Abstract P3-01-22: Isolation and expansion the circulating tumor cells from patients' peripheral blood <i>in vitro</i>
Authors: W Zhou, C Zou, P Zhao, C Liu, H Zhang, H Hu, C Zhong, Y Zhang, D Zhou
Journal: Cancer Research (2019)

Abstract Background: Circulating tumor cells (CTCs) are rare tumor cells disseminated in peripheral blood from either original sites or metastatic sites. Current studies on CTCs are mainly focused on its diagnosis and prognosis significances by liquid biopsy, although controversial results were reported elsewhere. Actually, as a few studies showed that CTCs would be a kind of intriguing drug targets and more beneficial for cancer patients if there genomic and proteomic information could be deciphered dynamically. However, the rarity, challenging isolation as well as the primary culture procedures limited the analysis of CTCs insightfully. Methods: All patients' samples were referred to the regulation of ethnic commission of Shenzhen People's Hospital and informed parental consent was then collected also for this study. The density gradient centrifugation was used to obtain PBMCs. Cells adherent and suspension culture for collected the circulating tumor cells. Immunofluorescence, kayyotypeing, mude mouse tumorigenesis were verify the tumor cells. Also characteristic and functional of CTCs were analyzed. Results: It was showed that the morphology of CTC-3 is clustered growth and the CTCs were DAPI-positive, Cytokeratin-positive, and CD45-negative with the immunofluorescence identification. The karyotyping results showed that the chromosome number of CTC-3 varied from 56-60 and the chromosomes were aneuploid. This cell line, which was named as CTC-3, formed clones and displayed stable epithelial phenotype and showed stem-cell like properties. CTC-3 had larger nuclear/cytoplasmic ratio and limitless replicative potential. The growth curve of CTC-3 was similar with that of MCF-7, and CTC-3 grew faster than MCF-7 since the fifth day. Furthermore, the sphere formation ability of CTC-3 was significantly higher than MCF-7 in vitro. Besides, the tumorigenesis of CTC-3 cells was almost the same as MCF-7 in vivo by the analysis of xenografting in immunodeficient mice. STR analysis showed that the similarity of CTC-3 and MCF-7 on the standard site alignment is 20%, which indicated that CTC-3 was a totally new breast cancer cell line. For drug resistance, CTC-3 cells were more resistant to anti-tumor drugs after 3D culture compared with 2D, survive rates of 3D-CTC-3 cells after treatment were more than 80%. Finally, the RT-PCR results shows that the expression of Nanog, E-cadherin and ABCG2 were significantly higher in CTC3, also Oct3/4, CD44, CD133, Vimentin, MMP2, MMP9 no significant differences were found between other genes. Conclusion: We have successfully isolated and expanted a new breast cancer cell line from the breast cancer patient, which could provide more drug sensitivity information of the patient and open up novel avenues towards the understanding of breast cancer. Citation Format: Zhou W, Zou C, Zhao P, Liu C, Zhang H, Hu H, Zhong C, Zhang Y, Zhou D. Isolation and expansion the circulating tumor cells from patients' peripheral blood in vitro [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-01-22.