Use of mutation profiles to refine the classification of endometrial carcinomas

Abstract

The classification of endometrial carcinomas is based on pathological assessment of tumour cell type; the different cell types (endometrioid, serous, carcinosarcoma, mixed, undifferentiated, and clear cell) are associated with distinct molecular alterations. This current classification system for high-grade subtypes, in particular the distinction between high-grade endometrioid (EEC-3) and serous carcinomas (ESC), is limited in its reproducibility and prognostic abilities. Therefore, a search for specific molecular classifiers to improve endometrial carcinoma subclassification is warranted. We performed target enrichment sequencing on 393 endometrial carcinomas from two large cohorts, sequencing exons from the following nine genes: ARID1A, PPP2R1A, PTEN, PIK3CA, KRAS, CTNNB1, TP53, BRAF, and PPP2R5C. Based on this gene panel, each endometrial carcinoma subtype shows a distinct mutation profile. EEC-3s have significantly different frequencies of PTEN and TP53 mutations when compared to low-grade endometrioid carcinomas. ESCs and EEC-3s are distinct subtypes with significantly different frequencies of mutations in PTEN, ARID1A, PPP2R1A, TP53, and CTNNB1. From the mutation profiles, we were able to identify subtype outliers, ie cases diagnosed morphologically as one subtype but with a mutation profile suggestive of a different subtype. Careful review of these diagnostically challenging cases suggested that the original morphological classification was incorrect in most instances. The molecular profile of carcinosarcomas suggests two distinct mutation profiles for these tumours: endometrioid-type (PTEN, PIK3CA, ARID1A, KRAS mutations) and serous-type (TP53 and PPP2R1A mutations). While this nine-gene panel does not allow for a purely molecularly based classification of endometrial carcinoma, it may prove useful as an adjunct to morphological classification and serve as an aid in the classification of problematic cases. If used in practice, it may lead to improved diagnostic reproducibility and may also serve to stratify patients for targeted therapeutics.

Figure 1. Mutation profiles of endometrial subtypes

A. Low-grade endometrioid carcinoma, including grade 1 and 2 tumours (n=276); B. High-grade endometrioid carcinoma, grade 3 tumours (n=30); C. Serous carcinoma (n=37); D. Carcinosarcoma (n=42), (+) indicates carcinosarcomas with heterologous differentiation elements; E. Undifferentiated and mixed histology subtypes, (a) undifferentiated carcinomas, (b) mixed low-grade EEC with serous carcinoma, (c) mixed endometrioid and clear cell carcinoma, (d) mixed serous and clear cell carcinoma. Rows indicate genes, columns represent tumour cases. Coloured bars indicate mutations’ including; missense, truncating, indels and splice site mutations. Grey bars indicate no mutations were detected. (*) indicates serous carcinoma outliers with ARID1A mutations; (#) indicates low-grade EECs and EEC-3s mutation outliers with serous-type mutations (TP53 or PPP2R1A).

Figure 2. A case originally diagnosed as serous carcinoma, but with an ARID1A mutation and no TP53 mutation, is a mixed low-grade endometrioid and serous carcinoma (case #1120)

A. A mix of a grade 1 endometrioid (left half ) and high-grade serous (right half) carcinoma, 40X magnification; B. High power (100X) image of histologically distinct low-grade endometrioid carcinoma; C. High power (100X) images of serous carcinoma component, of which the sampling of tumour was used for mutation sequencing; D. Atypical complex hyperplasia in the background endometrium 40X magnification.

Figure 3. Mutational analysis may be an effective tool to classify morphologically problematic cases into biologically relevant treatment groups

Intermediate high-grade cell types tend to be diagnostically challenging cases, often with multiple morphological features of endometrioid and/or serous carcinomas. The addition of mutation profiles can lead to reproducible diagnosis and the future of mutation-based treatment decisions for targeted therapeutics. Blue and red colours indicate distinct mutation profiles for low-grade EEC and serous carcinomas. Yellow indicates the cases were the mutational profiles will aid in separating out the appropriate histological subtype and dictate appropriate treatment options for patients.