Background & aims: Patients with intrahepatic cholangiocarcinoma (iCCA) respond poorly to immune checkpoint blockades (ICBs). In this study, we aimed to dissect the potential mechanisms underlying poor response to ICBs and explore a rational ICB-based combination therapy in iCCA.
Methods: scRNA-seq dataset GSE151530 was analyzed to investigate the differentially expressed genes in malignant cells following ICBs therapy. RNA-seq analysis and western blot assays were performed to examine the upstream and downstream signaling pathways of CD73. Subcutaneous tumor xenograft models were utilized to investigate the impact of CD73 on iCCA growth. Plasmid AKT/NICD-induced spontaneous murine iCCAs were used to explore the therapeutic efficacy of CD73 enzymatic inhibitor AB680 combined with PD-1 blockade. Time-of-flight mass cytometry (CyTOF) was conducted to identify the tumor-infiltrating immune cell populations and their functional changes in murine iCCAs treated with AB680 in combination with PD-1 antibody.
Results: scRNA-seq analysis identified elevated CD73 expression in malignant cells in response to ICBs therapy. Mechanistically, ICBs therapy upregulated CD73 expression in malignant cells via TNF-α/NF-κB signaling pathway. In vivo studies revealed that CD73 inhibition suppressed the growth of subcutaneous tumors, and achieved synergistic depression effects with gemcitabine and cisplatin (GC). Adenosine produced by CD73 activates AKT/GSK3β/β-catenin signaling axis in iCCA cells. CD73 inhibitor AB680 potentiates anti-tumor efficacy of PD-1 antibody in murine iCCAs. CyTOF analysis showed that AB680 combined with anti-PD-1 therapy promoted the infiltration of CD8+ T, CD4+ T cells, and NK cells in murine iCCAs, while simultaneously decreased the proportions of macrophages and neutrophils. Moreover, AB680 combined with anti-PD-1 significantly upregulated the expression of Granzyme B, Tbet and co-stimulatory molecule ICOS in infiltrating CD8+ T cells.
Conclusions: CD73 inhibitor AB680 limits tumor progression and potentiates therapeutic efficacy of GC chemotherapy or anti-PD-1 treatment in iCCA. AB680 combined with anti-PD-1 therapy effectively elicits anti-tumor immune response.
{"title":"Targeting CD73 limits tumor progression and enhances anti-tumor activity of anti-PD-1 therapy in intrahepatic cholangiocarcinoma.","authors":"Bao-Ye Sun, Dai Zhang, Wei Gan, Jing-Fang Wu, Zhu-Tao Wang, Guo-Qiang Sun, Jian Zhou, Jia Fan, Yong Yi, Bo Hu, Bo-Heng Zhang, Shuang-Jian Qiu","doi":"10.1007/s00432-024-05869-1","DOIUrl":"10.1007/s00432-024-05869-1","url":null,"abstract":"<p><strong>Background & aims: </strong>Patients with intrahepatic cholangiocarcinoma (iCCA) respond poorly to immune checkpoint blockades (ICBs). In this study, we aimed to dissect the potential mechanisms underlying poor response to ICBs and explore a rational ICB-based combination therapy in iCCA.</p><p><strong>Methods: </strong>scRNA-seq dataset GSE151530 was analyzed to investigate the differentially expressed genes in malignant cells following ICBs therapy. RNA-seq analysis and western blot assays were performed to examine the upstream and downstream signaling pathways of CD73. Subcutaneous tumor xenograft models were utilized to investigate the impact of CD73 on iCCA growth. Plasmid AKT/NICD-induced spontaneous murine iCCAs were used to explore the therapeutic efficacy of CD73 enzymatic inhibitor AB680 combined with PD-1 blockade. Time-of-flight mass cytometry (CyTOF) was conducted to identify the tumor-infiltrating immune cell populations and their functional changes in murine iCCAs treated with AB680 in combination with PD-1 antibody.</p><p><strong>Results: </strong>scRNA-seq analysis identified elevated CD73 expression in malignant cells in response to ICBs therapy. Mechanistically, ICBs therapy upregulated CD73 expression in malignant cells via TNF-α/NF-κB signaling pathway. In vivo studies revealed that CD73 inhibition suppressed the growth of subcutaneous tumors, and achieved synergistic depression effects with gemcitabine and cisplatin (GC). Adenosine produced by CD73 activates AKT/GSK3β/β-catenin signaling axis in iCCA cells. CD73 inhibitor AB680 potentiates anti-tumor efficacy of PD-1 antibody in murine iCCAs. CyTOF analysis showed that AB680 combined with anti-PD-1 therapy promoted the infiltration of CD8<sup>+</sup> T, CD4<sup>+</sup> T cells, and NK cells in murine iCCAs, while simultaneously decreased the proportions of macrophages and neutrophils. Moreover, AB680 combined with anti-PD-1 significantly upregulated the expression of Granzyme B, Tbet and co-stimulatory molecule ICOS in infiltrating CD8<sup>+</sup> T cells.</p><p><strong>Conclusions: </strong>CD73 inhibitor AB680 limits tumor progression and potentiates therapeutic efficacy of GC chemotherapy or anti-PD-1 treatment in iCCA. AB680 combined with anti-PD-1 therapy effectively elicits anti-tumor immune response.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11246275/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141603619","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-13DOI: 10.1007/s00432-024-05876-2
Junhao Zhang, Ruiqing Liu, Xujian Wang, Shiwei Zhang, Lizhi Shao, Junheng Liu, Jiahui Zhao, Quan Wang, Jie Tian, Yun Lu
Purpose: Neoadjuvant chemoradiotherapy has been the standard practice for patients with locally advanced rectal cancer. However, the treatment response varies greatly among individuals, how to select the optimal candidates for neoadjuvant chemoradiotherapy is crucial. This study aimed to develop an endoscopic image-based deep learning model for predicting the response to neoadjuvant chemoradiotherapy in locally advanced rectal cancer.
Methods: In this multicenter observational study, pre-treatment endoscopic images of patients from two Chinese medical centers were retrospectively obtained and a deep learning-based tumor regression model was constructed. Treatment response was evaluated based on the tumor regression grade and was defined as good response and non-good response. The prediction performance of the deep learning model was evaluated in the internal and external test sets. The main outcome was the accuracy of the treatment prediction model, measured by the AUC and accuracy.
Results: This deep learning model achieved favorable prediction performance. In the internal test set, the AUC and accuracy were 0.867 (95% CI: 0.847-0.941) and 0.836 (95% CI: 0.818-0.896), respectively. The prediction performance was fully validated in the external test set, and the model had an AUC of 0.758 (95% CI: 0.724-0.834) and an accuracy of 0.807 (95% CI: 0.774-0.843).
Conclusion: The deep learning model based on endoscopic images demonstrated exceptional predictive power for neoadjuvant treatment response, highlighting its potential for guiding personalized therapy.
{"title":"Deep learning model based on endoscopic images predicting treatment response in locally advanced rectal cancer undergo neoadjuvant chemoradiotherapy: a multicenter study.","authors":"Junhao Zhang, Ruiqing Liu, Xujian Wang, Shiwei Zhang, Lizhi Shao, Junheng Liu, Jiahui Zhao, Quan Wang, Jie Tian, Yun Lu","doi":"10.1007/s00432-024-05876-2","DOIUrl":"10.1007/s00432-024-05876-2","url":null,"abstract":"<p><strong>Purpose: </strong>Neoadjuvant chemoradiotherapy has been the standard practice for patients with locally advanced rectal cancer. However, the treatment response varies greatly among individuals, how to select the optimal candidates for neoadjuvant chemoradiotherapy is crucial. This study aimed to develop an endoscopic image-based deep learning model for predicting the response to neoadjuvant chemoradiotherapy in locally advanced rectal cancer.</p><p><strong>Methods: </strong>In this multicenter observational study, pre-treatment endoscopic images of patients from two Chinese medical centers were retrospectively obtained and a deep learning-based tumor regression model was constructed. Treatment response was evaluated based on the tumor regression grade and was defined as good response and non-good response. The prediction performance of the deep learning model was evaluated in the internal and external test sets. The main outcome was the accuracy of the treatment prediction model, measured by the AUC and accuracy.</p><p><strong>Results: </strong>This deep learning model achieved favorable prediction performance. In the internal test set, the AUC and accuracy were 0.867 (95% CI: 0.847-0.941) and 0.836 (95% CI: 0.818-0.896), respectively. The prediction performance was fully validated in the external test set, and the model had an AUC of 0.758 (95% CI: 0.724-0.834) and an accuracy of 0.807 (95% CI: 0.774-0.843).</p><p><strong>Conclusion: </strong>The deep learning model based on endoscopic images demonstrated exceptional predictive power for neoadjuvant treatment response, highlighting its potential for guiding personalized therapy.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11246300/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141603616","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-11DOI: 10.1007/s00432-024-05841-z
Valeria Merz, Francesca Maines, Stefano Marcucci, Chiara Sartori, Michela Frisinghelli, Chiara Trentin, Dzenete Kadrija, Francesco Giuseppe Carbone, Andrea Michielan, Armando Gabbrielli, Davide Melisi, Mattia Barbareschi, Alberto Brolese, Orazio Caffo
Background: Therapeutic approach used for pancreatic ductal adenocarcinoma is usually translated also for the rarer acinar counterpart, which shows a different mutational landscape nevertheless. While dMMR/MSI-H status is rare in the ductal histotype, it appears to be more prevalent in pancreatic acinar cell carcinoma (PACC).
Case presentation: We report the case of a patient with locally advanced MSI-H PACC in whom the treatment with the anti-PD-1 pembrolizumab, administered as third line, made possible surgical resection, achieving even an exceptional pathological complete response.
Conclusions: Treatment of PACC should be tailored based on the peculiar molecular features that distinguish PACC from ductal adenocarcinoma. Evaluation of potentially therapeutically targetable alterations should be mandatory in case of PACC diagnosis.
{"title":"Complete pathological response to pembrolizumab in pretreated pancreatic acinar cell carcinoma.","authors":"Valeria Merz, Francesca Maines, Stefano Marcucci, Chiara Sartori, Michela Frisinghelli, Chiara Trentin, Dzenete Kadrija, Francesco Giuseppe Carbone, Andrea Michielan, Armando Gabbrielli, Davide Melisi, Mattia Barbareschi, Alberto Brolese, Orazio Caffo","doi":"10.1007/s00432-024-05841-z","DOIUrl":"10.1007/s00432-024-05841-z","url":null,"abstract":"<p><strong>Background: </strong>Therapeutic approach used for pancreatic ductal adenocarcinoma is usually translated also for the rarer acinar counterpart, which shows a different mutational landscape nevertheless. While dMMR/MSI-H status is rare in the ductal histotype, it appears to be more prevalent in pancreatic acinar cell carcinoma (PACC).</p><p><strong>Case presentation: </strong>We report the case of a patient with locally advanced MSI-H PACC in whom the treatment with the anti-PD-1 pembrolizumab, administered as third line, made possible surgical resection, achieving even an exceptional pathological complete response.</p><p><strong>Conclusions: </strong>Treatment of PACC should be tailored based on the peculiar molecular features that distinguish PACC from ductal adenocarcinoma. Evaluation of potentially therapeutically targetable alterations should be mandatory in case of PACC diagnosis.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11239721/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141579759","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-09DOI: 10.1007/s00432-024-05860-w
Xiao-Hui Zhang, Shu-Wei Wu, Yi-Fan Feng, Yang-Qin Xie, Min Li, Ping Hu, Yunxia Cao
Background: Endometrial cancer (EC) is the sixth most frequent cancer in women worldwide and has higher fatality rates. The pathophysiology of EC is complex, and there are currently no reliable methods for diagnosing and treating the condition. Long non-coding RNA (lncRNA), according to mounting evidence, is vital to the pathophysiology of EC. HOTAIR is regarded as a significant prognostic indicator of EC. ZBTB7A decreased EC proliferation and migration, according to recent studies, however the underlying mechanism still needs to be clarified.
Methods: The research utilized RT-qPCR to measure HOTAIR expression in clinical EC tissues and various EC cell lines. Kaplan-Meier survival analysis was employed to correlate HOTAIR levels with patient prognosis. Additionally, the study examined the interaction between ZBTB7A and HOTAIR using bioinformatics tools and ChIP assays. The experimental approach also involved manipulating the expression levels of HOTAIR and ZBTB7A in EC cell lines and assessing the impact on various cellular processes and gene expression.
Results: The study found significantly higher levels of HOTAIR in EC tissues compared to adjacent normal tissues, with high HOTAIR expression correlating with poorer survival rates and advanced cancer characteristics. EC cell lines like HEC-1 A and KLE showed higher HOTAIR levels compared to normal cells. Knockdown of HOTAIR in these cell lines reduced proliferation, angiogenesis, and migration. ZBTB7A was found to be inversely correlated with HOTAIR, and its overexpression led to a decrease in HOTAIR levels and a reduction in malignant cell behaviors. The study also uncovered that HOTAIR interacts with ELAVL1 to regulate SOX17, which in turn activates the Wnt/β-catenin pathway, promoting malignant behaviors in EC cells.
Conclusion: HOTAIR is a critical regulator in EC, contributing to tumor growth and poor prognosis. Its interaction with ZBTB7A and regulation of SOX17 via the Wnt/β-catenin pathway underlines its potential as a therapeutic target.
{"title":"ZBTB7A regulates LncRNA HOTAIR-mediated ELAVL1/SOX17 axis to inhibit malignancy and angiogenesis in endometrial carcinoma.","authors":"Xiao-Hui Zhang, Shu-Wei Wu, Yi-Fan Feng, Yang-Qin Xie, Min Li, Ping Hu, Yunxia Cao","doi":"10.1007/s00432-024-05860-w","DOIUrl":"10.1007/s00432-024-05860-w","url":null,"abstract":"<p><strong>Background: </strong>Endometrial cancer (EC) is the sixth most frequent cancer in women worldwide and has higher fatality rates. The pathophysiology of EC is complex, and there are currently no reliable methods for diagnosing and treating the condition. Long non-coding RNA (lncRNA), according to mounting evidence, is vital to the pathophysiology of EC. HOTAIR is regarded as a significant prognostic indicator of EC. ZBTB7A decreased EC proliferation and migration, according to recent studies, however the underlying mechanism still needs to be clarified.</p><p><strong>Methods: </strong>The research utilized RT-qPCR to measure HOTAIR expression in clinical EC tissues and various EC cell lines. Kaplan-Meier survival analysis was employed to correlate HOTAIR levels with patient prognosis. Additionally, the study examined the interaction between ZBTB7A and HOTAIR using bioinformatics tools and ChIP assays. The experimental approach also involved manipulating the expression levels of HOTAIR and ZBTB7A in EC cell lines and assessing the impact on various cellular processes and gene expression.</p><p><strong>Results: </strong>The study found significantly higher levels of HOTAIR in EC tissues compared to adjacent normal tissues, with high HOTAIR expression correlating with poorer survival rates and advanced cancer characteristics. EC cell lines like HEC-1 A and KLE showed higher HOTAIR levels compared to normal cells. Knockdown of HOTAIR in these cell lines reduced proliferation, angiogenesis, and migration. ZBTB7A was found to be inversely correlated with HOTAIR, and its overexpression led to a decrease in HOTAIR levels and a reduction in malignant cell behaviors. The study also uncovered that HOTAIR interacts with ELAVL1 to regulate SOX17, which in turn activates the Wnt/β-catenin pathway, promoting malignant behaviors in EC cells.</p><p><strong>Conclusion: </strong>HOTAIR is a critical regulator in EC, contributing to tumor growth and poor prognosis. Its interaction with ZBTB7A and regulation of SOX17 via the Wnt/β-catenin pathway underlines its potential as a therapeutic target.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11233420/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141563449","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-09DOI: 10.1007/s00432-024-05866-4
Qiujin Chen, Kai Li, Yang Liu, Xiaozhai Yu, Fengrong Ou
Purpose: Colorectal cancer (CRC) is a common malignancy that affects adults worldwide, causing a high disease burden. Few studies have examined the relationship between body composition (BC) measures and the prevalence of CRC. Our purpose was to investigate the relationship between pertinent BC indicators and CRC.
Methods: Bioelectrical impedance analysis, laboratory test results, face-to-face questionnaire investigation, and nutritional risk assessment (Nutritional Risk Screening 2002 and Patient-Generated Subjective Global Assessment) were used in this case-control study. Bioelectrical impedance analysis in the case group was performed prior to antitumor therapy/surgery.
Results: From June 2018 to January 2019, a total of 303 cases and 286 controls were included. The results showed that low body fat percentage (BFP) and high visceral adiposity index (VAI) groups had a higher risk of developing CRC in comparison to the normal BFP and normal VAI groups. The risk of CRC decreased with the increase of BFP. The group with a normal BC had a lower risk of developing CRC compared to those with a greater VAI and a lower BFP, as indicated by the results of the pairwise and total combinations of VAI, fat-free mass index (FFMI), and BFP. Additionally, FFMI and VAI had positive correlations with prealbumin, serum albumin, and nutritional risk scores.
Conclusion: Low BFP and high VAI are associated with higher CRC risk. FFMI and VAI are positively correlated with prealbumin, serum albumin, and nutritional risk scores in CRC patients.
目的:结直肠癌(CRC)是一种常见的恶性肿瘤,影响着全世界的成年人,造成了沉重的疾病负担。很少有研究探讨身体成分(BC)指标与 CRC 发病率之间的关系。我们的目的是调查相关的 BC 指标与 CRC 之间的关系:这项病例对照研究采用了生物电阻抗分析、实验室检测结果、面对面问卷调查和营养风险评估(2002 年营养风险筛查和患者自发主观全面评估)。病例组的生物电阻抗分析在抗肿瘤治疗/手术前进行:2018年6月至2019年1月,共纳入303例病例和286例对照。结果显示,与正常体脂率(BFP)和正常内脏脂肪指数(VAI)组相比,低体脂率(BFP)和高内脏脂肪指数(VAI)组患 CRC 的风险更高。随着 BFP 的增加,患 CRC 的风险降低。VAI、去脂质量指数(FFMI)和BFP的成对组合和总组合结果表明,与VAI较高和BFP较低的人群相比,BC正常的人群患上CRC的风险较低。此外,FFMI 和 VAI 与前白蛋白、血清白蛋白和营养风险评分呈正相关:结论:低 BFP 和高 VAI 与较高的 CRC 风险相关。结论:低 BFP 和高 VAI 与较高的 CRC 风险有关,FFMI 和 VAI 与 CRC 患者的前白蛋白、血清白蛋白和营养风险评分呈正相关。
{"title":"Association of body composition indicators with colorectal cancer: a hospital-based case-control study.","authors":"Qiujin Chen, Kai Li, Yang Liu, Xiaozhai Yu, Fengrong Ou","doi":"10.1007/s00432-024-05866-4","DOIUrl":"10.1007/s00432-024-05866-4","url":null,"abstract":"<p><strong>Purpose: </strong>Colorectal cancer (CRC) is a common malignancy that affects adults worldwide, causing a high disease burden. Few studies have examined the relationship between body composition (BC) measures and the prevalence of CRC. Our purpose was to investigate the relationship between pertinent BC indicators and CRC.</p><p><strong>Methods: </strong>Bioelectrical impedance analysis, laboratory test results, face-to-face questionnaire investigation, and nutritional risk assessment (Nutritional Risk Screening 2002 and Patient-Generated Subjective Global Assessment) were used in this case-control study. Bioelectrical impedance analysis in the case group was performed prior to antitumor therapy/surgery.</p><p><strong>Results: </strong>From June 2018 to January 2019, a total of 303 cases and 286 controls were included. The results showed that low body fat percentage (BFP) and high visceral adiposity index (VAI) groups had a higher risk of developing CRC in comparison to the normal BFP and normal VAI groups. The risk of CRC decreased with the increase of BFP. The group with a normal BC had a lower risk of developing CRC compared to those with a greater VAI and a lower BFP, as indicated by the results of the pairwise and total combinations of VAI, fat-free mass index (FFMI), and BFP. Additionally, FFMI and VAI had positive correlations with prealbumin, serum albumin, and nutritional risk scores.</p><p><strong>Conclusion: </strong>Low BFP and high VAI are associated with higher CRC risk. FFMI and VAI are positively correlated with prealbumin, serum albumin, and nutritional risk scores in CRC patients.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11233301/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141563446","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-09DOI: 10.1007/s00432-024-05875-3
Ziwei Liang, Yang Ge, Jianjian Li, Yunting Bai, Zeru Xiao, Rui Yan, Guangyu An, Donglei Zhang
Background: Cholangiocarcinoma (CCA), characterized by high heterogeneity and extreme malignancy, has a poor prognosis. Doublecortin-like kinase 1 (DCLK1) promotes a variety of malignant cancers in their progression. Targeting DCLK1 or its associated regulatory pathways can prevent the generation and deterioration of several malignancies. However, the role of DCLK1 in CCA progression and its molecular mechanisms remain unknown. Therefore, we aimed to investigate whether and how DCLK1 contributes to CCA progression.
Methods: The expression of DCLK1 in CCA patients was detected using Immunohistochemistry (IHC). We established DCLK1 knockout and DCLK1 overexpression cell lines for Colony Formation Assay and Transwell experiments to explore the tumor-promoting role of DCLK1. RT-PCR, Western blot and multiple fluorescent staining were used to assess the association between DCLK1 and epithelial-mesenchymal transition (EMT) markers. RNA sequencing and bioinformatics analysis were performed to identify the underlying mechanisms by which DCLK1 regulates CCA progression and the EMT program.
Results: DCLK1 was overexpressed in CCA tissues and was associated with poor prognosis. DCLK1 overexpression facilitated CCA cell invasion, migration, and proliferation, whereas DCLK1 knockdown reversed the malignant tendencies of CCA cells, which had been confirmed both in vivo and in vitro. Furthermore, we demonstrated that DCLK1 was substantially linked to the advancement of the EMT program, which included the overexpression of mesenchymal markers and the downregulation of epithelial markers. For the underlying mechanism, we proposed that the PI3K/AKT/mTOR pathway is the key process for the role of DCLK1 in tumor progression and the occurrence of the EMT program. When administered with LY294002, an inhibitor of the PI3K/AKT/mTOR pathway, the tumor's ability to proliferate, migrate, and invade was greatly suppressed, and the EMT process was generally reversed.
Conclusions: DCLK1 facilitates the malignant biological behavior of CCA cells through the PI3K/AKT/mTOR pathway. In individuals with cholangiocarcinoma who express DCLK1 at high levels, inhibitors of the PI3K/AKT/mTOR signaling pathway may be an effective therapeutic approach.
{"title":"Targeting the PI3K/AKT/mTOR pathway offer a promising therapeutic strategy for cholangiocarcinoma patients with high doublecortin-like kinase 1 expression.","authors":"Ziwei Liang, Yang Ge, Jianjian Li, Yunting Bai, Zeru Xiao, Rui Yan, Guangyu An, Donglei Zhang","doi":"10.1007/s00432-024-05875-3","DOIUrl":"10.1007/s00432-024-05875-3","url":null,"abstract":"<p><strong>Background: </strong>Cholangiocarcinoma (CCA), characterized by high heterogeneity and extreme malignancy, has a poor prognosis. Doublecortin-like kinase 1 (DCLK1) promotes a variety of malignant cancers in their progression. Targeting DCLK1 or its associated regulatory pathways can prevent the generation and deterioration of several malignancies. However, the role of DCLK1 in CCA progression and its molecular mechanisms remain unknown. Therefore, we aimed to investigate whether and how DCLK1 contributes to CCA progression.</p><p><strong>Methods: </strong>The expression of DCLK1 in CCA patients was detected using Immunohistochemistry (IHC). We established DCLK1 knockout and DCLK1 overexpression cell lines for Colony Formation Assay and Transwell experiments to explore the tumor-promoting role of DCLK1. RT-PCR, Western blot and multiple fluorescent staining were used to assess the association between DCLK1 and epithelial-mesenchymal transition (EMT) markers. RNA sequencing and bioinformatics analysis were performed to identify the underlying mechanisms by which DCLK1 regulates CCA progression and the EMT program.</p><p><strong>Results: </strong>DCLK1 was overexpressed in CCA tissues and was associated with poor prognosis. DCLK1 overexpression facilitated CCA cell invasion, migration, and proliferation, whereas DCLK1 knockdown reversed the malignant tendencies of CCA cells, which had been confirmed both in vivo and in vitro. Furthermore, we demonstrated that DCLK1 was substantially linked to the advancement of the EMT program, which included the overexpression of mesenchymal markers and the downregulation of epithelial markers. For the underlying mechanism, we proposed that the PI3K/AKT/mTOR pathway is the key process for the role of DCLK1 in tumor progression and the occurrence of the EMT program. When administered with LY294002, an inhibitor of the PI3K/AKT/mTOR pathway, the tumor's ability to proliferate, migrate, and invade was greatly suppressed, and the EMT process was generally reversed.</p><p><strong>Conclusions: </strong>DCLK1 facilitates the malignant biological behavior of CCA cells through the PI3K/AKT/mTOR pathway. In individuals with cholangiocarcinoma who express DCLK1 at high levels, inhibitors of the PI3K/AKT/mTOR signaling pathway may be an effective therapeutic approach.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11233391/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141558833","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Purpose: Radical resection of retroperitoneal liposarcoma (RLPS) may necessitate vascular resection and reconstruction. The study was conducted to assess surgical outcomes of surgery for RLPS with major vascular involvement.
Methods: Patients with RLPS who underwent surgical resection at the Sarcoma Center of Peking University Cancer Hospital between April 2011 and December 2022 were identified from a prospectively maintained database. Patients were classified into two groups: vascular resection and non-vascular resection groups. A propensity score matching analysis was performed to eliminate baseline differences between the groups. Surgical details and postoperative outcomes were analyzed. Furthermore, prognostic factors for local recurrence-free survival (LRFS) and overall survival (OS) were assessed.
Results: Overall, 199 patients were identified and the median follow-up period was 48 (interquartile range [IQR] 45-69) months. Vascular resection was performed in 42 (21%) patients, 25 of whom had vascular infiltration. A total of 39 patients had vascular replacement and 3 patients underwent partial resection (side-wall resection). Vascular resection was burdened by higher rates of major morbidity (38% vs. 14%, p < 0.001) and 30-day mortality (7.1% vs. 1.3%, p = 0.005). After propensity-matched analysis, patients who underwent vascular resection had 5-year LRFS and OS rates comparable to those without vascular involvement. Major vascular resection was not an independent risk factor for LRFS or OS.
Conclusions: Although accompanied by increased risks of major morbidity and mortality, the major vascular resection enabled radical resection in patients with advanced RLPS, affording comparable 5-year LRFS and OS rates compared to those who did not.
{"title":"Surgical outcomes of major vascular resection for retroperitoneal liposarcoma from a high‑volume sarcoma center: a propensity score matching analysis.","authors":"Guoqiang Xue, Xiaopeng Wang, Bonan Liu, Chengpeng Li, Ang Lv, Xiuyun Tian, Jianhui Wu, Hui Qiu, Chunyi Hao","doi":"10.1007/s00432-024-05871-7","DOIUrl":"10.1007/s00432-024-05871-7","url":null,"abstract":"<p><strong>Purpose: </strong>Radical resection of retroperitoneal liposarcoma (RLPS) may necessitate vascular resection and reconstruction. The study was conducted to assess surgical outcomes of surgery for RLPS with major vascular involvement.</p><p><strong>Methods: </strong>Patients with RLPS who underwent surgical resection at the Sarcoma Center of Peking University Cancer Hospital between April 2011 and December 2022 were identified from a prospectively maintained database. Patients were classified into two groups: vascular resection and non-vascular resection groups. A propensity score matching analysis was performed to eliminate baseline differences between the groups. Surgical details and postoperative outcomes were analyzed. Furthermore, prognostic factors for local recurrence-free survival (LRFS) and overall survival (OS) were assessed.</p><p><strong>Results: </strong>Overall, 199 patients were identified and the median follow-up period was 48 (interquartile range [IQR] 45-69) months. Vascular resection was performed in 42 (21%) patients, 25 of whom had vascular infiltration. A total of 39 patients had vascular replacement and 3 patients underwent partial resection (side-wall resection). Vascular resection was burdened by higher rates of major morbidity (38% vs. 14%, p < 0.001) and 30-day mortality (7.1% vs. 1.3%, p = 0.005). After propensity-matched analysis, patients who underwent vascular resection had 5-year LRFS and OS rates comparable to those without vascular involvement. Major vascular resection was not an independent risk factor for LRFS or OS.</p><p><strong>Conclusions: </strong>Although accompanied by increased risks of major morbidity and mortality, the major vascular resection enabled radical resection in patients with advanced RLPS, affording comparable 5-year LRFS and OS rates compared to those who did not.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11233382/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141563448","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Purpose: To develop a deep learning (DL) model for differentiating between benign and malignant ovarian tumors of Ovarian-Adnexal Reporting and Data System Ultrasound (O-RADS US) Category 4 lesions, and validate its diagnostic performance.
Methods: A retrospective analysis of 1619 US images obtained from three centers from December 2014 to March 2023. DeepLabV3 and YOLOv8 were jointly used to segment, classify, and detect ovarian tumors. Precision and recall and area under the receiver operating characteristic curve (AUC) were employed to assess the model performance.
Results: A total of 519 patients (including 269 benign and 250 malignant masses) were enrolled in the study. The number of women included in the training, validation, and test cohorts was 426, 46, and 47, respectively. The detection models exhibited an average precision of 98.68% (95% CI: 0.95-0.99) for benign masses and 96.23% (95% CI: 0.92-0.98) for malignant masses. Moreover, in the training set, the AUC was 0.96 (95% CI: 0.94-0.97), whereas in the validation set, the AUC was 0.93(95% CI: 0.89-0.94) and 0.95 (95% CI: 0.91-0.96) in the test set. The sensitivity, specificity, accuracy, positive predictive value, and negative predictive values for the training set were 0.943,0.957,0.951,0.966, and 0.936, respectively, whereas those for the validation set were 0.905,0.935, 0.935,0.919, and 0.931, respectively. In addition, the sensitivity, specificity, accuracy, positive predictive value, and negative predictive value for the test set were 0.925, 0.955, 0.941, 0.956, and 0.927, respectively.
Conclusion: The constructed DL model exhibited high diagnostic performance in distinguishing benign and malignant ovarian tumors in O-RADS US category 4 lesions.
{"title":"Developing a deep learning model for predicting ovarian cancer in Ovarian-Adnexal Reporting and Data System Ultrasound (O-RADS US) Category 4 lesions: A multicenter study.","authors":"Wenting Xie, Wenjie Lin, Ping Li, Hongwei Lai, Zhilan Wang, Peizhong Liu, Yijun Huang, Yao Liu, Lina Tang, Guorong Lyu","doi":"10.1007/s00432-024-05872-6","DOIUrl":"10.1007/s00432-024-05872-6","url":null,"abstract":"<p><strong>Purpose: </strong>To develop a deep learning (DL) model for differentiating between benign and malignant ovarian tumors of Ovarian-Adnexal Reporting and Data System Ultrasound (O-RADS US) Category 4 lesions, and validate its diagnostic performance.</p><p><strong>Methods: </strong>A retrospective analysis of 1619 US images obtained from three centers from December 2014 to March 2023. DeepLabV3 and YOLOv8 were jointly used to segment, classify, and detect ovarian tumors. Precision and recall and area under the receiver operating characteristic curve (AUC) were employed to assess the model performance.</p><p><strong>Results: </strong>A total of 519 patients (including 269 benign and 250 malignant masses) were enrolled in the study. The number of women included in the training, validation, and test cohorts was 426, 46, and 47, respectively. The detection models exhibited an average precision of 98.68% (95% CI: 0.95-0.99) for benign masses and 96.23% (95% CI: 0.92-0.98) for malignant masses. Moreover, in the training set, the AUC was 0.96 (95% CI: 0.94-0.97), whereas in the validation set, the AUC was 0.93(95% CI: 0.89-0.94) and 0.95 (95% CI: 0.91-0.96) in the test set. The sensitivity, specificity, accuracy, positive predictive value, and negative predictive values for the training set were 0.943,0.957,0.951,0.966, and 0.936, respectively, whereas those for the validation set were 0.905,0.935, 0.935,0.919, and 0.931, respectively. In addition, the sensitivity, specificity, accuracy, positive predictive value, and negative predictive value for the test set were 0.925, 0.955, 0.941, 0.956, and 0.927, respectively.</p><p><strong>Conclusion: </strong>The constructed DL model exhibited high diagnostic performance in distinguishing benign and malignant ovarian tumors in O-RADS US category 4 lesions.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11233367/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141563447","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-08DOI: 10.1007/s00432-024-05850-y
Dag Rune Stormoen, Kristoffer Staal Rohrberg, Kent William Mouw, Katrine Ørum, Zoltan Szallasi, Maria Rossing, Frederik Otzen Bagger, Helle Pappot
Introduction: Urothelial tract cancer (UTC) ranks as the tenth most prevalent cancer and holds the seventh position in terms of mortality worldwide. Despite its prevalence and mortality ranking, there are still gaps in the knowledge of the mutational landscape in patients with advanced disease who have limited therapeutic options after multiple lines of prior treatment. This study compares the genomic and transcriptomic landscape, and targeted treatment options between metastatic UTC (mUTC) patients treated with multiple lines of therapy compared to newly diagnosed, untreated Muscle Invasive Bladder Cancer (MIBC).
Methods: We compared genomic and clinical data from two cohorts: mUTC patients who received multiple lines of therapy and were referred to the Copenhagen Prospective Personalized Oncology (CoPPO) project at Rigshospitalet, University of Copenhagen. Data for MIBC UTC patients were acquired from the Cancer Genome Atlas Bladder Cancer (TCGA BLCA) cohort. Biopsies in CoPPO were performed at the time of enrollment. 523 highly important cancer-related genes (TrueSight Oncology-500 targeted sequencing panel) were used from both cohorts for comparative analysis. Analyses included RNA count data to compare predicted molecular subtypes in each cohort separately.
Results: Patients from the CoPPO cohort had a lower median age at first-line treatment than the TCGA BLCA cohort, with no significant gender disparity. The predominant histology was urothelial cell carcinoma in both cohorts. Genomic analysis revealed no significant difference between the top mutated genes in the two cohorts, specifically looking into DNA damage repair genes. Molecular subtyping indicated a higher frequency of neuroendocrine differentiation in the CoPPO cohort. 13% of patients in the CoPPO cohort received targeted therapy based on genomic findings, and 16% received non-targeted treatment, totaling 29% receiving CoPPO treatment (9 patients). The remaining 71% received best supportive care. Kaplan-Meier analysis showed a non-significant survival benefit for the intervention group in the CoPPO cohort.
Conclusion: When focusing on 523 highly relevant cancer genes, the mutational profile of mUTC patients who have undergone numerous treatment lines resembles that of newly diagnosed MIBC. These alterations can be targeted, indicating the potential advantage of early genomic testing for personalized treatment within clinical trials.
{"title":"Similar genetic profile in early and late stage urothelial tract cancer.","authors":"Dag Rune Stormoen, Kristoffer Staal Rohrberg, Kent William Mouw, Katrine Ørum, Zoltan Szallasi, Maria Rossing, Frederik Otzen Bagger, Helle Pappot","doi":"10.1007/s00432-024-05850-y","DOIUrl":"10.1007/s00432-024-05850-y","url":null,"abstract":"<p><strong>Introduction: </strong>Urothelial tract cancer (UTC) ranks as the tenth most prevalent cancer and holds the seventh position in terms of mortality worldwide. Despite its prevalence and mortality ranking, there are still gaps in the knowledge of the mutational landscape in patients with advanced disease who have limited therapeutic options after multiple lines of prior treatment. This study compares the genomic and transcriptomic landscape, and targeted treatment options between metastatic UTC (mUTC) patients treated with multiple lines of therapy compared to newly diagnosed, untreated Muscle Invasive Bladder Cancer (MIBC).</p><p><strong>Methods: </strong>We compared genomic and clinical data from two cohorts: mUTC patients who received multiple lines of therapy and were referred to the Copenhagen Prospective Personalized Oncology (CoPPO) project at Rigshospitalet, University of Copenhagen. Data for MIBC UTC patients were acquired from the Cancer Genome Atlas Bladder Cancer (TCGA BLCA) cohort. Biopsies in CoPPO were performed at the time of enrollment. 523 highly important cancer-related genes (TrueSight Oncology-500 targeted sequencing panel) were used from both cohorts for comparative analysis. Analyses included RNA count data to compare predicted molecular subtypes in each cohort separately.</p><p><strong>Results: </strong>Patients from the CoPPO cohort had a lower median age at first-line treatment than the TCGA BLCA cohort, with no significant gender disparity. The predominant histology was urothelial cell carcinoma in both cohorts. Genomic analysis revealed no significant difference between the top mutated genes in the two cohorts, specifically looking into DNA damage repair genes. Molecular subtyping indicated a higher frequency of neuroendocrine differentiation in the CoPPO cohort. 13% of patients in the CoPPO cohort received targeted therapy based on genomic findings, and 16% received non-targeted treatment, totaling 29% receiving CoPPO treatment (9 patients). The remaining 71% received best supportive care. Kaplan-Meier analysis showed a non-significant survival benefit for the intervention group in the CoPPO cohort.</p><p><strong>Conclusion: </strong>When focusing on 523 highly relevant cancer genes, the mutational profile of mUTC patients who have undergone numerous treatment lines resembles that of newly diagnosed MIBC. These alterations can be targeted, indicating the potential advantage of early genomic testing for personalized treatment within clinical trials.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11230994/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141554876","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Objective: Glioma is a leading cause of mortality worldwide, its recurrence poses a major challenge in achieving effective treatment outcomes. Cancer stem cells (CSCs) have emerged as key contributors to tumor relapse and chemotherapy resistance, making them attractive targets for glioma cancer therapy. This study investigated the potential of FERMT1 as a prognostic biomarker and its role in regulating stemness through cell cycle in glioma.
Methods: Using data from TCGA-GBM, GSE4290, GSE50161 and GSE147352 for analysis of FERMT1 expression in glioma tissues. Then, the effects of FERMT1 knockdown on cell cycle, proliferation, sphere formation ability, invasion and migration were investigated. The influences of FERMT1 on expression of glycolysis-related proteins and levels of ATP, glucose, lactate and G6PDH were also explored. Furthermore, the effects of FERMT1 knockdown on cellular metabolism were evidenced.
Results: Significant upregulation of FERMT1 in glioma tissues was observed. Silencing FERMT1 not only affected the cell cycle but also led to a notable reduction in proliferation, invasion and migration. The expression of glycolysis-associated proteins including GLUT1, GLUT3, GLUT4, and SCO2 were reduced by FERMT1 knockdown, resulted in increased ATP and glucose as well as decreased lactic acid and G6PDH levels. FERMT1 knockdown also inhibited cellular metabolism. Moreover, FERMT1 knockdown significantly reduced sphere diameter, along with inhibiting the expression of transcription factors associated with stemness in glioma cells.
Conclusion: These findings demonstrated that FERMT1 could be an ideal target for the advancement of innovative strategies against glioma treatment via modulating cellular process involved in stemness regulation and metabolism.
{"title":"FERMT1 suppression induces anti-tumor effects and reduces stemness in glioma cancer cells.","authors":"Zhigang Pan, Chuhan Ke, Hanlin Zheng, Xiumei Guo, Wen Gao, Xinyue Huang, Chunhui Chen, Yu Xiong, Shuni Zheng, Feng Zheng, Weipeng Hu","doi":"10.1007/s00432-024-05859-3","DOIUrl":"10.1007/s00432-024-05859-3","url":null,"abstract":"<p><strong>Objective: </strong>Glioma is a leading cause of mortality worldwide, its recurrence poses a major challenge in achieving effective treatment outcomes. Cancer stem cells (CSCs) have emerged as key contributors to tumor relapse and chemotherapy resistance, making them attractive targets for glioma cancer therapy. This study investigated the potential of FERMT1 as a prognostic biomarker and its role in regulating stemness through cell cycle in glioma.</p><p><strong>Methods: </strong>Using data from TCGA-GBM, GSE4290, GSE50161 and GSE147352 for analysis of FERMT1 expression in glioma tissues. Then, the effects of FERMT1 knockdown on cell cycle, proliferation, sphere formation ability, invasion and migration were investigated. The influences of FERMT1 on expression of glycolysis-related proteins and levels of ATP, glucose, lactate and G6PDH were also explored. Furthermore, the effects of FERMT1 knockdown on cellular metabolism were evidenced.</p><p><strong>Results: </strong>Significant upregulation of FERMT1 in glioma tissues was observed. Silencing FERMT1 not only affected the cell cycle but also led to a notable reduction in proliferation, invasion and migration. The expression of glycolysis-associated proteins including GLUT1, GLUT3, GLUT4, and SCO2 were reduced by FERMT1 knockdown, resulted in increased ATP and glucose as well as decreased lactic acid and G6PDH levels. FERMT1 knockdown also inhibited cellular metabolism. Moreover, FERMT1 knockdown significantly reduced sphere diameter, along with inhibiting the expression of transcription factors associated with stemness in glioma cells.</p><p><strong>Conclusion: </strong>These findings demonstrated that FERMT1 could be an ideal target for the advancement of innovative strategies against glioma treatment via modulating cellular process involved in stemness regulation and metabolism.</p>","PeriodicalId":15118,"journal":{"name":"Journal of Cancer Research and Clinical Oncology","volume":null,"pages":null},"PeriodicalIF":2.7,"publicationDate":"2024-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11231014/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141554875","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}