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Updated: Jul 14, 2026
Single Droplet Digital Polymerase Chain Reaction for Comprehensive and Simultaneous Detection of Mutations in Hotspot Regions
Published on: September 25, 2018
A novel PIK3CA hot-spot mutation in breast cancer patients detected by HRM-COLD-PCR analysis
Saoussen Debouki-Joudi1, Wala Ben Kridis2, Fatma Trifa1
1Department of Cancer Genetics, Laboratory of Molecular Biotechnology of Eukaryotes, Center of Biotechnology of Sfax, University of Sfax, Sfax, Tunisia.
Background:
The PI3K protein is involved in the PI3K/AKT/mTOR pathway. Deregulation of this pathway through PIK3CA mutation is common in various tumors. The aim of this work is to identify hotspot mutation at exons 9 and 20 in Tunisian patients with sporadic or hereditary breast cancer.
Methods:
Hotspot mutations in exon 9 and exon 20 of the PIK3CA gene were identified by QPCR-High Resolution Melting followed by COLD-PCR and sequencing in 63 (42 sporadic cases and 21 hereditary cases) tumor tissues collected from Tunisian patient with breast cancer. MCF7, and BT20 breast cancer cell lines harboring the PIK3CA hotspot mutations E545K and H1047R in exon 9 and exon 20 respectively, were used as controls in HRM experiments.
Results:
PIK3CA hotspot mutations were detected in 66.7% (28 out of 42) of sporadic BC cases, and in 14.3% (3 out of 21) of hereditary BC. The E545K and the H1048Y were the most prevalent mutations identified in patients with sporadic and hereditary BC, whereas the H1047R hotspot mutation was not found in our patients. Statistical analysis showed that PIK3CA mutation associated with an aggressive behavior in patients with sporadic BC, while it's correlated with age, tumor stage and tumor size in the group patients with hereditary breast cancer.
Conclusions:
Our results showed a novel PIK3CA hotspot mutation in Tunisian breast cancer patients detected by HRM-COLD-PCR. Moreover, the absence of PIK3CA hotspot mutation associated with good prognosis.
Insights
PIK3CA hotspot mutations were identified in Tunisian breast cancer patients. These mutations were more common in sporadic cases and linked to aggressive behavior, while their absence indicated a good prognosis.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- The PI3K/AKT/mTOR pathway is crucial in cell signaling and frequently deregulated in cancer.
- Mutations in the PIK3CA gene, particularly in hotspot regions, are common drivers in various malignancies.
- Understanding PIK3CA mutation prevalence is vital for targeted therapies in breast cancer.
Purpose of the Study:
- To investigate the frequency and spectrum of PIK3CA hotspot mutations in exons 9 and 20.
- To analyze the association of these mutations with clinicopathological features in Tunisian breast cancer patients.
- To explore the prognostic significance of PIK3CA mutations in sporadic and hereditary breast cancer.
Main Methods:
- Utilized quantitative PCR-High Resolution Melting (QPCR-HRM) combined with COLD-PCR and sequencing.
- Analyzed 63 tumor tissue samples from Tunisian breast cancer patients (42 sporadic, 21 hereditary).
- Employed MCF7 and BT20 cell lines with known PIK3CA mutations as controls.
Main Results:
- PIK3CA hotspot mutations were detected in 66.7% of sporadic and 14.3% of hereditary breast cancer cases.
- The E545K and H1048Y mutations were most prevalent; H1047R was notably absent.
- PIK3CA mutations correlated with aggressive behavior in sporadic BC and with age, stage, and size in hereditary BC.
Conclusions:
- Identified novel PIK3CA hotspot mutations in Tunisian breast cancer using HRM-COLD-PCR.
- The absence of PIK3CA hotspot mutations was associated with a favorable prognosis.
- Findings highlight the importance of PIK3CA mutation analysis for risk stratification and treatment strategies.
