Lab Med Online.  2021 Oct;11(4):297-304. 10.47429/lmo.2021.11.4.297.

Detection of Circulating Tumor Cells Using Three-dimensional and Conditionally Reprogrammed Culture Methods

• Park C
• Jeong YJ
• Park SH
• Kim DD
• Jeon CH

• KMID: 2526092
• DOI: http://doi.org/10.47429/lmo.2021.11.4.297

Abstract

Background
Circulating tumor cells (CTCs) are rare in the circulation of patients with tumors and are very difficult to detect. In this study, we cultured CTCs in a three-dimensionally reprogrammed condition using the peripheral blood of patients with cancer.
Methods
We evaluated the sensitivity of a three-dimensional (3D) conditionally reprogrammed cell (CRC) culture method using SNU-484 gastric cancer cells. From April 2017 to January 2018, blood was collected from 30 patients diagnosed with colorectal or breast cancer at the Daegu Catholic University Medical Center. After lysis of the red blood cells, the blood samples were centrifuged to isolate the mononuclear cells. We used Matrigel to create 3D conditions and reprogrammed the isolated cells using a Rho-kinase inhibitor and J2 feeder cells. After 30 days of culture, the cells were collected and RNA was extracted and amplified for the expression of GAPD, hTERT, and MAGE A1-6 using real-time PCR.
Results
The SNU-484 cells were cultured from three cells using the 3D CRC culture method. The cultured blood samples showed some cells or few colonies. The positive rates of hTERT and MAGE A1-6 mRNA were 50% and 30%, respectively. The positive rates of both mRNAs were 60% and 20%, respectively. MAGE and both mRNAs showed significantly increased odds ratios at the tumor, nodes, and metastases (TNM), N, and M stages in all subjects.
Conclusions
As 60% of CTCs were detected in the blood of 30 patients with cancer, the 3D CRC culture method was very effective for CTC culture.

Keyword

Neoplastic cells; Circulating; Breast cancer; Colorectal cancer; Culture

Figure

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  Fig. 1 (A) Cultured cells from a patient with colorectal cancer, captured on day 18 of culture, at 300× magnification. (B) Cultured cells from a patient with breast cancer, captured on day 15 of culture, at 300× magnification. Colorectal tumor cells formed large undifferentiated aggregates while breast tumor cells formed a large sphere.

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