ObjectiveTo construct and evaluate a mouse model that simulates the pathological progression of immune checkpoint inhibitor-associated pneumonitis (CIP), in order to address the critical issue of the lack of standardized animal models in this research field.MethodsThirty-five SPF-grade 6-week-old male C57BL/6J mice were randomly assigned to a control group (n=5), a bleomycin (BLM) group (n=15), and a BLM+programmed death-1 (PD-1) monoclonal antibody (BLM+PD-1) group (n=15). Corresponding drugs were administered via intraperitoneal injection for intervention. The general condition, body weight changes, survival rate, and pulmonary histopathological alterations were systematically monitored.ResultsCompared with the BLM group, the BLM+PD-1 group exhibited significantly more severe pulmonary fibrosis, characterized by progressive body weight loss and more severe deterioration of general conditions. Histopathological examination showed that pulmonary inflammation was alleviated 4 weeks post-modeling in the BLM group. In contrast, pulmonary lesions continued to worsen in the BLM+PD-1 group, with no evidence of resolution of inflammation 28 days post-modeling. Quantitative immunohistochemical analysis revealed that the positive expression rate of interleukin-17A (IL-17A) in lung tissues reached 34.28% in the BLM+PD-1 group 14 days post-modeling, which was markedly higher than that in the BLM group (24.02%, P < 0.000 1). The positive expression rate of α-smooth muscle actin (α-SMA) was 12.40% in the BLM+PD-1 group 14 days post-modeling, nearly twice the level in the BLM group (5.87%). 28 days post-modeling, the positive expression rate of α-SMA remained at 7.72% in the BLM+PD-1 group, while it declined to 3.38% in the BLM group.ConclusionThe addition of a PD-1 inhibitor to bleomycin treatment accelerates the progression of pulmonary interstitial fibrosis and effectively recapitulates the pathological process of lung injury and fibrosis driven by aberrant immune activation during clinical immunotherapy. Accordingly, this study successfully establishes a dynamic CIP animal model recapitulating the progression from early inflammatory response to pulmonary fibrosis. This model addresses the critical shortage of animal models for this condition and offers an experimental platform for further mechanistic studies and the development of therapeutic strategies.

Based on the high mortality rate and the limitations of current therapeutic strategies for sepsis,a critical illness,this study innovatively explored the potential application value of nano-formulations based on Polygonati Rhizoma polysaccharides in sepsis prevention.Initially,crude polysaccharides were extracted from the Polygonati Rhizoma by using the conventional water extraction and ethanol precipitation method,followed by a qualitative analysis for their main saccharide components via high-performance liquid chromatography,thereby determining their polysaccharide structures.Subsequently,Polygonati Rhizoma polysaccharides nanoparticles(PSPs NPs)were successfully fabricated through a combination of controlled caramelization reaction and low-temperature grinding technology.The resulting nanoparticles exhibited uniform particle sizes,good dispersibility,and excellent stability,with an average particle size of approximately 150 nm.The morphology,particle sizes,and surface properties of the nanoparticles were systematically characterized by dynamic light scattering and transmission electron microscopy.In vitro study results demonstrate that Cy3 labeled PSPs NPs were efficiently endocytosed by RAW264.7 macrophages,with a significantly higher uptake efficiency than the free polysaccharides.Moreover,the CCK-8 assay demonstrates that PSPs NPs significantly enhance the viability of human monocytic THP-1 cells in a dose-dependent manner,indicating that nano-formulation effectively enhances the bioactivity and immunomodulatory potential of PSPs.In vivo,a mouse model of sepsis induced by intraperitoneal infection of Escherichia coli was established.The results show that prophylactic administration of PSPs NPs significantly improves the 48 h survival rates of mice,attenuates sepsis-associated body weight loss,and exerts significant protective effects against inflammatory infiltration and tissue damage in critical organs including the lung,liver,and kidney,as evidenced by HE staining.Collectively,this study not only establishes a safe and effective nano-delivery system for TCM polysaccharides but also verifies its multi-target and multi-organ protective effects in the early intervention of sepsis by enhancing innate immune responses and regulating the inflammatory reaction pathways.These findings provide a robust theoretical basis and practical pathway for the modern formulation development of active components from TCM and the development of prevention and treatment strategies for sepsis.

Objectives To conduct an umbrella review to extensively evaluate and summarise the evidence regarding the relationship between risk factors and the occurrence of brain metastasis in lung cancer.Design Umbrella review of systematic reviews and meta-analyses.Data sources Four databases (PubMed, EMBASE, Web of Science and Cochrane Library) were searched from inception to 10 November 2024.Eligibility criteria Systematic reviews and meta-analyses that assessed the relationship between risk factors and brain metastasis in lung cancer were included. Only English language studies were considered.Data extraction and synthesis Two authors independently extracted data and assessed the methodological quality and risk of bias of the included studies. Certainty of evidence was evaluated and summarised for each identified risk factor.Results Six systematic reviews/meta-analyses were included. The quality of these studies varied, with most having low or critically low methodological quality. Epidermal growth factor receptor mutations, female gender, lung adenocarcinoma and advanced tumour stage were associated with an increased risk of brain metastasis. Prophylactic cranial irradiation and older age were associated with reduced risk.Conclusions This umbrella review suggests that several risk factors may be associated with brain metastasis in lung cancer, but the overall quality of evidence is low. Future studies with improved methodologies are needed to validate these findings.PROSPERO registration number CRD42023484563

Hepatocellular carcinoma (HCC) is the most prevalent form of primary liver cancer, associated with high morbidity and mortality worldwide. Despite advancements in diagnostic methods and systemic treatments, including tyrosine kinase inhibitors (TKIs) and immune checkpoint inhibitors (ICIs), the development of drug resistance remains a significant challenge in HCC management. Traditional treatments such as surgical resection and transarterial chemoembolization offer limited efficacy, especially in advanced stages. Although novel therapies like lenvatinib, sorafenib, regorafenib, and ICIs have shown promise, their effectiveness is often hindered by primary and acquired resistance, leading to poor long-term survival outcomes. This review focuses on the molecular mechanisms underlying resistance to targeted therapies and immunotherapies in HCC. Key factors contributing to resistance include alterations in the tumor microenvironment (TME), immune evasion, hypoxia, changes in cellular metabolism, and genetic mutations. Additionally, molecular players such as ferroptosis, autophagy, apoptosis, endoplasmic reticulum stress, ABC transporters, and non-coding RNAs(ncRNAs) are discussed as contributors to drug resistance. Understanding these mechanisms is critical for the development of novel therapeutic strategies aimed at overcoming resistance, improving patient outcomes, and ultimately enhancing survival rates in HCC patients.

Background Fuzheng Kang'ai Formula (FZKA) is a clinically effective traditional Chinese medicine compound with demonstrated efficacy against non-small cell lung cancer (NSCLC). This study systematically investigated its anti-tumor mechanisms, focusing on the interplay between autophagy and ferroptosis. Purpose We aimed to determine whether FZKA suppresses NSCLC through autophagy-ferroptosis crosstalk and to identify key epigenetic regulators involved in this process. Study Design The study combined in vitro assays using NSCLC cell lines with in vivo validation via subcutaneous xenograft models. Methods : An Agilent 6546 LC/Q-TOF liquid chromatography-mass spectrometry system was used to detect the stability of component contents in FZKA across different batches. Autophagy was assessed through mRFP-GFP-LC3 adenovirus assay, transmission electron microscopy, and LC3B-II/I conversion. Ferroptosis was evaluated by measuring lipid peroxidation and intracellular Fe²⁺ levels. Crosstalk was examined using autophagy and ferroptosis inhibitors to monitor reciprocal effects on SLC7A11 and p-mTOR. EZH2 was investigated as an upstream regulator of mTOR and SLC7A11. Protein and mRNA expression were analyzed by Western blot and qRT-PCR. EZH2-overexpressing cells were established to evaluate functional rescue, and lysine methylation label-free quantitative proteomics was employed to identify downstream effectors. A nude mouse xenograft model was used for in vivo validation, with tumor weight and volume measured, and key proteins analyzed. Results The composition of FZKA remains stable across different batches. FZKA induced autophagy via mTOR suppression and Beclin1 upregulation, while promoting ferroptosis through SLC7A11 and GPX4 inhibition. Functional crosstalk was confirmed through inhibitor experiments showing mTOR and SLC7A11 as key mediators. FZKA significantly reduced EZH2 expression, and EZH2 overexpression partially reversed the effects of FZKA on mTOR and SLC7A11. Proteomic analysis revealed DOT1L/H3K79me2 as a downstream pathway of FZKA-mediated EZH2 regulation. DOT1L expression correlated with EZH2 levels, and EZH2 overexpression rescued DOT1L/H3K79me2 suppression. These results were consistently verified in vivo. Conclusion FZKA suppresses NSCLC by regulating the EZH2/DOT1L/H3K79me2 signaling to induce crosstalk between mTOR-mediated autophagy and SLC7A11-dependent ferroptosis, leading to extensive cell death. This study reveals a novel epigenetic mechanism coordinating two cell death pathways and supports FZKA's clinical application.

A variety of factors, such as dietary habits, the external environment, and individual genetic differences, can lead to the development of cancer. While chemotherapy and radiotherapy are commonly used for cancer treatment, drug resistance and side effects are prevalent issues. Therefore, there is an urgent need to find new treatment modalities. Studies have shown that radiotherapy and chemotherapy can lead to a significant increase in apoptotic cells (ACs) within the tumor microenvironment (TME). The process of phagocytosis helps maintain homeostasis by engulfing and removing these ACs from the organism promptly, which is referred to as efferocytosis. However, it has been observed that excessive efferocytosis can promote the formation of an immunosuppressive TME, which is detrimental to tumor therapy. Thus, inhibiting efferocytosis to enhance the formation of an immune microenvironment shows promise as a treatment direction for tumors. As researchers gradually uncover the molecular mechanisms of efferocytosis, various small-molecule inhibitors and monoclonal antibodies are actively being assessed in clinical trials. Targeting efferocytosis is anticipated to emerge as a promising approach in tumor treatment. In this review, the intricate steps involved in efferocytosis are explored and the current drugs that targeting this process for cancer treatment are outlined.

BACKGROUND:Polyphyllin I (PPI), the primary active component extracted from the commonly utilized drug Paris polyphylla, in clinical applications, exhibits exceptional anti-tumor activity. Therefore, the objective of this study is to delve into its antitumor properties by promoting ferroptosis and enhancing anti-tumor immunity. RESULTS:Our result demonstrate that PPI effectively suppresses the proliferation of NSCLC cells and triggers their demise through the induction of ferroptosis. Furthermore, the degradation of iron via autophagy is pivotal in mediating PPI-induced ferroptosis in NSCLC. Notably, EZH2 emerges as a potential key target of PPI, and its overexpression can significantly attenuate the therapeutic efficacy of PPI. Lastly, these mechanistic insights and pathways were corroborated in animal model, and we preliminarily revealing a stimulatory role of PPI in bolstering anti-tumor immunity. CONCLUSION:PPI triggers ferroptosis in NSCLC by downregulating EZH2, promoting NCOA4/Ferritin mediated ferritinophagy, and enhances anti-tumor immunity by promoting CD8 +T infiltration into tumor tissue.

Tumor-associated macrophages (TAMs) are crucial in hepatocellular carcinoma (HCC) development and invasion. This study explores monocyte/ macrophage-associated gene expression profiles in HCC, constructs a prognostic model based on these genes, and examines its relationship with drug resistance and immune therapy responses. Single-cell RNA sequencing(scRNA-seq) data from 10 HCC tissue biopsy samples, totaling 24,597 cells, were obtained from the GEO database to identify monocyte/macrophage-associated genes. A prognostic model was constructed and validated using external datasets and Western blot. Relationships between the model, clinical correlates, drug sensitivity, and immune therapy responses were investigated. From scRNA-seq data, 2,799 monocyte/macrophage marker genes were identified. Using the TCGA dataset, a prognostic model based on the single-gene UQCRH was constructed, stratifying patients into high-risk and low-risk groups based on overall survival rates. High-risk group patients showed reduced survival rates and higher UQCRH expression in tumor tissues. Western blot analysis further confirmed the elevated expression of UQCRH in HCC cell lines. Spatial transcriptomics analysis revealed that high UQCRH expression co-localized with malignant cells in the tumor tissue. Drug sensitivity analysis revealed that the high-risk group had lower sensitivity to sorafenib and axitinib. Immune therapy response analysis indicated poorer outcomes in the high-risk group, with more pronounced APC inhibition and a weaker IFN-II response. Clinical indicator analysis showed a positive correlation between high UQCRH expression and tumor invasion. Enrichment analysis of UQCRH and associated molecules indicated involvement in oxidative phosphorylation and mitochondrial electron transport. This study introduces a prognostic model for HCC patients based on monocyte/macrophage marker genes. The single-gene model predicts HCC patient survival and treatment outcomes, identifying high-risk individuals with varying drug sensitivities and immune suppression states.

Lung cancer remains one of the most prevalent and lethal malignancies worldwide, responsible for nearly 1.8 million deaths annually, which accounts for approximately 18.7

Moutan cortex has demonstrated antitumor properties attributed to its bioactive compound Paeoniflorigenone (PA). Nevertheless, there is limited research on the efficacy of PA in the prevention and treatment of hepatocellular carcinoma (HCC). We aimed to investigate the potential pharmacological mechanisms of PA in the treatment of Aflatoxin B1 (AFB1)-induced hepatocarcinogenesis using network pharmacology and bioinformatics analysis approaches. Through various databases and bioinformatics analysis approaches, 34 shared targets were identified as potential candidate genes for PA in fighting liver cancer caused by AFB1. Pathway analysis revealed involvement in cell cycle, HIF-1, and Rap1 pathways. A risk assessment model was developed using LASSO regression, showing an association between the identified genes and the tumor immune microenvironment. The genes within the risk model were found to be linked to the immune response in liver cancer. Molecular docking studies indicated that PA interacts with its targets through hydrogen bonding and hydrophobic interactions. This study provides insights into the possible mechanisms of PA in liver cancer treatment and offers a predictive model for assessing the risk level of individuals with liver cancer. These findings have significant implications for the therapeutic strategies in managing liver cancer patients.

OBJECTIVE:The purpose of this overview is to assess systematic reviews (SRs)/ meta-analyses (MAs) of Huachansu (HCS) combination chemotherapy for treating non-small cell lung cancer (NSCLC) and provide summarized evidence for clinical decision making.METHODS:From the creation of the database to JUNE 2023, 8 databases in English and Chinese were searched. SRs/MAs that met the inclusion and exclusion criteria were included. Two reviewers independently screened research, extracted data and assessed methodological quality, risk of bias, report quality and evidence quality by using relevant criteria from AMSTAR-2, ROBIS scale, PRISMA, and GRADE system.RESULTS:The short-term effect, long-term effect, quality of life improvement, safety and pain relief effect in 8 included SRs/MAs were assessed in this overview according to quantitative synthesis. Results assessed by AMSTAR-2, PRISMA, and ROBIS were generally unsatisfactory, with the results of the AMSTAR-2 assessment showing that all of them were of low or critically low quality; the number of items in the included research that were fully reported (compliance was 100%) by the PRISMA checklist was only 50%, while there were 38.10% of the research reporting less than 60% completeness; the ROBIS assessment showed a small number of systems to be low risk of bias. In addition, 26 items were rated as moderate quality, while 50.94% of items were rated as low or critically low quality by GRADE.CONCLUSION:HCS may be a promising adjuvant therapy for NSCLC. However, high-quality SRs/MAs and randomized control trials (RCTs) should be conducted to provide sufficient evidence so as to draw a definitive conclusion.

Ethnopharmacological relevance: Fuzheng Kang-Ai (FZKA) decoction is mainly composed of 12 components with different types of herbs. In the last decade, FZKA has been used as an adjuvant treatment for lung cancer in clinical practice. Our previous studies have confirmed that FZKA shows a strong anti-cancer activity, significantly increases the clinical efficacy of gefitinib and reverses gefitinib resistance in non-small cell lung cancer (NSCLC). However, the molecular mechanism still needs to be further elucidated. Aim of the study: The aim of this study was to investigate the role and mechanism by which FZKA inhibited the cell growth, proliferation and invasion of lung adenocarcinoma(LUAD) and reversed the acquired resistance of gefitinib for the therapy in LUAD. Materials and methods: Cell viability assay and EDU assay were used for detecting of cell viability and cell proliferation. Transwell assay was performed to measure cell invasion. Western Blot and qRT-PCR were used for protein and gene expression test. The gene promoter activity was determined by dul-luciferase reporter assay. The in situ expression of protein was measured by cell immunofluorescence. Stabilized cell lines were established for stable overexpression of EZH2. Transient transfection assay was used for gene silence and overexpression. Xenograft tumors and bioluminescent imaging were used for in vivo experiments. Results: FZKA significantly inhibited the cell viability, proliferation and cell invasion of LUAD, the combination of FZKA and gefitinib had a great synergy on the above processes. Moreover, FZKA significantly decreased EZH2 mRNA and protein expression, FZKA reversed the resistance of gefitinib by down-regulation of EZH2 protein. ERK1/2 kinase mediated the down-regulation of EZH2 reduced by FZKA. In addition, FZKA decreased the expression of Snail and EGFR by decreasing EZH2. Overexpression of Snail and EGFR significantly reversed the effect of FZKA-inhibited cell invasion and cell proliferation. More important, the combination of FZKA and gefitinib enhanced the inhibitory effect on EZH2, Snail and EGFR proteins. Furthermore, the growth inhibition and reversal of gefitinib resistance induced by FZKA were further validated in vivo. Finally, the expression and clinical correlation of EZH2,EGFR and Snail in cancer patients were further validated using bioinformatics analysis. Conclusions: FZKA significantly suppressed tumor progression and reversed gefitinib resistance by regulating the p-ERK1/2-EZH2-Snail/EGFR signaling pathway in LUAD.

Enhancer of zeste homolog 2 (EZH2), an epigenetic regulator that primarily inhibits downstream gene expression by tri-methylating histone H3, which is usually overexpressed in tumors and participates in many processes such as tumor occurrence and development, invasion, migration, drug resistance, and anti-tumor immunity as an oncogene, making it an important biomarker in cancer therapy. Collectively, several transcription factors and RNAs cooperate to facilitate the elevated expression of EZH2 in cancer. Although the significance of blocking EZH2 in cancer for inhibiting cancer progression is widely recognized, the clinical application of EZH2 inhibitors continues to encounter numerous challenges. In this review, drawing upon our comprehensive understanding of the factual underpinnings of EZH2's role in cancer, we aim to clarify the crucial importance of targeting EZH2 in cancer treatment. Furthermore, we summarize the current research landscape surrounding targeted EZH2 inhibitors and offer insights into potential future applications of these inhibitors.

Supplementary Figure 1: MiRs are expressed differently in CNE-2 and CNE-2R cells. Supplementary Figure 2: The relative percentage of methylation level between CNE-2 and CNE- 2R by cytosine-extension assay. Supplementary Figure 3: Genomic DNA methylation is altered between CNE-2 and CNE-2R cells. Supplementary Figure 4: MiR-24 promoters's CpG islands distribution. Supplementary Table 1: Information of 6 pairs NPC primary and recurrent patients. Supplementary Table 2: Radioresistant miRs signature between CNE-2 and CNE-2R. Supplementary Table 3: The numbers of genes altered on methylation between CNE-2 and CNE-2R.

BACKGROUND:Conflicting results about the effect of concomitant medications on immunotherapy in non-small cell lung cancer (NSCLC) were reported by many meta-analyses (MAs), and the certainty of evidence linking concomitant medications with immunotherapy efficacy has not been quantified, which may cause some evidence to be misinterpreted. METHODS:Four databases including Embase, Cochrane Library, PubMed, and Web of Science were searched from inception to January 2023 in English. Based on prospective or retrospective clinical controlled trials including immunotherapy with concomitant medications or not in NSCLC, quantitative MAs reporting the efficacy of immunotherapy with binary direct comparison and enough extractable data were collected. The methodological quality, reporting quality, and risk of bias of included MAs were evaluated respectively. New meta-analyses were conducted and their evidence certainty was classified as nonsignificant, weak, suggestive, highly suggestive, or convincing. RESULTS:Fifteen MAs with 5 medications were included. After being assessed by AMSTAR-2, PRISMA, and ROBIS, the major shortcomings were focused on the registration of protocol, literature retrieval or data extraction, implementation of sensitivity analysis or evidence certainty assessment, and incomplete reporting in the section of method and result. New pooled analyses indicated that antibiotics (HR = 1.545[1.318-1.811]), steroids (HR = 1.784[1.520-2.093]), proton pump inhibitors (PPIs) (HR = 1.303[1.048-1.621]) and opioids (HR = 1.910[1.213-3.006]) could shorten overall survival (OS) in patients with NSCLC receiving immunotherapy. Besides, antibiotics (HR = 1.285[1.129-1.462]) and steroids (HR = 1.613[1.315-1.979]) were harmful to progression-free survival (PFS) in these patients significantly. No negative effect was found in nonsteroidal anti-inflammatory drugs and the objective response rate of all medications. High-level evidence suggested that using PPIs before or after the initiation of immunotherapy and using steroids during the first-course immunotherapy could weaken the OS of patients with NSCLC. Meanwhile, the negative effects of antibiotics and opioids on OS or PFS were only supported by moderate or low-level evidence. CONCLUSIONS:The concurrent usage of PPIs or steroids adversely affects the survival of patients with NSCLC receiving immunotherapy. Future investigations are required to ascertain whether these adverse effects are primarily attributed to the comorbidities or the concurrent medications.

Abstract Background: Fuzheng Kang-Ai decoction (FZKA), which consists of twelve Chinese herbal medicines (CHM), has been extensively used in the treatment of Non-Small Cell Lung Cancer (NSCLC) patients. It was reported in our preceding study that the progression-free survival (PFS) could be prolonged and the toxic effect was reduced after the combination of FZKA and Gefitinib (GFTN) comparing to GFTN alone. Recent studies have shown that FZKA could promote ferroptosis in NSCLC cells. Hence, we assumed that FZKA might strengthen the effect of GFTN by regulating cell ferroptosis in NSCLC. Results: First of all, according to our results, FZKA increased the inhibition effect of NSCLC by GFTN in vitro. Next, we found that FZKA increased the sensitivity of GFTN on cell ferroptosis by testing lipid peroxidation and intracellular-free iron. Interestingly, the effect of FZKA on increasing sensitivity of GFTN in NSCLC cells could be reversed by blocking ferroptosis. Additionally, the results demonstrated that the expression of AMPKα was increased and SLC7A11 reduced in the combined treatment group, at both mRNA and protein levels. Notably, the inhibition of AMPKα reversed the effect of FZKA on increasing GFTN sensitivity. Finally, in vivoexperiments validated that FZKA strengthened the effect of GFTN by inducing NSCLC cell ferroptosis. Conclusions: Based on our findings, we conclude that FZKA increases the sensitivity of GFTN via inducing NSCLC ferroptosis through AMPK/SLC7A11 pathway, providing the solid evidences to the treatment of FZKA together with GFTN in NSCLC patients.

Lung cancer is the leading cause of cancer-related deaths worldwide. Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) showed great therapeutic efficacy for non-small cell lung cancer (NSCLC) patients. However, acquired resistance severely limits the clinical application and efficacy of EGFR-TKIs. In the current study, we found that solamargine (SM), a natural alkaloid derived from the fruit of Lycium tomato lobelia, has been found to inhibit the progression of NSCLC and enhance the anticancer effect of EGFR-TKIs. In brief, SM significantly inhibited the cell viability of NSCLC cells and enhanced the anticancer effect of gefitinib (GFTN) and erlotinib (ERL). Mechanistically, SM decreased the expression of MALAT1 and induced miR-141-3p, whereas reduced SP1 protein levels. Interestingly, both MALAT1 and Sp1 have classical and conservative binding sites of miR-141-3p in their 3'-UTR regions. Silence of MALAT1 and overexpression of miR-141-3p both decreased the protein expression of Sp1. Subsequently, promoter activity and protein expression of IGFBP1 were upregulated by SM, which was not observed in cells with SP1 overexpression. Moreover, the inhibitory effect of SM on cell growth was significantly blocked by knockdown of IGFBP1 expression. More importantly, the combination of SM and GFTN synergistically inhibited the progression of lung cancer. Similar results were observed in experiments in vivo. Finally, the clinical relevance of MALAT1, Sp1 and IGFBP1 was further validated using bioinformatics analysis. Taken together, we confirmed that SM significantly enhanced the anticancer effect of EGFR-TKIs by regulating the MALAT1/miR-141-3p/Sp1/IGFBP1 signaling pathway. This study unravels a novel mechanism and suggests a new potential NSCLC-associated therapy.

目的 研究hsa-miR-93在鼻咽癌放疗抗拒过程中的作用及分子机制.方法 以人鼻咽癌细胞株CNE-1、CNE-2、CNE-2R、HONE1、C666.1以及鼻咽上皮永生细胞NP460为研究对象,采用实时荧光定量PCR(qRT-PCR)法检测各组细胞经放疗后hsa-miR-93的表达;采用Western blot法检测各组细胞经放疗后STAT3蛋白的表达;通过miRWalk软件和RNAHybrid软件预测hsa-miR-93和STAT3的结合作用及结合位点;通过双荧光素酶报告基因法检测hsa-miR-93对STAT3靶向结合情况;瞬时转染小分子模拟物或抑制剂调控hsa-miR-93的表达后,采用qRT-PCR和Western blot法检测STAT3的表达;转染siSTAT3沉默STAT3的表达后,通过细胞集落形成实验检测鼻咽癌细胞经放疗后的集落形成能力.结果 与CNE-1、CNE-2细胞(相对放疗敏感)相比,hsa-miR-93在HONE1、CNE-2R细胞(相对放疗抗拒)中表达降低(P<0.001),且各组鼻咽癌细胞经放疗后hsa-miR-93的表达呈剂量及时间依赖性降低(P<0.05);与CNE-1、CNE-2细胞相比,STAT3在HONE1、CNE-2R细胞中表达升高,且各组鼻咽癌细胞经放疗后表达呈剂量及时间依赖性升高;经预测,hsa-miR-93可直接靶向结合STAT3.过表达hsa-miR-93可以在mRNA以及蛋白水平上显著抑制STAT3的表达(P<0.05),而抑制hsa-miR-93的表达可以显著上调STAT3的表达(P<0.05);过表达hsa-miR-93或沉默STAT3均使鼻咽癌细胞的集落形成显著减少(P<0.001);抑制hsa-miR-93减弱了鼻咽癌对放疗的敏感性,而同时抑制hsa-miR-93和STAT3后,可提高鼻咽癌对放疗的敏感性.结论 hsa-miR-93可能通过靶向STAT3提高鼻咽癌对放疗的敏感性.

Background: Current treatments for lung cancer have their own deficiencies, such as severe adverse effect. Therefore, more safe and effective drugs are needed. Purpose: Fuzheng Kang-Ai (FZKA for short) has been applied as an adjuvant treatment in advanced Non-Small Cell Lung Cancer (NSCLC) patients for decades in China, showing a definitive effect with minimal toxicities. However, the underlying mechanism is yet to be identified. Study design: Both in vitro and in vivo experiments were performed in this study to identify the exact mechanism by which FZKA inhibits NSCLC cell proliferation. Methods: MTT and CCK-8 assays were used to detect cell viability. Xenograft model was performed for in vivo experiments. CircRNA and miRNA sequencing were used to find the differentially expressed circRNAs and miRNAs, respectively. qRT-PCR was performed to check the expression levels of circRNA, miRNA and mRNA. BaseScope was carried out to observe the expression of circRNA in situ. Actinomycin D and RNase R experiments were done to show the stability of circRNA. Nuclear-cytoplasmic fractionation and FISH were used to identify the localization of circRNA and miRNA. Pull-down, RIP, and luciferase activity assays were performed to show the biding ability of circRNA, miRNA and target proteins. Flow cytometry was done to observe cell apoptosis. Western blot and IHC were done to detect the protein expression. TCGA database was used to analyze the survival rate. Results: FZKA inhibits NSCLC cell proliferation both in vitro and in vivo. Hsa_circ_0048091 and hsa-miR-378g were the most differentially expressed circRNA and miRNA, respectively, after FZKA treatment. Silencing hsa_circ_0048091 and overexpressing hsa-miR-378g promoted cell proliferation and reversed the inhibition effect of FZKA on NSCLC, respectively. Hsa-miR-378g was sponged by hsa_circ_0048091, and the overexpression of miR378g reversed the inhibition effect of hsa_ circ_0048091 on NSCLC. ARRDC3, as a target of hsa-miR-378g, was increased by FZKA treatment. Silencing ARRDC3 reversed both the inhibition effect of FZKA and miR-378g inhibitor on NSCLC. Conclusion: This study, for the first time, has established the function of hsa_circ_0048091, hsa- miR-378g, and ARRDC3 in lung cancer. It also shows that FZKA inhibits NSCLC cell proliferation through hsa_circ_0048091/hsamiR-378g/ARRDC3 pathway, uncovering a novel mechanism by which FZKA controls human NSCLC cell growth.

BACKGROUND:Kaempferol is extracted from Hedyotis diffusa, exerting an obvious anti-cancer effect. Here in the present study, we explored the anti-cancer effects and mechanism of kaempferol in non-small cell lung cancer cell (NSCLC).PURPOSE:Our objective is to figure out the molecular mechanism by which kaempferol promotes autophagy in NSCLC cells.STUDY DESIGN:A549 and H1299 NSCLC cell lines were used for in vitro experiments. And BALB/c nude mice of NSCLC were used to perform in vivo experiments.METHODS:For in vitro experiments, CCK-8 and EdU assay was used to observe the effect of kaempferol on NSCLC cell proliferation. Confocal microscopy of mCherry-EGFR-LC3 assay and electron microscopy assay were used to detect NSCLC cell autophagy. Protein expression was determined using Western blot, and mRNA expression was determined using qRT-PCR. Flow cytometry was performed to detect the cell apoptosis. For in vivo experiments, a subcutaneously implanted tumor model in BALB/C nude mice was performed using human NSCLC cell line A549-Luc. The kaempferol effect on NSCLC mice model was detected by measuring the tumor weight and bioluminescence intensity. Immunohistochemistry was done to measure the key protein expression from mice tumor tissues.RESULTS:Our results confirmed that kaempferol inhibited NSCLC cell proliferation significantly. And it promoted NSCLC cell autophagy, leading to NSCLC cell death. Interestingly, Met-was greatly inhibited at both protein and mRNA levels. Meanwhile, PI3K/AKT/mTOR signaling pathway was inhibited accordingly. Furthermore, overexpressing Met-reversed the effect of kaempferol on NSCLC cell viability and cell autophagy with significance. Finally, the above effect and pathway were validated using the xenograft model.CONCLUSION:Kaempferol may exert its anti-NSCLC effect by promoting NSCLC cell autophagy. Mechanistically, Met-and its downstream PI3K/AKT/mTOR signaling pathway were involved in the process, which provides a novel mechanism how kaempferol functions in inhibiting NSCLC.