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Biological characterization and clinical significance of cuproptosis-related genes in lung adenocarcinoma

BMC Pulmonary Medicine volume 25, Article number: 13 (2025) Cite this article

Abstract

Background

Lung cancer has high morbidity and mortality rates, which results in a poor prognosis. Cuproptosis is a novel cell death mechanism. The aim of this study was to examine the biological characteristics and clinical significance of genes associated with cuproptosis in lung adenocarcinoma (LUAD), and to understand the molecular mechanisms underlying the occurrence and progression of LUAD.

Methods

We targeted 10 cuproptosis-related genes from previous studies and used the datasets from GEO and TCGA databases to identify differential genes related to cuproptosis; then the data were analyzed by R package, Cytoscape, TISDB, cBioPortal, STRING, CancerSEA, and Disgenet; and finally, the data were detected by immunohistochemistry validation was performed.

Results

CDKN2A and MTF1 were cuproptosis-associated LUAD differential genes and were differentially expressed in immune subtypes. The expression of CDKN2A and MTF1 showed correlation with multiple functional states of LUAD.CDKN2A was negatively correlated with LUAD survival prognosis.

Conclusion

CDKN2A and MTF1 were correlated with the diagnosis of LUAD, and CDKN2A was negatively correlated with the survival and prognosis of LUAD. CDKN2A has the potential to contribute to the early diagnosis and prognosis analysis of LUAD.

Peer Review reports

Background

Lung adenocarcinoma (LUAD) is the most common subtype of lung cancer [1], which has the highest global incidence and mortality rates, posing a significant threat to human health. Due to the lack of typical clinical manifestations in the early stages of LUAD, many patients are already metastatic at the time of diagnosis [2, 3], missing the opportunity for surgery. In recent years, with the continuous development of medical care, molecular targeted therapy and immunotherapy have become routine treatments for LUAD, improving its prognosis. However, the overall 5-year survival rate remains low. Therefore, it is of great significance to search for new and effective diagnostic and prognostic biomarkers.

Recently, Tsvetkov and colleagues [4] revealed for the first time a novel mechanism of cell death—cuproptosis—following pyroptosis and ferroptosis. Cuproptosis is induced by copper ions targeting lipidated tricarboxylic acid (TCA) cycle proteins [5, 6]. Serum copper levels are higher in lung cancer patients than in healthy individuals [7]. Interestingly, studies have found that the expression of cuproptosis-related genes is dysregulated in various tumors, including lung cancer, and is associated with prognosis [8]. Moreover, copper-based complexes can be used as antitumor drugs and have the potential to be at least one component of multifunctional combinations in cancer treatment [9].However, current research on the correlation between cuproptosis-related genes and LUAD is limited and mostly confined to bioinformatics analysis; the diagnostic and prognostic value of cuproptosis-related genes in LUAD remains unclear.

This study first screened two copper death-related differentially expressed genes, CDKN2A and MTF1, based on the GEO database. Subsequently, data mining analysis was conducted across multiple bioinformatics databases.Finally, immunohistochemical detection was used to explore the biological characteristics and clinical significance of CDKN2A and MTF1 in LUAD, aiming to identify new diagnostic and prognostic markers for LUAD. The workflow of this study is shown in Fig. 1.

Fig. 1
figure 1

Workflow of this study

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Methods

Data collection

We gathered a set of 10 genes associated with cuproptosis from previous scholars, CDKN2A,DLD, DLAT, FDX1,GLS, LIAS, LIPT1, MTF1, PDHA1 and PDHB [10]. From the Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/GEO/), we acquired the datasets GSE140797 and GSE10072. Additionally, the LUAD dataset, comprising RNA sequencing data, raw counts, clinical information, and a total of 539 tumor tissue cases, was obtained from the Cancer Genome Atlas (TCGA) (https://portal.gdc.Cancer.gov/).

Differential analysis

We used R 4.2.1 for data analysis.From the GEO database, we utilized the GEOquery package to download the data. Probes associated with multiple molecules were eliminated, and in cases where multiple probes corresponded to the same molecule, only the one with the highest signal value was retained.Normalization was performed using the Variance Stabilizing Transformations (VST) method in the "DESeq2" package [11], and differential analysis was conducted using the same package. RNAseq data of 33 tumor items STAR process were downloaded and organized from TCGA database (https://portal.gdc.cancer.gov) in TPM format, and differential analysis was performed by package “stats[4.2.1] [12], car[3.1–0]”.Finally, box plots, PCA plots, and volcano plots were constructed using the "ggplot2 [3.3.6]" package [13].

Receiver Operator Characteristic (ROC) Curve and Survival Analysis

We conducted ROC analysis and ROC test on CDKN2A and MTF1 in lung adenocarcinoma samples using the "pROC [1.18.0]" package [14], calculating the Area under Curve (AUC). The closer the AUC is to 1, the better the diagnosis. Survival analysis of copper death-related differential genes was performed using the "survival [3.3.1]" package [14], and visualization was done through the "survminer" package.

Baseline Data Table.

We used the "stats [4.2.1]" package to create baseline information for high and low expression groups of CDKN2A and MTF1, based on clinical information from the TCGA database and collected samples.

Functional Enrichment Analysis and Gene Set Enrichment Analysis

Selected patients were divided into two groups based on CDKN2A expression, and target genes were extracted by utilizing package “DESeq2” difference analysis with |log2(FC)|> 0.5 and adjP value < 0.05 as screening criteria. (GO) including biological process (BP), cellular component (CC), molecular function (MF) and (KEGG) pathway enrichment analyses were performed using package “cluster Profiler [4.4.4]” [15],p < 0.05 was considered statistically significant.Gene Set Enrichment Analysis (GSEA) was conducted using the "clusterProfiler" package, and the top 4 enriched terms were displayed in a mountain plot. The GSEA results were visualized using the "ggplot2" package.

Construction of mRNA-miRNA regulatory network

We utilized the miRNet database (https://www.mirnet.ca/) to predict the association between differentially expressed mRNAs and miRNAs. Subsequently, an mRNA-miRNA regulatory network was constructed based on the obtained results.

Expression levels of CDKN2A and MTF1 in different immune subtypes of LUAD

We utilized the TISDB database to investigate the expression profiles of CDKN2A and MTF1 across various subtypes of LUAD, including C1 (wound healing), C2 ((IFN-γ dominant), C3 (inflammatory), C4 (lymphocyte deplete), C5 (immunologically quiet), and C6 (TGF-β dominant) [16]. Information on genetic variants of CDKN2A, MTF1 was retrieved through cBioPortal (https://www.cbioportal.org/). We explored the CancerSEA database to examine the average correlation between CDKN2A, MTF1, and the functional status across 18 different types of cancers.The potential respiratory diseases related to CDKN2A and MTF1 was analyzed through the Disgene website.

Protein–Protein Interaction (PPI) Network Analyses of CDKN2A, MTF1

Potential protein interactions with CDKN2A, MTF1 were collected and integrated through the STRING database (https://string-db.org/) and extracted the relevant genes from these interactions to conduct PPI network analysis.

Immunohistochemistry

Clinical data collection

Collect paraffin-embedded tissue samples (both adjacent normal and cancerous tissues) from patients diagnosed with LUAD via pathology, who underwent surgical treatment at Xiangtan Central Hospital (affiliated with Xiangtan Medicine & Health Vocational College) between June 2019 and August 2021. A total of 93 cases were included, comprising 36 males and 57 females. Inclusion criteria: (1) All patients were histopathologically diagnosed with LUAD; (2) None of the patients had received any antitumor treatment before surgery; (3) Ages ranged from 18 to 70 years; (4) At least one measurable lesion confirmed by imaging; (5) No significant abnormalities in routine blood tests, liver and kidney function, or electrocardiogram. If deceased, the cause of death was closely related to LUAD for all patients. Informed consent was obtained from all patients, and the study was approved by the Medical Ethics Committee.

Exclusion criteria: (1) Age > 70 or < 18 years; (2) Patients with severe cardiovascular and cerebrovascular diseases, organ failure; (3) Patients with other organ malignancies; (4) Patients who received radiotherapy, chemotherapy, or other tumor-specific treatments before diagnosis; (5) Patients with poor compliance unable to participate in follow-up.

Immunohistochemical testing

The specimens were fixed in formalin, dehydrated, embedded, and sectioned into 4-μm continuous slices. Immunohistochemical analysis was performed using the MaxVision method. The CDKN2A antibody was purchased from ABclonal; the MTF1 antibody was purchased from ProteinTech; The universal immunohistochemistry MaxVision kit and DAB staining solution were purchased from Celnovte. The working concentration of the CDKN2A antibody was 1:100, and the working concentration of the MTF1 antibody was 1:200. The specific staining steps were strictly followed according to the instructions provided with the kit.

Interpretation of immunohistochemistry results

The Immuno-Reactive Score (IRS) was used to evaluate the expression scores of CDKN2A and MTF1. IRS is calculated by multiplying the cell staining intensity score with the percentage of positive cells. The cell staining intensity is scored on a 4-point scale: negative = 0, weak positive = 1, positive = 2, strong positive = 3; the percentage of positive cells is also scored on a 4-point scale: ≤ 25% = 1, 26%−50% = 2, 51%−75% = 3, > 75% = 4 [17]. An IRS < 6 is considered low expression, while an IRS ≥ 6 is considered high expression [18].

Results

Differentially expressed genes of cuproptosis-related genes in normal tissues and LUAD based on GEO database analysis

In this study, we first identified the LUAD gene set GSE140797 and normalized the expression matrix of this gene set, as shown in Fig. 2A. Subsequently, we performed Principal Component Analysis (PCA), as shown in Fig. 2B. We found that there were significant differences among the various sample groups in GSE140797. Based on the screening criteria of |log2(FC)|> 0.5 and P-value < 0.05, 6,519 differentially expressed genes (3,223 upregulated and 3,296 downregulated) were identified and represented as a volcano plot (Fig. 2C).In this study, 10 cuproptosis-related genes were collected from previous investigators, namely CDKN2A,DLD, DLAT, FDX1,GLS, LIAS, LIPT1, MTF1, PDHA1 and PDHB.Next, we intersected the differentially expressed genes of GSE140797 with cuproptosis-related genes, resulting in 5 common genes: MTF1, CDKN2A, DLD, FDX1, and PDHB, which were presented as a Venn diagram (Fig. 2D). Among them, MTF1, DLD, FDX1, and PDHB were down-regulated genes, while CDKN2A was an up-regulated gene. Subsequently, we introduced GSE10072 for validation, ultimately identifying the down-regulated gene MTF1 and the up-regulated gene CDKN2A (Fig. 2E).

Fig. 2
figure 2

Identification and validation of cuproptosis-related differential genes: A Normalized expression matrices of the GSE14079 datasets. B PCA diagrams of the GSE14079 datasets. C Volcano plots of the GSE14079 datasets, including 6,519 differentially expressed genes (3,223 up-regulated, 3,296 down-regulated). D Comparison of expression levels of MTF1, CDKN2A, DLD, FDX1, and PDHB in normal individuals and lung adenocarcinoma patients in the GSE10072 dataset

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Prediction of target miRNAs of CDKN2A, MTF1 based on miRNet tool

The study predicted he target miRNAs of CDKN2A and MTF1 by miRNet tool, and the final result (Fig. 3A), there are 150 target miRNAs, and 158 pairs of mRNA-miRNAs.Among them, CDKN2A is regulated by 31 miRNAs, and MTF1 is regulated by 127 miRNAs. Among them, hsa-mir-191-5p, hsa-let-7b-5p, hsa-mir-155-5p, hsa-mir-16-5p, hsa-mir-125b-5p, hsa-mir-522-5p, hsa-mir-423-3p, and hsa-mir-24-3p co-regulated CDKN2A, MTF1. This study further explored the diagnostic value of these 8 shared miRNAs in LUAD patients, such as Fig. 3B showed that hsa-mir-191-5p, hsa-mir-423-3p had the greatest diagnostic value. The study further browsed the PubMed database (https://PubMed.ncbi.nlm.nih.gov/), a total of 7 miRNAs related to LUAD were found:hsa-mir-191-5p [19], hsa-let-7b-5p [20],hsa-mir-155-5p [21],hsa-mir-16-5p [22],hsa-mir-125b-5p [23], hsa-mir-423-3p [24], hsa-mir-24-3p [25].

Fig. 3
figure 3

Prediction of common miRNAs related to cuproptosis-related differential genes and their diagnostic value: A Prediction of 8 miRNAs through the mRNA-miRNA regulatory network. B ROC analysis of 7 miRNAs obtained from PubMed search, all of which have certain diagnostic value

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Analysis of CDKN2A, MTM1 and LUAD based on TCGA

The analysis by TCGA combined with GTEx yielded that CDKN2A was expressed at the highest level in CESC, OV, and UCS, while MTF1 was expressed at the lowest level in LIHC and UVM (Fig. 4A, B). Compared with normal tissues, the expression level of CDKN2A was significantly up-regulated in the tissues of LUAD samples (P < 0.05), while the expression level of MTF1 was slightly down-regulated, with a statistically significant difference, as in (Fig. 4C). In paired samples, MTF1 was down-regulated and CDKN2A was up-regulated in LUAD patients (Fig. 4D, E). ROC curves were constructed using the LUAD dataset in the TCGA database (Fig. 4F), in which CDKN2A had high diagnostic value and MTF1 had some diagnostic value. In addition, MTF1 levels were down-regulated and CDKN2A levels were up-regulated in LUAD patients who smoked compared to LUAD patients who did not smoke (Fig. 4G).

Fig. 4
figure 4

Pan-cancer and LUAD expression analysis: A-B Comparison of CDKN2A and MTF1 expression in 33 cancer types and normal tissues using unpaired sample analysis. C Comparison of CDKN2A and MTF1 expression in LUAD and normal tissues using unpaired sample analysis, where the expression level of MTF1 is down-regulated, and the expression level of CDKN2A is up-regulated. D In the paired sample analysis, the expression level of CDKN2A is up-regulated in LUAD compared to normal tissues. E In the paired sample analysis, the expression level of MTF1 is down-regulated in LUAD compared to normal tissues. F In ROC curve, CDKN2A has a higher diagnostic value, while MTF1 has a lower diagnostic value. G Expression levels of CDKN2A and MTF1 between normal subjects, nonsmoking lung adenocarcinoma patients, and smoking lung adenocarcinoma patients

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CDKN2A expression in LUAD tissues was correlated with patients' overall survival (OS), disease-specific survival (DSS), and progression-free interval (PFI) (P = 0.007, P = 0.008, and P = 0.028, respectively); OS, DSS, and PFI were all better in low CDKN2A-expressing than in high CDKN2A-expressing patients, as shown in Fig. 5A, B, and C. While MTF1 protein there was no significant correlation between expression and prognosis (Fig. 5D). Moreover, we constructed baseline data tables for the high and low expression groups of CDKN2A and MTF1 using the TCGA database(Table 1, 2).Our findings indicated no significant associations between the expression levels of CDKN2A and MTF1 and the stages of T, N, and M.

Fig. 5
figure 5

Prognostic analysis: A-C The OS, DSS, and PFI of LUAD patients in the high CDKN2A expression group are shorter compared to those in the low expression group, with P < 0.05. D The expression level of MTF1 is not significantly associated with prognosis, P > 0.05

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Table 1 Table of baseline information of MTF1 high and low expression groups constructed based on TCGA database
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Table 2 Table of baseline information of CDKN2A high and low expression groups constructed based on TCGA database
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Expression of CDKN2A and MTF1 in different immune subtypes of LUAD

In different immune subtypes of LUAD, the expression levels of CDKN2A and MTF1 were significantly different (Fig. 6A, B).Among them, CDKN2A has the lowest expression levels in C3 and C6, while MTF1 has the highest expression in C3.

Fig. 6
figure 6

A CDKN2A expression correlates with immune subtypes in LUAD; B MTF1 expression correlates with immune subtypes in LUAD

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Genetic alterations of CDKN2A, MTF1

CDKN2A and MTF1 gene mutations in cancer were analyzed through the cBioPortal website, which included 32 studies and 10,967 samples (Fig. 7A, B). CDKN2A had a total of 57 mutation sites, including 49 missense mutations, 8 In-frame mutations, of which R80Q was the most common mutation site, with a mutation type of MTF1 has a total of 143 mutation sites, including 120 missense mutations, 16 truncating, 2 splices, 5 SV/fusion, of which R251Q is the most common mutation site, the mutation type is missense. The mutation rate of CDKN2A and MTF1 in LUAD lung tissues is in the 32 cancer tissues ranked high (Fig. 7C, D). In contrast, the counts of CDKN2A and MTF1 mutations in LUAD were lower than those in Lung Squamous Cell Carcinoma(LSCC) (Fig. 7E,F).

Fig. 7
figure 7

Gene mutation analysis of CDKN2A and MTF1: A Mutation diagram of CDKN2A across protein domains, where R80Q is the most common mutation site. B Mutation diagram of MTF1 across protein domains, where R251Q is the most common mutation site. C CDKN2A mutation map based on TCGA PanCancer Atlas Studies; D MTF1 mutation map based on TCGA PanCancer Atlas Studies. E The mutation count of CDKN2A in LSCC is higher than that in LUAD. F The mutation count of MTF1 in LSCC is higher than that in LUAD

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PPI network and enrichment analysis

The protein–protein interaction network of CDKN2A and MTF1 was searched and constructed with a high confidence level (0.7) as the screening condition and the threshold value of top 50 (Fig. 8A and B). The CDKN2A, MTF1 interacting proteins were combined and analyzed by GO/KEGG enrichment, and the results were as shown in Fig. 8C renal filtration cell differentiation (BP), glomerular visceral epithelial cell differentiation (BP), glomerular epithelial cell differentiation (BP), mitochondrial inner membrane (CC), and heat shock protein binding (MF). KEGG enrichment pathway was mainly Prion disease. The GSEA enrichment results for CDKN2A are shown in Fig. 8D. The common results were REACTOME_SYNTHESIS_OF_DNA,CELL_CYCLE,REACTOME_MITOTIC_G1_PHASE_AND_G1_S_TRANSITION,RETINOBLASTOMA_GENE_IN_CANCER.The results of GSEA enrichment for MTF1 are shown in Fig. 8E,with common results for RIBOSOME,SRP_DEPENDENT_COTRANSLATIONAL_PROTEIN_.

Fig. 8
figure 8

A The PPI network of CDKN2A; B The PPI network of MTF1; C GO/KEGG pathway enrichment of CDKN2A and MTF1 interacting proteins.D GSEA functional enrichment pathways of CDKN2A, and the top 5 pathways were selected.E GSEA functional enrichment pathways of MTF1, and the top 5 pathways were selected

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TARGETING_TO_MEMBRANE,RESPONSE_OF_EIF2AK4_GCN2_TO_AMINO_ACID_DEFICIENCY,EUKARYOTIC_TRANSLATION_ELONGATION,CYTOPLASMIC_RIBOSOMAL_PROTEINSCYTOPLASMIC_RIBOSOMAL_PROTEINS.

Functional status of CDKN2A, MTF1 in LUAD

The functional states of CDKN2A, MTF1 in various cancer types were explored through CancerSEA website (Fig. 9A, B), especially the correlation with various functional states in LUAD. Among them, CDKN2A was weakly positively correlated with Apoptosis (Fig. 9C), while MTF1 was positively correlated with DNA damage, DNA repair Invasion, and CellCycle (Fig. 9D).

Fig. 9
figure 9

The correlation between CDKN2A, MTF1 and tumor functional morphology: A The interactive bubble chart present correlation of CDKN2A with functional state in 16 cancers. B The interactive bubble chart present correlation of MTF1 with functional state in 16 cancers. C The correlation of CDKN2A with functional state in LUAD.D The correlation of MTF1 with functional state in LUAD

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Potential association of CDKN2A, MTF1 with other respiratory diseases

CDKN2A was analyzed through the Disgenet website and found to be associated with 49 respiratory diseases such as Neurogenic tumour, Dysplasia of larynx, and others (Fig. 10A), whereas MTF1 was only associated with Malignant neoplasm of lung (Fig. 10B).

Fig. 10
figure 10

Potential association of CDKN2A, MTF1 with other respiratory diseases: A CDKN2A-associated respiratory disease(including 49 diseases). B MTF1-associated respiratory disease (including 1 disease)

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CDKN2A, MTF1 expression in tissues of LUAD patients

In this study, a total of 93 cases of lung adenocarcinoma were examined. Immunohistochemistry was used to detect the expression of CDKN2A and MTF1 proteins in adenocarcinoma and adjacent tissues. The results suggest that the expression levels of CDKN2A and MTF1 in lung adenocarcinoma tissues are significantly higher than those in adjacent tissues (Fig. 11A-F). CDKN2A is not significantly expressed in the nucleus or cell membrane but is primarily expressed in the cytoplasm, with no significant expression in stromal cells; MTF1 shows no significant expression in the nucleus, weak expression in the cell membrane, and is mainly expressed in the cytoplasm, with no significant expression in stromal cells (Fig. 11G-H).

Fig. 11
figure 11

CDKN2A and MTF1 immunohistochemistry results: A High expression of CDKN2A in LUAD cancer tissues (× 200). B Low expression of CDKN2A in paracancerous tissues (× 200). C High expression of MTF1 in LUAD cancer tissues (× 200). D Low expression of MTF1 in paracancerous tissues (× 200). E High expression of CDKN2A in LUAD cancer tissues (left) and low expression in adjacent paracancerous tissues (right) (× 200). F High expression of MTF1 in LUAD cancer tissues (left) and low expression in adjacent paracancerous tissues (right) (× 200). G Negative expression of CDKN2A in normal tissues adjacent to LUAD cancer (× 200). H Negative expression of MTF1 in normal tissues adjacent to LUAD cancer (× 200)

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Relative quantification of immunohistochemistry results shows that compared to normal tissues, the expression levels of CDKN2A and MTF1 in LUAD tissues are significantly upregulated(Fig. 12A).

Fig. 12
figure 12

A The expression levels of CDKN2A and MTF1 in LUAD tissues are significantly up-regulated

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Discussions

Lung cancer is the most prevalent and lethal type of cancer worldwide, with lung adenocarcinoma being the predominant histological subtype among lung cancer patients [1, 2].Cuproptosis, identified as a novel form of cell death, exhibits close associations with tumor progression, growth, metastasis, and patient prognosis [26].In this study,CDKN2A and MTF1 differential genes associated with cuproptosis in lung adenocarcinoma were identified through bioinformation analysis, and immunohistochemical verification was performed through collected clinical specimens.

This study screened two copper death-related differential genes, CDKN2A and MTF1, through GSE140797. Subsequently, a miRNA interaction network relationship between CDKN2A and MTF1 was constructed, ultimately identifying eight common targeted miRNAs. Further searching the PubMed database for literature related to miRNA and LUAD, and excluding non-human sample studies, it was found that among the eight targeted miRNAs, seven met the relevant criteria. Among them, hsa-mir-191-5p and hsa-mir-423-3p had the highest diagnostic value for LUAD. Previous literature has shown that hsa-mir-191-5p is significantly dysregulated in the serum and lung tissues of patients with LUAD [19], and hsa-mir-423-3p drives the EMT process and tumor growth of LUAD by targeting CYBRD1 and activating the FAK signaling pathway [25], which may be related to the molecular mechanisms of lung adenocarcinoma.

Both MTF1 and CDKN2A are negative regulators among copper death-related genes [27]. CDKN2A is a reverse regulator of the cell cycle G1/S checkpoint. The protein transcribed by CDKN2A can stably bind to E3 ubiquitin ligase and effectively inhibit the expression of P53 in tumor cells [28]. CDKN2A is highly expressed in most tumor cells [27] and is associated with poorer prognosis. Research by WU Changwu et al.[8]found that the expression of CRGs is dysregulated in various tumors, including lung cancer, and is related to prognosis. In this study, bioinformatics analysis showed that compared to normal tissues, the gene expression level of CDKN2A was significantly up-regulated in LUAD samples. Patients with low CDKN2A expression had better OS, DSS, and PFI than those with high expression. Immunohistochemical detection showed that CDKN2A protein expression level was significantly higher in LUAD cancer tissues than in paracancerous tissues, and there was no significant correlation between CDKN2A expression level and T, N, and M stages, which verified the results of bioinformatics analysis.This study suggests that CDKN2A has high diagnostic value for LUAD, but its prognostic value in LUAD needs further clinical trial validation.

MTF1 may influence cellular sensitivity to copper death by affecting the intracellular levels of copper-binding substances such as glutathione and metallothionein [29]. In this study, bioinformatics analysis showed that compared to normal tissues, the gene expression level of MTF1 was slightly downregulated in LUAD samples, and constructing ROC curves suggested that MTF1 has certain diagnostic value for LUAD. Immunohistochemistry detection showed that the protein expression level of MTF1 in LUAD cancer tissues was significantly higher than that in adjacent non-cancerous tissues. The trend between mRNA levels and protein levels was not consistent. After excluding experimental operation reasons, it is considered that one or more mechanisms may be at play, such as potential post-transcriptional regulation mediated by m6A, post-transcriptional regulation related to alternative splicing, and post-translational modification-related regulation, which may affect the genetic variation buffering the protein level at the mRNA level. Yang Y's research showed that after sulfate treatment, the abundance of m6A in MTF1 mRNA was significantly reduced [30]. In experiments by Guoan Chen et al., it was found that only a portion of proteins were significantly correlated with messenger ribonucleic acid abundance in the same LUAD, where the expression of individual isoforms of a single protein might be affected by other post-translational mechanisms, thereby altering the isomer abundance in tissue and cancer [31]. Therefore, the mechanism behind the inconsistency between mRNA and protein levels of MTF1 remains to be further studied.

Some studies have shown that the frequency of CDKN2A deletion mutations significantly increases in LUAD patients with brain metastasis [32], and may also be involved in the construction of different immune phenotypes of LUAD [33]. The mutation of CDKN2A will significantly promote its RNA expression. In this study, the mutation rates of CDKN2A and MTF1 in LUAD tissues were ranked eleventh and ninth among 32 cancers, respectively, suggesting that mutations in MTF1 and CDKN2A may play a certain role in the development and metastasis of LUAD. This study showed that the mutation counts of CDKN2A and MTF1 in LUAD are lower than those in lung squamous cell carcinoma, consistent with the results of Yanhong Shang et al. [34], suggesting that these two genes may have widespread effects in both lung adenocarcinoma and lung squamous cell carcinoma.Ferroptosis, a form of iron-dependent programmed cell death,is marked by the accumulation of lipid peroxides [35].In previous studies, concurrent mutations in STK11 and KEAP1 were found to induce the expression of ferroptosis-protective genes. This leads to an increase in the levels of protective gene expression, thereby allowing tumor cells to evade the ferroptosis apoptotic mechanism. This could potentially promote the proliferation of tumor cells in vitro and their growth in vivo [36].Additionally, the high expression of SLC7A11 inhibits the level of ferroptosis in human LUAD cell lines A549, thereby promoting the proliferation and accelerated migration of tumor cells [37].Therefore, both copper death and ferroptosis play a certain role in the growth and migration of lung adenocarcinoma.

This study still has some limitations. Firstly, as a novel mode of cell death, the genes involved in copper death and their mechanisms of action in lung tissue need further exploration, and the intervention strategy for LUAD targeting CDKN2A and MTF1 as well as the possible combined treatment options will be the focus of the next study. Secondly, the follow-up time for clinical samples in this study is relatively short, and further follow-up is needed to investigate the relationship between CDKN2A, MTF1 and the prognosis of LUAD patients.

In summary, this study explored the correlation between CDKN2A and MTF1 genes and lung adenocarcinoma, which may contribute to the early diagnosis and prognostic analysis of LUAD.

Data availability

Publicly available datasets were analyzed in this study. This data can be found as follow: https://www.ncbi.nlm.nih.gov/geo/,with the accession number GSE140797. The other data used during the current study would be available from the corresponding authors on reasonable request.

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Acknowledgements

Not applicable.

Funding

This work was supported by the National Natural Science Foundation of China [Grant numbers 81870033]; the Six Talent Peaks Project of Jiangsu Province [Grant number WSN-106]; the Medical Scientific Research Foundation of Jiangsu Province of China [Grant numbers QNRC2016340]; and the Foundation for High-level Talents during the 13th Five-year Plan Period of Yangzhou, China [Grant numbers ZDRC201866].

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Author notes

  1. Meilin Li and Yu Tan contributed equally to this work.

Authors and Affiliations

  1. Xiangtan Medicine & Health Vocational College, Xiangtan, China

    Meilin Li

  2. Northern Jiangsu People’s Hospital Affiliated to Yangzhou University, Yangzhou, China

    Yu Tan & Lingfeng Min

  3. Department of Pathology, Xiangtan Central Hospital, Xiangtan, China

    Zhixin Li

Authors
  1. Meilin Li
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  2. Yu Tan
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  3. Zhixin Li
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  4. Lingfeng Min
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Contributions

M.L.L, Y.T and L.F.M designed this study.Y.T contributed to the bioinformatics analysis. M.L.L and Z.X. L collected clinical data, pathological sections and performed immunohistochemistry. M.L.L, Y.T and L.F.M drafted and revised the manuscript. All authors finalized the manuscript.

Corresponding author

Correspondence to Lingfeng Min.

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Ethics approval and consent to participate

The study was approved by the Medical Ethics Committee of the Northern Jiangsu People's Hospital Affiliated to Yangzhou University(No. 2021ky053). All participants were informed and signed a consent form.

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The authors declare no competing interests.

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Li, M., Tan, Y., Li, Z. et al. Biological characterization and clinical significance of cuproptosis-related genes in lung adenocarcinoma. BMC Pulm Med 25, 13 (2025). https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12890-025-03477-4

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  • DOI: https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12890-025-03477-4

Keywords

BMC Pulmonary Medicine

ISSN: 1471-2466

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