Host immunity plays a central and complex role in tumour progression, and the quality and quantity of immune infiltrate at the tumour site is used as a prognostic marker of cancer progression. Therefore, detailed single-cell analysis of tumor infiltrating immune cells, and ultimately liquid biopsy of circulating immune cells, is of vital importance for the development of new and improved immunotherapy strategies, and for diagnosis, prognosis, patient stratification and follow-up. We proposed in OncoSeq to adapt the droplet barcoding microfluidics system, an output from the ERC EVOEVO project, to develop a novel “molecular cytometer” for high-throughput transcriptomic and phenotypic analysis, at the single-cell level, of tumor infiltrating or circulating immune cells. Readout of genotype and phenotype is entirely based on next-generation sequencing (NGS) allowing higher phenotypic multiplexing capacity than fluorescence or mass spectrometry based approaches and simultaneous RNA-sequencing, allowing the creation of high-resolution phenotype-transcriptome maps for tens of thousands of single cells. A first set of proof of concept (PoC) experiments has successfully validated the developed molecular tools, demonstrating that it is possible to discriminate between two cell lines based on their sequencing profiles. Further work is being carried out to finalise the development of this system for use as a powerful tool for cancer prognosis, clinical decision making and monitoring of disease progression, especially in the context of immunotherapy.
