Microcystic elongated and fragmented (MELF) pattern of invasion: Molecular features and prognostic significance in the PORTEC-1 and -2 trials.

OBJECTIVE: Microcystic, elongated fragmented (MELF) pattern of myometrial invasion is a distinct histologic feature occasionally seen in low-grade endometrial carcinomas (EC). The prognostic relevance of MELF invasion was uncertain due to conflicting data, and it had not yet appropriately been studied in the context of the molecular EC classification. We aimed to determine the relation of MELF invasion with clinicopathological and molecular characteristics, and define its prognostic relevance in early-stage low/intermediate risk EC. METHODS: Single whole tumor slides of 979 (85.8%) out of 1141 (high)intermediate-risk EC of women who participated in the PORTEC-1/-2 trials were available for review. Clinicopathological and molecular features were compared between MELF invasion positive and negative cases. Time-to-event analyses were done by Kaplan-Meier method, log-rank tests and Cox' proportional hazards models. RESULTS: MELF invasion was found in 128 (13.1%) cases, and associated with grade 1-2 histology, deep myometrial invasion and substantial lymph-vascular space invasion (LVSI). 85.6% of MELF invasion positive tumors were no-specific-molecular-profile (NSMP) EC. NSMP EC with MELF invasion were CTNNB1 wild type in 92.2% and KRAS mutated in 24.4% of cases. Risk of recurrence was lower for MELF invasion positive as compared to MELF invasion negative cases (4.9% vs. 12.7%, p = 0.026). However, MELF invasion had no independent impact on risk of recurrence (HR 0.65, p = 0.30) after correction for clinicopathological and molecular factors. CONCLUSIONS: MELF invasion has no independent impact on risk of recurrence in early-stage EC, and is frequently observed in low-grade NSMP tumors. Routine assessment of MELF invasion has no clinical implications and is not recommended.

Ten-year results of the PORTEC-2 trial for high-intermediate risk endometrial carcinoma: improving patient selection for adjuvant therapy.

BACKGROUND: PORTEC-2 was a randomised trial for women with high-intermediate risk (HIR) endometrial cancer, comparing pelvic external beam radiotherapy (EBRT) with vaginal brachytherapy (VBT). We evaluated long-term outcomes combined with the results of pathology review and molecular analysis. METHODS: 427 women with HIR endometrial cancer were randomised between 2002-2006 to VBT or EBRT. Primary endpoint was vaginal recurrence (VR). Pathology review was done in 97.4%, combined with molecular analysis. RESULTS: Median follow-up was 116 months; 10-year VR was 3.4% versus 2.4% for VBT vs. EBRT (p = 0.55). Ten-year pelvic recurrence (PR) was more frequent in the VBT group (6.3% vs. 0.9%, p = 0.004), mostly combined with distant metastases (DM). Ten-year isolated PR was 2.5% vs. 0.5%, p = 0.10, and DM 10.4 vs. 8.9% (p = 0.45). Overall survival for VBT vs. EBRT was 69.5% vs. 67.6% at 10 years (p = 0.72). L1CAM and p53-mutant expression and substantial lymph-vascular space invasion were risk factors for PR and DM. EBRT reduced PR in cases with these risk factors. CONCLUSION: Long-term results of the PORTEC-2 trial confirm VBT as standard adjuvant treatment for HIR endometrial cancer. Molecular risk assessment has the potential to guide adjuvant therapy. EBRT provided better pelvic control in patients with unfavourable risk factors.

Markers of the p53 pathway further refine molecular profiling in high-risk endometrial cancer: A TransPORTEC initiative.

BACKGROUND: The morphological classification of high-risk endometrial cancer is of limited prognostic value. Recent attempts to stratify tumours according to molecular signatures have shown considerable promise. Here we attempted to further refine molecular classifications using markers of the p53 pathway. METHODS: We analysed the expression of p53 as well as three downstream markers of the p53 pathway, p21, mdm2 and phospho-p63 (pp63), by immunohistochemistry in a series of 114 endometrial cancers (86 endometrioid, 28 non-endometrioid subtype) with high-risk features (such as high tumour grade and deep myometrial invasion) and correlated results with clinical outcome. The Cancer Genome Atlas (TCGA) data were used to analyse TP63 mutations and copy-number alterations using cBioPortal. TP53 was silenced in two endometrial cancer cell lines to study its effect on p21 and p63. RESULTS: About half of the tumours showed a p53 mutant phenotype and there was a strong negative correlation with p21 expression. Being marker positive for pp63 or mdm2 was associated with a significantly increased likelihood of dying, [hazard ratios 5.93 (95% CI 2.37-7.27) and 7.48 (95% CI 3.04-9.39), respectively]. These findings were seen in both p53 wildtype and p53 mutant tumours. Only 11% of TCGA endometrial cancers had a functional TP63 alteration. Upon silencing of TP53, p21 expression was decreased in one cell line, but no effects on p63 were observed. CONCLUSION: Markers of the p53 pathway improve stratification of endometrial cancers and provide novel insights into the role of this pathway in the disease.

Practical guidance for mismatch repair-deficiency testing in endometrial cancer.

Background: Mismatch repair (MMR)-deficiency analysis is increasingly recommended for all endometrial cancers, as it identifies Lynch syndrome patients, and is emerging as a prognostic classifier to guide adjuvant treatment. The aim of this study was to define the optimal approach for MMR-deficiency testing and to clarify discrepancies between microsatellite instability (MSI) analysis and immunohistochemical (IHC) analysis of MMR protein expression. Patients and methods: Six hundred ninety- six endometrial cancers were analyzed for MSI (pentaplex panel) and MMR protein expression (IHC). Agreement between methodologies was calculated using Cohen's Kappa. MLH1 promoter hypermethylation, dinucleotide microsatellite markers and somatic MMR and POLE exonuclease domain (EDM) gene variants (using next-generation/Sanger sequencing) were analyzed in discordant cases. Results: MSI was found in 180 patients. Complete loss of expression of one or more MMR proteins was observed in 196 cases. A PMS2- and MSH6-antibody panel detected all cases with loss of MMR protein expression. The results of MSI and MMR protein expression were concordant in 655/696 cases (kappa = 0.854, P < 0.001). Ambiguous cases (n = 41, 6%) included: subclonal loss of MMR protein expression (n = 18), microsatellite stable or MSI-low cases with loss of MMR protein expression (n = 20), and MSI-low or MSI-high cases with retained MMR protein expression (n = 3). Most of these cases could be explained by MLH1 promoter hypermethylation. Five of seven cases with solitary loss of PMS2 or MSH6 protein expression carried somatic gene variants. Two MSI-high cases with retained MMR protein expression carried a POLE-EDM variant. Conclusion: MSI and IHC analysis are highly concordant in endometrial cancer. This holds true for cases with subclonal loss of MMR protein expression. Discordant MMR-proficient/MSI-high cases (<1%), may be explained by POLE-EDM variants.

The anticancer gene ORCTL3 targets stearoyl-CoA desaturase-1 for tumour-specific apoptosis.

ORCTL3 is a member of a group of genes, the so-called anticancer genes, that cause tumour-specific cell death. We show that this activity is triggered in isogenic renal cells upon their transformation independently of the cells' proliferation status. For its cell death effect ORCTL3 targets the enzyme stearoyl-CoA desaturase-1 (SCD1) in fatty acid metabolism. This is caused by transmembrane domains 3 and 4, which are more efficacious in vitro than a low molecular weight drug against SCD1, and critically depend on their expression level. SCD1 is found upregulated upon renal cell transformation indicating that its activity, while not impacting proliferation, represents a critical bottleneck for tumourigenesis. An adenovirus expressing ORCTL3 leads to growth inhibition of renal tumours in vivo and to substantial destruction of patients' kidney tumour cells ex vivo. Our results indicate fatty acid metabolism as a target for tumour-specific apoptosis in renal tumours and suggest ORCTL3 as a means to accomplish this.

L1 cell adhesion molecule is a strong predictor for distant recurrence and overall survival in early stage endometrial cancer: pooled PORTEC trial results.

BACKGROUND: L1 cell adhesion molecule (L1CAM) expression has been implicated as risk factor for disease recurrence in endometrial cancer (EC), most likely due to its role in promoting tumour cell motility. We tested the performance of L1CAM expression in predicting the risk of recurrence in the randomised post operative radiation therapy in endometrial carcinoma (PORTEC)-1 and -2 trials. METHODS: In the PORTEC trials, stage I EC patients were randomised to external beam radiotherapy (EBRT) versus no additional treatment (PORTEC-1, n=714), or to EBRT versus vaginal brachytherapy (PORTEC-2, n=427). Tumour samples of 865 (75.8%) patients were available for L1CAM expression analysis by immunohistochemistry. An established scoring system for EC was used, with >10% L1CAM staining defined as positive. RESULTS: Positive L1CAM expression was significantly correlated with risk of distant recurrence, with a hazard ratio (HR) of 5.1 (95% confidence interval (CI) 3.1-8.7) but not with vaginal relapse, while a trend for pelvic nodal relapse was found. Tumours with the highest expression levels (>50% positive) had the strongest risk of distant recurrence (HR 5.3, CI 2.7-10.4). In multivariate Cox analysis with the risk factors age, depth of invasion, grade, lympho-vascular space invasion (LVSI) and treatment, L1CAM expression remained an independent prognostic factor for distant recurrence (HR 3.5, CI 1.92-6.30) and overall survival (HR 2.1, CI 1.41-2.98). CONCLUSION: L1CAM expression is a strong independent predictor for distant recurrence and overall survival in stage I endometrial cancer. These results warrant prospective validation of L1CAM as marker for selecting patients who could benefit from more extensive diagnostic and/or therapeutic procedures.