Blood-based molecular and cellular biomarkers of early response to neoadjuvant PD-1 blockade in patients with non-small cell lung cancer.

BACKGROUND: Despite the improved survival observed in PD-1/PD-L1 blockade therapy, a substantial proportion of cancer patients, including those with non-small cell lung cancer (NSCLC), still lack a response. METHODS: Transcriptomic profiling was conducted on a discovery cohort comprising 100 whole blood samples, as collected multiple times from 48 healthy controls (including 43 published data) and 31 NSCLC patients that under treatment with a combination of anti-PD-1 Tislelizumab and chemotherapy. Differentially expressed genes (DEGs), simulated immune cell subsets, and germline DNA mutational markers were identified from patients achieved a pathological complete response during the early treatment cycles. The predictive values of mutational markers were further validated in an independent immunotherapy cohort of 1661 subjects, and then confirmed in genetically matched lung cancer cell lines by a co-culturing model. RESULTS: The gene expression of hundreds of DEGs (FDR p < 0.05, fold change < -2 or > 2) distinguished responders from healthy controls, indicating the potential to stratify patients utilizing early on-treatment features from blood. PD-1-mediated cell abundance changes in memory CD4 + and regulatory T cell subset were more significant or exclusively observed in responders. A panel of top-ranked genetic alterations showed significant associations with improved survival (p < 0.05) and heightened responsiveness to anti-PD-1 treatment in patient cohort and co-cultured cell lines. CONCLUSION: This study discovered and validated peripheral blood-based biomarkers with evident predictive efficacy for early therapy response and patient stratification before treatment for neoadjuvant PD-1 blockade in NSCLC patients.

miR-31-5p modulates cell progression in lung adenocarcinoma through TNS1/p53 axis.

OBJECTIVE: To clarify the modulatory mechanism of miR-31-5p in lung adenocarcinoma (LUAD) progression in vivo and in vitro. METHODS: The Cancer Genome Atlas (TCGA) database was employed to access LUAD-related miRNA and mRNA expression data. Downstream targets of miR-31-5p were predicted by public databases. The interaction between miR-31-5p and TNS1 was determined by dual-luciferase reporter assay. Quantitative real-time polymerase chain reaction (qRT-PCR) was utilized to measure miR-31-5p and TNS1 expression levels in LUAD cells. Western blot was introduced to test protein expression levels of TNS1, p53, and apoptosis-related proteins. In-vitro functional assays were conducted to evaluate the biological effects of miR-31-5p on cell proliferation, colony formation, migration, and apoptosis. In-vivo tumor xenograft experiment was applied to examine the effects of miR-31-5p on LUAD tumor growth, followed by immunochemistry assays for assessing TNS1 and p53 expression levels in the tumor tissue. RESULTS: miR-31-5p was prominently upregulated in LUAD tissue and was identified to present a similar trend in LUAD cell lines H1299, H23, and A549. miR-31-5p overexpression exerted an active role in cell proliferation and migration, but it suppressed cell apoptosis. Additionally, a reverse correlation between miR-31-5p and TNS1 regarding the expression level was identified, and TNS1 was verified to be a direct target of miR-31-5p. Besides, it was further validated by the rescue experiments that the tumor-promoting effects of miR-31-5p on LUAD cell functions were attenuated by TNS1 overexpression to some extent. The results based on the tumor xenograft experiment revealed that LUAD cell growth could be facilitated by miR-31-5p via the TNS1/p53 axis. CONCLUSION: miR-31-5p facilitates LUAD cell progression mediated by the TNS1/p53 axis.

A prognostic nomogram based on a new classification of combined micropapillary and solid components for stage IA invasive lung adenocarcinoma.

BACKGROUND: We aimed to develop a prognostic nomogram based on a new classification of combined micropapillary and solid components in pathological stage IA invasive lung adenocarcinoma (LUAD). METHODS: According to the total proportion of solid and micropapillary components (TPSM), the X-tile software was applied to classify patients into the following three groups: TPSM-low (TPSM-L), TPSM-middle (TPSM-M), and TPSM-high (TPSM-H). The postoperative survival was compared among the three groups. The multivariate Cox regression analysis was performed to identify independent prognostic factors for survival. According to these factors, a nomogram model was developed to provide a personalized prognostic evaluation. RESULTS: A total of 595 patients with pathological stage IA invasive LUAD were included in our study. The 5-year disease-free survival and overall survival rates in patients with TPSM-H and TPSM-M were significantly lower than those with TPSM-L. The multivariate Cox regression analysis revealed that the TPSM classification was an independent prognostic factor for survival. According to TPSM classification, we developed a nomogram model which had good calibration and reliable discrimination ability to evaluate survival. CONCLUSIONS: The nomogram based on the combination of micropapillary and solid components has good prognostic value in predicting postoperative recurrence and survival of patients with pathological stage IA invasive LUAD.

miR-423-3p activates FAK signaling pathway to drive EMT process and tumor growth in lung adenocarcinoma through targeting CYBRD1.

BACKGROUND: Lung adenocarcinoma (LUAD) is a malignant tumor with a high fatality rate and poor overall survival, while molecular targets diagnosing and alleviating lung cancer remain inadequate. METHODS: In this article, we highlighted the upregulation of microRNA-423-3p (miR-423-3p) in LUAD, especially in smokers aged over 40, and revealed that the high expression of miR-423-3p was significantly associated with smoker, age, and pathologic stage of LUAD patients. RESULTS: Moreover, overexpressing miR-423-3p could facilitate LUAD cell proliferation, invasion, adhesion, and epithelial-mesenchymal transition (EMT) process, while depleted miR-423-3p caused repressive influence upon it. Mechanically, we identified that miR-423-3p could activate FAK signaling pathway through binding to the 3'-UTR of cytochrome B reductase 1 (CYBRD1). Furthermore, we demonstrated that CYBRD1 was lowly expressed in LUAD, and miR-423-3p overexpression could rescue the impairment of LUAD cell proliferation, invasion, adhesion, and EMT caused by CYBRD1 depletion. Noticeably, miR-423-3p depletion efficiently hindered LUAD tumor growth in vivo. CONCLUSION: Collectively, our findings demonstrated that miR-423-3p/CYBRD1 axis could be regarded as a promising biomarker to alleviate the poor LUAD prognosis.

The Role of Neutrophil-to-Lymphocyte Ratio in Predicting Pathological Response for Resectable Non-Small Cell Lung Cancer Treated with Neoadjuvant Chemotherapy Combined with PD-1 Checkpoint Inhibitors.

PURPOSE: The aim of our study was to investigate the value of baseline and preoperative neutrophil-to-lymphocyte ratio (NLR) in predicting the pathological response and disease-free survival (DFS) of neoadjuvant chemotherapy alone or combined with programmed cell death-1 (PD-1) checkpoint inhibitors in patients with resectable non‒small cell lung cancer (NSCLC). MATERIALS AND METHODS: Resectable NSCLC patients who underwent neoadjuvant chemotherapy alone or combined with PD-1 checkpoint inhibitors between January 2018 and January 2020 were included. Peripheral venous blood samples of the patients were collected within 3 days prior to the first neoadjuvant treatment and within 3 days prior to surgery. RESULTS: A total of 79 patients in neoadjuvant chemotherapy combined with PD-1 checkpoint inhibitors group and 89 patients in neoadjuvant chemotherapy alone group were included. Thirty-five point four percent of the patients achieved pathological complete response (pCR) in neoadjuvant chemotherapy combined with PD-1 checkpoint inhibitors group, whereas only 9.0% reached pCR in the group of neoadjuvant chemotherapy. High NLR level were correlated with poor pathological response and DFS in neoadjuvant chemotherapy or combined with PD-1 checkpoint inhibitors group. Multivariate analysis revealed that baseline NLR could independently predict pathological response and DFS in the neoadjuvant chemotherapy combined with PD-1 checkpoint inhibitors group. CONCLUSION: High NLR level were correlated with poor pathological response and shorter DFS in patients with NSCLC undergoing neoadjuvant chemotherapy or combined with PD-1 checkpoint inhibitors. Meanwhile, baseline NLR could independently predict response to pathological response and DFS, revealing its potential as a screening tool in NSCLC patients who received neoadjuvant chemotherapy combined with PD-1 checkpoint inhibitors.

Comprehensive analyses unveil novel genomic and immunological characteristics of micropapillary pattern in lung adenocarcinoma.

Lung adenocarcinoma (LUAD) usually contains heterogeneous histological subtypes, among which the micropapillary (MIP) subtype was associated with poor prognosis while the lepidic (LEP) subtype possessed the most favorable outcome. However, the genomic features of the MIP subtype responsible for its malignant behaviors are substantially unknown. In this study, eight FFPE samples from LUAD patients were micro-dissected to isolate MIP and LEP components, then sequenced by whole-exome sequencing. More comprehensive analyses involving our samples and public validation cohorts on the two subtypes were performed to better decipher the key biological and evolutionary mechanisms. As expected, the LEP and MIP subtypes exhibited the largest disease-free survival (DFS) differences in our patients. EGFR was found with the highest mutation frequency. Additionally, shared mutations were observed between paired LEP and MIP components from single patients, and recurrent mutations were verified in the Lung-Broad, Lung-OncoSG, and TCGA-LUAD cohorts. Distinct biological processes or pathways were involved in the evolution of the two components. Besides, analyses of copy number variation (CNV) and intratumor heterogeneity (ITH) further discovered the possible immunosurveillance escape, the discrepancy between mutation and CNV level, ITH, and the pervasive DNA damage response and WNT pathway gene alternations in the MIP component. Phylogenetic analysis of five pairs of LEP and MIP components further confirmed the presence of ancestral EGFR mutations. Through comprehensive analyses combining our samples and public cohorts, PTP4A3, NAPRT, and RECQL4 were identified to be co-amplified. Multi-omics data also demonstrated the immunosuppression prevalence in the MIP component. Our results uncovered the evolutionary pattern of the concomitant LEP and MIP components from the same patient that they were derived from the same initiation cells and the pathway-specific mutations acquired after EGFR clonal mutation could shape the subtype-specificity. We also confirmed the immunosuppression prevalence in the MIP subtype by multi-omics data analyses, which may have resulted in its unfavorable prognosis.

Plasma CXCL14 as a Candidate Biomarker for the Diagnosis of Lung Cancer.

Background: Effective biomarkers for early diagnosis of lung cancer are needed. Previous studies have indicated positive associations between abnormal circulating cytokines and the etiology of lung cancer. Methods: Blood samples were obtained from 286 patients with pretreatment lung cancer and 80 healthy volunteers. Circulating cytokine levels were detected with a Luminex assay and enzyme-linked immunosorbent assay (ELISA). Urine samples were obtained from 284 patients and 122 healthy volunteers. CXC chemokine ligand 14 (CXCL14) expression in tumors and nontumor regions of lung tissues from 133 lung cancer cases was detected by immunohistochemical (IHC) staining and immunofluorescence (IF) staining of formalin fixed paraffin-embedded (FFPE) tissues. Results: Compared with healthy volunteers, a 65.7-fold increase was observed in the level of CXCL14 in the plasma of lung cancer patients, and a 1.7-fold increase was observed in the level of CXCL14 in the urine of lung cancer patients, achieving a 0.9464 AUC (area under the curve) value and a 0.6476 AUC value for differentiating between lung cancer patients and healthy volunteers, respectively. Stromal CXCL14 expression was significantly associated with advanced pathologic stage (P<0.001), pathologic N stage (P<0.001), and recurrence and metastasis (P=0.014). Moreover, multivariate analysis suggested stromal CXCL14 expression as an independent predictor of DFS and OS. Conclusions: Our study demonstrates that CXCL14 might serve as a potential diagnostic and prognostic biomarker in patients with lung cancer. Impact: CXCL14 might serve as a potential diagnostic and prognostic biomarker in patients with lung cancer.

Linc8087 predicts favorable prognosis and inhibits cell migration and invasion in NSCLC.

BACKGROUND: Non-small cell lung cancer (NSCLC) is the most common cancer and has poor prognosis. Long non-coding RNA(LncRNA) plays important roles in the regulation of cell migration in various types of cancer. In this study, we aimed to demonstrate the function of linc8087 in regulating cell migration and invasion in NSCLC cells. METHODS: A lncRNA microarray was used to identify differentially expressed lncRNAs between NSCLC tissues and normal tissues. RT-qPCR was used to confirm the expression of linc8087 in tumor tissues. The association between linc8087 expression and clinicopathological characteristics was analyzed. RNA fluorescence in situ hybridization (FISH) was performed to observe the subcellular localization of linc8087. We investigated the effects of linc8087 expression on cell migration and invasion by wound healing assay, Transwell and invasion assays. The Human Tumor Metastasis RT2 Profiler PCR Array was used to detect and analyze the mRNA levels of 84 genes involved in metastasis. RESULTS: We found that linc8087 expression was obviously decreased in both NSCLC tissues and cell lines compared with paired normal tissues and a normal bronchial epithelium cell line. Low expression of linc8087 was significantly associated with poor survival. In addition, linc8087 was an independent risk factor for survival. Overexpressed linc8087 inhibited cell migration and invasion in A549 and PC9 cell lines. Knockdown of linc8087 promoted cell migration and invasion. The result of RT2 Profiler PCR Array showed that overexpressed linc8087 upregulated the expression of the COL4A2, CST7 and FAT1 genes and led to the downregulation of SERPINE1. CONCLUSIONS: These results indicate that linc8087 plays a key role in the progression of NSCLC, and it may serve as a meaningful prognostic biomarker as well as a latent therapeutic target in NSCLC patients.

Bioinformatic analysis of PLOD family member expression and prognostic value in non-small cell lung cancer.

BACKGROUND: Procollagen-lysine, 2-oxoglutarate 5-dioxygenases (PLODs) are a group of enzymes that can mediate the hydroxylation of lysyl to hydroxylysine and participate in the formation of stabilized collagen. Evidence has demonstrated that PLODs are involved in the steps of tumor progression, including proliferation, invasion, and metastasis. However, limited information is available on the function of PLOD1/2/3 in lung cancer. In this study, we investigated the expression patterns and prognostic values of PLODs in patients with lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC). METHODS: The Oncomine database and UALCAN were used to analyze the mRNA expression levels of PLOD family members in non-small cell lung cancer (NSCLC). The prognostic values of PLODs were investigated by the Kaplan-Meier Plotter database. We collected 33 patients with lung cancer to further verify the expression profiles and prognostic values of PLODs. The Kaplan-Meier method was used to perform survival curves, and the log-rank test was performed to evaluate the differences in survival. According to the GSE31210 databset, univariate and multivariate analyses were performed to identify whether PLODs were independent prognostic indicators for survival. Meanwhile, we investigated the mutations, potential biological functions and immune relevance of PLODs on the basis of the cBioPortal, Metascape and TIMER databases respectively. RESULTS: We found that the mRNA and protein expression levels of PLODs in NSCLC tissues were higher than those in normal lung tissues. High PLOD1/2/3 expression had significant relevance to poor survival in LUAD but not in LUSC. In addition, the GSE31210 dataset showed that PLOD1 and PLOD3 were independent risk factors for relapse-free survival and overall survival (OS) in LUAD. We observed a high alteration rate of PLODs in LUSC patients, and the genetic alterations of PLODs had significant relevance to favorable OS. Furthermore, we observed that PLODs were significantly associated with tumor immunity in lung cancer. The enrichment analysis of the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway showed that the functions of the PLODs focused on cell cycle, DNA replication, and glycolysis/gluconeogenesis in LUAD. CONCLUSIONS: These results indicated that PLODs were highly expressed in lung cancer and may be suitable prognostic markers.

The Prognostic Value of Non-Predominant Micropapillary Pattern in a Large Cohort of Resected Invasive Lung Adenocarcinoma Measuring ≤3 cm.

The aim of this study was to analyze the influence of non-predominant micropapillary pattern in small sized invasive lung adenocarcinoma. A total of 986 lung adenocarcinoma patients with tumor size ≤3 cm were identified and classified according to the IALSC/ATS/ERS classification. Emphasis was placed on the impact of non-predominant micropapillary pattern on disease-free survival (DFS) and overall survival (OS). The relationship between lung adenocarcinoma subtype and lymph node involvement, EGFR mutation and KRAS mutation was also evaluated. A nomogram was developed to predict the probability of 3- and 5-year OS for these patients. The concordance index and calibration plot were used to validate this model. Among all 986 patients, the percentages of lymph node involvement were: 58.1, 50.0, 33.5, 21.4, 21.1, 10.9, 0, and 0% for micropapillary predominant, solid predominant, acinar predominant, papillary predominant, invasive mucinous adenocarcinoma (IMA), lepidic predominant, minimally invasive adenocarcinoma (MIA), adenocarcinoma in situ (AIS), respectively. The frequency of EGFR mutation in the cases of lepidic predominant, acinar predominant, MIA, micropapillary predominant, papillary predominant, solid predominant, IMA, and AIS were 51.1, 45.2, 44.4, 36.8, 29.3, 26.8, 8.3, and 0%, respectively. A non-predominant micropapillary pattern was observed in 344 (38.4%) invasive adenocarcinoma (IAC), and its presence predicted a poorer DFS (median: 56.0 months vs. 66.0 months, P <0.001) and OS (median: 61.0 months vs. 70.0 months, P <0.001). After propensity score matching, non-predominant micropapillary pattern retained its unfavorable effect on DFS (P = 0.007) and OS (P = 0.001). Multivariate analysis showed that non-predominant micropapillary pattern was identified as an independent prognostic factor for DFS (P = 0.003) and OS (P <0.001) in IAC. The nomogram showed good calibration and reliable discrimination ability (C-index = 0.775) to evaluated the 3- and 5-year OS. This retrospective analysis of patients with small sized IAC suggests the value of non-predominant micropapillary pattern to predict poor prognosis. A reliable nomogram model was constructed to provide personalized survival predictions.

Prognostic significance of combined fibrinogen concentration and neutrophil-to-lymphocyte ratio in patients with resectable non-small cell lung cancer.

OBJECTIVE: Cancer-associated inflammation and coagulation cascades play vital roles in cancer progression and survival. In this study, we investigated the significance of the combination of preoperative fibrinogen and the neutrophil-to-lymphocyte ratio (NLR) in predicting the survival of patients with non-small cell lung cancer (NSCLC). METHODS: We retrospectively enrolled 589 patients with NSCLC who underwent surgery. The univariate and multivariate Cox survival analyses were used to evaluate the prognostic indicators, including the combination of fibrinogen and NLR (F-NLR). The cut-off values for fibrinogen, NLR, and clinical laboratory variables were defined by the receiver operating characteristic (ROC) curve analysis. According to the ROC curve, the recommended cut-off values for fibrinogen and the NLR were 3.48 g/L and 2.30, respectively. Patients with both a high NLR (≥ 2.30) and hyperfibrinogenemia (≥ 3.48 g/L) were given a score of 2, whereas those with one or neither were scored as 1 or 0, respectively. RESULTS: Our results showed that F-NLR was an independent prognostic indicator for disease-free survival (DFS) [hazard ratio (HR), 1.466; 95% confidence interval (CI), 1.243-1.730; P < 0.001] and overall survival (OS) (HR, 1.512; 95% CI, 1.283-1.783; P < 0.001). The five-year OS rates were 66.1%, 53.5%, and 33.3% for the F-NLR = 0, F-NLR = 1, and F-NLR = 2, respectively ( P < 0.001). Correspondingly, their five-year DFS rates were 62.2%, 50.3%, and 30.4%, respectively ( P < 0.001). In the subgroup analyses of the pathological stages, the F-NLR level was significantly correlated with DFS and OS in stage I and IIIA cancers. CONCLUSIONS: Preoperative F-NLR score can be used as a valuable prognostic marker for patients with resectable early-stage NSCLC.

Prognostic value of the combined preoperative plasma levels of fibrinogen and lym-phocyte to monocyte ratio (F-LMR) in patients with non-small cell lung cancer / 中国肿瘤临床

Objective:This study aims to evaluate the correlation of combined preoperative plasma levels of fibrinogen (Fbg) and lym-phocyte to monocyte ratio (LMR) (F-LMR) with the prognosis of patients with non-small cell lung cancer (NSCLC) after complete resec-tion. Methods:The clinical data of 589 patients with NSCLC who underwent complete resection in our hospital were retrospectively analyzed. Receiver operating characteristic curve (ROC) analysis was used to select the cut-off values of Fbg and LMR. Based on the cri-teria of F-LMR, we divided the patients into three groups:F-LMR 0 score, F-LMR 1 score, and F-LMR 2 score. The association between F-LMR and the clinicopathological characteristics was analyzed by theχ2 test. Kaplan-Meier analysis was used to analyze the prognostic factors, and the log-rank test was used to determine the differences in survival rates. Prognostic factors were assessed by univariate and multivariate analyses (Cox's proportional hazards regression model). Results:According to the ROC curve, the cut-off values of Fbg and LMR were 3.48 g/L and 3.23, respectively. F-LMR 0 score had n=215, F-LMR 1 score had n=228, and F-LMR 2 score had n=146. Pre-operative F-LMR was closely related to age, gender, smoking history, tumor location, surgical type, pathological stage, pathological type, and tumor size (P<0.05). Univariate analysis showed that tumor location, surgical type, pathological stage, tumor size, F-LMR score, LMR, and Fbg were associated with survival (P<0.05). Multivariate analysis showed that the pathological stage [disease-free sur-vival (DFS): hazard ratio (HR)=1.700, 95%confidence interval (CI)=1.483-1.950, P<0.001;overall survival (OS):HR=1.703, 95%CI=1.486-1.952, P<0.001] and F-LMR score (DFS:HR=1.264, 95%CI=1.077-1.484, P=0.004;OS:HR=1.301, 95%CI=1.107-1.528, P=0.001) were the independent prognostic factors of NSCLC patients. Conclusion:The preoperative F-LMR score may be a useful blood marker for predicting the prognosis of patients with NSCLC with radical resection.

Prognostic value of the combined preoperative plasma levels of fibrinogen and lym-phocyte to monocyte ratio (F-LMR) in patients with non-small cell lung cancer / 中国肿瘤临床

Objective:This study aims to evaluate the correlation of combined preoperative plasma levels of fibrinogen (Fbg) and lym-phocyte to monocyte ratio (LMR) (F-LMR) with the prognosis of patients with non-small cell lung cancer (NSCLC) after complete resec-tion. Methods:The clinical data of 589 patients with NSCLC who underwent complete resection in our hospital were retrospectively analyzed. Receiver operating characteristic curve (ROC) analysis was used to select the cut-off values of Fbg and LMR. Based on the cri-teria of F-LMR, we divided the patients into three groups:F-LMR 0 score, F-LMR 1 score, and F-LMR 2 score. The association between F-LMR and the clinicopathological characteristics was analyzed by theχ2 test. Kaplan-Meier analysis was used to analyze the prognostic factors, and the log-rank test was used to determine the differences in survival rates. Prognostic factors were assessed by univariate and multivariate analyses (Cox's proportional hazards regression model). Results:According to the ROC curve, the cut-off values of Fbg and LMR were 3.48 g/L and 3.23, respectively. F-LMR 0 score had n=215, F-LMR 1 score had n=228, and F-LMR 2 score had n=146. Pre-operative F-LMR was closely related to age, gender, smoking history, tumor location, surgical type, pathological stage, pathological type, and tumor size (P<0.05). Univariate analysis showed that tumor location, surgical type, pathological stage, tumor size, F-LMR score, LMR, and Fbg were associated with survival (P<0.05). Multivariate analysis showed that the pathological stage [disease-free sur-vival (DFS): hazard ratio (HR)=1.700, 95%confidence interval (CI)=1.483-1.950, P<0.001;overall survival (OS):HR=1.703, 95%CI=1.486-1.952, P<0.001] and F-LMR score (DFS:HR=1.264, 95%CI=1.077-1.484, P=0.004;OS:HR=1.301, 95%CI=1.107-1.528, P=0.001) were the independent prognostic factors of NSCLC patients. Conclusion:The preoperative F-LMR score may be a useful blood marker for predicting the prognosis of patients with NSCLC with radical resection.