RNA-based next-generation sequencing (NGS) has been recommended as a method for detecting fusion genes in non-small cell lung cancer (NSCLC) according to clinical practice guidelines and expert consensus. The primary targetable alterations in NSCLC consist of gene mutations and fusions, making the detection of gene mutations and fusions indispensable for assessing the feasibility of targeted therapies. Currently, the integration of DNA-based NGS and RNA-based NGS allows for simultaneous detection of gene mutations and fusions and has been partially implemented in clinical practice. However, standardized guidelines and criteria for the significance, application scenarios, and quality control of RNA-based NGS in fusion gene detection are still lacking in China. This consensus aims to provide further clarity on the practical significance, application scenarios, and quality control measures of RNA-based NGS in fusion gene detection. Additionally, it offers guiding recommendations to facilitate the clinical implementation of RNA-based NGS in the diagnosis and treatment of NSCLC, ultimately maximizing the benefits for patients from fusion gene detection.
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Objective Machine learning was used to screen the key characteristic genes of nasopharyngeal carcinoma (NPC) and analyze their correlation with immune cells. Methods Download the NPC training datasets (GSE12452 and GSE13597) and the validation dataset (GSE53819) from the Gene Expression Omnibus (GEO). Firstly, the training data sets were merged and screened for differentially expressed genes (DEGs); Secondly, the DEGs were analyzed by gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), gene set enrichment analysis (GSEA), and immune cell infiltration analysis. Next, the least absolute shrinkage and selection operator (LASSO) and support vector machine (SVM) algorithms were used to identify NPC-related genes in the training datasets and examined in the validation dataset, to further identify key genes using the area under curve (AUC) of receiver operating characteristic curve (ROC); Finally, the correlation between the key genes and immune cells was analyzed. Results A total of 55 DEGs were obtained, including 43 down-regulated genes and 12 up-regulated genes. The GO functions were enriched in humoral immune response, cell differentiation, neutrophil activation and chemokine receptor binding. The KEGG were mainly enriched in the IL-17 signaling pathway. The GSEA was enriched in cell cycle, extracellular matrix receptor interactions, cancer pathways and DNA replication. Immune infiltration analysis showed that the expression of naive B cells, memory B cells, and resting memory CD4+ T cells was significantly lower in NPC, while CD8+ T cells, naive CD4+ T cells, activated memory CD4+ T cells, follicular helper T cells, M0 macrophages and M1 macrophages were highly expressed in NPC. Among the feature genes screened by LASSO and SVM, only CCDC19, LAMB1, SPAG6 and RAD51AP1 genes' AUC were greater than 0.9 in both the training and validation datasets and were closely associated with immune cell infiltration. Conclusion The key genes CCDC19, LAMB1, SPAG6 and RAD51AP1 in NPC development are screened by machine learning algorithms, and are closely associated with immune cell infiltration.

To investigate the expression levels of LINC00342 in gastric cancer (GC) tissues and cells and the pathways mediating its effects on biological behaviors of GC cells.

To explore the radiosensitizing effect of icaritin on nasopharyngeal carcinoma (NPC) cells and the underlying mechanism.

Biliary tract cancer is characterized by high invasiveness, occult early clinical manifestations and rapid progression. Surgical resection typically fails to achieve satisfactory outcomes. Biliary tract cancer exhibits low sensitivity to radiotherapy and chemotherapy. The prognosis of patients is extremely poor. Genomics research based on next-generation sequencing technology has made some advances. The gene mutation spectrum of biliary tract cancer has been preliminarily revealed, which lays a foundation for the study of molecular typing. This review summarizes the research progress and clinical application of gene mutation spectrum of biliary tract cancer in recent years, aiming to provide reference for the clinical diagnosis, treatment and basic research.

Objective To explore the cell subsets and characteristics related to the prognosis of osteosarcoma by analyzing the cellular composition of tumor tissue samples from different osteosarcoma patients.Methods The single-cell sequencing data and bulk sequencing data of different osteosarcoma patients were downloaded.We extracted the information of cell samples for dimensionality reduction,annotation,and cell function analysis,so as to identify the cell subsets and clarify the cell characteristics related to the prognosis of osteosarcoma.The development trajectory of macrophages with prognostic significance was analyzed,and the prognostic model of osteosarcoma was established based on the differentially expressed genes of macrophage differentiation.Results The cellular composition presented heterogeneity in the patients with osteosarcoma.The infiltration of mononuclear phagocytes in osteosarcoma had prognostic significance(P=0.003).Four macrophage subsets were associated with prognosis,and their signature transcription factors included RUNX3(+),ETS1(+),HOXD11(+),ZNF281(+),and PRRX1(+).Prog_Macro2 and Prog_Macro4 were located at the end of the developmental trajectory,and the prognostic ability of macrophage subsets increased with the progression of osteosarcoma.The prognostic model established based on the differentially expressed genes involved in macrophage differentiation can distinguish the survival rate of osteosarcoma patients with different risks(P<0.001).Conclusion Macrophage subsets are closely related to the prognosis of osteosarcoma and can be used as the key target cells for the immunotherapy of osteosarcoma.

Objective To investigate the expression level of serine/threonine phosphoprotein phosphatase 4C(PPP4C)in gastric cancer,and analyze its relationship with prognosis and the underlying regulatory mechanism.Methods The clinical data of 104 gastric cancer patients admitted to the First Affiliated Hospital of Bengbu Medical College between January 2012 and August 2016 were collected.Immunohistochemical staining was employed to determine the expression levels of PPP4C and Ki-67 in the gastric cancer tissue.The gastric cancer cell lines BGC823 and HGC27 were cultured and transfected with the vector for PPP4C knockdown,the vector for PPP4C overexpression,and the lentiviral vector(control),respectively.The effects of PPP4C on the cell cycle and proliferation were analyzed and the possible regulatory mechanisms were explored.Results PPP4C was highly expressed in gastric cancer(P<0.001),and its expression promoted malignant progression of the tumor(all P<0.01).Univariate and Cox multivariate analysis clarified that high expression of PPP4C was an independent risk factor affecting the 5-year survival rate of gastric cancer patients(P=0.003).Gene ontology and Kyoto encyclopedia of genes and genomes enrichment analysis suggested that PPP4C may be involved in the cell cycle.The correlation analysis showed that the expression of PPP4C was positively correlated with that of Ki-67 in gastric cancer(P<0.001).The up-regulation of PPP4C expression increased the proportion of tumor cells in the S phase,alleviated the G2/M phase arrest,and promoted the proliferation of gastric cancer cells and the expression of cyclin D1 and cyclin-dependent kinase 6(CDK6)(all P<0.05).The down-regulation of PPP4C decreased the proportion of gastric cancer cells in the S phase,promoted G2/M phase arrest,and inhibited cell proliferation and the expression of cyclin D1,CDK6,and p53(all P<0.05).p53 inhibitors promoted the proliferation of BGC823 and HGC27 cells in the PPP4C knockdown group(P<0.001,P<0.001),while p53 activators inhibited the proliferation of BGC823 and HGC27 cells in the PPP4C overexpression group(P<0.001,P=0.002).Conclusions PPP4C is highly expressed in gastric cancer and affects the prognosis of the patients.It may increase the proportion of gastric cancer cells in the S phase and alleviate the G2/M phase arrest by inhibiting p53 signaling,thereby promoting cell proliferation.

Objective To investigate the impacts of forkhead box M1(FOXM1)on the proliferation,invasion,and drug resistance of gastric cancer cells by regulating the circular RNA circ_NOTCH1.Methods Western blotting and real-time quantitative PCR were performed to determine the expression of FOXM1 protein and circ_NOTCH1,respectively,in the gastric cancer tissue,para-carcinoma tissue,human normal gastric mucosa epithelial cell line GES-1 and gastric cancer cell lines MGC-803,HGC-27,and BGC-823.BGC-823 cells were classified into the following groups:control,short hairpin RNA FOXM1(sh-FOXM1)and negative control(sh-NC),small interfering RNA circ_NOTCH1(si-circ_NOTCH1)and negative control(si-NC),and sh-FOXM1+circ_NOTCH1 overexpression plasmid(sh-FOXM1+pcDNA-circ_NOTCH1)and sh-FOXM1+negative control(sh-FOXM1+pcDNA).CCK-8 assay and clone formation assay were employed to measure the cell proliferation,and Transwell assay to measure cell invasion.After treatment with 1.0 mg/L adriamycin for 48 h,the cell resistance in each group was analyzed.Western blotting was employed to determine the expression levels of FOXM1,proliferating cell nuclear antigen(PCNA),Bax,multi-drug resistance-associated protein 1(MRP1),and multi-drug resistance gene 1(MDR1).RNA pull-down and RNA immunoprecipitation were employed to examine the binding of circ_NOTCH1 to FOXM1 protein.Results Compared with those in the para-carcinoma tissue,the expression levels of FOXM1 protein and circ_NOTCH1 in the gastric cancer tissue were up-regulated(all P<0.001).Compared with GES-1 cells,MGC-803,HGC-27,and BGC-823 cells showed up-regulated expression levels of FOXM1 protein and circ_NOTCH1(all P<0.001).Compared with the control group and sh-NC group,the sh-FOXM1 group with down-regulated expression of FOXM1 protein and circ_NOTCH1 showed decreased optical density value,clone formation rate,cell invasion number,and cell viability,down-regulated expression of PCNA,MRP1,and MDR1,and up-regulated expression of Bax protein in BGC-823 cells(all P<0.001).Compared with the control group and the si-NC group,the si-circ_NOTCH1 group with down-regulated expression of circ_NOTCH1 showed decreased optical density value,clone formation rate,cell invasion number,and cell viability,down-regulated expression of PCNA,MRP1,and MDR1,and up-regulated expression of Bax protein in BGC-823 cells(all P<0.001).Compared with sh-FOXM1 group and sh-FOXM1+pcDNA group,the sh-FOXM1+pcDNA-circ_NOTCH1 group with up-regulated expression of circ_NOTCH1 showed increased optical density value,clone formation rate,cell invasion number,and cell viability,up-regulated expression of PCNA,MRP1,and MDR1,and down-regulated expression of Bax protein(all P<0.001).FOXM1 protein was able to interact with circ_NOTCH1.Conclusion Interference with FOXM1 may inhibit the proliferation,invasion,and drug resistance of gastric cancer cells by silencing circ_NOTCH1 expression.

Objective: To investigate the expression of neuraminidase-1 (NEU1) in Ewing sarcoma (ES) tissue and its effect on the proliferation and migration of ES cells. Methods: To obtain datasets of ES from the National Center for Biotechnology Information's High-Throughput Gene Expression Omnibus (GEO) for the analysis of NEU1 expression in ES; to acquire ES patient dataset from the International Cancer Genome Consortium (ICGC) database and apply Kaplan-Meier survival analysis to investigate the relationship between NEU1 and the prognosis of ES patients; adopting both univariate and multivariate Cox regression analysis to determine whether NEU1 is a prognostic factor for ES; adopting the Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation to analyze the potential mechanism of NEU1 in regulating the malignant biological behavior of ES; adopting the real-time fluorescence quantitative polynucleotide chain reaction (RT-qPCR) to verify the expression of NEU1 in the human bone marrow mesenchymal stem cells (hBMSC) and the ES cell line RD-ES; adopting the transfection technology to knock down the expression of NEU1 in ES cell lines and divide them into two groups: shRNA-NEU1 and shRNA-NC to explore the effects of altered NEU1 expression on ES malignant behavior; adopting the cell counting kit (CCK-8) and cell clone formation experiment to detect the proliferation ability of two groups of cells; adopting the scratch healing experiment to test the cell migration ability of the two groups. Results: We retrieved and analyzed data from the GEO database, including GSE17674 (44 ES tissues and 18 normal tissues) and GSE17679 (87 ES tissues and 18 normal tissues), and found that NEU1 expression was significantly higher in ES tissues compared to normal control tissues (P<0.001). The complete gene expression and clinical information of 56 ES patients obtained from the ICGC database revealed that the ES patients with high NEU1 expression (n=28) had a significantly lower overall survival rates at different time points compared to those with low NEU1 expression (n=28) (HR=2.830, 95%CI:1.324-6.051, P=0.005). Univariate analysis indicated that NEU1 could impact ES patient prognosis (HR=1.049, 95%CI: 1.008-1.092, P=0.019), and multivariate analysis further suggested that NEU1 could serve as a risk factor for ES prognosis (HR=1.087, 95%CI: 1.028-1.148, P=0.003). KEGG results show that MAPK signaling pathway and cell adhesion molecule signaling pathway were potential mechanisms regulating the malignant process of ES. The RT-qPCR results showed that the expression level of NEU1 in the RD-ES cell line is significantly higher than that in the control cell hBMSC (2 184.23±527.32 vs 1.00±0.08, P<0.001). The CCK-8 experiment results show that the proliferation of RD-ES cells in the NEU1 knockdown group was lower than that in the control group at 24, 48, and 72 hours (0.494±0.126 vs 0.696±0.118, 0.657±0.096 vs 1.142±0.182, 1.053±0.064 vs 1.980±0.146, all P<0.001). The results of single cell clone formation experiment show that the number of colony formation in the low expression NEU1 group was significantly lower than that in the control group (184.2±123.9 vs 362.8±78.0, P=0.021). The cell scratch healing experiment finds that the average scratch distance of the NEU1 knockdown group was significantly lower than that of the control group (19.6%±5.7% vs 56.0%±7.6%, P<0.001). Conclusion: NEU1 may be a prognostic factor in ES, and its abnormal expression in ES can affect the proliferation and migration ability of the ES cells, leading to poor prognosis in ES patients.

To carry out combined genetic analysis on two patients suspected for Burkitt lymphoma to facilitate their diagnosis and treatment.

To explore the clinical and laboratory characteristics of hematological tumors with different types of abnormalities in platelet derived growth factor β (PDGFRβ) gene.

Circulating tumor DNA (ctDNA) is emerging as a novel biomarker for tumor evaluation, offering advantages such as high sensitivity and specificity, minimal invasiveness, and absence of radiation. Currently, various techniques including gene sequencing and PCR are employed for ctDNA detection. The utilization of ctDNA for monitoring minimal residual disease (MRD) enables comprehensive assessment of tumor status and early identification of tumor recurrence, achieving a remarkable detection sensitivity of 0.01%. Therefore, ctDNA holds promise as a biomarker for early diagnosis, treatment response monitoring, and prognosis prediction in solid tumors. This article reviews the commonly used methods for detecting ctDNA and their advantages in evaluating tumor MRD and guiding clinical diagnosis and treatment.

To investigate the effects and mechanisms of deubiquitinating enzyme Josephin domain containing 2 (JOSD2) on susceptibility of non-small cell lung carcinoma (NSCLC) cells to anti-cancer drugs.

Objective: To investigate the clinicopathological features of perivascular epithelioid cell tumor (PEComa) of the lung. Methods: Eight PEComa cases of the lung diagnosed at the First Affiliated Hospital of Soochow University, Suzhou, China from July 2008 to December 2021 were collected and subject to immunohistochemical staining, fluorescence in situ hybridization and next generation sequencing. The relevant literature was reviewed and the clinicopathological features were analyzed. Results: There were 5 males and 3 females, aged from 18 to 70 years (mean 39 years). There were 3 cases of the right upper lung, 3 cases of the left lower lung, 1 case of the left upper lung and 1 case of the right middle lung. Seven cases were solitary and 1 case was multifocal (4 lesions). Seven cases were benign while one was malignant. The tumors were all located in the peripheral part of the lung, with a maximum diameter of 0.2-4.0 cm. Grossly, they were oval and well circumscribed. Microscopically, the tumor cells were oval, short spindle-shaped, arranged in solid nests, acinar or hemangiopericytoma-like patterns, with clear or eosinophilic cytoplasm. The stroma was rich in blood vessels with hyalinization. Coagulated necrosis and high-grade nuclei were seen in the malignant case, and calcification was seen in 2 cases. Immunohistochemically, the tumor cells were positive for Melan A (8/8), HMB45 (7/8), CD34 (6/8), TFE3 (4/7), and SMA (3/8). All cases were negative for CKpan and S-100. TFE3 (Xp11.2) gene fusion was examined using the TFE3 break-apart fluorescence in situ hybridization in 5 cases, in which only the malignant case was positive. The next generation sequencing revealed the SFPQ-TFE3 [t(X;1)(p11.2;p34)] fusion. Follow-up of the patients ranged from 12 to 173 months while one patient was lost to the follow-up. The malignant case had tumor metastasis to the brain 4 years after the operation and then received radiotherapy. Other 6 cases had no recurrence and metastasis, and all the 7 patients survived. Conclusions: Most of the PEComas of the lung are benign. When there are malignant morphological features such as necrosis, high-grade nuclei or SFPQ-TFE3 gene fusion, close follow-up seems necessary.

Objective: To investigate the clinicopathological features, immunophenotype, molecular features and differential diagnosis of primary synovial sarcoma of the lung (PSSL). Methods: Twelve cases of PSSL were collected at Henan Provincial People's Hospital, during May 2010 and April 2021, and their clinicopathological parameters were summarized. SS18-SSX, H3K27Me3, and SOX2 were added to the original immunomarkers to evaluate their diagnostic value for PSSL. Results: The age of 12 patients when diagnosed ranged from 32 to 75 years (mean of 50 years). There were 7 males and 5 females, 2 left lung cases and 10 right lung cases. Of the 6 patients who underwent surgical resection, five cases were confined to lung tissue (T1), one case had mediastinal invasion (T3), two cases had regional lymph node metastasis (N1), and none had distal metastasis. Microscopically, 11 cases showed monophasic spindle cell type and one case showed biphasic type composed of mainly epithelial cells consisting of cuboidal to columnar cells with glandular and cribriform structures. It was difficult to make the diagnosis by using the biopsy specimens. Immunohistochemistry (IHC) showed CKpan expression in 8 of 12 cases; EMA expression in 11 of 12 case; TLE1 expression in 8 of 12 cases; S-100 protein expression in two of 12 cases; various expression of bcl-2 and vimentin in 12 cases, but no expression of SOX10 and CD34 in all the cases. The Ki-67 index was 15%-30%. The expression of SS18-SSX fusion antibody was diffusely and strongly positive in all 12 cases. SOX2 was partially or diffusely expressed in 8 of 12 cases, with strong expression in the epithelial component. H3K27Me3 was absent in 3 of 12 cases. SS18 gene translocation was confirmed by fluorescence in situ hybridization (FISH) test in all 12 samples. Six cases underwent surgery and postoperative chemotherapy, while the other six cases had chemotherapy alone. Ten patients were followed up after 9-114 months, with an average of 41 months and a median of 26 months. Five patients survived and five died of the disease within two years. Conclusions: PSSL is rare and has a broad morphological spectrum. IHC and molecular tests are needed for definitive diagnosis. Compared with current commonly used IHC markers, SS18-SSX fusion antibody has better sensitivity to PSSL, which could be used as an alternative for FISH, reverse transcription-polymerase chain reaction or next generation sequencing in the diagnosis of PSSL.

Medical studies have found that tumor mutation burden (TMB) is positively correlated with the efficacy of immunotherapy for non-small cell lung cancer (NSCLC), and TMB value can be used to predict the efficacy of targeted therapy and chemotherapy. However, the calculation of TMB value mainly depends on the whole exon sequencing (WES) technology, which usually costs too much time and expenses. To deal with above problem, this paper studies the correlation between TMB and slice images by taking advantage of digital pathological slices commonly used in clinic and then predicts the patient TMB level accordingly. This paper proposes a deep learning model (RCA-MSAG) based on residual coordinate attention (RCA) structure and combined with multi-scale attention guidance (MSAG) module. The model takes ResNet-50 as the basic model and integrates coordinate attention (CA) into bottleneck module to capture the direction-aware and position-sensitive information, which makes the model able to locate and identify the interesting positions more accurately. And then, MSAG module is embedded into the network, which makes the model able to extract the deep features of lung cancer pathological sections and the interactive information between channels. The cancer genome map (TCGA) open dataset is adopted in the experiment, which consists of 200 pathological sections of lung adenocarcinoma, including 80 data samples with high TMB value, 77 data samples with medium TMB value and 43 data samples with low TMB value. Experimental results demonstrate that the accuracy, precision, recall and F1 score of the proposed model are 96.2%, 96.4%, 96.2% and 96.3%, respectively, which are superior to the existing mainstream deep learning models. The model proposed in this paper can promote clinical auxiliary diagnosis and has certain theoretical guiding significance for TMB prediction.

Objective: To explore the molecular markers related to lymph node metastasis of prostate cancer (PCa) based on bioinformatics technology and carry out clinical verification. Methods: The differentially expressed genes of PCa with lymph node metastasis were screened from geo data, and the hub genes of the gene co expression network were constructed. The hub genes were incorporated into the support vector machine model to evaluate its prediction efficiency. The hub genes were verified in the TCGA data set and analyzed for immune infiltration. The clinical data of 80 patients with prostate cancer in the Fourth Hospital of Hebei Medical University from January 2019 to December 2022 were collected. The logistic risk model was used to evaluate the prediction efficiency of hub gene metastasis. Results: Five hub genes (GSK3B, TP53, PSMC6, SUMO1, PIK3CA) were identified, and the support vector machine model constructed by them had good diagnostic value (the accuracy rate was 83.87%). TCGA validation results showed that only PSMC6 was significantly differentially expressed in PCa tissues with lymph node metastasis (P<0.001). The results of immune infiltration analysis showed that the expression of PSMC6 was significantly correlated with 9 kinds of immune cells (B cells, DC, IDC, etc.). Clinical information analysis showed that the expression of PSMC6 was significantly correlated with lymph node metastasis, PSA value, T stage and Gleason score (P<0.01). Univariate logistic results showed that T stage (OR=3.230, 95%CI:1.192-8.757, P=0.021), Gleason score (OR=4.627, 95%CI:2.212-9.677, P<0.001), PSMC6 (OR=25.235, 95%CI:5.326-119.560, P<0.001) could be used as predictors of lymph node metastasis. Multivariate logistic analysis showed that PSMC6 (OR=16.537, 95%CI:2.928-93.393, P=0.001) could be used as an independent risk factor for predicting lymph node metastasis. Conclusion: PSMC6 may be used as a potential molecular marker for judging lymph node metastasis in patients with PCa.

The incidence and mortality of lung cancer rank first among malignant tumors in China. Among them, non-small cell lung cancer (NSCLC) is the most important pathological type, accounting for 80%-85% of lung cancer patients, including adenocarcinoma, squamous cell carcinoma and other pathological subtypes. In recent years, with the discovery of epidermal growth factor receptor (EGFR) gene and the successful development of tyrosine kinase inhibitors (TKIs), the treatment efficacy of stage Ⅳ NSCLC patients with EGFR gene sensitive mutation has been greatly improved. As of August 23, 2023, the first generation EGFR-TKIs, gefitinib, icotinib, and erlotinib; the second generation EGFR-TKIs, afatinib and dacomitinib; and the third generation EGFR-TKIs, osimertinib, almonertinib, furmonertinib and befotertinib were all approved for marketing by China National Medical Products Administration (NMPA). In addition, multiple domestic third-generation EGFR-TKIs are undergoing clinical trials, such as rezivertinib (BPI-7711), limertinib (ASK120067), and oritinib (SH-1028). Meanwhile, mobocertinib and sunvozertinib, which targets EGFR 20ins mutations, were also approved by NMPA. With the increasing variety of EGFR-TKIs approved for marketing subsequently, it brings confusion to clinicians when choosing specific medications, and there is an urgent need to develop relevant treatment guidelines. Hence, the Medical Oncology Branch of China International Exchange and Promotive Association for Medical and Health Care and the Chinese Association for Clinical Oncologists convened experts to integrate the research results of various EGFR-TKIs, and proposed the "China clinical practice guideline for epidermal growth factor receptor tyrosine kinase inhibitors in stage Ⅳ non-small cell lung cancer (version 2023)", to provide reference for better clinical practice.