Abstract A022: Assessment of clonal and subclonal <i>PIK3CA</i> mutations in patients with breast cancer

Abstract Background: PIK3CA missense mutations are commonly present in HR+HER2- breast cancer (BC) tumors, and these mutations vary in frequency. When common, mutations arise early in tumor progression and are deemed to be clonal events. In contrast, rarer subclonal mutations are present in a small subset of tumor cells. BC tumors with two or more PIK3CA mutations appear to respond better to PI3K inhibitors, though PI3K pathway activation may also confer resistance to HER2-targeted therapy in HER2+ BC patients. This study investigates the clonality of PIK3CA double mutations in BC patients to better understand tumor heterogeneity for this important therapy target. Methods: BC samples were analyzed using the OncoExTraTM assay, which identifies somatic mutations and gene fusions through whole-exome, whole-transcriptome sequencing with tumor-normal pairing. PIK3CA clonality was determined by calculating mutation frequency divided by tumor purity for samples with ≥20% tumor purity. Mutations with an adjusted mutation frequency ≥0.25 were classified as clonal, and those &amp;lt;0.25 as subclonal. We classified double PIK3CA mutations into three subgroups: both clonal (CC), both subclonal (SS) or one clonal and one subclonal (CS), and used Fisher’s Exact Test (p≤0.05) to test associations with other biomarkers. Results: Among 2161 BC samples, 739 (34.2%) had a single PIK3CA mutation and 87 (4.0%) had two or more PIK3CA mutations, of which 81 had tumor purity &amp;gt;20%. Among these 81 patient samples, 32 (39.5%) were CC, 20 (24.7%) were SS, and 29 (35.8.%) were CS. Most (72, 88.8%) samples were from primary tumors. The frequency of multiple PIK3CA mutations was 5.4% in HR+HER2- BC, 2.0% in HER2+ BC, and 0.3% in TNBC. The most common double PIK3CA mutations were in exon 20 and exon 9 (n=25, 30.9%), and in exon 20 and exon 1 (n=11, 13.6%). Examination of other biomarkers revealed that none were significantly associated with any of the three subgroups. We also investigated co-occurring alterations in the PI3K/PTEN/AKT/mTOR pathway. Among the 81 patient samples, three had a PTEN and one had an AKT1 mutation. Two of the three PTEN and the single AKT1 mutations were subclonal. Conclusion: Two or more PIK3CA mutations were present in 4.0% of BC patient samples, and in 39.5% of these both mutations were clonal, suggesting these tumors may be particularly sensitive to PI3K inhibitors. In addition, in this cohort 2% of HER2+ BC samples carried two or more PIK3CA mutations, suggesting strong PI3K pathway activation, and possible concomitant resistance to HER2-targeted treatments. Citation Format: Sameer S Udhane, Fadel Alyaqoub, Pawan Noel, Ariane Kemkes, Cynthia A Flannery, Paige E Innis, David W Hall, Snehal G Thakkar, Gargi Basu, Joyce O'Shaughnessy. Assessment of clonal and subclonal PIK3CA mutations in patients with breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A022.