PIK3CA mutational status in circulating tumor cells can change during disease recurrence or progression in patients with breast cancer.
OBJECTIVE
Molecular characterization of circulating tumor cells (CTC) is crucial for the investigation of molecular-targeted therapies while PIK3CA somatic mutations play a crucial role in therapy response. We investigated the presence of PIK3CA mutations in CTC and whether this is associated with clinical outcome.


METHODS
We developed and validated an ultrasensitive methodology for the detection of PIK3CA mutations that is based on a combination of allele-specific, asymmetric rapid PCR and melting analysis. We analyzed PIK3CA hotspot mutations in: (i) a training group consisting of EpCAM-positive CTC fraction from 37 patients with clinically confirmed metastasis, and 26 healthy female volunteers and 15 primary breast tumor tissues and (ii) an independent group consisting of EpCAM-positive CTC fraction from 57 metastatic and 118 operable breast cancer patients and 76 corresponding primary tumors.


RESULTS
The assay could detect 0.05% of mutated dsDNA in the presence of 99.95% wtDNA for both exons (9 and 20) and was highly specific (0/26 healthy donors). PIK3CA mutations were identified in EpCAM-positive CTC in 20 of 57(35.1%) and in 23 of 118 (19.5%) patients with metastatic and operable breast cancer, and in 45 of 76(59.2%) corresponding FFPEs. Our data indicate that PIK3CA mutational status in CTCs can change during disease progression and is associated with worse survival (P = 0.047).


CONCLUSIONS
PIK3CA hotspot mutations are present at a relatively high frequency in CTCs and their presence is associated with worse survival in patients with breast cancer with metastasis. Evaluation of PIK3CA mutational status in CTCs is a strategy with potential clinical application.
Analysis of Circulating Tumor Cells, Laboratory of Analytical Chemistry, Department of Chemistry, University of Athens, Athens, Greece.
Athina
A
Markou
Analysis of Circulating Tumor Cells, Laboratory of Analytical Chemistry, Department of Chemistry, University of Athens, Athens, Greece.
Sofia
S
Farkona
Analysis of Circulating Tumor Cells, Laboratory of Analytical Chemistry, Department of Chemistry, University of Athens, Athens, Greece.
Christina
C
Schiza
Analysis of Circulating Tumor Cells, Laboratory of Analytical Chemistry, Department of Chemistry, University of Athens, Athens, Greece.
Tonia
T
Efstathiou
Oncology Unit and Pathology Department, Helena Venizelou Hospital, Athens, Greece.
Sophia
S
Kounelis
Oncology Unit and Pathology Department, Helena Venizelou Hospital, Athens, Greece.
Nikos
N
Malamos
Laboratory of Tumor Cell Biology, Medical School, University of Crete, Heraklion, Greece.
Vassilis
V
Georgoulias
Analysis of Circulating Tumor Cells, Laboratory of Analytical Chemistry, Department of Chemistry, University of Athens, Athens, Greece. lianidou@chem.uoa.gr.
Evi
E
Lianidou
D001943:Breast Neoplasms / Q000175:diagnosis / Q000235:genetics* / Q000401:mortality / Q000473:pathology*; D016022:Case-Control Studies; D045744:Cell Line, Tumor; D058534:Class I Phosphatidylinositol 3-Kinases; D004252:DNA Mutational Analysis / Q000379:methods; D018450:Disease Progression; D005091:Exons; D005260:Female; D006801:Humans; D009154:Mutation; D009364:Neoplasm Recurrence, Local; D009367:Neoplasm Staging; D009360:Neoplastic Cells, Circulating / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics*; D011379:Prognosis; D012680:Sensitivity and Specificity
D019869:Phosphatidylinositol 3-Kinases; D058534:Class I Phosphatidylinositol 3-Kinases; C484760:PIK3CA protein, human