Advances in recombinant DNA technology, genomics, and immunology have greatly influenced the vaccine development process. The genetic manipulation of viruses makes them suitable as vectors for gene delivery, providing a novel and unique opportunity for vaccine design. Viruses survive and replicate by invading their host’s cells and hijacking their protein-making machinery. These virus particles contain antigens, molecules that can trigger an immune response. A similar principle underpins viral vector vaccines, though in this case the host cells only receive code to make antigens. The viral vector acts as a delivery system, taking advantage of the virus’s inherent ability to infect and insert the code for a different virus’ antigens (the pathogen you’re trying to vaccinate against), and finally transducing cells to produce the desired immunized outcome. The virus itself is harmless, and by getting the cells only to produce antigens the body can mount an immune response safely, without developing disease.
With the in-depth study of various viruses, viral vectors are becoming an important immuno-prophylactic tool in modern medical arsenals. Among various vaccine vectors, adenovirus vectors are being extensively investigated and show great promise as vaccine vectors. Currently, there are three approved and widely administered adenovirus vector vaccines for COVID 19 worldwide; AstraZeneca/Oxford’s Covishield, Gamelya Research Institute’s Sputnik V, and Johnson & Johnson/Jansen’s vaccine. They are made of replication-deficient adenovirus serotypes loaded with a transgene encoding the spike, S protein of the coronavirus.
Adenoviruses (AdVs) are non-enveloped icosahedral viruses ranging from 70-90 nm in size with an outer protein shell surrounding an inner nucleoprotein core that belong to the adenoviridae virus family. Named for the adenoid (tonsil) tissue from which they were first identified, AdVs are now known to infect a broad range of tissues, causing mostly benign infections such as nasal congestion and rhinorrhea (runny nose), cough and conjunctivitis. Adenoviruses are double-stranded DNA viruses with a genome of 34~43 kb, a size that is amenable for easy manipulation. The facets of the viral capsid are composed primarily of trimers of hexon protein and other minor components. The apex of the capsid is composed of a penta-base, which acts to anchor fibrin and is responsible for attaching the virion to the cell surface. The genome encodes five early gene transcription units and a late transcription unit. The early transcription units play a key role in viral DNA replication and evasion of host immunosurveillance, while the late transcription units primarily encode viral structural components.
Malvern Panalytical Technology Solutions
As the demand for viral vectors in gene therapy and vaccines development grows, robust and reproducible platforms are needed. Monitoring critical quality attributes (CQAs) is important in the early development phase all the way up to the late scale-up manufacturing phase to ensure reproducibility. These CQAs include viral potency, identity, quantity, process residuals, aggregation, empty capsids, protein content and product safety. There are different characterization assays available and their applicability depends on the type of virus purified as well as the expression system used. Malvern Panalytical offers rapid and comprehensive technology solutions for assay development to monitor several of these CQAs. Here is a list of the CQAs and the corresponding applicable technology solutions:
Physical Viral Titre or Concentration
NanoSight 300 is a real-time nanoparticle visualization system based on Nanoparticle Tracking Analysis, where a laser-illuminated microscope detects Brownian motion of and counts virus particles in solution. Another technology solution for viral titre determination of viruses with size range < 40 nm, like adeno-associated virus (AAV), is the Zetasizer Ultra, which is based on Multi Angle Dynamic Light Scattering, MADLS to count photons, and calculates particle concentration from other measured or known sample properties, including size, refractive index, viscosity, and buffer scatter rate.
Formulation Stability and Aggregate Formation
Zetasizer Advance Range with Dynamic and Electrophoretic Light Scattering Capabilities to probe the presence of unwanted aggregates and measure surface charge and zeta potential to assess formulation stability.
Capsid Content: Empty vs Full
Malvern Panalytical’s OMNISEC is a Gel Permeation Chromatography (GPC) / Size Exclusion Chromatography (SEC) system, comprised of OMNISEC RESOLVE and OMNISEC REVEAL suites, for the measurement of absolute molecular weight, molecular size, intrinsic viscosity, branching and other parameters. Its multi-detector suite, OMNISEC REVEAL, is compatible with any Size Exclusion Chromatography system, providing comprehensive Compositional Analysis of viral vectors for full capsid content ratios in a sample.
The MicroCal PEAQ-DSC is a great tool for understanding conformational and thermal stability by obtaining multiple descriptors of thermal transitions and obtaining fingerprints of Higher Order Structure.
- Biophysical tools to accelerate the development of safe and effective vaccines
- Orthogonal and complementary nanoparticle characterization techniques
- Total virus titer in minutes
- Measuring the concentration of Adeno-Associated Virus (AAV) with multi-angle dynamic light scattering (MADLS)
- The importance of Multi-Detection SEC in gene therapy
- Focus on Vaccine Development 4 – Virus: Vaccine and VLP Characterization with Nano-particle Tracking Analysis (NTA