The project generated multicomponent nanovaccines targeting tumour associated antigens by encapsulating different peptides (NY-ESO1, MSLN and KRAS), as well as different adjuvants such as the TLR3 agonist Poly(I:C; pIC) and the dual TLR7/8 activator Resiquimod. Covalent attachment of polyethylene glycol chains increased the solubility, bulk amount, and recovery of most peptides. Several of the synthesized peptides were encapsulated into polymer PLGA-NPs, that were coated with chitosan to prevent aggregation and increase the affinity to tumour cells. Nanovaccines were partly labeled with fluorescent dyes, for visual tracking in both in vitro and in vivo assays. These vaccines were designed to stimulate a robust immune response, particularly by activating T-cells against PDAC cells.
Recognizing the limitations of existing preclinical models, PAVE developed new models, including a porcine (pig) model that replicates human PDAC more accurately than mouse models. This "oncopig" with specific genetic mutations relevant to PDAC, provides a valuable tool for testing the efficacy of new therapies. In addition, PAVE succeeded in developing a toolbox of pig-specific immunology reagents and assays including bioactive immune proteins, cloned cytokines and chemokines, and monoclonal antibodies to the proteins and their receptors for the porcine model.
PAVE researchers developed several innovative imaging tools, such as near-infrared fluorescent biosensors, transgenic mice with bioluminescent and fluorescent gene reporters, and optoacoustic imaging techniques. These tools enable for example real-time tracking of vaccine biodistribution, T-cell activation, and tumour progression in vivo, enhancing the ability to monitor treatment responses and thus optimize therapy.
The project made significant progress in understanding the immunosuppressive nature of the PDAC tumour microenvironment (TME). This included the identification of immunoregulatory genes (IRGs) whose knockdown improved T-cell mediated tumor killing, the characterization of the heterogeneity of cancer associated fibroblasts (CAFs) in human PDAC, and the development of a novel platform for constructing immune component-based antigens targeting key PDAC proteins such as MSLN and KRAS.
Research on the role of extracellular vesicles (EVs) in PDAC revealed their strong thrombotic potential, which could contribute to the understanding of the high incidence of thrombosis in PDAC patients. This finding opens the door to use EVs as biomarkers for assessing thrombosis risk in these patients.
PAVE demonstrated that the MSLN-based nanovaccines could effectively reduce tumor growth and metastasis in PDAC mouse models. These nanovaccines further showed potential synergistic effects when combined with other therapies, such as immune checkpoint inhibitors and conventional chemotherapy in human PDAC organoid co-cultures. Given age-related immune decline, the project also investigated whether senolysis before vaccination could enhance immune responses in aged mice and whether there is an age-related difference in immune response to the MSLN nanovaccine. No significant difference was observed between young and old mice in their response.
The project also advanced methods for analyzing tumor heterogeneity using single-cell data, which is crucial for understanding the complex interactions of tumour, immune cells and fibroblasts within the TME and for identifying new therapeutic targets.
