Updated nanoscale tweezers for single-cell biopsies applied to pre-adapted dormant breast cancer cells.

Abstract

The nanotweezers are a spatially controlled minimally invasive nanoscale tweezer that can extract biomolecules with single-molecule precision from individual living cells, using dielectrophoresis (DEP) to trap molecules by subjecting them to a non-uniform electric field. Our system avoids aspirating cytoplasmic fluid, allowing for the preconcentration of the analyte in real time. The aim was to create a step changing technology for the next generation of minimally invasive biopsy tools for high throughput RNA sampling and sequencing whilst keeping the cell alive. In theme with keeping the system as minimally invasive as possible, and since the electrochemical conditions could not be altered without causing unwanted electrothermal heating, the surface glass on the nanotweezers were modified to contain a poly-Thymine oligomer (Poly T). Poly T was chosen to act as a broad-spectrum tool to enhance the overall trapping capacity per biopsy. The modification was successful and significantly increased the trapping capacity per biopsy. Therefore, the updated nanotweezers were used to acquire insight into the transcriptional changes that occur in treatment resistant luminal breast cancer cells. These cells have a higher probability to survive exposure to endocrine therapies, surgical resection, and start of adjuvant treatment. Of these cells, only a handful of plastic cells survive and become enriched with the transcriptional signature of pre-adaptation previously identified in our group. These cells can then accumulate genetic changes and further transcriptional re-wiring seen in completely treatment resistant cells, eventually leading to metastasis. The updated nanotweezers were used to give a continuous transcriptional dataset for 10 of those cells over a period of 28-days observing 7 genes of interest. We propose our modified nanotweezers as a novel system extracting cytoplasmic mRNA for continuous live single-cell RNA sequencing of luminal breast cancer cells.