Overexpression SPRY2 Suppresses Esophageal Squamous Cell Carcinoma Progression Via Inactivation of ERK/AKT Signaling / La Sobreexpresión SPRY2 Suprime la Evolución del Carcinoma de Células Escamosas de Esófago Mediante la Inactivación de la Señalización ERK/AKT

SUMMARY: Esophageal cancer is one of the most aggressive gastrointestinal cancers. Invasion and metastasis are the main causes of poor prognosis of esophageal cancer. SPRY2 has been reported to exert promoting effects in human cancers, which controls signal pathways including PI3K/AKT and MAPKs. However, the expression of SPRY2 in esophageal squamous cell carcinoma (ESCC) and its underlying mechanism remain unclear. In the present study, we aimed to investigate the detailed role of SPRY2 in the regulation of cell proliferation, invasion and ERK/AKT signaling pathway in ESCC. It was identified that the expression level of SPRY2 in ESCC was remarkably decreased compared with normal tissues, and it was related to clinicopathologic features and prognosis ESCC patients. The upregulation of SPRY2 expression notably inhibited the proliferation, migration and invasion of Eca-109 cells. In addition, the activity of ERK /AKT signaling was also suppressed by the SPRY2 upregulation in Eca-109 cells. Our study suggests that overexpression of SPRY2 suppress cancer cell proliferation and invasion of by through suppression of the ERK/AKT signaling pathways in ESCC. Therefore, SPRY2 may be a promising prognostic marker and therapeutic target for ESCC.

El cáncer de esófago es uno de los cánceres gastrointestinales más agresivos. La invasión y la metástasis son las principales causas de mal pronóstico del cáncer de esófago. Se ha informado que SPRY2 ejerce efectos promotores en los cánceres humanos, que controla las vías de señales, incluidas PI3K/AKT y MAPK. Sin embargo, la expresión de SPRY2 en el carcinoma de células escamosas de esófago (ESCC) y su mecanismo subyacente aún no están claros. En el presente estudio, nuestro objetivo fue investigar el papel detallado de SPRY2 en la regulación de la proliferación celular, la invasión y la vía de señalización ERK/AKT en ESCC. Se identificó que el nivel de expresión de SPRY2 en ESCC estaba notablemente disminuido en comparación con los tejidos normales, y estaba relacionado con las características clínico-patológicas y el pronóstico de los pacientes con ESCC. La regulación positiva de la expresión de SPRY2 inhibió notablemente la proliferación, migración e invasión de células Eca-109. Además, la actividad de la señalización de ERK/AKT también fue suprimida por la regulación positiva de SPRY2 en las células Eca-109. Nuestro estudio sugiere que la sobreexpresión de SPRY2 suprime la proliferación y la invasión de células cancerosas mediante la supresión de las vías de señalización ERK/AKT en ESCC. Por lo tanto, SPRY2 puede ser un marcador de pronóstico prometedor y un objetivo terapéutico para la ESCC.

Effect of Survivin Down-Regulation by Egr1-Survivin shRNA Combined with Radiotherapy on Radiosensitivity of Esophageal Squamous Carcinoma / Efecto de la Regulación Negativa de Survivina por shRNA de Egr1-Survivina Combinado con Radioterapia sobre la Radiosensibilidad del Carcinoma Escamoso de Esófago

SUMMARY: This study is to investigate the effect of survivin down-regulation by Egr1-survivin shRNA combined with radiotherapy on the apoptosis and radiosensitivity of esophageal squamous cell carcinoma ECA109 and KYSE150 cells. ECA109 and KYSE150 cells were transfected with Egr1-survivin shRNA, and then treated with radiotherapy. After 24 h, the mRNA and protein levels of Egr1-survivin were detected by qPCR and Western-Blot. Cell cycle and apoptosis were detected by flow cytometry. Western blot also detected levels of cleavaged Caspase 3 and Caspase 9. YM155 was used as a positive control to inhibit survivin expression. The levels of survivin mRNA and protein in ECA109 and KYSE150 cells treated with Egr1-survivin shRNA combined with radiotherapy were significantly lower than those of the blank control group, the empty vector control group, and, the YM155 + radiotherapy group (P<0.05). Meanwhile, after survivin down-regulation, the ratio of G2 to S phase of ECA109 and KYSE150 cells increased significantly, leading to significant G2 and S phase arrest. Additionally, apoptosis of ECA109 and KYSE150 cells increased significantly (P <0.01). Further, protein levels of cleavaged Caspase 3 and Caspase 9 significantly increased in Egr1-survivin shRNA combined with radiotherapy group. Egr1-survivin shRNA combined with radiotherapy can down-regulate survivin expression, which further increases the apoptosis, and enhances the radiosensitivity of ECA109 and KYSE150 cells.

Este estudio tuvo como objetivo investigar el efecto de la regulación negativa de survivina por el shRNA de Egr1-survivina combinado con radioterapia sobre la apoptosis y la radiosensibilidad del carcinoma de células escamosas de esófago Células ECA109 y KYSE150. Las células ECA109 y KYSE150 se transfectaron con shRNA de survivina Egr1 y luego se trataron con radioterapia. Después de 24 h, los niveles de ARNm y proteína de Egr1-survivina se detectaron mediante qPCR y Western-Blot. El ciclo celular y la apoptosis se detectaron mediante citometría de flujo. La transferencia Western también detectó niveles de Caspasa 3 y Caspasa 9 escindidas. Se usó YM155 como control positivo para inhibir la expresión de survivina. Los niveles de ARNm y proteína de survivina en células ECA109 y KYSE150 tratadas con shRNA de survivina Egr1 combinado con radioterapia fueron significativamente más bajos que los del grupo control en blanco, el grupo control de vector vacío y el grupo de radioterapia YM155 + (P <0,05). Mientras tanto, después de la regulación negativa de survivina, la proporción entre las fases G2 y S de las células ECA109 y KYSE150 aumentó significativamente, lo que llevó a una detención significativa de las fases G2 y S. Además, la apoptosis de las células ECA109 y KYSE150 aumentó significativamente (P <0,01). Además, los niveles de proteína de Caspasa 3 y Caspasa 9 escindidas aumentaron significativamente en el shRNA de Egr1- survivina combinado con el grupo de radioterapia. El shRNA de survivina de Egr1 combinado con radioterapia puede regular negativamente la expresión de survivina, lo que aumenta aún más la apoptosis y mejora la radiosensibilidad de las células ECA109 y KYSE150.

Therapeutic Effect of Reibaisan (WTMCGEP) in Patients with Advanced Cancer / 日本東洋医学雑誌

Here we report 3 cases of advanced cancer using multidisciplinary treatment including reibaisan (WTMCGEP, a dry extract of Wisteria floribunda, Trapa natans, Myristica fragrans, Coix semen, Ganoderma lucidum, Elfvingia applanata, Punica granatum). Case 1 : 87-year-old man, suffering from stage IV esophageal squamous cell carcinoma (ESCC) with aortic and bronchial invasion, was referred to our clinic for palliative care. He had radiotherapy and chemotherapy. Only one course of chemotherapy was performed due to its intolerable side effects. The treatment with reibaisan started 11 months after the diagnosis. ESCC disappeared after 17 months of reibaisan treatment, and no relapse was observed for 66 months after the diagnosis. Case 2 : 79-year-old man, suffering from stage III ESCC, was initially scheduled for surgery after preoperative chemotherapy. Only one course of preoperative chemotherapy was performed because of its intolerable side effects. Therefore, radiotherapy combined with reibaisan followed. ESCC disappeared 6 months later, and no relapse was observed for 33 months after the diagnosis. Case 3 : 73-year-old woman, suffering from stage IV pancreatic cancer with systemic metastasis (brain, lung, and peritoneum). She initially showed Trousseau syndrome and was treated with low-molecular-weight heparin for multiple cerebral infarctions. One-month palliative chemotherapy and reibaisan resulted in a rapid reduction of ascites and improvement of neurological symptoms. Her progression-free survival period was 7 months. She lived 13 months thereafter. This suggests that reibaisan, which contains crude drugs that have been shown to have antitumor effects, may be another promising treatment for advanced cancers.

Sarcopenia Before Concurrent Chemoradiotherapy May Develop Severe Adverse Reactions in Patients with Esophageal Squamous Cell Carcinoma / 肿瘤防治研究

Objective To investigate whether sarcopenia before concurrent chemoradiotherapy is prone to grade≥3 acute adverse reactions (AAR) and shorten survival in patients with advanced esophageal squamous cell carcinoma (ESCC). Methods Data of advanced patients with pathologically diagnosed ESCC and CCRT (FP regimen) from August 2018 to July 2022 were reviewed retrospectively.Skeletal muscle mass and body composition were measured using pre-treatment CT images, and patients were divided into sarcopenia and non-sarcopenia groups.Grade≥3 AAR was diagnosed based on CTCAE5.0 and acute radiation injury criteria of the US RTOG.Risk factors for developing grade≥3 AAR were analyzed, and survival rates were calculated by Kaplan-Meier method. Results Among 132 patients with ESCC (87 in the sarcopenia group and 45 in the non-sarcopenia group), 23(17.4%) experienced grade≥3 AAR.In multivariate regression analysis, independent risk factors for grade ≥3 AAR included the following: sarcopenia (OR: 6.034, 95%CI: 1.206-30.190, P=0.029), decreased SMD (OR: 0.693, 95%CI: 0.492-0.976, P=0.036), decreased SMI (OR: 0.841, 95%CI: 0.721-0.982, P=0.028), and increased FMI (OR: 2.433, 95%CI: 1.194-4.958;P=0.014).The OS rates were 16.01 months (95%CI: 14.89-17.13) in the sarcopenia group and 19.27 months (95%CI: 14.45-24.09) in the non-sarcopenia group (χ2=5.326, P=0.021) as well as 14.86 months (95%CI: 11.30-18.42) for patients with grade≥3 AAR and 16.67 months (95%CI: 14.91-18.43) for patients with grade 0-2 AAR (χ2=5.47, P=0.019).Among patients with grade≥3 AAR, the OS rates were 12.13 months (95%CI: 10.15-14.11) in the sarcopenia group and 18.69 months (95%CI: 12.85-21.88) in the non-sarcopenia group (χ2=4.466, P=0.035). Conclusion Sarcopenia, decreased skeletal muscle density, and increased fat mass are important predictors of grade≥3 AAR.The OS of patients with sarcopenia and grade≥3 AAR may be reduced.

Artesunate inhibits development of esophageal squamous cell carcinoma via regulating expression of Skp2 and P21 / 中国药理学通报

Aim To clarify the regulatory effect of Artesunate(ART) on tumor cell function and cell cycle in the pathological process of esophageal squamous cell carcinoma(ESCC). Methods KYSE450 and TE14 cells were treated with different concentrations of ART. The cells treated with 0 mg •L

Study on the prognostic influencing factors of esophageal squamous cell carcinoma and the predictive value of inflammatory reaction indexes on its postoperative recurrence / 中华肿瘤杂志

Objective: To explore the influence factors of poor prognosis of esophageal squamous cell carcinoma (ESCC) and the predictive value of inflammatory reaction indexes including neutrophils and lymphocytes ratio (NLR), platelet and lymphocyte ratio (PLR), monocyte and lymphocyte ratio (MLR) provision and differentiation degree, infiltration depth, lymph node metastasis number on the postoperative recurrence of ESCC. Methods: A total of 130 patients with ESCC who underwent radical resection from February 2017 to February 2019 in Nanyang Central Hospital were selected and divided into good prognosis group (66 cases) and poor prognosis group (64 cases) according to the prognostic effect. The clinical data and follow-up data were collected. Multivariate logistic regression analysis was used to determine the independent influencing factors of poor prognosis. Spearman correlation analysis was used to determine the correlation between preoperative NLR, PLR and MLR with the degree of differentiation, depth of invasion and number of lymph node metastases. Receiver operating characteristic (ROC) curve analysis was used to evaluate the efficacy of NLR, PLR and MLR in predicting poor prognosis of ESCC. Results: Univariate analysis showed that the degree of differentiation, the degree of invasion and the number of lymph node metastasis were related to the prognoses of patients with ESCC (P<0.05). Multivariate logistic regression analysis showed that the degree of differentiation, depth of invasion and number of lymph node metastases were independent influencing factors for poor prognosis of patients with ESCC, moderate differentiation (OR=2.603, 95% CI: 1.009-6.715) or low differentiation (OR=9.909, 95% CI: 3.097-31.706), infiltrating into fibrous membrane (OR=14.331, 95% CI: 1.333-154.104) or surrounding tissue (OR=23.368, 95% CI: 1.466-372.578), the number of lymph node metastases ≥ 3 (OR=9.225, 95% CI: 1.693-50.263) indicated poor prognosis. Spearman correlation analysis showed that NLR was negatively correlated with the degree of differentiation and the number of lymph node metastases (r=-0.281, P=0.001; r=-0.257, P=0.003), PLR was negatively correlated with the degree of differentiation, depth of invasion and number of lymph node metastasis (r=-0.250, P=0.004; r=0.197, P=0.025; r=-0.194, P=0.027), MLR was positively correlated with the degree of differentiation and the number of lymph node metastasis (r=0.248, P=0.004; r=0.196, P=0.025). ROC curve analysis showed that the areas under the curve of NLR, PLR and MLR in predicting poor prognosis of ESCC were 0.971, 0.925 and 0.834, respectively. The best cut-off value of NLR was 2.87. The sensitivity and specificity of NLR in predicting poor prognosis of ESCC were 90.6% and 87.9%, respectively. The optimal cut-off value of PLR was 141.75. The sensitivity and specificity for predicting poor prognosis of ESCC were 92.2% and 87.9%, respectively. The best cut-off value of MLR was 0.40. The sensitivity and specificity of MLR in predicting poor prognosis of esophageal squamous cell carcinoma were 54.7% and 100.0%, respectively. Conclusions: The degree of differentiation, the degree of invasion and the number of lymph node metastases are closely related to the poor prognosis of patients with esophageal squamous cell carcinoma. NLR, PLR and MLR can provide important information for predicting the poor prognosis of esophageal squamous cell carcinoma.

Analysis of risk factors for depth of invasion and angiolymphatic invasion for circumferential superficial esophageal squamous cell carcinoma and precancerous lesion / 中华肿瘤杂志

Objective: To analyze clinicopathological features of circumferential superficial esophageal squamous cell carcinoma and precancerous lesions and investigate the risk factors for deep submucosal invasion and angiolymphatic invasion retrospectively. Methods: A total of 116 cases of esophageal squamous epithelial high-grade intraepithelial neoplasia or squamous cell carcinoma diagnosed by gastroscopy, biopsy pathology and endoscopic resection pathology during November 2013 to October 2021 were collected, and their clinicopathological features were analyzed. The independent risk factors of deep submucosal invasion and angiolymphatic invasion were analyzed by logistic regression model. Results: The multivariate logistic regression analysis showed that drinking history (OR=3.090, 95% CI: 1.165-8.200; P<0.05), The AB type of intrapapillary capillary loop (IPCL) (OR=11.215, 95% CI: 3.955-31.797; P<0.05) were the independent risk factors for the depth of invasion. The smoking history (OR=5.824, 95% CI: 1.704-19.899; P<0.05), the presence of avascular area (AVA) (OR=3.393, 95% CI: 1.285-12.072; P<0.05) were the independent factors for the angiolymphatic invasion. Conclusions: The risk of deep submucosal infiltration is greater for circumferential superficial esophageal squamous cell carcinoma patients with drinking history and IPCL type B2-B3 observed by magnifying endoscopy, while the risk of angiolymphatic invasion should be vigilant for circumferential superficial esophageal squamous cell carcinoma patients with smoking history and the presence of AVA observed by magnifying endoscopy. Ultrasound endoscopy combined with narrowband imagingand magnification endoscopy can improve the accuracy of preoperative assessment of the depth of infiltration of superficial squamous cell carcinoma and precancerous lesions and angiolymphaticinvasion in the whole perimeter of the esophagus, and help endoscopists to reasonably grasp the indications for endoscopic treatment.

Cost-utility analysis of tislelizumab monotherapy for second-line treatment of advanced or metastatic esophageal squamous cell carcinoma / 中国药房

OBJECTIVE To evaluate the cost-effectiveness of tislelizumab monotherapy in the second-line treatment of advanced or metastatic esophageal squamous cell carcinoma （ESCC），so as to provide reference for rational use of drug in clinic. METHODS A three-state partitioned survival model was constructed from the perspective of China’s health system， based on the data of RATIONALE-302 study，with simulation time limit of 10 years， cycle period of 1 month. The incremental cost-effectiveness ratio （ICER） was calculated with quality-adjusted life year （QALY） as utility index. The cost-effectiveness of tislelizumab monotherapy was compared with that of chemotherapy for second-line treatment of advanced or metastatic ESCC by cost-utility analysis. The stability of basic analysis results was validated through sensitivity analysis and scenario analysis. RESULTS The results of basic analysis showed that compared with chemotherapy group， incremental cost per capita of tislelizumab group was 35 025.32 yuan，and incremental utility per capita was 2.71 QALYs； ICER was 12 892.31 yuan/QALY， which was far lower than the willingness-to-pay （WTP） threshold of 3 times of China’s per capita gross domestic product （GDP） 242 928 yuan in 2021. The results of univariate sensitivity analysis showed that parameters such as the cost of apatinib， the utility value of disease progression status and the cost of adverse reactions in the chemotherapy group had a great impact on the ICER value， but these parameters could not cause the reversal of the basic analysis results. Probabilistic sensitivity analysis showed that WTP threshold was higher than 80 000 yuan/QALY，the probability of tislelizumab monotherapy possessed cost-effectiveness was 100%. Results of scenario analysis showed that in which model simulation time lasted for 5 or 20 years，ICER of tislelizumab was 8 331.00 yuan/QALY and 12 981.00 yuan/QALY， which were less than 3 times of China’s per capita GDP in 2021 as WTP threshold. CONCLUSIONS When three times of China’s GDP per capita in 2021 is taken as the WTP threshold， the second-line treatment of tirelizumab monotherapy for advanced or metastatic ESCC is more cost-effective than chemotherapy.

Cost-utility analysis of sintilimab combined with chemotherapy in first-line treatment of advanced，recurrent or metastatic esophageal squamous cell carcinoma / 中国药房

OBJECTIVE To evaluate the cost-effectiveness of sintilimab combined with chemotherapy than single-use chemotherapy in the first-line treatment of advanced， recurrent or metastatic esophageal squamous cell carcinoma （ESCC） from the perspective of health system of our country， and provide reference for rational use of drug in clinic. METHODS Based on ORIENT-15 study data， TreeAge Pro 2011 software was used to establish a three-state Markov model of non-progressive survival （PFS）， disease progression and death for cost-utility analysis. The model period was 3 weeks， the research time limit was 10 years， and the discount rate was 5%. The main outputs of the model were total cost， quality-adjusted life year （QALY） and incremental cost-effectiveness ratio （ICER）. The 1-3 times of China’s GDP per capita in 2021 was taken as the threshold of willing- ness to pay （WTP）. The uncertainty of the parameters was analyzed by single factor sensitivity analysis and probability sensitivity analysis， and the cost-effectiveness of the two schemes was discussed under three situations： different discount rates， comparison with other similar treatment schemes and charitable drug donation schemes. RESULTS The results of basic analysis showed that compared with chemotherapy plan alone， the ICER of sintilimab combined with chemotherapy was 64 208.75 yuan/QALY， which was less than WTP threshold. The results of single factor sensitivity analysis show that PFS state utility value， cycle cost of sintilimab and discount rate had relatively great influence on the results. Probability sensitivity analysis showed that when WTP≥120 000 yuan， the economic probability of sintilimab combined with chemotherapy plan was 100%. The results of situational analysis showed that sintilimab combined chemotherapy was more cost- effective than single-use chemotherapy. CONCLUSIONS Sintilimab combined with chemotherapy is more cost-effective than single-use chemotherapy in the first-line treatment of advanced， recurrent or metastatic ESCC.

Cost-effectiveness analysis of serplulimab combined with chemotherapy as first-line treatment for advanced esophageal squamous cell carcinoma / 中国药房

OBJECTIVE To evaluate the cost-effectiveness of serplulimab combined with chemotherapy as first-line treatment for advanced esophageal squamous cell carcinoma from the perspective of the Chinese healthcare system. METHODS A partitioned survival model with three health states was constructed for cost-effectiveness analysis. Clinical data were extracted from ASTRUM- 007. Information on parameters such as cost and health utility was derived from related websites and published literature. The quality-adjusted life years （QALYs） was used as the output index to calculate the incremental cost-effectiveness ratio （ICER）， and then compared with three times the per capita gross domestic product （GDP） in China to judge whether it was cost-effective. One- way sensitivity analysis and probabilistic sensitivity analysis were performed to evaluate the robustness of the model； the cost- effectiveness of applying this plan to subgroup patients with programmed cell death-ligand 1 combined positive score （PD-L1 CPS） ≥10 and the scheme in the context of charitable drug donations was explored. RESULTS Among advanced or metastatic oesophageal squamous cell carcinoma patients and patients with PD-L1 CPS ≥10， serplulimab combined with chemotherapy could improve health outcomes with an augmentation of cost， compared with placebo combined with chemotherapy，resulting the ICERs were 599 623.64 yuan/QALY and 629 121.57 yuan/QALY， respectively. Therefore， serplulimab combined with chemotherapy was not cost-effective. Sensitivity analysis of single factor showed that the costs of serplulimab were the crucial factor affecting the ICER； probabilistic sensitivity analysis demonstrated basic analysis results were relatively robust. The results of scenario analysis showed that when all patients met the requirements of the charitable drug donation program， serplulimab combined with chemotherapy was cost-effective； the economic outcome of this scheme was reversed compared with the results of the basic analysis. CONCLUSIONS From Chinese healthcare perspective， first-line treatment with serplulimab in combination with chemotherapy is not a cost-effective option for patients with advanced esophageal squamous cell carcinoma， but it may be an economic option to implement a charitable drug donation program for all patients or if the price of serplulimab is significantly reduced.

Immune-related Features of Esophageal Squamous Cell Carcinoma：A Multi-omics Perspective / 中山大学学报(医学科学版)

Esophageal squamous cell carcinoma is a common malignancy in the Asia-Pacific region， especially in China， where the morbidity remains high in spite of the improved overall survival due to advances in medical technology. Immunotherapy becomes a hot spot in recent tumor research when it has provided significant survival benefits in patients with advanced malignant tumors， such as lung cancer， breast cancer， colon cancer， etc. In esophageal squamous cell carcinoma， immunotherapy promotes survival benefit as well. However， because of the complex and changeable biological functions and gene expression regulation of malignant tumors， the conclusions based on a single-omics analysis are often incomprehensive. Currently， most of the immune-related studies on esophageal squamous cell carcinoma are still confined to a single-omics study like genomics， with limitations and one-sidedness. Since multi-omics analysis helps us better understand tumors from a wider and deeper perspective， this review explores and summarizes immune-related features of esophageal squamous cell carcinoma from a multi-omics perspective.

Clinical Significance of High Expression of Endoplasmic Reticulum Stress-related Proteins Glucose-regulating Protein 78 and X-box Binding Protein 1 induced by Fusobacterium Nucleatum in Esophageal Squamous Cell Carcinoma / 中山大学学报(医学科学版)

ObjectiveTo analyze the induction effect of Fusobacterium nucleatum （Fn） on endoplasmic reticulum stress-related proteins Glucose-regulating protein 78（GRP78） and X-box binding protein 1（XBP1） in esophageal squamous cell carcinoma （ESCC）， and to explore its potential mechanism and clinical significance. MethodsESCC cells KYSE150 and KYSE140 were infected with Fn for 12 h， 24 h and 48 h. The oxidative stress indexes （ROS， MDA and SOD） and the expression of GRP78 and XBP1 in each group were detected by oxidative stress index kit and Western blot. The experiment was divided into Fn groups， Fn+siNC1 groups， Fn+siGRP78 groups， Fn+siNC2 groups and Fn+siXBP1 groups； the oxidative stress indexes， paclitaxel （PTX） response efficacy， abilities of proliferation， invasion and metastasis in each group were compared. The infection of Fn and the expression of GRP78 and XBP1 in 234 ESCC and paracancerous tissues were detected by RNA scope and immunohistochemistry. The correlation between each factor and clinicopathological characteristics of patients was analyzed by Chi-square test. The influence of each factor on the survival of patients was compared by Kaplan-meier survival estimate. ResultsCompared with Fn uninfected KYSE150 and KYSE140 cells， the content of ROS and MDA was gradually increased， the activity of SOD was gradually decreased， and the expression of GRP78 and XBP1 was gradually increased in Fn infected groups （12 h， 24 h and 48 h） （P < 0.05）. Compared with Fn groups， Fn+siNC1 groups， and Fn+siNC2 groups， ROS and MDA contents were decreased， SOD activity was increased， PTX response efficacy was enhanced， and abilities of proliferation， invasion and metastasis were decreased in Fn+siGRP78 and Fn+siXBP1 groups （P < 0.05）. The rates of Fn， GRP78 and XBP1 in ESCC tissues were 43.16%， 69.66% and 60.68%， respectively. And the three indexes were significantly consistent （P < 0.05）. The patients with positive Fn infection and high expression of GRP78 and XBP1 were mostly males with a history of smoking and drinking， and the tumor differentiation degree was low， the invasion degree was deep， the lymph node metastasis rate was high， and the clinical stage was mostly stage Ⅲ/Ⅳ. The 5-year survival time of patients with above positive indexes was shortened （P < 0.05）. ConclusionsFn could induce endoplasmic reticulum stress by inducing the high expression of GRP78 and XBP1， and promote the malignant evolution of ESCC.

PAFR/Stat3 axis maintains the symbiotic ecosystem between tumor and stroma to facilitate tumor malignancy

Stroma surrounding the tumor cells plays crucial roles for tumor progression. However, little is known about the factors that maintain the symbiosis between stroma and tumor cells. In this study, we found that the transcriptional regulator-signal transducer and activator of transcription 3 (Stat3) was frequently activated in cancer-associated fibroblasts (CAFs), which was a potent facilitator of tumor malignancy, and formed forward feedback loop with platelet-activating factor receptor (PAFR) both in CAFs and tumor cells. Importantly, PAFR/Stat3 axis connected intercellular signaling crosstalk between CAFs and cancer cells and drove mutual transcriptional programming of these two types of cells. Two central Stat3-related cytokine signaling molecules-interleukin 6 (IL-6) and IL-11 played the critical role in the process of PAFR/Stat3 axis-mediated communication between tumor and CAFs. Pharmacological inhibition of PAFR and Stat3 activities effectively reduced tumor progression using CAFs/tumor co-culture xenograft model. Our study reveals that PAFR/Stat3 axis enhances the interaction between tumor and its associated stroma and suggests that targeting this axis can be an effective therapeutic strategy against tumor malignancy.

GDF15 negatively regulates chemosensitivity via TGFBR2-AKT pathway-dependent metabolism in esophageal squamous cell carcinoma / 医学前沿

Treating patients with esophageal squamous cell carcinoma (ESCC) is challenging due to the high chemoresistance. Growth differentiation factor 15 (GDF15) is crucial in the development of various types of tumors and negatively related to the prognosis of ESCC patients according to our previous research. In this study, the link between GDF15 and chemotherapy resistance in ESCC was further explored. The relationship between GDF15 and the chemotherapy response was investigated through in vitro and in vivo studies. ESCC patients with high levels of GDF15 expression showed an inferior chemotherapeutic response. GDF15 improved the tolerance of ESCC cell lines to low-dose cisplatin by regulating AKT phosphorylation via TGFBR2. Through an in vivo study, we further validated that the anti-GDF15 antibody improved the tumor inhibition effect of cisplatin. Metabolomics showed that GDF15 could alter cellular metabolism and enhance the expression of UGT1A. AKT and TGFBR2 inhibition resulted in the reversal of the GDF15-induced expression of UGT1A, indicating that TGFBR2-AKT pathway-dependent metabolic pathways were involved in the resistance of ESCC cells to cisplatin. The present investigation suggests that a high level of GDF15 expression leads to ESCC chemoresistance and that GDF15 can be targeted during chemotherapy, resulting in beneficial therapeutic outcomes.

The role of B3GNT3 as an oncogene in the growth, invasion and migration of esophageal cancer cells

Purpose: To investigate the role and mechanism of ß1,3-N-acetylglucosaminyltransferase-3 gene (B3GNT3) in esophageal cancer (ESCA). Methods: The starBase database was used to evaluate the expression of B3GNT3. B3GNT3 function was measured using KYSE-30 and KYSE-410 cells of esophageal squamous cell carcinoma (ESCC) cell lines. The mRNA levels were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Cell counting kit-8, clone formation assay and transwell assay were used to detect the changes of proliferation, invasion and migration. Results: B3GNT3 expression was higher in ESCA tissues than in normal tissues. The overall survival rate of ESCA patients with high B3GNT3 expression was lower than that of ESCA patients with low B3GNT3 expression. In vitro functional experiments showed that the proliferation ability, migration and invasion ability of KYSE-30 and KYSE-410 cells with B3GNT3 interference were lower than those of the control, and the overexpression of B3GNT3 had the opposite effect. After silencing B3GNT3 expression in ESCC cell lines, the growth of both cell lines was inhibited and the invasiveness was decreased. Knockdown of B3GNT3 reduced the growth rate and Ki-67 expression level. Conclusion: B3GNT3, as an oncogene, may promote the growth, invasion and migration of ESCC cell.

Effect of Small-molecule Inhibitor CIL56 on Death of Esophageal Squamous Cell Carcinoma Cells / 肿瘤防治研究

Objective To investigate the role and mechanism of the small-molecule inhibitor CIL56 in the death of esophageal squamous cell carcinoma cells. Methods SRB method and plate-cloning method were used to detect the effect of CIL56 on the proliferation of esophageal squamous cell carcinoma. The effect of CIL56 on the migration of esophageal squamous cell carcinoma cells was investigated by scratch-healing test. The effect of CIL56 on the concentration of iron ions in esophageal squamous cell carcinoma was detected with an iron-detection kit. A total glutathione test kit was used to examine the effect of CIL56 on glutathione concentration in esophageal squamous cell carcinoma. Western blot was used to investigate the effect of CIL56 on the expression of xCT and GPX4 proteins related to iron death, as well as YAP1 protein, in esophageal squamous cell carcinoma. Results CIL56 could significantly inhibit the proliferation (P < 0.05) and migration (P < 0.001) of esophageal squamous cell carcinoma. With the increased CIL56 concentration, the iron concentration in esophageal squamous cell carcinoma increased (P < 0.05). CIL56 could reduce the glutathione content in esophageal squamous cell carcinoma (P < 0.01). CIL56 could reduce the expression of xCT and GPX4 proteins related to iron death and decrease the level of YAP1 protein in esophageal squamous cell carcinoma (both P < 0.001). Conclusion The small-molecule inhibitor CIL56 can significantly inhibit the proliferation and migration of esophageal squamous cell carcinoma cells and reduce the expression of the iron-death-related proteins xCT and GPX4, as well as YAP1 protein.

Repurposed benzydamine targeting CDK2 suppresses the growth of esophageal squamous cell carcinoma / 医学前沿

Esophageal squamous cell carcinoma (ESCC) is one of the leading causes of cancer death worldwide. It is urgent to develop new drugs to improve the prognosis of ESCC patients. Here, we found benzydamine, a locally acting non-steroidal anti-inflammatory drug, had potent cytotoxic effect on ESCC cells. Benzydamine could suppress ESCC proliferation in vivo and in vitro. In terms of mechanism, CDK2 was identified as a target of benzydamine by molecular docking, pull-down assay and in vitro kinase assay. Specifically, benzydamine inhibited the growth of ESCC cells by inhibiting CDK2 activity and affecting downstream phosphorylation of MCM2, c-Myc and Rb, resulting in cell cycle arrest. Our study illustrates that benzydamine inhibits the growth of ESCC cells by downregulating the CDK2 pathway.

Comparison of short-term efficacy of neoadjuvant immunotherapy combined with chemotherapy and neoadjuvant chemoradiation for locally advanced resectable esophageal squamous cell carcinoma / 中国胸心血管外科临床杂志

@#Objective    To investigate the short-term therapeutic effect of neoadjuvant immunotherapy combined with chemotherapy in the locally advanced esophageal squamous cell carcinoma. Methods    The clinical data of patients with esophageal squamous cell carcinoma treated with neoadjuvant treatment in Gaozhou People's Hospital from August 2019 to October 2020 were retrospectively analyzed. According to the different treatments, the patients were divided into two groups: a neoadjuvant immunotherapy combined with chemotherapy group (NIC group) and a neoadjuvant chemoradiotherapy group (NC group). The baseline data, incidence of adverse events during treatment, perioperative indicators, postoperative pathological remission rate and incidence of postoperative complications were compared between the two groups. Results    Totally 33 patients were enrolled, including 15 males and 18 females, with an average age of 62.37±7.99 years. There were 17 patients in the NIC group and 16 patients in the NC group. In the NIC group, the carcinoma was mainly located in the middle and lower esophagus, with 5 paitents in stage Ⅱ, 9 patients in stage Ⅲ, and 3 patients in stage Ⅳa. In the NC group, the carcinoma was mainly located in the upper-middle esophagus, with 1 patient in stage Ⅱ and 15 patients in stage Ⅲ. During the neoadjuvant treatment, there was no significant difference in the occurrence of bone marrow suppression or gastrointestinal reactions between the two groups (P>0.05). There were 4 immune-related rashes in the NIC group and 1 esophageal perforation in the NC group. Fourteen (82.35%) patients in the NIC group and 12 (75.00%) patients in the NC group completed the operation on schedule. The postoperative ICU stay time and chest tube indwelling time in the NIC group were shorter than those in the NC group (P<0.05). There were 5 patients of complete remission in the NIC group, and 6 patients in the NC group. There was no significant difference in the pathological regression grade or residual tumor cells between the two groups (P>0.05). There was no significant difference in the incidence of anastomotic fistula, thoracic gastric fistula, bronchial mediastinal fistula, abdominal distension, pulmonary infection, stroke, or hoarseness during the perioperative period between the two groups of patients who completed the operation (P>0.05). In the NC group, 2 patients died during the perioperative period because of thoracic gastric fistula complicated by severe infection. Conclusion    Neoadjuvant immunotherapy combined with chemotherapy dose not significantly increase the occurrence of adverse events and shows a good rate of pathological remission, which indicates that the neoadjuvant immunotherapy combined with chemotherapy is a safe, feasible and potential new treatment model.

Analysis of immune microenvironment and potential sensitive drugs in esophageal squamous cell carcinoma based on GEO database and bioinformatics method / 中国胸心血管外科临床杂志

@#Objective    To construct a prognostic model of esophageal squamous cell carcinoma (ESCC) based on immune checkpoint-related genes and explore the potential relationship between these genes and the tumor microenvironment (TME). Methods     The transcriptome sequencing data and clinical information of immune checkpoint genes of samples from GSE53625 in GEO database were collected. The difference of gene expression between ESCC and normal paracancerous tissues was evaluated, and the drug sensitivity of differentially expressed genes in ESCC was analyzed. We then constructed a risk model based on survival-related genes and explored the prognostic characteristics, enriched pathway, immune checkpoints, immune score, immune cell infiltration, and potentially sensitive drugs of different risk groups. Results    A total of 358 samples from 179 patients were enrolled, including 179 ESCC samples and 179 corresponding paracancerous tissues. There were 33 males and 146 females, including 80 patients≤60 years and 99 patients>60 years. 39 immune checkpoint genes were differentially expressed in ESCC, including 14 low expression genes and 25 high expression genes. Drug sensitivity analysis of 8 highly expressed genes (TNFRSF8, CTLA4, TNFRSF4, CD276, TNFSF4, IDO1, CD80, TNFRSF18) showed that many compounds were sensitive to these immunotherapy targets. A risk model based on three prognostic genes (NRP1, ICOSLG, HHLA2) was constructed by the least absolute shrinkage and selection operator analysis. It was found that the overall survival time of the high-risk group was significantly lower than that of the low-risk group (P<0.001). Similar results were obtained in different ESCC subtypes. The risk score based on the immune checkpoint gene was identified as an independent prognostic factor for ESCC. Different risk groups had unique enriched pathways, immune cell infiltration, TME, and sensitive drugs. Conclusion     A prognostic model based on immune checkpoint gene is established, which can accurately stratify ESCC and provide potential sensitive drugs for ESCC with different risks, thus providing a possibility for personalized treatment of ESCC.

Relationship between DDX46 gene and invasion and metastasis behaviors of TE-1 esophageal squamous cell carcinoma cells / 中国胸心血管外科临床杂志

@#Objective    To investigate the relationship between DDX46 genes and invasion and migration of esophageal squamous cell carcinoma cells. Methods    Human esophageal squamous cell carcinoma cells TE-1 were transfected by fluorescent marker shRNA lentivirus (shDDX46 group), and an empty vector was transfected as a control (shCtrl group). The expression rate of green fluorescent protein under the microscope was used to evaluate the cell transfection efficiency. Real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR) and Western blotting (WB) detected the knockdown efficiency of the target gene at the mRNA and protein expression levels. Wound healing, invasion assay and migration assay detected the changes of invasion and metastasis ability. Classical pathway analysis was used to explore signaling pathway changes and the possible mechanism of DDX46 in the invasion and metastasis was explored by detecting fibronectin expression. Results    DDX46 gene at mRNA and protein levels was significantly inhibited after lentiviral transfection. Wound healing showed that after 8 h the cell mobility of TE-1 cells decreased significantly (P=0.001). Invasion assay showed that after 24 h the average cell metastasis rate of TE-1 cells was lower in the shDDX46 group than that in the shCtrl group (P<0.001). The cell metastasis rate in the shDDX46 group corresponding to observation points in the transwell assay was lower than that in the shCtrl group (P<0.001) after 24 h culture. The results of the classical pathway analysis showed that the integrin signaling pathway activity was inhibited, further exploration of the mechanism of action found that the expression of fibronectin associated with cell adhesion was decreased. Conclusion    DDX46 gene is related to the invasion and migration ability of esophageal squamous cell carcinoma cells. Knockdown of DDX46 genes may reduce cell adhesion by downregulating the integrin pathway signaling.