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Dinosaur tracks have a long history of discovery and study in the UK, but track sites for sauropodomorph dinosaurs-the group that included the giant, graviportal herbivorous sauropods-are comparatively rare. Here, we provide a description of a sauropod dinosaur track site at Spyway Quarry in Dorset, southern England. The tracks at Spyway were discovered in the late 1990s and occur in the Stair Hole Member of the Durlston Formation in the Purbeck Limestone Group, of earliest Cretaceous age. More than 130 individual tracks of large sauropod dinosaurs are present at the site, but they are generally poorly preserved and do not form clear trackways, although it is likely that they represent multiple individuals. They provide further evidence for sauropods living in or passing through coastal lagoonal environments. Although poorly preserved, Spyway represents the largest in situ dinosaur track site currently accessible within the Purbeck Group, with considerable potential for further discoveries through ongoing quarrying in the surrounding area.
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Introduction: The ability to modulate and enhance the anti-tumor immune responses is critical in developing novel therapies in cancer. The Tumor Necrosis Factor (TNF) Receptor Super Family (TNFRSF) are potentially excellent targets for modulation which result in specific anti-tumor immune responses. CD40 is a member of the TNFRSF and several clinical therapies are under development. CD40 signaling plays a pivotal role in regulating the immune system from B cell responses to myeloid cell driven activation of T cells. The CD40 signaling axis is well characterized and here we compare next generation HERA-Ligands to conventional monoclonal antibody based immune modulation for the treatment of cancer. Methods & results: HERA-CD40L is a novel molecule that targets CD40 mediated signal transduction and demonstrates a clear mode of action in generating an activated receptor complex via recruitment of TRAFs, cIAP1, and HOIP, leading to TRAF2 phosphorylation and ultimately resulting in the enhanced activation of key inflammatory/survival pathway and transcription factors such asNFkB, AKT, p38, ERK1/2, JNK, and STAT1 in dendritic cells. Furthermore, HERA-CD40L demonstrated a strong modulation of the tumor microenvironment (TME) via the increase in intratumoral CD8+ T cells and the functional switch from pro-tumor macrophages (TAMs) to anti-tumor macrophages that together results in a significant reduction of tumor growth in a CT26 mouse model. Furthermore, radiotherapy which may have an immunosuppressive modulation of the TME, was shown to have an immunostimulatory effect in combination with HERA-CD40L. Radiotherapy in combination with HERA-CD40L treatment resulted in an increase in detected intratumoral CD4+/8+ T cells compared to RT alone and, additionally, the repolarization of TAMs was also observed, resulting in an inhibition of tumor growth in a TRAMP-C1 mouse model. Discussion: Taken together, HERA-CD40L resulted in activating signal transduction mechanisms in dendritic cells, resulting in an increase in intratumoral T cells and manipulation of the TME to be pro-inflammatory, repolarizing M2 macrophages to M1, enhancing tumor control.
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Ligando de CD40 , Neoplasias , Animales , Ratones , Antígenos CD40 , Células Presentadoras de Antígenos , Macrófagos , Neoplasias/radioterapia , Modelos Animales de Enfermedad , Microambiente TumoralRESUMEN
Digital interventions can increase physical activity (PA) levels in adults. However, the COVID-19 pandemic highlighted the complexities faced when guiding people to start or return to PA following illness or inactivity. A digital tool, Movement Foundations, was developed to provide remote guidance on building strength and capacity across functional movement patterns, with graduated progression based on user responses and input. This qualitative study aimed to explore the perceived impacts of using the tool. Nine participants aged over 35 years from the healthcare and academic healthcare sectors were recruited to use it and were subsequently interviewed. Thematic analysis identified three themes falling under the overarching concept of 'Capability, Opportunity and Motivation-Behaviour (COM-B) Plus', encompassing: skills and capacity for movement; opportunities, motivations and barriers for movement; and a personalised, safe space in which to develop. Participants felt that the digital tool increased their capacity and confidence in movement and positively impacted their daily activities. External factors such as illness and stress clouded perceptions of the impacts of PA. Time, work pressures and needing equipment were still considered significant barriers to PA. Still, participants appreciated the flexibility and non-prescriptive nature of the tool and felt that it helped movement to become opportunistic and habitual. Increased capacity for PA and feeling the subsequent physical and mental effects positively influenced motivation. Structure and guidance, with graduated progress, were seen as protective. Guided self-reflection helped participants understand their capacity and limitations with regard to movement and promoted motivation. Although acquiring technical skills to guide movement may be important for those recovering from illness, participants found that a structure promoting individualised guidance, graduated progression and guided self-reflection were important motivational factors for continuing use. Digital interventions should consider these aspects when seeking to promote habitual PA.
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Cell signaling is initiated by characteristic protein patterns in the plasma membrane, but tools to decipher their molecular organization and activation are hitherto lacking. Among the well-known signaling pattern is the death inducing signaling complex with a predicted hexagonal receptor architecture. To probe this architecture, DNA origami-based nanoagents with nanometer precise arrangements of the death receptor ligand FasL are introduced and presented to cells. Mimicking different receptor geometries, these nanoagents act as signaling platforms inducing fastest time-to-death kinetics for hexagonal FasL arrangements with 10 nm inter-molecular spacing. Compared to naturally occurring soluble FasL, this trigger is faster and 100× more efficient. Nanoagents with different spacing, lower FasL number or higher coupling flexibility impede signaling. The results present DNA origami as versatile signaling scaffolds exhibiting unprecedented control over molecular number and geometry. They define molecular benchmarks in apoptosis signal initiation and constitute a new strategy to drive particular cell responses.
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Apoptosis , Receptor fas , Proteínas Portadoras/metabolismo , ADN , Transducción de Señal , Receptor fas/metabolismoRESUMEN
TNF Receptor Superfamily (TNF-R-SF) signaling is a structurally well-defined event that requires proper receptor clustering and trimerization. While the TNF-SF ligands naturally exist as trivalent functional units, the receptors are usually separated on the cell surface. Critically, receptor assembly into functional trimeric signaling complexes occurs through binding of the natural ligand unit. TNF-R-SF members, including CD40, have been key immunotherapeutic targets for over 20 years. CD40, expressed by antigen-presenting cells, endothelial cells, and many tumors, plays a fundamental role in connecting innate and adaptive immunity. The multiple investigated strategies to induce CD40 signaling can be broadly grouped into antibody-based or CD40L-based approaches. Currently, seven different antibodies and one CD40L-based hexavalent fusion protein are in active clinical trials. In this review, we describe the biology and structural properties of CD40, requirements for agonistic signal transduction through CD40 and summarize current attempts to exploit the CD40 signaling pathway for the treatment of cancer.
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Células Endoteliales , Neoplasias , Antígenos CD40 , Ligando de CD40 , Humanos , Neoplasias/terapia , Receptores del Factor de Necrosis Tumoral , Transducción de SeñalRESUMEN
BACKGROUND: Glucocorticoid-induced TNFR-related protein (TNFRSF18, GITR, CD357), expressed by T cells, and its ligand (TNFSF18, GITRL), expressed by myeloid populations, provide co-stimulatory signals that boost T cell activity. Due to the important role that GITR plays in regulating immune functions, agonistic stimulation of GITR is a promising therapeutic concept. Multiple strategies to induce GITR signaling have been investigated. The limited clinical efficacy of antibody-based GITR agonists results from structural and functional characteristics of antibodies that are unsuitable for stimulating the well-defined trimeric members of the TNFRSF. METHODS: To overcome limitations of antibody-based TNFRSF agonists, we have developed HERA-GITRL, a fully human hexavalent TNF receptor agonist (HERA) targeting GITR and mimicking the natural signaling concept. HERA-GITRL is composed of a trivalent but single-chain GITRL-receptor-binding-domain (scGITRL-RBD) unit fused to an IgG1 derived silenced Fc-domain serving as dimerization scaffold. A specific mouse surrogate, mmHERA-GITRL, was also generated to examine in vivo activity in respective mouse tumor models. RESULTS: For functional characterization of HERA-GITRL in vitro, human immune cells were isolated from healthy-donor blood and stimulated with anti-CD3 antibody in the presence of HERA-GITRL. Consistently, HERA-GITRL increased the activity of T cells, including proliferation and differentiation, even in the presence of regulatory T cells. In line with these findings, mmHERA-GITRL enhanced antigen-specific clonal expansion of both CD4+ (OT-II) and CD8+ (OT-I) T cells in vivo while having no effect on non-specific T cells. In addition, mmHERA-GITRL showed single-agent anti-tumor activity in two subcutaneous syngeneic colon cancer models (CT26wt and MC38-CEA). Importantly, this activity is independent of its FcγR-binding functionality, as both mmHERA-GITRL with a functional Fc- and a silenced Fc-domain showed similar tumor growth inhibition. Finally, in a direct in vitro comparison to a bivalent clinical benchmark anti-GITR antibody and a trivalent GITRL, only the hexavalent HERA-GITRL showed full biological activity independent of additional crosslinking. CONCLUSION: In this manuscript, we describe the development of HERA-GITRL, a true GITR agonist with a clearly defined mechanism of action. By clustering six receptor chains in a spatially well-defined manner, HERA-GITRL induces potent agonistic activity without being dependent on additional FcγR-mediated crosslinking.
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Receptores del Factor de Necrosis Tumoral/agonistas , Anticuerpos de Cadena Única/administración & dosificación , Linfocitos T Reguladores/inmunología , Factores de Necrosis Tumoral/química , Animales , Línea Celular Tumoral , Humanos , Fragmentos Fc de Inmunoglobulinas/inmunología , Activación de Linfocitos , Macaca fascicularis , Ratones , Proteínas Recombinantes de Fusión/inmunología , Transducción de Señal , Anticuerpos de Cadena Única/inmunología , Factores de Necrosis Tumoral/metabolismoRESUMEN
CD40 ligand (TNFSF5/CD154/CD40L), a member of the tumor necrosis factor (TNF) superfamily is a key regulator of the immune system. The cognate receptor CD40 (TNFRSF5) is expressed broadly on antigen-presenting cells and many tumor types, and has emerged as an attractive target for immunologic cancer treatment. Most of the CD40 targeting drugs in clinical development are antibodies which display some disadvantages: their activity typically depends on Fcγ receptor-mediated crosslinking, and depletion of CD40-expressing immune cells by antibody-dependent cellular cytotoxicity compromises an efficient antitumor response. To overcome the inadequacies of antibodies, we have developed the hexavalent receptor agonist (HERA) Technology. HERA compounds are fusion proteins composed of 3 receptor binding domains in a single chain arrangement, linked to an Fc-silenced human IgG1 thereby generating a hexavalent molecule. HERA-CD40L provides efficient receptor agonism on CD40-expressing cells and, importantly, does not require FcγR-mediated crosslinking. Strong activation of NFκB signaling was observed upon treatment of B cells with HERA-CD40L. Monocyte treatment with HERA-CD40L promoted differentiation towards the M1 spectrum and repolarization of M2 spectrum macrophages towards the M1 spectrum phenotype. Treatment of in vitro co-cultures of T and B cells with HERA-CD40L-triggered robust antitumor activation of T cells, which depended upon direct interaction with B cells. In contrast, bivalent anti-CD40 antibodies and trivalent soluble CD40L displayed weak activity which critically depended on crosslinking. In vivo, a murine surrogate of HERA-CD40L-stimulated clonal expansion of OT-I-specific murine CD8 T cells and showed single agent antitumor activity in the CD40 syngeneic MC38-CEA mouse model of colorectal cancer, suggesting an involvement of the immune system in controlling tumor growth. We conclude that HERA-CD40L is able to establish robust antitumor immune responses both in vitro and in vivo.
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Antineoplásicos Inmunológicos/farmacología , Antígenos CD40/antagonistas & inhibidores , Ligando de CD40/farmacología , Inmunoglobulina G/farmacología , Proteínas Recombinantes de Fusión/farmacología , Animales , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Femenino , Humanos , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/inmunología , Macrófagos/citología , Macrófagos/efectos de los fármacos , Ratones Endogámicos C57BL , FN-kappa B/inmunologíaRESUMEN
Tumor necrosis factor receptor superfamily member 7 (TNFRSF7, CD27), expressed primarily by T cells, and its ligand CD27L (TNFSF7, CD70) provide co-stimulatory signals that boost T cell activation, differentiation, and survival. Agonistic stimulation of CD27 is therefore a promising therapeutic concept in immuno-oncology intended to boost and sustain T cell driven anti-tumor responses. Endogenous TNFSF/TNFRSF-based signal transmission is a structurally well-defined event that takes place during cell-to-cell-based contacts. It is well-established that the trimeric-trivalent TNFSF-receptor binding domain (TNFSF-RBD) exposed by the conducting cell and the resulting multi-trimer-based receptor clustering on the receiving cell are essential for agonistic signaling. Therefore, we have developed HERA-CD27L, a novel hexavalent TNF receptor agonist (HERA) targeting CD27 and mimicking the natural signaling concept. HERA-CD27L is composed of a trivalent but single-chain CD27L-receptor-binding-domain (scCD27L-RBD) fused to an IgG1 derived silenced Fc-domain serving as dimerization scaffold. The hexavalent agonist significantly boosted antigen-specific T cell responses while having no effect on non-specific T cells and was superior over stabilized recombinant trivalent CD27L. In addition, HERA-CD27L demonstrated potent single-agent anti-tumor efficacy in two different syngeneic tumor models, MC38-CEA and CT26wt. Furthermore, the combination of HERA-CD27L and an anti-PD-1 antibody showed additive anti-tumor effects highlighting the importance of both T cell activation and checkpoint inhibition in anti-tumor immunity. In this manuscript, we describe the development of HERA-CD27L, a true CD27 agonist with a clearly defined forward-signaling mechanism of action.
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Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has raised attention as a novel anticancer therapeutic as it induces apoptosis preferentially in tumor cells. However, first-generation TRAIL-receptor agonists (TRAs), comprising recombinant TRAIL and agonistic receptor-specific antibodies, have not demonstrated anticancer activity in clinical studies. In fact, cancer cells are often resistant to conventional TRAs. Therefore, in addition to TRAIL-sensitizing strategies, next-generation TRAs with superior apoptotic activity are warranted. APG350 is a novel, highly potent TRAIL-receptor agonist with a hexavalent binding mode allowing the clustering of six TRAIL-receptors per drug molecule. Here we report on preclinical in vitro and in vivo studies testing the activity of APG350 on pancreatic ductal adenocarcinoma (PDAC) cells. We found that APG350 potently induced apoptosis of Colo357, PancTuI and Panc89 cells in vitro. In addition, APG350 treatment activated non-canonical TRAIL signaling pathways (MAPK, p38, JNK, ERK1/ERK2 and NF-κB) and induced the secretion of IL-8. Stable overexpression of Bcl-xL inhibited APG350-induced cell death and augmented activation of non-canonical pathways. Intriguingly, pre-treatment of Bcl-xL-overexpressing cells with the BH3-mimic Navitoclax restored their sensitivity to APG350. To study the effects of APG350 on PDAC cells in vivo, we applied two different orthotopic xenotransplantation mouse models, with and without primary tumor resection, representing adjuvant and palliative treatment regimes, respectively. APG350 treatment of established tumors (palliative treatment) significantly reduced tumor burden. These effects, however, were not seen in tumors with enforced overexpression of Bcl-xL. Upon primary tumor resection and subsequent APG350 treatment (adjuvant therapy), APG350 limited recurrent tumor growth and metastases. Importantly, therapeutic efficacy of APG350 treatment was more effective compared with treatment with soluble TRAIL in both models. In conclusion, APG350 represents a promising next-generation TRA for the treatment of PDAC. Moreover, our results suggest that combining APG350 with Navitoclax might be a succesfull strategy for cancers harboring mitochondrial apoptosis resistance.
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Carcinoma Ductal Pancreático , Proteínas de Neoplasias , Neoplasias Pancreáticas , Proteínas Recombinantes de Fusión/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF , Animales , Carcinoma Ductal Pancreático/tratamiento farmacológico , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patología , Línea Celular Tumoral , Femenino , Humanos , Ratones , Ratones SCID , Proteínas de Neoplasias/agonistas , Proteínas de Neoplasias/metabolismo , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Ligando Inductor de Apoptosis Relacionado con TNF/agonistas , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
In recent years, new treatment options for malignant melanoma patients have enhanced the overall survival for selected patients. Despite new hope, most melanoma patients still relapse with drug-resistant tumors or experience intrinsic resistance to the therapy. Therefore, novel treatment modalities beneficial for subgroups of patients are needed. TRAIL receptor agonists have been suggested as promising candidates for use in cancer treatment as they preferentially induce apoptosis in cancer cells. Unfortunately, the first generation of TRAIL receptor agonists showed poor clinical efficacy. hvTRA is a second-generation TRAIL receptor agonist with improved composition giving increased potency, and in the present study, we showed hvTRA-induced activation of apoptosis leading to an efficient and sustained reduction in melanoma cell growth in cell lines and xenograft models. Furthermore, the potential of hvTRA in a clinical setting was demonstrated by showing efficacy on tumor cells harvested from melanoma patients with lymph node metastasis in an ex vivo drug sensitivity assay. Inhibition of mutated BRAF has been shown to regulate proteins in the intrinsic apoptotic pathway, making the cells more susceptible for apoptosis induction. In an attempt to increase the efficacy of hvTRA, combination treatment with the mutated BRAF inhibitor vemurafenib was investigated. A synergistic effect by the combination was observed for several cell lines in vitro, and an initial cytotoxic effect was observed in vivo. Unfortunately, the initial increased reduction in tumor growth compared with hvTRA mono treatment was not sustained, and this was related to downregulation of the DR5 level by vemurafenib. Altogether, the presented data imply that hvTRA efficiently induce apoptosis and growth delay in melanoma models and patient material, and the potential of this TRAIL receptor agonist should be further evaluated for treatment of subgroups of melanoma patients.
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Amyotrophic lateral sclerosis (ALS) is a devastating progressive neurodegenerative disease affecting primarily the upper and lower motor neurons. A common feature of all ALS cases is a well-characterized neuroinflammatory reaction within the central nervous system (CNS). However, much less is known about the role of the peripheral immune system and its interplay with CNS resident immune cells in motor neuron degeneration. Here, we characterized peripheral monocytes in both temporal and spatial dimensions of ALS pathogenesis. We found the circulating monocytes to be deregulated in ALS regarding subtype constitution, function and gene expression. Moreover, we show that CNS infiltration of peripheral monocytes correlates with improved motor neuron survival in a genetic ALS mouse model. Furthermore, application of human immunoglobulins or fusion proteins containing only the human Fc, but not the Fab antibody fragment, increased CNS invasion of peripheral monocytes and delayed the disease onset. Our results underline the importance of peripheral monocytes in ALS pathogenesis and are in agreement with a protective role of monocytes in the early phase of the disease. The possibility to boost this beneficial function of peripheral monocytes by application of human immunoglobulins should be evaluated in clinical trials.
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Esclerosis Amiotrófica Lateral/patología , Sistema Nervioso Central/metabolismo , Leucocitos Mononucleares/metabolismo , Monocitos/metabolismo , Sistema Mononuclear Fagocítico/metabolismo , Neuronas Motoras/patología , Médula Espinal/patología , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Modelos Animales de Enfermedad , Humanos , Ratones Endogámicos C57BL , Médula Espinal/metabolismoRESUMEN
Glioblastoma is a disease characterized by rapid invasive tumour growth. Studies on the proapoptotic CD95/CD95L signalling pathway recently suggested a significant contribution of CD95 signalling towards the high degree of motility in glioma cells. Apogenix has developed APG101, a clinical phase II compound designed to bind and neutralize CD95L, and thus to interfere with CD95/CD95L-based signalling. APG101 has shown clinical efficacy in a controlled randomized phase II trial in patients with recurrent glioma. Because APG101 is not cytotoxic to tumour cells in vitro, we postulated that the anti-invasive function of APG101 is the main mechanism of action for this compound. Using three-dimensional spheroid invasion assays in vitro and in murine brain tissue cultures, we found that knockdown of endogenous CD95L reduced the invasive phenotype in our two glioblastoma model cell lines U87-MG and U251-MG. Invasion was restored in CD95L knockdown cells upon the addition of soluble recombinant CD95L and this effect was inhibited by APG101. We conclude that CD95L from autocrine and paracrine sources contributes towards the invasive phenotype of glioblastoma cells and that APG101 acts as a suppressor of proinvasive signalling by the CD95/CD95L pathway in glioblastoma.
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Neoplasias Encefálicas/patología , Proteína Ligando Fas/metabolismo , Glioblastoma/patología , Inmunoglobulina G/farmacología , Proteínas Recombinantes de Fusión/farmacología , Esferoides Celulares/efectos de los fármacos , Receptor fas/metabolismo , Animales , Apoptosis/efectos de los fármacos , Neoplasias Encefálicas/metabolismo , Línea Celular Tumoral , Movimiento Celular , Resistencia a Antineoplásicos , Técnicas de Silenciamiento del Gen , Glioblastoma/metabolismo , Humanos , Ratones Endogámicos C57BL , Invasividad Neoplásica , Transducción de Señal , Esferoides Celulares/patología , Esferoides Celulares/fisiología , Receptor fas/genética , Receptor fas/farmacologíaRESUMEN
Neuroinflammation is increasingly recognized as a hallmark of neurodegeneration. Activated central nervous system-resident microglia and infiltrating immune cells contribute to the degeneration of dopaminergic neurons (DNs). However, how the inflammatory process leads to neuron loss and whether blocking this response would be beneficial to disease progression remains largely unknown. CD95 is a mediator of inflammation that has also been proposed as an apoptosis inducer in DNs, but previous studies using ubiquitous deletion of CD95 or CD95L in mouse models of neurodegeneration have generated conflicting results. Here we examine the role of CD95 in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridin (MPTP)-induced neurodegeneration using tissue-specific deletion of CD95 or CD95L. We show that DN death is not mediated by CD95-induced apoptosis because deletion of CD95 in DNs does not influence MPTP-induced neurodegeneration. In contrast, deletion of CD95L in peripheral myeloid cells significantly protects against MPTP neurotoxicity and preserves striatal dopamine levels. Systemic pharmacological inhibition of CD95L dampens the peripheral innate response, reduces the accumulation of infiltrating myeloid cells, and efficiently prevents MPTP-induced DN death. Altogether, this study emphasizes the role of the peripheral innate immune response in neurodegeneration and identifies CD95 as potential pharmacological target for neurodegenerative disease.
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Apoptosis/inmunología , Neuronas Dopaminérgicas/inmunología , Proteína Ligando Fas/inmunología , Inmunidad Innata , Células Mieloides/inmunología , Trastornos Parkinsonianos/inmunología , Animales , Apoptosis/genética , Cuerpo Estriado/inmunología , Cuerpo Estriado/patología , Dopamina/genética , Dopamina/inmunología , Neuronas Dopaminérgicas/patología , Proteína Ligando Fas/antagonistas & inhibidores , Proteína Ligando Fas/genética , Inflamación , Ratones , Ratones Noqueados , Células Mieloides/patología , Trastornos Parkinsonianos/genética , Trastornos Parkinsonianos/patología , Receptor fas/inmunologíaRESUMEN
Cancer cells can be specifically driven into apoptosis by activating Death-receptor-4 (DR4; TRAIL-R1) and/or Death-receptor-5 (DR5; TRAIL-R2). Albeit showing promising preclinical efficacy, first-generation protein therapeutics addressing this pathway, especially agonistic anti-DR4/DR5-monoclonal antibodies, have not been clinically successful to date. Due to their bivalent binding mode, effective apoptosis induction by agonistic TRAIL-R antibodies is achieved only upon additional events leading to antibody-multimer formation. The binding of these multimers to their target subsequently leads to effective receptor-clustering on cancer cells. The research results presented here report on a new class of TRAIL-receptor agonists overcoming this intrinsic limitation observed for antibodies in general. The main feature of these agonists is a TRAIL-mimic consisting of three TRAIL-protomer subsequences combined in one polypeptide chain, termed the single-chain TRAIL-receptor-binding domain (scTRAIL-RBD). In the active compounds, two scTRAIL-RBDs with three receptor binding sites each are brought molecularly in close proximity resulting in a fusion protein with a hexavalent binding mode. In the case of APG350-the prototype of this engineering concept-this is achieved by fusing the Fc-part of a human immunoglobulin G1 (IgG1)-mutein C-terminally to the scTRAIL-RBD polypeptide, thereby creating six receptor binding sites per drug molecule. In vitro, APG350 is a potent inducer of apoptosis on human tumor cell lines and primary tumor cells. In vivo, treatment of mice bearing Colo205-xenograft tumors with APG350 showed a dose-dependent antitumor efficacy. By dedicated muteins, we confirmed that the observed in vivo efficacy of the hexavalent scTRAIL-RBD fusion proteins is-in contrast to agonistic antibodies-independent of FcγR-based cross-linking events.
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Antineoplásicos/farmacología , Fragmentos de Péptidos/farmacología , Receptores de IgG/metabolismo , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Proteínas Recombinantes de Fusión/farmacología , Animales , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales Humanizados , Antineoplásicos/administración & dosificación , Antineoplásicos/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Ratones , Modelos Biológicos , Fragmentos de Péptidos/administración & dosificación , Fragmentos de Péptidos/química , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/agonistas , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/antagonistas & inhibidores , Proteínas Recombinantes de Fusión/administración & dosificación , Proteínas Recombinantes de Fusión/química , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The participants were 256 African-American students between the ages of 18 and 25, from two historically Black universities. The purpose of this study was to see how dimensions of religiosity and spirituality influenced the HIV risk behavior in African-American college students. Each participant completed the Expressions of Spirituality Inventory (ESI) and a survey of sexual attitudes, beliefs, and behaviors. The data were analyzed using a series of ANOVAs, t tests, and correlations. The results from the study confirmed that there was a relationship between religiosity/spirituality and one's tendency to engage in HIV risk behaviors in the population of African-American college students. Interestingly, this study was able to reveal that traditional indicators of religiosity, such as association and church attendance, were not predictors of any of the risky sexual behaviors or attitudes. The portions of religiosity with the greatest impact on these behaviors were the Experiential/Phenomenological, the Existential Well-being, and the Cognitive dimensions, with high scores on each indicative of less likelihood of engaging in risky sexual behaviors.
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Negro o Afroamericano/psicología , Infecciones por VIH/etnología , Infecciones por VIH/transmisión , Religión y Medicina , Estudiantes/psicología , Sexo Inseguro/etnología , Sexo Inseguro/psicología , Sexo Inseguro/estadística & datos numéricos , Adolescente , Adulto , Femenino , Infecciones por VIH/prevención & control , Infecciones por VIH/psicología , Conocimientos, Actitudes y Práctica en Salud , Encuestas Epidemiológicas , Humanos , Masculino , Inventario de Personalidad/estadística & datos numéricos , Psicometría , Espiritualidad , Estadística como Asunto , Trastornos Relacionados con Sustancias/etnología , Trastornos Relacionados con Sustancias/psicología , Adulto JovenRESUMEN
Injury to the central nervous system initiates an uncontrolled inflammatory response that results in both tissue repair and destruction. Here, we showed that, in rodents and humans, injury to the spinal cord triggered surface expression of CD95 ligand (CD95L, FasL) on peripheral blood myeloid cells. CD95L stimulation of CD95 on these cells activated phosphoinositide 3-kinase (PI3K) and metalloproteinase-9 (MMP-9) via recruitment and activation of Syk kinase, ultimately leading to increased migration. Exclusive CD95L deletion in myeloid cells greatly decreased the number of neutrophils and macrophages infiltrating the injured spinal cord or the inflamed peritoneum after thioglycollate injection. Importantly, deletion of myeloid CD95L, but not of CD95 on neural cells, led to functional recovery of spinal injured animals. Our results indicate that CD95L acts on peripheral myeloid cells to induce tissue damage. Thus, neutralization of CD95L should be considered as a means to create a controlled beneficial inflammatory response.
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Movimiento Celular , Proteína Ligando Fas/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Células Mieloides/metabolismo , Peritonitis/inmunología , Proteínas Tirosina Quinasas/metabolismo , Animales , Células Cultivadas , Proteína Ligando Fas/genética , Proteína Ligando Fas/inmunología , Humanos , Inflamación , Metaloproteinasa 9 de la Matriz/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células Mieloides/inmunología , Células Mieloides/patología , Peritoneo/inmunología , Peritoneo/patología , Peritonitis/inducido químicamente , Fosfatidilinositol 3-Quinasas/metabolismo , Transducción de Señal , Médula Espinal/inmunología , Médula Espinal/patología , Quinasa Syk , Tioglicolatos/administración & dosificaciónRESUMEN
For a series of beta-homophenylalanine based inhibitors of dipeptidyl peptidase IV ADME properties were improved by the incorporation of amide replacements. These efforts led to a novel series of potent and selective inhibitors of DPP-4 that exhibit an attractive pharmacokinetic profile and show excellent efficacy in an animal model of diabetes.
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Amidas/química , Aminobutiratos/química , Inhibidores de la Dipeptidil-Peptidasa IV , Relación Estructura-Actividad , Amidas/antagonistas & inhibidores , Células CACO-2 , Línea Celular Tumoral , Química Farmacéutica , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Inhibidores de la Dipeptidil-Peptidasa IV/uso terapéutico , Diseño de Fármacos , Humanos , Hipoglucemiantes/antagonistas & inhibidores , Hipoglucemiantes/uso terapéutico , Inhibidores de Proteasas/farmacocinética , Inhibidores de Proteasas/farmacología , Canales de Sodio/metabolismoRESUMEN
Adult neurogenesis persists in the subventricular zone and the dentate gyrus and can be induced upon central nervous system injury. However, the final contribution of newborn neurons to neuronal networks is limited. Here we show that in neural stem cells, stimulation of the "death receptor" CD95 does not trigger apoptosis but unexpectedly leads to increased stem cell survival and neuronal specification. These effects are mediated via activation of the Src/PI3K/AKT/mTOR signaling pathway, ultimately leading to a global increase in protein translation. Induction of neurogenesis by CD95 was further confirmed in the ischemic CA1 region, in the naive dentate gyrus, and after forced expression of CD95L in the adult subventricular zone. Lack of hippocampal CD95 resulted in a reduction in neurogenesis and working memory deficits. Following global ischemia, CD95-mediated brain repair rescued behavioral impairment. Thus, we identify the CD95/CD95L system as an instructive signal for ongoing and injury-induced neurogenesis.
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Células Madre Adultas/metabolismo , Isquemia Encefálica/metabolismo , Encéfalo/metabolismo , Proteína Ligando Fas/metabolismo , Neurogénesis/fisiología , Receptor fas/metabolismo , Células Madre Adultas/trasplante , Animales , Isquemia Encefálica/terapia , Femenino , Expresión Génica/fisiología , Memoria/fisiología , Ratones , Ratones Endogámicos C57BL , Neuronas/metabolismo , Proteínas Quinasas/metabolismo , Transducción de Señal/fisiología , Trasplante de Células Madre , Serina-Treonina Quinasas TORRESUMEN
A series of highly potent and selective inhibitors of DPP-4 was optimized for ADMET properties. The effort resulted in the discovery of inhibitor 1g, that exhibits excellent efficacy in an oral glucose tolerance test and an attractive pharmacokinetic profile.
Asunto(s)
Aminobutiratos/química , Inhibidores de la Dipeptidil-Peptidasa IV , Hidrocarburos Fluorados/química , Hipoglucemiantes/química , Inhibidores de Proteasas/química , Pirrolidinas/química , Administración Oral , Aminobutiratos/síntesis química , Aminobutiratos/farmacocinética , Animales , Células CACO-2 , Línea Celular Tumoral , Dipeptidil Peptidasa 4/metabolismo , Perros , Humanos , Hidrocarburos Fluorados/síntesis química , Hidrocarburos Fluorados/farmacología , Hipoglucemiantes/síntesis química , Hipoglucemiantes/farmacocinética , Inhibidores de Proteasas/síntesis química , Inhibidores de Proteasas/farmacocinética , Pirrolidinas/síntesis química , Pirrolidinas/farmacología , Ratas , Relación Estructura-ActividadRESUMEN
Modifications of DPP-4 inhibitor 5, that was discovered by structure based design, are described and structure-activity relationships discussed. With analogue 7k one of the most potent non-covalent inhibitors of DPP-4 reported to date (IC(50)=0.38nM) was discovered. X-ray structure of inhibitor 7k bound to DPP-4 revealed a hydrogen bonding interaction with Q553. First successful efforts in balancing overall properties, as demonstrated by improved metabolic stability, highlight the potential of this series.