Protocol

Expansion of patient-derived circulating tumor cells from liquid biopsies using a CTC microfluidic culture device

  • Nature Protocols volume 13, pages 3458 (2018)
  • doi:10.1038/nprot.2017.125
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Abstract

The development of personalized cancer therapy depends on a robust system to monitor the patient's individual response to anticancer treatment. Anticancer drug efficacy has been tested on circulating tumor cells (CTCs) derived from patient blood samples after ex vivo expansion into CTC clusters. Current attempts to culture these primary cancer cells focus on long-term maintenance under growth factor supplements into cell lines, which usually takes >6 months and results in a CTC expansion efficiency of <20%. We recently developed a simple but unique microfluidics-based culture approach that requires minimal preprocessing (30 min) and does not require prior enrichment of CTCs or depend on the use of growth factor supplements. The approach capitalizes on co-culture of immune cells from the same patient blood sample within specially designed microwells that promote CTC cluster formation within 2 weeks, with an overall cluster formation success rate of 50%. Drug screening is facilitated by the incorporation of a gradient generator for parallel exposure to two or more drugs at various concentrations. Owing to the cost-effectiveness and less-invasive nature of this procedure, routine monitoring of disease progression can be achieved. The described microfluidics system can be operated with a single syringe pump to introduce drug compounds (which takes 6 min), followed by incubation of the CTC clusters for 48 h before analysis. In addition to its applications in biomedical research, the rapid readout of our platform will enable clinicians to assess or predict a patient's response to various therapeutic strategies, so as to enable personalized or precision therapy.

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Acknowledgements

We express our sincere gratitude to all volunteers who participated in this trial and donated blood samples for characterization of our device. The clinical samples and data collection were supported by a Singapore National Medical Research Council (NMRC) grant. This work was also supported by the Mechanobiology Institute and the Singapore–MIT Alliance for Research and Technology (SMART) BioSystems and Micromechanics (BioSyM) Interdisciplinary Research Groups (IRGs), which are funded by the National Research Foundation, Prime Minister's Office, Singapore, under CREATE.

Author information

Author notes

    • Gianluca Grenci
    •  & Ying Bena Lim

    These authors contributed equally to this work.

Affiliations

  1. BioSystems and Micromechanics (BioSyM) IRG, Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore, Singapore.

    • Bee Luan Khoo
    • , Jongyoon Han
    •  & Chwee Teck Lim
  2. Mechanobiology Institute, National University of Singapore, Singapore, Singapore.

    • Gianluca Grenci
    •  & Chwee Teck Lim
  3. Department of Biomedical Engineering, National University of Singapore, Singapore, Singapore.

    • Gianluca Grenci
    • , Ying Bena Lim
    •  & Chwee Teck Lim
  4. Department of Hematology–Oncology, National University Cancer Institute, National University Hospital, Singapore, Singapore.

    • Soo Chin Lee
  5. Cancer Science Institute of Singapore, National University of Singapore, Singapore, Singapore.

    • Soo Chin Lee
  6. Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.

    • Jongyoon Han
  7. Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.

    • Jongyoon Han
  8. Department of Mechanical Engineering, National University of Singapore, Singapore, Singapore.

    • Chwee Teck Lim

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Contributions

B.L.K. and G.G. conceived and designed the experiments. B.L.K., G.G. and Y.B.L. performed the experiments. C.T.L., S.C.L. and J.H. contributed by providing reagents, materials and analysis tools. B.L.K., G.G., Y.B.L., S.C.L., J.H. and C.T.L. wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Bee Luan Khoo or Jongyoon Han or Chwee Teck Lim.

Integrated supplementary information

Supplementary information

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    Supplementary Figures and Tables

    Supplementary Figures 1–12 and The Supplementary Tutorial.

Excel files

  1. 1.

    Supplementary Table

    Supplementary Table 1

Zip files

  1. 1.

    Supplementary Data 1

    Design of photolithography mask for gradient generator layer.

  2. 2.

    Supplementary Data 2

    Design of photolithography mask for microwell layer.

  3. 3.

    Supplementary Data 3

    CAD file for aluminum mold for barrier layer.

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