Vaccines are often regarded as the most important biomedical advance in the fight against infectious disease. The ability of vaccines to elicit a protective immune response is also being exploited in the fight against cancer and Alzheimer’s disease. While conventional vaccines against infectious diseases are based on inactivated or live attenuated pathogens, peptide-based vaccines have many advantages, including fully-defined composition with no biological contamination, the possibility to customize, and the possibility of cost-effective large-scale production. Success in the rational design, development and production of peptide vaccines depends on many factors, not least the ability to synthesize often complex peptides efficiently and quickly.
Solid-phase peptide synthesis makes it all possible
Advances in solid-phase peptide synthesis (SPPS) now enable the large-scale synthesis of complex peptides with high purity and yield, which has powered the development of new approaches to the rational design of vaccines. How this can be achieved will be the subject of the final article in this series, ‘How to ensure optimal purity and yield in solid-phase peptide synthesis’ .
Peptide vaccines against infectious and chronic diseases
Peptide-based vaccines are under development against a number of pathogens, including the parasite causing malaria, Hepatitis C virus, influenza virus, and HIV-1. Peptides are also featuring in the cutting-edge R&D efforts to fight the COVID-19 pandemic. This includes investigations into the potency of peptides as therapeutics1 and also as vaccines2. Another path involves a novel ’antidote-vaccine’ that could act as a prophylactic, immune-stimulant and therapeutic against the SARS-CoV-2 virus3.
Peptides are also being used in vaccines in cancer, in the form of neoantigens that are normally presented on the surface of tumor cells and can be targeted by T cells as foreign, leading to cancer cell death4. Another application is the use of HER2-related peptides to increase T-cell helper response in the fight against HER2-expressing tumor cells5. Alzheimer’s disease is also being addressed with peptide-based vaccines that target the aberrant aggregation of two proteins, Aβ and Tau5.
Vaccines must be shown to be safe, pure, potent, and effective for regulatory approval, and this process can normally take 10–15 years. Added to that, developing vaccines is a complex risk-intensive process requiring substantial investment and with a high attrition rate; fewer than 10% of vaccine candidates ever reach the market6. Let’s look at how vaccines work in general, and the challenges in peptide vaccine design, synthesis and production in particular, aimed at presenting a correctly-conformed epitope to the immune system that elicits a stable immune response in a safe manner.
How to stimulate an immune response
Vaccines against infectious diseases are designed to preempt attack from a pathogen by exposing the immune system of a naïve host to pathogen epitopes to induce a protective immune response. Vaccines directed against chronic diseases such as cancer and Alzheimer’s disease target ‘self’ antigens.
The immune response essentially involves two types of response that are relevant in vaccine design:
- B cell – Vaccines can stimulate the production of epitope-specific antibodies by B cells. Neutralizing antibodies can stop infection by blocking host cell attachment, the entry of pathogens, or by inducing pathogen−antibody immune complexes that are cleared systemically.
- T cell – Stimulating epitope-specific T cells can lead to rapid destruction and clearance of the pathogen, or pathogen-infected host cells. In the case of immunooncology, tumors frequently evade immune surveillance by local downregulation of cancer-specific T-cells. Immunotherapies can aim at globally upregulating T cells.
Most conventional vaccines against infectious diseases are based on inactivated or live attenuated pathogens. These contain both B- and T-cell epitopes presented in a conformation that is relevant to the pathogen and therefore stimulate a robust immune response. An alternative is a subunit vaccine that consists primarily of peptides or proteins that can stimulate long-lasting protection, and this has become a tendency in vaccine development. Interest in peptide-based vaccines is increasing since peptides have a number of advantages that support rational vaccine design:
- Fully-defined composition
- Large-scale production is affordable
- Water-soluble, stable in storage, can be freeze-dried
- No biological contamination
- Minimal allergic and autoimmune response
- Can be customized
- Different sequences can be quickly synthesized in parallel
Choosing the right epitope
The immunogenicity of regions of antigens varies for both B- and T-cell epitopes. This immunodominance is particularly important for peptide vaccines that target only a single or a few critical epitopes. Choosing the most effective binding region or regions to focus on is therefore vital.
The art of mimicry
The peptide of a vaccine must mimic the epitope presented by a structure that is often more complex. Larger structural epitopes are generally conformation-dependent, with recognition based on residues that can be close or distant in primary protein sequence and brought together by globular folding. These larger epitopes have been mimicked by, for example, peptide engineering using phage display involving selection from a naïve peptide library of candidates that bind the antibody.
In contrast, conformation-independent epitopes have low complexity and are formed by linear stretches of residues that may adopt local secondary structure once they are bound to the antibody. These linear epitopes are generally found in protein loops and are prime candidates for peptide vaccine design based on peptide synthesis.
Improving the performance of peptides
While peptide-based vaccines can be effective and have a promising future, there are some disadvantages that must be addressed.
- Poor immunogenicity – The simplicity of peptides becomes a disadvantage due to the limitations of the epitope that can be presented to the immune system. Immunogenicity can be augmented through linkage to short sequences that are known to stimulate an immune response. Epitopes can be presented in multimeric formats based on virus-like particles (VLPs) or nanoparticles. Another common approach is the inclusion of immunostimulatory adjuvants.
- Unstable in vivo – The degradation of peptides can be reduced by binding to biopolymer conjugates or nanoparticles, which can also act as adjuvants.
- Loss of native conformation – The structure of the peptide can be stabilized by adding flanking sequences, cyclization, stapling etc.
- Effective for a limited population – The immunostimulatory power of peptide-based vaccines can be improved by multi-epitope constructs.
Challenges in peptide synthesis for vaccine R&D and production
Ensuring the immunogenicity of a peptide-based vaccine often depends on the ability to synthesize complex peptides:
- Efforts to improve the rigidity, stability and resistance to degradation of therapeutic peptides have resulted in synthesis of phospho-peptides, and cyclic peptides that are stapled, disulfide-bridged, bicyclic, or cyclized head-to-tail.
- Longer peptides, for example >30 amino acids, are a challenge to synthesize.
- Beta-amyloids involved in Alzheimer’s disease and used in vaccine research and development are highly hydrophobic.
- Peptides can be branched, include side-chain modifications, or be cysteine rich.
- Post-translational modifications (PTMs) of native peptides may need to be mimicked to achieve maximum effect, for example in neoantigens.
Meeting these challenges means using a peptide synthesizer that can handle complex chemistries while minimizing cross-contamination, dead volumes, and reagent carryover. This is especially important for the synthesis of long sequences, in which even small amounts of impurities, side products, and incomplete reactions over many cycles can drastically reduce the final purity and yield of desired peptides. Aggregation, secondary structure, steric hindrance, and conformational effects can still pose challenges in synthesis, and real-time deprotection monitoring optimizes reaction times to ensure complete deprotection.
In the case of neoantigens, parallel peptide synthesis is also important to quickly generate the pool of neoantigens needed. Speed is a critical factor since timing the delivery of a vaccine can make all the difference to the outcome. Neoantigens may be further altered through posttranslational modifications (PTMs) that occur in malignant but not healthy cells and are therefore an additional source of unique antigens that are specific to the individual patient.
Peptides hold promise as the basis for rational design and development of vaccines, but challenges with immunogenicity, stability and conformation demand advanced solid-phase peptide synthesis that enables the rapid and efficient synthesis of complex peptides.
You can find out more about how neoantigen vaccines promise to bring personalized cancer therapy to a new level by reading the Case Study, ‘Cancer is a personal issue – a worldwide scientific effort focuses on making the treatment match the individual’
1 Synthetic peptides are promising therapeutic candidates for Covid-19. GlobalData Healthcare 26 May 2020
2 A candidate multi‑epitope vaccine against SARS‑CoV‑2. Kar, T et al. Scientific Reports. (2020) 10:10895.
3 Peptide antidotes to SARS-CoV-2 (COVID-19). Watson, A et al. bioRxiv, August 6, 2020. doi.
4 Personalized neoantigen vaccination with synthetic long peptides: recent advances and future perspectives. Chen, X et al. Theranostics 2020; 10(13): 6011-6023. doi: 10.7150/thno.38742
5 Peptide-based vaccines: Current progress and future challenges. Malonis, RJ et al. Chem. Rev. 2020, 120, 3210−3229.
6 Risk in vaccine research and development quantified. Pronker ES et al (2013) PLoS ONE 8(3): e57755. doi:10.1371/journal.pone.0057755