Small cell lung cancer (SCLC) represents one of the most aggressive malignancies among cancer types. Not only tumor sample availability is limited, but also the ability for tumor cells to rapidly acquire drug resistance are the rate-limiting bottlenecks for overall survival in current clinical settings. A liquid biopsy capable of capturing and enriching circulating tumor cells (CTCs), together with the possibility of drug screening, is a promising solution. Here, we illustrate the development of a highly efficient ex vivo CTC expansion system based on binary colloidal crystals substrate. Clinical samples were enrolled from 22 patients with SCLC in the study. The CTCs were enriched and expanded from the collected peripheral blood samples. Expanded cells were analyzed for protein expression and observed for drug sensitivity with the use of immunofluorescence and ATP titer evaluation, respectively. Successful CTC spheroid proliferation was established after 4 weeks within 82% of all the collected peripheral blood samples from enrolled patients. Upon immunofluorescence analysis, the enriched cells showed positive markers for EpCAM, TTF-1, synaptophysin and negative for CD45. Additionally, the expanded CTCs demonstrated marked heterogeneity in the expression of E-cadherin and N-cadherin. In a preliminary case series, the drug sensitivity of patient-derived CTC to cisplatin and etoposide was studied to see the correlation with the corresponding therapeutic outcome. In conclusion, our study demonstrates that it is possible to efficiently expand CTCs from SCLC within a clinically relevant time frame; the biomarker information generated from enriched CTCs can assist the selection of effective drugs and improve disease outcome.

Non-small-cell lung cancer (NSCLC) is a typical inflammation-associated cancer, and lung adenocarcinoma (LUAD) is the most common pathological subtype. Epidermal growth factor (EGF) receptor (EGFR) mutations are the most common driver mutations of LUAD, and they have been identified as important therapeutic targets by EGFR-tyrosine kinase inhibitors (TKIs). The proinflammatory cytokine, interleukin (IL)-17A, and IL-17A-producing cells were reported to be elevated in the tumor microenvironment and peripheral blood of NSCLC patients and to be correlated with tumor progression and poor prognoses. However, the pathophysiological role of IL-17A in NSCLC remains unclear, although some studies suggested its involvement in cancer cell invasion and metastasis. Herein, we observed that expressions of IL-17A and its receptor, IL-17 receptor C (IL-17RC), were elevated in LUAD tissues and were correlated with poor survival in different lung cancer cohorts. In LUAD cells with mutant EGFR, the IL-17A/IL-17RC axis was shown to enhance phosphorylation of EGFR and Met, thereby promoting proliferation and resistance to EGFR-TKIs such as afatinib. In LUAD cells with wild-type (WT) EGFR, we found that the IL-17A/IL-17RC axis enhanced EGF-induced EGFR activation and cell proliferation through causing impairment of EGF-induced EGFR lysosomal degradation. Collectively, our results indicated diverse impacts of the IL-17A/IL-17RC axis on EGFR activation in LUAD cells with WT and mutant EGFR and suggested that developing therapeutic strategies against IL-17A/IL-17RC would be valuable for LUAD treatment.

Lung adenocarcinoma (LADC) is a major subtype of lung cancer, particularly among populations of East Asia. The epidermal growth factor receptor (EGFR) is the most frequently mutated oncogene promoting LADC progression and can serve as a therapeutic target in LADC. The tissue inhibitor of metalloproteinases (TIMP)-3 is a major regulator of extracellular matrix turnover via targeting of matrix metalloproteinases (MMPs), and thus, plays a critical role in tumor development and progression. The purpose of this study was to investigate potential associations among TIMP-3 genetic polymorphisms, EGFR statuses, and cancer clinicopathologic development in patients with LADC. In this study, 277 LADC patients with different EGFR statuses were recruited to dissect the allelic discrimination of TIMP-3 -1296 T>C (rs9619311), TIMP3 249T>C (rs9862), and TIMP3 261C>T (rs11547635) polymorphisms using a TaqMan allelic discrimination assay. Our data showed that compared to those LADC patients with wild-type CC homozygotes of TIMP-3 rs9862, patients harboring TT homozygotes of rs9862 were at a higher risk of developing mutant EGFR (adjusted odds ratio (AOR) = 2.530; 95% confidence interval (CI): 1.230–5.205; p = 0.012), particularly the EGFR L858R point mutation (AOR = 2.975; 95% CI: 1.182–7.488; p = 0.021). Moreover, we observed that TIMP-3 TT homozygotes of rs9862 were correlated with the incidence of EGFR mutations in patients with a smoking habit (p = 0.045). Within male patients harboring a mutant EGFR, TIMP-3 rs9862 T (CT+TT) allele carriers were at higher risk of developing an advanced stage (p = 0.025) and lymph node metastasis (p = 0.043). Further analyses of clinical datasets revealed correlations of TIMP-3 expression with a favorable prognosis in patients with LADC. In conclusion, the data suggest that TIMP-3 rs9862 polymorphisms may contribute to identify subgroups of lung cancer patients at high risk for tumor progression, among carriers of LADC-bearing mutant EGFR.

Non-small cell lung cancer (NSCLC) is a typical inflammation-associated cancer, and lung adenocarcinoma (LUAD) is the most common histopathological subtype. Epidermal growth factor receptor (EGFR) mutations are the most common driver mutations of LUAD, and they have been identified as important therapeutic targets by EGFR tyrosine kinase inhibitors. Interleukin (IL)-17A secreted by T-helper 17 lymphocytes is a proinflammatory cytokine that plays an important role in cancer pathogenesis. The present study was designed to investigate the possible associations among IL-17A genetic polymorphisms, EGFR mutation status, and the clinicopathologic development of LUAD in a Taiwanese population. Our study population consisted of 277 LUAD patients harboring the wild-type (WT) EGFR or a mutant (MT) EGFR. Four single-nucleotide polymorphisms (SNPs) of IL-17A in the peripheral blood, including rs8193036(C > T), rs8193037(G > A), rs2275913(G > A), and rs3748067(C > T) loci, were genotyped using a TaqMan allelic discrimination assay. Our results showed that none of these IL-17A SNPs were correlated with the risk of developing mutant EGFR. However, patients with a smoking habit who carried the GA genotype of IL-17A rs8193037 had a significantly lower susceptibility to EGFR mutations (adjusted odds ratio (AOR): 0.225; 95% confidence interval (CI): 0.056~0.900, p = 0.035). Moreover, compared to individuals carrying the CC genotype of rs8193036 at IL-17A, T-allele carriers (CT + TT) were at higher risk of developing more-advanced stages (stage III or IV; p = 0.020). In the WT EGFR subgroup analysis, IL-17A rs8193036 T-allele carriers had higher risks of developing an advanced tumor stage (p = 0.016) and lymphatic invasion (p = 0.049). Further analyses of clinical datasets revealed correlations of IL-17 receptor A (IL-17RA) and IL-17RC expressions with a poor prognosis of LUAD patients with a smoking history or with higher levels of tumor-infiltrating lymphocytes. In conclusion, our results suggested that two functional promoter polymorphisms of IL-17A, i.e., rs8193036 and rs8193037, were associated with the EGFR mutation status and progression in LUAD patients, indicating that these two genetic variants might act as possible markers for predicting patients’ clinical prognoses.

Toll-like receptor (TLR) is essential for the immune response to Mycobacterium tuberculosis (MTB) infection. However, the mechanism whereby TLR mediates the MTB-induced pleural mesothelial hyperpermeability in tuberculous pleural effusion (TBPE) remains unclear. Pleural effusion size and pleural fluid levels of vascular endothelial growth factor (VEGF) and soluble TLR2 (sTLR2) in patients with TBPE (n = 36) or transudative pleural effusion (TPE, n = 16) were measured. The effects of MTB H37Ra (MTBRa) on pleural mesothelial permeability and the expression of VEGF and zonula occludens (ZO)-1 in human pleural mesothelial cells (PMCs) were assessed. Levels of VEGF and sTLR2 were significantly elevated in TBPE compared to TPE. Moreover, effusion VEGF levels correlated positively, while sTLR2 values correlated negatively, with pleural effusion size in TBPE. In human PMCs, MTBRa substantially activated JNK/AP-1 signaling and upregulated VEGF expression, whereas knockdown of TLR2 remarkably inhibited MTBRa-induced JNK phosphorylation and VEGF overexpression. Additionally, both MTBRa and VEGF markedly reduced ZO-1 expression and induced pleural mesothelial permeability, while TLR2 silencing or pretreatment with anti-VEGF antibody significantly attenuated the MTBRa-triggered effects. Collectively, TLR2 mediates VEGF overproduction and downregulates ZO-1 expression in human PMCs, leading to mesothelial hyperpermeability in TBPE. Targeting TLR2/VEGF pathway may confer a potential treatment strategy for TBPE.