Adenovirus was originally used as a vector for gene therapy. In recent years, with the development of the next-generation vectors with increased safety and high immunogenicity to transgene products, its utility as a vaccine vector has continued to increase. Adenovirus-based vaccines are currently being tested not only to prevent various infectious diseases but also to be applied as cancer vaccines.

Introduction

Adenovirus was initially mainly used as a delivery vector for specific genes, but as its highly immunogenic properties were elucidated, it was recognized as a vaccine vector capable of inducing various immune responses. In humans, more than 50 serotypes have been found to cause illnesses, from mild respiratory infections to life-threatening multi-organ diseases. Among them, human adenovirus serotypes 1, 2 and 5 (HAd1, HAd2, and HAd5, respectively) mainly cause mild respiratory diseases, and human adenovirus serotypes 4 and 7 (HAd4 and HAd7, respectively) may cause more severe pneumonia.

Considering the advantages of adenovirus as a vector, first, the structure of the genome is well understood, and it can be easily manipulated. Thus, it is relatively easy to insert foreign genes that are unable to replicate. Through this, the desired gene transfer or Ag expression can be easily achieved. Second, adenovirus has a very broad spectrum of host cell tropism, so it can infect host cells regardless of cell division, and this is an excellent characteristic of a delivery vector. Furthermore, it has been shown that the recombinant genome is stably maintained through continuous passages, and it can be quickly and massively produced. In addition to these advantages, when a recombinant adenovirus-based vaccine is used via a mucosal route, high transduction efficiency can be expected because mucosal infection is an inherent characteristic of adenoviruses. Thus, adenoviral vectors have emerged as very promising platforms for vaccines due to their documented immunogenicity and ability to induce host protection in multiple species, including humans.

To use adenovirus vectors safely and effectively in humans, the adenovirus genome has been improved through genetic engineering. First-generation adenoviral vectors are developed by E1/E3 deletion for transgene insertion, replication-defective properties, and improvement of immunogenicity (Fig. 1). However, in this generation, replication-competent adenovirus (RCA) is likely to occur due to the homology of the vector genome and the E1 part inserted into the packaging cell line HEK293. To resolve this shortcoming, cell lines with minimal homology, such as PERC.6, can be used in production. In second-generation adenoviral vectors, the transgene capacity is further increased by additionally deleting the E2/E4 site, and the likelihood of RCA formation might be decreased. However, the overall production yield is lower than that of first-generation adenoviral vectors due to decreased replication ability in producer cell lines. Third-generation adenoviral vectors are also called helper-dependent or gutless adenovirus vectors because they are created by deleting almost all genomic sequences except for sequences that are essential for packaging, such as inverted terminal repeat sequences. In addition, it boasts high capacity as it can insert multiple transgene expression cassettes, but it is more difficult to manufacture, its immunogenicity is lower than that of previous generations, and there is still the possibility of helper virus contamination.

Fig. 1 Schematic representation of the adenovirus genome and adenovirus-based vectors.

Interactions Between Adenoviral Vectors and The Immune System

Adenovirus stimulates innate immune cells by activating several innate immune signaling pathways and induces secretion of various proinflammatory cytokines and chemokines. This altered innate immune environment effectively induces robust adaptive humoral and cellular immune responses. In mice, intramuscular immunization with adenoviral vectors leads to efficient and sustained Ag expression in muscle tissues. Under these conditions, direct and cross-presentation of Ags to CD8 T cells occurs with high efficiency. To remove intracellular pathogens such as viruses, CD8 CTL responses are essential. Since transgene Ags derived from adenovectors are effectively presented to T cells by MHC class I molecules, efficient and robust CTL responses can be induced by adenovector vaccines. In particular, because of the diverse tropism of adenovirus, transgene Ags can be expressed in various cell types, which is advantageous for Ag presentation through MHC molecules. Although CTLs cannot provide sterilizing protection, their potential to limit the viral replication and reduce disease severity is another advantage.

Adenovector vaccines have been also shown to induce strong and durable humoral responses in animal models and humans. For example, the single injection of replication-defective adenovirus serotype 5 vectors elicit durable Ab responses in nonhuman primates and in humans. These properties allow efficient induction of robust and durable cell-mediated and humoral immunity and make adenoviral vectors promising vaccine vectors.

Fig. 2 The interaction between adenovirus and the innate and adaptive immune environment.

Current Status of Adenovirus-Based Vaccine Development

• Ebola/COVID-19 vaccine

The development of adenovirus-based vaccines has a long history. The first adenovirus-based vaccine approved for use in humans for preventive purposes was the Ebola vaccine named Ad26.ZEBOV. For fatal diseases such as Ebola, research using adenovector technology has been intensively conducted because of the urgent need for a vaccine. Of the 2 components of the new Ebola vaccine approved in Europe in 2020, Ad26.ZEBOV is used in the first dose.

Regarding a vaccine for COVID-19, which is currently being evaluated with large-scale clinical trials, a significant proportion of the lead candidates uses recombinant adenovector platform accounts. For example, vaccines using platforms such as HAd5 (nonreplicating), HAd26 (nonreplicating), and ChAdOx-1 (nonreplicating) are being developed through large-scale clinical trials. Most of these vaccine candidates are designed to express the S protein or RBD of SARS-CoV-2. Adenovirus-based vaccines can induce both strong cellular and humoral immunity, and the natural tropism of adenoviruses for the respiratory mucosa is an advantage of adenoviral vector-based vaccines for COVID-19-associated respiratory infections.

• Influenza vaccine

Studies of influenza vaccines based on adenovectors are actively being conducted because there is a great need for new vaccine development due to the various shortcomings of the currently used trivalent inactivated vaccine. In particular, as adenovectors can stimulate humoral immunity and induce broadly cross-reactive T-cell immunity against conserved Ags, many studies have employed the adenovector platform for universal influenza vaccines. As a typical example of universal influenza vaccines that are being developed, Abs can be induced by targeting the HA stalk region or the ectodomain of the M2 ion channel with relatively high conservancy, or broadly cross-reactive T cells can be induced to the NP or M1 as the target. Several human clinical trials have been conducted with HAd5-based influenza vaccines expressing HA, and their safety and immunogenicity were well proven in most cases. A dose-escalating phase II trial is currently being conducted to develop an intranasal spray-type influenza vaccine with HAd5-based vaccines expressing H1HA (https://clinicaltrials.gov/ct2/show/results/NCT03232567; Single-Ascending-Dose Study of the Safety and Immunogenicity of NasoVAX-Study Results. Available online). An optimal influenza vaccine should induce both broad cross-reactive T-cell immunity and neutralizing Abs, and the protective immunity should be sustained for a long period of time. In this regard, influenza vaccines using adenovector platform are attractive choices.

• HIV-1 vaccine

The high immunogenicity of adenovirus vectors has resulted in excellent efficacy of HIV-1 vaccines. Indeed, experiments conducted on nonhuman primates have shown very good efficacy in preventing infection. However, the subsequent large-scale STEP trial with a HAd5-based vaccine in humans did not show the expected efficacy in the vaccination group. Rather, HIV-1 infections tended to increase in HAd5-seropositive volunteers. Further research is needed to elucidate the mechanism in detail, but it is likely that vaccination-induced activation of CD4 T cells contributed to increased HIV-1 infection. Research on adenovector-based HIV-1 vaccines with a new strategy considering this hypothesis is currently being actively conducted. For example, to avoid pre-existing immunity and take advantage of the high immunogenicity of adenovectors, one clinical trial adopted a strategy using adenovectors of different serotypes for priming and boosting.

Creative Biogene offers a wide range of premaed adenovirus particles to aid our customers' research in gene therapy and vaccine development. Our products include reporter adenovirus particles, control adenovirus particles, iPS adenovirus particles, recombinase adenovirus particles, gene-specific recombinant adenoviruses and microRNA adenovirus particles.

View more of our premade adenovirus particles.