Abstract
There is an urgent need for the development of vaccine thermostabilisation methodologies as the maintenance of a continuous and reliable cold chain remains a major hurdle to the global distribution of safe and effective vaccines. Ensilication, a method that encases proteins in a resistant silica cage has been shown to physically prevent the thermal denaturation of a number of model proteins. In this study we investigate the utility of this promising approach in improving the thermal stability of antigens and vaccine conjugates highly relevant to the development of candidate tuberculosis vaccines, including antigen 85b conjugated with the Staphylococcus aureus-protein based adjuvant Sbi. Here we analyse the sensitivity of these constructs to thermal denaturation and demonstrate for the first time the benefits of ensilication in conferring these vaccine-relevant proteins with protection against temperature-induced loss of structure and function without the need for refrigeration. Our results reveal the potential of ensilication in facilitating the storage and transport of vaccines at ambient temperatures in the future and therefore in delivering life-saving vaccines globally, and in particular to remote areas of developing countries where disease rates are often highest.
Highlights
Given the barriers associated with the maintenance of a continuous and reliable cold chain, and lyophilisation, research on novel methods to thermally stabilise vaccines has risen substantially in the last decade
antigen 85b (Ag85b) co-incubated with or conjugated to Sbi III-IV is rapidly opsonised by activation products of the central complement component C3, including the natural molecular adjuvant C3d, which can lead to C3d-complement receptor 2 (CR2) and antigen-B cell receptor (BCR) co-ligation in vivo
In order to gain a better understanding of the temperature sensitivity of Ag85b, the Sbi III-IV-Ag85b vaccine conjugate and Sbi III-IV adjuvant, and the utility of ensilication in improving their thermal stability, these vaccine-relevant proteins were recombinantly expressed and purified (Fig. 1, Supplementary Table S1, Fig. S1), ensilicated and subjected to harsh short-term conditions of thermal denaturation or longer-term aging in the form of an ensilicated powder
Summary
Given the barriers associated with the maintenance of a continuous and reliable cold chain, and lyophilisation, research on novel methods to thermally stabilise vaccines has risen substantially in the last decade. Ensilication, a method recently developed by our group, is based on the solution-gelation process whereby negatively-charged silanol groups associate with charged amino acid residues through non-covalent electrostatic interactions[14] This results in growth of a protective silica cage around protein molecules[14,15] which is subsequently vacuum-filtered and dried. Potential Ag85b-containing vaccines being investigated are viral vector-based[18,19], contain other tuberculosis antigens including ESAT-620–23 and TB 10.424 and/or incorporate adjuvants such as toll-like receptor agonists (IC31)[25,26,27,28,29,30,31] and liposomes (CAF01)[32,33,34,35,36,37,38] In this regard, our group has recently revealed that domains III and IV of the Staphylococcus aureus immunomodulator Sbi can enhance the immunogenicity of Ag85b. The functionality of Sbi III-IV as an adjuvant compound has been investigated, its stability and temperature sensitivity, on its own or as part of a vaccine conjugate with Ag85b, remains unexplored
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