ABSTRACT
Delivery of biologics via cerebrospinal fluid (CSF) has demonstrated potential to access the tissues of the central nervous system (CNS) by circumventing the blood-brain barrier and blood-CSF barrier. Developing an effective CSF drug delivery strategy requires optimization of multiple parameters, including choice of CSF access point, delivery device technology, and delivery kinetics to achieve effective therapeutic concentrations in the target brain region, whereas also considering the biologic modality, mechanism of action, disease indication, and patient population. This review discusses key preclinical and clinical examples of CSF delivery for different biologic modalities (antibodies, nucleic acid-based therapeutics, and gene therapy) to the brain via CSF or CNS access routes (intracerebroventricular, intrathecal-cisterna magna, intrathecal-lumbar, intraparenchymal, and intranasal), including the use of novel device technologies. This review also discusses quantitative models of CSF flow that provide insight into the effect of fluid dynamics in CSF on drug delivery and CNS distribution. Such models can facilitate delivery device design and pharmacokinetic/pharmacodynamic translation from preclinical species to humans in order to optimize CSF drug delivery to brain regions of interest.
Subject(s)
Biological Products , Brain , Biological Transport/physiology , Blood-Brain Barrier , Central Nervous System , HumansABSTRACT
Monoclonal antibodies (mAbs), which are now more frequently administered by subcutaneous (SC) injection rather than intravenously, have become a tremendously successful drug format across a wide range of therapeutic areas. Preclinical evaluations of mAbs to be administered by SC injection are typically performed in species such as mice, rats, minipigs, and cynomolgus monkeys to obtain critical information regarding formulation performance and prediction of PK/PD outcomes needed to select clinical doses for first-in-human studies. Despite extensive efforts, no preclinical model has been identified to date that accurately predicts clinical outcomes for these SC injections. We have addressed this deficiency with a novel in vitro instrument, termed Scissor, to model events occurring at the SC injection site and now further validated this approach using a set of eight mAbs for which clinical PK/PD outcomes have been obtained. Diffusion of these mAbs from the Scissor system injection cartridge into a large volume physiological buffer, used to emulate mAb movement from the SC injection site into the systemic circulation, provided distinct profiles when monitored over a 6h period. Curve-fitting analysis of these profiles using the Hill equation identified parameters that were used, along with physicochemical properties for each mAb, in a partial least squares analysis to define a relationship between molecule and formulation properties with clinical PK outcomes. The results demonstrate that parameters of protein charge at neutral pH and isoelectric point (pI) along with combined formulation properties such as viscosity and mAb concentration can dictate the movement of the mAb from the injection cartridge to infinite sink compartment. Examination of profile characteristics of this movement provided a strong predictive correlation for these eight mAbs. Together, this approach demonstrates the feasibility of this in vitro modelling strategy as a tool to identify drug and formulation properties that can define the performance of SC injected medicines and provide the potential for predicting clinical outcomes that could be useful for formulation selection and a first-in-human clinical dosing strategy.
Subject(s)
Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal/pharmacokinetics , Drug Delivery Systems , Models, Biological , Animals , Biological Availability , Humans , Injections, Subcutaneous , Macaca fascicularis , Mice , Rats , Swine , Swine, MiniatureABSTRACT
BACKGROUND AND PURPOSE: The potential for therapeutic antibody treatment of neurological diseases is limited by poor penetration across the blood-brain barrier. I.c.v. delivery is a promising route to the brain; however, it is unclear how efficiently antibodies delivered i.c.v. penetrate the cerebrospinal spinal fluid (CSF)-brain barrier and distribute throughout the brain parenchyma. EXPERIMENTAL APPROACH: We evaluated the pharmacokinetics and pharmacodynamics of an inhibitory monoclonal antibody against ß-secretase 1 (anti-BACE1) following continuous infusion into the left lateral ventricle of healthy adult cynomolgus monkeys. KEY RESULTS: Animals infused with anti-BACE1 i.c.v. showed a robust and sustained reduction (~70%) of CSF amyloid-ß (Aß) peptides. Antibody distribution was near uniform across the brain parenchyma, ranging from 20 to 40 nM, resulting in a ~50% reduction of Aß in the cortical parenchyma. In contrast, animals administered anti-BACE1 i.v. showed no significant change in CSF or cortical Aß levels and had a low (~0.6 nM) antibody concentration in the brain. CONCLUSION AND IMPLICATIONS: I.c.v. administration of anti-BACE1 resulted in enhanced BACE1 target engagement and inhibition, with a corresponding dramatic reduction in CNS Aß concentrations, due to enhanced brain exposure to antibody.
Subject(s)
Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid beta-Peptides/metabolism , Antibodies, Monoclonal/pharmacology , Antibodies, Monoclonal/pharmacokinetics , Aspartic Acid Endopeptidases/antagonists & inhibitors , Amyloid Precursor Protein Secretases/immunology , Amyloid beta-Peptides/blood , Amyloid beta-Peptides/cerebrospinal fluid , Animals , Antibodies, Monoclonal/blood , Antibodies, Monoclonal/cerebrospinal fluid , Aspartic Acid Endopeptidases/immunology , Brain/metabolism , Female , Infusions, Intraventricular , Macaca fascicularisABSTRACT
Purpose: Poly(lactic-co-glycolic) acid (PLGA) inserts have been successfully developed for the treatment of posterior eye disease as a means of reducing injection frequency of intravitreally administered therapeutics. PLGA microspheres are also of interest for the delivery of intravitreal drugs, since they offer the advantage of being easily injected without surgical procedures or large injectors. Methods: In the current study, the toxicity of PLGA microspheres and rods was investigated in nonhuman primates (NHPs) and rabbits. An in vitro assessment of cytokine responses to PLGA in peripheral blood mononuclear cells (PBMCs) and macrophages was also performed. Results: Intravitreal administration of 3, 10, or 12.5 mg/eye of PLGA microspheres in NHPs resulted in a severe immune response characterized by a foreign body response. Follow-up studies in the rabbit confirmed this finding for PLGA microspheres ranging in size from 20 to 100 µm. In contrast, administration of PLGA rod implants with a similar PLGA mass did not elicit a significant immune response. In vitro assays in PBMCs and macrophages confirmed proinflammatory cytokine release upon treatment with PLGA microspheres but not PLGA rods. Conclusions: These data demonstrate a lack of tolerability of PLGA microspheres upon intravitreal injection, and suggest that the size, shape, and/or surface area of PLGA depots are critical attributes in determining ocular toxicity.
Subject(s)
Biocompatible Materials/toxicity , Drug Delivery Systems/adverse effects , Lactic Acid/toxicity , Microspheres , Polyglycolic Acid/toxicity , Animals , Biocompatible Materials/administration & dosage , Cytokines/metabolism , Disease Models, Animal , Foreign-Body Reaction/metabolism , Foreign-Body Reaction/pathology , Intravitreal Injections , Lactic Acid/administration & dosage , Macrophages/metabolism , Polyglycolic Acid/administration & dosage , Polylactic Acid-Polyglycolic Acid Copolymer , Rabbits , Retina/drug effects , Retina/pathology , Vitreous Body/drug effects , Vitreous Body/pathologyABSTRACT
Subcutaneous (SC) injection is becoming a more common route for the administration of biopharmaceuticals. Currently, there is no reliable in vitro method that can be used to anticipate the in vivo performance of a biopharmaceutical formulation intended for SC injection. Nor is there an animal model that can predict in vivo outcomes such as bioavailability in humans. We address this unmet need by the development of a novel in vitro system, termed Scissor (Subcutaneous Injection Site Simulator). The system models environmental changes that a biopharmaceutical could experience as it transitions from conditions of a drug product formulation to the homeostatic state of the hypodermis following SC injection. Scissor uses a dialysis-based injection chamber, which can incorporate various concentrations and combinations of acellular extracellular matrix (ECM) components that may affect the release of a biopharmaceutical from the SC injection site. This chamber is immersed in a container of a bicarbonate-based physiological buffer that mimics the SC injection site and the infinite sink of the body. Such an arrangement allows for real-time monitoring of the biopharmaceutical within the injection chamber, and can be used to characterize physicochemical changes of the drug and its interactions with ECM components. Movement of a biopharmaceutical from the injection chamber to the infinite sink compartment simulates the drug migration from the injection site and uptake by the blood and/or lymph capillaries. Here, we present an initial evaluation of the Scissor system using the ECM element hyaluronic acid and test formulations of insulin and four different monoclonal antibodies. Our findings suggest that Scissor can provide a tractable method to examine the potential fate of a biopharmaceutical formulation after its SC injection in humans and that this approach may provide a reliable and representative alternative to animal testing for the initial screening of SC formulations.
Subject(s)
Biological Products/administration & dosage , Biological Products/pharmacokinetics , Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/pharmacokinetics , Bicarbonates/chemistry , Biological Products/chemistry , Buffers , Chemistry, Pharmaceutical , Dialysis , Drug Design , Extracellular Matrix/metabolism , Humans , Hyaluronic Acid/metabolism , Injections, Subcutaneous , Lymph/metabolism , Subcutaneous Tissue/metabolismABSTRACT
Subcutaneous (SC) delivery is a common route of administration for therapeutic monoclonal antibodies (mAbs) with pharmacokinetic (PK)/pharmacodynamic (PD) properties requiring long-term or frequent drug administration. An ideal in vivo preclinical model for predicting human PK following SC administration may be one in which the skin and overall physiological characteristics are similar to that of humans. In this study, the PK properties of a series of therapeutic mAbs following intravenous (IV) and SC administration in Göttingen minipigs were compared with data obtained previously from humans. The present studies demonstrated: (1) minipig is predictive of human linear clearance; (2) the SC bioavailabilities in minipigs are weakly correlated with those in human; (3) minipig mAb SC absorption rates are generally higher than those in human and (4) the SC bioavailability appears to correlate with systemic clearance in minipigs. Given the important role of the neonatal Fc-receptor (FcRn) in the PK of mAbs, the in vitro binding affinities of these IgGs against porcine, human and cynomolgus monkey FcRn were tested. The result showed comparable FcRn binding affinities across species. Further, mAbs with higher isoelectric point tended to have faster systemic clearance and lower SC bioavailability in both minipig and human. Taken together, these data lend increased support for the use of the minipig as an alternative predictive model for human IV and SC PK of mAbs.
Subject(s)
Antibodies, Monoclonal/pharmacology , Antibodies, Monoclonal/pharmacokinetics , Models, Immunological , Administration, Intravenous , Animals , Antibodies, Monoclonal/immunology , Female , Humans , Injections, Subcutaneous , Male , Swine , Swine, MiniatureABSTRACT
Local delivery of therapeutic angiogenic agents that stimulate blood vessel formation represents a promising strategy for the treatment of peripheral vascular disease (PVD). At present, requirements for temporal and spatial parameters for localized delivery are unclear, with a variety of sustained delivery approaches being examined. Two polymer-based sustained formulations containing the 165 amino acid isoform of human recombinant vascular endothelial growth factor-A (rhVEGF(165)) were evaluated for their potential application in the treatment of PVD following intramuscular injection. Microspheres prepared from a 50:50 ratio of polylactic-co-glycolic acid (PLGA) and a gel of PLGA polymer solubilized in N-methyl pyrrolidone (PLGA:NMP) were each loaded with rhVEGF(165) and tested in vitro and in vivo. PLGA microspheres averaged â¼30 µm in diameter and contained 8.9% (w/w) rhVEGF(165), while the PLGA:NMP gel was formulated with varying amounts of spray freeze-dried rhVEGF(165) to result in final gel formulations having concentrations of 0.36, 0.72, or 3.6 mg/mL rhVEGF(165). In vitro release of rhVEGF(165) from PLGA microspheres showed â¼10% cumulative release by day 6, whereas the cumulative release of rhVEGF(165) from the PLGA:NMP gel matrices (0.65% w/w loading) was less than 0.25% at this same time point. While the in vitro release characteristics of these two sustained release formulations were broadly different, the plasma rhVEGF(165) concentration-time profiles following hind-limb intramuscular (IM) injection of these formulations in non-compromised rats revealed similar in vivo pharmacokinetics. Three-dimensional resin casts of vascular architecture were prepared at days 3, 7, 14, 21, 28, 60, and 75 following a single IM dosing of these sustained release microsphere and gel matrix formulations in the gastrocnemius muscle of immune-compromised mice. Scanning electron microscopic visualization of these vascular casts demonstrated spatial arrangement of capillary sprouts and vessel enlargement consistent with profound vascular changes occurring within 3 days of dosing that persisted for 2 months, approximately 1 month beyond the anticipated completion of rhVEGF(165) release from these sustained delivery formulations. Vascular re-modeling events were correlated with histological and immunohistochemical parameters attributed to known biological actions of rhVEGF(165) signaling. Together, these pharmacokinetic and pharmacodynamic results support the use of sustained release PLGA-based formulations for the local delivery of rhVEGF(165) to achieve a durable vascular re-modeling response.
Subject(s)
Disease Models, Animal , Neovascularization, Pathologic/prevention & control , Vascular Endothelial Growth Factor A/pharmacology , Animals , Female , Humans , Mice , Mice, Nude , Recombinant Proteins/chemistry , Recombinant Proteins/pharmacology , Vascular Endothelial Growth Factor A/chemistryABSTRACT
Vascular endothelial growth factor (VEGF) is an extracellular matrix (ECM)-binding growth factor capable of driving neovascularization. VEGF can potentially be applied clinically via intramuscular (IM) injection to correct local ischemia associated with peripheral artery disease (PAD). As interactions with ECM elements and cognate receptors at the site of an IM injection define the local biology of VEGF and previous studies have only focused on systemic distribution measurements, we have established a method to monitor the local VEGF distribution and fate. Fluorescent-labeled VEGF was prepared that bound to ECM and activated a cognate receptor similarly to VEGF. Beginning by 2 h and becoming complete by 12 h following injection, fluorescence microscopy demonstrated the transition of labeled VEGF from an initial extensive interaction with ECM components to a focused labeling of vascular endothelial cells. Biochemical characterization verified the association of VEGF with ECM components and modification of endothelial cell function associated with vascular permeability changes known to accompany VEGF actions. Our data provide information concerning temporal and spatial VEGF fate and actions at the site of an IM injection that can help guide decisions regarding the identification of acceptable formulation strategies.
Subject(s)
Endothelium, Vascular/metabolism , Extracellular Matrix/metabolism , Vascular Endothelial Growth Factor A/pharmacokinetics , Animals , Female , Injections, Intramuscular , Microscopy, Fluorescence , Permeability , Rats , Rats, Sprague-Dawley , Time Factors , Tissue DistributionABSTRACT
PURPOSE: To quantify spontaneous and therapeutic arteriogenesis in vivo in a murine model of peripheral arterial disease using magnetic resonance angiography. MATERIALS AND METHODS: Male, 8-12-week-old, C57/BL6 mice underwent femoral artery ligation; 21 days later, 2 mg/kg recombinant murine VEGF165, formulated for slow release, was injected into the ipsilateral gastrocnemius. The spontaneous (following ligation) and therapeutic (following vascular endothelial growth factor (VEGF)) formation of collateral vessels was quantified using 3D magnetic resonance angiography on a small-bore 4.7T system. Therapeutically induced angiogenesis and blood flow were quantified using an in situ anti-platelet endothelial cell adhesion molecule (PECAM) 1 radioimmunoassay and radiolabeled microsphere deposition, respectively. RESULTS: Spontaneous arteriogenesis was visible in all animals five days after ligation. VEGF treatment doubled the arteriogenic response five days after treatment compared to vehicle (cross-sectional area of vessels: 0.96 vs. 0.46 mm2, P<0.01). VEGF also induced angiogenesis (PECAM1 levels 191% of vehicle, P<0.05) and increased blood flow specific to the injection site (57 vs. 7 mL/minute/100 g, P<0.05). CONCLUSION: The presented methodology allowed in vivo quantification of spontaneous arteriogenesis in a murine model of peripheral arterial disease and demonstrated that therapeutic enlargement of collateral vessels is possible with VEGF.
Subject(s)
Blood Flow Velocity/drug effects , Disease Models, Animal , Magnetic Resonance Angiography/methods , Neovascularization, Physiologic/drug effects , Peripheral Vascular Diseases/drug therapy , Peripheral Vascular Diseases/pathology , Vascular Endothelial Growth Factor A/administration & dosage , Animals , Collateral Circulation/drug effects , Male , Mice , Mice, Inbred C57BL , Prognosis , Treatment OutcomeABSTRACT
Antibodies can have exquisite specificity of target recognition and thus generate highly selective outcomes following their systemic administration. While antibodies can have high specificity, the doses required to treat patients, particularly for a chronic condition, are typically large. Fortunately, advances in production and purification capacities have allowed for the exceptionally large amounts of highly purified monoclonal antibodies to be produced. Additionally, genetic engineering of antibodies has provided a stable of antibody-like proteins that can be easier to prepare. Together, these advances have made antibody-based therapies one of the most commonly pursued pharmaceuticals in biotechnology pipelines. With this success, however, has come a series of technical challenges in the formulation of antibody-based materials to maintain sufficient stability in a variety of configurations and sometimes at particularly high concentrations. This review focuses on issues related to identifying and verifying stable antibody-based formulations.
Subject(s)
Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal/therapeutic use , Chemistry, Pharmaceutical , Animals , Drug Delivery Systems , Drug Stability , HumansABSTRACT
In the past decade, genomic research and the nascent field of proteomics have exponentially increased the number of potential protein therapeutic molecules for treating medical needs that were previously unmet. To realise the full clinical potential of many of the novel protein drug entities arising from these intense research efforts, emerging protein delivery technologies may be required. Advanced delivery technologies may offer the ability to overcome biochemical and anatomical barriers to protein drug transport, without incurring adverse events, to deliver the agent(s) at a certain desired rate and duration, to protect therapeutic macromolecules from in situ or systemic degradation, as well as increase their therapeutic index by targeting the drug action to a specific site. This review will cover a myriad of novel and emerging technologies that are directed at bypassing biological barriers and that have shown promise in advancing the therapeutic potential of protein drugs.
Subject(s)
Proteins/administration & dosage , Proteins/therapeutic use , Proteomics/trends , Administration, Cutaneous , Administration, Inhalation , Administration, Oral , Animals , Blood-Air Barrier , Blood-Brain Barrier , Drug Delivery Systems , Humans , Peripheral Nervous System/physiologyABSTRACT
Several growth factors are expressed in distinct temporal and spatial patterns during fracture repair. Of these, vascular endothelial growth factor, VEGF, is of particular interest because of its ability to induce neovascularization (angiogenesis). To determine whether VEGF is required for bone repair, we inhibited VEGF activity during secondary bone healing via a cartilage intermediate (endochondral ossification) and during direct bone repair (intramembranous ossification) in a novel mouse model. Treatment of mice with a soluble, neutralizing VEGF receptor decreased angiogenesis, bone formation, and callus mineralization in femoral fractures. Inhibition of VEGF also dramatically inhibited healing of a tibial cortical bone defect, consistent with our discovery of a direct autocrine role for VEGF in osteoblast differentiation. In separate experiments, exogenous VEGF enhanced blood vessel formation, ossification, and new bone (callus) maturation in mouse femur fractures, and promoted bony bridging of a rabbit radius segmental gap defect. Our results at specific time points during the course of healing underscore the role of VEGF in endochondral vs. intramembranous ossification, as well as skeletal development vs. bone repair. The responses to exogenous VEGF observed in two distinct model systems and species indicate that a slow-release formulation of VEGF, applied locally at the site of bone damage, may prove to be an effective therapy to promote human bone repair.
Subject(s)
Bone and Bones/injuries , Bone and Bones/metabolism , Endothelial Growth Factors/pharmacology , Femoral Fractures/physiopathology , Fracture Healing/drug effects , Lymphokines/pharmacology , Neovascularization, Physiologic/drug effects , Osteoblasts/physiology , Animals , Bone and Bones/blood supply , Cells, Cultured , Humans , Male , Mice , Mice, Inbred C57BL , Osteoblasts/drug effects , Rabbits , Radius/injuries , Tibia/diagnostic imaging , Tibia/injuries , Tomography, X-Ray Computed , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth FactorsABSTRACT
Several studies have demonstrated that the biological properties of secreted bacterial toxins could be harnessed for the induction of mucosal and systemic immunity following application at epithelial surfaces. Although the properties and potential application of several of these toxins will be discussed in this review, special focus will be placed on Pseudomonas aeruginosa exotoxin A (PE). A non-toxic form of PE (ntPE) into which antigenic epitopes can be integrated appears to be a particularly promising vaccination tool, which is able to cross the polarized epithelia of the gastrointestinal, respiratory and reproductive tracts and selectively target macrophages and dendritic cells.