MicroRNA-122-5p inhibits cell proliferation, migration and invasion by targeting CCNG1 in pancreatic ductal adenocarcinoma
MicroRNA-122-5p 通过靶向 CCNG1 抑制胰腺导管腺癌细胞增殖、迁移和侵袭
- 作者列表："Chen Dai","Yan Zhang","Zhihua Xu","Mengxian Jin
Abstract Background Pancreatic ductal adenocarcinoma (PDAC) is a lethal human malignancy, and previous researches support the contribution of microRNA (miRNA) to cancer progression. MiR-122-5p is reported to participate in the regulation of various cancers, while the function of miR-122-5p in PDAC remains unclear. In this study, we investigated the precise mechanism of miR-122-5p involved in PDAC pathogenesis. Methods The expression levels of miR-122-5p were detected in human PDAC tissues and cell lines by miRNA RT-PCR. The effects of miR-122-5p on cell proliferation were explored by MTT assays, colony formation assays and flow cytometry assays. The ability of migration and invasion was determined by transwell assays. Dual Luciferase reporter assay was performed to validate the direct interaction between miR-122-5p and its target gene. The related molecules of cell cycle, apoptosis and epithelial–mesenchymal transition (EMT) were examined with qRT-PCR and western blot. In addition, xenograft mouse models were applied to explore the effects of miR-122-5p in vivo. Results MiR-122-5p was underexpressed, while CCNG1 was highly expressed in PDAC tissues and cells. MiR-122-5p was negatively correlated with TNM stage, tumor size and lymph node metastasis in PDAC patients. Overexpression of miR-122-5p suppressed the proliferation, migration and invasion in vitro and inhibited tumorigenesis in vivo. Furthermore, CCNG1 was a direct target of miR-122-5p. Upregulated CCNG1 could partially reverse the effects caused by miR-122-5p. Moreover, miR-122-5p inhibited EMT through downregulation of CCNG1. Conclusion Overexpression of miR-122-5p could inhibit cell proliferation, migration, invasion, and EMT by downregulating CCNG1 in PDAC, suggesting a potential therapeutic target for PDAC.
背景胰腺导管腺癌 (PDAC) 是一种致死性人类恶性肿瘤，既往研究支持 microRNA (miRNA) 在癌症进展中的作用。据报道 MiR-122-5p 参与多种癌症的调节，而 PDAC 中 miR-122-5p 的功能仍不清楚。在本研究中，我们探讨了 miR-122-5p 参与 PDAC 发病的确切机制。方法采用 miRNA RT-PCR 方法检测人 PDAC 组织和细胞系中 miR-122-5p 的表达水平。采用 MTT 法、集落形成法及流式细胞仪检测 miR-122-5p 对细胞增殖的影响。通过 transwell 试验确定迁移和侵袭的能力。通过双荧光素酶报告基因检测来验证 miR-122-5p 与其靶基因之间的直接相互作用。用 qRT-PCR 和 western blot 检测细胞周期、凋亡和上皮间质转化 (EMT) 的相关分子。此外，应用异种移植小鼠模型，探讨 miR-122-5p 在体内的作用。结果 MiR-122-5p 表达不足，而 CCNG1 在 PDAC 组织和细胞中高表达。PDAC 患者 MiR-122-5p 与 TNM 分期、肿瘤大小、淋巴结转移呈负相关。过表达 miR-122-5p 抑制体外增殖、迁移和侵袭，抑制体内肿瘤发生。此外，CCNG1 是 miR-122-5p 的直接靶点。上调的 CCNG1 可部分逆转 miR-122-5p 引起的效应。此外，miR-122-5p 通过下调 ccng1 抑制 EMT。结论过表达 miR-122-5p 可通过下调 CCNG1 抑制 PDAC 细胞的增殖、迁移、侵袭和 EMT，提示其可能是 PDAC 的潜在治疗靶点。
METHODS::Pancreatic ductal adenocarcinoma (PDAC) is a disease of aging. The TP53 gene product regulates cell growth, aging, and cancer. To determine the important targets of TP53 in PDAC, we examined the expression of 440 proteins on a reverse phase protein array (RPPA) in PDAC-derived MIA-PaCa-2 cells which either had WT-TP53 or lacked WT-TP53. MIA-PaCa-2 cells have a TP53 mutation as well as mutant KRAS and represent a good in vitro model to study PDAC. RPPA analysis demonstrated expression of tumor promoting proteins in cells that lacked WT-TP53; and this feature could be reversed significantly when the cells were transfected with vector encoding WT-TP53 or treated with berberine or a modified berberine (BBR). Expression of miR-34a-associated signaling was elevated in cells expressing WT-TP53 compared to cells expressing mTP53. Results from in vivo studies using human PDAC specimens confirmed the in vitro results as the expression of miR-34a and associated signaling was significantly decreased in PDAC specimens compared to non-cancerous tissues. This study determined SERPINE1 as a miR-34a target with relevance to the biology of PDAC. Thus, we have identified a key target (SERPINE1) of the TP53/miR-34a axis that may serve as a potential biomarker for early detection of pancreatic cancer.
METHODS::Background: SLC6A14 (ATB0,+), a Na+/Cl-coupled transporter for neutral/cationic amino acids, is overexpressed in many cancers; It has been investigated as a target for improved liposomal drug delivery to treat liver cancer.Research design and methods: Here we explored the mechanism of ATB0,+-mediated entry of such liposomes. As ATB0,+ is highly-expressed in pancreatic cancer, we also examined the therapeutic utility of ATB0,+-targeted liposomal drug delivery to treat this cancer.Results: The uptake of lysine-conjugated liposomes (LYS-LPs) was greater in ATB0,+-positive MCF7 cells. The uptake process consisted of two steps: binding and internalization. The binding of LYS-LPs to MCF7 cells was higher than that of bare liposomes, and the process was dependent on Na+ and Cl-, and inhibitable by ATB0,+ substrates or blocker. In contrast, the internalization step was independent of lysine. The cellular entry of LYS-LPs facilitated by ATB0,+ occurred via endocytosis with transient endosomal degradation of ATB0,+ protein with subsequent recovery. Moreover, LYS-LPs also enhanced the uptake and cytotoxicity of gemcitabine in these cells in an ATB0,+-dependent manner.Conclusions: We conclude that ATB0,+ could be exploited for targeted drug delivery in the form of lysine-conjugated liposomes and that the approach represents a novel strategy for enhanced pancreatic cancer therapy.
METHODS:PURPOSE:Pre-operative prediction of histological response to neoadjuvant therapy aids decisions regarding surgical management of borderline resectable pancreatic cancer (BRPC). We elucidate correlation between pre-/post-treatment whole-tumor apparent diffusion coefficient (ADC) value and rate of tumor cell destruction. We newly verify whether post-treatment ADC value at the site of vascular contact predicts R0 resectability of BRPC. METHODS:We prospectively reviewed 28 patients with BRPC who underwent diffusion-weighted magnetic resonance imaging before neoadjuvant chemotherapy and surgery. Correlation between the percentage of tumor cell destruction and various parameters was analyzed. Strong parameters were assessed for their ability to predict therapeutic histological response and R0 resectability. RESULTS:Pre-/post-treatment whole-tumor ADC value correlated with tumor cell destruction rate by all parameters (R = 0.630/0.714, P 50% was determined at 1.40 × 10-3 mm2/s. It predicts histological response with 100% sensitivity, 81% specificity, and 89% accuracy. It predicts R0 with 88% sensitivity, 70% specificity, and 75% accuracy. CONCLUSIONS:Post-treatment whole-tumor ADC value may be a predictor of R0 resectability in patients with BRPC. Tumor cell destruction rate is indicated by the difference between pre-/post-treatment ADC values. This difference is strongly affected by the pre-treatment ADC value. The cutoff value of ADC at the site of vascular contact could not discriminate R0 resectability.