Supplementary Figure S5 from Evolution and Co-occurrence of PI3K Pathway Gene Mutations in Endometrial Carcinoma Molecular Subtypes at the Single-Cell Level

Da Cruz Paula, Arnaud; Zhu, Yingjie; Brown, David N.; Issa Bhaloo, Shirin; Pareja, Fresia; Hoang, Timothy J.; Green, Hunter; Basili, Thais; Dopeso, Higinio; Selenica, Pier; da Silva, Edaise M.; Riaz, Nadeem; Chandarlapaty, Sarat; Abu-Rustum, Nadeem R.; Reis-Filho, Jorge S.; Weigelt, Britta

doi:10.1158/1078-0432.30617836

Supplementary Figure S5 from Evolution and Co-occurrence of PI3K Pathway Gene Mutations in Endometrial Carcinoma Molecular Subtypes at the Single-Cell Level

https://doi.org/10.1158/1078-0432.30617836

journal contribution

posted on 2025-11-14, 08:21 authored by Arnaud Da Cruz Paula, Yingjie Zhu, David N. Brown, Shirin Issa Bhaloo, Fresia Pareja, Timothy J. Hoang, Hunter Green, Thais Basili, Higinio Dopeso, Pier Selenica, Edaise M. da Silva, Nadeem Riaz, Sarat Chandarlapaty, Nadeem R. Abu-Rustum, Jorge S. Reis-Filho, Britta Weigelt

Supplementary Figure S5. p53 and PTEN protein expression in the POLE-mutant EEC131 assessed by immunohistochemistry. Immunohistochemical analysis of case EEC131 of POLE molecular subtype revealed subclonal p53 overexpression (left) and subclonal PTEN loss of expression (right), consistent with the mutational findings by single cell sequencing. Magnification, 10x.

Funding

National Institutes of Health (NIH)

Cycle for Survival

Breast Cancer Research Foundation (BCRF)

History

Related Materials

1.
DOI - Is supplement to Evolution and Co-occurrence of PI3K Pathway Gene Mutations in Endometrial Carcinoma Molecular Subtypes at the Single-Cell Level

ARTICLE ABSTRACT

The PI3K pathway is altered in >85% of endometrioid endometrial carcinomas (EEC), with multiple mutations commonly co-occurring. Yet, the therapeutic effects of single-agent PI3K pathway inhibitors have been limited. We used single-cell sequencing to determine whether co-occurring PTEN, PIK3CA, and/or PIK3R1 somatic mutations in EECs stratified by molecular subtype originated through convergent or linear evolution. Banked frozen EECs with co-occurring PI3K pathway mutations of no specific molecular profile (NSMP; n = 5), mismatch repair–deficient (MMRd; n = 3), and POLE (n = 3) subtypes were selected for single-nucleus DNA sequencing targeting hotspot variants of 64 cancer-related genes and the PTEN, PIK3R1, and PIK3CA coding sequences. EEC cell lines and nonmalignant samples were used to define error rates and filter false-positive calls. Single-nucleus analyses (n = 50,009 cells) revealed that in NSMP EECs, the co-occurring PIK3CA, PIK3R1, and/or PTEN mutations affected nearly all cells through linear evolution. MMRd EECs displayed higher levels of genetic heterogeneity, harboring PI3K pathway gene mutations in subsets of cells ranging from 3.9% to 96%. POLE EECs had the highest level of clonal diversity and harbored multiple, minor subclonal structures in all cases, through convergent evolution. We found a clear distinction between nearly clonal PI3K pathway gene alterations (>95%) and multiple, minor mutually exclusive subclones only affecting 1.4% to 27% of the tumor cells sequenced. Our exploratory, hypothesis-generating analysis suggests that PI3K pathway alterations evolve distinctly in MMRd/POLE compared with NSMP EECs, which may have therapeutic consequences. Further studies on the signaling output and PI3K pathway inhibitor response in EECs with subclonal PI3K pathway alterations are warranted.