CISD2 promotes lung squamous carcinoma cell migration and invasion via the TGF-ß1-induced Smad2/3 signaling pathway.

BACKGROUND: Although aberrant expression of CDGSH iron sulfur domain 2 (CISD2) contributes to the tumorigenesis and progression of numerous human cancers, the biological function of CISD2 and its specific prognostic value in lung squamous cell carcinoma (LUSC) have yet to be comprehensively explored. The current study aimed to elucidate the role of CISD2 in LUSC as well as the underlying molecular mechanisms. METHODS: Immunohistochemistry was conducted to detect the protein expression of CISD2 and analyze whether high expression of CISD2 affects the overall survival (OS) of LUSC patients. Cell proliferation, colony formation, wound healing and Transwell invasion assays were performed to clarify whether CISD2 contributes to LUSC cell proliferation and disease progression. Quantitative real-time reverse transcription-PCR and western blot assays were used to detect the levels of transcription factors and key epithelial-mesenchymal transition (EMT)-related markers in LUSC cells after CISD2 knockdown and overexpression to determine whether CISD2 regulates transforming growth factor-beta (TGF-ß)-induced EMT in LUSC. RESULTS: Immunohistochemistry of human tissue microarrays containing 90 pairs of adjacent and cancerous tissues revealed that CISD2 is considerably overexpressed in LUSC and strongly linked to poor OS. Functional experiments suggested that silencing endogenous CISD2 inhibited the growth, colony formation, migration, and invasion of H2170 and H226 cell lines. Exogenous overexpression of CISD2 facilitated these phenotypes in SK-MES-1 and H2170 cells. Furthermore, CISD2 promoted EMT progression by increasing the expression of mesenchymal markers (N-cadherin, vimentin, Snail, and Slug) as well as SMAD2/3 and reducing the expression of the epithelial marker E-cadherin. Mechanistically, our studies provide the first evidence that CISD2 can promote EMT by enhancing TGF-ß1-induced Smad2/3 expression in LUSC cells. CONCLUSION: In conclusion, our research illustrates that CISD2 is highly expressed in LUSC and may facilitate LUSC proliferation and metastasis. Thus, CISD2 may serve as an independent prognostic marker and possible treatment target for LUSC.

The second-generation tyrosine kinase inhibitor afatinib inhibits IL-1ß secretion via blocking assembly of NLRP3 inflammasome independent of epidermal growth factor receptor signaling in macrophage.

Chronic inflammation might lead to many malignancies, and inadequate resolution could play a crucial role in tumor invasion, progression and metastases. Afatinib is a second-generation tyrosine kinase inhibitor targeting epidermal growth factor receptor in non-small cell lung cancer. Few studies showed the correlation of afatinib and the innate immune system especially macrophage. Our study showed that afatinib could block the activation of NLRP3 inflammasome in a dose-dependent manner in macrophage, and that afatinib could prevent the assembly of NLRP3 inflammasome. Besides, afatinib could inhibit NLRP3 inflammasome activation independent of EGFR signaling. Moreover, afatinib was able to alleviate the LPS-induced sepsis in vivo. These investigations provide significant experimental evidence in afatinib as therapeutic drug for non-small cell lung cancer or other tumors and NLRP3-related diseases, and explore new target for afatinib in macrophage.

Integrated Analysis of Genomic and Immunological Features in Lung Adenocarcinoma With Micropapillary Component.

BACKGROUND: Micropapillary adenocarcinoma is one of the most aggressive histologic subtypes of lung adenocarcinoma (LADC), and even a minor proportion of micropapillary component (MPC) within the LADC could contribute to poor prognosis. Comprehensive analysis of genetic and immunological features of LADC with different percentages of MPC would help better understand cancer biology of this LADC subtype and direct future treatments. METHODS: We performed next-generation sequencing (NGS) for a discovery cohort of 43 LADC patients whose tumors were micro-dissected to separate MPC and non-MPC lesions and a reference cohort of 113 LADC patients. MPC-enriched genetic alterations that were detected in the discovery cohort were then confirmed using a validation cohort of 183 LADC patients. Immunological staining was also conducted on the MPC-containing samples in the discovery cohort. RESULTS: Tumors with a higher percentage of MPC tended to harbor more tumor mutation burdens (TMBs) and chromosome instability (CIN). Some rare genetic events may serve as the genetic landscape to drive micropapillary tumor progression. Specifically, alterations in transcription termination factor 1 (TTF1), brain-specific angiogenesis inhibitor 3 (BAI3), mammalian target of rapamycin (MTOR), and cyclin-dependent kinase inhibitor 2A (CDKN2A) were cross-validated to be enriched in MPC-contained LADC. Additionally, tumors with a higher percentage of MPC were associated with a higher percentage of CD4+, CD8+, and PD-L1+ staining, and some genetic changes that were enriched in MPC, including MET amplification and MTOR mutation, were correlated with increased PD-L1 expression. CONCLUSION: We identified multiple novel MPC-enriched genetic changes that could help us understand the nature of this aggressive cancer subtype. High MPC tumors tended to have elevated levels of TMBs, T cell infiltration, and immunosuppression than low MPC tumors, implying the potential link between MPC content and sensitivity to immunotherapy.

Thoracic radiotherapy and concurrent almonertinib for unresectable stage III EGFR-mutated non-small-cell lung cancer: a phase 2 study.

BACKGROUND: Concurrent chemo-radiotherapy remains the standard treatment in unresectable stage III non-small-cell lung cancer (NSCLC) patients. Several studies have shown a potential value of concurrent epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) with thoracic radiotherapy in EGFR-mutated population, but a high risk of radiation pneumonitis raised a major concern. This study intends to explore the safety and efficacy of concurrent almonertinib, a new third-generation EGFR-TKI, with radiotherapy in locally advanced EGFR-mutated NSCLC patients. METHODS: Locally advanced NSCLC patients harboring sensitive EGFR mutation will be included in this study. A radiotherapy plan will be made for each patient before treatment, and the lung V20 will be calculated. Patients with lung V20 ≥ 28% were enrolled in induction group (arm A), which almonertinib was given for 2 months followed by concurrent radiotherapy; patients with lung V20 < 28% were enrolled in concurrent group (arm B), which almonertinib was given concurrent with thoracic radiotherapy. The primary endpoint is the incidence of grade ≥ 3 radiation pneumonitis within 6 months post-radiotherapy, and the secondary endpoints are local control rate, progression-free survival, and overall survival. DISCUSSION: The safety and efficacy of third-generation EGFR-TKI concurrent with thoracic radiotherapy in locally advanced EGFR-mutated NSCLC is still unknown. We propose to conduct this phase 2 study evaluating the safety especially the radiation pneumonitis within 6 months post-radiotherapy. This trial protocol has been approved by the Ethics committee of Hangzhou cancer hospital. The ethics number is HZCH-2020-030. TRIAL REGISTRATION: clinicaltrials.gov, NCT04636593 . Registered 19 November 2020 - Retrospectively registered.

Inhibition of histamine receptor H3 suppresses the growth and metastasis of human non-small cell lung cancer cells via inhibiting PI3K/Akt/mTOR and MEK/ERK signaling pathways and blocking EMT.

Recent evidence shows that the expression levels of histamine receptor H3 (Hrh3) are upregulated in several types of cancer. However, the role of Hrh3 in non-small cell lung cancer (NSCLC) has not been elucidated. In the present study, we showed that the expression levels of Hrh3 were significantly increased in NSCLC samples, and high levels of Hrh3 were associated with poor overall survival (OS) in NSCLC patients. In five human NSCLC cell lines tested, Hrh3 was significantly upregulated. In NSCLC cell lines H1975, H460, and A549, Hrh3 antagonist ciproxifan (CPX, 10-80 µM) exerted moderate and concentration-dependent inhibition on the cell growth and induced apoptosis, whereas its agonist RAMH (80 µM) reversed these effects. Furthermore, inhibition of Hrh3 by CPX or siRNA retarded the migration and invasion of NSCLC cells through inhibiting epithelial-mesenchymal transition (EMT) progression via reducing the phosphorylation of PI3K/Akt/mTOR and MEK/ERK signaling pathways. In nude mice bearing H1975 cell xenograft or A549 cell xenograft, administration of CPX (3 mg/kg every other day, intraperitoneal) significantly inhibited the tumor growth with increased E-cadherin and ZO-1 expression and decreased Fibronectin expression in tumor tissue. In conclusion, this study reveals that Hrh3 plays an important role in the growth and metastasis of NSCLC; it might be a potential therapeutic target against the lung cancer.

[Pathological and Molecular Features of Lung Micropapillary Adenocarcinoma].

Lung micropapillary adenocarcinoma is characterized by frequent metastasis, lymph node infiltration, high recurrence rate and low overall survival rate as a high-grade lung adenocarcinoma. Special oncogenic pathway is activated and immune microenvironment is established in this subtype of tumor. This article reviews the Pathological phenomena and molecular features of micropapillary adenocarcinoma studied in recent years, aiming to deepen the understanding of micropapillary lesions and lay the foundation for formulating specific treatment strategies.
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Identification of somatic copy number variations in plasma cell free DNA correlating with intrinsic resistances to EGFR targeted therapy in T790M negative non-small cell lung cancer.

BACKGROUND: About 20-30% EGFR-mutant non-small lung cancer show intrinsic resistance to EGFR targeted therapies. Compared to T790M positive in acquired resistance patients, little is known about EGFR-TKI intrinsic resistance for T790M negative patients. METHODS: Thirty-one patients with advanced stage lung cancer, including 18 patients with intrinsic resistance (PFS <6 months) and 13 patients with acquired resistance (PFS >36 months) but are negative for plasma T790M were recruited in the study. Plasma cell free DNA was profiled by low coverage whole genome sequencing with median genome coverage of 1.86X by Illumina X10. Sequencing coverage across chromosomes was summarized by samtools, and normalized by segmentation analysis as provided by R package 'DNACopy'. RESULTS: The most frequent chromosomal changes were found on chr7, chr1 and chr8. Among them, chr7p gains were found in 12 (66.7%) intrinsic resistance and 4 (30.7%) acquired resistance patients. The gene EGFR was found located on the focal amplification peak of chr7p. The performance of 7p gain to predict intrinsic resistance reaches AUC =0.902. Similarly, focal amplifications were also found on chromosome 5, 16 and 22, where tumor related gene PCDHA@, ADAMTS18 and CRKL were located. Focal deletions were also found in chr1, 8, 10 and 16, where genes SFTPA1/2, DLC1, PTEN and CDH1 are located. CONCLUSIONS: The results suggest cell free DNA copy number might be a useful peripheral blood tumor biomarker for predicting intrinsic resistance of EGFR targeted therapy and prognosis.

Chalcomoracin inhibits cell proliferation and increases sensitivity to radiotherapy in human non-small cell lung cancer cells via inducing endoplasmic reticulum stress-mediated paraptosis.

Chalcomoracin (CMR) is a kind of Diels-Alder adduct extracted from the mulberry leaves. Recent studies showed that CMR has a broad spectrum of anticancer activities and induces paraptosis in breast cancer and prostate cancer cells. In this study, we investigated the effects of CMR against human non-small cell lung cancer cells and the underlying mechanisms. We found that CMR dose-dependently inhibited the proliferation of human lung cancer H460, A549 and PC-9 cells. Furthermore, exposure to low and median doses of CMR induced paraptosis but not apoptosis, which was presented as the formation of extensive cytoplasmic vacuolation with increased expression of endoplasmic reticulum stress markers, Bip and Chop, as well as activation of MAPK pathway in the lung cancer cells. Knockdown of Bip with siRNA not only reduced the cell-killing effect of CMR, but also decreased the percentage of cytoplasmic vacuoles in H460 cells. Moreover, CMR also increased the sensitivity of lung cancer cells to radiotherapy through enhanced endoplasmic reticulum stress. In lung cancer H460 cell xenograft nude mice, combined treatment of CMR and radiation caused greatly enhanced tumor growth inhibition with upregulation of endoplasmic reticulum stress proteins and activation of pErk in xenograft tumor tissue. These data demonstrate that the anticancer activity and radiosensitization effect of CMR result from inducing paraptosis, suggesting that CMR could be considered as a potential anticancer agent and radiation sensitizer in the future cancer therapeutics.

Genome-wide profiling reveals alternative polyadenylation of mRNA in human non-small cell lung cancer.

BACKGROUND: Lung cancer is the second most common cancer with an extremely poor overall survival rate. Post-transcriptional regulation of gene expression play many important roles in human cancer, and one of the potential mechanisms underlying this is alternative mRNA maturation at its 3' untranslated regions (3'-UTRs). METHODS: Cancer tissues and paired adjacent normal lung tissues from 26 patients diagnosed with non-small cell lung cancer (NSCLC) were analyzed by in vitro transcription-sequencing alternative polyadenylation sites (IVT-SAPAS). 41,773,101 reads in average were obtained from each paired sample. A potential regulation of Cleavage Stimulation Factor Subunit 2 (CSTF2) on 3'UTR length of genes was tested in H460 cells. RESULTS: 1439 (10.26%) genes showed up-regulated expression and 1364 (9.72%) genes showed down-regulated expression in lung cancer tissue versus normal lung tissue, and shorten 3'UTR in cancer tissue was detected in cancer tissues collected from 96.2% (25/26) patients, indicating lung cancer tend to have shortened 3'UTRs of these identified genes. KEGG analysis showed 1855 genes with shorten 3'UTR were enriched in mTOR signaling, ubiquitin mediated proteolysis and RNA degradation. Knocking down CSTF2 expression in H460 cells results in 3'UTR elongation of genes that was identified to be with shortened length in cancer tissues. CONCLUSION: Alternative polyadenylation (APA) site-switching of 3'UTRs is prevalent in NSCLC, and CSTF2 may serve as an oncogene regulates the 3'UTR length of cancer related genes in NSCLC.