Identification and Validation of a Three Pyroptosis-Related lncRNA Signature for Prognosis Prediction in Lung Adenocarcinoma

Abstract

Pyroptosis, defined as programmed cell death, results in the release of inflammatory mediators. Recent studies have revealed that pyroptosis plays essential roles in antitumor immunity and immunotherapy efficacy. Long noncoding RNAs (lncRNAs) are involved in a variety of biological behaviors in tumor cells, although the roles and mechanisms of lncRNAs in pyroptosis are rarely studied. Our study aimed to establish a novel pyroptosis-related lncRNA signature as a forecasting tool for predicting prognosis and ascertaining immune value. Based on lung adenocarcinoma (LUAD) patients from The Cancer Genome Atlas (TCGA), we performed Pearson's correlation analysis to identify pyroptosis-related lncRNAs. After differentially expressed gene analysis and univariate Cox regression analysis, we selected prognosis-related and differentially expressed lncRNAs. Finally, we performed multivariate Cox regression analysis to establish the three pyroptosis-related lncRNA signature. Kaplan-Meier (KM) survival analyses and receiver operating characteristic (ROC) curves indicated the excellent performance for predicting the prognosis of LUAD patients. At the same time, we applied multidimensional approaches to further explore the functional enrichment, tumor microenvironment (TME) landscape, and immunotherapy efficacy among the different risk groups. A nomogram was constructed by integrating risk scores and clinical characteristics, which was validated using calibrations and ROC curves. Three lncRNAs, namely, AC090559.1, AC034102.8, and AC026355.2, were involved in this signature and used to classify LUAD patients into low- and high-risk groups. Overall survival time (OS) was higher in the low-risk group than in the high-risk group, which was also validated in our LUAD cohort from Shandong Provincial Hospital. TME landscape analyses revealed that a higher abundance of infiltrating immune cells and a greater prevalence of immune-related events existed in the low-risk group. Meanwhile, higher expression of immune checkpoint (ICP) genes, higher immunophenoscore (IPSs), and greater T cell dysfunction in the low-risk group demonstrated a better response to immunotherapy than the high-risk group. Combined with predictions from the Tumor Immune Dysfunction and Exclusion (TIDE) website, we found that LUAD patients in the low-risk group significantly benefited from programmed cell death 1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 (CTLA4) immune checkpoint blockade (ICB) therapy compared with those in the high-risk group. Furthermore, drug susceptibility analysis identified potential sensitive chemotherapeutic drugs for each risk group. In this study, a novel three pyroptosis-related lncRNA signature was constructed, which could accurately predict the immunotherapy efficacy and prognosis in LUAD patients.

Keywords: immunotherapy; long noncoding RNA; lung adenocarcinoma; prognosis; pyroptosis.

FIGURE 1

Landscape of expression, genetic variation, and functional enrichment of pyroptosis-related genes in LUAD. (A) Expression of pyroptosis-related genes between normal tissues and LUAD tissues (Wilcoxon test, *p < 0.05; **p < 0.01; ***p < 0.001; ns, not statistically significant). (B) Landscape of mutation profiles in LUAD patients from the TCGA cohort. (C) CNV frequency of 33 pyroptosis-related genes. (D) Location of CNV alternation of pyroptosis-related genes in the chromosome. The red dots represent more samples with increased copy number gains than samples with copy number losses, while the blue dots are the opposite. The black dot means the two are equal. (E) Enrichment analysis of GO biological process, cellular component, and molecular function. LUAD, lung adenocarcinoma; CNV, copy number variation.

FIGURE 2

Identification and characteristics of pyroptosis-related lncRNAs. (A) Volcano plot showing the differently expressed pyroptosis-related lncRNAs. (B) Heat map visualizes the differential expression of prognostic pyroptosis-related lncRNAs between normal and LUAD. (C) Forest plot showing the result of univariate Cox regression analysis for screening prognosis-related lncRNAs (p < 0.05). (D) Interaction network of the 17 prognostic prognosis-related lncRNAs–mRNAs. (E) Visualization of prognosis-related lncRNA–mRNA correlation. ***p < 0.001, LUAD, lung adenocarcinoma.

FIGURE 3

Construction of risk model and clinical correlation of high- and low-risk groups. (A) Three lncRNAs were identified in multivariate Cox regression analysis for model construction. (B) Differential expression of pyroptosis-related genes between high- and low-risk groups (Wilcoxon test, *p < 0.05; **p < 0.01; ***p < 0.001). (C) Kaplan–Meier curve of high- and low-risk groups. (D) PCA of high and low-risk groups. (E) Risk curve based on the risk score of each sample. Scatterplot showing the survival status of LUAD patients. Heat map showing the expression of identified lncRNAs in high- and low-risk groups. (F) Relationship between tumor stage and risk score. (G) Time-dependent ROC curves of OS at 1, 3, and 5 years. (H) Comparison of the risk model with four published lncRNA signatures. OS, overall survival; PCA, principal component analysis.

FIGURE 4

Construction and validation of a nomogram. Univariate (A) and multivariate (B) Cox regression analyses of age, gender, stage, and risk score. (C) Nomogram for predicting the OS of LUAD patients at 1, 3, and 5 years. (D) Calibration curves of the nomogram for OS prediction at 1, 3, and 5 years. OS, overall survival; LUAD, lung adenocarcinoma.

FIGURE 5

Differences in landscape of the TME between high- and low-risk groups. (A) GSVA enrichment analysis of tumor hallmark pathways. (B–E) Differences in immune-related functions, TME infiltrating immune cells, other tumor-related functions, and HLA-related gene expression in the high- and low-risk groups. (F) Comparison of tumor purity, immune score, stromal score, and ESTIMATE score between high- and low-risk groups. TME, tumor microenvironment, *p < 0.05; **p < 0.01; ***p < 0.001; ns, not statistically significant.

FIGURE 6

Prediction of response to ICB therapy. (A) The expression of ICPs between high- and low-risk groups (Wilcox test, *p < 0.05; **p < 0.01; ***p < 0.001). The response to combined PD1 and CTLA4 blockade therapy (B) and PD1 monotherapy (C) between high- and low- risk groups. Differences in T cell status, including T cell dysfunction (D) and T cell exclusion (E), in high and low risk groups. (F–I) Prediction of the “responder” and “no benefit” of PD1 and CTLA4 blockade therapy in TCGA- LUAD patients from TIDE website. ICB immune checkpoint blockade, ICP immune checkpoint, PD1 programmed cell death protein 1, TCGA The Cancer Genome Atlas, LUAD lung adenocarcinoma.

FIGURE 7

Validation of risk score using clinical specimens of LUAD patients. (A) Kaplan–Meier curve of high- and low-risk groups in clinical LUAD patients. (B) Time-dependent ROC curves of OS at 1, 3, and 5 years. (C) The correlation between 3 identified lncRNAs, risk score and immune checkpoints. LUAD, lung adenocarcinoma; OS, overall survival.