Development and preclinical evaluation of an oral therapeutic colorectal cancer vaccine
Colorectal cancer (CRC) has become the second most common cause of cancer death worldwide, indicating the need for novel therapies to reduce mortality rates and the global burden of this disease. Therapeutic subunit vaccines are a safe and promising method for the treatment of established cancers. These vaccines aim to stimulate the patient’s immune system to recognize and eradicate malignant cells. Robust immune stimulation is necessary to maximize the therapeutic potential of the cancer vaccines and in the case of CRC, this can be achieved via oral vaccine administration. Anti-tumour immune responses can then be initiated at the tumour location, specifically, at the lymphoid tissues of the gastrointestinal tract. However, vaccines delivered orally require protection from the harsh conditions found in the gastrointestinal tract. This protection can be provided through the use of appropriate delivery systems, which also facilitate the addition of vaccine adjuvants. The hypothesis of this research was that orally delivered therapeutic subunit cancer vaccines in lipid-based formulations stimulate local and systemic anti-tumour immune responses that would lead to CRC elimination in a preclinical orthotopic mouse model. The hypothesis was tested initially with oral subunit vaccine formulations composed of a peptide containing epitopes for CD8+ and CD4+ T cells, in combination with the Toll-like receptor 2 (TLR2)-stimulating adjuvant, Pam2Cys. These formulations were encapsulated in liposomes and in water-in-oil-in-water (W/O/W) double emulsions. Following optimisation of the formulations, the emulsion vaccine induced local immune cell accumulation and systemic lymphocyte activation after oral vaccination. Furthermore, both emulsion and liposome vaccines demonstrated therapeutic potential by significantly reducing the growth of CT26 tumours injected intracaecally into mice. The findings confirmed that positive therapeutic effect can be achieved with oral subunit vaccines delivered in lipid-based formulations. Although both vaccine candidates demonstrated promising treatment outcomes, they were not able to completely eliminate the tumours. Therefore, a number of modifications were made to the vaccine and/or the therapy including; using self-adjuvanting peptides, modifying liposomes for targeted delivery, and combination therapy with a nonsteroidal anti-inflammatory drug (NSAID) with or without an immune checkpoint inhibitor. The self-adjuvanting construct was not effective when delivered orally or with lipid-based formulations, underlining the need to further investigate the immunostimmulatory activity of these constructs as oral vaccines. Targeting of liposome to M cells with the Ulex europaeus agglutinin I (UEAI) lectin also did not improve therapeutic outcomes. It was not clear if this was due to the lectin not improving liposome uptake by the M cells or if improved uptake had occurred but was not translated into therapeutic effect. Combination therapy with the NSAID licofelone was the most promising strategy with a partial response to vaccination and reduced tumour growth. The addition of the checkpoint inhibitor anti-programmed cell death protein 1 (PD-1) was of no benefit in this mouse model. This is likely due to insufficient lymphocyte infiltration into the tumour. A novel technique utilising bacterial ‘microswimmers’ to improve the delivery and immunogenicity of the oral vaccines, was explored and adapted for the preclinical evaluation of immunotherapeutic CRC treatment. Escherichia coli (E. coli) were attached to liposomes or emulsion vaccine particles and used as an oral vaccine microsystem. Microswimmer vaccines were not toxic and demonstrated favourable immunostimulatory profiles in cell culture, as expected due to the natural pathogen associated molecular patterns (PAMP), which could interact with pattern recognition receptors (PRR) on dendritic cells (DC). Improved therapeutic outcomes were observed after treatment with emulsion microswimmers in mice, confirming that bacteria can be a powerful immunostimulatory agent and boost immune responses to orally delivered vaccine antigens. Although tumour growth was reduced with the emulsion microswimmers, the liposome microswimmers did not demonstrate the same effect. The likely reason for this is the stability of the bacterial attachment. The study evaluated strategies for the treatment of established colorectal tumours with oral subunit vaccines administered in lipid-based formulations. The preclinical vaccine development study provided important insights for the future investigation of therapeutic oral CRC vaccines. The findings confirmed the potential of these vaccines to be used in human therapy as the vaccine candidates demonstrated immunostimulatory and therapeutic capacity to induce multiple anti-tumour immune responses and reduce tumour growth. The emulsion vaccine candidate is being further investigated for use in clinical trials as it produced the best results and is easier to make, facilitating its transition into the clinic.
Advisor: Hook, Sarah; Kemp, Roslyn
Degree Name: Doctor of Philosophy
Degree Discipline: School of Pharmacy
Publisher: University of Otago
Keywords: New Zealand; oral; vaccine; therapeutic; colorectal; cancer; liposomes; emulsion; orthotopic
Research Type: Thesis