A propensity score-matched analysis of neoadjuvant chemoimmunotherapy versus surgery alone for locally advanced esophageal squamous cell carcinoma.

BACKGROUND: The aim of this study was to evaluate the safety, efficacy, and oncologic outcomes of neoadjuvant immunotherapy combined with chemotherapy (NICT) group and surgery alone group in the treatment of patients with locally advanced esophageal squamous cell carcinoma (ESCC). METHODS: A series of 232 consecutive patients who underwent surgery with or without NICT from June 2019 to August 2022 were evaluated. We performed propensity score matching between the NICT and surgery alone groups on the basis of estimated propensity scores for each patient. RESULTS: After propensity score matching, data of 137 patients with clinical stages II-IV ESCC, including 85 receiving surgery alone and 52 receiving NICT, were analyzed. Compared with the surgery alone group (301.7 ± 94.4 min), the operation time was significantly longer in the NICT group (333.4 ± 79.7 min). However, there was no significant difference between the two groups in the postoperative complications, intraoperative blood loss, thoracic fluid volume, chest tube duration, lengths of intensive care unit stay and postoperative hospitalization. Additionally, 90-day mortality rate and 30-day readmission were similar in both groups. CONCLUSIONS: Overall, NICT followed by esophagectomy appears to be safe and feasible for locally advanced ESCC. However, further multicenter prospective clinical trials are needed to validate our results.

Identification of hub genes in the subacute spinal cord injury in rats.

BACKGROUND: Spinal cord injury (SCI) is a common trauma in clinical practices. Subacute SCI is mainly characterized by neuronal apoptosis, axonal demyelination, Wallerian degeneration, axonal remodeling, and glial scar formation. It has been discovered in recent years that inflammatory responses are particularly important in subacute SCI. However, the mechanisms mediating inflammation are not completely clear. METHODS: The gene expression profiles of GSE20907, GSE45006, and GSE45550 were downloaded from the GEO database. The models of the three gene expression profiles were all for SCI to the thoracic segment of the rat. The differentially expressed genes (DEGs) and weighted correlation network analysis (WGCNA) were performed using R software, and functional enrichment analysis and protein-protein interaction (PPI) network were performed using Metascape. Module analysis was performed using Cytoscape. Finally, the relative mRNA expression level of central genes was verified by RT-PCR. RESULTS: A total of 206 candidate genes were identified, including 164 up-regulated genes and 42 down-regulated genes. The PPI network was evaluated, and the candidate genes enrichment results were mainly related to the production of tumor necrosis factors and innate immune regulatory response. Twelve core genes were identified, including 10 up-regulated genes and 2 down-regulated genes. Finally, seven hub genes with statistical significance in both the RT-PCR results and expression matrix were identified, namely Itgb1, Ptprc, Cd63, Lgals3, Vav1, Shc1, and Casp4. They are all related to the activation process of microglia. CONCLUSION: In this study, we identified the hub genes and signaling pathways involved in subacute SCI using bioinformatics methods, which may provide a molecular basis for the future treatment of SCI.

Methylation of neurofilament light polypeptide promoter is associated with cell invasion and metastasis in NSCLC.

The role of NEFL in NSCLC remains largely unknown. Immunohistochemistry was performed to investigate the expression of NEFL in 108 lung cancer specimens. NEFL expression was associated with decreased lymph node metastases and favorable prognosis. Furthermore, real-time PCR and Western blot were used to investigate the expression of the NEFL gene in NSCLC cell lines. Subsequently, lentivirus-mediated RNA interference and overexpression were used to demonstrate that knocked-down of NEFL enhanced the invasion and migration of A549 and H460 NSCLC cells, whereas NEFL overexpression resulted in a suppression of the invasion and migration of GLC-82 and L78 cells in vitro. In addition, bisulfite sequence PCR assay demonstrated that NEFL downregulation was associated with promoter methylation, and NEFL expression was restored after treatment with 5-Aza-dC. Finally, we demonstrated that NEFL inhibited the NF-κB pathway, thereby suppressing the expression of uPA and decreasing NSCLC invasiveness and migration. Our studies suggest that NEFL methylation is a novel mechanism for NSCLC invasion and metastasis and that NEFL may represent a candidate biomarker for recurrence and survival in patients with NSCLC.

Anti-PD-L1 immune checkpoint inhibitors in combination with etoposide and platinum for extensive-stage small cell lung cancer: a case report.

The conventional etoposide-platinum (EP) regimen and adjuvant radiotherapy remain the gold-standard treatment for small cell lung cancer (SCLC). However, most patients already have multiple metastases when they are first diagnosed with SCLC. The objective response rate (ORR) and 1-year survival rate are low in these patients despite active radiotherapy and chemotherapy. SCLC is oncologically featured by the high tumor mutational burden (TMB) of multiple genes, which makes immunotherapy a possible new treatment strategy for SCLC. New data from the IMpower133 and CASPIAN trials will shed new light on the treatment of SCLC. In 2020, the results from the phase 3 CASPIAN trial have already suggested that programmed cell death-ligand 1 (PD-L1) inhibitors may represent breakthroughs in the management of SCLC. Here, we report a patient with extensive-stage SCLC (ES-SCLC) treated with first-line anti-PD-L1 immune checkpoint inhibitor (PD-L1 inhibitor) (i.e., durvalumab) combined with the EP regimen for 6 cycles. The patient consistently achieved partial response (PR) [nearly complete response (CR)], and no immune-related adverse events were noted during this period. The Karnofsky performance status (PS) score maintained at 1-2 points. We further review the history of SCLC treatment and elucidate the role of combination with immunotherapy in treating SCLC in the coming years.

Detection of circulating genetically abnormal cells using 4-color fluorescence in situ hybridization for the early detection of lung cancer.

PURPOSE: Available biomarkers lack sensitivity for an early lung cancer. Circulating genetically abnormal cells (CACs) occur early in tumorigenesis. To determine the diagnostic value of CACs in blood detected by 4-color fluorescence in situ hybridization (FISH) for lung cancer. METHODS: This was a prospective study of patients with pulmonary nodules ≤ 30 mm detected between 10/2019 and 01/2020 at four tertiary hospitals in China. All patients underwent a pathological examination of lung nodules found by imaging and were grouped as malignant and benign. CACs were detected by 4-color FISH. Patients were divided into the training and validation cohorts. Receiver operating characteristics analysis was used to analyze the diagnosis value of CACs. RESULTS: A total of 205 participants were enrolled. Using a cut-off value of ≥ 3, blood CACs achieved areas under the curve (AUCs) of 0.887, 0.823, and 0.823 for lung cancer in the training and validation cohorts, and all patients, respectively. CACs had high diagnostic values across all tumor sizes and imaging lesion types. CACs were decreased after surgery (median, 4 vs. 1, P < 0.001) in the validation set. The CAC status between blood and tissues was highly consistent (kappa = 0.909, P < 0.001). The AUC of CAC (0.823) was higher than that of CEA (0.478), SCC (0.516), NSE (0.506), ProGRP (0.519), and CYFRA21-1 (0.535) (all P < 0.001). CONCLUSION: CACs might have a high value for the early diagnosis of lung cancer. These findings might need to be validated in future studies. Evidence suggested homology in genetic aberrations between the CACs and the tumor cells.

miR-21 Overexpression Promotes Esophageal Squamous Cell Carcinoma Invasion and Migration by Repressing Tropomyosin 1.

The migration and invasion of esophageal squamous cell carcinoma are associated with clinical outcomes, however, the mechanisms remain poorly understood. Here, we found that miR-21 is significantly overexpressed in ESCC, lung cancer, and bladder cancer compared with the adjacent normal tissue. MiR-21 and TPM1 expressions were analyzed by RT-qPCR and WB in 30 ESCC, 10 lung cancer, and 10 bladder cancer clinical specimens, each with matched adjacent normal tissue. Knockdown and overexpression of miR-21 as well as knockdown of TPM1 in ESCC cell lines were performed using synthetic oligonucleotides. TPM1 3'UTR luciferase reporter constructs were used to investigate targeting of TPM1 by miR-21. ESCC migration and invasion were assessed using transwell migration and invasion assays. Inhibition of miR-21 reduced migration and invasion in two ESCC cell lines, and overexpression of miR-21 promoted migration and invasion in vitro. Interestingly, TPM1 exhibited inverse patterns of expression compared with miR-21 in tissues and cell lines. Luciferase reporter assays demonstrated that TPM1 was directly regulated by miR-21. Moreover, the forced overexpression of miR-21 repressed the TPM1 expression, while silencing of miR-21 restored the TPM1 expression in ESCC cell lines. What is more, simultaneous silencing of miR-21 and TPM1 expressions did not alter the migratory and invasive characteristics demonstrating that the effects of miR-21 were mediated through TPM1. In conclusion, the aberrant overexpression of miR-21 is common in cancer and promotes the migration and invasion of ESCC through inhibiting the TPM1 expression. These results suggest that miR-21 may be a novel predictive marker and therapeutic target for treatment of ESCC.

Glutathione Peroxidase 1 Promotes NSCLC Resistance to Cisplatin via ROS-Induced Activation of PI3K/AKT Pathway.

PURPOSE: Reactive oxygen species (ROS)-induced cytotoxicity is an important mechanism by which cisplatin kills tumor cells. Glutathione peroxidase family (GPXs) is an important member of antioxidant system which metabolizes intracellular ROS and maintains homeostasis of cells. Altered expressions of GPXs enzymes, especially GPX1, have been described in a variety of human cancers. However, their functional roles in cisplatin-based chemoresistance in human malignancies including non-small cell lung cancer have never been explored. METHODS: A panel of NSCLC cell lines were selected for this study. GPX1 expression was detected using quantitative RT-PCR and Western blot. Cisplatin-induced cell killing was analyzed by CCK8 assay. Intracellular ROS levels were detected by fluorescence-based flow cytometry analysis. In vitro overexpression and knockdown of GPX1 expression were performed using GPX1 expression vector and siRNA approaches. Protein levels of PTEN, NF-κB, BCL2, Bax, and phosphorylated AKT were detected with western blot analysis using specific antibodies. RESULTS: GPX1 expression was upregulated in a subset of NSCLC cell lines resistant to cisplatin treatment. Expression vector-mediated forced overexpression of GPX1 significantly increased cisplatin resistance in NSCLC cell lines, whereas RNA inference-mediated downregulation of GPX1 could restore sensitivity to cisplatin. Overexpression of GPX1 significantly suppressed elevation of intracellular ROS and activation of AKT pathway when NSCLC cell lines were exposed to different concentrations of cisplatin. Activation of the AKT pathway inhibited proapoptotic cascade and subsequently led to cisplatin resistance in NSCLC cells. Inhibition of NF-κB by its chemical inhibitor BAY can significantly downregulate GPX1 expression and restore the cisplatin sensitivity of the cell lines resistant to cisplatin. CONCLUSIONS: Our findings suggested that overexpression of GPX1 is a novel molecular mechanism for cisplatin-based chemoresistance in NSCLC. GPX1 overexpression blocks cisplatin-induced ROS intracellular accumulation, activates PI3K-AKT pathway by increased AKT phosphorylation, and further leads to cisplatin resistance in NSCLC cells. Inhibition of NF-κB signaling may be an alternative approach for restoring cisplatin sensitivity for NSCLC cells resistant to cisplatin-based chemotherapy.

[Applicable Value of AMSS-PCR in Lung Cancer Gene Mutation Detection].

BACKGROUND: The detection of driver oncogenes of lung cancer is of great importance. There are various gene detection techniques nowadays which are different from each other. We carried out this study to investigate the specificity and sensitivity of assay panels based on an Amplification Refractory Mutation System-polymerase chain reaction (ARMS-PCR) technique of Amplification Mutation Specific System (AMSS) in detection of lung cancer gene mutation. To estimate the applicable value of assay panels in clinical settings. METHODS: We collected cancer tissue specimens or fluid specimens from 309 patients. Mutation results were presented for those samples previously detected by ARMS-PCR. In comparison, we carried out AMSS-PCR using (epidermal growth factor receptor, EGFR) assay panel and Six-Alliance assay panel as well as Sanger sequencing. Software SPSS 22.0 (SPSS IBM) was used for statistical analysis. RESULTS: The rates of consistency between the results by assay panels and Sanger sequencing or ARMS-PCR were 97.41% and 97.73%, respectively. Besides, EGFR assay panel had higher consistency rates with other detection methods than Six-Alliance assay panel. As for consistency test, the Kappa values of assay panels with Sanger sequencing, assay panels with ARMS-PCR, and ARMS-PCR with Sanger sequencing were 0.946, 0.953, and 0.913, respectively. The receiver operating characteristic curve (ROC) area under curve (AUC) of assay panels was 0.976 referring to Sanger sequencing, and 0.975 as ARMS-PCR was referred to. CONCLUSIONS: AMSS-PCR can make an optimal cancer gene mutation detection method for clinical settings.

Hypoxia-induced microRNA-301b regulates apoptosis by targeting Bim in lung cancer.

OBJECTIVES: Worldwide, lung cancer accounts for the majority of cancer-related deaths. Aberrant expression of miRNAs has increasingly been reported to be associated with tumour progression. This study aimed to explore the role of miR-301b in regulating apoptosis in lung cancer. MATERIALS AND METHODS: Expression of miR-301b was assessed by real-time PCR in cell lines, human patient tissues and cells treated under hypoxia and DMOG. Scramble siRNA, miR-301b inhibitor and miR-301b mimics were transfected into lung cancer cells to determine their effects on apoptosis. Additionally, a mouse xenograft model was used to explore functions of miR-301b on apoptosis, in vivo. Finally, relationships between Bim and miR-301b levels were explored by luciferase reporter assay and Western blotting. RESULTS: We found that miR-301b was highly expressed in lung cancer tissues and cell lines. Expression of miR-301b was induced by hypoxia, and miR-301b suppressed expression of Bim by targeting its 3'UTR. Functionally, ectopic expression of miR-301b enhanced cell population growth, reduced apoptosis and reduced sensitivity of cells to chemotherapy. In the xenograft model, overexpression of miR-301b promoted tumour growth. Additionally, miR-301b and Bim expression were inversely correlated in clinical lung cancer samples. CONCLUSIONS: This study provides new insights into the function of miRNA-301b in lung cancer and suggests that miRNA-301b could be a potential molecular target for chemotherapy.

Left- and right-sided video-assisted thoracoscopic thymectomy exhibit similar effects on myasthenia gravis.

BACKGROUND: Unilateral video-assisted thoracoscopic (VATS) thymectomy features less operative trauma, improved cosmesis, and similar efficiency compared with transsternal (TS) thymectomy for treatment of patients with myasthenia gravis (MG). Unilateral VATS thymectomy can be easily performed from either side of the thorax, because thymus is located in the middle of mediastinum. Nevertheless, the side that provides better outcomes remains controversial. This study presents our experience on treatments for MG and reveals the differences between the unilateral VATS thymectomy performed on each side. METHODS: Eighty-one consecutive patients with MG who underwent TS or VATS thymectomy on either side between January 2003 and December 2012 were enrolled in the study. Clinicopathologic data and surgical outcomes were retrospectively analyzed and compared among different surgical approaches. RESULTS: TS thymectomy was administered in 50 patients, whereas unilateral VATS approaches were performed on the remaining 31 patients, 15 on the left side and 16 on the right side. The VATS group exhibited a significantly shorter surgery duration (P<0.001), less intraoperative blood loss (P=0.009), shorter postoperative hospital stay (P=0.025), smaller thoracic drainage volume (P=0.033), shorter thoracic drainage duration (P=0.006), and less postoperative complications (P<0.001) compared with the TS group. However, disease remission rates did not significantly differ among the groups (P=0.988). The left-sided group exhibited considerably longer thoracic drainage duration than the right-sided group (P=0.041). Moreover, surgical time (P=0.736), intraoperative blood loss (P=0.281), postoperative hospital stay (P=0.599), thoracic drainage volume (P=0.571), postoperative complications (P=0.742) and therapeutic effect (P=1.000) did not significantly differ among the groups. Multivariate analysis revealed that the ocular type of MG is the only independent factor for clinical remission (P=0.002). CONCLUSIONS: Unilateral VATS thymectomy can reduce surgical risks and shorten hospitalization duration without threatening the therapeutic effect. This technique can be safely and effectively performed by experienced surgeons in either side of the thorax.

DNA promoter hypermethylation contributes to down-regulation of galactocerebrosidase gene in lung and head and neck cancers.

Galactocerebrosidase (GALC) is a lysosomal enzyme responsible for glycosphingolipids degradation byproducts of which are important for synthesis of apoptosis mediator ceramide. Reduced expression of GALC has been identified in human malignancies; however, molecular mechanisms underlying down-regulation of GALC expression in cancer remain unknown. We performed methylation and expression analysis on GALC gene in a panel of head and neck cancer (HNC) and lung cancer cell lines, attempting to understand the regulation of GALC in human cancer. QRT-PCR and western blot analysis were performed to detect the expression of GALC in HNC. Bisulfite DNA sequencing and real-time qMSP were used to detect the methylation of GALC in HNC and lung cancer cell lines. 5aza-dC treatment assay was used to analysis the functional effect of GALC methylation on GALC expression in HNC. Reduction or complete absence of GALC expression was observed in more than a half of the tested HNC cell lines (8/14). 7 out of 8 cell lines with down-regulated expression harbored heavy CpG island methylation, while all cell lines with abundant expression of the gene contained no methylation. Hypermethylation was also found in primary HNC tumor tissues and lung cancer cell lines whereas absent in normal oral mucosa tissues. Demethylating treatment demonstrated that 5aza-dC significantly restored GALC expression in cell lines with methylated promoter while showed no effect on cell lines without promoter hypermethylation. Our findings for the first time demonstrated that promoter hypermethylation contributed to down-regulation of GALC Gene, implicating epigenetic inactivation of GALC may play a role in tumorigenesis of cancer.

High GPX1 expression promotes esophageal squamous cell carcinoma invasion, migration, proliferation and cisplatin-resistance but can be reduced by vitamin D.

Esophageal cancer is one of the most common cancers worldwide. Despite recent progress in the development of novel therapies, esophageal carcinoma remains an aggressive cancer associated with a poor prognosis. The glutathione peroxidase 1 (GPX1) gene located on chromosome 3p21.3 is associated with the cancer of several organs. According to available information, GPX1, a gene downstream of NF-κB, is considered to exert adverse effects on tumour progression and enhance malignancy in some cancers but has not been reported in esophageal cancer. It is also reported that vitamin D (Vit. D), a widely used drug in the clinical setting, could suppress GPX1 expression through the NF-κB pathway. Thus, it is speculated that Vit. D could reduce malignancy in esophageal cancer by altering the NF-κB pathway. In this study, we confirmed our speculation by finding that Vit. D, through the inhibition of GPX1, decreased the migratory, invasive and proliferative capabilities, as well as cisplatin resistance, in esophageal cancer cells. Furthermore, when invasion and migration were reduced in the GPX1-inhibited cells, the expression of urokinase type plasminogen activator (uPA) and matrix metalloproteinase-2 (MMP2) was also suppressed correspondingly. Therefore, we believe that, in esophageal cancer cells, the expression of GPX1 can promote invasion, migration, proliferation and cisplatin resistance, and Vit. D can reduce the associated malignancy through the NF-κB pathway. The Vit. D- and NF-κB-mediated decrease in GPX1 expression resulted in a decrease in MMP2- and uPA-mediated invasion and migration.

Role of neurofilament light polypeptide in head and neck cancer chemoresistance.

Resistance to cisplatin-based chemotherapy is responsible for therapeutic failure of many common human cancers including cancer of head and neck (HNC). Mechanisms underlying cisplatin resistance remain unclear. In this study, we identified neurofilament light polypeptide (NEFL) as a novel hypermethylated gene associated with resistance to cisplatin-based chemotherapy in HNC. Analysis of 14 HNC cell lines revealed that downregulation of NEFL expression significantly correlated with increased resistance to cisplatin. Hypermethylation of NEFL promoter CpG islands was observed in cell lines as examined by bisulfite DNA sequencing and methylation-specific PCR (MSP) and tightly correlated with reduced NEFL mRNA and protein expression. Furthermore, in patient samples with HNC (n = 51) analyzed by quantitative MSP, NEFL promoter hypermethylation was associated with resistance to cisplatin-based chemotherapy [relative risk (RR), 3.045; 95% confidence interval (CI), 1.459-6.355; P = 0.007] and predicted diminished overall and disease-free survival for patients treated with cisplatin-based chemotherapy. Knockdown of NEFL by siRNA in the highly cisplatin-sensitive cell line PCI13 increased (P < 0.01) resistance to cisplatin. In cisplatin-resistant O11 and SCC25cp cells, restored expression of NEFL significantly increased sensitivity to the drug. Furthermore, NEFL physically associated with tuberous sclerosis complex 1 (TSC1), a known inhibitor of the mTOR pathway, and NEFL downregulation led to functional activation of mTOR pathway and consequentially conferred cisplatin resistance. This is the first study to show a role for NEFL in HNC chemoresistance. Our findings suggest that NEFL methylation is a novel mechanism for HNC chemoresistance and may represent a candidate biomarker predictive of chemotherapeutic response and survival in patients with HNC.

GPx3 promoter hypermethylation is a frequent event in human cancer and is associated with tumorigenesis and chemotherapy response.

Glutathione peroxidase 3 (GPx3), a plasma antioxidant enzyme, maintains genomic integrity by inactivating reactive oxygen species (ROS), known DNA-damaging agents and mediators of cancer chemotherapy response. In this study, we demonstrate that loss of GPx3 expression by promoter hypermethylation is frequently observed in a wide spectrum of human malignancies. Furthermore, GPx3 methylation correlates with head and neck cancer (HNC) chemoresistance and may serve as a potential prognostic indicator for HNC patients treated with cisplatin-based chemotherapy. Our findings support the hypothesis that defects in the antioxidant system may contribute to tumorigenesis of a wide spectrum of human malignancies. GPx3 methylation may have implications in chemotherapy response and clinical outcome of HNC patients.

[Clinical significance of tumor interstitial T lymphocyte subset activity in non-small-cell lung cancer].

OBJECTIVE: To study the relation of tumor interstitial T lymphocyte subset activity to the clinical staging of non-small-cell lung cancer (NSCLC) and the immune response. METHODS: Immunohistochemical staining for CD4(+), CD8(+) and CD4(+)CD25(+) Foxp3(+) (regulatory T cells, Treg) T cells was performed on paraffin-embedded tissues from 60 NSCLC cases. RESULTS: Compared to stage I/II NSCLC patients, patients in stage III/IV showed a significant decrease in the percentage of CD4(+) and CD4(+)/CD8(+) T cells (P<0.05) and an increase in CD8(+) and CD4(+)CD25(+) Foxp3(+) T cells (P<0.05). Treg cells were enriched in the tumor tissue as compared with those in the adjacent tissues. CONCLUSIONS: The proportion of CD4(+)CD25(+) Foxp3(+) Treg cells is positively correlated to the clinical staging of NSCLC, in which T cell-mediated immune response is suppressed.