Tumor cell-based liquid biopsy using high-throughput microfluidic enrichment of entire leukapheresis product

Avanish Mishra; Shih-Bo Huang; Taronish Dubash; Risa Burr; Jon F Edd; Ben S Wittner; Quinn E Cunneely; Victor R Putaturo; Akansha Deshpande; Ezgi Antmen; Kaustav A Gopinathan; Keisuke Otani; Yoshiyuki Miyazawa; Ji Eun Kwak; Sara Y Guay; Justin Kelly; John Walsh; Linda T Nieman; Isabella Galler; PuiYee Chan; Michael S Lawrence; Ryan J Sullivan; Aditya Bardia; Douglas S Micalizzi; Lecia V Sequist; Richard J Lee; Joseph W Franses; David T Ting; Patricia A R Brunker; Shyamala Maheswaran; David T Miyamoto; Daniel A Haber; Mehmet Toner

doi:10.1038/s41467-024-55140-x

Tumor cell-based liquid biopsy using high-throughput microfluidic enrichment of entire leukapheresis product

Nat Commun. 2025 Jan 2;16(1):32. doi: 10.1038/s41467-024-55140-x.

Authors

Avanish Mishra^#^{1

2}, Shih-Bo Huang^#^{2

3}, Taronish Dubash², Risa Burr², Jon F Edd^{1

2}, Ben S Wittner², Quinn E Cunneely^{1

2}, Victor R Putaturo^{1

2}, Akansha Deshpande^{1

2}, Ezgi Antmen^{1

2}, Kaustav A Gopinathan^{1

2}, Keisuke Otani^{2

4}, Yoshiyuki Miyazawa^{2

4}, Ji Eun Kwak², Sara Y Guay², Justin Kelly^{2

4}, John Walsh^{1

2}, Linda T Nieman², Isabella Galler⁵, PuiYee Chan⁵, Michael S Lawrence^{2

6

7}, Ryan J Sullivan⁵, Aditya Bardia^{5

8}, Douglas S Micalizzi^{2

5}, Lecia V Sequist⁵, Richard J Lee⁵, Joseph W Franses⁵, David T Ting^{2

5}, Patricia A R Brunker⁶, Shyamala Maheswaran², David T Miyamoto^{9

10

11}, Daniel A Haber^{12

13

14}, Mehmet Toner^{15

16}

Affiliations

¹ Center for Engineering in Medicine and Surgery, Massachusetts General Hospital and Harvard Medical School, Boston, MA, 02114, USA.
² Krantz Family Center for Cancer Research, Massachusetts General Hospital Cancer Center and Harvard Medical School, Charlestown, MA, 02129, USA.
³ Howard Hughes Medical Institute, Bethesda, MD, 20815, USA.
⁴ Department of Radiation Oncology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, 02114, USA.
⁵ Division of Hematology Oncology, Massachusetts General Hospital Cancer Center and Harvard Medical School, Boston, MA, 02114, USA.
⁶ Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, 02114, USA.
⁷ Broad Institute of MIT and Harvard, Cambridge, MA, 02142, USA.
⁸ Hematology/Oncology, University of California, Los Angeles, USA.
⁹ Krantz Family Center for Cancer Research, Massachusetts General Hospital Cancer Center and Harvard Medical School, Charlestown, MA, 02129, USA. [email protected].
¹⁰ Department of Radiation Oncology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, 02114, USA. [email protected].
¹¹ Broad Institute of MIT and Harvard, Cambridge, MA, 02142, USA. [email protected].
¹² Krantz Family Center for Cancer Research, Massachusetts General Hospital Cancer Center and Harvard Medical School, Charlestown, MA, 02129, USA. [email protected].
¹³ Howard Hughes Medical Institute, Bethesda, MD, 20815, USA. [email protected].
¹⁴ Division of Hematology Oncology, Massachusetts General Hospital Cancer Center and Harvard Medical School, Boston, MA, 02114, USA. [email protected].
¹⁵ Center for Engineering in Medicine and Surgery, Massachusetts General Hospital and Harvard Medical School, Boston, MA, 02114, USA. [email protected].
¹⁶ Shriners Children's Boston, Boston, MA, 02114, USA. [email protected].

^# Contributed equally.

Abstract

Circulating Tumor Cells (CTCs) in blood encompass DNA, RNA, and protein biomarkers, but clinical utility is limited by their rarity. To enable tumor epitope-agnostic interrogation of large blood volumes, we developed a high-throughput microfluidic device, depleting hematopoietic cells through high-flow channels and force-amplifying magnetic lenses. Here, we apply this technology to analyze patient-derived leukapheresis products, interrogating a mean blood volume of 5.83 liters from seven patients with metastatic cancer. High CTC yields (mean 10,057 CTCs per patient; range 100 to 58,125) reveal considerable intra-patient heterogeneity. CTC size varies within patients, with 67% overlapping in diameter with WBCs. Paired single-cell DNA and RNA sequencing identifies subclonal patterns of aneuploidy and distinct signaling pathways within CTCs. In prostate cancers, a subpopulation of small aneuploid cells lacking epithelial markers is enriched for neuroendocrine signatures. Pooling of CNV-confirmed CTCs enables whole exome sequencing with high mutant allele fractions. High-throughput CTC enrichment thus enables cell-based liquid biopsy for comprehensive monitoring of cancer.

MeSH terms

Aneuploidy
Biomarkers, Tumor / genetics
Biomarkers, Tumor / metabolism
Exome Sequencing
Female
High-Throughput Nucleotide Sequencing / methods
Humans
Leukapheresis* / instrumentation
Leukapheresis* / methods
Liquid Biopsy / methods
Male
Microfluidic Analytical Techniques / instrumentation
Microfluidic Analytical Techniques / methods
Microfluidics / instrumentation
Microfluidics / methods
Neoplasms / genetics
Neoplasms / pathology
Neoplastic Cells, Circulating* / metabolism
Neoplastic Cells, Circulating* / pathology
Prostatic Neoplasms / genetics
Prostatic Neoplasms / pathology
Single-Cell Analysis / methods

Substances

Biomarkers, Tumor

Abstract

MeSH terms

Substances

Grants and funding