Aptamer-Driven Toxin Gene Delivery in U87 Model Glioblastoma Cells.
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ABSTRACT:
SUBMITTER: di Leandro L
PROVIDER: S-EPMC8082512 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Glioblastoma is the most aggressive primary brain tumor leading to death in most of patients. It comprises almost 50-55% of all gliomas with an incidence rate of 2-3 per 100,000. Despite its rarity, overall mortality of glioblastoma is comparable to the most frequent tumors. The current standard treatment combines surgical resection, radiotherapy and chemotherapy with temozolomide. In spite of this aggressive multimodality protocol, prognosis of glioblastoma is poor and the median survival remains about 12-14.5 months. In this regard, new therapeutic approaches should be developed to improve the life quality and survival time of the patient after the initial diagnosis. Before switching to clinical trials in humans, all innovative therapeutic methods must be studied first on a relevant animal model in preclinical settings. In this regard, we validated the feasibility of intratumoral delivery of a holmium (Ho) microparticle suspension to an induced U87 glioblastoma model. Among the different radioactive beta emitters, 166Ho emits high-energy β(-) radiation and low-energy γ radiation. β(-) radiation is an effective means for tumor destruction and γ rays are well suited for imaging (SPECT) and consequent dosimetry. In addition, the paramagnetic Ho nucleus is a good asset to perform MRI imaging. In this study, five minipigs, implanted with our glioblastoma model were used to test the injectability of 165Ho (stable) using a bespoke injector and needle. The suspension was produced in the form of Ho microparticles and injected inside the tumor by a technique known as microbrachytherapy using a stereotactic system. At the end of this trial, it was found that the 165Ho suspension can be injected successfully inside the tumor with absence or minimal traces of Ho reflux after the injections. This injection technique and the use of the 165Ho suspension needs to be further assessed with radioactive 166Ho in future studies.
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Project description:How cancer cells adapt to hypoxia during tumor development remains an important question. The hypothesis tested in the present study was that tumor cell-derived exosome vesicles (also known as microvesicles or extracellular vesicles) are mediators of hypoxia-dependent intercellular signaling in glioblastoma (GBM), i.e. highly aggressive brain tumors characterized by hypoxia and a vascular density that is among the highest of all human malignancies. In vitro hypoxia experiments and studies with patient materials reveal the enrichment in exosomes of hypoxia-regulated mRNAs and proteins, several of which were associated with poor patient prognosis. We show that cancer cell exosomes mediate hypoxia-dependent, phenotypic modulation of stromal cells in vitro and ex vivo, resulting in accelerated GBM tumor angiogenesis and growth in mice. These data suggest that exosomes constitute potent mediators of hypoxia-driven tumor development, and circulating multiparameter biomarkers of tumor hypoxia. U87 MG glioblastoma cells were grown at normoxic (21% oxygen) or hypoxic (1% oxygen) conditions for 48 hours. Conditioned media from normoxic and hypoxic cells were then used to isolate exosomes by differential centrifugation. Both cells and exosomes were lysed in Trizol reagent, and RNA was isolated.Total RNA from all samples (four types of samples in three biological repilicates) was subjected to genome-wide transcriptional analysis with Illumina HumanHT-12 V3.0 expression beadchip. Gene expression profile obtained from hypoxic U87 MG glioblastoma cells was compared to the profile of normoxic control cells. Analogically, gene expression profile obtained from hypoxic U87 MG cells was compared to the profile of exosomes secreted by normoxic U87 MG cells.
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Project description:Neocarzinostatin (NCS) is an anti-tumor DNA damaging agent. Conjugating NCS with EpCAM Aptamer will direct the toxin pay loads to the EpCAM positive cancer cells as a targeted therapy We used microarrays to detail the global gene expression to understand the pathways involved in EpCAM-mediated NCS drug delivery in breast cancer cells.
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Project description:Discovering the underlying signalling pathways that control cancer cells is crucial for understanding their biology and to develop therapeutic regimens. Thus, the aim of the present study was to determine the effect of Cripto-1 on pathways controlling glioblastoma (GBM) cell function. To this end, changes in protein phosphorylation in cells overexpressing Cripto-1 were analysed using the Kyoto Encyclopedia of Genes and Genomes pathway analysis tool, as well as the Uniprot resource to identify the functions of Cripto-1-dependent phosphorylated proteins. This revealed that proteins affected by Cripto-1 overexpression are involved in multiple signalling pathways. The mitogen-activated protein kinase (MAPK), focal adhesion (FA) and ErbB pathways were found to be enriched by Cripto-1 overexpression with 35, 27 and 24% of pathway proteins phosphorylated, respectively. These pathways control important cellular processes in cancer cells that correlate with the observed functional changes described in earlier studies. More specifically, Cripto-1 may regulate MAPK cellular proliferation and survival pathways by activating epithelial growth factor receptor (EGFR; Ser1070) or fibroblast GFR1 (Tyr654). Its effect on cellular proliferation and survival could be mediated through Src (Tyr418), FA kinase (FAK; Tyr396), p130CAS (Tyr410), c-Jun (Ser63), Paxillin (PXN; Tyr118) and BCL2 (Thr69) of the FA pathway. Cripto-1 may also control cellular motility and invasion by activating Src (Tyr418), FAK (Tyr396) and PXN (Tyr118) of the FA pathway. However, Cripto-1 regulation of cellular invasion and migration might be not limited to the FA pathway, it may also control these cellular mechanisms through signalling via EGFR (Ser1070)/Her2 (Tyr877) to mediate the Src (Tyr418) and FAK (Tyr396) cascade activation of the ErbB signalling pathway. Angiogenesis could be mediated by Cripto-1 by activating c-Jun (Ser63) through EGFR (Ser1070)/Her2 (Tyr877) of the ErbB pathway. To conclude, the present study has augmented and enriched our current knowledge on the crucial roles that Cripto-1 may play in controlling different cellular mechanisms in GBM cells.
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