A live attenuated RSV vaccine, process development studies

Respiratory Syncytial Virus (RSV) is the leading cause of lower respiratory tract disease in infants and young children. A vaccine to prevent the high burden of disease caused by RSV is urgently needed, but not available. A live attenuated respiratory syncytial virus (RSV) vaccine for intranasal delivery is currently under development at Intravacc. The vaccine concept comprises a live Glycoprotein-complemented RSVΔG virus. This G-RSVΔG virus is generated by proliferation of an RSVΔG on G-expressing Vero cells. The vaccine thus contains virus particles that have the G-protein on their surface but not in their RNA genomes. This recombinant virus is highly attenuated compared to wild type RSV and therefore presents a live attenuated vaccine candidate for RSV infection. A vaccine production process has been setup for the production of Phase I clinical lots. In short, the production process steps are: cell and virus culture, clarification, continuous flow density gradient ultracentrifugation, ultra/diafiltration, filling and lyophilization. An example of process development is the design of the cell and virus culture method. Using the statistical design of experiment approach the virus culture has been optimized to both virus yield and harvest quality. As RSV is a filamentous virus, the optimization of harvest quality with respect to purification opportunities is pivotal. This DoE was done at lab-scale bioreactors (2-L) and the chosen conditions were successfully scaled-up to 50-L single use bioreactors. Preparation of preclinical and clinical lots is done at this scale. The pre-clinical studies were successful. In the cotton rat model, the G-RSVΔG vaccine is safe, immunogenic and protects against challenge with wild type RSV. The following step, a clinical Phase I study, is planned

The effect of formulation on spray dried Sabin inactivated polio vaccine

The objective of this study was to develop a stable spray dried formulation, containing the three serotypes of Sabin inactivated polio vaccine (sIPV), aiming for minimal loss of native conformation (D-antigen) during drying and subsequent storage. The influence of atomization and drying stress during spray drying on trivalent sIPV was investigated. This was followed by excipient screening, in which monovalent sIPV was formulated and spray dried. Excipient combinations and concentrations were tailored to maximize both the antigen recovery of respective sIPV serotypes after spray drying and storage (T = 40 °C and t = 7 days). Furthermore, a fractional factorial design was developed around the most promising formulations to elucidate the contribution of each excipient in stabilizing D-antigen during drying. Serotype 1 and 2 could be dried with 98% and 97% recovery, respectively. When subsequently stored at 40 °C for 7 days, the D-antigenicity of serotype 1 was fully retained. For serotype 2 the D-antigenicity dropped to 71%. Serotype 3 was more challenging to stabilize and a recovery of 56% was attained after drying, followed by a further loss of 37% after storage at 40 °C for 7 days. Further studies using a design of experiments approach demonstrated that trehalose/monosodium glutamate and maltodextrin/arginine combinations were crucial for stabilizing serotype 1 and 2, respectively. For sIPV serotype 3, the best formulation contained Medium199, glutathione and maltodextrin. For the trivalent vaccine it is therefore probably necessary to spray dry the different serotypes separately and mix the dry powders afterwards to obtain the trivalent vaccine.</p

Developments in the formulation and delivery of spray dried vaccines

Spray drying is a promising method for the stabilization of vaccines, which are usually formulated as liquids. Usually, vaccine stability is improved by spray drying in the presence of a range of excipients. Unlike freeze drying, there is no freezing step involved, thus the damage related to this step is avoided. The edge of spray drying resides in its ability for particles to be engineered to desired requirements, which can be used in various vaccine delivery methods and routes. Although several spray dried vaccines have shown encouraging preclinical results, the number of vaccines that have been tested in clinical trials is limited, indicating a relatively new area of vaccine stabilization and delivery. This article reviews the current status of spray dried vaccine formulations and delivery methods. In particular it discusses the impact of process stresses on vaccine integrity, the application of excipients in spray drying of vaccines, process and formulation optimization strategies based on Design of Experiment approaches as well as opportunities for future application of spray dried vaccine powders for vaccine delivery

Development of a thermostable spray dried outer membrane vesicle pertussis vaccine for pulmonary immunization

Worldwide resurgence of whooping cough calls for improved, next-generation pertussis vaccines that induce broad and long-lasting immunity. A mucosal pertussis vaccine based on outer membrane vesicles (omvPV) is a promising candidate. Further, a vaccine that is stable outside the cold chain would be of substantial advantage for worldwide distribution and application. A vaccine formulated as a powder could both stabilize the vaccine as well as make it suitable for pulmonary vaccination. To that end, we developed a spray dried omvPV with improved stability compared to the liquid omvPV formulation. Spray drying did not affect the structural integrity of the omvPV. The antigenicity of Vag8, a major antigen in omvPV was diminished slightly and an altered tryptophan fluorescence indicated some changes in protein structure. However, when administered via the pulmonary route in mice after reconstitution, spray dried omvPV showed comparable immune responses and protection against challenge with live B. pertussis as liquid omvPV. Mucosal IgA and Th17 responses were established in addition to broad systemic IgG and Th1/Th17 responses, indicating the induction of an effective immunity profile. Overall, a spray dried omvPV was developed that maintained effective immunogenic properties and has an improved storage stability

Application and comparison of lyophilisation protocols to enhance stable long-term storage of filovirus pseudotypes for use in antibody neutralisation tests

Filoviruses encompass highly pathogenic viruses placing significant public health burden on countries affected. Efforts for improved diagnostics and surveillance are needed. The requirement for high-containment can be circumvented by using pseudotype viruses (PV), which can be handled safely, in tropism, drug screening, vaccine evaluation and serosurveillance studies. We assessed stability and functionality after long-term storage of lyophilised filovirus pseudotypes for use in neutralisation assay