RESUMEN
This manuscript is focussed on the development of pentablock (PB) copolymer based sustained release formulation for the treatment of posterior segment ocular diseases. We have successfully synthesised biodegradable and biocompatible PB copolymers for the preparation of nanoparticles (NPs) and thermosensitive gel. Achieving high drug loading with hydrophilic biotherapeutics (peptides/proteins) is a challenging task. Moreover, small intravitreal injection volume (≤100 µL) requires high loading to develop a long term (six months) sustained release formulation. We have successfully investigated various formulation parameters to achieve maximum peptide/protein (octreotide, insulin, lysozyme, IgG-Fab, IgG, and catalase) loading in PB NPs. Improvement in drug loading can facilitate delivery of larger doses of therapeutic proteins via limited injection volume. A composite formulation comprised of NPs in gel system exhibited sustained release (without burst effect) of peptides and proteins, may serve as a platform technology for the treatment of posterior segment ocular diseases.
Asunto(s)
Preparaciones de Acción Retardada/química , Oftalmopatías/tratamiento farmacológico , Nanopartículas/química , Péptidos/administración & dosificación , Polímeros/química , Proteínas/administración & dosificación , Animales , Catalasa/administración & dosificación , Pollos , Sistemas de Liberación de Medicamentos , Geles/química , Humanos , Inmunoglobulina G/administración & dosificación , Insulina/administración & dosificación , Muramidasa/administración & dosificación , Octreótido/administración & dosificaciónRESUMEN
IgG-Fab fragment, a model antibody protein was hydrophobically modified by a novel approach of ion-pairing complexation. Three different sulphated ion-pairing agents were utilised including sodium dodecyl sulphate, taurocholic acid and dextran sulphate (DS). The formations of hydrophobic ion-pairing (HIP) complexes were dependant on pH and molar ratio of ion-pairing agent to Fab. Aqueous solubilities of HIP complexes were very low compared to Fab alone. In particular, when dextran sulphate was added as ion-pairing agent, formed Fab:DS HIP complexes were least soluble in water. Further, nanoparticles (NPs) loaded with drug and Fab:DS HIP complex were prepared and characterised with respect to encapsulation efficiency and size. We observed significant improvement in encapsulation efficiency for Fab:DS HIP complex-loaded nanoparticles. This study demonstrates a novel approach of formulating antibody-loaded nanoparticles which can also be employed for delivery of large antibodies.
Asunto(s)
Portadores de Fármacos/química , Fragmentos Fab de Inmunoglobulinas/química , Inmunoglobulina G/química , Nanoestructuras/química , Sulfato de Dextran/química , Interacciones Hidrofóbicas e Hidrofílicas , Dodecil Sulfato de Sodio/química , Ácido Taurocólico/químicaRESUMEN
Polysorbate (PS) 20 and 80 are the most common surfactants in monoclonal antibody (mAb) drug product (DP) formulations. Residual host cell proteins (HCP) present at extremely low concentrations in DP formulations can maintain enough enzymatic activity to degrade PS surfactants. Over time, the hydrolysis of surfactant causes the accumulation of minimally soluble free fatty acids resulting in precipitation and formation of subvisible and visible particulates. This manuscript summarizes the investigation of a batch of high concentration (>100 mg/mL) mAb DP where subvisible particles formed abruptly after prolonged storage at 5C°. The work also summarizes the effectiveness of different strategies for managing host cell proteins and fatty acid particles. The concentration and fatty acid composition of polysorbates were found to be significant factors in particle development. Solubilizers and alternative surfactants were all shown to be effective means of preventing particle formation. Lipase inhibitors proved to be a simple means to identify the problem but are more difficult to utilize as a solution.
Asunto(s)
Anticuerpos Monoclonales , Polisorbatos , Química Farmacéutica , Ácidos Grasos , TensoactivosRESUMEN
Formulation development was performed with the live, attenuated, human neonatal rotavirus vaccine candidate (RV3-BB) with three main objectives to facilitate use in low- and middle- income countries including (1) a liquid, 2-8°C stable vaccine, (2) no necessity for pre-neutralization of gastric acid prior to oral administration of a small-volume dose, and (3) a low-cost vaccine dosage form. Implementation of a high-throughput RT-qPCR viral infectivity assay for RV3-BB, which correlated well with traditional FFA assays in terms of monitoring RV3-BB stability profiles, enabled more rapid and comprehensive formulation development studies. A wide variety of different classes and types of pharmaceutical excipients were screened for their ability to stabilize RV3-BB during exposure to elevated temperatures, freeze-thaw and agitation stresses. Sucrose (50-60% w/v), PEG-3350, and a solution pH of 7.8 were selected as promising stabilizers. Using a combination of an in vitro gastric digestion model (to mimic oral delivery conditions) and accelerated storage stability studies, several buffering agents (e.g., succinate, adipate and acetate at ~200 to 400 mM) were shown to protect RV3-BB under acidic conditions, and at the same time, minimize virus destabilization during storage. Several optimized RV3-BB candidate formulations were identified based on negligible viral infectivity losses during storage at 2-8°C and -20°C for up to 12 months, as well as by relative stability comparisons at 15°C and 25°C (up to 12 and 3 months, respectively). These RV3-BB stability results are discussed in the context of stability profiles of other rotavirus serotypes as well as future RV3-BB formulation development activities.
Asunto(s)
Infecciones por Rotavirus , Rotavirus , Anticuerpos Antivirales , Países en Desarrollo , Estabilidad de Medicamentos , Humanos , Recién Nacido , Rotavirus/genética , Infecciones por Rotavirus/prevención & control , Vacunas AtenuadasRESUMEN
In this work, two different in vitro gastric digestion models were used to evaluate the stability of a live attenuated rotavirus vaccine candidate (RV3-BB) under conditions designed to mimic oral delivery in infants. First, a forced-degradation model was established at low pH to assess the buffering capacity of formulation excipients and to screen for RV3-BB stabilizers. Second, a sequential-addition model was implemented to examine RV3-BB stability under conditions more representative of oral administration to infants. RV3-BB rapidly inactivated at < pH 5.0 (37 °C, 1 h) as measured by an infectivity RT-qPCR assay. Pre-neutralization with varying volumes of infant formula (Enfamil®) or antacid (Mylanta®) conferred partial to full protection of RV3-BB. Excipients with sufficient buffering capacity to minimize acidic pH inactivation of RV3-BB were identified (e.g., succinate, acetate, adipate), however, they concomitantly destabilized RV3-BB in accelerated storage stability studies. Both effects were concentration dependent, thus excipient optimization was required to design candidate RV3-BB formulations which minimize acid-induced viral inactivation during oral delivery while not destabilizing the vaccine during long-term 2-8 °C storage. Finally, a statistical Design -of-Experiments (DOE) study examining RV3-BB stability in the in vitro sequential-addition model identified key formulation parameters likely affecting RV3-BB stability during in vivo oral delivery.
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Infecciones por Rotavirus , Vacunas contra Rotavirus , Rotavirus , Anticuerpos Antivirales , Digestión , Humanos , Lactante , Infecciones por Rotavirus/prevención & control , Vacunas AtenuadasRESUMEN
A combination experimental approach, utilizing semi-empirical excipient screening followed by statistical modeling using design of experiments (DOE), was undertaken to identify stabilizing candidate formulations for a lyophilized live attenuated Flavivirus vaccine candidate. Various potential pharmaceutical compounds used in either marketed or investigative live attenuated viral vaccine formulations were first identified. The ability of additives from different categories of excipients, either alone or in combination, were then evaluated for their ability to stabilize virus against freeze-thaw, freeze-drying, and accelerated storage (25°C) stresses by measuring infectious virus titer. An exploratory data analysis and predictive DOE modeling approach was subsequently undertaken to gain a better understanding of the interplay between the key excipients and stability of virus as well as to determine which combinations were interacting to improve virus stability. The lead excipient combinations were identified and tested for stabilizing effects using a tetravalent mixture of viruses in accelerated and real time (2-8°C) stability studies. This work demonstrates the utility of combining semi-empirical excipient screening and DOE experimental design strategies in the formulation development of lyophilized live attenuated viral vaccine candidates.
Asunto(s)
Excipientes/química , Vacunas Atenuadas/química , Vacunas Virales/química , Flavivirus , LiofilizaciónRESUMEN
In this study, we investigated analytical challenges associated with the formulation of 2 anti-HIV broadly neutralizing antibodies (bnAbs), 3BNC117 and PGT121, both separately at 100 mg/mL and together at 50 mg/mL each. The bnAb formulations were characterized for relative solubility and conformational stability followed by accelerated and real-time stability studies. Although the bnAbs were stable during 4°C storage, incubation at 40°C differentiated their stability profiles. Specific concentration-dependent aggregation rates at 30°C and 40°C were measured by size exclusion chromatography for the individual bnAbs with the mixture showing intermediate behavior. Interestingly, although the relative ratio of the 2 bnAbs remained constant at 4°C, the ratio of 3BNC117 to PGT121 increased in the dimer that formed during storage at 40°C. A mass spectrometry-based multiattribute method, identified and quantified differences in modifications of the Fab regions for each bnAb within the mixture including clipping, oxidation, deamidation, and isomerization sites. Each bnAb showed slight differences in the levels and sites of lysine residue glycations. Together, these data demonstrate the ability to differentiate degradation products from individual antibodies within the bnAb mixture, and that degradation rates are influenced not only by the individual bnAb concentrations but also by the mixture concentration.
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Anticuerpos Neutralizantes/química , Anticuerpos Anti-VIH/química , Anticuerpos Monoclonales Humanizados , Anticuerpos Neutralizantes/inmunología , Anticuerpos ampliamente neutralizantes , Composición de Medicamentos , Anticuerpos Anti-VIH/inmunología , Infecciones por VIH/inmunología , VIH-1/inmunología , Humanos , Oxidación-Reducción , Agregado de Proteínas , Conformación Proteica , Estabilidad Proteica , SolubilidadRESUMEN
Discovery of insulin in the early 1900s initiated the research and development to improve the means of therapeutic protein delivery in patients. In the past decade, great emphasis has been placed on bringing protein and peptide therapeutics to market. Despite tremendous efforts, parenteral delivery still remains the major mode of administration for protein and peptide therapeutics. Other routes such as oral, nasal, pulmonary and buccal are considered more opportunistic rather than routine application. Improving biological half-life, stability and therapeutic efficacy is central to protein and peptide delivery. Several approaches have been tried in the past to improve protein and peptide in vitro/in vivo stability and performance. Approaches may be broadly categorized as chemical modification and colloidal delivery systems. In this review we have discussed various chemical approaches such as PEGylation, hyperglycosylation, mannosylation, and colloidal carriers including microparticles, nanoparticles, liposomes, carbon nanotubes and micelles for improving protein and peptide delivery. Recent developments on in situ thermosensitive gel-based protein and peptide delivery have also been described. This review summarizes recent developments on some currently existing approaches to improve stability, bioavailability and bioactivity of peptide and protein therapeutics following parenteral administration.
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Péptidos/administración & dosificación , Proteínas/administración & dosificación , Animales , Disponibilidad Biológica , Química Farmacéutica , Formas de Dosificación , Sistemas de Liberación de Medicamentos , Estabilidad de Medicamentos , Excipientes/química , Humanos , Infusiones Parenterales , Péptidos/química , Péptidos/farmacocinética , Proteínas/química , Proteínas/farmacocinética , Tecnología Farmacéutica/métodosRESUMEN
Proteins and peptides are widely indicated in many diseased states. Parenteral route is the most commonly em- ployed method of administration for therapeutic proteins and peptides. However, requirement of frequent injections due to short in vivo half-life results in poor patient compliance. Non-invasive drug delivery routes such as nasal, transdermal, pulmonary, and oral offer several advantages over parenteral administration. Intrinsic physicochemical properties and low permeability across biological membrane limit protein delivery via non-invasive routes. One of the strategies to improve protein and peptide absorption is by delivering through nanostructured delivery carriers. Among nanocarriers, polymeric nanoparticles (NPs) have demonstrated significant advantages over other delivery systems. This article summarizes the application of polymeric NPs for protein and peptide drug delivery following oral, nasal, pulmonary, parenteral, transder mal, and ocular administrations.
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Vías de Administración de Medicamentos , Sistemas de Liberación de Medicamentos , Nanopartículas , Péptidos , Polímeros , Proteínas , Animales , Humanos , Ratones , Nanopartículas/administración & dosificación , Nanopartículas/química , Péptidos/administración & dosificación , Péptidos/química , Polímeros/administración & dosificación , Polímeros/química , Proteínas/administración & dosificación , Proteínas/químicaRESUMEN
The major challenge faced by today's pharmacologist and formulation scientist is ocular drug delivery. Topical eye drop is the most convenient and patient compliant route of drug administration, especially for the treatment of anterior segment diseases. Delivery of drugs to the targeted ocular tissues is restricted by various precorneal, dynamic and static ocular barriers. Also, therapeutic drug levels are not maintained for longer duration in target tissues. In the past two decades, ocular drug delivery research acceleratedly advanced towards developing a novel, safe and patient compliant formulation and drug delivery devices/techniques, which may surpass these barriers and maintain drug levels in tissues. Anterior segment drug delivery advances are witnessed by modulation of conventional topical solutions with permeation and viscosity enhancers. Also, it includes development of conventional topical formulations such as suspensions, emulsions and ointments. Various nanoformulations have also been introduced for anterior segment ocular drug delivery. On the other hand, for posterior ocular delivery, research has been immensely focused towards development of drug releasing devices and nanoformulations for treating chronic vitreoretinal diseases. These novel devices and/or formulations may help to surpass ocular barriers and associated side effects with conventional topical drops. Also, these novel devices and/or formulations are easy to formulate, no/negligibly irritating, possess high precorneal residence time, sustain the drug release, and enhance ocular bioavailability of therapeutics. An update of current research advancement in ocular drug delivery necessitates and helps drug delivery scientists to modulate their think process and develop novel and safe drug delivery strategies. Current review intends to summarize the existing conventional formulations for ocular delivery and their advancements followed by current nanotechnology based formulation developments. Also, recent developments with other ocular drug delivery strategies employing in situ gels, implants, contact lens and microneedles have been discussed.
RESUMEN
The objective of this study is to delineate whether overexpression of human efflux transporters (P-gp, MRP2, and BCRP) in transfected MDCK cells affect the functional activities, and gene and protein expression of endogenous influx peptide transporter system (PepT). Real-time PCR, immunoblotting, uptake and permeability studies of [(3)H]Gly-Sar were conducted on transfected MDCKII and wild-type cells to investigate functional differences. Cellular [(3)H]Gly-Sar accumulation was significantly lower in transfected MDCKII cell lines compared to wild-type cells. Transport efficiency of apical peptide transporters was markedly reduced to around 25%, 30%, and 40% in P-gp-, MRP2-, and BCRP-overexpressed MDCK cell lines, respectively. With ascending cell-passage, transport efficiency was enhanced. A significantly higher Gly-Sar permeability was observed across parental cell-monolayers over transfected cells at all pHs. Levels of mRNA for both canine PepT1 and PepT2 were substantially reduced when efflux transporters overexpressed but enhanced when mRNA-levels of efflux genes diminished with ascending cell-passage of transfected cells. An inverse correlation was evident between endogenous PepT and exogenous efflux transporters in transfected MDCKII cells. Results of protein expression also supported these findings. Overexpression of MDR genes can affect endogenous PepT function which might be due to the phenomenon of transporter-compensation resulting in down-regulation of endogenous genes.
Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Transportadoras de Casetes de Unión a ATP/genética , Proteínas de Transporte de Membrana/genética , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Proteínas de Neoplasias/genética , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Animales , Transporte Biológico , Western Blotting , Dipéptidos/metabolismo , Perros , Regulación hacia Abajo , Expresión Génica , Humanos , Concentración de Iones de Hidrógeno , Células de Riñón Canino Madin Darby , Proteína 2 Asociada a Resistencia a Múltiples Medicamentos , Transportador de Péptidos 1 , Reacción en Cadena en Tiempo Real de la Polimerasa , Simportadores/genética , TransfecciónRESUMEN
Oral delivery of peptide and protein drugs faces immense challenge partially due to the gastrointestinal (GI) environment. In spite of considerable efforts by industrial and academic laboratories, no major breakthrough in the effective oral delivery of polypeptides and proteins has been accomplished. Upon oral administration, gastrointestinal epithelium acts as a physical and biochemical barrier for absorption of proteins resulting in low bioavailability (typically less than 1-2%). An ideal oral drug delivery system should be capable of (a) maintaining the integrity of protein molecules until it reaches the site of absorption, (b) releasing the drug at the target absorption site, where the delivery system appends to that site by virtue of specific interaction, and (c) retaining inside the gastrointestinal tract irrespective of its transitory constraints. Various technologies have been explored to overcome the problems associated with the oral delivery of macromolecules such as insulin, gonadotropin-releasing hormones, calcitonin, human growth factor, vaccines, enkephalins, and interferons, all of which met with limited success. This review article intends to summarize the physiological barriers to oral delivery of peptides and proteins and novel pharmaceutical approaches to circumvent these barriers and enhance oral bioavailability of these macromolecules.
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Péptidos/farmacocinética , Proteínas/farmacocinética , Administración Oral , Animales , Disponibilidad Biológica , Química Farmacéutica , Humanos , Absorción Intestinal , Péptidos/administración & dosificación , Péptidos/química , Proteínas/administración & dosificación , Proteínas/químicaRESUMEN
One of the most challenging areas of pharmaceutical research is ocular drug delivery. The unique anatomy and physiology of the eye impedes drug permeation to deeper ocular tissues. Nanosized carrier systems such as nanoparticles, liposomes, suspensions, dendrimers, and nanomicelles are being explored for ocular drug delivery. In this review, we have focused on application of emerging nanomicellar carrier systems in ocular drug delivery. Nanomicelles are nanosized vesicular carriers formed from amphiphilic monomer units. Surfactant and polymeric micellar nanocarriers provide an amenable means to improve drug solubilization, develop clear aqueous formulations and deliver drugs to anterior and posterior ocular tissues. Nanomicelles due to their amphiphilic nature encapsulate hydrophobic drugs and aid in drug delivery. Various methods are employed to develop nanosized micellar formulations depending upon the physicochemical properties of the drug. Nanomicellar carriers appear to be promising vehicles with potential applications in ocular drug delivery. In this review, we attempted to discuss about the progress in ocular drug delivery research using nanomicelles as carriers from the published literature and issued patents. Also, with regards to ocular static and dynamic barriers which prevent drug permeation, a brief discussion about nanomicelles, types of nanomicelles, their methods of preparation and micellar strategy to overcome ocular barriers, delivering therapeutic levels of drugs to anterior and posterior ocular tissues are discussed.
RESUMEN
Advancements in the field and rising interest among pharmaceutical researchers have led to the development of new molecules with enhanced therapeutic activity. Design of new drugs which can target a particular pathway and/or explore novel targets is of immense interest to ocular pharmacologists worldwide. Delivery of suitable pharmacologically active agents at proper dose (within the therapeutic window) to the target tissues without any toxicity to the healthy ocular tissues still remain an elusive task. Moreover, the presence of static and dynamic barriers to drug absorption including the corneal epithelium (lipophilic), corneal and scleral stroma (hydrophilic), conjunctival lymphatics, choroidal vasculature and the blood-ocular barriers also pose a significant challenge for achieving therapeutic drug concentrations at the target site. Although many agents are currently available, new compounds are being introduced for treating various ocular diseases. Deeper understanding of the etiology and complex mechanisms associated with the disease condition would aid in the development of potential therapeutic candidates. Novel small molecules as well as complex biotechnology derived macromolecules with superior efficacy, safety and tolerability are being developed. Therefore, this review article provides an overview of existing drugs, treatment options, advances in emerging therapeutics and related recent patents for the treatment of ocular disorders such as glaucoma, age related macular degeneration (AMD) and uveitis.