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IMPAKT 2015 News: A Circulating Tumour Cell Liquid Biopsy May Be Used for Non-Invasive Mutational Analysis in Patients with Breast Cancer

Circulating tumour cells in the blood of patients with hormone receptor positive breast cancer contain both wild-type and mutated PIK3CA
07 May 2015
Breast cancer;  Translational research
Abstract 41O IMPAKT 2015

PIK3CA mutational analysis in CTCs of patients with metastatic hormone receptor-positive breast cancer reveals intra-patient homo- and heterogeneity.
© Bram De Laere

A novel methodology was used successfully in the non-invasive evaluation of the mutation frequency of the PIK3CA gene in circulating tumour cells (CTCs) found in the blood of patients with metastatic hormone receptor-positive breast cancer, according to findings presented during a Best abstracts session at the IMPAKT Breast Cancer Conference held 7–9 May, 2015 in Brussels, Belgium.

PIK3CA is frequently mutated in hormone receptor-positive metastatic breast cancer and may be a marker for disease progression.

Bram De Laere, headed a research team from the Center for Oncological Research - Campus GZA Sint Augustinus, University of Antwerp, Antwerp, Belgium in conducting this study to determine the PIK3CA genotype in circulating tumour cells (CTCs) at the single cell level. CTCs were detected in the blood of patients with metastatic hormone-receptor positive breast cancer. The investigators also compared the frequency of PIK3CA mutations in CTCs with primary tumour samples to establish whether CTCs could be used to depict the PIK3CA mutational status of the primary tumour.

CellSearch and DEPArray techniques were used to purify 249 single CTCs and 148 groups of CTCs (range: 5 to 120 cells per group) and 58 white blood cells (WBC; range: 4 - 20) from the peripheral blood of 29 patients with metastatic oestrogen receptor-positive/progesterone receptor-positive/HER2-negative breast cancer. The investigators also purified circulating cell-free DNA (cfDNA) from cell preparation tube (CPT)-collected plasma and genomic DNA from archival formalin-fixed paraffin embedded (FFPE) tissue sections from primary tumour as comparators. Mutation analysis was performed by targeted 454 pyrosequencing.

Mutation analysis in this study demonstrates a clinical application of a semi-automated methodology

PIK3CA mutations were present in 59.2% of archival primary tumour samples and showed poor agreement with 21 cfDNA samples (42.8% disparity; κ=0.113) and fair agreement with 22 CTCs (27.2% disparity; κ=0,394). Comparison of 22 each of CTCs and temporally matched cfDNA samples revealed moderate agreement (22.7% disparity; κ=0.409). A concordant PIK3CA status across all compartments was observed in 55.5% of 18 samples. At the used sequencing depth, cfDNA failed to detect PIK3CA mutations in 4 (22,2%) cases.

Gain of mutation was observed in 4 (22.2%) of 18 patients having a wild-type primary tumour plus mutant CTCs and cfDNA upon disease progression.

A wild-type sequence found in the WBCs of all patients, which indicated a high specificity with the detected variants and indicated the tumorigenic nature of the variants. In depth analysis performed among the CTCs revealed PIK3CA mutations that demonstrated both homo- and heterogeneity with cases, which suggested the presence of both mutant and wild-type CTCs in the population. Additionally, unique double-mutated CTCs were detected in 10 (38.4%) out of 26 patients. This analysis showed intra-patient heterogeneity with cases of mutation persistence and discordance when comparing to the temporally unmatched primary tumour.

Prof. W. Fraser Symmans from the University of Texas M.D. Anderson Cancer Center, Houston, USA, who discussed the study results, said that feasibility was demonstrated for single CTC measurements. PIK3CA genomic status in tumour tissue appears to be more concordant with status in CTCs than with status in cfDNA. Sometimes the PIK3CA genomic status was different in the circulation from the status in the tumour tissue. It was observed more often if tumour tissue had wild-type status. It may be more common with cfDNA than with CTCs. Prof. Symmans looks forward to a larger study with matched primary and metastatic tissues, and blood samples.


PIK3CA status may change between primary (early) and metastatic (advanced) disease. It emphasises the necessity of assessing the PIK3CA status in a real-time manner. In this study, disparity in PIK3CA status between early and advanced disease was found in 4 patients.

Analysis of CTCs was more useful than plasma analysis to assess PIK3CA mutational status in a non-invasive manner in women with advanced hormone receptor positive breast cancer. CTCs showed better agreement with primary tumour sample DNA in terms of mutational status.

The authors concluded that this study demonstrated the utilisation of a CTC liquid biopsy, thereby paving the way towards the application of a more personalised medicine in the management of patients with metastatic cancer.


41O Exploring the intra-patient PIK3CA mutational heterogeneity of circulating tumour cells by massive parallel sequencing in patients with metastatic hormone receptor-positive breast cancer

Last update: 07 May 2015

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