What Genetically engineered subunit vaccine are being developed?

Introduction to Subunit Vaccines

Definition and Types
Subunit vaccines are a class of immunizations that contain only specific antigenic components of a pathogen rather than the whole infectious organism. Rather than using live or inactivated whole pathogens, these vaccines comprise purified proteins, peptides, polysaccharides, or even virus‐like particles (VLPs) that are known to trigger protective immune responses. The selection of individual antigens is based on the identification of immunodominant regions or epitopes that can induce strong B-cell and T-cell responses. As a consequence, subunit vaccines can be classified into protein-based vaccines, peptide vaccines, polysaccharide vaccines, conjugate vaccines, and VLPs. This method of isolating the fundamental protective component from the pathogenic microorganism ensures that the vaccine is defined at the molecular level and does not include unnecessary or potentially reactogenic components.

Advantages and Limitations
There are several advantages to subunit vaccines. First, they have an enhanced safety profile because they do not involve whole pathogens, eliminating risks such as reversion to virulence or infection in immunocompromised individuals. Because the antigen is a well-defined component, there is also less potential for unwanted side effects or local reactogenicity compared to vaccines containing live attenuated or inactivated organisms. Additionally, subunit vaccines can be manufactured using recombinant DNA technologies that are well standardized and commonly used in biopharmaceutical production. However, a significant limitation of subunit vaccines is their relatively low immunogenicity when administered alone. Since they lack the full complement of pathogen-associated molecular patterns (PAMPs), the ability of these vaccines to stimulate innate immunity is reduced. Consequently, they often require the addition of adjuvants and booster doses to achieve long-lasting, protective immunity. Moreover, the production process itself—including the correct folding, post-translational modifications, and assembly—can be challenging, particularly when using different expression systems which impact the antigen’s structural integrity and immunogenicity.

Genetically Engineered Subunit Vaccines

Current Technologies and Methods
Genetically engineered subunit vaccines harness the power of recombinant DNA technology to produce antigenic protein components in various heterologous expression systems. The central strategy involves identifying the gene sequences that encode protective antigens, cloning these sequences into appropriate expression vectors, and then producing the recombinant proteins in systems such as bacteria (e.g., Escherichia coli), yeast (e.g., Pichia pastoris), insect cells (via baculovirus systems) or mammalian cells (e.g., CHO or HEK293).

For instance, modern approaches use advanced gene cloning and expression optimization techniques. One patent outlines a genetic engineering subunit vaccine against goat hydatidosis using an optimized version of the Eg95 protein. In this case, the Eg95 protein gene is codon-optimized for high expression in mammalian systems such as Chinese hamster ovary (CHO) cells, ensuring proper glycosylation, solubility, and yield of the vaccine antigen. In another example, genetic engineering methods are applied to develop an H9N2 subtype avian influenza vaccine by intercepting the extracellular region of the hemagglutinin protein, constructing a shuttle vector incorporating this gene, and then transfecting insect cell lines (such as hi5 cells) to produce the secreted protein antigen at high levels, an approach that circumvents the costs and limitations of egg-based systems.

Additional improvements in recombinant technologies include the use of mammalian stable expression cell lines (for example, HEK-293 T cells) that have been engineered to stably produce antigens like the E2 glycoprotein for classical swine fever vaccines. Genetic engineering not only optimizes the expression but also facilitates the incorporation of adjuvants or other immunomodulatory sequences to boost immune responses. Some systems even employ viral vectors (such as baculovirus) to ensure proper glycosylation of the expressed proteins, which is crucial as the immune system often responds better to proteins that closely mimic their native structure. Overall, these genetic engineering strategies permit rapid design and production of subunit vaccines with defined antigenic components while allowing for large-scale production in bioreactors, including serum-free processes that lower production costs and enhance safety profiles.

Examples of Vaccines in Development
Several genetically engineered subunit vaccines are at various stages of development, both for human and veterinary applications. Some notable examples include:

1. Goat Hydatidosis Vaccine:
A genetic engineering approach has been used to produce an optimized version of the Eg95 protein—an antigen derived from Echinococcus granulosus. This subunit vaccine is designed to prevent hydatidosis in goats. The technology involves high-level expression in CHO cells, where glycosylation and folding mimic the natural protein, leading to improved immunogenicity at a lower dosage.

2. H9N2 Subtype Avian Influenza Vaccine:
Another example is the genetic engineering subunit vaccine developed for the H9N2 avian influenza virus. Researchers isolate the extracellular fragment of the hemagglutinin (HA) protein, and through the use of recombinant baculovirus vectors in insect cells (hi5 cells), they achieve high quantities of secreted HA protein antigen. This approach simplifies production, reduces dependence on chick embryo cultures, and enhances the scale-up for mass vaccination programs in poultry.

3. Classical Swine Fever (CSF) Vaccine:
An E2 subunit vaccine for classical swine fever has also been produced using recombinant technologies. A stable HEK-293 T cell line has been developed that expresses the E2 protein consistently. Immunization studies in rabbits and pigs have demonstrated that the recombinant E2 subunit vaccine can induce robust neutralizing antibody responses and offer protection against classical swine fever virus challenges.

4. Feline Leukaemia Virus (FeLV) Vaccine:
A recombinant subunit vaccine against FeLV has been designed using an E. coli expression system to produce non-glycosylated forms of envelope proteins. The protein, expressed as rgp70D, includes key antigenic regions necessary for inducing both humoral and cellular immune responses. The vaccine formulation, when combined with novel adjuvants, has shown promising protective efficacy in experimental studies.

5. Rabies Subunit Vaccine:
In the evolution of rabies vaccines, the G protein is a primary antigen targeted for subunit vaccine development. Initial attempts using bacterial expression systems were hampered by poor immunogenicity due to aberrant glycosylation. Progress has been made by expressing the G protein in yeast and insect cells, with baculovirus expression systems providing a closer mimic to the native glycoprotein structure and thereby offering improved immunogenicity and protective neutralizing antibody responses.

6. Feline Leukaemia from External Sources:
In addition to the peer-reviewed studies, there are also product developments summarised on websites where genetically engineered subunit vaccines, for feline leukaemia among other viral targets, are being pursued by biopharmaceutical companies. These products leverage recombinant protein expression platforms to achieve consistent and scalable vaccine production.

These examples illustrate that genetically engineered subunit vaccines are being developed for both human and veterinary pathogens using a variety of production platforms, each chosen based on the antigen’s complexity and the necessity for post-translational modifications. The choice of expression system—whether bacterial, yeast, insect, or mammalian—plays a pivotal role in the functional and immunogenic properties of the final vaccine product.

Potential Benefits and Challenges

Efficacy and Safety Considerations
Genetically engineered subunit vaccines offer major benefits, primarily in the area of safety. Because they only include specific antigenic segments rather than whole organisms, the risk of adverse reactions such as reversion to a virulent form is virtually eliminated. This safety profile is particularly important when vaccinating vulnerable populations such as immunocompromised individuals, pregnant women, and neonates. Moreover, the defined composition allows for rigorous characterization of the vaccine product, ensuring consistency across lots and lowering the risk of batch-to-batch variation.

However, the inherent purity and defined nature of subunit vaccines often result in lower immunogenicity. Without the presence of multiple pathogen-associated molecular patterns (PAMPs), these vaccines may not stimulate a sufficiently robust innate immune response on their own. As a consequence, genetically engineered subunit vaccines frequently require the inclusion of adjuvants to provoke a more potent and durable adaptive immune response. Safe adjuvants, such as alum or novel TLR ligands, have been used in various studies to augment the immune response while maintaining an acceptable safety profile. The balance between achieving adequate immune responses and maintaining safety is a core focus of current research in subunit vaccine development.

Another challenge relates to the structural fidelity of the recombinant antigen. The expression system selected can influence the glycosylation, folding, and even the conformation of the antigen, all of which are critical determinants of immunogenicity. For example, the same rabies virus G protein expressed in different systems may result in disparate glycosylation patterns; only those expressed in systems that can mimic native glycosylation (such as yeast or insect cells) have demonstrated the desired immune responses in experimental models. Therefore, both the choice of adjuvant and the expression platform are critical in ensuring that the vaccine is both safe and efficacious.

Manufacturing and Distribution Challenges
From a manufacturing standpoint, genetically engineered subunit vaccines benefit from established recombinant protein production technologies, which leverage controlled bioreactor conditions for large-scale production. Systems such as CHO cells or hi5 insect cells are already widely used in biopharmaceutical manufacturing, enabling cost-effective and scalable production. However, the requirement for extensive protein purification—often to a higher standard than traditional vaccines—can increase production complexity and cost. The formulation process must also ensure that the vaccine product is stable, particularly in scenarios where cold chain maintenance may be challenging.

In addition, the choice of expression system not only impacts the immunogenic profile of the vaccine but also its stability and ease of downstream processing. For example, while bacterial expression systems like E. coli can produce high yields rapidly, the lack of post-translational modifications such as glycosylation may require additional steps to reconstitute the antigen’s native conformation, potentially affecting the vaccine’s potency. Conversely, mammalian systems produce proteins that more closely resemble native antigens, but these platforms are generally more expensive and time-intensive in terms of development and production.

Distribution challenges also persist even after an effective vaccine has been manufactured. For instance, the necessity for an adjuvant and possible booster doses complicates administration protocols, and the sensitive nature of protein antigens can require strict temperature-controlled storage conditions. Innovative formulation techniques, such as lyophilization (freeze-drying) or nanoparticle encapsulation, are being explored to address these issues and improve the vaccine’s shelf life and ease of distribution, particularly in low-resource settings.

Future Directions and Innovations

Emerging Research and Technologies
Innovations in vaccine design are rapidly emerging, promising to address many of the current limitations of genetically engineered subunit vaccines. One of the most exciting areas is the integration of nanotechnology into vaccine delivery systems. Nanoparticle carriers—including liposomes, polymeric nanoparticles, and even ferritin-based nanoparticles—are being engineered to deliver antigens in a controlled and sustained manner. These carriers can protect the antigen from degradation, enhance cellular uptake, and even serve as built-in adjuvants by mimicking the pathogen’s size and shape. The use of nanoparticles thus represents an innovative strategy that could significantly improve the immunogenicity of subunit vaccines while maintaining safety.

Another emerging field is that of genetic code expansion. Recent advances allow for the incorporation of noncanonical amino acids into recombinant proteins, which can help in fine-tuning the antigen’s immunogenic properties or even creating replication-incompetent virus structures for vaccine use. Additionally, mRNA vaccine technology, which has gained unprecedented attention during the COVID-19 pandemic, offers lessons and approaches that could be applied to subunit vaccine development—for example, mRNA encoding a stabilized antigen may be used to drive high-fidelity expression of vaccine proteins in vivo with rapid production timelines.

Advances in bioinformatics and reverse vaccinology are also playing a crucial role. These methods enable researchers to screen pathogen genomes rapidly and identify promising antigen candidates even before laboratory experiments begin. This approach significantly shortens the development cycle and may lead to multivalent vaccine designs that cover a broader spectrum of antigenic variants, thereby enhancing cross-protection against different strains. In this context, combination subunit vaccines—where multiple recombinant antigens are assembled into a single formulation—are emerging as a promising strategy to tackle pathogens with high mutation rates or multiple serotypes.

Regulatory and Ethical Considerations
As genetically engineered subunit vaccines transition from bench to bedside, regulatory and ethical challenges must be carefully managed. The production and quality control of recombinant proteins require rigorous adherence to current Good Manufacturing Practices (cGMP) and extensive validation to ensure consistent safety and efficacy. Regulatory agencies such as the FDA require thorough preclinical studies followed by phased clinical trials for any new vaccine candidate. Given that subunit vaccines are relatively new in this context, there is an evolving landscape of guidelines on acceptable purity levels, adjuvant safety, and immunogenicity endpoints.

Ethically, the use of genetically engineered vaccines necessitates transparency regarding potential risks, benefits, and uncertainties. Informed consent becomes particularly important when new adjuvants or nanoparticle delivery platforms are included, as long-term effects remain under investigation. Moreover, integrating these vaccines into global immunization programs must balance the need for rapid and widespread distribution with concerns about equitable access and affordability. For instance, while high-income countries may be able to adopt cutting-edge subunit vaccines quickly, cost and infrastructure barriers may limit access in low-resource settings. This creates an imperative for international collaboration in regulatory harmonization and funding support to ensure that innovative vaccines serve global public health needs.

Conclusion
Genetically engineered subunit vaccines represent a transformative approach in modern vaccinology that underscores safety, specificity, and scalability. In summary, these vaccines are built on carefully selected antigenic components that, when produced through advanced recombinant DNA technologies, provide a well-defined product with reduced risk of adverse events compared to whole-pathogen vaccines. The current technologies and methods range from high-yield expression in CHO cells or insect cells to the development of stable cell lines that can produce complex proteins with native-like glycosylation patterns. Examples in development include vaccines against goat hydatidosis, H9N2 avian influenza, classical swine fever, feline leukaemia, and improved rabies vaccines, each tailored to optimize immune response while addressing the inherent limitations of subunit formulations.

From an efficacy and safety perspective, genetically engineered subunit vaccines offer enhanced safety because they only incorporate necessary antigenic determinants, thereby avoiding the risk of reversion to virulence. However, their lower inherent immunogenicity necessitates the use of adjuvants and sophisticated delivery systems to elicit robust immune responses. Manufacturing challenges—such as ensuring correct folding, glycosylation, stability, and cost-effective purification—remain, yet advances in bioprocessing and formulation technologies are steadily addressing these issues.

Looking ahead, emerging research avenues such as nanoparticle-encapsulated antigens, genetic code expansion techniques, and mRNA-inspired approaches promise to revolutionize the subunit vaccine field. These innovations, coupled with robust bioinformatics and reverse vaccinology strategies, are paving the way for multivalent and broadly protective vaccines that can adapt to the challenges posed by rapidly evolving pathogens. Simultaneously, regulatory and ethical considerations remain central to ensuring that these new vaccine platforms are both safe for individual use and viable for public health deployment, particularly in resource-limited settings.

In conclusion, the ongoing development of genetically engineered subunit vaccines is a dynamic and multi-disciplinary field that is poised to address some of the most persistent challenges in vaccine design. By leveraging modern genetic engineering methods, innovative antigen delivery systems, and advanced manufacturing technologies, these vaccines offer the promise of safer, more effective, and more accessible immunization strategies for both human and veterinary diseases. Continued investment in research, coupled with international regulatory collaboration and ethical oversight, will be vital in realizing the full potential of these next-generation vaccines.