Supervisors info:
Ευρύκλεια Λιανίδου, Καθηγήτρια Τμήματος Χημείας ΕΚΠΑ Επιβλέπουσα, Κωνσταντίνα Γαλανοπούλου, Αναπληρώτρια Καθηγήτρια Τμήματος Χημείας ΕΚΠΑ, Χρήστος Κρούπης, Επίκουρος Καθηγητής ΕΚΠΑ
Summary:
Mutations in the PIK3CA gene have recently been described in a number of
cancers.
In the present study we used High Resolution Melting Analysis (HRMA), method
that was previously developed in our laboratory1, for the detection of two hot
spot mutations in exon 9 of the PIK3CA gene, in breast cancer patients with.
The HRMA was applied in: a) 106 samples of genomic DNA (gDNA) isolated from
FFPEs of patients with breast cancer who had received Trastuzumab treatment.
This method detected PIK3CA mutations in exon 9 in 12% (13/106) of these
samples. b) samples of gDNA isolated from circulating tumor cells (CTCs) of
patients with operable breast cancer, patients with metastatic breast cancer
and healthy donors (57, 34, 26 respectively) c) samples of cDNA of patients of
operable breast cancer, patients with metastatic breast cancer and healthy
donors (57, 9, 21 respectively) and d) samples of cfDNA isolated from
peripheral blood of patients with operable breast cancer, patients with
metastatic breast cancer and healthy donors (25, 45, 12 respectively). This
method detected PIK3CA mutations in exon 9 in 4.4 % (2/45) of cfDNA samples of
patients with metastatic breast cancer as well as in 4 % (1/25) of cfDNA
samples of patients with operable breast cancer. No mutation was detected in
all samples of gDNA and cDNA.
We then made an attempt to develop a new methodology based on the combination
of ARMS-PCR with HRMA. After the optimization of all steps the final protocol
had a sensitivity of 0.25 %. However, the application of this protocol in DNA
isolated from FFPE samples, which was previously analysed by DNA Sequencing,
indicated that the specificity of this methodology was not satisfying enough in
order to be applied in clinical samples.
Keywords:
PIK3CA, HRMA, circulating tumor cells (ctc), ARMS-PCR, Exon 9
