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Tracking the expression of therapeutic protein targets in rare cells by antibody-mediated nanoparticle labelling and magnetic sorting

Abstract

Molecular-level features of tumours can be tracked using single-cell analyses of circulating tumour cells (CTCs). However, single-cell measurements of protein expression for rare CTCs are hampered by the presence of a large number of non-target cells. Here, we show that antibody-mediated labelling of intracellular proteins in the nucleus, mitochondria and cytoplasm of human cells with magnetic nanoparticles enables analysis of target proteins at the single-cell level by sorting the cells according to their nanoparticle content in a microfluidic device with cell-capture zones sandwiched between arrays of magnets. We used the magnetic labelling and cell-sorting approach to track the expression of therapeutic protein targets in CTCs isolated from blood samples of mice with orthotopic prostate xenografts and from patients with metastatic castration-resistant prostate cancer. We also show that mutated proteins that are drug targets or markers of therapeutic response can be directly identified in CTCs, analysed at the single-cell level and used to predict how mice with drug-susceptible and drug-resistant pancreatic tumour xenografts respond to therapy.

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Fig. 1: The single-cell intracellular protein analysis approach.
Fig. 2: Intracellular protein analysis and the sensitivity of the approach.
Fig. 3: Analysis of clinically relevant intracellular proteins.
Fig. 4: Analysis of c-Myc and vimentin in xenografts and clinical samples.
Fig. 5: Analysis of mutated proteins relevant for therapeutic selection.

Data availability

The main data supporting the results in this study are available within the paper and its Supplementary Information. The raw and analysed datasets generated during the study are too large to be publicly shared, but are available for research purposes from the corresponding authors on reasonable request.

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Acknowledgements

This work was supported by the Canadian Institutes of Health Research (grant no. FDN-148415), the Natural Sciences and Engineering Research Council of Canada (grant no. 2016-06090), the Province of Ontario though the Ministry of Research, Innovation and Science (grant no. RE05-009) and the National Cancer Institute of the National Institutes of Health (grant no. 1R33CA204574). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health or the other funding agencies. We thank A. Joshua for providing the clinical specimens.

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M.L., S.O.K. and E.H.S. conceived and designed the experiments. M.L., Z.W., S.U.A., R.M.M., B.D. and B.G. performed the experiments and analysed the data. All authors discussed the results and contributed to the preparation and editing of the manuscript.

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Correspondence to Shana O. Kelley.

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Labib, M., Wang, Z., Ahmed, S.U. et al. Tracking the expression of therapeutic protein targets in rare cells by antibody-mediated nanoparticle labelling and magnetic sorting. Nat Biomed Eng 5, 41–52 (2021). https://doi.org/10.1038/s41551-020-0590-1

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