSS-Palm results showed Cys¹⁵ was the S-palmitoylation site of INCENP. Interestingly, Cys¹⁵ locates in the 1–26 aa sequence of INCENP, and is a conserved site across species. As INCENP is a nuclear protein, we predicted that the nuclear localization signal of ZDHHC5 was specific to the importin αβ pathway, and the result of immunofluorescence proves that ZDHHC5 is located in the nucleoplasm, in addition to the plasma membrane. Therefore, our study indicates the S-palmitoylation of INCENP mediated by ZDHHC5 as a potential mechanism of S-palmitoylation to modulate CSCs in LUAD.

Increasing evidence suggests that the glutathione peroxidase 2 may actually play important roles in tumorigenesis and progression in various human cancers such as colorectal carcinomas and lung adenocarcinomas. However, the role of glutathione peroxidase 2 in gastric carcinoma remains to be determined. In this study, the expression and prognostic significance of glutathione peroxidase 2 in gastric carcinoma were investigated and the well-known prognostic factor Ki-67 labeling index was also assessed as positive control. Glutathione peroxidase 2 expression levels in the tumor tissue specimens, the matched adjacent normal tissue specimens, and the lymph node metastases of 176 patients with gastric carcinoma were evaluated by quantitative polymerase chain reaction, western blotting, and immunohistochemical staining. The associations between glutathione peroxidase 2 expression levels, as determined by immunohistochemical staining, and multiple clinicopathological characteristics were determined by Pearson’s chi-square test and Spearman’s correlation analysis. The relationships between glutathione peroxidase 2 expression and other clinicopathological variables and patient prognoses were analyzed further by the Kaplan–Meier method, the log-rank test, and Cox multivariate regression. The quantitative polymerase chain reaction, western blotting, and immunohistochemical staining results showed that glutathione peroxidase 2 expression levels were upregulated in both the primary tumor foci and the lymph node metastases of patients with gastric carcinoma (all p values < 0.05). Furthermore, Pearson’s chi-square tests, as well as Spearman’s correlation analysis, revealed that glutathione peroxidase 2 expression levels were strongly correlated with the Ki-67 labeling index, differentiation, histological patterns, Lauren classifications, lymph node metastasis, vascular invasion, tumor–node–metastasis stages, Helicobacter pylori infection, and overall survival (all p values < 0.05). Kaplan–Meier analysis, as well as the log-rank test and multivariate Cox regression analysis, showed that multiple clinicopathological risk factors and glutathione peroxidase 2 expression were novel independent prognostic factors for gastric carcinoma (all p values < 0.05). Glutathione peroxidase 2 expression is a novel independent prognostic biomarker for gastric carcinoma that may be used to devise personalized therapeutic regimens and precision treatments for this disease.