Background:Drug resistance is the main factor contributing to cancer recurrence and poor prognosis.Exploration of drug resistance-related mechanisms and effective therapeutic targets are the aim of molecular targeted ...Background:Drug resistance is the main factor contributing to cancer recurrence and poor prognosis.Exploration of drug resistance-related mechanisms and effective therapeutic targets are the aim of molecular targeted therapy.In our study,the role of long non-coding RNA(lncRNA)AFAP1-AS1 in gemcitabine resistance and related mechanisms were explored in cervical cancer cells.Methods:Gemcitabine-resistant cervical cancer cell lines HT-3-Gem and SW756-Gem were constructed using the gemcitabine concentration gradient method.The overall survival rates and recurrence-free survival rates were evaluated by Kaplan-Meier analysis.The interaction was verified through a Dual-luciferase reporter gene assay and a Biotinylated RNA pull-down assay.Cell proliferation ability was assessed through methyl-thiazolyl-tetrazolium(MTT),soft agar,and colony formation experiments.Cell cycle and apoptosis were detected byflow cytometry.Results:Up-regulation of AFAP1-AS1 in cervical cancer predicted a poor prognosis.Besides,patients in the gemcitabine-resistance group had higher levels of AFAP1-AS1 than the gemcitabine-sensitive group.AFAP1-AS1 promoted tumor growth and induced gemcitabine tolerance of cervical cancer cells.In addition,AFAP1-AS1 mediated epidermal growth factor receptor(EGFR)expression by serving as a molecular sponge for microRNA-7a-5p(miR-7-5p).This present study also proved that the knockdown of EGFR or overexpression of miR-7a-5p abolished the accelerative role of AFAP1-AS1 overexpression in cancer progression and gemcitabine tolerance.Conclusions:In general,the AFAP1-AS1/miR-7-5p/EGFR axis was tightly related to the progression and gemcitabine tolerance of cervical cancer,providing potential targets for the management of cervical cancer.展开更多
目的:探讨血浆长链非编码RNA(long non-coding RNA,lncRNA)肌动蛋白纤维相关蛋白1-反义RNA1(actin filament-associated protein 1-antisense RNA1,AFAP1-AS1)和Y染色体性别决定基因簇2重叠转录本(sex-determining region of Y chromoso...目的:探讨血浆长链非编码RNA(long non-coding RNA,lncRNA)肌动蛋白纤维相关蛋白1-反义RNA1(actin filament-associated protein 1-antisense RNA1,AFAP1-AS1)和Y染色体性别决定基因簇2重叠转录本(sex-determining region of Y chromosome-box2 overlapping transcript,SOX2OT)表达水平对于非小细胞肺癌(non-small cell lung cancer,NSCLC)的临床诊断价值。方法:留取48例NSCLC患者(NSCLC组)和48例良性肺病患者(对照组)血液标本,采用实时定量PCR检测lncRNA AFAP1-AS1和lncRNA SOX2OT相对表达水平,评价二者诊断NSCLC的可行性及效能,同时检测血清癌胚抗原水平。结果:NSCLC患者血浆lncRNA AFAP1-AS1和lncRNA SOX2OT表达水平显著高于对照组(t=6.236,5.680,P均<0.01),lncRNA AFAP1-AS1和lncRNA SOX2OT鉴别诊断NSCLC与良性肺病的受试者工作特征曲线下面积(AUC)分别为0.875(95%CI:0.804~0.947,P<0.05)和0.787(95%CI:0.691~0.883,P<0.05)。二者联合血清癌胚抗原可提高诊断的AUC。结论:NSCLC患者血浆中lncRNA AFAP1-AS1和lncRNA SOX2OT表达明显上调,二者联合血清癌胚抗原对NSCLC的诊断效能优于单独应用。展开更多
目的:探讨沉默GCB弥漫型大B细胞淋巴瘤(diffuse large B cell lymphoma,DLBCL)中AFAP1-AS1的表达对细胞增殖和凋亡的影响。方法:培养GCB-DLBCL细胞至对数生长期后转染OCI-Ly1细胞系,建立的GCB-DLBCL细胞系对AFAP1-AS1表达进行沉默;实验...目的:探讨沉默GCB弥漫型大B细胞淋巴瘤(diffuse large B cell lymphoma,DLBCL)中AFAP1-AS1的表达对细胞增殖和凋亡的影响。方法:培养GCB-DLBCL细胞至对数生长期后转染OCI-Ly1细胞系,建立的GCB-DLBCL细胞系对AFAP1-AS1表达进行沉默;实验设立3组,实验组为腺病毒感染细胞,sh-NC无关序列腺病毒感染细胞组为无关序列对照组,未感染腺病毒细胞组为空白组,应用PCR法检测AFAP1-AS1表达水平、CCK-8法测定细胞增殖情况、流式细胞术检测细胞凋亡情况,对比检测结果。结果:AFAP1-AS1表达水平检测结果显示:三种shRNA序列干扰效率均较无关序列对照组(sh-NC)强,差异有统计学意义(P<0.05);采用CCK-8法检测各组细胞凋亡情况,结果显示:经腺病毒sh3-AFAP1-AS1感染后,OCI-Ly1细胞系中实验组细胞吸光度较无关序列对照组和空白组显著降低,下调AFAP1-AS1可抑制GCB-DLBCL细胞的增殖(P<0.05);采用流式细胞仪检测各组细胞凋亡情况,结果显示:经sh3-AFAP1-AS1和sh-NC转染后,OCI-Ly1细胞实验组凋亡率明显高于无关序列对照组和空白组,下调AFAP1-AS1可诱导GCB-DLBCL细胞凋亡(P<0.05)。结论:沉默GCB-DLBCL细胞中的AFAP1-AS1表达能有效抑制细胞增殖,诱导细胞凋亡,或可作为GCB-DLBCL治疗的靶目标。展开更多
Triple-negative breast cancer(TNBC)is characterized by fast growth,high metastasis,high invasion,and a lack of therapeutic targets.Mitosis and metastasis of TNBC cells are two important biological behaviors in TNBC ma...Triple-negative breast cancer(TNBC)is characterized by fast growth,high metastasis,high invasion,and a lack of therapeutic targets.Mitosis and metastasis of TNBC cells are two important biological behaviors in TNBC malignant progression.It is well known that the long noncoding RNA AFAP1-AS1 plays a crucial role in various tumors,but whether AFAP1-AS1 is involved in the mitosis of TNBC cells remains unknown.In this study,we investigated the functional mechanism of AFAP1-AS1 in targeting Polo-like Kinase 1(PLK1)activation and participating in mitosis of TNBC cells.We detected the expression of AFAP1-AS1 in the TNBC patient cohort and primary cells by in situ hybridization(ISH),northern blot,fluorescent in situ hybridization(FISH)and cell nucleus/cytoplasm RNA fraction isolation.High AFAP1-AS1 expression was negatively correlated with overall survival(OS),disease-free survival(DFS),metastasis-free survival(MFS)and recurrence-free survival(RFS)in TNBC patients.We explored the function of AFAP1-AS1 by transwell,apoptosis,immunofluorescence(IF)and patient-derived xenograft(PDX)models in vitro and in vivo.We found that AFAP1-AS1 promoted TNBC primary cell survival by inhibiting mitotic catastrophe and increased TNBC primary cell growth,migration and invasion.Mechanistically,AFAP1-AS1 activated phosphorylation of the mitosis-associated kinase PLK1 protein.Elevated levels of AFAP1-AS1 in TNBC primary cells increased PLK1 pathway downstream gene expression,such as CDC25C,CDK1,BUB1 and TTK.More importantly,AFAP1-AS1 increased lung metastases in a mouse metastasis model.Taken together,AFAP1-AS1 functions as an oncogene that activates the PLK1 signaling pathway.AFAP1-AS1 could be used as a potential prognostic marker and therapeutic target for TNBC.展开更多
文摘Background:Drug resistance is the main factor contributing to cancer recurrence and poor prognosis.Exploration of drug resistance-related mechanisms and effective therapeutic targets are the aim of molecular targeted therapy.In our study,the role of long non-coding RNA(lncRNA)AFAP1-AS1 in gemcitabine resistance and related mechanisms were explored in cervical cancer cells.Methods:Gemcitabine-resistant cervical cancer cell lines HT-3-Gem and SW756-Gem were constructed using the gemcitabine concentration gradient method.The overall survival rates and recurrence-free survival rates were evaluated by Kaplan-Meier analysis.The interaction was verified through a Dual-luciferase reporter gene assay and a Biotinylated RNA pull-down assay.Cell proliferation ability was assessed through methyl-thiazolyl-tetrazolium(MTT),soft agar,and colony formation experiments.Cell cycle and apoptosis were detected byflow cytometry.Results:Up-regulation of AFAP1-AS1 in cervical cancer predicted a poor prognosis.Besides,patients in the gemcitabine-resistance group had higher levels of AFAP1-AS1 than the gemcitabine-sensitive group.AFAP1-AS1 promoted tumor growth and induced gemcitabine tolerance of cervical cancer cells.In addition,AFAP1-AS1 mediated epidermal growth factor receptor(EGFR)expression by serving as a molecular sponge for microRNA-7a-5p(miR-7-5p).This present study also proved that the knockdown of EGFR or overexpression of miR-7a-5p abolished the accelerative role of AFAP1-AS1 overexpression in cancer progression and gemcitabine tolerance.Conclusions:In general,the AFAP1-AS1/miR-7-5p/EGFR axis was tightly related to the progression and gemcitabine tolerance of cervical cancer,providing potential targets for the management of cervical cancer.
文摘目的:探讨沉默GCB弥漫型大B细胞淋巴瘤(diffuse large B cell lymphoma,DLBCL)中AFAP1-AS1的表达对细胞增殖和凋亡的影响。方法:培养GCB-DLBCL细胞至对数生长期后转染OCI-Ly1细胞系,建立的GCB-DLBCL细胞系对AFAP1-AS1表达进行沉默;实验设立3组,实验组为腺病毒感染细胞,sh-NC无关序列腺病毒感染细胞组为无关序列对照组,未感染腺病毒细胞组为空白组,应用PCR法检测AFAP1-AS1表达水平、CCK-8法测定细胞增殖情况、流式细胞术检测细胞凋亡情况,对比检测结果。结果:AFAP1-AS1表达水平检测结果显示:三种shRNA序列干扰效率均较无关序列对照组(sh-NC)强,差异有统计学意义(P<0.05);采用CCK-8法检测各组细胞凋亡情况,结果显示:经腺病毒sh3-AFAP1-AS1感染后,OCI-Ly1细胞系中实验组细胞吸光度较无关序列对照组和空白组显著降低,下调AFAP1-AS1可抑制GCB-DLBCL细胞的增殖(P<0.05);采用流式细胞仪检测各组细胞凋亡情况,结果显示:经sh3-AFAP1-AS1和sh-NC转染后,OCI-Ly1细胞实验组凋亡率明显高于无关序列对照组和空白组,下调AFAP1-AS1可诱导GCB-DLBCL细胞凋亡(P<0.05)。结论:沉默GCB-DLBCL细胞中的AFAP1-AS1表达能有效抑制细胞增殖,诱导细胞凋亡,或可作为GCB-DLBCL治疗的靶目标。
基金supported by the Natural Science Foundation of China(Nos.82002782,82202657)the Guangdong Basic and Applied Basic Research Foundation(2022A1515012021,2020A1515110930).
文摘Triple-negative breast cancer(TNBC)is characterized by fast growth,high metastasis,high invasion,and a lack of therapeutic targets.Mitosis and metastasis of TNBC cells are two important biological behaviors in TNBC malignant progression.It is well known that the long noncoding RNA AFAP1-AS1 plays a crucial role in various tumors,but whether AFAP1-AS1 is involved in the mitosis of TNBC cells remains unknown.In this study,we investigated the functional mechanism of AFAP1-AS1 in targeting Polo-like Kinase 1(PLK1)activation and participating in mitosis of TNBC cells.We detected the expression of AFAP1-AS1 in the TNBC patient cohort and primary cells by in situ hybridization(ISH),northern blot,fluorescent in situ hybridization(FISH)and cell nucleus/cytoplasm RNA fraction isolation.High AFAP1-AS1 expression was negatively correlated with overall survival(OS),disease-free survival(DFS),metastasis-free survival(MFS)and recurrence-free survival(RFS)in TNBC patients.We explored the function of AFAP1-AS1 by transwell,apoptosis,immunofluorescence(IF)and patient-derived xenograft(PDX)models in vitro and in vivo.We found that AFAP1-AS1 promoted TNBC primary cell survival by inhibiting mitotic catastrophe and increased TNBC primary cell growth,migration and invasion.Mechanistically,AFAP1-AS1 activated phosphorylation of the mitosis-associated kinase PLK1 protein.Elevated levels of AFAP1-AS1 in TNBC primary cells increased PLK1 pathway downstream gene expression,such as CDC25C,CDK1,BUB1 and TTK.More importantly,AFAP1-AS1 increased lung metastases in a mouse metastasis model.Taken together,AFAP1-AS1 functions as an oncogene that activates the PLK1 signaling pathway.AFAP1-AS1 could be used as a potential prognostic marker and therapeutic target for TNBC.
