RESUMEN
The ability to deliver stable and active dried protein therapeutics from biopharmaceutical drug delivery systems is critical for solid dosage formulation development. Spray dried formulations with carefully selected excipients provide a unique opportunity in amorphous phase stabilization of the therapeutic proteins. Herein, we discuss the role of hydroxypropyl methylcellulose acetate succinate (HPMCAS) derivatives as polymeric excipients for stabilizing a model fragment antibody (Fab2) during high temperature processing and in possible low pH environments of a drug delivery platform. The effects of high temperature processing and microenvironmental pH sensitivity are of particular interest to us due to their adverse impact on stability of molecules that demonstrate temperature and pH dependent inactivation within drug delivery devices. It appears in solid state at 90 °C and 37 °C and within low pH micro-environment HPMCAS protects protein against aggregation. The high temperature performance of HPMCAS is comparable to that of a disaccharide excipient like trehalose in spray dried protein powder. Simultaneously, inside a poly(lactic-co-glycolic acid) (PLGA) based delivery system HPMCAS provides protection to a pH sensitive protein against acidic degradation products from aqueous hydrolysis of PLGA.
Asunto(s)
Excipientes , Metilcelulosa , Anticuerpos , Estabilidad de Medicamentos , Excipientes/química , Concentración de Iones de Hidrógeno , Derivados de la Hipromelosa , Metilcelulosa/química , TemperaturaRESUMEN
An extensive data set has been developed and used to further the progress of a model-informed design of controlled drug release. An improved drug-release model with mechanistic modeling of hydrolytic polymer degradation is used and validated by comparing model predictions to in vitro experiments. Combining parameter estimates from the literature with model fits to the data set, this study can aid in achieving a priori design of controlled drug release from a model PLGA release system. A systematic series of model release systems were formulated with FITC-labeled dextran, as a surrogate for biopharmaceuticals, in PLGA rods over a broad range of compositions. While general comparisons between the model and experiments were favorable, important discrepancies were identified for several formulations with significant first-phase drug release. Supported by cross-sectional fluorescence microscopy images of the FITC-dextran distribution within the rods, this first-phase release was attributed to a combination of two main factors: (1) percolation of the drug particles and (2) swelling of and pore formation in the rods due to water uptake. These observations indicate the importance of careful selection of the PLGA polymer grade when designing drug release systems but also reflect a need for better understanding of phenomena such as pore formation. Adapting model parameters, without modifying the physical processes included in the model, enabled accurate fitting of the experimental data for all formulations, highlighting the applicability of the model.
Asunto(s)
Ácido Láctico , Ácido Poliglicólico , Estudios Transversales , Preparaciones de Acción Retardada , Sistemas de Liberación de Medicamentos/métodos , Liberación de Fármacos , Microesferas , Tamaño de la Partícula , Copolímero de Ácido Poliláctico-Ácido PoliglicólicoRESUMEN
Long-acting implants are typically formulated using carrier(s) with specific physical and chemical properties, along with the active pharmaceutical ingredient (API), to achieve the desired daily exposure for the target duration of action. In characterizing such formulations, real-time in-vitro and in-vivo experiments that are typically used to characterize implants are lengthy, costly, and labor intensive as these implants are designed to be long acting. A novel characterization technique, combining high resolution three-dimensional X-Ray microscopy imaging, image-based quantification, and transport simulation, has been employed to provide a mechanistic understanding of formulation and process impact on the microstructures and performance of a polymer-based implant. Artificial intelligence-based image segmentation and image data analytics were used to convert morphological features visualized at high resolution into numerical microstructure models. These digital models were then used to calculate key physical parameters governing drug transport in a polymer matrix, including API uniformity, API domain size, and permeability. This powerful new tool has the potential to advance the mechanistic understanding of the interplay between drug-microstructure and performance and accelerate the therapeutic development long-acting implants.
Asunto(s)
Inteligencia Artificial , Polímeros , Liberación de Fármacos , Microscopía , Rayos XRESUMEN
The port delivery system with ranibizumab (PDS) is an investigational long-acting drug delivery system for the continuous release of ranibizumab, an anti-VEGF biologic, in the vitreous humor. The efficacy of the PDS implant relies on the maintenance of long-term drug stability under physiological conditions. Herein, the long-term stability of three anti-VEGF biologics - ranibizumab, bevacizumab and aflibercept - was investigated in phosphate buffered saline (PBS) at 37 °C for several months. Comparison of stability profiles shows that bevacizumab and aflibercept are increasingly prone to aggregation whereas ranibizumab undergoes minimal aggregation. Ranibizumab also shows the smallest loss in antigen binding capacity after long-term incubation in PBS. Even though the aggregated forms of bevacizumab and aflibercept bind to VEGF, the consequences of aggregation on immunogenicity, implant function and efficacy are unknown. These results highlight the importance of maintaining long-term drug stability under physiologically relevant conditions which is necessary for achieving efficacy with an in vivo continuous drug delivery device such as the PDS implant.
Asunto(s)
Productos Biológicos , Factor A de Crecimiento Endotelial Vascular , Inhibidores de la Angiogénesis , Bevacizumab , Inyecciones Intravítreas , Ranibizumab , Proteínas Recombinantes de FusiónRESUMEN
Maintenance of protein stability during manufacture, storage, and delivery is necessary for the successful development of a drug product. Herein, the utility of two compatible solutes-ectoine and hydroxyectoine-in stabilizing a model protein labeled Fab2 has been investigated. Specifically, the performance of ectoine and hydroxyectoine in stabilizing Fab2 in a spray-dried formulation at elevated temperature and after multiple freeze/thaw cycles has been compared with the performance of a formulation containing trehalose and a formulation containing no excipient as controls. In the solid state at 90 and 37 °C and in freeze concentrate systems, ectoine and hydroxyectoine suppress protein aggregation. Like trehalose, hydroxyectoine also limits N-terminal pyroglutamate formation in Fab2 in the solid state. The extent of protein stabilization is dependent on the excipient concentration in the formulation, but at a 1:1 excipient to protein mass ratio, hydroxyectoine is better than trehalose in stabilizing Fab2. The results presented here suggest that ectoine and hydroxyectoine are effective excipients for stabilizing therapeutic antibodies.
Asunto(s)
Aminoácidos Diaminos/química , Anticuerpos/química , Química Farmacéutica/métodos , Estabilidad de Medicamentos , Excipientes/química , Liofilización/métodos , Congelación , Estabilidad Proteica/efectos de los fármacos , Proteínas/química , Temperatura , Trehalosa/químicaRESUMEN
PURPOSE: To evaluate the stability of a model Mab1 and Fab1 under in vitro vitreal conditions in the presence of various metabolites and in the presence of light at λ > 400 nM. METHODS: A full length IgG1 monoclonal antibody (Mab1) and a fab fragment (Fab1) were formulated with various metabolites typically found in the vitreous humor and subjected to visible light treatment. Both proteins were analyzed using a variety of biochemical techniques. Furthermore, Fab1 was also tested for antigen binding ability using surface plasmon resonance. RESULTS: Our data shows that some vitreal metabolites such as riboflavin and ascorbic acid affect protein stability, via formation of reactive oxygen species (ROS) and advanced glycation end products (AGE) s respectively. In contrast, metabolites such as glutathione may protect these proteins from light-induced degradation to some extent. CONCLUSIONS: Ascorbic acid and riboflavin were found to photosensitize therapeutic proteins especially when exposed to light. Ascorbic acid reacted with proteins even in the absence of light. Antioxidants such as glutathione helped limit photooxidation under ambient or blue light exposure. Since antioxidant capacity in the eye decreases with age we recommend understanding long term stability, including photooxidation and photosensitization, of new candidate proteins in the context of controlled or sustained release technologies for ocular diseases.
Asunto(s)
Anticuerpos Monoclonales/efectos de la radiación , Oftalmopatías/metabolismo , Fragmentos Fab de Inmunoglobulinas/efectos de la radiación , Inmunoglobulina G/metabolismo , Antioxidantes/farmacología , Ácido Ascórbico , Luz , Estabilidad Proteica/efectos de la radiación , Especies Reactivas de Oxígeno/metabolismo , RiboflavinaRESUMEN
Treatment of ocular diseases associated with neovascularization currently requires frequent intravitreal injections of antivascular endothelial growth factor (anti-VEGF) therapies. Reducing the required frequency of anti-VEGF injections and associated clinical visits may improve patient adherence to the prescribed treatment regimen and improve outcomes. Herein, we explore conjugation of rabbit and fragment antibodies (Fab) to the biopolymer hyaluronic acid (HA) as a half-life modifying strategy, and assess the impact on Fab biophysical properties and vitreal pharmacokinetics. HA-Fab conjugates of three distinct molecular weights and hydrodynamic radii (RH) were assessed for in vivo pharmacokinetic performance relative to unconjugated Fab after intravitreal injection in rabbits. Covalent conjugation to HA did not significantly alter the thermal stability or secondary or tertiary structure, or diminish the potency of the Fab, thereby preserving its pharmacological properties. Conjugation to HA did significantly slow the in vivo clearance of Fab from the rabbit vitreous in an RH-dependent manner. Compared to free Fab (observed vitreal half-life of 2.8 days), HA-Fab conjugates cleared with observed half-lives of 7.6, 10.2, and 18.3 days for 40 kDa, 200 kDa, and 600 kDa HA conjugates, respectively. This work elucidates a possible strategy for long-acting delivery of proteins intended for the treatment of chronic posterior ocular diseases.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacocinética , Ácido Hialurónico/inmunología , Fragmentos Fab de Inmunoglobulinas/inmunología , Cuerpo Vítreo/metabolismo , Animales , Anticuerpos Monoclonales/administración & dosificación , Humanos , Inyecciones Intravítreas , Conejos , Distribución Tisular , Cuerpo Vítreo/inmunologíaRESUMEN
Mathematical modeling of drug release can aid in the design and development of sustained delivery systems, but the parameter estimation of such models is challenging owing to the nonlinear mathematical structure and complexity and interdependency of the physical processes considered. Highly parameterized models often lead to overfitting, strong parameter correlations, and as a consequence, inaccurate model predictions for systems not explicitly part of the fitting database. Here, we show that an efficient stochastic optimization algorithm can be used not only to find robust estimates of global minima to such complex problems but also to generate metadata that allow quantitative evaluation of parameter sensitivity and correlation, which can be used for further model refinement and development. A practical methodology is described through the analysis of a predictive drug release model on published experimental data sets. The model is then used to design a zeroth-order release profile in an experimental system consisting of an antibody fragment in a poly(lactic-co-glycolic acid) solvent depot, which is validated experimentally. This approach allows rational decision-making when developing new models, selecting models for a specific application, or designing formulations for experimental trials.
Asunto(s)
Preparaciones de Acción Retardada/química , Preparaciones Farmacéuticas/química , Sistemas de Liberación de Medicamentos/métodos , Liberación de Fármacos/efectos de los fármacos , Modelos Teóricos , Copolímero de Ácido Poliláctico-Ácido Poliglicólico/química , Solventes/químicaRESUMEN
The preparation of PLGA rods for sustained release applications via a hot-melt extrusion process employs heat and mechanical shear. Understanding protein stability and degradation mechanisms at high temperature in the solid state is therefore important for the preparation of protein-loaded PLGA rods. The stability of a model protein, labeled Fab2, has been investigated in solid-state formulations containing trehalose at elevated temperatures. Spray-dried formulations containing varying levels of trehalose were exposed to temperatures ranging from 90 to 120 °C. Measurement of aggregation and chemical degradation rates suggests that trehalose limits Fab2 degradation in a concentration-dependent manner, but the effect tends to saturate when the mass ratio of trehalose to protein is around 1 in the solid formulation. The Fab2 secondary structure and spray-dried particle morphology were studied using circular dichroism and scanning electron microscopy techniques, respectively. On the basis of temperature and trehalose-dependent aggregation kinetics as well as changes in spray-dried particle morphology, a mechanism is proposed for the trehalose stabilization of proteins in solid state at elevated temperatures. The results reported here suggest that when fragment antibodies in the solid state are formulated with trehalose as excipient, a high temperature process such as hot-melt extrusion can be successfully accomplished with minimal degradation.
Asunto(s)
Anticuerpos/química , Excipientes/química , Trehalosa/química , Estabilidad de Medicamentos , Copolímero de Ácido Poliláctico-Ácido Poliglicólico/química , TemperaturaRESUMEN
Lyophilized and spray-dried biopharmaceutical formulations are used to provide long-term stability for storage and transport, but questions remain about the molecular structure in these solid formulations and how this structure may be responsible for protein stability. Small-angle neutron scattering with a humidity control environment is used to characterize protein-scale microstructural changes in such solid-state formulations as they are humidified and dried in situ. The findings indicate that irreversible protein aggregates of stressed formulations do not form within the solid-state but do emerge upon reconstitution of the formulation. After plasticization of the solid-state matrix by exposure to humidity, the formation of reversibly self-associating aggregates can be detected in situ. The characterization of the protein-scale microstructure in these solid-state formulations facilitates further efforts to understand the underlying mechanisms that promote long-term protein stability.
Asunto(s)
Anticuerpos Monoclonales/química , Estabilidad de Medicamentos , Liofilización , Estabilidad ProteicaRESUMEN
To date, ocular antibody therapies for the treatment of retinal diseases rely on injection of the drug into the vitreous chamber of the eye. Given the burden for patients undergoing this procedure, less frequent dosing through the use of long-acting delivery (LAD) technologies is highly desirable. These technologies usually require a highly concentrated formulation and the antibody must be stable against extended exposure to physiological conditions. Here we have increased the potential of a therapeutic antibody antigen-binding fragment (Fab) for LAD by using protein engineering to enhance the chemical and physical stability of the molecule. Structure-guided amino acid substitutions in a negatively charged complementarity determining region (CDR-L1) of an anti-factor D (AFD) Fab resulted in increased chemical stability and solubility. A variant of AFD (AFD.v8), which combines light chain substitutions (VL-D28S:D30E:D31S) with a substitution (VH-D61E) to stabilize a heavy chain isomerization site, retained complement factor D binding and inhibition potency and has properties suitable for LAD. This variant was amenable to high protein concentration (>250 mg/mL), low ionic strength formulation suitable for intravitreal injection. AFD.v8 had acceptable pharmacokinetic (PK) properties upon intravitreal injection in rabbits, and improved stability under both formulation and physiological conditions. Simulations of expected human PK behavior indicated greater exposure with a 25-mg dose enabled by the increased solubility of AFD.v8.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Fragmentos Fab de Inmunoglobulinas/inmunología , Ingeniería de Proteínas/métodos , Enfermedades de la Retina/inmunología , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Animales , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/farmacocinética , Afinidad de Anticuerpos/inmunología , Factor D del Complemento/inmunología , Regiones Determinantes de Complementariedad/genética , Regiones Determinantes de Complementariedad/inmunología , Sistemas de Liberación de Medicamentos , Estabilidad de Medicamentos , Humanos , Fragmentos Fab de Inmunoglobulinas/química , Fragmentos Fab de Inmunoglobulinas/genética , Modelos Moleculares , Conformación Proteica , Conejos , Enfermedades de la Retina/tratamiento farmacológico , Enfermedades de la Retina/metabolismoRESUMEN
Chemically cross-linked hydrogels are promising systems for protein delivery applications, but their utility may be limited due to the possibility of protein reaction with hydrogel precursors. Herein, a catalyst-free inverse-demand Diels-Alder reaction between tetrazine and norbornene groups was used to demonstrate the bio-orthogonal nature of cross-linking chemistry that is chemically inert to proteins. Tetrazine-modified hyaluronic acid and norbornene-modified polyethylene glycol were used as hydrogel precursors for in situ encapsulation of a model protein, Fab1. Measurement of gelation kinetics demonstrates that network formation and gel stiffness are temperature-dependent but independent of Fab1 concentration. In vitro release testing shows that Fab1 is completely released from the hydrogel matrix over a period of several weeks. Analytical characterization suggests that Fab1 is released without any physical or chemical modifications and retains its antigen binding capacity. Thus, the bio-orthogonal and catalyst-free aqueous phase chemistry enables efficient in situ protein encapsulation in a single step and provides sustained protein release.
Asunto(s)
Ácido Hialurónico/química , Hidrogeles/química , Proteínas/química , Química Clic/métodos , Preparaciones de Acción Retardada , Cinética , Estabilidad ProteicaRESUMEN
Filamentous viruses are common in nature and efficiently deliver - sometimes via aerosol - genetic material, viral proteins, and other factors to animals and plants. Aerosolization can be a severe physicochemical test of the stability of any filamentous assembly whether it is made from natural polymers such as viral proteins or synthetic polymers. Here, worm-like "filomicelles" that self-assemble in water from amphiphilic block copolymers were investigated as aerosolized delivery vehicles. After spraying and drying, fluorophore-loaded filomicelles that were originally ~10-20µm long could be imaged as 2-5µm long fragments that survived rehydration on natural and artificial surfaces (i.e. plant leaves and glass). As a functional test of delivery, the hydrophobic pesticide bifenthrin was loaded into filomicelles (up to 25% w/w) and sprayed onto plants infested with two agricultural pests, beet army worm or two-spotted spider mites; pesticidal efficacy exceeded that of commercial formulations. Persistent delivery by the filomicelle formulation was especially notable and broadly consistent with previous intravenous delivery of other drugs and dyes with the highly elongated filomicelles.
Asunto(s)
Portadores de Fármacos , Micelas , Aerosoles , Animales , Portadores de Fármacos/administración & dosificación , Portadores de Fármacos/química , Insecticidas/administración & dosificación , Insecticidas/química , Larva/efectos de los fármacos , Mariposas Nocturnas , Polímeros/administración & dosificación , Polímeros/química , Piretrinas/administración & dosificación , Piretrinas/química , TetranychidaeRESUMEN
Protein-stabilizer microheterogeneity is believed to influence long-term protein stability in solid-state biopharmaceutical formulations and its characterization is therefore essential for the rational design of stable formulations. However, the spatial distribution of the protein and the stabilizer in a solid-state formulation is, in general, difficult to characterize because of the lack of a functional, simple, and reliable characterization technique. We demonstrate the use of confocal fluorescence microscopy with fluorescently labeled monoclonal antibodies (mAbs) and antibody fragments (Fabs) to directly visualize three-dimensional particle morphologies and protein distributions in dried biopharmaceutical formulations, without restrictions on processing conditions or the need for extensive data analysis. While industrially relevant lyophilization procedures of a model IgG1 mAb generally lead to uniform protein-excipient distribution, the method shows that specific spray-drying conditions lead to distinct protein-excipient segregation. Therefore, this method can enable more definitive optimization of formulation conditions than has previously been possible.
Asunto(s)
Excipientes/química , Proteínas/química , Anticuerpos Monoclonales/química , Biofarmacia/métodos , Química Farmacéutica/métodos , Estabilidad de Medicamentos , Liofilización/métodos , Inmunoglobulina G/química , Microscopía Confocal/métodos , Microscopía Fluorescente/métodos , Estabilidad ProteicaRESUMEN
Achieving long-term drug release from polymer-based delivery systems continues to be a challenge particularly for the delivery of large hydrophilic molecules such as therapeutic antibodies and proteins. Here, we report on the utility of an in situ-forming and injectable polymer-solvent system for the long-term release of a model antibody fragment (Fab1). The delivery system was prepared by dispersing a spray-dried powder of Fab1 within poly(lactide-co-glycolide) (PLGA)-triacetin solution. The formulation viscosity was within the range 1.0 ± 0.3 Pa s but it was injectable through a 27G needle. The release profile of Fab1, measured in phosphate-buffered saline (PBS), showed a lag phase followed by sustained-release phase for close to 80 days. Antibody degradation during its residence within the depot was comparable to its degradation upon long-term incubation in PBS. On the basis of temporal changes in surface morphology, stiffness, and depot mass, a mechanism to account for the drug release profile has been proposed. The unprecedented release profile and retention of greater than 80% of antigen-binding capacity even after several weeks demonstrates that PLGA-triacetin solution could be a promising system for the long-term delivery of biologics.
Asunto(s)
Fragmentos de Inmunoglobulinas/química , Triacetina/química , Química Farmacéutica , Preparaciones de Acción Retardada , Desecación , Portadores de Fármacos , Sistemas de Liberación de Medicamentos , Estabilidad de Medicamentos , Concentración de Iones de Hidrógeno , Fragmentos Fab de Inmunoglobulinas/administración & dosificación , Fragmentos Fab de Inmunoglobulinas/química , Fragmentos de Inmunoglobulinas/administración & dosificación , Ácido Láctico/química , Tamaño de la Partícula , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Propiedades de Superficie , Triacetina/administración & dosificación , Viscosidad , Agua/análisisRESUMEN
Polymer implants are promising systems for sustained release applications but their utility for protein delivery has been hindered because of concerns over drug stability at elevated temperatures required for processing. Using bovine serum albumin (BSA) as a model, we have assessed whether proteins can be formulated for processing at elevated temperatures. Specifically, the effect of trehalose and histidine-HCl buffer on BSA stability in a spray-dried formulation has been investigated at temperatures ranging from 80°C to 110°C. When both the sugar and buffer are present, aggregation is suppressed even when exposed to 100°C, the extrusion temperature of poly(lactide-co-glycolide) (PLGA), a bioresorbable polymer. Estimation of aggregation rate constants (k) indicate that though both trehalose and histidine-HCl buffer contribute to BSA stability, the effect because of trehalose alone is more pronounced. BSA-loaded PLGA implants were prepared using hot-melt extrusion process and in vitro release was conducted in phosphate buffered saline at 37°C. Comparison of drug released from implants prepared using four different formulations confirmed that maximal release was achieved from the formulation in which BSA was least aggregated. These studies demonstrate that when trehalose and histidine-HCl buffer are included in spray-dried formulations, BSA stability is maintained both during processing at 100°C and long-term residence within implants.
Asunto(s)
Preparaciones de Acción Retardada/química , Ácido Láctico/química , Ácido Poliglicólico/química , Albúmina Sérica Bovina/administración & dosificación , Trehalosa/química , Implantes Absorbibles , Animales , Bovinos , Desecación , Calor , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Estabilidad Proteica , Albúmina Sérica Bovina/químicaRESUMEN
Nanoparticles that are made by self-assembly into non-spherical shapes are promising as drug delivery vehicles. This review focuses on flexible and fragmentable filamentous micelles referred to as filomicelles made of degradable block copolymer amphiphiles. They are inspired by filoviruses and also by tubular proplatelets that break up into smaller platelets in blood flow. The synthesis and assembly of the constituent block copolymers are described together with ligand targeting and fragmentation as well as drug release in therapeutic applications to model tumors and most recently brain tumors.
RESUMEN
Tissue cells lack the ability to see or hear but have evolved mechanisms to feel into their surroundings and sense a collective stiffness. A cell can even sense the effective stiffness of rigid objects that are not in direct cellular contact - like the proverbial princess who feels a pea placed beneath soft mattresses. How deeply a cell feels into a matrix can be measured by assessing cell responses on a controlled series of thin and elastic gels that are affixed to a rigid substrate. Gel elasticity E is readily varied with polymer concentrations of now-standard polyacrylamide hydrogels, but to eliminate wrinkling and detachment of thin gels from an underlying glass coverslip, vinyl groups are bonded to the glass before polymerization. Gel thickness is nominally specified using micron-scale beads that act as spacers, but gels swell after polymerization as measured by z-section, confocal microscopy of fluorescent gels. Atomic force microscopy (AFM) is used to measure E at gel surfaces, employing stresses and strains that are typically generated by cells and yielding values for E that span a broad range of tissue microenvironments. To illustrate cell sensitivities to a series of thin-to-thick gels, the adhesive spreading of mesenchymal stem cells was measured on gel mimics of a very soft tissue (eg. brain, E ~ 1 kPa). Initial results show that cells increasingly respond to the rigidity of an underlying 'hidden' surface starting at about 10-20 microm gel thickness with a characteristic tactile length of less than about 5 microm.
Asunto(s)
Resinas Acrílicas/química , Materiales Biomiméticos/química , Adhesión Celular/fisiología , Adhesiones Focales/fisiología , Mecanotransducción Celular/fisiología , Células Madre Mesenquimatosas/fisiología , Resistencia al Corte/fisiología , Animales , Módulo de Elasticidad , Ratones , Estrés Mecánico , Propiedades de Superficie , ViscosidadRESUMEN
Crystallization processes are in general sensitive to detailed conditions, but our present understanding of underlying mechanisms is insufficient. A crystallizable chain within a diblock copolymer assembly is expected to couple curvature to crystallization and thereby impact rigidity as well as preferred morphology, but the effects on dispersed phases have remained unclear. The hydrophobic polymer polycaprolactone (PCL) is semi-crystalline in bulk (T(m) = 60°C) and is shown here to generate flexible worm micelles or rigid vesicles in water from several dozen polyethyleneoxide-based diblocks (PEO-PCL). Despite the fact that `worms' have a mean curvature between that of vesicles and spherical micelles, `worms' are seen only within a narrow, process-dependent wedge of morphological phase space that is deep within the vesicle phase. Fluorescence imaging shows worms are predominantly in one of two states - either entirely flexible with dynamic thermal undulations or fully rigid; only a few worms appear rigid at room temperature (T << T(m)), indicating suppression of crystallization by both curvature and PCL hydration. Worm rigidification, which depends on molecular weight, is also prevented by copolymerization of caprolactone with just 10% racemic lactide that otherwise has little impact on bulk crystallinity. In contrast to worms, vesicles of PEO-PCL are always rigid and typically leaky. Defects between crystallite domains induce dislocation-roughening with focal leakiness although select PEO-PCL - which classical surfactant arguments would predict make worms - yield vesicles that retain encapsulant and appear smooth, suggesting a gel or glassy state. Hydration in dispersion thus tends to selectively soften high curvature microphases.
RESUMEN
Selective binding of multivalent ligands within a mixture of polyvalent amphiphiles provides, in principle, a simple mechanism for driving domain formation in self-assemblies. Divalent cations are shown here to crossbridge polyanionic amphiphiles, which thereby demix from neutral amphiphiles and form spots or rafts within vesicles as well as stripes within cylindrical micelles. Calcium- and copper-crossbridged domains of synthetic block copolymers or natural lipid (phosphatidylinositol-4,5-bisphosphate) possess tunable sizes, shapes and/or spacings that can last for years. Lateral segregation in these 'ligand-responsive Janus assemblies' couples weakly to curvature and proves to be restricted within phase diagrams to narrow regimes of pH and cation concentration that are centred near the characteristic binding constants for polyacid interactions. Remixing at high pH is surprising, but a theory for strong lateral segregation shows that counterion entropy dominates electrostatic crossbridges, thus illustrating the insights gained into ligand-induced pattern formation within self-assemblies.