Isolation of viable CTCs from leukapharesis product via Parsortix system enables subsequent culture

Background:

Solid tumors are constantly releasing cells into the circulatory system. They are genotypically and phenotypically different from the primary tumor. Viable CTCs are of high interest to obtain therapeutically relevant information. Their extremely low frequency is one of the main limiting factors to use them for further characterization.

To overcome this challenge we tested to isolate viable CTCs from leukapharesis products obtained from breast cancer patients.

Material/methods:

31 leukapharesis samples and matched peripheral blood samples were collected and CTC numbers were determined by CellSearch® analysis. Viable cells were enriched from DLA product with Parsortix system. Genomic DNA of single cultured CTCs was isolated and amplified by whole genome amplification. Array-based comparative genomic hybridization of single cells was performed to analyze for genomic aberrations. Resulting profiles were compared to genomic aberration profiles of CTCs isolated before in vitro culture.

Results:

CTCs were detected in eleven peripheral blood and matched leukapharesis samples (1/14 UICC 1 – 2 and 10/17 UICC 3 – 4) and in ten more leukapharesis sample whose corresponding blood sample was CTC-negative (5/13 UICC 1 – 2 and 5/7 UICC 3 – 4).

We could grow viable CTCs from two out of four CTC-positive leukapharesis samples from metastasized patients as determined by immune fluorescence analysis.

The growing cells harbor genomic anomalies confirming their malignant origin. Most aberrations are widely identical to the aberrations detected in uncultured CTCs.

Conclusions:

Leukapharesis provides greater numbers of viable CTCs which can be enriched with Parsortix system in order to enable their in vitro cultivation. This workflow will allow functional studies.

No conflict of interest has been declared by the author(s).