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Gemcitabine-induced heparanase promotes aggressiveness of pancreatic cancer cells via activating EGFR signaling.

ABSTRACT: Pancreatic cancer (PC), characterized by aggressive local invasion and metastasis, is one of the most malignant cancers. Gemcitabine is currently used as the standard drug for the treatment of advanced and metastatic PC, but with limited efficacy. In this study, we demonstrated that gemcitabine increased the expression of heparanase (HPA1), the only known mammalian endoglycosidase capable of cleaving heparan sulfate, both in vitro and in vivo. Furthermore, overexpression of HPA1 in PC cell lines enhanced proliferation and invasion, accompanied with elevated phosphorylation of EGFR. In addition, we showed that the NF-?B pathway mediated the gemcitabine-induced HPA1 expression. Importantly, we found that an HPA1 inhibitor attenuated gemcitabine-induced invasion of PC cells. Finally, we showed that HPA1 was of negative prognostic value for PC patients. Taken together, our results demonstrated that gemcitabine-induced HPA1 promotes proliferation and invasion of PC cells through activating EGFR, implying that HPA1 may serve as promising therapeutic target in the treatment of PC.

SUBMITTER: Song JW

PROVIDER: S-EPMC5601663 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Gemcitabine-induced heparanase promotes aggressiveness of pancreatic cancer cells via activating EGFR signaling.

Song Jin-Wen JW Tan Ying-Xia YX Li Su-Bo SB Zhang Shi-Kun SK Wan Lu-Ming LM Ji Shou-Ping SP Zhou Hong H Zhou Zhi-Hang ZH Gong Feng F

Oncotarget 20170407 35

Pancreatic cancer (PC), characterized by aggressive local invasion and metastasis, is one of the most malignant cancers. Gemcitabine is currently used as the standard drug for the treatment of advanced and metastatic PC, but with limited efficacy. In this study, we demonstrated that gemcitabine increased the expression of heparanase (HPA1), the only known mammalian endoglycosidase capable of cleaving heparan sulfate, both <i>in vitro</i> and <i>in vivo</i>. Furthermore, overexpression of HPA1 in ...[more]

PMID: 28938567

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Project description:BackgroundPancreatic cancer (PC) is a highly lethal malignancy regarding digestive system, which is the fourth leading factor of cancer-related mortalities in the globe. Prognosis is poor due to diagnosis at advanced disease stage, low rates of surgical resection, and resistance to traditional radiotherapy and chemotherapy. In order to develop novel therapeutic strategies, further elucidation of the molecular mechanisms underlying PC chemoresistance is required. Ribosomal RNA biogenesis has been implicated in tumorigenesis. Small nucleolar RNAs (snoRNAs) is responsible for post-transcriptional modifications of ribosomal RNAs during biogenesis, which have been identified as potential markers of various cancers. Here, we investigate the U3 snoRNA-associated protein RRP9/U3-55 K along with its role in the development of PC and gemcitabine resistance.MethodsqRT-PCR, western blot and immunohistochemical staining assays were employed to detect RRP9 expression in human PC tissue samples and cell lines. RRP9-overexpression and siRNA-RRP9 plasmids were constructed to test the effects of RRP9 overexpression and knockdown on cell viability investigated by MTT assay, colony formation, and apoptosis measured by FACS and western blot assays. Immunoprecipitation and immunofluorescence staining were utilized to demonstrate a relationship between RRP9 and IGF2BP1. A subcutaneous xenograft tumor model was elucidated in BALB/c nude mice to examine the RRP9 role in PC in vivo.ResultsSignificantly elevated RRP9 expression was observed in PC tissues than normal tissues, which was negatively correlated with patient prognosis. We found that RRP9 promoted gemcitabine resistance in PC in vivo and in vitro. Mechanistically, RRP9 activated AKT signaling pathway through interacting with DNA binding region of IGF2BP1 in PC cells, thereby promoting PC progression, and inducing gemcitabine resistance through a reduction in DNA damage and inhibition of apoptosis. Treatment with a combination of the AKT inhibitor MK-2206 and gemcitabine significantly inhibited tumor proliferation induced by overexpression of RRP9 in vitro and in vivo.ConclusionsOur data reveal that RRP9 has a critical function to induce gemcitabine chemoresistance in PC through the IGF2BP1/AKT signaling pathway activation, which might be a candidate to sensitize PC cells to gemcitabine. Video abstract.

Project description:BackgroundThe prognosis of human glioma is poor, and the highly invasive nature of the disease represents a major impediment to current therapeutic modalities. The oncoprotein B-cell-specific Moloney murine leukemia virus integration site 1 protein (Bmi-1) has been linked to the development and progression of glioma; however, the biological role of Bmi-1 in the invasion of glioma remains unclear.MethodsA172 and LN229 glioma cells were engineered to overexpress Bmi-1 via stable transfection or to be silenced for Bmi-1 expression using RNA interfering method. Migration and invasiveness of the engineered cells were assessed using wound healing assay, Transwell migration assay, Transwell matrix penetration assay and 3-D spheroid invasion assay. MMP-9 expression and activity were measured using real-time PCR, ELISA and the gelatin zymography methods. Expression of NF-kappaB target genes was quantified using real-time PCR. NF-kappaB transcriptional activity was assessed using an NF-kappaB luciferase reporter system. Expression of Bmi-1 and MMP-9 in clinical specimens was analyzed using immunohistochemical assay.ResultsEctopic overexpression of Bmi-1 dramatically increased, whereas knockdown of endogenous Bmi-1 reduced, the invasiveness and migration of glioma cells. NF-kappaB transcriptional activity and MMP-9 expression and activity were significantly increased in Bmi-1-overexpressing but reduced in Bmi-1-silenced cells. The reporter luciferase activity driven by MMP-9 promoter in Bmi-1-overexpressing cells was dependent on the presence of a functional NF-kappaB binding site, and blockade of NF-kappaB signaling inhibited the upregulation of MMP-9 in Bmi-1 overexpressing cells. Furthermore, expression of Bmi-1 correlated with NF-kappaB nuclear translocation as well as MMP-9 expression in clinical glioma samples.ConclusionsBmi-1 may play an important role in the development of aggressive phenotype of glioma via activating the NF-kappaB/MMP-9 pathway and therefore might represent a novel therapeutic target for glioma.

Project description:BackgroundGlioma is the most frequent and lethal primary brain malignancy. Amounting evidence has highlighted the importance of exosomal microRNAs (miRNAs or miRs) in this malignancy. This study aimed to investigate the regulatory role of exosomal miR-148a-3p in glioma.MethodsBioinformatics analysis was firstly used to predict the target genes of miR-148a-3p. Exosomes were then extracted from normal human astrocytes and glioma cells. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was applied to determine the expression patterns of miR-148a-3p and ERBB receptor feedback inhibitor 1 (ERRFI1). Dual-luciferase reporter gene assay was applied to verify the direct binding between miR-148a-3p and ERRFI1. Cell counting kit-8 and tube formation assays were further conducted to assess the proliferation and angiogenic properties of human umbilical vein endothelial cells (HUVECs) in the co-culture system with exosomes. Lastly, glioma tumor models were established in BALB/c nude mice to study the role of exosomal miR-148a-3p in vivo.ResultsmiR-148a-3p was highly expressed, while ERRFI1 was poorly expressed in glioma. miR-148a-3p was found to be enriched in glioma cells-derived exosomes and could be transferred to HUVECs via exosomes to promote their proliferation and angiogenesis. ERRFI1 was identified as a target gene of miR-148a-3p. In addition, miR-148a-3p activated the epidermal growth factor receptor (EGFR)/mitogen-activated protein kinase (MAPK) signaling pathway by inhibiting ERRFI1. In the co-culture system, our data demonstrated that glioma cells-derived exosomal miR-148a-3p down-regulated ERRFI1 and activated the EGFR/MAPK signaling pathway, so as to promote cell proliferation and angiogenesis. In vivo experimentation further demonstrated that this mechanism was responsible for the promotive role of exosomal miR-148a-3p in tumorigenesis and angiogenesis.ConclusionTaken together, glioma-derived exosomal miR-148a-3p promoted tumor angiogenesis through activation of the EGFR/MAPK signaling pathway by ERRFI1 inhibition.

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Gemcitabine-induced heparanase promotes aggressiveness of pancreatic cancer cells via activating EGFR signaling.

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Gemcitabine-induced heparanase promotes aggressiveness of pancreatic cancer cells via activating EGFR signaling.

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