RESUMEN
Oxaliplatin (cyclohexane-1,2-diamine; oxalate; platinum [2+]) is a third-generation chemotherapeutic drug with anticancer effects. Oxaliplatin has a role in the treatment of several cancers. It is one of the few drugs which can eliminate the neoplastic cells of colorectal cancer. Also, it has an influential role in breast cancer, lung cancer, bladder cancer, prostate cancer, and gastric cancer. Although oxaliplatin has many beneficial effects in cancer treatment, resistance to this drug is in the way to cure neoplastic cells and reduce treatment efficacy. microRNAs are a subtype of small noncoding RNAs with â¼22 nucleotides that exist among species. They have diverse roles in physiological processes, including cellular proliferation and cell death. Moreover, miRNAs have essential roles in resistance to cancer treatment and can strengthen sensitivity to chemotherapeutic drugs and regimens. In colorectal cancer, the co-treatment of oxaliplatin with anti-miR-19a can partially reverse the oxaliplatin resistance through the upregulation of phosphatase and tensin homolog (PTEN). Moreover, by preventing the spread of gastric cancer cells and downregulating glypican-3 (GPC3), MiR-4510 may modify immunosuppressive signals in the tumor microenvironment. Treatment with oxaliplatin may develop into a specialized therapeutic drug for patients with miR-4510 inhibition and glypican-3-expressing gastric cancer. Eventually, miR-122 upregulation or Wnt/ß-catenin signaling suppression boosted the death of HCC cells and made them more sensitive to oxaliplatin. Herein, we have reviewed the role of microRNAs in regulating cancer cells' response to oxaliplatin, with particular attention to gastrointestinal cancers. We also discussed the role of these noncoding RNAs in the pathophysiology of oxaliplatin-induced neuropathic pain.
Asunto(s)
Carcinoma Hepatocelular , Neoplasias Colorrectales , Neoplasias Hepáticas , MicroARNs , Neoplasias Gástricas , Masculino , Humanos , MicroARNs/metabolismo , Oxaliplatino/farmacología , Glipicanos/metabolismo , Glipicanos/farmacología , Glipicanos/uso terapéutico , Neoplasias Gástricas/patología , Apoptosis , Resistencia a Antineoplásicos , Línea Celular Tumoral , Neoplasias Hepáticas/genética , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Regulación Neoplásica de la Expresión Génica , Proliferación Celular , Microambiente TumoralRESUMEN
Several studies including ours reported the detrimental effects of extracellular tau oligomers (ex-oTau) on glutamatergic synaptic transmission and plasticity. Astrocytes greatly internalize ex-oTau whose intracellular accumulation alters neuro/gliotransmitter handling thereby negatively affecting synaptic function. Both amyloid precursor protein (APP) and heparan sulfate proteoglycans (HSPGs) are required for oTau internalization in astrocytes but the molecular mechanisms underlying this phenomenon have not been clearly identified yet. Here we found that a specific antibody anti-glypican 4 (GPC4), a receptor belonging to the HSPG family, significantly reduced oTau uploading from astrocytes and prevented oTau-induced alterations of Ca2+-dependent gliotransmitter release. As such, anti-GPC4 spared neurons co-cultured with astrocytes from the astrocyte-mediated synaptotoxic action of ex-oTau, thus preserving synaptic vesicular release, synaptic protein expression and hippocampal LTP at CA3-CA1 synapses. Of note, the expression of GPC4 depended on APP and, in particular, on its C-terminal domain, AICD, that we found to bind Gpc4 promoter. Accordingly, GPC4 expression was significantly reduced in mice in which either APP was knocked-out or it contained the non-phosphorylatable amino acid alanine replacing threonine 688, thus becoming unable to produce AICD. Collectively, our data indicate that GPC4 expression is APP/AICD-dependent, it mediates oTau accumulation in astrocytes and the resulting synaptotoxic effects.
Asunto(s)
Precursor de Proteína beta-Amiloide , Glipicanos , Animales , Ratones , Péptidos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Astrocitos/metabolismo , Glipicanos/metabolismo , Glipicanos/farmacología , Neuronas/metabolismo , Transmisión Sináptica/fisiologíaRESUMEN
Constantly stimulated by the tumor microenvironment (TME), programmed death 1 (PD1) is elevated, and it interacts with PD ligand 1 (PDL1), rendering chimeric antigen receptor (CAR)T cells dysfunctional. Hence, CART cells immune to PD1induced immunosuppression were constructed to improve the function of CART cells in hepatocellular carcinoma (HCC). Doubletarget CART cells, targeting glypican3 (GPC3) [a tumour-associated antigen (TAA)] and hindering PD1PDL1 binding, were established. The expression of GPC3, PDL1, and inhibitory receptors was measured using flow cytometry. The cytotoxicity, cytokine release, and differentiation level of CART cells were determined using lactate dehydrogenase release assay, enzymelinked immunosorbent assay, and flow cytometry, respectively. HCC cells were targeted and eliminated by doubletarget CART cells. These doubletarget CART cells limit PD1PDL1 binding and sustain cytotoxicity to PDL1+ HCC cells. The relatively low IR expression and differentiation level in doubletarget CART cells in tumour tissues induced tumoursuppression and extended survival in PDL1+ HCC TX models, as opposed to their singletarget counterparts. The results of the present study suggested that the newly constructed doubletarget CART cells exhibit stronger tumoursuppressing effects in HCC than their singletarget counterparts, which are common, suggesting the potential of strengthening CART cell activity in HCC treatment.
Asunto(s)
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/patología , Antígeno B7-H1/metabolismo , Receptor de Muerte Celular Programada 1/metabolismo , Glipicanos/metabolismo , Glipicanos/farmacología , Linfocitos T , Microambiente TumoralRESUMEN
During placentation, cytotrophoblasts differentiate and fuse to form multinucleated cells (syncytiotrophoblasts) in a process that involves autophagy. Appropriate syncytial differentiation is essential for establishing a healthy pregnancy. In this study, we evaluated the effect of two chloroquine compounds, hydroxychloroquine (HCQ) and chloroquine (CQ), on syncytial differentiation and autophagy in cultured primary human trophoblasts (PHTs). PHT cells were isolated from the human term placenta. Bafilomycin, a well-known autophagy inhibitor, was used as a positive control. Biochemical and morphological differentiation was assessed in syncytiotrophoblasts, and autophagy-related proteins and genes were evaluated. Affymetrix Human Gene 2.0 ST Array profiling was used to identify genes affected by HCQ during syncytial differentiation. Chloroquine compounds lowered the production of beta-human chorionic gonadotropin (ß-hCG) and the fusion index in PHTs. Syncytial differentiation in PHT was associated with the increased expression of ATG4C mRNA (autophagy-related gene), and this expression was affected by CQ but not by HCQ. Microarray analysis revealed that HCQ or CQ affected several genes (MMP15, GPC3, CXCL10, TET-1, and S100A7) during syncytial differentiation, which were different from that of the syncytial differentiation suppression (Ham's/Waymouth media) or autophagy inhibition (bafilomycin treatment). Using Kyoto Encyclopedia of Genes and Genomes analysis we identified that HCQ might affect JAK2 signaling in the syncytial differentiation of PHT. In conclusion, chloroquine compounds could mitigate biochemical and morphological syncytial trophoblast differentiation in cultured PHT cells through the JAK signaling pathway rather than the inhibition of autophagic activity.
Asunto(s)
Cloroquina , Hidroxicloroquina , Autofagia , Diferenciación Celular , Cloroquina/farmacología , Femenino , Glipicanos/metabolismo , Glipicanos/farmacología , Humanos , Hidroxicloroquina/farmacología , Embarazo , Trofoblastos/metabolismoRESUMEN
Pancreatic ductal adenocarcinoma (PDAC) is a stroma-rich cancer. Extracellular matrix proteins produced by cancer-associated fibroblasts (CAFs) found in tumor stroma that impedes effective delivery of chemotherapeutic agents results in poor response in patients with PDAC. Previously, our group reported that glypican-1 (GPC1) was overexpressed in human PDAC and negatively correlated with patient survival. Immunohistochemical analysis of 25 patients with PDAC tumor specimens revealed elevated expression of GPC1 in stromal cells and pancreatic cancer cells in 80% of patients. Interestingly, GPC1 was expressed on CAFs in PDAC. We generated a GPC1 antibody-drug conjugate conjugated with monomethyl auristatin E [GPC1-ADC(MMAE)] and evaluated its preclinical antitumor activity by targeting GPC1-positive CAF and cancer cells in PDAC. GPC1-ADC(MMAE) inhibited the growth of GPC1-positive PDAC cell lines in vitro Furthermore, GPC1-ADC(MMAE) showed a potent antitumor effect in the PDAC patient-derived tumor xenograft (PDX) model against GPC1-positive CAF and heterogeneous GPC1-expressing cancer cells. Notably, GPC1-ADC(MMAE) showed robust preclinical efficacy against GPC1 in a stroma-positive/cancer-negative PDAC PDX model. GPC1-ADC(MMAE) was delivered and internalized to CAFs. Although apoptosis was not observed in CAFs, the released MMAE from CAFs via MDR-1 induced apoptosis of cancer cells neighboring CAFs and efficiently inhibited PDAC tumor growth. GPC1-ADC(MMAE) exhibited potent and unique antitumor activity in GPC1-positive PDAC PDX models, which suggests that GPC1 is a novel therapeutic target in PDAC and other stromal GPC1-positive solid tumors. These findings show that targeting GPC1 on CAF using GPC1-ADC(MMAE) is a useful approach in case of stroma-rich tumors such as PDAC.
Asunto(s)
Adenocarcinoma/tratamiento farmacológico , Carcinoma Ductal Pancreático/tratamiento farmacológico , Glipicanos/uso terapéutico , Inmunoconjugados/uso terapéutico , Animales , Glipicanos/farmacología , Humanos , Inmunoconjugados/farmacología , Ratones , Ratones Endogámicos NODRESUMEN
Hepatocellular carcinoma (HCC) is the most frequent primary liver cancer but has shown limited success to date in the treatment of advanced stage. Recruitment of T cells for cancer treatment is a rapidly growing strategy in immunotherapy such as chimeric antigen receptor T cells and bispecific antibodies. However, unwanted aggregations, structural instability or short serum half-life are major challenges of bispecific antibodies. Here, we developed a new format of T cell-redirecting antibody that is bispecific for membrane proteoglycans GPC3 of HCC and the T-cell-specific antigen CD3, which demonstrated to be favorable stability and productivity. Cross-linking of T cells with GPC3 positive tumor cells by the anti-GPC3/CD3 bispecific antibody-mediated potent GPC3-dependent and concentration-dependent cytotoxicity in vitro. Administration of the bispecific antibody with different concentrations in murine xenograft models of human HCC significantly inhibited tumor growth. In addition, no effects on tumor growth were observed in the absence of human effector cells or the bispecific antibody. Taken together, the anti-GPC3/CD3 bispecific antibody might be a potential therapeutic treatment for HCC.
Asunto(s)
Anticuerpos Biespecíficos/metabolismo , Carcinoma Hepatocelular/genética , Glipicanos/uso terapéutico , Inmunoterapia/métodos , Neoplasias Hepáticas/genética , Animales , Carcinoma Hepatocelular/patología , Femenino , Glipicanos/farmacología , Humanos , Neoplasias Hepáticas/patología , Ratones , Ratones Endogámicos NODRESUMEN
Craniosynostosis is a complex disease condition, which involves premature fusion of cranial vault sutures and lacks desirable treatment. Previous studies have demonstrated decreased proliferation rate of osteoblasts and downregulated expression of glypican 3 (GPC3) in syndromic craniosynostosis patients. In this study, quantitative and qualitative analysis were utilized to assess the effect of GPC3 in human fetal osteoblastic cell line, hFOB 1.19. Lentiviral transfection efficiency with green fluorescent protein images was obtained after 72âhours. Western Blot and quantitative real-time polymerase chain reaction analysis results indicated that GPC3 was overexpressed in hFOB 1.19 cells transfected with recombinant lentivirus LV-GPC3-GFP. Cell proliferation was assessed by CCK-8 assay and cell cycle progression and apoptosis were analyzed by flow cytometric assay. Results revealed that GPC3 promoted cell viability, induced cell cycle entry into S phase, and inhibited cell apoptosis. These findings provide novel ideas in understanding the pathogenesis of craniosynostosis. It also provides novel insights in the treatment of craniosynostosis by targeting GPC3.
Asunto(s)
Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Glipicanos/genética , Glipicanos/farmacología , Osteoblastos , Línea Celular , Glipicanos/análisis , Glipicanos/metabolismo , Humanos , Osteoblastos/citología , Osteoblastos/efectos de los fármacos , TransfecciónRESUMEN
Glypicans are multifunctional proteoglycans with regulatory roles in several intercellular signaling pathways. Here, we examine the functional requirements for glypican regulation of bone morphogenetic protein (BMP)-mediated body length in C. elegans. We provide evidence that two parts of C. elegans glypican LON-2 can independently inhibit BMP signaling in vivo: the N-terminal furin protease product and the C-terminal region containing heparan sulfate attachment sequences. While the C-terminal protease product is dispensable for LON-2 minimal core protein activity, it does affect the localization of LON-2. Cleavage of LON-2 into two parts at the conserved furin protease site is not required for LON-2 to inhibit BMP-like signaling. The glycosyl-phosphatidylinositol (GPI) membrane anchor is also not absolutely required for LON-2 activity. Finally, we show that an RGD protein-protein interaction motif in the LON-2 N-terminal domain is necessary for LON-2 core protein activity, suggesting that LON-2 inhibits BMP signaling by acting as a scaffold for BMP and an RGD-binding protein.
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Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/crecimiento & desarrollo , Glipicanos/metabolismo , Transducción de Señal/fisiología , Animales , Tamaño Corporal/genética , Tamaño Corporal/fisiología , Pesos y Medidas Corporales , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/farmacología , Glicosilfosfatidilinositoles/metabolismo , Glipicanos/farmacología , Microscopía Confocal , Estructura Terciaria de Proteína/fisiología , Transducción de Señal/efectos de los fármacosRESUMEN
Obesity, especially visceral obesity, is associated with insulin resistance and metabolic syndrome. We previously identified the cell surface proteoglycan glypican-4 as differentially expressed in subcutaneous versus visceral white fat depots. Here we show that glypican-4 is released from cells and adipose tissue explants of mice, and that circulating glypican-4 levels correlate with BMI and insulin sensitivity in humans. Furthermore, glypican-4 interacts with the insulin receptor, enhances insulin receptor signaling, and enhances adipocyte differentiation. Conversely, depletion of glypican-4 results in reduced activation of the insulin receptor and prevents adipocyte differentiation in vitro by inhibiting insulin-mediated C/EBPß phosphorylation. These functions of glypican-4 are independent of its glycosylphosphatidylinositol membrane anchorage, as a nonmembrane-bound mutant of glypican-4 phenocopies the effects of native glypican-4 overexpression. In summary, glypican-4 is a novel circulating insulin sensitizing adipose-derived factor that, unlike other insulin sensitizers, acts directly on the insulin receptor to enhance signaling.
Asunto(s)
Glipicanos/farmacología , Receptor de Insulina/efectos de los fármacos , Adipocitos , Animales , Diferenciación Celular/efectos de los fármacos , Glipicanos/biosíntesis , Humanos , Grasa Intraabdominal/metabolismo , Ratones , Receptor de Insulina/metabolismoRESUMEN
In the developing central nervous system (CNS), the control of synapse number and function is critical to the formation of neural circuits. We previously demonstrated that astrocyte-secreted factors powerfully induce the formation of functional excitatory synapses between CNS neurons. Astrocyte-secreted thrombospondins induce the formation of structural synapses, but these synapses are postsynaptically silent. Here we use biochemical fractionation of astrocyte-conditioned medium to identify glypican 4 (Gpc4) and glypican 6 (Gpc6) as astrocyte-secreted signals sufficient to induce functional synapses between purified retinal ganglion cell neurons, and show that depletion of these molecules from astrocyte-conditioned medium significantly reduces its ability to induce postsynaptic activity. Application of Gpc4 to purified neurons is sufficient to increase the frequency and amplitude of glutamatergic synaptic events. This is achieved by increasing the surface level and clustering, but not overall cellular protein level, of the GluA1 subunit of the AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) glutamate receptor (AMPAR). Gpc4 and Gpc6 are expressed by astrocytes in vivo in the developing CNS, with Gpc4 expression enriched in the hippocampus and Gpc6 enriched in the cerebellum. Finally, we demonstrate that Gpc4-deficient mice have defective synapse formation, with decreased amplitude of excitatory synaptic currents in the developing hippocampus and reduced recruitment of AMPARs to synapses. These data identify glypicans as a family of novel astrocyte-derived molecules that are necessary and sufficient to promote glutamate receptor clustering and receptivity and to induce the formation of postsynaptically functioning CNS synapses.
Asunto(s)
Astrocitos/metabolismo , Potenciales Postsinápticos Excitadores/fisiología , Glipicanos/metabolismo , Receptores AMPA/metabolismo , Sinapsis/metabolismo , Animales , Astrocitos/citología , Cerebelo/citología , Cerebelo/metabolismo , Medios de Cultivo Condicionados/metabolismo , Medios de Cultivo Condicionados/farmacología , Femenino , Glipicanos/deficiencia , Glipicanos/farmacología , Hipocampo/citología , Hipocampo/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratas , Ratas Sprague-Dawley , Células Ganglionares de la Retina/citología , Células Ganglionares de la Retina/efectos de los fármacos , Células Ganglionares de la Retina/metabolismo , Sinapsis/efectos de los fármacos , Sinapsis/patologíaRESUMEN
Physical and chemical constraints imposed by the periinfarct glial scar may contribute to the limited clinical improvement often observed after ischemic brain injury. To investigate the role of some of these mediators in outcome from cerebral ischemia, we treated rats with the growth-inhibitory chondroitin sulfate proteoglycan neurocan, the growth-stimulating heparan sulfate proteoglycan glypican, or the chondroitin sulfate proteoglycan-degrading enzyme chondroitinase ABC. Neurocan, glypican, or chondroitinase ABC was infused directly into the infarct cavity for 7 d, beginning 7 d after middle cerebral artery occlusion. Glypican and chondroitinase ABC reduced glial fibrillary acidic protein immunoreactivity and increased microtubule-associated protein-2 immunoreactivity in the periinfarct region, and glypican- and chondroitinase ABC-treated rats showed behavioral improvement compared with neurocan- or saline-treated rats. Glypican and chondroitinase ABC also increased neurite extension in cortical neuron cultures. Glypican increased fibroblast growth factor-2 expression and chondroitinase ABC increased brain-derived neurotrophic factor expression in these cultures, whereas no such effects were seen following neurocan treatment. Thus, treatment with glypican or enzymatic disruption of neurocan with chondroitinase ABC improves gross anatomical, histological, and functional outcome in the chronic phase of experimental stroke in rats. Changes in growth factor expression and neuritogenesis may help to mediate these effects.
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Condroitina ABC Liasa/farmacología , Glipicanos/farmacología , Neurocano/farmacología , Accidente Cerebrovascular/tratamiento farmacológico , Animales , Western Blotting , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Condroitina ABC Liasa/administración & dosificación , Condroitina ABC Liasa/uso terapéutico , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Proteína Ácida Fibrilar de la Glía/inmunología , Glipicanos/administración & dosificación , Glipicanos/uso terapéutico , Inmunohistoquímica , Infusiones Intraarteriales , Proteínas Asociadas a Microtúbulos/inmunología , Neuritas/efectos de los fármacos , Neurocano/administración & dosificación , Neurocano/uso terapéutico , Desempeño Psicomotor/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Accidente Cerebrovascular/enzimologíaRESUMEN
OBJECTIVE: To construct glypican-3 (GPC3)-green fluorescent protein eukaryotic expression vector pEGFP-c3-GPC3, and analyze the effect of GPC3 on the proliferation of human hepatoma cell line MHCC-97L. METHODS: The eukaryotic expression vector pEGFP-c3-GPC3 was constructed with recombinant DNA technique and transfected into MHCC-97L cells via Lipofectamine 2000. The cells stably expressing GPC3 were screened by flow cytometry and G418. The mRNA expression of GPC3 was detected by RT-QPCR method, and the protein expression by Western blotting and fluorescence microscope. The effect of GPC3 gene on the growth of the cells was examined by MTT assay. RESULTS: Restriction endonuclease analysis and DNA sequencing verified correct construction of the recombinant plasmid. The green fluorescence was detected in the transfected MHCC-97L cells under fluorescence microscope. RT-QPCR and Western blotting both confirmed successful expression of GPC3 in MHCC-97L cells. The growth curve showed a significant acceleration of the proliferation of the transfected MHCC97-Lsol;GPC3 cells as compared with MHCC97-L and MHCC97-L/C3 cells (P<0.001). CONCLUSION: We have successfully constructed the eukaryotic expression vector pEGFR-c3-GPC3, which allows stable GPC3 expression in MHCC97-L/GPC3 cells. The upregulation of GPC3 expression can stimulate the growth of hepatoma cell line MHCC97-L in vitro.
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Proliferación Celular/efectos de los fármacos , Vectores Genéticos , Glipicanos/farmacología , Transfección , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , Proteínas Fluorescentes Verdes/genética , Humanos , Neoplasias Hepáticas/patología , PlásmidosAsunto(s)
Carcinoma Hepatocelular/metabolismo , Glipicanos/metabolismo , Neoplasias Hepáticas/metabolismo , Carcinoma Hepatocelular/patología , Proliferación Celular/efectos de los fármacos , Citometría de Flujo , Glipicanos/farmacología , Células HEK293 , Humanos , Neoplasias Hepáticas/patología , Proteínas RecombinantesRESUMEN
Simpson-Golabi-Behmel syndrome (SGBS) is an X-linked condition shown to be the result of deletions of the glypican-3 (GPC3) gene. GPC3 is a proteoglycan localized to the cell membrane via a glycosylphosphatidyl-inositol (GPI) anchor. To further elucidate the GPC3 function(s), we have screened various cell lines for proteins that interact with GPC3, resulting in the isolation of a 115 kDa protein, identified as CD26. The interaction occurred with both the glycosylated and unglycosylated forms of GPC3 and led to the inhibition of CD26 peptidase activity. Moreover, introduction of CD26 into Cos-1 cells was accompanied by the up-regulation of cell growth, while inclusion of recombinant GPC3 in the media reduced the growth of CD26 transfected Cos-1 cells, drastically. Furthermore, HepG2 C3A cells containing CD26 underwent apoptosis in the presence of recombinant GPC3 in both concentration and time-dependant manner. In light of the fact that inhibition of CD26 reduces the rate of cell proliferation, we propose that a number of physical findings observed in SGBS patients may be a consequence of a direct interaction of GPC3 with CD26. Furthermore, GPC3 without the GPI anchor is capable of inducing apoptosis indicating that neither the GPI anchor nor the membrane attachment is required for apoptosis induction.