Development of a Stable Respiratory Syncytial Virus Pre-Fusion Protein Powder Suitable for a Core-Shell Implant with a Delayed Release in Mice: A Proof of Concept Study.

Currently, there is an increasing interest to apply pre-fusion (pre-F) protein of respiratory syncytial virus (RSV) as antigen for the development of a subunit vaccine. A pre-F-containing powder would increase the flexibility regarding the route of administration. For instance, a pre-F-containing powder could be incorporated into a single-injection system releasing a primer, and after a lag time, a booster. The most challenging aspect, obtaining the booster after a lag time, may be achieved by incorporating the powder into a core encapsulated by a nonporous poly(dl-lactic-co-glycolic acid) (PLGA) shell. We intended to develop a stable freeze-dried pre-F-containing powder. Furthermore, we investigated whether incorporation of this powder into the core-shell implant was feasible and whether this system would induce a delayed RSV virus-neutralizing antibody (VNA) response in mice. The developed pre-F-containing powder, consisting of pre-F in a matrix of inulin, HEPES, sodium chloride, and Tween 80, was stable during freeze-drying and storage for at least 28 days at 60 °C. Incorporation of this powder into the core-shell implant was feasible and the core-shell production process did not affect the stability of pre-F. An in vitro release study showed that pre-F was incompletely released from the core-shell implant after a lag time of 4 weeks. The incomplete release may be the result of pre-F instability within the core-shell implant during the lag time and requires further research. Mice subcutaneously immunized with a pre-F-containing core-shell implant showed a delayed RSV VNA response that corresponded with pre-F release from the core-shell implant after a lag time of approximately 4 weeks. Moreover, pre-F-containing core-shell implants were able to boost RSV VNA titers of primed mice after a lag time of 4 weeks. These findings could contribute to the development of a single-injection pre-F-based vaccine containing a primer and a booster.

Towards the Oral Treatment of Ileo-Colonic Inflammatory Bowel Disease with Infliximab Tablets: Development and Validation of the Production Process.

Infliximab (IFX) is an intravenously administered monoclonal antibody antagonizing the effects of tumor necrosis factor-alpha (TNF) systemically and is efficacious in the treatment of inflammatory bowel disease (IBD). However, studies suggest that the anti-inflammatory effects result from local immunomodulation in the inflamed regions. Furthermore, topical inhibition of TNF in IBD ameliorates inflammation. We therefore hypothesized that orally administered IFX targeted to the ileo-colonic region in IBD may be an efficacious new treatment option. This study describes the development and validation of the production process of ileo-colonic-targeted 5 mg IFX tablets (ColoPulse-IFX) intended for the oral treatment of IBD by means of producing three consecutive validation batches (VAL1, VAL2, and VAL3, respectively). UV-VIS spectroscopy, HPLC-SEC analysis (content, fragments, aggregates), fluorescence spectroscopy (tertiary protein structure), and ELISA (potency) showed no noticeable deviations of IFX compounded to ColoPulse-IFX compared to fresh IFX stock. The average ± SD (n = 10) IFX content of VAL1, VAL2, and VAL3 was 96 ± 2%, 97 ± 3%, and 96 ± 2%, respectively, and complied with the European Pharmacopeia (Ph. Eur.) requirements for Content Uniformity. The average ± SD (n = 3) ColoPulse-IFX potency was 105 ± 4%, 96 ± 4%, and 97 ± 5%, respectively, compared to fresh IFX stock. The IFX release profile from the tablet core was complete (≥85%) after 10 min in simulated ileum medium. The in vitro coating performance of ColoPulse-IFX showed that the formulation was targeted to the simulated ileo-colonic region. Stability data showed that ColoPulse-IFX was stable for up to 6 months stored at 25 °C/60% RH. Based on these results, the production process can be considered validated and its application is discussed in light of the rationale and available evidence for the topical treatment of IBD with IFX.

Respiratory syncytial virus subunit vaccines based on the viral envelope glycoproteins intended for pregnant women and the elderly.

Introduction: Respiratory syncytial virus (RSV) causes high morbidity and mortality rates among infants, young children, and the elderly worldwide. Unfortunately, a safe and effective vaccine is still unavailable. In 1966, a formalin-inactivated RSV vaccine failed and resulted in the death of two young children. This failure shifted research toward the development of subunit-based vaccines for pregnant women (to passively vaccinate infants) and the elderly. Among these subunit-based vaccines, the viral envelope glycoproteins show great potential as antigens. Areas covered: In this review, progress in the development of safe and effective subunit RSV vaccines based on the viral envelope glycoproteins and intended for pregnant women and the elderly, are reviewed and discussed. Studies published in the period 2012-2018 were included. Expert opinion: Researchers are close to bringing safe and effective subunit-based RSV vaccines to the market using the viral envelope glycoproteins as antigens. However, it remains a major challenge to elicit protective immunity, with a formulation that has sufficient (storage) stability. These issues may be overcome by using the RSV fusion protein in its pre-fusion conformation, and by formulating this protein as a dry powder. It may further be convenient to administer this powder via the pulmonary route.

The mechanism behind the biphasic pulsatile drug release from physically mixed poly(dl-lactic(-co-glycolic) acid)-based compacts.

Successful immunization often requires a primer, and after a certain lag time, a booster administration of the antigen. To improve the vaccinees' comfort and compliance, a single-injection vaccine formulation with a biphasic pulsatile release would be preferable. Previous work has shown that such a release profile can be obtained with compacts prepared from physical mixtures of various poly(dl-lactic(-co-glycolic) acid) types (Murakami et al., 2000). However, the mechanism behind this release profile is not fully understood. In the present study, the mechanism that leads to this biphasic pulsatile release was investigated by studying the effect of the glass transition temperature (Tg) of the polymer, the temperature of compaction, the compression force, the temperature of the release medium, and the molecular weight of the incorporated drug on the release behavior. Compaction resulted in a porous compact. Once immersed into release medium with a temperature above the Tg of the polymer, the drug was released by diffusion through the pores. Simultaneously, the polymer underwent a transition from the glassy state into the rubbery state. The pores were gradually closed by viscous flow of the polymer and further release was inhibited. After a certain period of time, the polymer matrix ruptured, possibly due to a build-up in osmotic pressure, resulting in a pulsatile release of the remaining amount of drug. The compression force and the molecular weight of the incorporated drug did not influence the release profile. Understanding this mechanism could contribute to further develop single-injection vaccines.

Ovalbumin-containing core-shell implants suitable to obtain a delayed IgG1 antibody response in support of a biphasic pulsatile release profile in mice.

A single-injection vaccine formulation that provides for both a prime and a boost immunization would have various advantages over a multiple-injection regime. For such a vaccine formulation, it is essential that the booster dose is released after a certain, preferably adjustable, lag time. In this study we investigated whether a core-shell based implant, containing ovalbumin as core material and poly(DL-lactic-co-glycolic acid) of various monomer ratios as shell material can be used to obtain such a booster release. An in vitro release study showed that the lag time after which the ovalbumin was released from the core-shell implant increased with increasing lactic to glycolic acid ratio of the polymer and ranged from 3-6 weeks. Fluorescence spectroscopy showed minimal differences between native ovalbumin and ovalbumin from core-shell implants that were incubated until just before the observed in vitro release. In addition, mice immunized with a subcutaneous inserted core-shell implant containing ovalbumin showed an ovalbumin-specific IgG1 antibody response after a lag time of 4 or 6-8 weeks. Moreover, delayed release of ovalbumin caused higher IgG1 antibody titers than conventional subcutaneous vaccination with ovalbumin dissolved in PBS. Collectively, these findings could contribute to the further development of a single-injection vaccine, making multiple injections of the vaccine superfluous.