Collection of a blood sample defined by the term “blood liquid biopsy” is commonly used to detect diagnostic, prognostic, and therapeutic decision‐making markers of metastatic tumors including circulating tumor cells (CTCs). Many tumors also release CTCs and other markers into lymph fluid, but the utility of lymphatic markers largely remains unexplored. Here, we introduce lymph liquid biopsy through collection of peripheral (afferent) and central (thoracic duct [TD]) lymph samples and demonstrates its feasibility for detection of stem‐like CTCs potentially responsible for metastasis development and tumor relapse. Stemness of lymphatic CTCs (L‐CTCs) was determined by spheroid‐forming assay in vitro. Simultaneously, we tested blood CTCs by conventional blood liquid biopsy, and monitored the primary tumor size, early metastasis in a sentinel lymph node (SLN) and distant metastasis in lungs. Using a mouse model at early melanoma stage with no distant metastasis, we identified stem‐like L‐CTCs in lymph samples from afferent lymphatic vessels. Since these vessels transport cells from the primary tumor to SLN, our finding emphasizes the significance of the lymphatic pathway in development of SLN metastasis. Surprisingly, in pre‐metastatic disease, stem‐like L‐CTCs were detected in lymph samples from the TD, which directly empties lymph into blood circulation. This suggests a new contribution of the lymphatic system to initiation of distant metastasis. Integration of lymph and blood liquid biopsies demonstrated that all mice with early melanoma had stem‐like CTCs in at least one of three samples (afferent lymph, TD lymph, and blood). At the stage of distant metastasis, spheroid‐forming L‐CTCs were detected in TD lymph, but not in afferent lymph. Altogether, our results demonstrated that lymph liquid biopsy and testing L‐CTCs holds promise for diagnosis and prognosis of early metastasis. © 2020 International Society for Advancement of Cytometry
Most cancer patients die from metastatic disease as a result of a circulating tumor cell (CTC) spreading from a primary tumor through the blood circulation to distant organs. Many studies have demonstrated the tremendous potential of using CTC counts as prognostic markers of metastatic development and therapeutic efficacy. However, it is only the viable CTCs capable of surviving in the blood circulation that can create distant metastasis. To date, little progress has been made in understanding what proportion of CTCs is viable and what proportion is in an apoptotic state. Here, we introduce a novel approach toward in situ characterization of CTC apoptosis status using a multicolor in vivo flow cytometry platform with fluorescent detection for the real‐time identification and enumeration of such cells directly in blood flow. The proof of concept was demonstrated with two‐color fluorescence flow cytometry (FFC) using breast cancer cells MDA‐MB‐231 expressing green fluorescein protein (GFP), staurosporine as an activator of apoptosis, Annexin‐V apoptotic kit with orange dye color, and a mouse model. The future application of this new platform for real‐time monitoring of antitumor drug efficiency is discussed. © 2019 International Society for Advancement of Cytometry
more » « less- NSF-PAR ID:
- 10458005
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- Cytometry Part A
- Volume:
- 95
- Issue:
- 6
- ISSN:
- 1552-4922
- Page Range / eLocation ID:
- p. 664-671
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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