Synthetic bacterial vesicles combined with tumour extracellular vesicles as cancer immunotherapy

Bacterial outer membrane vesicles (OMV) have gained attention as a promising new cancer vaccine platform for efficiently provoking immune responses. However, OMV induce severe toxicity by activating the innate immune system. In this study, University of Gothenburg researchers applied a simple isolation approach to produce artificial OMV that they have named Synthetic Bacterial Vesicles (SyBV) that do not induce a severe toxic response. The researchers also explored the potential of SyBV as an immunotherapy combined with tumour extracellular vesicles to induce anti-tumour immunity. Bacterial SyBV were produced with high yield by a protocol including lysozyme and high pH treatment, resulting in pure vesicles with very few cytosolic components and no RNA or DNA. These SyBV did not cause systemic pro-inflammatory cytokine responses in mice compared to naturally released OMV. However, SyBV and OMV were similarly effective in activation of mouse bone marrow-derived dendritic cells. Co-immunization with SyBV and melanoma extracellular vesicles elicited tumour regression in melanoma-bearing mice through Th-1 type T cell immunity and balanced antibody production. Also, the immunotherapeutic effect of SyBV was synergistically enhanced by anti-PD-1 inhibitor. Moreover, SyBV displayed significantly greater adjuvant activity than other classical adjuvants. Taken together, these results demonstrate a safe and efficient strategy for eliciting specific anti-tumour responses using immunotherapeutic bacterial SyBV.

Production and characterization of E. coli-derived SyBV

(a) Schematic diagram of the isolation of bacterial SyBV. (b) Representative TEM images of natural OMV and SyBV (three biological replicates for each sample and 10 pictures collected for each). Scale bars, 200 nm. (c, d) The number of particles derived from 1 ml culture media (c) and the number of particles per one microgram of vesicular proteins (d) from OMV and SyBV. Data are presented as the mean ± S.E.M. from five independent experiments. ^***P < 0.001 by unpaired two-tailed Student’s t-test. (e, f) Representative electropherograms of DNA (e) and RNA (f) molecules isolated from SyBV in comparison to those from OMV. Three independent experiments, and filled triangles indicate internal markers. (g) Venn diagram of the OMV and SyBV proteomes. The common proteins are divided into three groups – a 1.5-fold increase, a 1.5-fold decrease, and no change – based on relative protein abundance. Two biological replicates were performed per sample, and three technical replicates were performed for each biological replicate. (h) Plot of the log2 value of relative protein abundance from OMV and SyBV. The solid line and dotted lines indicate no change and 1.5-fold changes, respectively. (i) Proteomes of OMV-enriched proteins and SyBV-enriched proteins were analysed by GO cellular component annotations. Note that proteins normally have several GO annotations