Correction to "Circulating Levels of Galectin-9 Are a Potential Biomarker of Survival in Advanced Non-Small-Cell Lung Cancer".

Lung Cancer in Germany

Identification of secreted proteins of lung cancer could provide new candidates of serum biomarkers for cancer diagnosis or targets for therapeutic intervention. In this study, we developed a novel strategy that combined functional monoclonal antibody library screening technique and mass spectrometry to identify functional secreted proteins. BALB/c mice were immunized with cancer cells isolated from fresh human lung cancer tissues. The monoclonal antibody library containing 1160 mAbs was established with the mouse spleen cells, whose serum had most anti-proliferative effect on lung cancer cells. Monoclonal antibodies were subjected to an immunoreactive and functional screen and monoclonal antibodies that reacted strongly with secreted proteins in condition medium and lung cancer tissues with high inhibotion of cell proliferation were selected. Antigens that recognized by antibodies were obtained by immunoprecipitation and then identified by mass spectrometry. Mac-2-binding protein (Mac-2BP), the antigen of 13H3 antibody, was identified using this approach. Functional studies demonstrated that the 13H3 antibody suppressed lung cancer cell lines ANIP-973 and A549 proliferation in vitro and inhibit ANIP973 xenograft tumors growth in vivo by inducing cell-cycle arrest at G1 phase, with up-regulation of p27 and down-regulation of cyclin D1. Moreover, the serum level of Mac-2BP was significantly higher in lung cancer patients than healthy controls. At a cutoff value of 6 μg/ml, Mac-2BP might be a diagnostic biomarker of lung cancer, especially for SCLC. Mac-2BP concentrations of 6 μg/ml or higher was associated with poor overall survival in univariate analysis, and was an independent predictor in the multivariate COX analysis. Together, these results firstly demonstrated that Mac-2BP can be used as a therapeutic target and potential biomarker for lung cancer. Our strategy is feasible, which may facilitate the identification of novel secreted biomarkers of lung cancer.

Lung cancer is often asymptomatic or causes only nonspecific symptoms in its early stages. Early detection represents one of the most promising approaches to reduce the growing lung cancer burden. Human saliva is an attractive diagnostic fluid because its collection is less invasive than that of tissue or blood. Profiling of proteins in saliva over the course of disease progression could reveal potential biomarkers indicative of oral or systematic diseases, which may be used extensively in future medical diagnostics. There were 72 subjects enrolled in this study for saliva sample collection according to the approved protocol. Two-dimensional difference gel electrophoresis combined with MS was the platform for salivary proteome separation, quantification, and identification from two pooled samples. Candidate proteomic biomarkers were verified and prevalidated by using immunoassay methods. There were 16 candidate protein biomarkers discovered by two-dimensional difference gel electrophoresis and MS. Three proteins were further verified in the discovery sample set, prevalidation sample set, and lung cancer cell lines. The discriminatory power of these candidate biomarkers in lung cancer patients and healthy control subjects can reach 88.5% sensitivity and 92.3% specificity with AUC = 0.90. This preliminary data report demonstrates that proteomic biomarkers are present in human saliva when people develop lung cancer. The discriminatory power of these candidate biomarkers indicate that a simple saliva test might be established for lung cancer clinical screening and detection.

Previous studies have shown that asthma is a risk factor for lung cancer, while the mechanisms involved remain unclear. We attempted to further explore the association between asthma and non-small cell lung cancer (NSCLC) via bioinformatics analysis. We obtained GSE143303 and GSE18842 from the GEO database. Lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC) groups were downloaded from the TCGA database. Based on the results of differentially expressed genes (DEGs) between asthma and NSCLC, we determined common DEGs by constructing a Venn diagram. Enrichment analysis was used to explore the common pathways of asthma and NSCLC. A protein-protein interaction (PPI) network was constructed to screen hub genes. KM survival analysis was performed to screen prognostic genes in the LUAD and LUSC groups. A Cox model was constructed based on hub genes and validated internally and externally. Tumor Immune Estimation Resource (TIMER) was used to evaluate the association of prognostic gene models with the tumor microenvironment (TME) and immune cell infiltration. Nomogram model was constructed by combining prognostic genes and clinical features. 114 common DEGs were obtained based on asthma and NSCLC data, and enrichment analysis showed that significant enrichment pathways mainly focused on inflammatory pathways. Screening of 5 hub genes as a key prognostic gene model for asthma progression to LUAD, and internal and external validation led to consistent conclusions. In addition, the risk score of the 5 hub genes could be used as a tool to assess the TME and immune cell infiltration. The nomogram model constructed by combining the 5 hub genes with clinical features was accurate for LUAD. Five-hub genes enrich our understanding of the potential mechanisms by which asthma contributes to the increased risk of lung cancer.

ABSTRACTRecently, microbiota dysbiosis in lung cancer has attracted immense attention. Studies on lung microbes are mostly based on sequencing, which has left the potentially functional bacteria with extremely low abundance uncovered. In this study, we characterized and compared the lung and oral cavity microbiotas using culturomics and 16S rRNA gene sequencing. Of the 198 bacteria identified at the species level from bronchoalveolar lavage fluid (BALF) samples, Firmicutes was predominant (39.90%). Twenty bacterial species isolated from BALF samples were present in at least half of the patients and were also highly abundant in oral samples. Of all isolated strains, Streptococcus and Veillonella were highly dominant. The abundance of Prevotella and Veillonella decreased from the oral cavity to the lung, whereas that of Pseudomonas increased. Linear discriminant analysis effect size demonstrated that Prevotella was more abundant in the healthy samples than in the cancerous ones, which is in accordance with the isolation of Prevotella oralis only from the healthy group using culturomics. Moreover, Gemella sanguinis and Streptococcus intermedius were isolated only from the non-small-cell lung cancer (NSCLC) group, and 16S rRNA gene sequencing showed that they were higher in the NSCLC than in the small-cell lung cancer group. Furthermore, while Bacillus and Castellaniella were enriched in lung adenocarcinoma, Brucella was enriched in lung squamous cell carcinoma. Overall, alterations were observed in the microbial community of patients with lung cancer, whose diversity might be site and pathology dependent. Using culturomics and 16S rRNA gene amplicon sequencing, this study has provided insights into pulmonary and oral microbiota alterations in patients with lung cancer.IMPORTANCE The relationship between lung microbiota and cancer has been explored based on DNA sequencing; however, culture-dependent approaches are indispensable for further studies on the lung microbiota. In this study, we applied a comprehensive approach combining culturomics and 16S rRNA gene amplicon sequencing to detect members of the microbiotas in saliva and BALF samples from patients with unilateral lobar masses. We found alterations in the microbial community of patients with lung cancer, whose diversity might be site and pathology dependent. These features may be potential bacterial biomarkers and new targets for lung cancer diagnosis and treatment. In addition, a lung and oral microbial biobank from lung cancer patients was established, which represents a useful resource for studies of host-microbe interactions.

583P - Serum miRNA Assays for detection and management of gastric and lung cancers

Early stage lung cancer detection is the first step toward successful clinical therapy and increased patient survival. Clinicians monitor cancer progression by profiling tumor cell proteins in the blood plasma of afflicted patients. Blood plasma, however, is a difficult cancer protein assessment medium because it is rich in albumins and heterogeneous protein species. We report herein a method to detect the proteins released into the circulatory system by tumor cells. Initially we analyzed the protein components in the conditioned medium (CM) of lung cancer primary cell or organ cultures and in the adjacent normal bronchus using one-dimensional PAGE and nano-ESI-MS/MS. We identified 299 proteins involved in key cellular process such as cell growth, organogenesis, and signal transduction. We selected 13 interesting proteins from this list and analyzed them in 628 blood plasma samples using ELISA. We detected 11 of these 13 proteins in the plasma of lung cancer patients and non-patient controls. Our results showed that plasma matrix metalloproteinase 1 levels were elevated significantly in late stage lung cancer patients and that the plasma levels of 14-3-3 sigma, beta, and eta in the lung cancer patients were significantly lower than those in the control subjects. To our knowledge, this is the first time that fascin, ezrin, CD98, annexin A4, 14-3-3 sigma, 14-3-3 beta, and 14-3-3 eta proteins have been detected in human plasma by ELISA. The preliminary results showed that a combination of CD98, fascin, polymeric immunoglobulin receptor/secretory component and 14-3-3 eta had a higher sensitivity and specificity than any single marker. In conclusion, we report a method to detect proteins released into blood by lung cancer. This pilot approach may lead to the identification of novel protein markers in blood and provide a new method of identifying tumor biomarker profiles for guiding both early detection and therapy of human cancer.

Cancers of the lungs and head and neck are one of the leading causes of death, particularly in Kentucky, which leads the nation in multiple categories related to smoking. Thus, there is an urgent need for new biomarkers and therapeutic targets for airway cancers. The sigma-2 receptor is an intracellular receptor for numerous drugs and metabolites and was recently found to be identical to Pgrmc1 (progesterone receptor membrane component 1), a cytochrome-related protein implicated in metabolism and hormone signaling. S2RPgrmc1 is up-regulated in multiple types of cancer and is required for tumor cell proliferation and motility in vitro and tumor formation and metastasis in vivo. Furthermore, small molecule inhibitors of S2RPgrmc1 suppressed the growth of lung, breast and cervical cancer cell lines. In the present study, we have analyzed S2RPgrmc1 levels in clinical tumor samples from squamous cell lung cancers (SqCLC), lung adenocarcinomas, head and neck cancers and others. S2RPgrmc1 levels were increased in tumors throughout and correlated with survival in lung cancer (n=43, p=0.02) and with grade in head and neck cancer (higher in grade 2 versus 1, p=0.007). One of the cancer types with highest S2RPgrmc1 expression was squamous cell lung cancer (SqCLC), and treatment options for SqCLC are limited. S2RPgrmc1 was highly expressed in SqCLC cell lines, and SqCLC cell survival was inhibited by siRNA knockdown of S2RPgrmc1 or the S2RPgrmc1 inhibitor AG-205. S2RPgrmc1 is part of a family of secreted cytochromes, and we found that S2RPgrmc1 was significantly elevated in the plasma of 42 lung cancer patients compared to non-cancer patients and was significantly higher in the plasma of stage I lung cancer patients compared to stage 2-4 patients. Purified, recombinant S2RPgrmc1 increased the proliferation of NSCLC cell lines, suggesting that secreted S2RPgrmc1 may contribute to lung tumor growth. Together, the results demonstrate that S2RPgrmc1 correlates with key clinical endpoints in tumors and is a therapeutic target in some cancers with few therapeutic options, particularly squamous cell lung cancer. Furthermore, S2RPgrmc1 is a potential plasma biomarker for early stage lung cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3604. doi:1538-7445.AM2012-3604

Identification of lung cancer breath biomarkers based on perioperative breathomics testing: A prospective observational study

SummaryLung cancer is an extremely heterogeneous disease, and its treatment remains one of the most challenging tasks in medicine. Few existing laboratory lung cancer models can faithfully recapitulate the diversity of the disease and predict therapy response. Here, we establish 12 patient-derived organoids from the most common lung cancer subtype, lung adenocarcinoma (LADC). Extensive gene and histopathology profiling show that the tumor organoids retain the histological architectures, genomic landscapes, and gene expression profiles of their parental tumors. Patient-derived lung cancer organoids are amenable for biomarker identification and high-throughput drug screening in vitro. This study should enable the generation of patient-derived lung cancer organoid lines, which can be used to further the understanding of lung cancer pathophysiology and to assess drug response in personalized medicine.

Gut microbiota, especially human pathogens, has been shown to be involved in the occurrence and development of cancer. Esophageal squamous cell carcinoma and lung cancer are two malignant cancers, and their relationship with gut microbiota is still unclear. Virulence factor database (VFDB) is an integrated and comprehensive online resource for curating information about human pathogens. Here, based on VFDB database, we analyzed the differences of bacteria at genus level in the gut of patients with esophageal squamous cell carcinoma, lung cancer, and healthy controls. We proposed the possible cancer-associated bacteria in gut and put forward their possible effects. Apart from this, principal coordinate analysis (PCoA) and analysis of similarities (ANSOIM) suggested that some bacteria in the gut can be used as potential biomarkers to screen esophageal squamous cell carcinoma and lung cancer, and their effectiveness was preliminary verified. The relative abundance of Klebsiella and Streptococcus can be used to distinguish patients with esophageal squamous cell carcinoma and lung cancer from healthy controls. The absolute abundance of Klebsiella can further distinguish patients with esophageal squamous cell carcinoma from patients with lung cancer. In particular, the relative abundance of Fusobacterium can directly distinguish between patients with esophageal squamous cell carcinoma and healthy controls. Additionally, the absolute abundance of Haemophilus can distinguish lung cancer from healthy controls. Our study provided a new way based on VFDB database to explore the relationship between gut microbiota and cancer, and initially proposed a feasible cancer screening method.

We read with great interest the article, published in Cell Biochem Biophys, showing that serum level of miR-21 was significantly elevated in lung cancer patients compared with controls. Receiver operating characteristic (ROC) curve analysis showed that miR-21 could differentiate lung cancer from healthy controls with an AUC (the area under the curve) of 0.85, and the optimal sensitivity and specificity were 85.7 and 86.5 %, respectively [1]. These findings suggest that miR-21 can be a potential non-invasive biomarker for lung cancer patients and miR-21 may be a potential therapeutic target for lung cancer. Lung cancer is one of the most common malignant tumors and is the leading cause of cancer mortality in the world [2]. Non-small cell lung cancer (NSCLC) accounts for approximately 80–85 % of all cases of lung cancer [3]. Despite much advances in lung cancer therapy, the survival rate for lung cancer patients remains very poor [4]. Recent advances in functional genomics provide an increasingly comprehensive portrayal of lung tumorigenesis, but the molecular pathogenesis of lung cancer remains poorly understood. These findings highlight the need for a better understanding of novel molecular mechanisms to guide treatment of lung cancer. MicroRNAs (miRNAs) are small non-coding RNAs that repress target gene expression by a combination of mRNA degradation and translation inhibition [5]. Recently, miR-21 expression was found to be higher in lung cancer serum samples than in control serum samples [3, 6–9]. High serum miR-21 was significantly correlated with tumor node metastases stage and lymph node metastasis of NSCLC patients [3, 5]. Overexpression of miR-21 showed a highly discriminative ROC curve profile, clearly distinguishing cancer patients from cancer-free subjects, and the detection of miR-21 expression yielded high sensitivity and specificity in the diagnosis of lung cancer [7–9]. Furthermore, miR-21 expression was elevated in human lung cancer cell lines (NCI-H446 and NCI-H460) [2]. Human mutS homolog 2 (hMSH2), one of the core mismatch repair genes, is affected in lung cancer development. Zhong et al. [2] found that hMSH2 was directly regulated by miR-21, and miR-21 expression was inversely correlated with hMSH2 expression in human lung cancer cell lines [2]. Notably, upregulation of miR-21 markedly promoted cell proliferation and revealed a higher proportion of cells at S phase. However, knockdown of miR-21 expression led to cell cycle arrest at G2/M phase and inhibited cell proliferation [2]. Similarly, tumor tissues showed an inverse correlation between high expression of miR-21 and low amounts of PTEN protein, where miR-21 suppresses PTEN by direct binding to the 30-UTR of PTEN [5]. Lung cancer cell lines (A549, H1703, and 801D) transfected with miR-21 inhibitor showed decreased endogenous miR-21 levels but significantly increased PTEN protein expression [5]. In addition, A549 and H1703 cells transfected with miR-21 inhibitor could restrain the growth of A549 and H1703 cells, and downregulation of miR-21 with miR-21 inhibitor in 801D cells could restrain its invasion ability [5]. Collectively, available evidence suggests that miR-21 may be a good biomarker for detection of lung cancer, and miR-21 may play an important role in lung cancer. However, further studies, especially in human systems, are needed to comprehensively explore the role of miR-21 in lung cancer, and miR-21 may be a potential therapeutic target for lung cancer. S.-C. Wang (&) Y.-F. Zhang Q. Xie PET/CT Center, Anhui Provincial Hospital, 17 Lujiang Road, Hefei 230001, Anhui, People’s Republic of China e-mail: wangsc3329@163.com

Circulating tumor cells (CTCs) have emerged as potential biomarkers in several cancers such as colon, prostate, and breast carcinomas, with a correlation between CTC number and patient prognosis being established by independent research groups. The detection and enumeration of CTCs, however, is still a developing field, with no universal method of detection suitable for all types of cancer. CTC detection in lung cancer in particular has proven difficult to perform, as CTCs in this type of cancer often present with nonepithelial characteristics. Moreover, as many detection methods rely on the use of epithelial markers to identify CTCs, the loss of these markers during epithelial-to-mesenchymal transition in certain metastatic cancers can render these methods ineffective. The development of personalized medicine has led to an increase in the advancement of molecular characterization of CTCs. The application of techniques such as FISH and RT-PCR to detect EGFR, HER2, and KRAS abnormalities in lung, breast, and colon cancer, for example, could be used to characterize CTCs in real time. The use of CTCs as a ‘liquid biopsy’ is therefore an exciting possibility providing information on patient prognosis and treatment efficacy. This review summarizes the state of CTC detection today, with particular emphasis on lung cancer, and discusses the future applications of CTCs in helping the clinician to develop new strategies in patient treatment.

In 2024, an estimated 234,580 patients will be diagnosed with lung cancer and 125,070 are expected to die, making lung cancer the leading cause of cancer-related deaths. Early detection of lung cancer is critical for improving prognosis, hence the need for early detection biomarkers. Fat mass and obesity-associated gene (FTO) had earlier been linked with obesity, however current studies indicate it may have a role in tumor development and could be a potential biomarker. In this study, we aimed to identify a possible link between FTO and non-small cell lung cancer (NSCLC), which accounts for 85% of lung cancer diagnosis. We hypothesize FTO is involved in increasing tumorigenicity and mortality among lung cancer patients. The role of FTO was studied in tumor progression, patient survival analysis, and smoking induced lung cancer, which is not established. Using immunohistochemistry, 115 tumor sections of NSCLC patients were studied; of these patients, 64 were early-stage NSCLC and 51 were late-stage. Patient tissues were obtained from local hospital pathology archives. Staining was completed using FTO primary antibody and a biotinylated secondary antibody. FTO brightness scores were generated using the BZX-800 analyzer software. FTO expression was also calculated using an H-score with the help of a pathologist. Both imaging studies and pathology grading indicated that late-stage lung cancer tissues had significantly (p<0.05) higher FTO expression compared to early-stage cases. When subdivided into low and high expression levels, 69% of early-stage tissue samples exhibited low expression levels and 55% of late-stage tumor tissues had high FTO expression. To assess the role of FTO expression in lung cancer survival, a Kaplan-Meier analysis was done with 47 patients. Results showed that NSCLC patients with higher expression had a shorter median survival time (14 months) compared to NSCLC patients with low expression (27 months) (p=0.03). These results suggest that patients with higher FTO expression may have poorer prognosis. Since 80-90% of lung cancer patients had a history of smoking, we studied the connection between FTO expression and smoking associated lung cancer. 50 NSCLC tumor sections were stained, 28 being smokers and 22 being non-smokers. The average FTO expression level was 24% higher in smokers compared to non-smokers (p<0.02). Using H-scores, smokers were divided into low and high expression levels. 93% of smokers had a high FTO expression compared to 7% in non-smokers indicating that smokers had higher expression of FTO compared to non-smokers (P=0.001). In conclusion, FTO may have a role in tumor progression as it is highly expressed in late-stage lung tumor tissues. High expression of FTO is also a potential indicator of patient survival, as patients who had an upregulation of FTO had shorter survival time. Finally, we found a possible link between high FTO expression and smoking-induced lung cancer. Citation Format: Rachel Campoli, Rong Qiu, Robert Hillwig, Neelu Puri. FTO in smoking-associated lung cancer and lung cancer tumor progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3355.

Pleural effusion (PE), a tumor-proximal body fluid, may be a promising source for biomarker discovery in human cancers. Because a variety of pathological conditions can lead to PE, characterization of the relative PE proteomic profiles from different types of PEs would accelerate discovery of potential PE biomarkers specifically used to diagnose pulmonary disorders. Using quantitative proteomic approaches, we identified 772 nonredundant proteins from six types of exudative PEs, including three malignant PEs (MPE, from lung, breast, and gastric cancers), one lung cancer paramalignant PE, and two benign diseases (tuberculosis and pneumonia). Spectral counting was utilized to semiquantify PE protein levels. Principal component analysis, hierarchical clustering, and Gene Ontology of cellular process analyses revealed differential levels and functional profiling of proteins in each type of PE. We identified 30 candidate proteins with twofold higher levels (q<0.05) in lung cancer MPEs than in the two benign PEs. Three potential markers, MET, DPP4, and PTPRF, were further verified by ELISA using 345 PE samples. The protein levels of these potential biomarkers were significantly higher in lung cancer MPE than in benign diseases or lung cancer paramalignant PE. The area under the receiver-operator characteristic curve for three combined biomarkers in discriminating lung cancer MPE from benign diseases was 0.903. We also observed that the PE protein levels were more clearly discriminated in effusions in which the cytological examination was positive and that they would be useful in rescuing the false negative of cytological examination in diagnosis of nonsmall cell lung cancer-MPE. Western blotting analysis further demonstrated that MET overexpression in lung cancer cells would contribute to the elevation of soluble MET in MPE. Our results collectively demonstrate the utility of label-free quantitative proteomic approaches in establishing differential PE proteomes and provide a new database of proteins that can be used to facilitate identification of pulmonary disorder-related biomarkers.