1P - Detection of heterozygous mutations in circulating tumor cells separated by lateral magnetophoretic microseparator

Date 04 May 2017
Event IMPAKT 2017
Session Welcome reception and Poster Walk
Topics Breast Cancer
Imaging, Diagnosis and Staging
Pathology/Molecular Biology
Presenter Hyungseok Cho
Authors H. Cho1, J. Kim1, C.W. Jeon2, K. Han1
  • 1Nano Science And Engineering, 197 Inje-ro Gimhae Gyeongsangnam-do Inje University, 50834 - Gimhae/KR
  • 2Department Of Surgery, Gospel Hospital, 262 Gamcheon-ro, Busan, 49267 - Busan/KR

Abstract

Body

CTCs get a lot of attention because provide genetic information that enables monitoring of the evolution of cancer and selection of therapeutic strategies that cannot be obtained from tumor biopsy. Despite their importance, current techniques for separating CTCs are limited in terms of their ability to yield high quality CTCs from peripheral blood for use in profiling genetic mutations by DNA sequencing technologies. This paper presents somatic mutation detection from circulating tumor cells separated by the lateral magnetophoretic microseparator (named ‘CTC-µChip’).

To sensitively detect somatic mutations with NGS, purity of CTCs should be higher than 5%. Since previously reported technologies mostly result in purities below 1%. The new technology, which can separate CTCs with a high-purity of larger than 10%, is therefore required. Our group has previously reported the CTC-µChip for separating CTCs with a high-purity. Therefore, we demonstrate that the CTC-µChip can be used for separating CTCs from peripheral blood with a high-purity of > 25% and somatic mutations could be detected from the separated CTCs with NGS.

The nucleated cells in blood are first extracted using RBC lysis and treated by immunomagnetic nanobeads, which can specifically bind on CTCs. Next, CTCs in the nucleated cells are isolated by the CTC-µChip. Then, genomic DNA extracted from the isolated CTCs was multiplied by a commercialized whole genome amplification (WGA) kit and the product was used for detecting somatic mutation with NGS. Analytical experiments using 10, 30, 50 MCF7 breast cancer cell lines spiked into 5 ml peripheral blood showed that CTC recovery of the CTC-µChip is over 90%. Then, the number of residual WBCs in the isolated sample was an average 53. The isolated CTCs results in purities from 6.9% to 67.9%. Using NGS, heterogeneous somatic mutations (PIK3CA and APC) of MCF7 cell line were detected from the isolated sample. This results explained that mutation detection is possible from as few as 2 CTCs per 1 ml of blood.

Consequently, the experiments demonstrated that the CTC-µChip is a suitable device for detecting somatic mutations from CTCs, thereby allowing real-time monitoring of the evolution of cancer.

Clinical trial identification