Serum HE4 predicts progestin treatment response in endometrial cancer and atypical hyperplasia: A prognostic study.

OBJECTIVE: To investigate serum human epididymis-4 (HE4) as a predictive biomarker of intrauterine progestin response in endometrial cancer and atypical endometrial hyperplasia (AEH). DESIGN: Prospective prognostic factor study. SETTING: Consecutive sample of women attending a tertiary gynaecological oncology centre in northwest England. POPULATION: Women with AEH or early-stage, low-grade endometrial cancer who were unfit for or declined primary surgical management. METHODS: A total of 76 women, 32 with AEH and 44 with endometrial cancer, were treated with a levonorgestrel intrauterine system (LNG-IUS) for 12 months. Endometrial biopsies and imaging were performed to assess treatment response. Pretreatment serum HE4 was analysed by chemiluminescence immunoassay and diagnostic accuracy and logistic regression analyses were performed. MAIN OUTCOME MEASURES: Progestin response at 12 months defined by histology and imaging. RESULTS: The median age and body mass index (BMI) of the final cohort were 52 years (interquartile range [IQR] 33-62 years) and 46 kg/m2 (IQR 38-54 kg/m2 ), respectively. Baseline serum HE4 was significantly higher in non-responders than responders (119.2 pmol/L, IQR 94.0-208.4 pmol/L versus 71.8 pmol/L, IQR 56.1-84.2 pmol/L, p < 0.001). Older age (odds ratio [OR] 0.96, 95% CI 0.93-0.99, p = 0.02), baseline serum HE4 (OR 0.97, 95% CI 0.96-0.99, p = 0.001) and endometrial cancer histology (OR 0.22, 95% CI 0.72-0.68, p = 0.009) were associated with a lower likelihood of progestin treatment response. Serum HE4 remained independently associated with progestin treatment failure when adjusted for age and histology (adjusted hazard ratio 0.97, 95% CI 0.96-0.99, p = 0.008). CONCLUSION: Serum HE4 shows promise as a predictive biomarker of progestin treatment response in endometrial cancer and AEH.

The impact of obesity and bariatric surgery on the immune microenvironment of the endometrium.

BACKGROUND: The incidence of endometrial cancer is rising in parallel with the obesity epidemic. Obesity increases endometrial cancer risk and weight loss is protective, but the underlying mechanisms are incompletely understood. We hypothesise that the immune microenvironment may influence susceptibility to malignant transformation in the endometrium. The aim of this study was to measure the impact of obesity and weight loss on the immunological landscape of the endometrium. METHODS: We conducted a prospective cohort study of women with class III obesity (body mass index, BMI ≥ 40 kg/m2) undergoing bariatric surgery or medically-supervised low-calorie diet. We collected blood and endometrial samples at baseline, and two and 12 months after weight loss intervention. Serum was analysed for inflammatory markers CRP, IL-6 and TNF-α. Multiplex immunofluorescence was used to simultaneously identify cells positive for immune markers CD68, CD56, CD3, CD8, FOXP3 and PD-1 in formalin-fixed paraffin-embedded endometrial tissue sections. Kruskal-Wallis tests were used to determine whether changes in inflammatory and immune biomarkers were associated with weight loss. RESULTS: Forty-three women with matched serum and tissue samples at all three time points were included in the analysis. Their median age and BMI were 44 years and 52 kg/m2, respectively. Weight loss at 12 months was greater in women who received bariatric surgery (n = 37, median 63.3 kg) than low-calorie diet (n = 6, median 12.8 kg). There were significant reductions in serum CRP (p = 3.62 × 10-6, r = 0.570) and IL-6 (p = 0.0003, r = 0.459), but not TNF-α levels, with weight loss. Tissue immune cell densities were unchanged except for CD8+ cells, which increased significantly with weight loss (p = 0.0097, r = -0.323). Tissue CD3+ cell density correlated negatively with systemic IL-6 levels (p = 0.0376; r = -0.318). CONCLUSION: Weight loss is associated with reduced systemic inflammation and a recruitment of protective immune cell types to the endometrium, supporting the concept that immune surveillance may play a role in endometrial cancer prevention.

Advances in genetic technologies result in improved diagnosis of mismatch repair deficiency in colorectal and endometrial cancers.

BACKGROUND: Testing cancers for mismatch repair deficiency (dMMR) by immunohistochemistry (IHC) is a quick and inexpensive means of triaging individuals for germline Lynch syndrome testing. The aim of this study was to evaluate tumour dMMR and the prevalence of Lynch syndrome in patients referred to the Manchester Centre for Genomic Medicine, which serves a population of 5.6 million. METHODS: Tumour testing used IHC for MMR proteins with targeted BRAF and MLH1 promotor methylation testing followed by germline mutation and somatic testing as appropriate. RESULTS: In total, 3694 index tumours were tested by IHC (2204 colorectal cancers (CRCs), 739 endometrial cancers (ECs) and 761 other), of which 672/3694 (18.2%) had protein loss, including 348 (9.4%) with MLH1 loss. MLH1 loss was significantly higher for 739 ECs (15%) vs 2204 CRCs (10%) (p=0.0003) and was explained entirely by higher rates of somatic MLH1 promoter hypermethylation (87% vs 41%, p<0.0001). Overall, 65/134 (48.5%) patients with MLH1 loss and no MLH1 hypermethylation or BRAF c.1799T>A had constitutional MLH1 pathogenic variants. Of 456 patients with tumours showing loss of MSH2/MSH6, 216 (47.3%) had germline pathogenic variants in either gene. Isolated PMS2 loss was most suggestive of a germline MMR variant in 19/26 (73%). Of those with no germline pathogenic variant, somatic testing identified likely causal variants in 34/48 (71%) with MLH1 loss and in MSH2/MSH6 in 40/47 (85%) with MSH2/MSH6 loss. CONCLUSIONS: Reflex testing of EC/CRC leads to uncertain diagnoses in many individuals with dMMR following IHC but without germline pathogenic variants or MLH1 hypermethylation. Tumour mutation testing is effective at decreasing this by identifying somatic dMMR in >75% of cases.

Weight Loss During Intrauterine Progestin Treatment for Obesity-associated Atypical Hyperplasia and Early-Stage Cancer of The Endometrium.

Intrauterine progestin is a treatment option for women with atypical hyperplasia or low-risk endometrial cancer who wish to preserve their fertility, or whose poor surgical fitness precludes safe hysterectomy. We hypothesized that in such women with obesity, weight loss during progestin treatment may improve oncological outcomes. We conducted a prospective nonrandomized study of women with obesity and atypical hyperplasia or low-grade stage 1a endometrial cancer undergoing progestin treatment. Women with a body mass index (BMI) ≥ 35 kg/m2 were offered bariatric surgery; those who declined and those with a BMI of 30 to 34.9 kg/m2 were encouraged to lose weight by low-calorie diet. We assessed uptake of bariatric surgery; weight lost during progestin treatment; and the impact of more than 10% total body weight loss on progestin treatment response at 12 months. 71 women [median age 58 years (interquartile range; IQR 35-65); mean BMI 48 kg/m2 (SD 9.3)] completed the study. Twenty-three women (32%) had bariatric surgery, on average 5 months (IQR 3-8) after progestin treatment commenced. Weight change during progestin treatment was -33.4 kg [95% confidence interval (CI) -42.1, -24.7] and -4.6 kg (95% CI -7.8, -1.4) in women receiving bariatric surgery and low-calorie diet, respectively (P < 0.001). Forty-three women (61%) responded to progestin, while 23 (32%) showed stabilized and 5 (7%) progressive disease. Response at 12 months was not predicted by age or baseline BMI, but women who lost more than 10% of their total body weight were more likely to respond to progestin than those who did not (adjusted odds ratio 3.95; 95% CI 1.3, 12.5; P = 0.02). Thus weight loss may improve oncological outcomes in women with obesity-associated endometrial neoplastic abnormalities treated with progestin. PREVENTION RELEVANCE: This study found that weight loss improves response rates in women with obesity and atypical hyperplasia or low-risk endometrial cancer undergoing conservative management with intrauterine progestin. Given the additional benefits of weight loss for fertility, cardiovascular health and quality of life, future research should focus on how best to accomplish it.

Assessment of mismatch repair deficiency in ovarian cancer.

BACKGROUND: Hereditary causes of ovarian cancer include Lynch syndrome, which is due to inherited pathogenic variants affecting one of the four mismatch repair genes involved in DNA repair. The aim of this study was to evaluate tumour mismatch repair deficiency and prevalence of Lynch syndrome in high-risk women referred to the Manchester Centre for Genomic Medicine with ovarian cancer over the past 20 years. METHODS: Women with ovarian cancer diagnosed before the age of 35 years and/or with a suggestive personal or family history of Lynch syndrome cancers underwent tumour testing with immunohistochemistry for mismatch repair deficiency and, where indicated, MLH1 promoter methylation testing followed by constitutional testing for Lynch syndrome. RESULTS: In total, 261 ovarian cancers were tested and 27 (10.3%; 95% CI 6.9% to 14.7%) showed mismatch repair deficiency by immunohistochemistry. Three of 7 with MLH1 loss showed MLH1 promoter hypermethylation, and 18 of the remaining 24 underwent constitutional testing for Lynch syndrome. A further 15 women with mismatch repair proficient tumours underwent constitutional testing because of a strong family history of Lynch syndrome cancers. Pathogenic variants were identified in 9/33 (27%) women who underwent constitutional testing, aged 33-59 years (median 48 years), including one whose tumour was mismatch repair proficient. Most Lynch syndrome tumours were of endometrioid histological subtype. CONCLUSIONS: Tumour mismatch repair deficiency identified by immunohistochemistry is a useful prescreen for constitutional testing in women with ovarian cancer with personal or family histories suggestive of Lynch syndrome.

The proportion of endometrial tumours associated with Lynch syndrome (PETALS): A prospective cross-sectional study.

BACKGROUND: Lynch syndrome (LS) predisposes to endometrial cancer (EC), colorectal cancer, and other cancers through inherited pathogenic variants affecting mismatch-repair (MMR) genes. Diagnosing LS in women with EC can reduce subsequent cancer mortality through colonoscopic surveillance and aspirin chemoprevention; it also enables cascade testing of relatives. A growing consensus supports LS screening in EC; however, the expected proportion of test positives, and optimal testing strategy is uncertain. Previous studies from insurance-based healthcare systems were limited by narrow selection criteria, failure to apply reference standard tests consistently, and poor conversion to definitive testing. The aim of this study was to establish the prevalence of LS and the diagnostic accuracy of LS testing strategies in an unselected EC population. METHODS AND FINDINGS: This was a prospective cross-sectional study carried out at a large United Kingdom gynaecological cancer centre between October 2015 and January 2017. Women diagnosed with EC or atypical hyperplasia (AH) were offered LS testing. Tumours underwent MMR immunohistochemistry (IHC), microsatellite instability (MSI), and targeted MLH1-methylation testing. Women <50 years, with strong family histories and/or indicative tumour molecular features, underwent MMR germline sequencing. Somatic MMR sequencing was performed when indicative molecular features were unexplained by LS or MLH1-hypermethylation. The main outcome measures were the prevalence of LS in an unselected EC population and the diagnostic accuracy of clinical and tumour testing strategies for risk stratifying women with EC for MMR germline sequencing. In total, 500 women participated in the study; only 2 (<1%) declined. Germline sequencing was indicated and conducted for 136 and 135 women, respectively. A total of 16/500 women (3.2%, 95% CI 1.8% to 5.1%) had LS, and 11 more (2.2%) had MMR variants of uncertain significance. Restricting testing to age <50 years, indicative family history (revised Bethesda guidelines or Amsterdam II criteria) or endometrioid histology alone would have missed 9/16 (56%), 8/13 (62%) or 9/13 (69%), and 5/16 (31%) cases of LS, respectively. In total 132/500 tumours were MMR deficient by IHC of which 83/132 (63%) had MLH1-hypermethylation, and 16/49 (33%) of the remaining patients had LS (16/132 with MMR deficiency, 12%). MMR-IHC with targeted MLH1-methylation testing was more discriminatory for LS than MSI with targeted methylation testing, with 100% versus 56.3% (16/16 versus 9/16) sensitivity (p = 0.016) and equal 97.5% (468/484) specificity; 64% MSI-H and 73% MMR deficient tumours unexplained by LS or MLH1-hypermethylation had somatic MMR mutations. The main limitation of the study was failure to conduct MMR germline sequencing for the whole study population, which means that the sensitivity and specificity of tumour triage strategies for LS detection may be overestimated, although the risk of LS in women with no clinical or tumour predictors is expected to be extremely low. CONCLUSIONS: In this study, we observed that age, family history, and histology are imprecise clinical correlates of LS-EC. IHC outperformed MSI for tumour triage and reliably identified both germline and somatic MMR mutations. The 3.2% proportion of LS-EC is similar to colorectal cancer, supporting unselected screening of EC for LS.

Detecting Endometrial Cancer by Blood Spectroscopy: A Diagnostic Cross-Sectional Study.

Endometrial cancer is the sixth most common cancer in women, with a rising incidence worldwide. Current approaches for the diagnosis and screening of endometrial cancer are invasive, expensive or of moderate diagnostic accuracy, limiting their clinical utility. There is a need for cost-effective and minimally invasive approaches to facilitate the early detection and timely management of endometrial cancer. We analysed blood plasma samples in a cross-sectional diagnostic accuracy study of women with endometrial cancer (n = 342), its precursor lesion atypical hyperplasia (n = 68) and healthy controls (n = 242, total n = 652) using attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy and machine learning algorithms. We show that blood-based infrared spectroscopy has the potential to detect endometrial cancer with 87% sensitivity and 78% specificity. Its accuracy is highest for Type I endometrial cancer, the most common subtype, and for atypical hyperplasia, with sensitivities of 91% and 100%, and specificities of 81% and 88%, respectively. Our large-cohort study shows that a simple blood test could enable the early detection of endometrial cancer of all stages in symptomatic women and provide the basis of a screening tool in high-risk groups. Such a test has the potential not only to differentially diagnose endometrial cancer but also to detect its precursor lesion atypical hyperplasia-the early recognition of which may allow fertility sparing management and cancer prevention.

Baseline Serum HE4 But Not Tissue HE4 Expression Predicts Response to the Levonorgestrel-Releasing Intrauterine System in Atypical Hyperplasia and Early Stage Endometrial Cancer.

The levonorgestrel-releasing intrauterine system (LNG-IUS) is a conservative management option for atypical hyperplasia (AH) and low grade early stage endometrial cancer (EEC), but around 1 in 3 patients fail to respond to treatment. The aim of this study was to investigate if serum and/or tissue HE4 expression could predict response to LNG-IUS therapy. Patients with AH or presumed Stage I EEC had serum and endometrial samples taken at baseline and at 3-month intervals over 12 months post-insertion of LNG-IUS. 74 patients were recruited and baseline demographics recorded. Of 57 patients for whom response was histologically determinable, 39 (68%) were responders and 18 (32%) non-responders. Mean baseline serum HE4 was significantly lower in responders (62.1 ± 1.1 pM, 95% confidence interval (CI) 52.7-73.2), compared to non-responders (125.6 ± 1.3 pM, 95% CI 74.5-211.7, p = 0.014), including when considering age, BMI, menopausal status, smoking status, and histological grade as covariables (p = 0.005). Baseline tissue HE4 expression was not significantly different in responders compared to non-responders (p = 0.999). Responders showed a significant mean reduction (-9.8 ± 3.4%, 95% CI -16.7 to -2.8%, p = 0.008) in serum HE4 between baseline and 3 months (p = 0.008), whereas non-responders showed no significant change (p = 0.676). Neither responders nor non-responders showed a significant percentage change in serum HE4 from baseline beyond 3 months (p > 0.05). Change in serum HE4 between baseline and 3 and 6 months and tissue HE4 tissue expression between baseline and 3, 6, and 12 months was not significantly different in responders compared to non-responders (p > 0.05). This study suggests that baseline serum HE4, but not baseline tissue HE4 expression, is independently predictive of response to the LNG-IUS and could be used to guide management decisions.

The impact of obesity and bariatric surgery on circulating and tissue biomarkers of endometrial cancer risk.

Obesity is the strongest risk factor for endometrial cancer (EC). To inform targeted screening and prevention strategies, we assessed the impact of obesity and subsequent bariatric surgery-induced weight loss on endometrial morphology and molecular pathways implicated in endometrial carcinogenesis. Blood and endometrial tissue were obtained from women with class III-IV obesity (body mass index ≥40 and ≥50 kg/m2 , respectively) immediately prior to gastric bypass or sleeve gastrectomy, and at two and 12 months' follow up. The endometrium underwent pathological examination and immunohistochemistry was used to quantify proliferation (Ki-67), oncogenic signaling (PTEN, pAKT, pERK) and hormone receptor (ER, PR) expression status. Circulating biomarkers of insulin resistance, reproductive function and inflammation were also measured at each time point. Seventy-two women underwent bariatric surgery. At 12 months, the mean change in total and excess body weight was -32.7 and -62.8%, respectively. Baseline endometrial biopsies revealed neoplastic change in 10 women (14%): four had EC, six had atypical hyperplasia (AH). After bariatric surgery, most cases of AH resolved (5/6) without intervention (3/6) or with intrauterine progestin (2/6). Biomarkers of endometrial proliferation (Ki-67), oncogenic signaling (pAKT) and hormone receptor status (ER, PR) were significantly reduced, with restoration of glandular PTEN expression, at 2 and 12 months. There were reductions in circulating biomarkers of insulin resistance (HbA1c, HOMA-IR) and inflammation (hsCRP, IL-6), and increases in reproductive biomarkers (LH, FSH, SHBG). We found an unexpectedly high prevalence of occult neoplastic changes in the endometrium of women undergoing bariatric surgery. Their spontaneous reversal and accompanying down-regulation of PI3K-AKT-mTOR signaling with weight loss may have implications for screening, prevention and treatment of this disease.

Monocarboxylate Transporter 1 (MCT1) is an independent prognostic biomarker in endometrial cancer.

BACKGROUND: Endometrial cancer (EC) is a major health concern due to its rising incidence. Whilst early stage disease is generally cured by surgery, advanced EC has a poor prognosis with limited treatment options. Altered energy metabolism is a hallmark of malignancy. Cancer cells drive tumour growth through aerobic glycolysis and must export lactate to maintain intracellular pH. The aim of this study was to evaluate the expression of the lactate/proton monocarboxylate transporters MCT1 and MCT4 and their chaperone CD147 in EC, with the ultimate aim of directing future drug development. METHODS: MCT1, MCT4 and CD147 expression was examined using immunohistochemical analysis in 90 endometrial tumours and correlated with clinico-pathological characteristics and survival outcomes. RESULTS: MCT1 and MCT4 expression was observed in the cytoplasm, the plasma membrane or both locations. CD147 was detected in the plasma membrane and associated with MCT1 (p = 0.003) but not with MCT4 (p = 0.207) expression. High MCT1 expression was associated with reduced overall survival (p = 0.029) and remained statistically significant after adjustment for survival covariates (p = 0.017). CONCLUSION: Our data suggest that MCT1 expression is an important marker of poor prognosis in EC. MCT1 inhibition may have potential as a treatment for advanced or recurrent EC.

Ancillary testing in liquid based cytology to distinguish cervical glandular neoplasia from tuboendometrial metaplasia in a young woman.

A 33-year old woman had a cervical sample taken at colposcopy clinic. Seven years prior to this, at the age of 26, she had had a cytological diagnosis of cervical glandular neoplasia (cytology descriptor indicated cells suspicious of endocervical neoplasia) and severe dyskaryosis. Confirmation and treatment were by LLETZ and knife cone, and, in keeping with England and Wales National Health Service guidelines, this woman was under follow-up by the colposcopy clinic. Intervening cytological follow-up included a number of negative cytological samples interspaced with one equivocal report. A recent repeat cytology which was rather cellular contained several hyperchromatic crowded cell groups (HCCG's). Careful examination revealed benign endometrial clusters, LUS, TEM and endocervical cells in strips showing pseudostratification and loss of polarity. Following an agar block, there was positive staining for p16 and Ki-67 in the abnormal groups whilst the benign TEM cells stained positive for bcl-2.

How long is urinary cytology abnormal after flexible cystoscopy?

OBJECTIVE: To examine the urinary cytological changes caused by flexible cystoscopy and provide clinical guidelines for a reliable time interval for urinary cytological examination after flexible cystoscopy. PATIENTS AND METHODS: Forty-eight patients attending for flexible cystoscopy were recruited into the study. Each patient was asked to provide eight urine samples before, immediately after and at 1, 2, 7, 14 and 28 days after cystoscopy. Cytospin preparations of the urine samples were made and slides stained using the Papanicolaou stain. Cytology was analysed while unaware of sample origin, by three different cytopathologists. RESULTS: The cytological changes were characterized by a striking increase in cellularity immediately after flexible cystoscopy, mostly accounted for by urothelial cells. Consistent morphological changes included the formation of 'columnar' cells, papillary clusters, increased nucleo-cytoplasmic ratio and nuclear atypia. These changes were transient, with most disappearing within a day of flexible cystoscopy. CONCLUSIONS: There are cytological changes, on voided urine cytology, after flexible cystoscopy but they were transient, and urine sent more than a day after flexible cystoscopy should be free from artefactual change caused by instrumentation. These results suggest that clinicians sending urine for cytological analysis should provide information about the nature and timing of any endoscopy so as to avoid false-positive interpretations of urine cytology by the cytopathologist.

Immunological and clinical responses in women with vulval intraepithelial neoplasia vaccinated with a vaccinia virus encoding human papillomavirus 16/18 oncoproteins.

This study assessed the immunological and clinical responses of women with human papillomavirus (HPV) 16-associated high-grade vulval intraepithelial neoplasia (VIN) vaccinated with TA-HPV, a recombinant vaccinia virus encoding modified HPV 16 and 18 E6 and E7. Eighteen women with HPV 16-positive high-grade VIN were vaccinated with TA-HPV. The extent of their baseline disease was compared after 24 weeks by lesion measurements and histological analysis. Viral load was assessed pre- and postvaccination by real time PCR. Cell-mediated immunity to HPV 16 E6 and/or E7 peptides (HLA-A2 epitopes) or vaccinia-infected cell lysates was determined by IFN-gamma enzyme-linked immunospot (ELISPOT) and T cell proliferation using an HPV 16 L2E6E7 fusion protein. Antibodies were measured by ELISA using vaccinia-infected cell lysates or HPV 16 and 18 E6 and E7 glutathione S-transferase-fusion proteins. Lesion-infiltrating CD4(+), CD8(+), CD1a(+), and CD68(+) immune cells were assessed by immunohistochemistry. The single vaccination with TA-HPV was well tolerated, and all patients showed an increased ELISPOT and/or antibody response to vaccinia. There were significant differences in HPV-16 E7-specific ELISPOT and L2E6E7 proliferative responses in the patients at one or more time points postvaccination as compared with the prevaccination status; two patients showed transient increased antibody responses. Overall, 13 women showed an increased HPV 16-specific immune response by one or more methodologies after immunization. Eight patients demonstrated a reduction in lesion diameter of at least 50% and a further four patients showed significant symptom relief. Viral load was reduced or cleared in six of eight lesion responders but also in six of ten nonresponders. Before vaccination, clinical responders had significantly higher levels of lesion-associated CD4(+), CD8(+), and CD1a(+)-immune cells than nonresponders. There were no differences in CD68 (macrophages) between responders and nonresponders before or after vaccination. Nonresponders did show a significant increase in CD4(+)- and CD8(+)- but not CD1a(+)-immune cells postvaccination but at lower levels overall than responder patients. Local immune infiltration may be a critical factor in potential responsiveness to vaccine therapy in HPV-associated neoplasia and should be carefully monitored in future placebo-controlled trials of immunotherapy for VIN.

Carcinoid tumor originating in a horseshoe kidney.

An atypical carcinoid tumor originating in the horseshoe kidney of a 39-year-old man is reported. The tumor showed cytological atypia, increased mitotic rate and histological signs of mucinous differentiation and osseous metaplasia. Even though there were distant metastases, the patient survived partial nephrectomy including resection of the primary tumor for four years. The review of literature shows that carcinoid tumors originate some 60 to 85 times more often in horseshoe kidneys than in normal kidneys. The clinical course of renal carcinoid tumors is highly variable.