https://orcid.org/0000-0001-7176-5201
Figures
Abstract
Background
The aim of this study was to analyze PIK3CA mutations in exons 9 and 20 in breast cancers (BCs) and their association with clinicopathological characteristics.
Methods
Mutational analysis of PIK3CA exon 9 and 20 was performed by Sanger sequencing in 54 primary BCs of Tunisian women. The associations of PIK3CA mutations with clinicopathological characteristics were analyzed.
Results
Fifteen exon 9 and exon 20 PIK3CA variants were identified in 33/54 cases (61%). PIK3CA mutations including pathogenic (class 5/Tier I) or likely pathogenic (class 4/Tier II) occurred in 24/54 cases (44%): 17/24 cases (71%) in exon 9, 5/24 cases (21%) in exon 20 and 2/24 cases (8%) in both exons. Of these 24 cases, 18 (75%) carried at least one of the three hot spot mutations: E545K (in 8 cases), H1047R (in 4 cases), E542K (in 3 cases), E545K/E542K (in one case), E545K/H1047R (in one case) and P539R/H1047R (in one case). Pathogenic PIK3CA mutations were associated with negative lymph node status (p = 0.027). Age distribution, histological SBR tumor grading, estrogen and progesterone receptors, human epidermal growth factor receptor 2, and molecular classification were not correlated with PIK3CA mutations (p > 0.05).
Citation: Ben Rekaya M, Sassi F, Saied E, Bel Haj Kacem L, Mansouri N, Zarrouk S, et al. (2023) PIK3CA mutations in breast cancer: A Tunisian series. PLoS ONE 18(5): e0285413. https://doi.org/10.1371/journal.pone.0285413
Editor: Avaniyapuram Kannan Murugan, King Faisal Specialist Hospital and Research Center, SAUDI ARABIA
Received: September 19, 2022; Accepted: April 23, 2023; Published: May 17, 2023
Copyright: © 2023 Ben Rekaya et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Data Availability: All relevant data are within the manuscript.
Funding: This study was supported by the Tunisian-Algerian project funded by the Tunisian Ministry of high education and research entitled: “Study of tumor biomarkers for theranostic purposes with the aim of setting up validated molecular tests in clinical practice”: PRD/TN/DZ/21/08. The authors state that the funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
Introduction
Breakthroughs in molecular biology have clearly established that breast cancer (BC) is a cell signaling disease. Phosphatidylinositol 3-kinase/lipid kinase B/mammalian target of the rapamycin (PI3K/AKT/mTOR) pathway is one of the major deregulated pathways in human cancers, especially in BC [1,2].
The p110α catalytic subunit (PI3K p110α) oncogene encoded by PIK3CA is a lipid kinase which regulates cell proliferation, catabolism, cell adhesion and apoptosis. Single base and insertions/deletions (indels) are the most frequent PIK3CA alterations observed in 13% of solid tumors [3]. However, the frequency of PIK3CA mutations differs among populations and varies among cancer types, stages and ethnicity. The role of ethnicity in frequency rate disparity of PIK3CA mutations has been showed in head and neck squamous cell carcinomas [4,5].
PIK3CA gene is mutated in 18–45% of BCs [5], with over 80% of mutations clustering within three hot spots: two of the helical domain (exon 9, commonly E542 and E545) and one of the kinase domain (exon 20, commonly H1047) [5,6].
Two-third of BCs express estrogen and progesterone receptors (ER/PR) and lack human epidermal growth factor receptor 2 (HER2) overexpression, for which endocrine therapy is the primary drug option. However, approximately 30% of BC patients carry mutations in the PIK3CA gene, which are associated with resistance to endocrine therapy [7]. This is due to multiple mechanisms including dysregulated PI3K/AKT/mTOR signaling [8]. With the emergence of PI3K inhibitors, it is important to identify patients who may benefit from this therapy [8]. Alpelisib is an oral alpha-specific PI3K inhibitor administred in combination with fulvestrant for the treatment of postmenopausal women with hormone receptors positive and HER2 negative, PIK3CA-mutated, advanced or metastatic BC with progression after endocrine therapy [9]. Conflicting correlations between PIK3CA mutations and clinicopathological data have been reported [10]. In early-stage disease, PIK3CA mutations are significantly associated with better invasive disease-free, distant disease-free, and overall survivals [11].
To our knowledge, this study is the first to analyze PIK3CA mutations in exon 9 and exon 20 in BCs of Tunisian patients.
Material and methods
Patients, tissue samples, DNA extraction and PCR reaction
Fifty-four Formalin-Fixed Paraffin-Embedded (FFPE) primitive BC specimens were selected from the pathology departments of Military and Charles Nicolle hospitals (Tunis). All the samples were obtained from women who did not receive preoperative treatment. The diagnosis of BC was made on core biopsies in 7 cases (13%), lumpectomies in 15 cases (28%) and mastectomies in 32 cases (59%).
Clinicopathological data were obtained from pathology records. A pathologist reviewed slides and selected areas rich in tumor cells (at least 20%) avoiding poorly fixed and necrotic areas. Selected areas from FFPE tissues of the 47 surgical specimens (lumpectomy/mastectomy) were manually macrodissected using a mechanical punch and were recuperated in sterile Eppendorf tubes. For core biopsies (n = 7), 4 or 5 FFPE sections (5-6μm thickness) were obtained. Blocks of each case were cut with a new blade to avoid carry-over contamination and the bloc holder and the plate of the microtome were disinfected. Only tissue enriched on tumor cells have been collected without healthy tissue or blood samples.
The total DNA was extracted using QIAamp FFPE kit (Qiagen, Germany) according to the manufacturer’s instructions. The nucleic acid concentration and DNA purity were measured using a NanoDrop 1000 (Thermofisher Scientific, Waltman, MA, USA). The double strand DNA was measured by the Denovix fluorometer with dsDNA Broad Range Assay having a standard detection range from 2 to 2000 ng total mass in 200 μl volumes.
Primer pair have been designed using the Primer 3 version 4 software to amplify exon 20 and exon 9 of PIK3CA gene, avoiding the frequent cross-amplification of chromosome 22q (a known PIK3CA pseudogene). Primer sequences, annealing temperatures (Ta) and product lengths are listed in Table 1. A final concentration of 0.4 μM of each primer and 20 to 50 ng of template DNA were used per reaction. Amplification conditions involved a heat-activation step of 15 min at 95°C, followed by 35 cycles of 94°C for 30 seconds, 55°C for 30 seconds and 72°C for 30 seconds followed by a final extension step at 72°C for 30 min to perform entire elongation of all neosynthesized DNA strands. The amplified fragments were visualized in 2% agarose gel stained with EasyStain I (Biomatik).
Ethical considerations
Ethical approval was obtained from the medical Ethics Committee of Charles Nicolle Hospital of Tunis. All patients gave written informed consent for publication of clinical and laboratory data. Patients were fully anonymized.
Sanger sequencing and variant analysis
To remove unused dNTPs and primers, PCR products were purified using the enzymatic method Exo-SAP PCR Product Cleanup Reagent. PCR sequencing has been performed using the Big Dye Terminator Kit V.3.1 (Applied Biosystems, Foster City, CA, USA). To remove both unlabeled and labeled dye, sequencing reactions were purified using BigDye Xterminator Purification (Life Technologies). Sequences analysis was performed in the Applied Biosystem 3500 Genetic Analyzer. Sequence reading was performed using the BioEdit sequence alignment editor. Variants including those with unknown function were annotated using Mutation Taster, Polyphen and SIFT tools and Sequence Variant Nomenclature was performed according to the guidelines of the Human Genome Variation Society (HGVS) using the Mutalyzer program and the reference sequence NM_006218.4. Free variants databases: dbSNP, ClinVar, ClinGen, COSMIC, and the Clinical Knowledge base were queried to identify known variants and published data on clinical significance and to collect the reference identifier (rs) and the cosmic ID of each variant. Alamut Plus tool (from SOPHiA GENETICS) has been used to check annotation and function prediction. Variants classification has been performed according to ACMG/AMP guidelines for somatic sequence variant interpretation [12].
Immunohistochemistry
Data including ER, PR and HER2 immunohistochemical expression were obtained from pathology records. Molecular subtypes were defined based on the St Gallen’s criteria [13]. Chromogenic In Situ Hybridization (CISH) was performed for HER2 score 2+. All HER2 test results were classified according to 2018 ASCO/CAP HER2 testing recommendations [14].
Statistical analysis
Statistical analyses were performed using SPSS 26.0. The χ2 and Fisher’s tests were used to determine associations between pathogenic and likely pathogenic mutations (class 5/Tier I and class 4/Tier II) of PIK3CA gene and clinicopathological features of BCs (Age, lymph node status, ER, PR, HER2 and molecular classification). P-values <0.05 were considered significant.
Results
Clinicopathological characteristics of cases
The mean age of patients was 51 years (31–84). Median tumor size was 3 cm (0.8cm-10 cm). Invasive ductal carcinoma was diagnosed in 50/54 cases (93%) and invasive lobular carcinoma in the remaining 4 cases (7%). Based on the SBR grading system, 4 cases (8%) were grade I, 25 (46%) were grade II, and 25 (46%) were grade III. Lymph node status was known for 45 cases, with 29 cases (64%) having metastatic BC. Immunohistochemically, 38 cases (70%) were ER positive, 36 (67%) were PR positive, and 4 cases (7%) showed overexpression of HER2. BCs were classified as luminal A in 40 cases (74%), luminal B in 5 cases (9%), HER2+ in 4 cases (8%), and triple negative in the remaining 5 cases (9%).
Mutational analysis
Fifteen exon 9 and exon 20 PIK3CA variants were identified in 33 cases (61%): 24/54 (44%) had pathogenic mutations classified as class 5 or 4 and 9/54 (17%) had mutations with uncertain significance (Figs 1 and 2).
Electropherograms of the 15 PIK3CA variants identified in breast cancers (A) Known PIK3CA mutations (B) Novel variants. All sequences are in sense strand. Red rectangle box shows the position of the mutations. The major peak corresponds to the normal nucleotide and the minor peak corresponds to the mutant nucleotide. Altered nucleotide and amino acid positions and related codon substitution are shown of each corresponding sequence.
Pathogenic mutations were observed in 17/24 cases (71%) in exon 9, in 5/24 cases (21%) in exon 20 and in 2/24 cases (8%) in both exons. Of these cases, 18 (75%) carried at least one of the three hot spot mutations: E545K (in 8 cases), H1047R (in 4 cases), E542K (in 3 cases), E545K/E542K (in one case), E545K/H1047R (in one case) and P539R/H1047R (in one case). H1047L was not found. PIK3CA mutations were class 3 in 10/54 cases (19%).
Three cases (13%) had concomitance of class 5 and class 4 mutations; 6 cases (25%) had concomitance of class 5 and class 3 mutations and 15 (62%) had a single pathogenic mutation.
Among the 15 identified PIK3CA variants, 7 were class 5 (tier I): 5 variants (p.P539R; p.E542K; p.E545K; p.H1047R; p.E545A) had clinical evidence and 2 (p.Q546E and p.E542G) had experimental evidence of PIK3CA gain of function. One variant (M1004V) was class 4 with likely PIK3CA gain of function. Five variants had been previously described in databases (H994R, F998C, D1056N, S514G and E1012K) with unknown clinical significance, and one variant L997H had never been previously described (Fig 2). The S514G had been described in a germinal state, and E1012K had been described in melanoma and lung cancers (Table 2). All the six variants were predicted to be pathogenic by at least one In silico analysis software. One variant (E547K) was likely neutral, classified class 2 with unknown function in therapeutic response (Figs 2 and 3 and Table 3).
(A) Frequencies of the fifteen identified PIK3CA variants among breast cancer samples. (B) Proportion of samples with class 5 and 4 mutations. (C) Proportion of samples with concomitant or single mutations. (D) Proportion of samples with pathogenic class 5 and 4 PIK3CA mutations.
A significant association was found between PIK3CA mutations and negative lymph node status (p = 0.027). No association was found between PIK3CA mutations and age, SBR grade, ER and PR status, HER2 overexpression, and molecular classification (Table 4).
Discussion
In this study, we investigated the distribution of PIK3CA mutations in BCs of Tunisian women. We found a high frequency of PIK3CA mutations, particularly in exon 9. The PIK3CA mutated status was associated with negative lymph node status.
In our series, pathogenic and likely pathogenic PIK3CA mutations were identified in 44% of BCs. This rate is higher than that reported in The Cancer Genome Atlas (TCGA) (34.18%) [30]. The frequency of PIK3CA mutations varies according to the population studied and environmental factors. For example, it is reported in 29% of BCs in Japan [31] and in 34% of BCs in India [32]. It also varies with the number of exons analyzed and the techniques used. The highest rate of PIK3CA mutations detected by Sanger sequencing was 46.5% reported in a Chinese series using Sanger sequencing of 8 exons (1, 2, 4, 7, 9, 13, 18, and 20) of the PIK3CA gene [33].
Currently, targeted next generation sequencing (NGS) has become the most commonly used technique. It is recommended by ASCO for the detection of PIK3CA mutations for treatment eligibility for alpelisib among patients with luminal subtype BC [34]. Data concerning the concordance between Sanger sequencing and NGS are limited. Arsenic et al. [35] compared Sanger sequencing and NGS for the detection of PIK3CA hotspot mutations in exon 9 and exon 20 in 184 BCs and reported a concordance rate of 98.4%. NGS allows the identification of multiple mutations simultaneously, avoiding the need to perform sequential individual tests. Sanger sequencing is more expensive and labor-intensive [35,36]. However, it has the advantage of identifying novel variants. Our study identified six novel variants (H994R, L997H, F998C, D1056N, S514G, and E1012K) that have not been previously described in BCs. The E1012K variant has been described in lung cancer [26] and in melanoma [27] with an unknown functional effect.
In BCs, PIK3CA mutations are more commonly clustered in exon 20 than in exon 9, as reported in previous studies [26,28–30]. However, in our study, the prevalence of pathogenic PIK3CA mutations in exon 9 (17/24) was found to be higher than in exon 20 (5/24). Nevertheless, this result needs to be confirmed in larger series. The distribution of hotspot PIK3CA mutations in exon 9 and exon 20 accounted for 75% of class 5 and class 4 PIK3CA mutations in our series, which is similar to the rate reported in the literature [37,38]. The distribution of these PIK3CA hotspot mutations was found to be 55% for H1047R, 20% for E545K, and 11% for E542K [39].
Concomitant PIK3CA mutations are not uncommon in BCs. In our series, 13% of BCs had concomitant class 5 and class 4 PIK3CA mutations, and 25% had concomitance of class 5 and class 3 mutations. Lian et al. [40] reported one case of PIK3CA mutations in both exon 9 and exon 20 (E545K+H1047L) out of 43 mutated BCs. In a Chinese study, 17 of 537 (3.2%) BCs carried two mutations. Two of them had H1047R simultaneous with E542K or E545K [41]. Vasan et al. [42] demonstrated that the presence of double PIK3CA mutations on the same allele increases PI3K activity, which leads to enhanced downstream signal transduction, cell proliferation, and tumor growth. Concomitant mutations could be present in different tumor clones or may be present in the same tumor cell. Heterogeneity of PIK3CA mutational status has been previously described at the single cell level in circulating tumor cells from the same BC’s patient [43].
The association of PIK3CA mutations in BCs with clinicopathological features is controversial. In the present study, PIK3CA mutations were associated with negative nodal status but not associated with age, SBR grade, ER and PR status, HER2 overexpression and molecular classification. However, data from literature are conflicting. In some studies, PIK3CA mutations are associated with positive ER and PR status, negative HER2 expression [44–46], and negative nodal status [45,47,48]. The negative association between PIK3CA mutations and lymph node metastasis may be explained by the fact that actionable mutations in PIK3CA display constitutive activation of Akt [49]. Once Akt is activated, it promotes carcinogenesis in the early stages while suppressing tumor invasion and metastatic potential. It has been demonstrated that bitransgenic mice that express both activated Akt and ErbB2 in the mammary epithelium show increased breast tumor growth and a significant reduction in lung metastasis when compared to transgenic mice that express only activated ErbB2 [50]. Other reports found an association of PIK3CA mutations with positive lymph node status suggesting that activation of the PI3K/Akt pathway may increase the invasion of cancer cells into the lymph nodes [35,51–53].
Conclusion
Our study shows a high frequency of PIK3CA mutations in BCs of Tunisian women, especially in exon 9. PIK3CA mutated status is associated with negative lymph node status. Further investigations should be undertaken in larger series exploring other exons and using more sensitive techniques such as NGS.
Acknowledgments
The authors would like to thank Dr Maher Kharrat and Mrs Ons Azaiez from the research platform of the Faculty of medicine for their help in Sanger sequencing.
References
- 1.
Landscape of Phosphatidylinositol-3-Kinase Pathway Alterations Across 19 784 Diverse Solid Tumors | Cancer Biomarkers | JAMA Oncology | JAMA Network n.d. https://jamanetwork.com/journals/jamaoncology/fullarticle/2532351 (accessed April 29, 2022).
- 2. Murugan AK. Special issue: PI3K/Akt signaling in human cancer. Semin Cancer Biol 2019;59:1–2. pmid:31689493
- 3. Millis SZ, Jardim DL, Albacker L, Ross JS, Miller VA, Ali SM, et al. Phosphatidylinositol 3-kinase pathway genomic alterations in 60,991 diverse solid tumors informs targeted therapy opportunities. Cancer 2019;125:1185–99. pmid:30582752
- 4. Murugan AK, Hong NT, Fukui Y, Munirajan AK, Tsuchida N. Oncogenic mutations of the PIK3CA gene in head and neck squamous cell carcinomas. Int J Oncol 2008;32:101–11. pmid:18097548
- 5. Harlé A, Lion M, Lozano N, Husson M, Harter V, Genin P, et al. Analysis of PIK3CA exon 9 and 20 mutations in breast cancers using PCR-HRM and PCR-ARMS: Correlation with clinicopathological criteria. Oncology Reports 2013;29:1043–52. pmid:23314198
- 6. Mollon LE, Anderson EJ, Dean JL, Warholak TL, Aizer A, Platt EA, et al. A Systematic Literature Review of the Prognostic and Predictive Value of PIK3CA Mutations in HR+/HER2- Metastatic Breast Cancer. Clin Breast Cancer 2020;20:e232–43. https://doi.org/10.1016/j.clbc.2019.08.011.
- 7. Schwartzberg LS, Vidal GA. Targeting PIK3CA Alterations in Hormone Receptor-Positive, Human Epidermal Growth Factor Receptor-2–Negative Advanced Breast Cancer: New Therapeutic Approaches and Practical Considerations. Clinical Breast Cancer 2020;20:e439–49. https://doi.org/10.1016/j.clbc.2020.02.002.
- 8. Hempel D, Ebner F, Garg A, Trepotec Z, Both A, Stein W, et al. Real world data analysis of next generation sequencing and protein expression in metastatic breast cancer patients. Sci Rep 2020;10:10459. pmid:32591580
- 9. Chang D-Y, Ma W-L, Lu Y-S. Role of Alpelisib in the Treatment of PIK3CA-Mutated Breast Cancer: Patient Selection and Clinical Perspectives. TCRM 2021;17:193–207. pmid:33707948
- 10. Tserga A, Chatziandreou I, Michalopoulos NV, Patsouris E, Saetta AA. Mutation of genes of the PI3K/AKT pathway in breast cancer supports their potential importance as biomarker for breast cancer aggressiveness. Virchows Arch 2016;469:35–43. pmid:27059323
- 11. Fan H, Li C, Xiang Q, Xu L, Zhang Z, Liu Q, et al. PIK3CA mutations and their response to neoadjuvant treatment in early breast cancer: A systematic review and meta-analysis. Thorac Cancer 2018;9:571–9. pmid:29575819
- 12. Li MM, Datto M, Duncavage EJ, Kulkarni S, Lindeman NI, Roy S, et al. Standards and Guidelines for the Interpretation and Reporting of Sequence Variants in Cancer: A Joint Consensus Recommendation of the Association for Molecular Pathology, American Society of Clinical Oncology, and College of American Pathologists. J Mol Diagn 2017;19:4–23. pmid:27993330
- 13.
St Gallen molecular subtypes in primary breast cancer and matched lymph node metastases—aspects on distribution and prognosis for patients with luminal A tumours: results from a prospective randomised trial—PMC n.d. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4222553/ (accessed April 5, 2022).
- 14. Wolff AC, Hammond MEH, Allison KH, Harvey BE, Mangu PB, Bartlett JMS, et al. Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer: American Society of Clinical Oncology/College of American Pathologists Clinical Practice Guideline Focused Update. J Clin Oncol 2018;36:2105–22. pmid:29846122
- 15. Ng PK-S, Li J, Jeong KJ, Shao S, Chen H, Tsang YH, et al. Systematic Functional Annotation of Somatic Mutations in Cancer. Cancer Cell 2018;33:450–462.e10. pmid:29533785
- 16. Nykamp K, Anderson M, Powers M, Garcia J, Herrera B, Ho Y-Y, et al. Sherloc: a comprehensive refinement of the ACMG-AMP variant classification criteria. Genet Med 2017;19:1105–17. pmid:28492532
- 17. Gymnopoulos M, Elsliger M-A, Vogt PK. Rare cancer-specific mutations in PIK3CA show gain of function. Proc Natl Acad Sci U S A 2007;104:5569–74. pmid:17376864
- 18. Mankoo PK, Sukumar S, Karchin R. PIK3CA somatic mutations in breast cancer: Mechanistic insights from Langevin dynamics simulations. Proteins 2009;75:499–508. pmid:18951408
- 19. Samuels Y, Wang Z, Bardelli A, Silliman N, Ptak J, Szabo S, et al. High frequency of mutations of the PIK3CA gene in human cancers. Science 2004;304:554. pmid:15016963
- 20. Abubaker J, Jehan Z, Bavi P, Sultana M, Al-Harbi S, Ibrahim M, et al. Clinicopathological analysis of papillary thyroid cancer with PIK3CA alterations in a Middle Eastern population. J Clin Endocrinol Metab 2008;93:611–8. pmid:18000091
- 21. Konstantinova D, Kaneva R, Dimitrov R, Savov A, Ivanov S, Dyankova T, et al. Rare mutations in the PIK3CA gene contribute to aggressive endometrial cancer. DNA Cell Biol 2010;29:65–70. pmid:19839777
- 22. Dogruluk T, Tsang YH, Espitia M, Chen F, Chen T, Chong Z, et al. Identification of Variant-Specific Functions of PIK3CA by Rapid Phenotyping of Rare Mutations. Cancer Res 2015;75:5341–54. pmid:26627007
- 23. Li VSW, Wong CW, Chan TL, Chan ASW, Zhao W, Chu K-M, et al. Mutations of PIK3CA in gastric adenocarcinoma. BMC Cancer 2005;5:29. pmid:15784156
- 24. Loibl S, Majewski I, Guarneri V, Nekljudova V, Holmes E, Bria E, et al. PIK3CA mutations are associated with reduced pathological complete response rates in primary HER2-positive breast cancer: pooled analysis of 967 patients from five prospective trials investigating lapatinib and trastuzumab. Ann Oncol 2016;27:1519–25. pmid:27177864
- 25. Beca F, Krings G, Chen Y-Y, Hosfield EM, Vohra P, Sibley RK, et al. Primary mammary angiosarcomas harbor frequent mutations in KDR and PIK3CA and show evidence of distinct pathogenesis. Mod Pathol 2020;33:1518–26. pmid:32123305
- 26. Capodanno A, Boldrini L, Alì G, Pelliccioni S, Mussi A, Fontanini G. Phosphatidylinositol-3-kinase α catalytic subunit gene somatic mutations in bronchopulmonary neuroendocrine tumours. Oncol Rep 2012;28:1559–66. https://doi.org/10.3892/or.2012.2017.
- 27. Shull AY, Latham-Schwark A, Ramasamy P, Leskoske K, Oroian D, Birtwistle MR, et al. Novel somatic mutations to PI3K pathway genes in metastatic melanoma. PLoS One 2012;7:e43369. pmid:22912864
- 28. Politz O, Siegel F, Bärfacker L, Bömer U, Hägebarth A, Scott WJ, et al. BAY 1125976, a selective allosteric AKT1/2 inhibitor, exhibits high efficacy on AKT signaling-dependent tumor growth in mouse models. Int J Cancer 2017;140:449–59. pmid:27699769
- 29. Juric D, Krop I, Ramanathan RK, Wilson TR, Ware JA, Sanabria Bohorquez SM, et al. Phase I Dose-Escalation Study of Taselisib, an Oral PI3K Inhibitor, in Patients with Advanced Solid Tumors. Cancer Discov 2017;7:704–15. pmid:28331003
- 30. Pereira B, Chin S-F, Rueda OM, Vollan H-KM, Provenzano E, Bardwell HA, et al. The somatic mutation profiles of 2,433 breast cancers refines their genomic and transcriptomic landscapes. Nat Commun 2016;7:11479. pmid:27161491
- 31. Maruyama N, Miyoshi Y, Taguchi T, Tamaki Y, Monden M, Noguchi S. Clinicopathologic analysis of breast cancers with PIK3CA mutations in Japanese women. Clin Cancer Res 2007;13:408–14. pmid:17202311
- 32. Sudhakar N, Priya Doss CG, Thirumal Kumar D, Chakraborty C, Anand K, Suresh M. Deciphering the impact of somatic mutations in exon 20 and exon 9 of PIK3CA gene in breast tumors among Indian women through molecular dynamics approach. J Biomol Struct Dyn 2016;34:29–41. pmid:25679319
- 33. Deng L, Zhu X, Sun Y, Wang J, Zhong X, Li J, et al. Prevalence and Prognostic Role of PIK3CA/AKT1 Mutations in Chinese Breast Cancer Patients. Cancer Res Treat 2019;51:128–40. https://doi.org/10.4143/crt.2017.598.
- 34. Burstein HJ, Somerfield MR, Barton DL, Dorris A, Fallowfield LJ, Jain D, et al. Endocrine Treatment and Targeted Therapy for Hormone Receptor-Positive, Human Epidermal Growth Factor Receptor 2-Negative Metastatic Breast Cancer: ASCO Guideline Update. Journal of Clinical Oncology 2021;39:3959–77. https://doi.org/10.1200/JCO.21.01392.
- 35. Arsenic R, Treue D, Lehmann A, Hummel M, Dietel M, Denkert C, et al. Comparison of targeted next-generation sequencing and Sanger sequencing for the detection of PIK3CA mutations in breast cancer. BMC Clin Pathol 2015;15:20. pmid:26587011
- 36.
A simple and robust real-time qPCR method for the detection of PIK3CA mutations—PubMed n.d. https://pubmed.ncbi.nlm.nih.gov/29523855/ (accessed February 7, 2022).
- 37. Lai Y-L, Mau B-L, Cheng W-H, Chen H-M, Chiu H-H, Tzen C-Y. PIK3CA Exon 20 Mutation is Independently Associated with a Poor Prognosis in Breast Cancer Patients. Ann Surg Oncol 2008;15:1064–9. pmid:18183466
- 38. Keraite I, Alvarez-Garcia V, Garcia-Murillas I, Beaney M, Turner NC, Bartos C, et al. PIK3CA mutation enrichment and quantitation from blood and tissue. Sci Rep 2020;10:17082. pmid:33051521
- 39. Schneck H, Blassl C, Meier-Stiegen F, Neves RP, Janni W, Fehm T, et al. Analysing the mutational status of PIK3CA in circulating tumor cells from metastatic breast cancer patients. Molecular Oncology 2013;7:976–86. pmid:23895914
- 40. Lian J, Xu E-W, Xi Y-F, Wang H-W, Bu P, Wang J, et al. Clinical-Pathologic Analysis of Breast Cancer With PIK3CA Mutations in Chinese Women. Technol Cancer Res Treat 2020;19:1533033820950832. pmid:33047659
- 41. Wu H, Wang W, Du J, Li H, Wang H, Huang L, et al. The distinct clinicopathological and prognostic implications of PIK3CA mutations in breast cancer patients from Central China. CMAR 2019;11:1473–92. https://doi.org/10.2147/CMAR.S195351.
- 42. Vasan N, Razavi P, Johnson JL, Shao H, Shah H, Antoine A, et al. Double PIK3CA mutations in cis increase oncogenicity and sensitivity to PI3Kα inhibitors. Science 2019;366:714–23. https://doi.org/10.1126/science.aaw9032.
- 43. Pestrin M, Salvianti F, Galardi F, De Luca F, Turner N, Malorni L, et al. Heterogeneity of PIK3CA mutational status at the single cell level in circulating tumor cells from metastatic breast cancer patients. Mol Oncol 2015;9:749–57. pmid:25539732
- 44. Saal LH, Holm K, Maurer M, Memeo L, Su T, Wang X, et al. PIK3CA mutations correlate with hormone receptors, node metastasis, and ERBB2, and are mutually exclusive with PTEN loss in human breast carcinoma. Cancer Res 2005;65:2554–9. pmid:15805248
- 45. Kalinsky K, Jacks LM, Heguy A, Patil S, Drobnjak M, Bhanot UK, et al. PIK3CA mutation associates with improved outcome in breast cancer. Clin Cancer Res 2009;15:5049–59. pmid:19671852
- 46. Buttitta F, Felicioni L, Barassi F, Martella C, Paolizzi D, Fresu G, et al. PIK3CA mutation and histological type in breast carcinoma: high frequency of mutations in lobular carcinoma. J Pathol 2006;208:350–5. pmid:16353168
- 47. Cho YA, Ko SY, Suh YJ, Kim S, Park JH, Park H-R, et al. PIK3CA Mutation as Potential Poor Prognostic Marker in Asian Female Breast Cancer Patients Who Received Adjuvant Chemotherapy. Curr Oncol 2022;29:2895–908. pmid:35621626
- 48. Deng L, Chen J, Zhong XR, Luo T, Wang YP, Huang HF, et al. Correlation between Activation of PI3K/AKT/mTOR Pathway and Prognosis of Breast Cancer in Chinese Women. PLoS One 2015;10:e0120511. pmid:25816324
- 49. Isakoff SJ, Engelman JA, Irie HY, Luo J, Brachmann SM, Pearline RV, et al. Breast Cancer–Associated PIK3CA Mutations Are Oncogenic in Mammary Epithelial Cells. Cancer Research 2005;65:10992–1000. pmid:16322248
- 50. Hutchinson JN, Jin J, Cardiff RD, Woodgett JR, Muller WJ. Activation of Akt-1 (PKB-alpha) can accelerate ErbB-2-mediated mammary tumorigenesis but suppresses tumor invasion. Cancer Res 2004;64:3171–8. https://doi.org/10.1158/0008-5472.can-03-3465.
- 51. Palimaru I, Brügmann A, Wium-Andersen MK, Nexo E, Sorensen BS. Expression of PIK3CA, PTEN mRNA and PIK3CA mutations in primary breast cancer: association with lymph node metastases. Springerplus 2013;2:464. pmid:24083111
- 52. Elwy F, Helwa R, El Leithy AA, Shehab El din Z, Assem MM, Hassan NHA. PIK3CA mutations in HER2-positive Breast Cancer Patients; Frequency and Clinicopathological Perspective in Egyptian Patients. Asian Pac J Cancer Prev 2017;18:57–64. pmid:28240010
- 53. Aleskandarany MA, Rakha EA, Ahmed MAH, Powe DG, Paish EC, Macmillan RD, et al. PIK3CA expression in invasive breast cancer: a biomarker of poor prognosis. Breast Cancer Res Treat 2010;122:45–53. pmid:19701705