Interpretation of somatic POLE mutations in endometrial carcinoma

Abstract

Pathogenic somatic missense mutations within the DNA polymerase epsilon (POLE) exonuclease domain define the important subtype of ultramutated tumours (‘POLE‐ultramutated’) within the novel molecular classification of endometrial carcinoma (EC). However, clinical implementation of this classifier requires systematic evaluation of the pathogenicity of POLE mutations. To address this, we examined base changes, tumour mutational burden (TMB), DNA microsatellite instability (MSI) status, POLE variant frequency, and the results from six in silico tools on 82 ECs with whole‐exome sequencing from The Cancer Genome Atlas (TCGA). Of these, 41 had one of five known pathogenic POLE exonuclease domain mutations (EDM) and showed characteristic genomic alterations: C>A substitution > 20%, T>G substitutions > 4%, C>G substitutions < 0.6%, indels < 5%, TMB > 100 mut/Mb. A scoring system to assess these alterations (POLE‐score) was developed; based on their scores, 7/18 (39%) additional tumours with EDM were classified as POLE‐ultramutated ECs, and the six POLE mutations present in these tumours were considered pathogenic. Only 1/23 (4%) tumours with non‐EDM showed these genomic alterations, indicating that a large majority of mutations outside the exonuclease domain are not pathogenic. The infrequent combination of MSI‐H with POLE EDM led us to investigate the clinical significance of this association. Tumours with pathogenic POLE EDM co‐existent with MSI‐H showed genomic alterations characteristic of POLE‐ultramutated ECs. In a pooled analysis of 3361 ECs, 13 ECs with DNA mismatch repair deficiency (MMRd)/MSI‐H and a pathogenic POLE EDM had a 5‐year recurrence‐free survival (RFS) of 92.3%, comparable to previously reported POLE‐ultramutated ECs. Additionally, 14 cases with non‐pathogenic POLE EDM and MMRd/MSI‐H had a 5‐year RFS of 76.2%, similar to MMRd/MSI‐H, POLE wild‐type ECs, suggesting that these should be categorised as MMRd, rather than POLE‐ultramutated ECs for prognostication. This work provides guidance on classification of ECs with POLE mutations, facilitating implementation of POLE testing in routine clinical care. © 2019 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Clinicopathological and molecular characterisation of ‘multiple‐classifier’ endometrial carcinomas

“…Our initial cohort comprised 3518 tumours, 3353 being classified to a single molecular subtype. Of the remaining 167 tumours, 30 were classified as MMRd– POLE mut ECs and will be reported separately (León‐Castillo et al ), leaving 138 tumours (3.9%) that were assigned a provisional status of multiple‐classifier with abnormal p53 (mutant p53 expression by IHC or a mutation in TP53 ). Stringent quality control including central pathological review resulted in the exclusion of 35 of these ECs for the following reasons: (1) reassignment of p53 immunostaining from mutant to wild‐type pattern ( n = 27, 21 being initially evaluated using tissue microarrays); (2) non‐pathogenic POLE variant ( n = 3); and (3) lack of evidence of MMRd on IHC review ( n = 1) (Figure ).…”

“…A larger number of these triple-classifier cases will be required to study the biological behaviour of these ECs. Until then, considering the results reported by León-Castillo et al [18] on MMRd-POLEmut ECs and the present study, we suggest that these triple-classifiers be classified as POLEmut ECs if they have a pathogenic POLE EDM based on whole-exome sequencing (WES) data or, in the case of absence of WES data, if the POLE EDM corresponds to one of the 11 pathogenic mutations described by León-Castillo et al [18]. In tumours in which the triple-classifier ECs carry a POLE EDM that does not comply with the previous criteria, we recommend that they be classified as MMRd ECs, as discussed in depth by León-Castillo et al [18].…”

“…A total of 2988 ECs had been molecularly classified in previous studies [10,12,13,16,17], in which MMR and p53 status were determined by immunohistochemistry (IHC) and POLE variants by Sanger sequencing or NGS of the complete exonuclease domain (exons [9][10][11][12][13][14] or targeted sequencing of exons 9, 13, and 14. For this current study, all POLE variants were reviewed and tumours were excluded when the POLE variant was not considered pathogenic following the recommended approach presented by León-Castillo et al [18]. Additionally, following the criteria mentioned previously, four tumours reported as having all three molecular features (POLEmut-MMRd-p53abn) but with a non-pathogenic POLE EDM based on review were classified as MMRd-p53abn ECs.…”

p53 immunohistochemistry is an accurate surrogate for TP53 mutational analysis in endometrial carcinoma biopsies

“…Results of the central reference laboratory p53 IHC were compared with results of TP53 mutation status for all cases and after removal of POLE mut and MMRd tumours . Binary analysis was carried out to assess concordance between p53 IHC and TP53 mutation by combining overexpression, complete absence and cytoplasmic patterns into a single abnormal/mutant (p53abn) score.…”

Prognostic Biomarkers in Endometrial Cancer: A Systematic Review and Meta-Analysis

“…In order to translate this system into clinical practice, the ProMisE decision-tree and the TransPORTEC classifications emerged, mainly based on the assessment of three surrogate biomarker tests: (i) POLE sequencing; (ii) immunohistochemistry of mismatch repair proteins (MMR-IHQ); (iii) immunohistochemistry of the p53 protein [17][18][19]. These surrogate biomarkers categorize most ECs according to their molecular classification, but the method is limited by (i) systematic evaluation of the pathogenicity of POLE mutations is required [20]; (ii) p53 immunohistochemistry does not perfectly correlate with TP53 copy-number alterations; (iii) tumors harboring more than one classifying genomic aberration are difficult to classify; (iv) the algorithms do not include the evaluation of the significant heterogeneity seen in the copy-number-low group [21]. Despite these limitations, the scientific and clinical community supports incorporating the TCGA molecular classification in daily clinical practice for classifying EC patients.…”