RESUMO
Effective delivery of mRNA or small molecule drugs to the brain is a significant challenge in developing treatment for acute ischemic stroke (AIS). To address the problem, we have developed targeted nanomedicine to increase drug concentrations in endothelial cells of the blood-brain barrier (BBB) of the injured brain. Inflammation during ischemic stroke causes continuous neuronal death and an increase in the infarct volume. To enable targeted delivery to the inflamed BBB, we conjugated lipid nanocarriers (NCs) with antibodies that bind cell adhesion molecules expressed at the BBB. In the transient middle cerebral artery occlusion mouse model, NCs targeted to vascular cellular adhesion molecule-1 (VCAM) achieved the highest level of brain delivery, nearly two orders of magnitude higher than untargeted ones. VCAM-targeted lipid nanoparticles with luciferase-encoding mRNA and Cre-recombinase showed selective expression in the ischemic brain. Anti-inflammatory drugs administered intravenously after ischemic stroke reduced cerebral infarct volume by 62% (interleukin-10 mRNA) or 35% (dexamethasone) only when they were encapsulated in VCAM-targeted NCs. Thus, VCAM-targeted lipid NCs represent a new platform for strongly concentrating drugs within the compromised BBB of penumbra, thereby ameliorating AIS.
Assuntos
Barreira Hematoencefálica , Modelos Animais de Doenças , AVC Isquêmico , Lipossomos , Nanopartículas , Molécula 1 de Adesão de Célula Vascular , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/efeitos dos fármacos , Animais , Camundongos , Molécula 1 de Adesão de Célula Vascular/metabolismo , Molécula 1 de Adesão de Célula Vascular/genética , Nanopartículas/química , AVC Isquêmico/metabolismo , AVC Isquêmico/tratamento farmacológico , Lipídeos/química , Sistemas de Liberação de Medicamentos/métodos , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/tratamento farmacológico , HumanosRESUMO
Thromboprophylaxis is indicated in patients at an elevated risk of developing thrombotic disorders, typically using direct oral anticoagulants or low-molecular-weight heparins. We postulated that transient thromboprophylaxis (days-weeks) could be provided by a single dose of an anticoagulant engineered for prolonged pharmacokinetics. In the present work, d-phenylalanyl-l-prolyl-l-arginine chloromethyl ketone (PPACK) was used as a model anticoagulant to test the hypothesis that conjugation of thrombin inhibitors to the surface of albumin would provide durable protection against thrombotic insults. Covalent conjugates were formed between albumin and PPACK using click chemistry, and they were tested in vitro using a thrombin activity assay and a clot formation assay. Thromboprophylactic efficacy was tested in mouse models of arterial thrombosis, both chemically induced (FeCl3) and following ischemia-reperfusion (transient middle cerebral artery occlusion; tMCAO). Albumin-PPACK conjugates were shown to have nanomolar potency in both in vitro assays, and following intravenous injection had prolonged circulation. Conjugates did not impact hemostasis (tail clipping) or systemic coagulation parameters in normal mice. Intravenous injection of conjugates prior to FeCl3-induced thrombosis provided significant protection against occlusion of the middle cerebral and common carotid arteries, and injection immediately following ischemia-reperfusion reduced stroke volume measured 3 days after injury by â¼40% in the tMCAO model. The data presented here provide support for the use of albumin-linked anticoagulants as an injectable, long-circulating, safe thromboprophylactic agent. In particular, albumin-PPACK provides significant protection against thrombosis induced by multiple mechanisms, without adversely affecting hemostasis.
Assuntos
Trombose , Tromboembolia Venosa , Humanos , Camundongos , Animais , Anticoagulantes/uso terapêutico , Trombina/uso terapêutico , Tromboembolia Venosa/tratamento farmacológico , Trombose/tratamento farmacológico , Trombose/prevenção & controle , Clorometilcetonas de Aminoácidos/farmacologia , Clorometilcetonas de Aminoácidos/uso terapêutico , IsquemiaRESUMO
Drug targeting to inflammatory brain pathologies such as stroke and traumatic brain injury remains an elusive goal. Using a mouse model of acute brain inflammation induced by local tumor necrosis factor alpha (TNFα), we found that uptake of intravenously injected antibody to vascular cell adhesion molecule 1 (anti-VCAM) in the inflamed brain is >10-fold greater than antibodies to transferrin receptor-1 and intercellular adhesion molecule 1 (TfR-1 and ICAM-1). Furthermore, uptake of anti-VCAM/liposomes exceeded that of anti-TfR and anti-ICAM counterparts by â¼27- and â¼8-fold, respectively, achieving brain/blood ratio >300-fold higher than that of immunoglobulin G/liposomes. Single-photon emission computed tomography imaging affirmed specific anti-VCAM/liposome targeting to inflamed brain in mice. Intravital microscopy via cranial window and flow cytometry showed that in the inflamed brain anti-VCAM/liposomes bind to endothelium, not to leukocytes. Anti-VCAM/LNP selectively accumulated in the inflamed brain, providing de novo expression of proteins encoded by cargo messenger RNA (mRNA). Anti-VCAM/LNP-mRNA mediated expression of thrombomodulin (a natural endothelial inhibitor of thrombosis, inflammation, and vascular leakage) and alleviated TNFα-induced brain edema. Thus VCAM-directed nanocarriers provide a platform for cerebrovascular targeting to inflamed brain, with the goal of normalizing the integrity of the blood-brain barrier, thus benefiting numerous brain pathologies.
Assuntos
Anticorpos/administração & dosagem , Barreira Hematoencefálica/efeitos dos fármacos , Encefalite/tratamento farmacológico , Endotélio Vascular/efeitos dos fármacos , Nanomedicina/métodos , Animais , Barreira Hematoencefálica/imunologia , Encefalite/genética , Encefalite/imunologia , Endotélio Vascular/imunologia , Humanos , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/imunologia , Camundongos , Receptores da Transferrina/genética , Receptores da Transferrina/imunologia , Trombomodulina/genética , Trombomodulina/imunologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Molécula 1 de Adesão de Célula Vascular/genética , Molécula 1 de Adesão de Célula Vascular/imunologiaRESUMO
Engineering drug delivery systems for prolonged pharmacokinetics (PK) has been an ongoing pursuit for nearly 50 years. The gold standard for PK enhancement is the coating of nanoparticles with polymers, namely polyethylene glycol (PEGylation), which has been applied in several clinically used products. In the present work, we utilize the longest circulating and most abundant component of bloodâthe erythrocyteâto improve the PK behavior of liposomes. Antibody-mediated coupling of liposomes to erythrocytes was tested in vitro to identify a loading dose that did not adversely impact the carrier cells. Injection of erythrocyte targeting liposomes into mice resulted in a â¼2-fold improvement in the area under the blood concentration versus time profile versus PEGylated liposomes and a redistribution from the plasma into the cellular fraction of blood. These results suggest that in vivo targeting of erythrocytes is a viable strategy to improve liposome PK relative to current, clinically viable strategies.
Assuntos
Lipossomos , Polietilenoglicóis , Animais , Sistemas de Liberação de Medicamentos , Eritrócitos , Lipossomos/farmacocinética , Camundongos , Polietilenoglicóis/farmacocinética , PolímerosRESUMO
The use of single-domain antibody fragments, or nanobodies, has gained popularity in recent years as an alternative to traditional monoclonal antibody-based approaches. Relatively little is known, however, about the utility of nanobodies as targeting agents for delivery of therapeutic cargoes, particularly to vascular epitopes or in the setting of acute inflammatory conditions. We used a nanobody (VCAMelid) directed against mouse vascular cell adhesion molecule 1 (VCAM-1) and techniques for site-specific radiolabeling and bioconjugation to measure targeting to sites of constitutive and inducible antigen expression and investigate the impact of various characteristics (affinity, valence, circulation time) on nanobody biodistribution and pharmacokinetics. Engineering of VCAMelid for bivalent binding (BiVCAMelid) increased affinity by an order of magnitude and provided 2.8- and 3.6-fold enhancements in splenic and brain targeting in naive mice, with a further 2.6-fold increase in brain uptake in the setting of focal CNS inflammation. In contrast, introduction of an albumin-binding arm (VCAM/ALB8) did not affect binding affinity, but its prolonged circulation time resulted in 3.5-fold and 17.4-fold increases in splenic and brain uptake at 20 min post-dose and remarkable 40-, 25-, and 15-fold enhancements in overall exposure of blood, spleen, and brain, respectively, relative to both VCAMelid and BiVCAMelid. Both therapeutic protein (superoxide dismutase, SOD-1) and nanocarrier (liposome) delivery were enhanced by conjugation to VCAM-1 targeted nanobodies. The bispecific VCAM/ALB8 maintained its superiority over VCAMelid in enhancing both circulation time and organ targeting of SOD-1, but its advantages were largely blunted by conjugation to liposomes.
Assuntos
Portadores de Fármacos/farmacocinética , Engenharia de Proteínas , Anticorpos de Domínio Único/genética , Anticorpos de Domínio Único/metabolismo , Animais , Transporte Biológico , Encéfalo/metabolismo , Portadores de Fármacos/metabolismo , Marcação por Isótopo , Camundongos , Anticorpos de Domínio Único/imunologia , Baço/metabolismo , Distribuição Tecidual , Molécula 1 de Adesão de Célula Vascular/imunologiaRESUMO
Target-mediated drug disposition (TMDD) is often observed for targeted therapeutics, and manifests as decreases in clearance and volume of distribution with increasing dose as a result of saturable, high affinity target binding. In the present work, we demonstrate that classically defined TMDD is just one of the characteristic features of the system. In fact, for molecules with rapid non-specific elimination relative to target-mediated elimination, binding to target may actually lead to improved exposure at sub-saturating doses. This feature, which we refer to as target-mediated exposure enhancement (TMEE), produces the opposite trend to classical TMDD, i.e., with increasing dose levels, clearance and volume of distribution will also increase. The general model of TMDD was able to well-characterize the pharmacokinetics of two molecules that display TMEE, ALX-0081 and linagliptin. Additional fittings using the commonly reported TMDD model approximations revealed that both the quasi-equilibrium and quasi-steady-state approximations were able to well-describe TMEE; however, the Michaelis-Menten approximation was unable to describe this behavior. With the development of next-generation therapeutics with high affinity for target and rapid non-specific elimination, such as antibody fragments and peptides, this previously unexplored limit of TMDD is anticipated to become increasingly relevant for describing pharmacokinetics of investigational therapeutics.
Assuntos
Inibidores da Dipeptidil Peptidase IV/farmacocinética , Linagliptina/farmacocinética , Modelos Biológicos , Anticorpos de Domínio Único/farmacologia , Fator de von Willebrand/metabolismo , Administração Intravenosa , Adulto , Animais , Conjuntos de Dados como Assunto , Dipeptidil Peptidase 4/metabolismo , Inibidores da Dipeptidil Peptidase IV/administração & dosagem , Drogas em Investigação/administração & dosagem , Drogas em Investigação/farmacocinética , Voluntários Saudáveis , Humanos , Linagliptina/administração & dosagem , Macaca fascicularis , Masculino , Dinâmica não Linear , Distribuição Tecidual , Fator de von Willebrand/antagonistas & inibidoresRESUMO
The use of drug delivery systems (DDS) is an attractive approach to facilitate uptake of therapeutic agents at the desired site of action, particularly when free drug has poor pharmacokinetics/biodistribution (PK/BD) or significant off-site toxicities. Successful translation of DDS into the clinic is dependent on a thorough understanding of the in vivo behavior of the carrier, which has, for the most part, been an elusive goal. This is, at least in part, due to significant differences in the mechanisms controlling pharmacokinetics for classic drugs and DDSs. In this review, we summarize the key physiologic mechanisms controlling the in vivo behavior of DDS, compare and contrast this with classic drugs, and describe engineering strategies designed to improve DDS PK/BD. In addition, we describe quantitative approaches that could be useful for describing PK/BD of DDS, as well as critical steps between tissue uptake and pharmacologic effect.
Assuntos
Sistemas de Liberação de Medicamentos/métodos , Farmacocinética , Farmacologia , Animais , Tratamento Farmacológico/métodos , Humanos , Distribuição TecidualRESUMO
Inflamed organs display marked spatial heterogeneity of inflammation, with patches of inflamed tissue adjacent to healthy tissue. To investigate how nanocarriers (NCs) distribute between such patches, we created a mouse model that recapitulates the spatial heterogeneity of the inflammatory lung disease ARDS. NCs targeting the epitope PECAM strongly accumulated in the lungs, but were shunted away from inflamed lung regions due to hypoxic vasoconstriction (HVC). In contrast, ICAM-targeted NCs, which had lower whole-lung uptake than PECAM/NCs in inflamed lungs, displayed markedly higher NC levels in inflamed regions than PECAM/NCs, due to increased regional ICAM. Regional HVC, epitope expression, and capillary leak were sufficient to predict intra-organ of distribution of NCs, antibodies, and drugs. Importantly, these effects were not observable with traditional spatially-uniform models of ARDS, nor when examining only whole-organ uptake. This study underscores how examining NCs' intra-organ distribution in spatially heterogeneous animal models can guide rational NC design.
Assuntos
Portadores de Fármacos/farmacocinética , Epitopos/imunologia , Inflamação/patologia , Pulmão/patologia , Nanopartículas/química , Animais , Anticorpos/química , Portadores de Fármacos/química , Epitopos/química , Hipóxia/fisiopatologia , Inflamação/metabolismo , Molécula 1 de Adesão Intercelular/imunologia , Pulmão/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Molécula-1 de Adesão Celular Endotelial a Plaquetas/imunologia , Síndrome do Desconforto Respiratório/metabolismo , Síndrome do Desconforto Respiratório/patologia , VasoconstriçãoRESUMO
Accurate prediction of the clinical pharmacokinetics of new therapeutic entities facilitates decision making during drug discovery, and increases the probability of success for early clinical trials. Standard strategies employed for predicting the pharmacokinetics of small-molecule drugs (e.g., allometric scaling) are often not useful for predicting the disposition monoclonal antibodies (mAbs), as mAbs frequently demonstrate species-specific non-linear pharmacokinetics that is related to mAb-target binding (i.e., target-mediated drug disposition, TMDD). The saturable kinetics of TMDD are known to be influenced by a variety of factors, including the sites of target expression (which determines the accessibility of target to mAb), the extent of target expression, the rate of target turnover, and the fate of mAb-target complexes. In most cases, quantitative information on the determinants of TMDD is not available during early phases of drug discovery, and this has complicated attempts to employ mechanistic mathematical models to predict the clinical pharmacokinetics of mAbs. In this report, we introduce a simple strategy, employing physiologically-based modeling, to predict mAb disposition in humans. The approach employs estimates of inter-antibody variability in rate processes of extravasation in tissues and fluid-phase endocytosis, estimates for target concentrations in tissues derived through use of categorical immunohistochemical scores, and in vitro measures of the turnover of target and target-mAb complexes. Monte Carlo simulations were performed for four mAbs (cetuximab, figitumumab, dalotuzumab, trastuzumab) directed against three targets (epidermal growth factor receptor, insulin-like growth factor receptor 1, human epidermal growth factor receptor 2). The proposed modeling strategy was able to predict well the pharmacokinetics of cetuximab, dalotuzumab, and trastuzumab at a range of doses, but trended towards underprediction of figitumumab concentrations, particularly at high doses. The general agreement between model predictions and experimental observations suggests that PBPK modeling may be useful for the a priori prediction of the clinical pharmacokinetics of mAb therapeutics.
Assuntos
Anticorpos Monoclonais/farmacocinética , Modelos Biológicos , Fenômenos Fisiológicos , Anticorpos Monoclonais/administração & dosagem , Simulação por Computador , Endocitose/fisiologia , Humanos , Método de Monte Carlo , Especificidade de Órgãos/fisiologia , Farmacologia Clínica , Pinocitose/fisiologia , Distribuição Tecidual/fisiologiaRESUMO
Preclinical assessment of monoclonal antibody (mAb) disposition during drug development often includes investigations in non-human primate models. In many cases, mAb exhibit non-linear disposition that relates to mAb-target binding [i.e., target-mediated disposition (TMD)]. The goal of this work was to develop a physiologically-based pharmacokinetic (PBPK) model to predict non-linear mAb disposition in plasma and in tissues in monkeys. Physiological parameters for monkeys were collected from several sources, and plasma data for several mAbs associated with linear pharmacokinetics were digitized from prior literature reports. The digitized data displayed great variability; therefore, parameters describing inter-antibody variability in the rates of pinocytosis and convection were estimated. For prediction of the disposition of individual antibodies, we incorporated tissue concentrations of target proteins, where concentrations were estimated based on categorical immunohistochemistry scores, and with assumed localization of target within the interstitial space of each organ. Kinetics of target-mAb binding and target turnover, in the presence or absence of mAb, were implemented. The model was then employed to predict concentration versus time data, via Monte Carlo simulation, for two mAb that have been shown to exhibit TMD (2F8 and tocilizumab). Model predictions, performed a priori with no parameter fitting, were found to provide good prediction of dose-dependencies in plasma clearance, the areas under plasma concentration versu time curves, and the time-course of plasma concentration data. This PBPK model may find utility in predicting plasma and tissue concentration versus time data and, potentially, the time-course of receptor occupancy (i.e., mAb-target binding) to support the design and interpretation of preclinical pharmacokinetic-pharmacodynamic investigations in non-human primates.
Assuntos
Anticorpos Monoclonais/farmacocinética , Animais , Anticorpos Monoclonais Humanizados/farmacocinética , Haplorrinos , Modelos Biológicos , Método de Monte Carlo , Distribuição Tecidual/fisiologiaRESUMO
A challenge in development of peptide and protein therapeutics is rapid elimination from the body, necessitating frequent dosing that may lead to toxicities and/or poor patient compliance. To solve this issue, there has been great investment into half-life extension of rapidly eliminated drugs using approaches such as albumin binding, fusion to albumin or Fc, or conjugation to polyethylene glycol. Despite clinical successes of half-life extension products, no clear relationship has been drawn between properties of drugs and the pharmacokinetic parameters of their half-life extended analogues. In this study, non-compartmentally derived pharmacokinetic parameters (half-life, clearance, volume of distribution) were collected for 186 half-life extended drugs and their unmodified parent molecules. Statistical testing and regression analysis was performed to evaluate relationships between pharmacokinetic parameters and a matrix of variables. Multivariate linear regression models were developed for each of the three pharmacokinetic parameters and model predictions were in good agreement with observed data with r2 values for each parameter being: half-life: 0.879, clearance: 0.820, volume of distribution: 0.937. Significant predictors for each parameter included the corresponding pharmacokinetic parameter of the parent drug and descriptors related to molecular weight. This model represents a useful tool for prediction of the potential benefits of half-life extension.
Assuntos
Algoritmos , Meia-Vida , Preparações Farmacêuticas/administração & dosagem , Preparações Farmacêuticas/química , Preparações Farmacêuticas/metabolismo , Humanos , Modelos Biológicos , Farmacocinética , Modelos LinearesRESUMO
RNA therapeutics are an emerging, powerful class of drugs with potential applications in a wide range of disorders. A central challenge in their development is the lack of clear pharmacokinetic (PK)-pharmacodynamic relationship, in part due to the significant delay between the kinetics of RNA delivery and the onset of pharmacologic response. To bridge this gap, we have developed a physiologically based PK/pharmacodynamic model for systemically administered mRNA-containing lipid nanoparticles (LNPs) in mice. This model accounts for the physiologic determinants of mRNA delivery, active targeting in the vasculature, and differential transgene expression based on nanoparticle coating. The model was able to well-characterize the blood and tissue PKs of LNPs, as well as the kinetics of tissue luciferase expression measured by ex vivo activity in organ homogenates and bioluminescence imaging in intact organs. The predictive capabilities of the model were validated using a formulation targeted to intercellular adhesion molecule-1 and the model predicted nanoparticle delivery and luciferase expression within a 2-fold error for all organs. This modeling platform represents an initial strategy that can be expanded upon and utilized to predict the in vivo behavior of RNA-containing LNPs developed for an array of conditions and across species.
RESUMO
For medical emergencies, such as acute ischemic stroke, rapid drug delivery to the target site is essential. For many small molecule drugs, this goal is unachievable due to poor solubility that prevents intravenous administration, and less obviously, by extensive partitioning to plasma proteins and red blood cells (RBCs), which greatly slows delivery to the target. Here we study these effects and how they can be solved by loading into nanoscale drug carriers. We focus on fingolimod, a small molecule drug that is FDA-approved for treatment of multiple sclerosis, which has also shown promise in the treatment of stroke. Unfortunately, fingolimod has poor solubility and very extensive partitioning to plasma proteins and RBCs (in whole blood, 86% partitions to RBCs, 13.96% to plasma proteins, and 0.04% is free). We develop a liposomal formulation that slows the partitioning of fingolimod to RBCs and plasma proteins, enables intravenous delivery, and additionally prevents fingolimod toxicity to RBCs. The liposomal formulation nearly completely prevented fingolimod adsorption to plasma proteins (association with plasma proteins was 98.4⯱â¯0.4% for the free drug vs. 5.6⯱â¯0.4% for liposome-loaded drug). When incubated with whole blood in vitro, the liposomal formulation greatly slowed partitioning of fingolimod to RBCs and also eliminated deleterious effects of fingolimod on RBC rigidity, morphology, and hemolysis. In vivo, the liposomal formulation delayed fingolimod partitioning to RBCs for over 30â¯min, a critical time window for stroke. Fingolimod-loaded liposomes showed improved efficacy in a mouse model of post-stroke neuroinflammation, completely sealing the leaky blood-brain barrier (114⯱â¯11.5% reduction in albumin leak into the brain for targeted liposomes vs. 38⯱â¯16.5% reduction for free drug). This effect was only seen for liposomes modified with antibodies to enable targeted delivery to the site of action, and not in unmodified, long-circulating liposomes. Thus, loading fingolimod into liposomes prevented partitioning to RBCs and associated toxicities and enabled targeted delivery. This paradigm can be used for tuning the blood distribution of small molecule drugs for the treatment of acute illnesses requiring rapid pharmacologic intervention.
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PD-1 checkpoint inhibitors have revolutionized the treatment of patients with different cancer histologies including melanoma, renal cell carcinoma, and non-small cell lung carcinoma. However, only a subset of patients show a dramatic clinical response to treatment. Despite intense biomarker discovery efforts, no single robust, prognostic correlation has emerged as a valid outcome predictor. Immune competent, pet dogs develop spontaneous tumors that share similar features to human cancers including chromosome aberrations, molecular subtypes, immune signatures, tumor heterogeneity, metastatic behavior, and chemotherapeutic response. As such, they represent a valuable parallel patient population in which to investigate predictive biomarkers of checkpoint inhibition. However, the lack of a validated, non-immunogenic, canine anti-PD-1 antibody for pre-clinical use hinders this comparative approach and prevents potential clinical benefits of PD-1 blockade being realized in the veterinary clinic. To address this, fully canine single-chain variable fragments (scFvs) that bind canine (c)PD-1 were isolated from a comprehensive canine scFv phage display library. Lead candidates were identified that bound with high affinity to cPD-1 and inhibited its interaction with canine PD-L1 (cPD-L1). The lead scFv candidate re-formatted into a fully canine IgGD reversed the inhibitory effects of cPD-1:cPD-L1 interaction on canine chimeric antigen receptor (CAR) T cell function. In vivo administration showed no toxicity and revealed favorable pharmacokinetics for a reasonable dosing schedule. These results pave the way for clinical trials with anti-cPD-1 in canine cancer patients to investigate predictive biomarkers and combination regimens to inform human clinical trials and bring a promising checkpoint inhibitor into the veterinary armamentarium.
Assuntos
Melanoma , Pesquisa Translacional Biomédica , Humanos , Cães , Animais , Receptor de Morte Celular Programada 1 , Anticorpos Monoclonais/uso terapêutico , Inibidores de Checkpoint Imunológico , Antígeno B7-H1RESUMO
Thorough characterization of the plasma pharmacokinetics (PK) is a critical step in clinical development of novel therapeutics and is routinely performed for small molecules and biologics. However, there is a paucity of even basic characterization of PK for nanoparticle-based drug delivery systems. This has led to untested generalizations about how nanoparticle properties govern PK. Here, we present a meta-analysis of 100 nanoparticle formulations following IV administration in mice to identify any correlations between four PK parameters derived by non-compartmental analysis (NCA) and four cardinal properties of nanoparticles: PEGylation, zeta potential, size, and material. There was a statistically significant difference between the PK of particles stratified by nanoparticle properties. However, single linear regression between these properties and PK parameters showed poor predictability (r2 < 0.10 for all analyses), while multivariate regressions showed improved predictability (r2 > 0.38, except for t1/2). This suggests that no single nanoparticle property alone is even moderately predictive of PK, while the combination of multiple nanoparticle features does provide moderate predictive power. Improved reporting of nanoparticle properties will enable more accurate comparison between nanoformulations and will enhance our ability to predict in vivo behavior and design optimal nanoparticles.
Assuntos
Nanopartículas , Animais , Camundongos , Composição de Medicamentos , FarmacocinéticaRESUMO
Intracerebral hemorrhage (ICH) is one of the most common causes of fatal stroke, yet has no specific drug therapies. Many attempts at passive intravenous (IV) delivery in ICH have failed to deliver drugs to the salvageable area around the hemorrhage. The passive delivery method assumes vascular leak through the ruptured blood-brain barrier will allow drug accumulation in the brain. Here we tested this assumption using intrastriatal injection of collagenase, a well-established experimental model of ICH. Fitting with hematoma expansion in clinical ICH, we showed that collagenase-induced blood leak drops significantly by 4 h after ICH onset and is gone by 24 h. We observed passive-leak brain accumulation also declines rapidly over â¼4 h for 3 model IV therapeutics (non-targeted IgG; a protein therapeutic; PEGylated nanoparticles). We compared these passive leak results with targeted brain delivery by IV monoclonal antibodies (mAbs) that actively bind vascular endothelium (anti-VCAM, anti-PECAM, anti-ICAM). Even at early time points after ICH induction, where there is high vascular leak, brain accumulation via passive leak is dwarfed by brain accumulation of endothelial-targeted agents: At 4 h after injury, anti-PECAM mAbs accumulate at 8-fold higher levels in the brain vs. non-immune IgG; anti-VCAM nanoparticles (NPs) deliver a protein therapeutic (superoxide dismutase, SOD) at 4.5-fold higher levels than the carrier-free therapeutic at 24 h after injury. These data suggest that relying on passive vascular leak provides inefficient delivery of therapeutics even at early time points after ICH, and that a better strategy might be targeted delivery to the brain endothelium, which serves as the gateway for the immune attack on the peri-hemorrhage inflamed brain region.
Assuntos
Encéfalo , Hemorragia Cerebral , Animais , Hemorragia Cerebral/tratamento farmacológico , Hemorragia Cerebral/induzido quimicamente , Hemorragia Cerebral/metabolismo , Encéfalo/metabolismo , Endotélio Vascular/metabolismo , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais/metabolismo , Colagenases/efeitos adversos , Colagenases/metabolismo , Imunoglobulina G/uso terapêutico , Modelos Animais de DoençasRESUMO
Diseases of the pulmonary alveolus, such as pulmonary fibrosis, are leading causes of morbidity and mortality, but exceedingly few drugs are developed for them. A major reason for this gap is that after inhalation, drugs are quickly whisked away from alveoli due to their high perfusion. To solve this problem, the mechanisms by which nano-scale drug carriers dramatically improve lung pharmacokinetics using an inhalable liposome formulation containing nintedanib, an antifibrotic for pulmonary fibrosis, are studied. Direct instillation of liposomes in murine lung increases nintedanib's total lung delivery over time by 8000-fold and lung half life by tenfold, compared to oral nintedanib. Counterintuitively, it is shown that pulmonary surfactant neither lyses nor aggregates the liposomes. Instead, each lung compartment (alveolar fluid, alveolar leukocytes, and parenchyma) elutes liposomes over 24 h, likely serving as "drug depots." After deposition in the surfactant layer, liposomes are transferred over 3-6 h to alveolar leukocytes (which take up a surprisingly minor 1-5% of total lung dose instilled) in a nonsaturable fashion. Further, all cell layers of the lung parenchyma take up liposomes. These and other mechanisms elucidated here should guide engineering of future inhaled nanomedicine for alveolar diseases.
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After more than 100 failed drug trials for acute ischemic stroke (AIS), one of the most commonly cited reasons for the failure has been that drugs achieve very low concentrations in the at-risk penumbra. To address this problem, here we employ nanotechnology to significantly enhance drug concentration in the penumbra's blood-brain barrier (BBB), whose increased permeability in AIS has long been hypothesized to kill neurons by exposing them to toxic plasma proteins. To devise drug-loaded nanocarriers targeted to the BBB, we conjugated them with antibodies that bind to various cell adhesion molecules on the BBB endothelium. In the transient middle cerebral artery occlusion (tMCAO) mouse model, nanocarriers targeted with VCAM antibodies achieved the highest level of brain delivery, nearly 2 orders of magnitude higher than untargeted ones. VCAM-targeted lipid nanoparticles loaded with either a small molecule drug (dexamethasone) or mRNA (encoding IL-10) reduced cerebral infarct volume by 35% or 73%, respectively, and both significantly lowered mortality rates. In contrast, the drugs delivered without the nanocarriers had no effect on AIS outcomes. Thus, VCAM-targeted lipid nanoparticles represent a new platform for strongly concentrating drugs within the compromised BBB of penumbra, thereby ameliorating AIS. Graphical abstract: Acute ischemic stroke induces upregulation of VCAM. We specifically targeted upregulated VCAM in the injured region of the brain with drug- or mRNA-loaded targeted nanocarriers. Nanocarriers targeted with VCAM antibodies achieved the highest brain delivery, nearly orders of magnitude higher than untargeted ones. VCAM-targeted nanocarriers loaded with dexamethasone and mRNA encoding IL-10 reduced infarct volume by 35% and 73%, respectively, and improved survival rates.
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Ex vivo-loaded white blood cells (WBC) can transfer cargo to pathological foci in the central nervous system (CNS). Here we tested affinity ligand driven in vivo loading of WBC in order to bypass the need for ex vivo WBC manipulation. We used a mouse model of acute brain inflammation caused by local injection of tumor necrosis factor alpha (TNF-α). We intravenously injected nanoparticles targeted to intercellular adhesion molecule 1 (anti-ICAM/NP). We found that (A) at 2 h, >20% of anti-ICAM/NP were localized to the lungs; (B) of the anti-ICAM/NP in the lungs >90% were associated with leukocytes; (C) at 6 and 22 h, anti-ICAM/NP pulmonary uptake decreased; (D) anti-ICAM/NP uptake in brain increased up to 5-fold in this time interval, concomitantly with migration of WBCs into the injured brain. Intravital microscopy confirmed transport of anti-ICAM/NP beyond the blood-brain barrier and flow cytometry demonstrated complete association of NP with WBC in the brain (98%). Dexamethasone-loaded anti-ICAM/liposomes abrogated brain edema in this model and promoted anti-inflammatory M2 polarization of macrophages in the brain. In vivo targeted loading of WBC in the intravascular pool may provide advantages of coopting WBC predisposed to natural rapid mobilization from the lungs to the brain, connected directly via conduit vessels.
Assuntos
Sistemas de Liberação de Medicamentos , Pulmão , Camundongos , Animais , Pulmão/metabolismo , Encéfalo/metabolismo , Lipossomos/metabolismo , Leucócitos/metabolismo , Molécula 1 de Adesão Intercelular/metabolismoRESUMO
Acute inflammation is a common dangerous component of pathogenesis of many prevalent conditions with high morbidity and mortality including sepsis, thrombosis, acute respiratory distress syndrome (ARDS), COVID-19, myocardial and cerebral ischemia-reperfusion, infection, and trauma. Inflammatory changes of the vasculature and blood mediate the course and outcome of the pathology in the tissue site of insult, remote organs and systemically. Endothelial cells lining the luminal surface of the vasculature play the key regulatory functions in the body, distinct under normal vs. pathological conditions. In theory, pharmacological interventions in the endothelial cells might enable therapeutic correction of the overzealous damaging pro-inflammatory and pro-thrombotic changes in the vasculature. However, current agents and drug delivery systems (DDS) have inadequate pharmacokinetics and lack the spatiotemporal precision of vascular delivery in the context of acute inflammation. To attain this level of precision, many groups design DDS targeted to specific endothelial surface determinants. These DDS are able to provide specificity for desired tissues, organs, cells, and sub-cellular compartments needed for a particular intervention. We provide a brief overview of endothelial determinants, design of DDS targeted to these molecules, their performance in experimental models with focus on animal studies and appraisal of emerging new approaches. Particular attention is paid to challenges and perspectives of targeted therapeutics and nanomedicine for advanced management of acute inflammation.