RESUMEN
CONTEXT: Rongjin Niantong Fang (RJNTF) is a Traditional Chinese Medicine formulation with a good therapeutic effect on osteoarthritis (OA). However, the underlying mechanisms remain unclear. OBJECTIVE: This study investigates whether RJNTF could delay OA cartilage degeneration by regulating the SDF-1/CXCR4-p38MAPK signalling pathway. MATERIALS AND METHODS: The Sprague-Dawley (SD) rats were used to establish the OA model by a modified Hulth's method. SD rats were divided into three groups (n = 10): blank group, model group (0.9% saline, 10 mL/kg/day), and treatment group (RJNTF, 4.5 g/kg/day). After 12 weeks of treatment, each group was analysed by H&E, Safranine-O solid green, ELISA, Immunohistochemistry, and Western blot. An in vitro model was induced with 100 ng/mL SDF-1 by ELISA, the blank group, model group, RJNTF group, and inhibitor group with intervention for 12 h, each group was analysed by Immunofluorescence staining and Western blot. RESULTS: SDF-1 content in the synovium was reduced in RJNTF treatment group compared to non-treatment model group (788.10 vs. 867.32 pg/mL) and down-regulation of CXCR4, MMP-3, MMP-9, MMP-13 protein expression, along with p38 protein phosphorylated were observed in RJNTF treatment group. In vitro results showed that RJNTF (IC50 = 8.925 mg/mL) intervention could down-regulate SDF-1 induced CXCR4 and p38 protein phosphorylated and reduce the synthesis of MMP-3, MMP-9, and MMP-13 proteins of chondrocytes from SD rat cartilage tissues. DISCUSSION AND CONCLUSION: RJNTF alleviates OA cartilage damage by SDF-1/CXCR4-p38MAPK signalling pathway inhibition. Our ongoing research focuses on Whether RJNTF treats OA through alternative pathways.
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Cartílago Articular , Osteoartritis , Ratas , Animales , Metaloproteinasa 3 de la Matriz/metabolismo , Metaloproteinasa 3 de la Matriz/farmacología , Metaloproteinasa 3 de la Matriz/uso terapéutico , Metaloproteinasa 9 de la Matriz/metabolismo , Metaloproteinasa 13 de la Matriz , Ratas Sprague-Dawley , Osteoartritis/tratamiento farmacológico , Receptores CXCR4/metabolismo , Receptores CXCR4/uso terapéuticoRESUMEN
Stromal cell-derived factor-1 alpha (SDF-1α, CXCL12) mediates the migration of circulating cells to desired sites for tissue development, homeostasis, and regeneration and can be used to promote cardiac regeneration by recruiting stem cells. However, the use of SDF-1α in the injured heart necessitates not only higher binding affinity to its receptor, CXCR4+, but also better robustness against enzymatic degradation than other SDF-1 isoforms. Here, we conduct a screening of SDF-1α analog peptides that were designed by structure-based drug design (SBDD), a type of computer-aided drug design (CADD). We have developed in vitro and in vivo methods that enable us to estimate the effect of peptides on the migration of human mesenchymal stem cells (hMSCs) and cardiac regeneration in acute myocardial infarction (AMI)-induced animals, respectively. We demonstrate that one type of SDF-1α analog peptide, SDP-4, among the four analog peptides preselected by SBDD, is more potent than native SDF-1α for cardiac regeneration in myocardial infarction. It is interesting to note that the migratory effects of SDP-4 determined by a wound healing assay, a Transwell assay, and a 2D migration assay are comparable to those of SDF-1α. These results suggest that in vivo, as well as in vitro, screening of peptides developed by SBDD is a quintessential process to the development of a novel therapeutic compound for cardiac regeneration. Our finding also has an implication that the SDP-4 peptide is an excellent candidate for use in the regeneration of an AMI heart.
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Quimiocina CXCL12 , Infarto del Miocardio , Animales , Movimiento Celular , Quimiocina CXCL12/química , Quimiocina CXCL12/farmacología , Quimiocina CXCL12/uso terapéutico , Diseño de Fármacos , Humanos , Infarto del Miocardio/tratamiento farmacológico , Infarto del Miocardio/metabolismo , Péptidos/farmacología , Péptidos/uso terapéutico , Receptores CXCR4/metabolismo , Receptores CXCR4/uso terapéuticoRESUMEN
Colorectal cancer (CRC) remains the third cause of cancer-related mortality in Western countries, metastases are the main cause of death. CRC treatment remains limited by systemic toxicity and chemotherapy resistance. Therefore, nanoparticle-mediated delivery of cytotoxic agents selectively to cancer cells represents an efficient strategy to increase the therapeutic index and overcome drug resistance. We have developed the T22-PE24-H6 therapeutic protein-only nanoparticle that incorporates the exotoxin A from Pseudomonas aeruginosa to selectively target CRC cells because of its multivalent ligand display that triggers a high selectivity interaction with the CXCR4 receptor overexpressed on the surface of CRC stem cells. We here observed a CXCR4-dependent cytotoxic effect for T22-PE24-H6, which was not mediated by apoptosis, but instead capable of inducing a time-dependent and sequential activation of pyroptotic markers in CRC cells in vitro. Next, we demonstrated that repeated doses of T22-PE24-H6 inhibit tumor growth in a subcutaneous CXCR4+ CRC model, also through pyroptotic activation. Most importantly, this nanoparticle also blocked the development of lymphatic and hematogenous metastases, in a highly aggressive CXCR4+ SW1417 orthotopic CRC model, in the absence of systemic toxicity. This targeted drug delivery approach supports for the first time the clinical relevance of inducing GSDMD-dependent pyroptosis, a cell death mechanism alternative to apoptosis, in CRC models, leading to the selective elimination of CXCR4+ cancer stem cells, which are associated with resistance, metastases and anti-apoptotic upregulation.
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Antineoplásicos , Neoplasias Colorrectales , Proteínas de Unión a Fosfato , Proteínas Citotóxicas Formadoras de Poros , Piroptosis , Receptores CXCR4 , Antineoplásicos/farmacología , Neoplasias Colorrectales/tratamiento farmacológico , Sistemas de Liberación de Medicamentos , Humanos , Metástasis de la Neoplasia/prevención & control , Proteínas de Unión a Fosfato/metabolismo , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Receptores CXCR4/metabolismo , Receptores CXCR4/uso terapéutico , Transducción de SeñalRESUMEN
Warts, Hypogammaglobulinemia, Infections and Myelokathexis (WHIM) is a primary immunodeficiency syndrome. Patients with WHIM syndrome are more susceptible to human papillomavirus (HPV) infections and commonly present to a dermatologist with recalcitrant to treatment warts. Other cardinal features of WHIM syndrome include recurrent sinopulmonary bacterial infections, neutropenia/lymphopenia, low levels of immunoglobulins (IgG, IgA, IgM) and myelokathexis. Research demonstrated that truncating gain-of-function mutations of the C-X-C chemokine receptor type 4 gene (CXCR4) are responsible for this disease. Plerixafor, a specific small molecule antagonist of CXCR4, is currently used for peripheral blood hematopoietic stem cell (HSC) mobilization in stem cell transplant recipients. It has recently shown promise for the treatment of WHIM syndrome in phase I/II clinical trials. In this paper we review the emerging patient clinical data for this medication and highlight the role of CXCR4 in other important skin diseases including keratinocyte carcinomas, psoriasis and cutaneous T-cell lymphoma.
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Agammaglobulinemia , Compuestos Heterocíclicos , Neutropenia , Infecciones por Papillomavirus , Verrugas , Agammaglobulinemia/tratamiento farmacológico , Bencilaminas , Ciclamas , Fantasía , Movilización de Célula Madre Hematopoyética , Compuestos Heterocíclicos/farmacología , Compuestos Heterocíclicos/uso terapéutico , Humanos , Neutropenia/tratamiento farmacológico , Enfermedades de Inmunodeficiencia Primaria , Receptores CXCR4/uso terapéutico , Síndrome , Verrugas/tratamiento farmacológico , Verrugas/patologíaRESUMEN
Temozolomide (TMZ) monotherapy is known to be insufficient for resistant/relapsed glioblastoma (GBM), thus seeking a sensitization agent for TMZ is necessary. It was found that regorafenib may improve the overall survival of relapsed GBM patients. We aimed to discover whether regorafenib can enhance the anti-GBM effects of TMZ, and elucidate underlying mechanism. Our analysis of The Cancer Genome Atlas database revealed that the increased expression of CXCR4 is linked to poor survival of GBM patients. Additionally, TMZ treatment may trigger CXCR4/CXCL12 axis of GBM. We used two GBM cell lines, two primary GBM cells, and animal model to identify underlying mechanism and treatment efficacy of regorafenib combined with TMZ by cytotoxicity, apoptosis, reporter gene and invasion/migration assays, chemokine array, Western blotting, MRI, microarray, and immunohistochemistry. We observed that the chemokine CXCL-12 and its receptor CXCR4 regulate the resistance to TMZ, whereas the inhibition of CXCL-12/CXCR4 signaling sensitizes GBM cells to TMZ. The TMZ-induced CXCL-12/CXCR4 signaling, phosphor-extracellular signal-regulated kinases 1 and 2 (ERK1/2) and nuclear factor kappa light chain enhancer of activated B cells (NF-κB), and NF-κB-related proteins can effectively diminish when combining with regorafenib. Regorafenib significantly enhanced the TMZ-induced extrinsic/intrinsic apoptotic pathways, and facilitated the suppression of invasion and migration potential in GBM. Orthotopic tumor experiments demonstrated tumor size reduction and prolonged survival in combination group even with half-dose of TMZ. Our findings provide promising evidence that regorafenib may sensitize GBM to TMZ treatment through inhibition of the CXCL12/CXCR4/ERK/NF-κB signaling.
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Neoplasias Encefálicas , Glioblastoma , Compuestos de Fenilurea , Piridinas , Animales , Apoptosis , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Línea Celular Tumoral , Quimiocina CXCL12/farmacología , Quimiocina CXCL12/uso terapéutico , Glioblastoma/tratamiento farmacológico , Glioblastoma/genética , Glioblastoma/metabolismo , Humanos , FN-kappa B/metabolismo , Compuestos de Fenilurea/farmacología , Compuestos de Fenilurea/uso terapéutico , Piridinas/farmacología , Piridinas/uso terapéutico , Receptores CXCR4/uso terapéutico , Temozolomida/farmacología , Temozolomida/uso terapéuticoRESUMEN
Rationale: Heart failure with preserved ejection fraction (HFpEF) can arise from hypertension-induced cardiac remodeling. Monocyte/macrophage accumulation and inflammation are crucial elements in the pathogenesis of hypertension-induced cardiac remodeling. The C-X-C chemokine receptor 4 (CXCR4) is a critical regulator of the macrophage-mediated immune response. Nevertheless, the contribution of CXCR4 to macrophage phenotype and function during the progression of HFpEF remains unclear. Herein, we aimed to determine the role of macrophagic CXCR4 in heart failure with preserved ejection fraction (HFpEF). Methods: As a HFpEF model, wild type mice and myeloid-specific CXCR4 deficiency mice were subjected to pressure overload for 30 days to assess the function of macrophagic CXCR4 on cardiac function. Medium from macrophages was used to treat cardiac fibroblasts to study macrophage-to-fibroblast signaling. Results: We found circulatory CXCR4+ immune cells, mainly monocytes, markedly increased in HFpEF patients with hypertension. In the experimental HFpEF mice model, macrophages but not neutrophils represent the main infiltrating inflammatory cells in the heart, abundantly expressing CXCR4. Myeloid-specific CXCR4 deficient impeded macrophage infiltration and inflammatory response in the heart of HFpEF mice, thus ameliorating cardiac fibrosis and improving cardiac diastolic function. Furthermore, transcriptomic profiling data revealed that CXCR4 loss in macrophages exhibited a decreased transcriptional signature associated with the regulation of inflammatory response. Notably, CXCR4 significantly augmented chemokine (CXC) motif ligand (CXCL3) expression, which at least partly contributed to fibrosis by promoting myofibroblast differentiation. Mechanistically, the increased production of pro-inflammatory cytokines in CXCR4 expressed macrophages could be attributed to the suppression of the peroxisome proliferator-activated receptor γ (PPARγ) activity. Conclusions: Collectively, our data supported that the infiltration of CXCR4+ macrophages in the heart exacerbates hypertension-induced diastolic function by promoting pro-inflammatory cytokines production and thus may serve as a potential therapeutic target for hypertension-induced HFpEF.
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Cardiomiopatías , Insuficiencia Cardíaca , Hipertensión , Animales , Citocinas , Fibroblastos/metabolismo , Hipertensión/complicaciones , Macrófagos/metabolismo , Ratones , Receptores CXCR4/genética , Receptores CXCR4/uso terapéutico , Volumen Sistólico/fisiología , Función Ventricular Izquierda , Remodelación Ventricular/fisiologíaRESUMEN
Insufficient T cell infiltration in triple-negative breast cancer (TNBC) has limited its response rate to immune checkpoint blockade (ICB) therapies and motivated the development of immunostimulatory approaches to enhance the ICB therapy. CXCR4 is a chemokine receptor highly upregulated both on cell surface and cytoplasm in tumor tissues. Activating CXCR4 has been associated with increased immunosuppression in the tumor microenvironment. Here, we developed a CXCR4-targeted liposomal formulation (Liposomal-AMD3100) to enhance therapeutic efficacy of AMD3100, a CXCR4 antagonist. Particularly, AMD3100 is not only encapsulated into the liposome but coated on the surface of the formulation to serve as a targeting moiety and a dual blocker capable of inhibiting CXCR4 activation extracellularly and intracellularly. The Liposomal-AMD3100 remodeled both immune and stromal microenvironment more efficiently compared with free AMD3100, indicating better pharmacodynamic profile of AMD3100 achieved by liposomal formulation. The combination of anti-PD-L1 with Liposomal-AMD3100 formulation exhibited an increased antitumor effect and prolonged survival time compared with monotherapies in a murine TNBC model (4T1). This work proves that immune activation via liposomal delivery of CXCR4 inhibitors has a great potential to expand ICB therapies to originally ICB-insensitive cancer types.
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Neoplasias de la Mama Triple Negativas , Animales , Quimiocina CXCL12/farmacología , Quimiocina CXCL12/uso terapéutico , Humanos , Inhibidores de Puntos de Control Inmunológico , Ratones , Receptores CXCR4/uso terapéutico , Transducción de Señal , Linfocitos T , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Microambiente TumoralRESUMEN
Mesenchymal stem cells (MSCs) are thought to have great potential in the therapy of acute liver injury. It is possible that these cells may be regulated by the stromal cell-derived factor-1 (SDF-1)/CXC chemokine receptor-4 (CXCR4) signaling axis, which has been shown to promote stem cells migration in the inflammation-associated diseases. However, the effects of SDF-1/CXCR4 axis on the MSCs-transplantation-based treatment for acute liver injury and the underlying mechanisms are largely unknown. In this study, we sought to determine whether SDF-1/CXCR4 would augment the therapeutic effect of bone marrow mesenchymal stem cells (BMSCs) by promoting their migration, which may result from activating the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway, in a rat acute liver injury model induced by lipopolysaccharide (LPS). We found that BMSCs transplantation markedly attenuated liver injury and improved the survival of LPS-treated rats. Of interest, overexpression of CXCR4 in BMSCs could substantially promote their migration both in vitro and in vivo, and result in even better therapeutic effects. This might be attributed to the activation of PI3K/Akt signaling pathway in BMSCs that is downstream of CXCR4, as demonstrated by the use of the CXCR4 antagonist AMD3100 and PI3K pathway inhibitor LY294002 assays in vitro and in vivo. Together, our results unraveled a novel molecular mechanism for the therapeutic effect of BMSCs for the treatment of acute liver injury, which may shed a new light on the clinical application of BMSCs for acute liver failure.
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Células de la Médula Ósea/metabolismo , Trasplante de Médula Ósea/métodos , Enfermedad Hepática Inducida por Sustancias y Drogas/tratamiento farmacológico , Lipopolisacáridos/efectos adversos , Células Madre Mesenquimatosas/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores CXCR4/uso terapéutico , Animales , Movimiento Celular , Femenino , Ratas , Ratas Sprague-Dawley , Transducción de SeñalRESUMEN
Glioblastoma multiforme (GBM) is the most common primary malignancy in the adult central nervous, and is characterized by high aggressiveness and a high mortality rate. The high mortality rate is largely due to the development of drug resistance. Temozolomide (TMZ) resistance is considered to be one of the major reasons responsible for GBM therapy failure. CXCL12/CXCR4 has been demonstrated to be involved in cell proliferation, migration, invasion, angiogenesis, and radioresistance in GBM. However, its role in TMZ resistance in GBM is unknown. In this study, we aimed to evaluate the role of CXCL12/CXCR4 in mediating the TMZ resistance to GBM cells and explore the underlying mechanisms. We found that the CXCL12/CXCR4 axis enhanced TMZ resistance in GBM cells. Further study showed that CXCL12/CXCR4 conferred TMZ resistance and promoted the migration and invasion of GBM cells by up-regulating FOXM1. This resistance was partially reversed by suppressing CXCL12/CXCR4 and FOXM1 silencing. Our study revealed the vital role of CXCL12/CXCR4 in mediating the resistance of GBM cells to TMZ, and suggested that targeting CXCL12/CXCR4 axis may attenuate the resistance to TMZ in GBM.
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Neoplasias Encefálicas , Glioblastoma , Antineoplásicos Alquilantes/farmacología , Antineoplásicos Alquilantes/uso terapéutico , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/genética , Línea Celular Tumoral , Quimiocina CXCL12/genética , Resistencia a Antineoplásicos/genética , Proteína Forkhead Box M1/genética , Proteína Forkhead Box M1/uso terapéutico , Glioblastoma/tratamiento farmacológico , Glioblastoma/genética , Humanos , Receptores CXCR4/genética , Receptores CXCR4/uso terapéutico , Temozolomida/farmacología , Temozolomida/uso terapéutico , Regulación hacia ArribaRESUMEN
BACKGROUND: Chronic myocardial infarction (CMI), represents a public health and a financial burden. Since stem cell transplant is used to regenerate cardiac tissue after acute myocardial infarction. AIM OF THE STUDY: To determine if autologous CXCR4 stem cells could restore damaged myocardial tissue in patients with CMI lesions. METHODS: 20 NYHA grade III male patients with CMI defined by clinical, biochemical, ECG and echocardiographic parameters were included. Patients were treated with G-CSF for 6 d before isolating their autologous stem cells from PBMCs. Cell phenotyping was done by cytofluorometry using monoclonal antibodies (anti-CXCR4, -CD34, -48, -117, -133, -Ki67, -SDF1 and CXCR4); CXCR4 cell subpopulations isolated by sorting were adjusted to 1 × 108 cells by subpopulation and injected in a circular pattern into the cicatrix previously defined by echocardiography. RESULTS: Patients were followed for 6 and 12 months. Six months after cell implant improvements in left ventricle ejection fraction (from 33-50%), stress rate values (from -3/-9% to -18/-22%), stress tests (from 4-12 METS), and the quantity of left ventricle affected segments (3-9) disappeared according to the G-SPECT images. 12 months evaluations did not show significant differences. Interestingly, 3 months after cell implant the ECG showed normal electrical activity in 9 patients whereas after 6 months it was normal in all the patients. CONCLUSIONS: These results ratify that locally injected autologous CXCR4+ bone marrow-derived stem cells have a physiological and a clinical impact in patients with CMI.
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Células Madre Hematopoyéticas/metabolismo , Infarto del Miocardio/terapia , Receptores CXCR4/uso terapéutico , Anciano , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Epilepsy is a neurological disorder often associated with seizure that affects â¼0.7% of pregnant women. During pregnancy, most epileptic patients are prescribed antiepileptic drugs (AEDs) such as valproic acid (VPA) to control seizure activity. Here, we show that prenatal exposure to VPA in mice increases seizure susceptibility in adult offspring through mislocalization of newborn neurons in the hippocampus. We confirmed that neurons newly generated from neural stem/progenitor cells (NS/PCs) are integrated into the granular cell layer in the adult hippocampus; however, prenatal VPA treatment altered the expression in NS/PCs of genes associated with cell migration, including CXC motif chemokine receptor 4 (Cxcr4), consequently increasing the ectopic localization of newborn neurons in the hilus. We also found that voluntary exercise in a running wheel suppressed this ectopic neurogenesis and countered the enhanced seizure susceptibility caused by prenatal VPA exposure, probably by normalizing the VPA-disrupted expression of multiple genes including Cxcr4 in adult NS/PCs. Replenishing Cxcr4 expression alone in NS/PCs was sufficient to overcome the aberrant migration of newborn neurons and increased seizure susceptibility in VPA-exposed mice. Thus, prenatal exposure to an AED, VPA, has a long-term effect on the behavior of NS/PCs in offspring, but this effect can be counteracted by a simple physical activity. Our findings offer a step to developing strategies for managing detrimental effects in offspring exposed to VPA in utero.
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Anticonvulsivantes/toxicidad , Neurogénesis/efectos de los fármacos , Efectos Tardíos de la Exposición Prenatal , Convulsiones/etiología , Ácido Valproico/toxicidad , Animales , Anticonvulsivantes/administración & dosificación , Anticonvulsivantes/farmacología , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Células Cultivadas , Giro Dentado/efectos de los fármacos , Giro Dentado/embriología , Giro Dentado/patología , Susceptibilidad a Enfermedades , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Edad Gestacional , Hipocampo/embriología , Hipocampo/patología , Hipocampo/fisiopatología , Ratones , Ratones Endogámicos C57BL , Proteínas del Tejido Nervioso/biosíntesis , Proteínas del Tejido Nervioso/genética , Células-Madre Neurales/efectos de los fármacos , Neuronas/patología , Esfuerzo Físico , Embarazo , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Distribución Aleatoria , Ratas , Receptores CXCR4/biosíntesis , Receptores CXCR4/genética , Receptores CXCR4/uso terapéutico , Convulsiones/inducido químicamente , Convulsiones/embriología , Transcriptoma , Ácido Valproico/administración & dosificación , Ácido Valproico/farmacologíaRESUMEN
AIM: Lactococcus lactis is a Gram-positive (endotoxin-free) food-grade bacteria exploited as alternative to Escherichia coli for recombinant protein production. We have explored here for the first time the ability of this platform as producer of complex, self-assembling protein materials. MATERIALS & METHODS: Biophysical properties, cell penetrability and in vivo biodistribution upon systemic administration of tumor-targeted protein nanoparticles produced in L. lactis have been compared with the equivalent material produced in E. coli. RESULTS: Protein nanoparticles have been efficiently produced in L. lactis, showing the desired size, internalization properties and biodistribution. CONCLUSION: In vitro and in vivo data confirm the potential and robustness of the production platform, pointing out L. lactis as a fascinating cell factory for the biofabrication of protein materials intended for therapeutic applications.
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Lactococcus lactis/genética , Nanopartículas/química , Neoplasias/tratamiento farmacológico , Proteínas Recombinantes/genética , Escherichia coli/genética , Humanos , Nanopartículas/administración & dosificación , Transporte de Proteínas/genética , Receptores CXCR4/química , Receptores CXCR4/genética , Receptores CXCR4/uso terapéutico , Proteínas Recombinantes/química , Proteínas Recombinantes/uso terapéutico , Distribución TisularRESUMEN
BACKGROUND: Receptor occupancy (RO) assays provide a means to measure the direct interaction of therapeutics with their cell surface targets. Free receptor assays quantify cell-surface receptors not bound by a therapeutic while total receptor assays quantify the amount of target on the cell surface. METHODS: We developed both a flow cytometry-based free RO assay to detect free surface CXCR4, and a total surface CXCR4 assay. In an effort to evaluate potential displacement interference, we performed in vitro experiments to compare on-cell affinity with the IC50 values from in vitro and in vivo from the free CXCR4 assay. We determined free and total surface CXCR4 on circulating blood cells in cynomolgus monkeys dosed with MEDI3185, a fully human monoclonal antibody to CXCR4. RESULTS: We devised an approach to evaluate displacement interference during assay development and showed that our free assay demonstrated little to no displacement interference. After dosing cynomolgus monkeys with MEDI3185, we observed dose-dependence in the magnitude and duration of receptor occupancy and found CXCR4 to increase on lymphocytes, monocytes, and granulocytes. In a multiple dose study, we observed time points where surface CXCR4 appeared fully occupied but MEDI3185 was not detectable in serum. These paradoxical results represented a type of assay interference, and by comparing pharmacokinetic, ADA and total CXCR4 results, the most likely reason for the free CXCR4 results was the emergence of neutralizing anti-drug antibodies (ADA). The total CXCR4 assay was unaffected by ADA and provided a reliable marker of target modulation in both in vivo studies.
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Anticuerpos Antiidiotipos/administración & dosificación , Anticuerpos Monoclonales/administración & dosificación , Citometría de Flujo , Receptores CXCR4/uso terapéutico , Animales , Anticuerpos Antiidiotipos/inmunología , Anticuerpos Monoclonales/inmunología , Granulocitos/efectos de los fármacos , Granulocitos/inmunología , Humanos , Inmunoglobulinas Intravenosas/administración & dosificación , Linfocitos/efectos de los fármacos , Linfocitos/inmunología , Macaca fascicularis/inmunología , Monocitos/efectos de los fármacos , Monocitos/inmunología , Receptores CXCR4/inmunologíaRESUMEN
Resting human CD4 T cells are highly resistant to transfection or infection with lentiviral vectors derived from the human immunodeficiency virus. We now describe a flexible and efficient approach involving virus-like particles containing simian immunodeficiency virus lentiviral gene product protein X and pseudotyping with CXCR4-tropic HIV Env. This method permits effective genetic manipulation of these cells while preserving their naturally quiescent state. This technology can also be extended to primary lymphoid cultures where authentic cellular composition and functional relationships are preserved.
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Linfocitos T CD4-Positivos/metabolismo , Técnicas de Transferencia de Gen , Terapia Genética , Proteínas Virales/genética , Animales , Linfocitos T CD4-Positivos/patología , Vectores Genéticos , VIH-1/genética , Humanos , Lentivirus/genética , Receptores CXCR4/genética , Receptores CXCR4/uso terapéutico , Virus de la Inmunodeficiencia de los Simios , Proteínas Virales/administración & dosificación , Proteínas Virales/uso terapéutico , Productos del Gen env del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen env del Virus de la Inmunodeficiencia Humana/uso terapéuticoRESUMEN
The therapeutic potential of bone marrow mesenchymal stem cells (MSCs) in kidney failure has been examined in some studies. However, recent findings indicate that after transplantation, these cells home to kidneys at very low levels. Interaction of stromal derived factor-1 (SDF-1) with its receptor, CXCR4, is of pivotal importance in migration and homing. Recently, CXCR7 has also been recognized as another SDF-1 receptor that interacts with CXCR4 and modulates its functions. In this study, CXCR4 and CXCR7 were separately and simultaneously overexpressed in BALB/c bone marrow MSCs by using a lentiviral vector system and the homing and renoprotective potentials of these cells were evaluated in a mouse model of cisplatin-induced acute kidney injury. Using flow cytometry, immunohistochemistry, and real-time PCR methods for detection of GFP-labeled MSCs, we found that although considerably entrapped in lungs, native MSCs home very rarely to kidneys and bone marrow and this rate cannot be significantly affected by CXCR4 and/or CXCR7 upregulation. Transplantation of neither native nor genetically engineered MSCs ameliorated kidney failure. We concluded that overexpression of CXCR4 and CXCR7 receptors in murine MSCs cannot improve the homing and therapeutic potentials of these cells and it can be due to severe chromosomal abnormalities that these cells bear during ex vivo expansion.
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Lesión Renal Aguda/terapia , Movimiento Celular , Terapia Genética , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/metabolismo , Receptores CXCR4/genética , Receptores CXCR/genética , Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/patología , Animales , Apoptosis , Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Separación Celular , Cisplatino , Modelos Animales de Enfermedad , Citometría de Flujo , Proteínas Fluorescentes Verdes/metabolismo , Inmunohistoquímica , Lentivirus/genética , Masculino , Ratones , Ratones Endogámicos BALB C , Receptores CXCR/uso terapéutico , Receptores CXCR4/uso terapéutico , Reproducibilidad de los Resultados , Análisis de Supervivencia , Transducción Genética , Regulación hacia Arriba/genéticaRESUMEN
The interaction between the chemokine stromal cell-derived factor 1 and its receptor CXCR4 plays an important role in GH production and cell proliferation in normal and tumorous pituitary somatotrope cells. Therefore, the chemokine receptor CXCR4 could be an attractive target for antitumor drugs in patients with acromegaly. A synthetic antagonist of CXCR4, cyclic pentapeptide d-Arg3FC131 (c[Gly1-d-Tyr2-d-Arg3-Arg4-Nal5]) significantly inhibited GH production and proliferation of GH3 somatotrope tumor cells in vitro. It also induced apoptosis of GH3 cells through activation of the caspase-3 pathway. Systemic administration of d-Arg3FC131 inhibited the growth of GH3 cell xenografts in immunodeficient nude mice by inducing apoptosis and suppressing the proliferation of tumor cells. These results indicate that d-Arg3FC131 might have potential for the treatment of pituitary tumors producing excess GH in patients with acromegaly.
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Apoptosis/efectos de los fármacos , Oligopéptidos/farmacología , Péptidos Cíclicos/farmacología , Neoplasias Hipofisarias/tratamiento farmacológico , Receptores CXCR4/antagonistas & inhibidores , Somatotrofos/efectos de los fármacos , Somatotrofos/patología , Adenoviridae , Animales , Western Blotting , Línea Celular , Citometría de Flujo , Vectores Genéticos/genética , Hormona del Crecimiento/metabolismo , Etiquetado Corte-Fin in Situ , Masculino , Ratones , Ratones Desnudos , Péptidos Cíclicos/uso terapéutico , Neoplasias Hipofisarias/metabolismo , Ratas , Receptores CXCR4/uso terapéutico , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Entry of HIV-1 virions into cells is a complex and dynamic process carried out by envelope (Env) glycoproteins on the surface of the virion that promote the thermodynamically unfavorable fusion of highly stable viral and target cell membranes. Insight gained from studies of the mechanism of viral entry allowed insight into the design of novel inhibitors of HIV-1 entry, several of which are now in clinical trials. This review highlights the mechanism by which viral and cellular proteins mediate entry of HIV-1 into permissive cells, with an emphasis on targeting this process in the design of novel therapies that target distinct steps of the entry process, including antagonizing receptor binding events and blocking conformational changes intimately involved in membrane fusion.