Abstract 900P
Background
Circulating tumour cells (CTCs) are a source of prognostic and predictive biomarkers across multiple cancers, including head and neck squamous cell carcinoma (HNSCC). Characterisation of CTCs has historically focussed on RNA expression. Although a powerful approach, important cellular changes can occur in the absence of transcriptional modification through processes such as alterations in protein stability, subcellular localisation or post-translational modification such as phosphorylation. To increase the number of protein markers that can be measured per cell we developed and validated a protocol to apply mass cytometry to CTCs enriched from blood using Parsortix technology.
Methods
Blood samples collected from a pilot cohort of advanced HNSCC patients and processed using the microfluidic-based Parsortix CTC enrichment device were used for panel validation. A 42-marker mass cytometry panel was developed and optimised using a HNSCC cell line model. The mass cytometry panel comprised of lineage markers and druggable targets such as PD-L1, EGFR and CTLA4. In addition, the activation status (phosphorylation) of key signalling pathways, including STAT, ERK, AKT and PARP, was assessed.
Results
The combination of Parsortix-enrichment and mass cytometric analysis allowed successful identification of HNSCC cell lines in spiked donor samples. In patient samples, CTCs were identified and grouped into phenotypic sub-populations. Phospho-markers allowed identification of ‘activated cells’.
Conclusions
We present novel data demonstrating the utility of mass cytometry to characterise CTCs in HNSCC; allowing a previously unseen view of functional marker expression at the single cell level. Our protocol was not only able to identify CTCs in patient samples, but also phenotypic sub-groups within the CTC population. Furthermore, we demonstrate how multiple druggable targets can be detected in these sub-groups. This approach has high clinical translational potential given the cost and complexity of alternative sequencing approaches. Our data serves as a foundation for further research and clinical trials.
Clinical trial identification
Editorial acknowledgement
Legal entity responsible for the study
The authors.
Funding
Cancer Research UK.
Disclosure
All authors have declared no conflicts of interest.