DNA ploidy in curettage specimens identifies high-risk patients and lymph node metastasis in endometrial cancer.

BACKGROUND: Preoperative risk stratification is essential in tailoring endometrial cancer treatment, and biomarkers predicting lymph node metastasis and aggressive disease are aspired in clinical practice. DNA ploidy assessment in hysterectomy specimens is a well-established prognostic marker. DNA ploidy assessment in preoperative curettage specimens is less studied, and in particular in relation to the occurrence of lymph node metastasis. METHODS: Curettage image cytometry DNA ploidy in relation to established clinicopathological variables and outcome was investigated in 785 endometrial carcinoma patients prospectively included in the MoMaTEC multicentre trial. RESULTS: Diploid curettage status was found in 72.0%, whereas 28.0% were non-diploid. Non-diploid status significantly correlated with traditional aggressive postoperative clinicopathological features, and was an independent predictor of lymph node metastasis among FIGO stage I-III patients in multivariate analysis (OR 1.94, P=0.033). Non-diploid status was related to shorter disease-specific survival (5-year DSS of 74.4% vs 88.8% for diploid curettage, P<0.001). When stratifying by FIGO stage and lymph node status, the prognostic effect remained. However, in multivariate regression analysis, preoperative histological risk classification was a stronger predictor of DSS than DNA ploidy. CONCLUSIONS: Non-diploid curettage is significantly associated with aggressive clinicopathological phenotype, lymph node metastasis, and poor survival in endometrial cancer. The prognostic effect was also observed among subgroups with (presumably) less aggressive traits, such as low FIGO stage and negative lymph node status. Our results indicate curettage DNA ploidy as a possible supplement to existing parameters used to tailor surgical treatment.

Fertility and gonadal function after adjuvant therapy in women diagnosed with a malignant ovarian germ cell tumor (MOGCT) during the "cisplatin era".

PURPOSE: By self-report and serum levels of anti-Mullerian hormone (AMH) this study aims to assess post-treatment fertility after modern treatment of women with malignant ovarian germ cell tumors (MOGCT). PATIENTS AND METHODS: In 2013 a questionnaire-based survey was performed in 61 MOGCT patients diagnosed at age <40years from 1980-2009. Forty-nine of them also attended the out-patient clinic. The event of first post-treatment pregnancy ("fertility") was documented as cumulative estimates for all 61 patients and within each of 4 treatment groups: Group 1: Surgery only (n=10); Group 2: ≤3cycles of cisplatin-based chemotherapy (CBCT) (n=20); Group 3: >3cycles of CBCT (n=15) and Group 4: other adjuvant treatment (n=16). AMH was determined in 22 women <40years at survey. Statistics were based on Kaplan Meier procedure, log-rank test and a significance level p<0.05. RESULTS: At least one post-treatment pregnancy was reported by 34 of 39 MOGCT survivors who attempted motherhood after treatment. The 15-year cumulative post-treatment fertility estimate was 28% (95% CI: 26-30) for all 61 survivors and was significantly higher in patients treated with 3 or fewer cycles of CBCT (53% [95% CI: 50-55]) than those treated with more than 3cycles (20% [95% CI: 17-22]) (P=0.03). Of 22 AMH levels, two were <3pmol/l, with one women being pregnant at survey. CONCLUSION: After fertility-sparing surgery and modern cisplatin-based chemotherapy, fertility is preserved in most MOGCT survivors though dependent on the number of cycles. AMH's role as a biomarker of gonadal function seems promising but requires further research.

Tumour apparent diffusion coefficient is associated with depth of myometrial invasion and is negatively correlated to tumour volume in endometrial carcinomas.

AIM: To explore possible correlations between tumour apparent diffusion coefficient (ADC), morphological tumour volume, and clinical and histological characteristics in endometrial carcinomas and to evaluate interobserver agreement for preoperative staging by MRI and for ADC measurements. MATERIALS AND METHODS: Preoperative conventional MRI including diffusion-weighted imaging (DWI) was performed in 105 endometrial carcinoma patients. Three radiologists independently reviewed the images for the presence of deep myometrial invasion, cervical stromal invasion, and lymph node metastases, and measured tumour ADC in regions of interest (ROIs). ADC values were analysed in relation to histomorphological characteristics and tumour volume. Kappa coefficients (κ) and intraclass correlation coefficients (ICC) for interobserver agreement for MRI staging results and ADC measurements, respectively, were calculated, and receiver operating characteristic (ROC) curves for identification of deep of myometrial invasion were generated. RESULTS: Mean tumour ADC was significantly lower in tumours with deep myometrial invasion (ADC = 0.75 × 10(-3) mm(2)/s) compared to tumours with superficial or no myometrial invasion (ADC = 0.85 × 10(-3) mm(2)/s; p < 0.001). ADC was negatively correlated to tumour size (p = 0.007). The interobserver agreement was fair (κ = 0.32) for depth of myometrial invasion, good for cervical stromal invasion (κ = 0.66), and moderate for lymph node metastases (κ = 0.54), and the interobserver variability for ADC value measurements was low (ICC = 0.60). CONCLUSION: Tumour ADC measurements may in the future provide an adjunct tool, aiding in the preoperative identification of high-risk patients with deep myometrial infiltration.

High level of HSF1 associates with aggressive endometrial carcinoma and suggests potential for HSP90 inhibitors.

BACKGROUND: Recent identification of a specific role of HSF1 in cancer progression has led to new relevance of HSF1 as both a prognostic and a predictive marker. The role of HSF1 in endometrial cancer has so far been unexplored. METHODS: A total of 823 lesions from endometrial carcinoma precursors, primary tumours and metastases were prospectively collected and explored for HSF1 protein expression in relation to established markers for aggressive disease and survival. Transcriptional alterations related to HSF1 protein level were investigated by microarray analysis for 224 freshly frozen samples in parallel. RESULTS: High expression of HSF1 protein in endometrial carcinoma is significantly associated with aggressive disease and poor survival (all P-values ≤ 0.02), also among ERα-positive patients presumed to have good prognosis. The HSF1-related gene signatures increase during disease progression and were also found to have prognostic value. Gene expression analyses identified HSP90 inhibition as a potential novel therapeutic approach for cases with high protein expression of HSF1. CONCLUSIONS: We demonstrate for the first time in endometrial cancer that high expression of HSF1 and measures for transcriptional activation of HSF1 associate with poor outcome and disease progression. The HSP90 inhibitors are suggested as new targeted therapeutics for patients with high HSF1 levels in tumour in particular.

Increased microvascular proliferation is negatively correlated to tumour blood flow and is associated with unfavourable outcome in endometrial carcinomas.

BACKGROUND: We aimed to study the angiogenic profile based on histomorphological markers in endometrial carcinomas in relation to imaging parameters obtained from preoperative dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and diffusion-weighted imaging (DWI) and to explore the potential value of these markers to identify patients with poor outcome. METHODS: In fifty-four surgically staged endometrial carcinoma patients, immunohistochemical staining with factor VIII and Ki67 allowed assessment of microvessel density (MVD) and microvascular proliferation reflecting tumour angiogenesis. In the same patients, preoperative pelvic DCE-MRI and DWI allowed the calculation of parameters describing tumour microvasculature and microstructure in vivo. RESULTS: Microvascular proliferation was negatively correlated to tumour blood flow (Fb) (r=-0.36, P=0.008), capillary permeability surface area product (PS) (r=-0.39, P=0.004) and transfer from the blood to extravascular extracellular space (EES) (Ktrans) (r=-0.40, P=0.003), and was positively correlated to tumour volume (r=0.34; P=0.004). High-tumour microvascular proliferation, low Fb and low Ktrans were all significantly associated with reduced progression/recurrence-free survival (P<0.05). CONCLUSION: Disorganised angiogenesis with coexisting microvascular proliferation and low tumour blood flow is a poor prognostic factor supporting that hypoxia is associated with progression and metastatic spread in endometrial carcinomas.

Endometrial Carcinoma Recurrence Score (ECARS) validates to identify aggressive disease and associates with markers of epithelial-mesenchymal transition and PI3K alterations.

PURPOSE: Our previously reported 29-gene expression signature identified an aggressive subgroup of endometrial cancer patients with PI3K activation. We here wanted to validate these findings by independent patient series. PATIENTS AND METHODS: The 29-gene expression signature was assessed in fresh frozen tumor tissue from 280 primary endometrial carcinomas (three independent cohorts), 19 metastatic lesions and in 333 primary endometrial carcinomas using TCGA data, and expression was related to clinico-pathologic features and survival. The 29-gene signature was assessed by real-time quantitative PCR, DNA oligonucleotide microarrays, or RNA sequencing. PI3K alterations were assessed by immunohistochemistry, DNA microarrays, DNA sequencing, SNP arrays or fluorescence in situ hybridization. A panel of markers of epithelial-mesenchymal transition (EMT) was also correlated to the 29-gene signature score. RESULTS: High 29-gene Endometrial Carcinoma Recurrence Score (ECARS) values consistently validated to identify patients with aggressive clinico-pathologic phenotype and reduced survival. Within the presumed favorable subgroups of low grade, endometrioid tumors confined to the uterus, high ECARS still predicted a poor prognosis. The score was higher in metastatic compared to primary lesions (P<0.001) and was significantly associated with potential measures of PI3K activation, markers of EMT and vascular invasion as an indicator of metastatic spread (all P<0.001). CONCLUSIONS: ECARS validates to identify aggressive endometrial carcinomas in multiple, independent patients cohorts. The higher signature score in metastatic compared to primary lesions, and the potential link to PI3K activation and EMT, support further studies of ECARS in relation to response to PI3K and EMT inhibitors in clinical trials of metastatic endometrial carcinoma.

Current status of molecular biomarkers in endometrial cancer.

In spite of the high and increasing incidence of endometrial cancer, our current models for prediction of prognosis and even more treatment response are suboptimal, and molecular biomarkers to assist clinical decision making are needed. In this review an overview is given of the currently known as well as promising prognostic and predictive biomarkers in endometrial carcinoma. Key clinical challenges, where use of molecular biomarkers can meet clinical needs, are highlighted. The current status for the presently most promising prognostic and predictive biomarkers in endometrial carcinoma is reviewed. DNA ploidy, p53 status, hormone receptor level, HER2, stathmin, L1 cell adhesion molecule expression and other biomarkers are discussed in relation to the scientific robustness of various essential steps in biomarker development and (current) clinical applicability for individualizing treatment strategies. Tumour heterogeneity and its consequences for biomarker assessment and the importance of developing standardised tests for implementation are discussed. To improve the development and clinical uptake of biomarkers, several strategies are proposed.

Loss of GPER identifies new targets for therapy among a subgroup of ERα-positive endometrial cancer patients with poor outcome.

BACKGROUND: The G protein-coupled oestrogen receptor, GPER, has been suggested as an alternative oestrogen receptor. Our purpose was to investigate the potential of GPER as a prognostic and predictive marker in endometrial carcinoma and to search for new drug candidates to improve treatment of aggressive disease. MATERIALS AND METHOD: A total of 767 primary endometrial carcinomas derived from three patient series, including an external dataset, were studied for protein and mRNA expression levels to investigate and validate if GPER loss identifies poor prognosis and new targets for therapy in endometrial carcinoma. Gene expression levels, according to ERα/GPER status, were used to search the connectivity map database for small molecular inhibitors with potential for treatment of metastatic disease for receptor status subgroups. RESULTS: Loss of GPER protein is significantly correlated with low GPER mRNA, high FIGO stage, non-endometrioid histology, high grade, aneuploidy and ERα loss (all P-values ≤0.05). Loss of GPER among ERα-positive patients identifies a subgroup with poor prognosis that until now has been unrecognised, with reduced 5-year survival from 93% to 76% (P=0.003). Additional loss of GPER from primary to metastatic lesion counterparts further supports that loss of GPER is associated with disease progression. CONCLUSION: These results support that GPER status adds clinically relevant information to ERα status in endometrial carcinoma and suggest a potential for new inhibitors in the treatment of metastatic endometrial cancers with ERα expression and GPER loss.

KRAS gene amplification and overexpression but not mutation associates with aggressive and metastatic endometrial cancer.

BACKGROUND: Three quarter of endometrial carcinomas are treated at early stage. Still, 15 to 20% of these patients experience recurrence, with little effect from systemic therapies. Homo sapiens v-Ki-ras2 Kirsten rat sarcoma viral oncogenes homologue (KRAS) mutations have been reported to have an important role in tumorigenesis for human cancers, but there is limited knowledge regarding clinical relevance of KRAS status in endometrial carcinomas. METHODS: We have performed a comprehensive and integrated characterisation of genome-wide expression related to KRAS mutations and copy-number alterations in primary- and metastatic endometrial carcinoma lesions in relation to clinical and histopathological data. A primary investigation set and clinical validation set was applied, consisting of 414 primary tumours and 61 metastatic lesions totally. RESULTS: Amplification and gain of KRAS present in 3% of the primary lesions and 18% of metastatic lesions correlated significantly with poor outcome, high International Federation of Gynaecology and Obstetrics stage, non-endometrioid subtype, high grade, aneuploidy, receptor loss and high KRAS mRNA levels, also found to be associated with aggressive phenotype. In contrast, KRAS mutations were present in 14.7% of primary lesions with no increase in metastatic lesions, and did not influence outcome, but was significantly associated with endometrioid subtype, low grade and obesity. CONCLUSION: These results support that KRAS amplification and KRAS mRNA expression, both increasing from primary to metastatic lesions, are relevant for endometrial carcinoma disease progression.

Revision of FIGO surgical staging in 2009 for endometrial cancer validates to improve risk stratification.

OBJECTIVE: Correct staging is a cornerstone in cancer treatment. The FIGO surgical staging for endometrial cancer was revised in 2009. We have evaluated if the revision improved stratification with respect to prognosis in a large prospective multicenter setting. METHODS: 1268 endometrial cancer patients have been prospectively recruited in the MoMaTEC study for the investigation of clinical and histopathological data. RESULTS: Restaging from FIGO 88 to FIGO 09 criteria increased the number of stage I cases from 932 to 979. The majority of the non-endometrioid tumors, down-staged to FIGO 09 stage I, were of serous histology. One third of the patients classified as stage II tumors based on FIGO 88 criteria (FIGO88 IIA) were down-staged to FIGO 09 IA (53%) and FIGO 09 IB (47%). The histological subtype for these cases was mainly endometrioid (86.1%) and high/intermediate grade (77.7%). Patients with FIGO 88 stages IA, IB, IIA and IIIA with positive cytology only, showed similar survival. In Cox multivariate survival analysis adjusting for histopathological variables we found that the revised FIGO 09 criteria improved prognostication. For FIGO stage I patients the adjusted HR was 3.9 (p=0.01, CI 1.35-11.36) for FIGO IB compared to FIGO IA. The independent prognostic impact for the FIGO 09 staging was also confirmed in a subset analysis of patients not subjected to lymphadenectomy and for the endometrioid subgroup. CONCLUSIONS: The FIGO 2009 staging system has improved prediction of prognosis, and is less complex, compared to earlier versions. Careful assessment of myometrial invasion seems particularly important for patients not subjected to lymphadenectomy.

Staging of endometrial carcinomas with MRI using traditional and novel MRI techniques.

Endometrial carcinoma is the most common gynaecological malignancy in industrialized countries. This review discusses the value of magnetic resonance imaging (MRI) and novel MRI techniques (diffusion, perfusion, spectroscopy, blood oxygen level-dependent (BOLD)-MRI, and MRI with new contrast agents) in endometrial carcinomas. Contrast-enhanced MRI is the imaging technique of choice, and diffusion-weighted MRI may help to identify malignant lesions and assess myometrial invasion. Novel MRI techniques may potentially increase diagnostic accuracy, enabling a refined, tailored surgical procedure and better prediction of treatment outcomes.

Integrated genomic profiling of endometrial carcinoma associates aggressive tumors with indicators of PI3 kinase activation.

Although 75% of endometrial cancers are treated at an early stage, 15% to 20% of these recur. We performed an integrated analysis of genome-wide expression and copy-number data for primary endometrial carcinomas with extensive clinical and histopathological data to detect features predictive of recurrent disease. Unsupervised analysis of the expression data distinguished 2 major clusters with strikingly different phenotypes, including significant differences in disease-free survival. To identify possible mechanisms for these differences, we performed a global genomic survey of amplifications, deletions, and loss of heterozygosity, which identified 11 significantly amplified and 13 significantly deleted regions. Amplifications of 3q26.32 harboring the oncogene PIK3CA were associated with poor prognosis and segregated with the aggressive transcriptional cluster. Moreover, samples with PIK3CA amplification carried signatures associated with in vitro activation of PI3 kinase (PI3K), a signature that was shared by aggressive tumors without PIK3CA amplification. Tumors with loss of PTEN expression or PIK3CA overexpression that did not have PIK3CA amplification also shared the PI3K activation signature, high protein expression of the PI3K pathway member STMN1, and an aggressive phenotype in test and validation datasets. However, mutations of PTEN or PIK3CA were not associated with the same expression profile or aggressive phenotype. STMN1 expression had independent prognostic value. The results affirm the utility of systematic characterization of the cancer genome in clinically annotated specimens and suggest the particular importance of the PI3K pathway in patients who have aggressive endometrial cancer.

High BMI is significantly associated with positive progesterone receptor status and clinico-pathological markers for non-aggressive disease in endometrial cancer.

BACKGROUND: Endometrial cancer incidence is increasing in industrialised countries. High body mass index (BMI, kg m(-2)) is associated with higher risk for disease. We wanted to investigate if BMI is related to clinico-pathological characteristics, hormone receptor status in primary tumour, and disease outcome in endometrial cancer. PATIENTS AND METHODS: In total, 1129 women primarily treated for endometrial carcinoma at Haukeland University Hospital during 1981-2009 were studied. Body mass index was available for 949 patients and related to comprehensive clinical and histopathological data, hormone receptor status in tumour, treatment, and follow-up. RESULTS: High BMI was significantly associated with low International Federation of Gynaecology and Obstetrics (FIGO) stage, endometrioid histology, low/intermediate grade, and high level of progesterone receptor (PR) mRNA by qPCR (n=150; P=0.02) and protein expression by immunohistochemistry (n=433; P=0.003). In contrast, oestrogen receptor (ERα) status was not associated with BMI. Overweight/obese women had significantly better disease-specific survival (DSS) than normal/underweight women in univariate analysis (P=0.035). In multivariate analysis of DSS adjusting for age, FIGO stage, histological subtype, and grade, BMI showed no independent prognostic impact. CONCLUSION: High BMI was significantly associated with markers of non-aggressive disease and positive PR status in a large population-based study of endometrial carcinoma. Women with high BMI had significantly better prognosis in univariate analysis of DSS, an effect that disappeared in multivariate analysis adjusting for established prognostic markers. The role of PR in endometrial carcinogenesis needs to be further studied.

Low BMI-1 expression is associated with an activated BMI-1-driven signature, vascular invasion, and hormone receptor loss in endometrial carcinoma.

We studied the expression of polycomb group (PcG) protein BMI-1 in a large population-based patient series of endometrial carcinomas in relation to clinical and molecular phenotype. Also, 57 fresh frozen endometrial carcinomas were studied for the relationship between BMI-1 protein expression, BMI-1 mRNA level, and activation of an 11-gene signature reported to represent a BMI-1-driven pathway. BMI-1 protein expression was significantly weaker in tumours with vascular invasion (P<0.0001), deep myometrial infiltration (P=0.004), and loss of oestrogen receptor (ER) (P<0.0001) and progesterone receptors (PR) (P=0.03). Low BMI-1 protein expression was highly associated with low BMI-1 mRNA expression (P=0.002), and similarly low BMI-1 mRNA expression correlated significantly with vascular invasion, ER and PR loss, and histologic grade 3. In contrast, activation of the reported 11-gene signature, supposed to represent a BMI-1-driven pathway, correlated with low mRNA expression of BMI-1 (P<0.001), hormone receptor loss, presence of vascular invasion, and poor prognosis. We conclude that BMI-1 protein and mRNA expression are significantly correlated and that BMI-1 expression is inversely associated with activation of the 11-gene signature. Loss of BMI-1 seems to be associated with an aggressive phenotype in endometrial carcinomas.

Frequency and prognostic impact of microsatellite instability in a large population-based study of endometrial carcinomas.

The replication error repair (RER) phenotype has been reported in 9-43% of sporadic endometrial carcinomas, but there are conflicting data about its effect on prognosis in this disease. This study was performed to establish the frequency of the RER phenotype and to determine its effect on prognosis in a population-based series of 259 endometrial carcinomas with long-term follow-up. Five mononucleotide and dinucleotide microsatellite markers on different chromosomes were analyzed, and tumors exhibiting microsatellite instability at two or more loci were classified as RER+. A total of 116 of 259 tumors (45%) were RER+. The 5-year survival rate for the RER- group was 76.2% compared with 79.6% for RER+ cases (P = 0.6). The 5-year recurrence-free survival rate among the 228 patients surgically treated for cure was 80.6% in the RER- group compared with 83.6% in the RER+ group (P = 0.6). The analysis indicates that the RER phenotype is common in endometrial carcinomas, but there is no association with prognosis in this large population-based series of endometrial carcinomas.

Prognostic significance of angiogenesis and Ki-67, p53, and p21 expression: a population-based endometrial carcinoma study.

PURPOSE: For endometrial carcinoma patients, there is a need for improved identification of high-risk groups that may benefit from postoperative adjuvant therapy. We therefore studied the prognostic impact of markers for cell proliferation, cell-cycle regulation, and angiogenesis among endometrial carcinoma patients in a population-based setting. PATIENTS AND METHODS: All patients diagnosed with endometrial carcinoma between 1981 and 1985 in Hordaland County, Norway, were studied. The median follow-up for the survivors was 11.5 years (range, 8 to 15 years), with no patient lost because of insufficient follow-up information. Paraffin-embedded tumor tissue, available in 96% of the cases (n = 142), was studied immunohistochemically for microvessel density (MVD) and expression of Ki-67, p53, and p21 proteins. We used the hot spot method for calculation of MVD, and expression of Ki-67 and p21 protein, because this approach may increase the probability of detecting small aggressive clones of possible prognostic relevance. The importance of these tumor markers was investigated in univariate survival analyses and Cox regression analysis. RESULTS: The majority of traditional clinicopathologic variables was significantly associated with the tumor biomarkers. Age, International Federation of Gynecology and Obstetrics (FIGO) stage, histologic type, histologic grade, MVD, as well as Ki-67, p53, and p21 protein expression, all significantly influenced survival in univariate analyses (P < or = .05). In the Cox regression analysis, age, FIGO stage, MVD, Ki-67 expression, and p53 expression were the only variables with independent prognostic impact (P < or = .05), whereas histologic type, histologic grade, and p21 expression had no independent influence. A group of high-risk patients with more than one unfavorable marker was identified. CONCLUSION: In addition to age and FIGO stage, MVD, Ki-67, and p53 protein expression showed an independent prognostic impact. Thus, information derived from routine histologic specimens identified a subgroup of high-risk endometrial carcinoma patients in this population-based study.

Loss of nuclear p16 protein expression is not associated with promoter methylation but defines a subgroup of aggressive endometrial carcinomas with poor prognosis.

The p16INK4aa (CDKN2) tumor suppressor gene is altered in several tumor types, but the frequency and mechanism of inactivation are largely unknown for endometrial carcinomas. We therefore wanted to assess the pattern and prognostic impact of p16 protein expression and promoter region methylation in a population-based series of 316 endometrial carcinoma patients with long-term and complete follow-up. Nuclear staining of p16 protein was related to clinicopathological variables, tumor markers, patient survival, and the presence of promoter region methylation. Absent/minimal nuclear staining for p16 protein was found in 14% of the tumors. Methylation of the p16 promoter region was found in only one tumor (0.7%) in a subset of 138 cases studied. This tumor lacked nuclear p16 protein expression as well. Loss of nuclear p16 staining was significantly associated with increased age, high FIGO (International Federation of Gynecology and Obstetrics) stage, serous papillary or clear cell histological types, high histological grade, aneuploidy, low estradiol and progesterone receptor concentrations, high expression of Ki-67, high intratumor microvessel density, and strong nuclear p53 protein expression. The 5-year survival was 47% for patients with absent/minimal nuclear p16 expression (n = 39) compared with 81% for patients with moderate/high nuclear p16 expression (n = 247; P < 0.0001). In Cox proportional hazards regression analysis, nuclear p16 expression showed an independent prognostic impact in addition to FIGO stage, age, Ki-67 expression, and microvessel density, with an adjusted hazard ratio of 2.9 (95% confidence interval, 1.3-6.5). The other variables lost their prognostic impact when nuclear p16 staining was added to the Cox model. In conclusion, loss of nuclear p16 protein expression was associated with aggressive endometrial carcinomas and high proliferative activity (Ki-67) and was found to represent a strong and independent prognostic indicator. Methylation of the promoter region seems to be an uncommon mechanism of p16 inactivation in endometrial carcinoma.

Identification of high-risk patients by assessment of nuclear Ki-67 expression in a prospective study of endometrial carcinomas.

For patients with localized endometrial carcinoma at primary operation, there is a definite need for more specific prognostic parameters to select patients for adjuvant therapy. The purpose of this study was to evaluate the applicability and prognostic significance of markers for tumor cell proliferation (S phase fraction and Ki-67 expression) among endometrial carcinoma patients, relative to the information derived from established clinicopathological variables including DNA and receptor analyses. In a prospective study of 115 patients treated for endometrial carcinoma, fresh tumor tissue was collected for assessments of ploidy, S phase fraction and hormone receptor concentrations. Ki-67 expression was assessed immunohistochemically on sections from formalin-fixed and paraffin-embedded tumor specimens. These variables were examined together with traditional clinicopathological features in univariate and multivariate (Cox proportional hazards regression model) survival analyses. The median follow-up time for the survivors was 9 years (range, 5-15), with no patient lost due to insufficient follow-up information. The expression of Ki-67 was significantly associated with International Federation of Gynecology and Obstetrics (FIGO) stage (P = 0.0004), histological type (P = 0.03), and histological grade (P = 0.0001). S phase fraction was significantly associated with histological grade (P = 0.02), whereas the association with histological type was of borderline significance (P = 0.07). In univariate analyses, survival was significantly influenced by FIGO stage (P <0.0001), histological type (P = 0.0002), histological grade (P = 0.002), ploidy (P = 0.015), S phase fraction (P = 0.017), progesterone receptor concentration (P = 0.02), and Ki-67 expression (P <0.0001). In Cox regression analysis, FIGO stage (hazard ratio, 11.7; 95% confidence interval, 4.3-32.1) and Ki-67 (hazard ratio, 8.7; 95% confidence interval, 3.0-25.2) were the only variables with independent prognostic impact. When patients whose tumors were confined to the uterus were analyzed separately, Ki-67 was the only variable with an independent prognostic importance. Ki-67 expression was superior to S phase fraction both in applicability and prognostic value. FIGO stage and Ki-67 expression were the only variables with independent prognostic impact in Cox regression analysis. Ki-67 expression is assessed on the histological specimens taken routinely, with no separate sampling technique; this should make it easy to use in a routine setting.

Methylation of hMLH1 in a population-based series of endometrial carcinomas.

Microsatellite instability (MSI) is a characteristic feature of hereditary nonpolyposis colorectal cancer and is also observed in sporadic colorectal and endometrial cancers. Alterations in the mismatch repair genes hMLH1 and hMSH2 are important for the development of MSI. It has recently been demonstrated that hypermethylation of the hMLH1 promoter region is associated with MSI and appears to be a common mechanism for gene inactivation. For endometrial carcinoma, however, previous studies have been relatively small and have not been population based. We therefore wanted to assess the frequency and prognostic significance of hypermethylation of the hMLH1 and hMSH2 genes in conjunction with hMLH1 protein expression in a prospective and population-based series of endometrial carcinoma patients with known MSI status and complete follow-up. A total of 138 patients were studied, and methylation of hMLH1 was found in 23% of tumors with conclusive results, whereas methylation of hMSH2 was seen in only 1% of tumors. Methylation of hMLH1 was significantly correlated with MSI (P < 0.001). Loss of nuclear staining of hMLH1 protein was seen in 14% of the cases and was significantly correlated with hMLH1 methylation and MSI (P < 0.001). Normal expression of hMLH1 was seen in all of the unmethylated tumors (100%). Of the 14 MSI-positive tumors that were also methylated, all but 1 (93%) showed a loss of nuclear expression of hMLH1. None of the tumors with loss of hMLH1 expression or hMLH1 methylation were aneuploid (P for both < or = 0.05), and loss of hMLH1 expression and hMLH1 methylation was significantly correlated with lack of p53 overexpression (P for both < or = 0.05). Nuclear hMLH1 staining and hMLH1 methylation did not significantly influence survival. In conclusion, hMLH1 methylation was common and was significantly correlated with loss of hMLH1 protein expression, MSI, diploid tumors, and lack of p53 overexpression. In contrast, hMSH2 methylation was infrequent in this prospective and population-based series of endometrial carcinomas.