Stabilization of HSV-2 viral vaccine candidate by spray drying.

This work demonstrates that an HSV-2 candidate vaccine can be thermostabilized by spray drying to reduce cold chain demands. This work is also to optimize the process responses by varying spray dry parameters for pre-screened suitable excipients; and to determine the validity of current prescreening techniques. Vaccine activity losses were measured by in vitro plaque forming assay with Vero cell line. An accelerated storage condition of 45 °C for 10 days was used to determine spray dried sample stability. Prescreening studies demonstrated that trehalose and sucrose were superior to other tested excipients spray dry thermal stabilization of HSV-2. Subsequent optimization by design of experiments (DOE) of activity responses to spray dry parameter changes demonstrated significant differences between trehalose and sucrose for stability of the viral vaccine. Model parameters included the drying conditions inlet temperature, spray gas flow rate, and solids concentration for the model responses of vaccine stabilization. Trehalose was an effective and robust stabilizing excipient for spray drying HSV-2 vaccine. In contrast, stabilization by sucrose was greatly dependent on the spray dry process parameters. These DOE differences indicated inadequate excipient selection by prescreening methods and the variability demonstrated current prescreening techniques may not be adequate for determining optimal excipients.

Consecutive Spray Drying to Produce Coated Dry Powder Vaccines Suitable for Oral Administration.

Current global vaccination programs are challenged by costs associated with vaccine cold chain storage and administration. A solid, thermally stable oral dosage form for vaccines would alleviate these costs but is difficult to produce due to general vaccine instability and the complication of bypassing the gastric barrier. We developed a novel consecutive spray drying method that is suitable for use with biologics and employs Eudragit L100 polymer as the enteric coating. More specifically, in step 1, recombinant replication deficient human type-5 adenovirus and vesicular stomatitis virus were encapsulated by spray drying with sugars from a water solution, and in step 2, the microparticles from step 1 were suspended in ethanol with Eudragit and spray dried again. Up to 25% of the starting material was fully encapsulated within the enteric coating, and encapsulation efficiency was largely dependent on spray gas flow rate and the solids concentration in the feed. After step 2, the coated vaccine-sugar particles maintained their thermostability and were slightly larger in size with a rugous surface morphology compared to the particles produced in step 1. The coated particles retained viral vector activity in vitro after 15 min incubation in 1 M HCl (simulating the stomach environment) and anhydrous ethanol (to dissolve the Eudragit outer shell). The production of dry, orally administered vaccine particles from consecutive spray drying offers the potential to remedy a number of vaccine storage, transportation, and administration limitations.

Spray dried human and chimpanzee adenoviral-vectored vaccines are thermally stable and immunogenic in vivo.

Cold chain-free vaccine technologies are needed to ensure effective vaccine delivery and coverage, particularly in resource-poor countries. However, the immunogenicity and thermostability of spray dried live viral vector-based vaccines such as recombinant adenoviral-vectored vaccines remain to be investigated. To address this issue, we have spray dried human adenoviral (AdHu5)- and chimpanzee adenoviral (AdCh68)-vectored tuberculosis vaccines in a mannitol and dextran matrix. Spray dried powders containing these two vaccines display the morphologic and chemical properties desired for long-term thermostability and vaccination. Upon reconstitution, they effectively transfected the cells in vitro with relatively small losses in viral infectivity related to the spray drying process. Following in vivo vaccination, AdHu5- and AdCh68-vectored vaccines were as immunogenic as the conventional fresh, cryopreserved liquid vaccine samples. Of importance, even after cold chain-free storage, at ambient temperatures and relatively low humidity for 30 and 90days, the vaccines retained their in vivo immunogenicity, while the liquid vaccine samples stored under the same conditions lost their immune-activating capability almost entirely. Our results support further development of our spray drying technologies for generating thermally stable adenoviral-vectored and other viral-vectored vaccines.

Optimization of Spray Drying Conditions for Yield, Particle Size and Biological Activity of Thermally Stable Viral Vectors.

PURPOSE: This work examines the relevance of viral activity in the optimization of spray drying process parameters for the development of thermally stable vaccine powders. In some instances, the actual active pharmaceutical ingredient (API) is not included in the process optimization as it is deemed too costly to use until the final selection of operating conditions, however, that approach is inappropriate for highly labile biopharmaceutics. We investigate the effects of spray drying parameters on i) yield, ii) particle size and iii) viral vector activity of a mannitol/dextran encapsulated recombinant human type 5 adenoviral vector vaccine, to demonstrate the effects and magnitude of each effect on the three responses, and further show that the API must be included earlier in the optimization. METHODS: A design of experiments approach was used with response surface methodology (RSM) to optimize parameters including inlet temperature, spray gas flow rate, liquid feed rate and solute concentration in the feed. RESULTS: In general, good conditions for maintaining viral activity led to reduced yield and fewer particles of the desired size. Within the range of parameters tested, the yield varied from 50 to 90%, the percentage of ideally size particles was 10-50%, and the viral vector titre loss was 0.25-4.0 log loss. CONCLUSIONS: RSM indicates that the most significant spray drying parameters are the inlet temperature and spray gas flow rate. It was not possible to optimize all three output variables with one set of parameters, indicating that there will only be one dominant criteria for processing which in the case of viral vaccines will likely be viral vector activity.

Evaluation of excipients for enhanced thermal stabilization of a human type 5 adenoviral vector through spray drying.

We have produced a thermally stable recombinant human type 5 adenoviral vector (AdHu5) through spray drying with three excipient formulations (l-leucine, lactose/trehalose and mannitol/dextran). Spray drying leads to immobilization of the viral vector which is believed to prevent viral protein unfolding, aggregation and inactivation. The spray dried powders were characterized by scanning electron microscopy, differential scanning calorimetry, Karl Fischer titrations, and X-ray diffraction to identify the effects of temperature and atmospheric moisture on the immobilizing matrix. Thermal stability of the viral vector was confirmed in vitro by infection of A549 lung epithelial cells. Mannitol/dextran powders showed the greatest improvement in thermal stability with almost no viral activity loss after storage at 20°C for 90days (0.7±0.3 log TCID50) which is a significant improvement over the current -80°C storage protocol. Furthermore, viral activity was retained over short term exposure (72h) to temperatures as high as 55°C. Conversely, all powders exhibited activity loss when subjected to moisture due to amplified molecular motion of the matrix. Overall, a straightforward method ideal for the production of thermally stable vaccines has been demonstrated through spray drying AdHu5 with a blend of mannitol and dextran and storing the powder under low humidity conditions.