Determining the changes induced in mitochondrial organisation by chemotherapeutic compounds that promote immunogenic cell death in tumour cells grown on physiological extracellular matrices

The objective of this proposal is to utilise high resolution imaging technologies to investigate the involvement of the extracellular matrix in the production of immunogenic cell death (ICD). ICD is a recently described, poorly understood process involving tumour cell death events that alter the tumour-derived secretome to act as activating signals for anti-tumour immune responses. Dendritic cells (DCs) are pivotal to ICD, as they alter their biology upon encountering specific molecular patterns, together with their ability to present antigens to T cells. T cell-derived extracellular vesicles (EVs) are taken up by DCs during cognate immune contacts and promote changes in DCs, such as a migratory phenotype and an increased ability to cross-present antigens. Our preliminary –OMICs data reveal that tumour-derived EVs during ICD contain mitochondrial and endolysosomal proteins relevant for mitochondrial structure and lipid balance, similar to T cell-derived EVs and inducing a similar immunogenic maturation of DCs. In addition, the adhesion of tumour cells to the extracellular matrix through integrins may regulate mitochondrial dynamics and thus EV production. We have identified reduced adhesion of tumour cells undergoing ICD versus non-ICD.

We want to use state-of-the-art cryocorrelative soft-X-ray tomography to study changes in metabolic organelles such as mitochondria and lipid droplets produced during ICD when tumour cells are grown on extracellular matrix-based substrates, mainly fibronectin, laminin and collagen, and treated with chemotherapeutic compounds leading to ICD vs non-ICD. Our ultimate goal is to discover specific patterns in the localisation and structure of organelles, which will be correlated with functional features and –OMICs, such as quantitative proteomic profiles of the secretome, isolated mitochondria and subcellular fractions from tumour cells upon ICD versus non-ICD. The image and –OMICs features found will determine which chemotherapeutics enhance immunogenicity, helping the discovery of effective therapies against cancer through combined immunotherapies.