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Volume 28,Issue 11,2021 Table of Contents
2021, 28(11):1053-1060.
Abstract:
Single-cell sequencing is a new technology for comprehensive analysis of genetic information at single cell level through high[1]throughput sequencing technology. Single-cell sequencing has become a powerful tool for in-depth research in modern biology. Single-cell sequencing technologies have enabled the research in biomedical field and clinical translation to achieve a high level of accuracy and sensitivity that traditional sequencing technology couldn’t achieve. At present, single-cell transcriptome sequencing (often referred to as “single-cell RNA sequencing”, scRNA-seq) has become the most widely used single-cell sequencing technology. In this paper, the development history, technical principles and methods of scRNA-seq are briefly introduced. In addition, the research achievements of scRNA-seq in the basic research fields of tumor heterogeneity, tumor metastasis, tumor immunity and tumor drug resistance, as well as its research progress on translational medicine and application in tumor precision medicine are systematically described, in hope to promote tumor precision medicine to a higher level through the development and application of single-cell sequencing technology.
Single-cell sequencing is a new technology for comprehensive analysis of genetic information at single cell level through high[1]throughput sequencing technology. Single-cell sequencing has become a powerful tool for in-depth research in modern biology. Single-cell sequencing technologies have enabled the research in biomedical field and clinical translation to achieve a high level of accuracy and sensitivity that traditional sequencing technology couldn’t achieve. At present, single-cell transcriptome sequencing (often referred to as “single-cell RNA sequencing”, scRNA-seq) has become the most widely used single-cell sequencing technology. In this paper, the development history, technical principles and methods of scRNA-seq are briefly introduced. In addition, the research achievements of scRNA-seq in the basic research fields of tumor heterogeneity, tumor metastasis, tumor immunity and tumor drug resistance, as well as its research progress on translational medicine and application in tumor precision medicine are systematically described, in hope to promote tumor precision medicine to a higher level through the development and application of single-cell sequencing technology.
2021, 28(11):1061-1067.
Abstract:
Objective: To investigate the expression and clinical significance of transducer of ERBB2.1 antisense RNA1 (TOB1-AS1)in epithelial ovarian cancer (EOC) tissues, and to preliminarily explore its effect on the proliferation, migration, and invasion of EOC cells in vitro. Methods: TOB1-AS1 expression in EOC tissues was analyzed using the TCGA database. The tumor tissues from 67 patients who underwent tumor resection and were pathologically confirmed as EOC in the Department of Gynecology, the Fourth Hospital of Hebei Medical University from July 2017 to January 2018 were collected for this study. In addition, 30 non-tumor ovarian tissues of patients with other gynecological diseases were collected as the control in the same period. qPCR was used to detect the expression of TOB1-AS1 in EOC tissues and non-tumor ovarian tissues. χ2 test was used to analyze the correlation between the expression of TOB1-AS1 and different clinicopathological factors of EOC patients. Kaplan-Meier method and Cox proportional hazard regression model were used to analyze the survival and potential influencing factors for prognosis in EOC patients. The effects of TOB1-AS1 knockdown on the proliferation, migration, and invasion of EOC SKOV3 and A2780 cells were detected by CCK-8 test,Wound-healing assay, and Transwell test, respectively. Results: TCGA database analysis and qPCR results showed the expression level of TOB1-AS1 in EOC tissues was significantly higher than that in non-tumor ovarian tissues (all P<0.01). The high expression of TOB1-AS1 was conspicuously correlated with advanced FIGO stage, poor tissue grade, lymph node metastasis, and peritoneal metastasis in patients with EOC (all P<0.05). Kaplan-Meier survival analysis showed that post-operative disease-free survival (DFS)and overall survival (OS) in the patients with high TOB1-AS1 expression were shorter than those with low TOB1-AS1 expression (all P<0.05). Cox proportional hazard regression model analysis showed that FIGO stage, lymph node metastasis, peritoneal metastasis, and TOB1-AS1 expression were independent prognostic factors in EOC patients (all P<0.05). Similarly, the expression level of TOB1-AS1 in SKOV3 and A2780 cells were significantly higher than that in normal epithelial ovarian IOSE80 cells (all P<0.01). Cell function experiments showed that TOB1-AS1 knockdown inhibited the proliferation, migration, and invasion of SKOV3 and A2780 cells (all P<0.05). Conclusion: TOB1-AS1 is highly expressed in EOC tissues, and it is significantly related to the poor prognosis of EOC patients.TOB1-AS1 may affect the malignant progression of EOC by promoting the cell proliferation, migration, and invasion of SKOV3 and A2780 cells.
Objective: To investigate the expression and clinical significance of transducer of ERBB2.1 antisense RNA1 (TOB1-AS1)in epithelial ovarian cancer (EOC) tissues, and to preliminarily explore its effect on the proliferation, migration, and invasion of EOC cells in vitro. Methods: TOB1-AS1 expression in EOC tissues was analyzed using the TCGA database. The tumor tissues from 67 patients who underwent tumor resection and were pathologically confirmed as EOC in the Department of Gynecology, the Fourth Hospital of Hebei Medical University from July 2017 to January 2018 were collected for this study. In addition, 30 non-tumor ovarian tissues of patients with other gynecological diseases were collected as the control in the same period. qPCR was used to detect the expression of TOB1-AS1 in EOC tissues and non-tumor ovarian tissues. χ2 test was used to analyze the correlation between the expression of TOB1-AS1 and different clinicopathological factors of EOC patients. Kaplan-Meier method and Cox proportional hazard regression model were used to analyze the survival and potential influencing factors for prognosis in EOC patients. The effects of TOB1-AS1 knockdown on the proliferation, migration, and invasion of EOC SKOV3 and A2780 cells were detected by CCK-8 test,Wound-healing assay, and Transwell test, respectively. Results: TCGA database analysis and qPCR results showed the expression level of TOB1-AS1 in EOC tissues was significantly higher than that in non-tumor ovarian tissues (all P<0.01). The high expression of TOB1-AS1 was conspicuously correlated with advanced FIGO stage, poor tissue grade, lymph node metastasis, and peritoneal metastasis in patients with EOC (all P<0.05). Kaplan-Meier survival analysis showed that post-operative disease-free survival (DFS)and overall survival (OS) in the patients with high TOB1-AS1 expression were shorter than those with low TOB1-AS1 expression (all P<0.05). Cox proportional hazard regression model analysis showed that FIGO stage, lymph node metastasis, peritoneal metastasis, and TOB1-AS1 expression were independent prognostic factors in EOC patients (all P<0.05). Similarly, the expression level of TOB1-AS1 in SKOV3 and A2780 cells were significantly higher than that in normal epithelial ovarian IOSE80 cells (all P<0.01). Cell function experiments showed that TOB1-AS1 knockdown inhibited the proliferation, migration, and invasion of SKOV3 and A2780 cells (all P<0.05). Conclusion: TOB1-AS1 is highly expressed in EOC tissues, and it is significantly related to the poor prognosis of EOC patients.TOB1-AS1 may affect the malignant progression of EOC by promoting the cell proliferation, migration, and invasion of SKOV3 and A2780 cells.
2021, 28(11):1068-1074.
Abstract:
Objective: To explore whether miR-496 affects the cycle, proliferation, migration, and invasion of cervical cancer HeLa cells and the growth of transplanted tumors in nude mice by targeting mTOR. Methods: The tumor specimens and para-cancerous tissue specimens of 74 patients with cervical cancer who underwent surgery at the Fourth Hospital of Hebei Medical University from December 2020 to December 2021 were selected. qPCR, WB, and immunofluorescence methods were used to detect the expression of miR-496 and mTOR at mRNA and protein levels in cervical cancer tissues. The TargetScan database was used to predict the target genes of miR-496, and the Dual-luciferase reporter gene experiment was further adopted to verify the targeting relationship. According to different transfectants, HeLa cells were divided into the control group, miR-496 mimic group, and miR-496 mimic+pMIR-mTOR group. CCK-8 assay, Flow cytometry, and Transwell experiment were used to detect the effects of miR-496 and mTOR on the proliferation, cell cycle, migration, and invasion of HeLa cells. The HeLa cells of the control group and miR-496 mimic group were inoculated subcutaneously into BALB/c nude mice to construct a transplanted tumor model of cervical cancer. After 21 d, the mice were sacrificed, and the tumor tissues of the mice were stripped and the tumor mass was weighed. Immunofluorescence and WB methods were used to detect the effect of miR-496 overexpression on the expression of mTOR and Ki67 in transplanted tumor tissues. Results: In cervical cancer tissues, miR-496 was lowly expressed, while mTOR was highly expressed at both mRNA and protein levels. miR- 496 could bind to the 3'-UTR sequence of mTOR mRNA. Compared with the control group and the miR-496 mimic+pMIR-mTOR group, the miR-496 level and S-phase cell ratio of HeLa cells in the miR-496 mimic group were significantly increased, while the proliferation level, the numbers of migrated cells and invaded cells were significantly reduced (all P<0.01). Nude mice transplanted tumor model of cervical cancer was successfully constructed. After 21 d of inoculation, the mass of transplanted tumors and the expression of mTOR and Ki67 in the transplanted tumor tissues in the miR-496 mimic group were significantly lower than those in the control group (all P<0.01). Conclusion: miR-496 is lowly expressed in cervical cancer tissues. Overexpression of miR-496 can inhibit the malignant biological behaviors of HeLa cells and the growth of transplanted tumors in nude mice through targeted regulation of mTOR.
Objective: To explore whether miR-496 affects the cycle, proliferation, migration, and invasion of cervical cancer HeLa cells and the growth of transplanted tumors in nude mice by targeting mTOR. Methods: The tumor specimens and para-cancerous tissue specimens of 74 patients with cervical cancer who underwent surgery at the Fourth Hospital of Hebei Medical University from December 2020 to December 2021 were selected. qPCR, WB, and immunofluorescence methods were used to detect the expression of miR-496 and mTOR at mRNA and protein levels in cervical cancer tissues. The TargetScan database was used to predict the target genes of miR-496, and the Dual-luciferase reporter gene experiment was further adopted to verify the targeting relationship. According to different transfectants, HeLa cells were divided into the control group, miR-496 mimic group, and miR-496 mimic+pMIR-mTOR group. CCK-8 assay, Flow cytometry, and Transwell experiment were used to detect the effects of miR-496 and mTOR on the proliferation, cell cycle, migration, and invasion of HeLa cells. The HeLa cells of the control group and miR-496 mimic group were inoculated subcutaneously into BALB/c nude mice to construct a transplanted tumor model of cervical cancer. After 21 d, the mice were sacrificed, and the tumor tissues of the mice were stripped and the tumor mass was weighed. Immunofluorescence and WB methods were used to detect the effect of miR-496 overexpression on the expression of mTOR and Ki67 in transplanted tumor tissues. Results: In cervical cancer tissues, miR-496 was lowly expressed, while mTOR was highly expressed at both mRNA and protein levels. miR- 496 could bind to the 3'-UTR sequence of mTOR mRNA. Compared with the control group and the miR-496 mimic+pMIR-mTOR group, the miR-496 level and S-phase cell ratio of HeLa cells in the miR-496 mimic group were significantly increased, while the proliferation level, the numbers of migrated cells and invaded cells were significantly reduced (all P<0.01). Nude mice transplanted tumor model of cervical cancer was successfully constructed. After 21 d of inoculation, the mass of transplanted tumors and the expression of mTOR and Ki67 in the transplanted tumor tissues in the miR-496 mimic group were significantly lower than those in the control group (all P<0.01). Conclusion: miR-496 is lowly expressed in cervical cancer tissues. Overexpression of miR-496 can inhibit the malignant biological behaviors of HeLa cells and the growth of transplanted tumors in nude mice through targeted regulation of mTOR.
2021, 28(11):1075-1080.
Abstract:
Objective: To investigate the expression level of miRNA let-7i-5p in renal cell carcinoma (RCC) tissues and to explore its effect on the proliferation, migration, invasion, and hyaluronan-binding protein 4 (HABP4) expression in human RCC 769-P cells.Methods: A meta-analysis of let-7i-5p expression in RCC tissues was performed using the TCGA RCC database and GEO database.The human RCC 769-P cells were routinely cultured for transfection in vitro, and were divided into three groups according to different transfection materials: overexpression group (transfected with let-7i-5p mimics), inhibition group (transfected with let-7i-5p inhibitors),and control group (transfected with NC sequence). CCK-8, cell scratch test, Transwell assay, and WB method were used to detect the effects of let-7i-5p on the proliferation, scratch healing rate, invaded cell numbers and HABP4 expression in 769-P cells. Dual[1]luciferase reporter gene assay was performed to demonstrate the targeting relationship between let-7i-5p and HABP4. Results: Data analysis of TCGA RCC database and 5 GEO data sets (GSE23085, GSE47582, GSE95385, GSE16441, and GSE71302) showed that the expression level of let-7i-5p was significantly higher in RCC tissues than in normal kidney tissues (all P <0.05). As shown by in vitro experiments, compared with the control group, the cell proliferation activity of the overexpression group was significantly increased at 24, 48, and 72 h, while that of the inhibition group was decreased (all P<0.01). The scratch healing rate [(37.276±2.058)% vs (15.663±2.949)%, P<0.01] and the invaded cell numbers [(377.000±34.044) vs (255.667±25.34), P<0.05] were significantly increased in the overexpression group, while the scratch healing rate [(8.791±2.568) % vs (15.663±2.949) %, P<0.05] and the invaded cell numbers [(170.333±14.978) vs (255.667±25.384), P<0.01] were significantly reduced in the inhibition group. The luciferase activity was significantly inhibited by the overexpression of let-7i-5p in the wild-type HABP4-3'URT plasmid group (P<0.01), while no statistical difference was observed in the mutant HABP4-3'URT plasmid group (P>0.05). WB results showed that compared with the control group, the expression of HABP4 and E-cadherin in the overexpression group was decreased (all P<0.01), and the expression of CDK2 was increased (P<0.01), while the inhibition group showed the opposite trend (all P<0.01). Conclusion: Let-7i-5p is highly expressed in RCC tissues and promotes the proliferation, migration, and invasion of 769-P cells possibly by targeting HABP4.
Objective: To investigate the expression level of miRNA let-7i-5p in renal cell carcinoma (RCC) tissues and to explore its effect on the proliferation, migration, invasion, and hyaluronan-binding protein 4 (HABP4) expression in human RCC 769-P cells.Methods: A meta-analysis of let-7i-5p expression in RCC tissues was performed using the TCGA RCC database and GEO database.The human RCC 769-P cells were routinely cultured for transfection in vitro, and were divided into three groups according to different transfection materials: overexpression group (transfected with let-7i-5p mimics), inhibition group (transfected with let-7i-5p inhibitors),and control group (transfected with NC sequence). CCK-8, cell scratch test, Transwell assay, and WB method were used to detect the effects of let-7i-5p on the proliferation, scratch healing rate, invaded cell numbers and HABP4 expression in 769-P cells. Dual[1]luciferase reporter gene assay was performed to demonstrate the targeting relationship between let-7i-5p and HABP4. Results: Data analysis of TCGA RCC database and 5 GEO data sets (GSE23085, GSE47582, GSE95385, GSE16441, and GSE71302) showed that the expression level of let-7i-5p was significantly higher in RCC tissues than in normal kidney tissues (all P <0.05). As shown by in vitro experiments, compared with the control group, the cell proliferation activity of the overexpression group was significantly increased at 24, 48, and 72 h, while that of the inhibition group was decreased (all P<0.01). The scratch healing rate [(37.276±2.058)% vs (15.663±2.949)%, P<0.01] and the invaded cell numbers [(377.000±34.044) vs (255.667±25.34), P<0.05] were significantly increased in the overexpression group, while the scratch healing rate [(8.791±2.568) % vs (15.663±2.949) %, P<0.05] and the invaded cell numbers [(170.333±14.978) vs (255.667±25.384), P<0.01] were significantly reduced in the inhibition group. The luciferase activity was significantly inhibited by the overexpression of let-7i-5p in the wild-type HABP4-3'URT plasmid group (P<0.01), while no statistical difference was observed in the mutant HABP4-3'URT plasmid group (P>0.05). WB results showed that compared with the control group, the expression of HABP4 and E-cadherin in the overexpression group was decreased (all P<0.01), and the expression of CDK2 was increased (P<0.01), while the inhibition group showed the opposite trend (all P<0.01). Conclusion: Let-7i-5p is highly expressed in RCC tissues and promotes the proliferation, migration, and invasion of 769-P cells possibly by targeting HABP4.
SUN Hui
,
HUA Weiwei
,
CHEN Xiwen
,
LI Yajuan
,
QIN Wei
,
YIN Zixin
,
ZHAO Ya
,
LIU Yanqing
,
QIAN Yayun
2021, 28(11):1081-1086.
Abstract:
Objective: To explore the synergistic effect of 28-hydroxy-3-oxoolean-12-en-2-oic acid and miR-451 on the proliferation and migration of human gastric cancer AGS cells and its possible molecular mechanism. Methods: AGS cells were infected with miR-451 overexpression lentivirus and induced with DOX (10 or 100 ng/ml) for 24 h to construct AGS/miR-451+ cells overexpressing miR-451. AGS/miR-451+ cells were treated with 28-hydroxy-3-oxoolean-12-en-2-oic acid of 10, 20, 40, 80, 160 μmol/L. The changes in proliferation and migration ability of cells were detected by the MTT method and the scratch test, respectively; WB method was used to detect the changes in the expression levels of mTOR signaling pathway-related and apoptosis-related proteins in cells. Results:AGS/miR-451+ cells overexpressing miR-451 were successfully constructed. Compared with the untreated control group, the proliferation inhibition rate of AGS/miR-451+ cells in the 28-hydroxy-3-oxoolean-12-en-2-oic acid treatment group increased in a time[1]and concentration-dependent manner (P<0.05 or P<0.01), while the cell migration rate was significantly reduced (P<0.05 or P<0.01). In the cells treated with 28-hydroxy-3-oxoolean-12-en-2-oic acid, the expression of mTOR signaling pathway-related proteins was reduced (P<0.05 or P<0.01); among the apoptosis pathway-related proteins, the expression of Bcl2 was decreased, but the expression of BAX,caspase-3, caspase-1, and cytochrome c was increased (P<0.05 or 0.01). Conclusion: The 28-hydroxy-3-oxoolean-12-en-2-oic and miR-451 can synergistically inhibit the proliferation and migration of human gastric cancer AGS cells. The mechanism may be related to their regulation of apoptosis signaling pathway- and mTOR signaling pathway-related proteins.
Objective: To explore the synergistic effect of 28-hydroxy-3-oxoolean-12-en-2-oic acid and miR-451 on the proliferation and migration of human gastric cancer AGS cells and its possible molecular mechanism. Methods: AGS cells were infected with miR-451 overexpression lentivirus and induced with DOX (10 or 100 ng/ml) for 24 h to construct AGS/miR-451+ cells overexpressing miR-451. AGS/miR-451+ cells were treated with 28-hydroxy-3-oxoolean-12-en-2-oic acid of 10, 20, 40, 80, 160 μmol/L. The changes in proliferation and migration ability of cells were detected by the MTT method and the scratch test, respectively; WB method was used to detect the changes in the expression levels of mTOR signaling pathway-related and apoptosis-related proteins in cells. Results:AGS/miR-451+ cells overexpressing miR-451 were successfully constructed. Compared with the untreated control group, the proliferation inhibition rate of AGS/miR-451+ cells in the 28-hydroxy-3-oxoolean-12-en-2-oic acid treatment group increased in a time[1]and concentration-dependent manner (P<0.05 or P<0.01), while the cell migration rate was significantly reduced (P<0.05 or P<0.01). In the cells treated with 28-hydroxy-3-oxoolean-12-en-2-oic acid, the expression of mTOR signaling pathway-related proteins was reduced (P<0.05 or P<0.01); among the apoptosis pathway-related proteins, the expression of Bcl2 was decreased, but the expression of BAX,caspase-3, caspase-1, and cytochrome c was increased (P<0.05 or 0.01). Conclusion: The 28-hydroxy-3-oxoolean-12-en-2-oic and miR-451 can synergistically inhibit the proliferation and migration of human gastric cancer AGS cells. The mechanism may be related to their regulation of apoptosis signaling pathway- and mTOR signaling pathway-related proteins.
2021, 28(11):1087-1097.
Abstract:
Objective: To investigate the effect of miR-133a-5p regulating serum exosomes derived fibronectin1 (FN1) on the proliferation, adhesion, and M1 macrophage polarization of gastric cardia adenocarcinoma (GCA) cells. Methods: The differentially expressed genes in GCA were predicted using the GEO database, and their functional enrichment analysis was performed. qPCR was used to detect the expression of FN1 in GCA tissue, serum and serum-derived exosomes of GCA patients, and GCA cells. The FN1 overexpression vector and its control plasmid were respectively transfected into the serum-derived exosomes of GCA patients. The miR-133a-5p mimics and its control mimics were transfected into HGC-27 cells. The transfected HGC-27 cells were co-cultured with transfected exosomes, and then the cells were co-cultured with THP-1 cells. CCK-8 and cell adhesion experiments were used to detect the proliferation and adhesion of HGC-27 cells in each group. WB method and ELISA were used to detect the levels of CD86 and iNOS in cells and the effects of transfection on the secretion of IL-6 and IL-1β from macrophages. Dual-luciferase reporter experiment was adopted to verify the interaction between FN1 mRNA and miR-133a-5p. Results: Compared with healthy controls, the expression levels of FN1 in GCA tissues, serum and serum-derived exosomes from GCA patients, and GCA cells were significantly up-regulated (all P<0.05). High expression of FN1 in serum exosomes was associated with the TNM stage (P=0.032 9) and lymph node metastasis (P=0.012 7) in GCA patients. FN1-riched exosomes could be internalized by GCA cells, and co-culture with FN1-riched exosomes could improve the proliferation and adhesion ability of GCA cells and inhibit the polarization of M1-type macrophages by blocking the expression of IL-6, IL-1β, CD86, and iNOS (P<0.05 or P<0.01). miR-133a-3p was lowly expressed in GCA tissues and cells and could negatively regulate the expression of FN1. Overexpression of miR-133a-5p could promote the expression of IL-6, IL-1β,CD86, and iNOS in M1 macrophages by reducing the proliferation and adhesion of GCA cells, and partially reversing the promotion effect of FN1 on the malignant behaviors of GCA cells (P<0.05 or P<0.01). Conclusion: miR-133a-5p inhibits the proliferation and adhesion of GCA cells via suppressing the secretion of FN1 by serum exosomes and promotes the polarization of M1 macrophages.
Objective: To investigate the effect of miR-133a-5p regulating serum exosomes derived fibronectin1 (FN1) on the proliferation, adhesion, and M1 macrophage polarization of gastric cardia adenocarcinoma (GCA) cells. Methods: The differentially expressed genes in GCA were predicted using the GEO database, and their functional enrichment analysis was performed. qPCR was used to detect the expression of FN1 in GCA tissue, serum and serum-derived exosomes of GCA patients, and GCA cells. The FN1 overexpression vector and its control plasmid were respectively transfected into the serum-derived exosomes of GCA patients. The miR-133a-5p mimics and its control mimics were transfected into HGC-27 cells. The transfected HGC-27 cells were co-cultured with transfected exosomes, and then the cells were co-cultured with THP-1 cells. CCK-8 and cell adhesion experiments were used to detect the proliferation and adhesion of HGC-27 cells in each group. WB method and ELISA were used to detect the levels of CD86 and iNOS in cells and the effects of transfection on the secretion of IL-6 and IL-1β from macrophages. Dual-luciferase reporter experiment was adopted to verify the interaction between FN1 mRNA and miR-133a-5p. Results: Compared with healthy controls, the expression levels of FN1 in GCA tissues, serum and serum-derived exosomes from GCA patients, and GCA cells were significantly up-regulated (all P<0.05). High expression of FN1 in serum exosomes was associated with the TNM stage (P=0.032 9) and lymph node metastasis (P=0.012 7) in GCA patients. FN1-riched exosomes could be internalized by GCA cells, and co-culture with FN1-riched exosomes could improve the proliferation and adhesion ability of GCA cells and inhibit the polarization of M1-type macrophages by blocking the expression of IL-6, IL-1β, CD86, and iNOS (P<0.05 or P<0.01). miR-133a-3p was lowly expressed in GCA tissues and cells and could negatively regulate the expression of FN1. Overexpression of miR-133a-5p could promote the expression of IL-6, IL-1β,CD86, and iNOS in M1 macrophages by reducing the proliferation and adhesion of GCA cells, and partially reversing the promotion effect of FN1 on the malignant behaviors of GCA cells (P<0.05 or P<0.01). Conclusion: miR-133a-5p inhibits the proliferation and adhesion of GCA cells via suppressing the secretion of FN1 by serum exosomes and promotes the polarization of M1 macrophages.
YANG Yanli
,
LIAO Li
,
ZHANG Jing
,
SUN Peng
,
TAKASHIMA Kenichi
,
ZHANG Fengchun
,
YANG Yuhui
,
ZHANG Bin
,
HU Liangding
2021, 28(11):1098-1106.
Abstract:
Objective: To evaluate whether adoptive immunotherapy (AIT) with activated autologous lymphocytes helps to improve the clinical efficacy of primary hepatocellular carcinoma. Methods: Sixty-four patients with primary hepatocellular carcinoma diagnosed at the Fifth Medical Center of Chinese PLA General Hospital from August 2016 to December 2018 were enrolled and divided into immunotherapy group (n=29) and control group (n=35) by the stratified randomized sampling method. 60 ml of peripheral blood was drawn from each patient in the immunotherapy group to prepare mononuclear cells, which were then activated and cultured in a medium containing OKT-3 and IL-2. A quality control test was done before blood re-transfusion. In the immunotherapy group,patients in stage Ⅰ-Ⅲ (n=14) received autologous lymphocyte infusion (6 infusions within 3 months) after first-line treatment, while patients in stage Ⅳ (n=15) only received autologous lymphocyte infusion therapy. Patients in the control group received other hepatocellular carcinoma-related treatments. The primary endpoint of the efficacy evaluation was the 2-year relapse-free survival (RFS), and the secondary endpoints were progression-free survival (PFS) and overall survival (OS). Results: The median follow-up time of the enrolled patients was 2.8 years (0.2-4.2 years). 29 patients in the immunotherapy group received a total of 167 scheduled lymphocyte infusions (with an average of 9.30×109 cells per patient, of which CD3+HLA-DR cells accounted for about 63%; 174 infusions were planned, with a completion rate of 96%). During the treatment period, no grade 3-4 adverse reactions were observed.Compared with the control group, patients in the immunotherapy group had a significantly increased 2-year RFS rate (62.1% vs 22.9%,OR=0.181, 95%CI: 0.06-0.54, P=0.002), and the median PFS (28 vs 8 months, P=0.004) and median OS (38 vs 34 months, P=0.915) were significantly prolonged. Among the patients at stage Ⅰ-Ⅲ, the PFS rate in the immunotherapy group was significantly higher than that in the control group (92.9% vs 33.3%, OR=0.38, 95%CI: 0.004-0.368, P=0.005), and the median PFS was significantly prolonged (38 vs 14.5 months, P=0.005), but there was no significant difference in OS between the two groups. In stage Ⅳ patients, there was no statistical difference in PFS (P=0.077) and OS (P=0.994) between the two groups. Conclusion: AIT with activated autologous lymphocytes is a safe and feasible adjuvant therapy for hepatocellular carcinoma, which can improve RFS rate after first-line treatment of stage Ⅰ-Ⅲ hepatocellular carcinoma and prolong the recurrence-free survival time of patients. It has no significant effect on PFS and OS in patients with advanced hepatocellular carcinoma.
Objective: To evaluate whether adoptive immunotherapy (AIT) with activated autologous lymphocytes helps to improve the clinical efficacy of primary hepatocellular carcinoma. Methods: Sixty-four patients with primary hepatocellular carcinoma diagnosed at the Fifth Medical Center of Chinese PLA General Hospital from August 2016 to December 2018 were enrolled and divided into immunotherapy group (n=29) and control group (n=35) by the stratified randomized sampling method. 60 ml of peripheral blood was drawn from each patient in the immunotherapy group to prepare mononuclear cells, which were then activated and cultured in a medium containing OKT-3 and IL-2. A quality control test was done before blood re-transfusion. In the immunotherapy group,patients in stage Ⅰ-Ⅲ (n=14) received autologous lymphocyte infusion (6 infusions within 3 months) after first-line treatment, while patients in stage Ⅳ (n=15) only received autologous lymphocyte infusion therapy. Patients in the control group received other hepatocellular carcinoma-related treatments. The primary endpoint of the efficacy evaluation was the 2-year relapse-free survival (RFS), and the secondary endpoints were progression-free survival (PFS) and overall survival (OS). Results: The median follow-up time of the enrolled patients was 2.8 years (0.2-4.2 years). 29 patients in the immunotherapy group received a total of 167 scheduled lymphocyte infusions (with an average of 9.30×109 cells per patient, of which CD3+HLA-DR cells accounted for about 63%; 174 infusions were planned, with a completion rate of 96%). During the treatment period, no grade 3-4 adverse reactions were observed.Compared with the control group, patients in the immunotherapy group had a significantly increased 2-year RFS rate (62.1% vs 22.9%,OR=0.181, 95%CI: 0.06-0.54, P=0.002), and the median PFS (28 vs 8 months, P=0.004) and median OS (38 vs 34 months, P=0.915) were significantly prolonged. Among the patients at stage Ⅰ-Ⅲ, the PFS rate in the immunotherapy group was significantly higher than that in the control group (92.9% vs 33.3%, OR=0.38, 95%CI: 0.004-0.368, P=0.005), and the median PFS was significantly prolonged (38 vs 14.5 months, P=0.005), but there was no significant difference in OS between the two groups. In stage Ⅳ patients, there was no statistical difference in PFS (P=0.077) and OS (P=0.994) between the two groups. Conclusion: AIT with activated autologous lymphocytes is a safe and feasible adjuvant therapy for hepatocellular carcinoma, which can improve RFS rate after first-line treatment of stage Ⅰ-Ⅲ hepatocellular carcinoma and prolong the recurrence-free survival time of patients. It has no significant effect on PFS and OS in patients with advanced hepatocellular carcinoma.
2021, 28(11):1107-1112.
Abstract:
Objective: To analyze the regulatory effects of miR-19 on PTEN and PI3K/Akt signaling pathway, and to reveal the effects of miR-19 and PTEN on invasion and migration of endometrial cancer KLE cells as well as its working mechanism. Methods: Tumor tissues and para-carcerous tissues of 74 patients with endometrial cancer were collected from the First Hospital of Hebei Medical University from May 2017 to August 2020. All patients did not receive radiotherapy or chemotherapy before surgery and were pathologically diagnosed as endometrial cancer. qPCR and WB were used to detect the effects of miR-19 mimics or inhibitors on PTEN expression in KLE cells. TargetScan and Dual-luciferase reporter gene assay were adopted to predict and verify whether PTEN was the target gene of miR-19. KLE cells were randomized into negative control (NC) group (transfected with miR-NC), miR-19 mimic group (transfected with miR-19 mimics) and miR-19+PTEN group (transfected with miR-19 mimics+PTEN overexpression vector).Transwell assay and Scratch assay were used to detect the effects of miR-19 and PTEN on KLE cell migration and invasion. WB was adopted to detect the expression of PI3K/Akt pathway-related proteins.Results: mRNA and protein expression of PTEN was obviously lower in endometrial cancer tissues compared with para-carcerous tissues (P<0.01). miR-19 could specifically bind to 3'-UTR of PTEN.Up-regulation of miR-19 could inhibit mRNA and protein expression of PTEN, and down-regulation of miR-19 showed an opposite result (P<0.01). Compared with miR-NC group, migratory cell count, invasive cell count, and scratch healing rate were significantly increased in miR-19 mimic group (all P<0.01). Compared with miR-19 mimic group, migratory cell count, invasive cell count and scratch healing rate were significantly decreased in miR-19+PTEN group (all P<0.01). WB analysis showed that, comparing with miR[1]NC group, PTEN protein was obviously decreased, while p-Akt 308 and p-Akt 473 protein was obviously elevated in miR-19 mimic group (P<0.01); however, Akt protein maintained unchanged. Additionally, the levels of p-Akt 308 and p-Akt 473 protein were obviously lower in miR-19+PTEN group comparing with miR-19 mimic group (P<0.01). Conclusion: miR-19 may activate PI3K/Akt signal pathway by inhibiting PTEN expression, thereby promoting migration and invasion of endometrial cancer KLE cells.
Objective: To analyze the regulatory effects of miR-19 on PTEN and PI3K/Akt signaling pathway, and to reveal the effects of miR-19 and PTEN on invasion and migration of endometrial cancer KLE cells as well as its working mechanism. Methods: Tumor tissues and para-carcerous tissues of 74 patients with endometrial cancer were collected from the First Hospital of Hebei Medical University from May 2017 to August 2020. All patients did not receive radiotherapy or chemotherapy before surgery and were pathologically diagnosed as endometrial cancer. qPCR and WB were used to detect the effects of miR-19 mimics or inhibitors on PTEN expression in KLE cells. TargetScan and Dual-luciferase reporter gene assay were adopted to predict and verify whether PTEN was the target gene of miR-19. KLE cells were randomized into negative control (NC) group (transfected with miR-NC), miR-19 mimic group (transfected with miR-19 mimics) and miR-19+PTEN group (transfected with miR-19 mimics+PTEN overexpression vector).Transwell assay and Scratch assay were used to detect the effects of miR-19 and PTEN on KLE cell migration and invasion. WB was adopted to detect the expression of PI3K/Akt pathway-related proteins.Results: mRNA and protein expression of PTEN was obviously lower in endometrial cancer tissues compared with para-carcerous tissues (P<0.01). miR-19 could specifically bind to 3'-UTR of PTEN.Up-regulation of miR-19 could inhibit mRNA and protein expression of PTEN, and down-regulation of miR-19 showed an opposite result (P<0.01). Compared with miR-NC group, migratory cell count, invasive cell count, and scratch healing rate were significantly increased in miR-19 mimic group (all P<0.01). Compared with miR-19 mimic group, migratory cell count, invasive cell count and scratch healing rate were significantly decreased in miR-19+PTEN group (all P<0.01). WB analysis showed that, comparing with miR[1]NC group, PTEN protein was obviously decreased, while p-Akt 308 and p-Akt 473 protein was obviously elevated in miR-19 mimic group (P<0.01); however, Akt protein maintained unchanged. Additionally, the levels of p-Akt 308 and p-Akt 473 protein were obviously lower in miR-19+PTEN group comparing with miR-19 mimic group (P<0.01). Conclusion: miR-19 may activate PI3K/Akt signal pathway by inhibiting PTEN expression, thereby promoting migration and invasion of endometrial cancer KLE cells.
2021, 28(11):1113-1118.
Abstract:
Objective: To explore the efficacy of PD-1 inhibitors in the treatment of patients with advanced lung cancer and its influence on the T lymphocyte subsets and cytokine levels in the peripheral blood of patients. Methods: A total of 50 patients with lung cancer admitted to Haikou Hospital from August 2018 to December 2020 were selected, and 50 healthy subjects were selected as control. The expression of PD-1 expression in lung cancer tissues was detected by immunohistochemistry. Lung cancer patients were treated with nivolumab or pembrolizumab, and venous blood was collected at one day before treatment, the end of treatment cycle 1,and the end of treatment cycle 4. After 4 cycles of treatment, CT or MRI was performed to evaluate the tumor size. Patients evaluated with complete response (CR), partial response (PR), and stable disease (SD) were classified as an immune responsive group, and patients evaluated with progressive disease (PD) were classified as an immune non-responsive group. The effect of PD-1 inhibitor treatment on T lymphocyte subsets (CD3+T, CD4+T, CD8+T, CD4+/CD8+T, Treg, and Th1/Th2 cells), NK cells, cytokines (IFN-γ, IL-2,IL-4, and IL-5) in peripheral blood of patients were evaluated. Results: Compared with healthy controls, the levels of CD3+T, CD4+T,CD4+/CD8+T, Th1/Th2 cells, IFN-γ, and IL-2 in peripheral blood of lung cancer patients were significantly decreased, while the levels of CD8+ T cells, Treg cells, NK cells, IL-4, and IL-5 were significantly increased (all P<0.05). Compared with pre-treatment, the levels of CD3+T, CD4+T, CD4+/CD8+T cells were significantly increased, while the levels of CD8+T cells, Treg cells, and NK cells were significantly decreased after 1 and 4 cycles of treatment (all P<0.05). After 4 cycles of treatment, there were 40 cases in the immune responsive group and 10 cases in the non-immune responsive group, with an effective rate of 80%. Compared with the immune non[1]responsive group, the levels of CD3+T, CD4+T, CD4+/CD8+T, and Th1/Th2 cells were significantly increased in the immune response group, while the levels of CD8+T cells, Treg cells, and NK cells were significantly decreased (all P<0.05); In the immune responsive group, there were 32 patients with high PD-L1 expression (≥50%) and 8 patients with low PD-L1 expression (<50%). Compared with patients with low PD-L1 expression, the levels of CD3+T, CD4+T, CD4+/CD8+T cells were significantly increased in patients with high PD-L1 expression, while the levels of CD8+T cells, Treg cells, and NK cells were significantly decreased (all P<0.05). Conclusion: Nivolumab or pembrolizumab treatment can affect the distribution of immune cells such as T lymphocyte subsets in patients with advanced lung cancer and improve the immune status of patients.
Objective: To explore the efficacy of PD-1 inhibitors in the treatment of patients with advanced lung cancer and its influence on the T lymphocyte subsets and cytokine levels in the peripheral blood of patients. Methods: A total of 50 patients with lung cancer admitted to Haikou Hospital from August 2018 to December 2020 were selected, and 50 healthy subjects were selected as control. The expression of PD-1 expression in lung cancer tissues was detected by immunohistochemistry. Lung cancer patients were treated with nivolumab or pembrolizumab, and venous blood was collected at one day before treatment, the end of treatment cycle 1,and the end of treatment cycle 4. After 4 cycles of treatment, CT or MRI was performed to evaluate the tumor size. Patients evaluated with complete response (CR), partial response (PR), and stable disease (SD) were classified as an immune responsive group, and patients evaluated with progressive disease (PD) were classified as an immune non-responsive group. The effect of PD-1 inhibitor treatment on T lymphocyte subsets (CD3+T, CD4+T, CD8+T, CD4+/CD8+T, Treg, and Th1/Th2 cells), NK cells, cytokines (IFN-γ, IL-2,IL-4, and IL-5) in peripheral blood of patients were evaluated. Results: Compared with healthy controls, the levels of CD3+T, CD4+T,CD4+/CD8+T, Th1/Th2 cells, IFN-γ, and IL-2 in peripheral blood of lung cancer patients were significantly decreased, while the levels of CD8+ T cells, Treg cells, NK cells, IL-4, and IL-5 were significantly increased (all P<0.05). Compared with pre-treatment, the levels of CD3+T, CD4+T, CD4+/CD8+T cells were significantly increased, while the levels of CD8+T cells, Treg cells, and NK cells were significantly decreased after 1 and 4 cycles of treatment (all P<0.05). After 4 cycles of treatment, there were 40 cases in the immune responsive group and 10 cases in the non-immune responsive group, with an effective rate of 80%. Compared with the immune non[1]responsive group, the levels of CD3+T, CD4+T, CD4+/CD8+T, and Th1/Th2 cells were significantly increased in the immune response group, while the levels of CD8+T cells, Treg cells, and NK cells were significantly decreased (all P<0.05); In the immune responsive group, there were 32 patients with high PD-L1 expression (≥50%) and 8 patients with low PD-L1 expression (<50%). Compared with patients with low PD-L1 expression, the levels of CD3+T, CD4+T, CD4+/CD8+T cells were significantly increased in patients with high PD-L1 expression, while the levels of CD8+T cells, Treg cells, and NK cells were significantly decreased (all P<0.05). Conclusion: Nivolumab or pembrolizumab treatment can affect the distribution of immune cells such as T lymphocyte subsets in patients with advanced lung cancer and improve the immune status of patients.
2021, 28(11):1119-1128.
Abstract:
RNA在多种生理和病理过程中起着至关重要的作用。在过去的几十年中，研究人员已经设计出多种类型的RNA及 其衍生体，并应用于不同的生物学领域。研究证实，RNA分子在肿瘤的生长、转移、代谢和免疫逃逸等方面发挥着重要作用。基 于肿瘤相关RNA的研究和相关技术的应用，RNA已经被设计应用于肿瘤免疫治疗之中。此外，作为肿瘤免疫治疗的工具，RNA 可以与其他类型的免疫疗法结合使用，例如免疫检查点抑制剂、树突状细胞回输治疗以及嵌合抗原受体T细胞免疫疗法等。在 综述中，论述了各种RNA工具（包括RNA适配体、mRNA疫苗和RNA溶瘤病毒）在肿瘤免疫治疗中应用的研究进展，总结了这些 RNA工具治疗肿瘤的优点和缺点，并展望了RNA及其衍生体未来在肿瘤免疫治疗中的潜在临床应用。
RNA在多种生理和病理过程中起着至关重要的作用。在过去的几十年中，研究人员已经设计出多种类型的RNA及 其衍生体，并应用于不同的生物学领域。研究证实，RNA分子在肿瘤的生长、转移、代谢和免疫逃逸等方面发挥着重要作用。基 于肿瘤相关RNA的研究和相关技术的应用，RNA已经被设计应用于肿瘤免疫治疗之中。此外，作为肿瘤免疫治疗的工具，RNA 可以与其他类型的免疫疗法结合使用，例如免疫检查点抑制剂、树突状细胞回输治疗以及嵌合抗原受体T细胞免疫疗法等。在 综述中，论述了各种RNA工具（包括RNA适配体、mRNA疫苗和RNA溶瘤病毒）在肿瘤免疫治疗中应用的研究进展，总结了这些 RNA工具治疗肿瘤的优点和缺点，并展望了RNA及其衍生体未来在肿瘤免疫治疗中的潜在临床应用。
2021, 28(11):1129-1134.
Abstract:
低氧微环境是大多数恶性肿瘤的重要特征。在低氧环境下，低氧诱导因子（hypoxia-inducible factors，HIF）高度表达， 通过诱导VEGF表达促进恶性肿瘤血管生成并增加血管通透性、通过调控免疫细胞作用促进免疫抑制和肿瘤细胞的免疫逃逸， 进而促进肿瘤的恶性进展。目前，已研究多种HIF的靶向抑制剂，其作用机制包括降低HIF转录活性、抑制HIF表达和诱导HIF 降解等，相关药物多处于临床前或临床研究阶段，走向临床应用的过程中仍有很多问题亟待解决，如HIF抑制剂单药治疗或与放 疗、化疗联合应用，特别是不同HIF抑制剂与免疫联合的疗效及安全性的探索等。
低氧微环境是大多数恶性肿瘤的重要特征。在低氧环境下，低氧诱导因子（hypoxia-inducible factors，HIF）高度表达， 通过诱导VEGF表达促进恶性肿瘤血管生成并增加血管通透性、通过调控免疫细胞作用促进免疫抑制和肿瘤细胞的免疫逃逸， 进而促进肿瘤的恶性进展。目前，已研究多种HIF的靶向抑制剂，其作用机制包括降低HIF转录活性、抑制HIF表达和诱导HIF 降解等，相关药物多处于临床前或临床研究阶段，走向临床应用的过程中仍有很多问题亟待解决，如HIF抑制剂单药治疗或与放 疗、化疗联合应用，特别是不同HIF抑制剂与免疫联合的疗效及安全性的探索等。
2021, 28(11):1135-1140.
Abstract:
在基础研究方面，业已证实lncRNA主要以癌基因或抑癌基因两种形式参与白血病细胞的增殖、分化或凋亡过程，是 白血病发生和发展的重要因素。临床研究表明，lncRNA与白血病的诊断、分型、风险分级和预后判断有关，具有成为临床病情监 测的生物标志物潜力。以lncRNA siRNA为代表的白血病治疗性分子产品也表现出了良好的应用前景。但是，lncRNA在AML 领域的研究依然面临着许多问题和挑战，需要进一步筛选特异性较高的关键标志性和功能性lncRNA，通过大型队列非干预性临 床试验验证其作为生物标志物或治疗靶点的可行性。
在基础研究方面，业已证实lncRNA主要以癌基因或抑癌基因两种形式参与白血病细胞的增殖、分化或凋亡过程，是 白血病发生和发展的重要因素。临床研究表明，lncRNA与白血病的诊断、分型、风险分级和预后判断有关，具有成为临床病情监 测的生物标志物潜力。以lncRNA siRNA为代表的白血病治疗性分子产品也表现出了良好的应用前景。但是，lncRNA在AML 领域的研究依然面临着许多问题和挑战，需要进一步筛选特异性较高的关键标志性和功能性lncRNA，通过大型队列非干预性临 床试验验证其作为生物标志物或治疗靶点的可行性。
2021, 28(11):1141-1147.
Abstract:
神经母细胞瘤是儿童最常见的实性颅外恶性肿瘤，常常发生转移性扩散，晚期预后差。靶向免疫治疗（如靶向GD2和 B7-H3）在神经母细胞瘤治疗中的效果已在临床对照试验中得到证实，美国FDA已批准靶向GD2的dinutuximab和naxitamab用 于治疗儿童神经母细胞瘤。最近一些研究表明，P物质（SP）通过神经激肽1受体（NK-1R）激活一系列信号通路，介导多种肿瘤的 发生发展。SP通过NK-1R在神经母细胞瘤细胞中发挥以下作用：促进细胞有丝分裂、促进细胞迁移（侵袭和转移）、抗凋亡和增 加糖酵解效率（肿瘤细胞因此增加代谢），而NK-1R拮抗剂会诱导肿瘤细胞的凋亡。因此，SP/NK-1R有成为神经母细胞瘤新的治 疗靶点的潜力。
神经母细胞瘤是儿童最常见的实性颅外恶性肿瘤，常常发生转移性扩散，晚期预后差。靶向免疫治疗（如靶向GD2和 B7-H3）在神经母细胞瘤治疗中的效果已在临床对照试验中得到证实，美国FDA已批准靶向GD2的dinutuximab和naxitamab用 于治疗儿童神经母细胞瘤。最近一些研究表明，P物质（SP）通过神经激肽1受体（NK-1R）激活一系列信号通路，介导多种肿瘤的 发生发展。SP通过NK-1R在神经母细胞瘤细胞中发挥以下作用：促进细胞有丝分裂、促进细胞迁移（侵袭和转移）、抗凋亡和增 加糖酵解效率（肿瘤细胞因此增加代谢），而NK-1R拮抗剂会诱导肿瘤细胞的凋亡。因此，SP/NK-1R有成为神经母细胞瘤新的治 疗靶点的潜力。
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Chinese Journal Of Cancer Biotheray ® 2025 All Rights Reserved
Supported by:Beijing E-Tiller Technology Development Co., Ltd.