Umeå University's logo

RNA modification pathways are often misregulated in various cancers, with N6-methyladenosine (m6A) having a pivotal role in cancer progression and metastasis. Methyltransferase-like 3 (METTL3), a core component of the m6A methyltransferase complex, not only functions as an m6A writer but also promotes tumorigenesis through m6A-independent mechanisms. Here, we show that METTL3 is mislocalized to the cytoplasm in breast cancer tumors from patients, contributing to the oncogenic phenotype. Cytoplasmic METTL3 interacts with EXOC7, a key regulator of exocytosis, promoting its stabilization. In addition, METTL3 regulates m6A-dependent alternative splicing of EXOC7. Silencing METTL3 impairs vesicle trafficking and the breast cancer secretome-effects that do not rely on its enzymatic activity but instead involve METTL3-mediated stabilization of EXOC7 and potentially other exocyst components. Furthermore, METTL3 knockdown impairs invadopodia formation, collagen matrix invasion, and focal adhesion morphology in vitro, while inhibition of METTL3 catalytic activity does not. Our findings uncover noncatalytic roles of METTL3 in regulating exocytosis and the cancer secretome.

Background: Nanoparticle immunoassays for CA125 and CA15-3 glycovariants are promising tools for epithelial ovarian cancer (EOC) and borderline ovarian tumor (BOT) early detection.

Method: This retrospective population-based study utilized prospective (< 3 years) plasma samples from 110 cases and 440 controls. Sialyl-Thomsen-nouveau (STn) antibody and macrophage galactose-type lectin (MGL) to detect cancer antigen 125 (CA125) and 15 − 3 (CA15-3) glycoforms were evaluated, using CA125, CA15-3, and HE4 enzyme immunoassay (EIA) references. The area under the receiver operating characteristic curve (AUC), partial AUC (pAUC) at specificities 0.9–1.0, and sensitivity (SN) at 0.98 specificity (SP) for detecting EOC/BOT were calculated.

Results: The single marker with the highest point estimate for pAUC and SN at 98% SP was CA125EIA (pAUC 0.71 (0.66, 0.76), SN at 98% SP 0.29 (0.17–0.42 combinations)). The CA15-3STn + CA125EIA and CA125EIA + HE4EIA improved the detection rate point estimates and provided the highest pAUC values (CA125EIA + CA15-3STn: 0.75 (0.70–0.80); CA125EIA + HE4EIA: 0.71 (0.66–0.77)). For 0–<1, 1–<2 and 2–<3 years lag time, CA125EIA and the CA125 glycovariants provided similar pAUC and SN at 98% SP, with largely overlapping confidence intervals.

Conclusion: This case-control study, which used plasma samples collected up to three years before diagnosis of EOC or BOT, did not provide evidence of a higher discriminatory capacity of the glycovariant biomarkers compared with the clinically established biomarker CA125EIA in asymptomatic women. Increased plasma volumes and larger cohorts are suggested in future studies.

Objective: To evaluate diagnostic accuracy of novel nanoparticle immunoassays across different histotypes of tubo-ovarian carcinoma (TOC) at diagnosis.

Method: This multicenter observational study consisted of consecutive patients (n = 1,312) having surgery due to suspected ovarian pathology. Serum were analyzed with Sialyl-Thomsen-nouveau (STn) antibody and Macrophage-Galactose-Lectin (MGL) for the detection of cancer antigen 125 (CA125) and 15-3 (CA15-3) glycoforms using CA125 enzyme immunoassay (EIA), CA15-3EIA and HE4EIA as references. Receiver operating characteristics (ROC) were applied and sensitivity at 75 % and 98 % specificity were calculated across histotypes.

Result: TOC was present in 596 patients and 716 had benign disease. CA125STn showed higher sensitivity at 98 % specificity compared with CA125EIA for high grade serous ovarian carcinoma (HGSC) (0.85 vs 0.62, p < 0.001), HGSC early-stage (0.66 vs 0.24, p = 0.003), and HGSC late-stage (0.90 vs 0.69, p < 0.001). CA15-3STn showed higher sensitivity at 98 % specificity compared to CA125EIA for mucinous ovarian carcinoma (0.50 vs 0.16, p = 0.038). No improvements were found for low grade serous carcinoma (LGSC), endometrioid and clear cell histotypes. The best performing combined biomarker test was CA125STn + HE4EIA with higher sensitivity at 98 % specificity for HGSC (0.93 vs 0.86, p < 0.001) and HGSC late-stage (0.97 vs 0.91p < 0.001) compared with CA125EIA + HE4EIA.

Conclusion: The STn glycovariants of CA125 and CA15-3 have improved sensitivity at high specificity for high grade serous and mucinous ovarian carcinoma and often perform better than the commonly used biomarker CA125EIA.

The resemblance between fallopian tube cells and serous borderline ovarian tumors (BOTs) suggests a potential origin link, with salpingitis proposed as a contributing factor in the pathogenesis of BOT. This study aimed to explore the potential association between pelvic inflammatory disease (PID) and the risk of developing BOT. A national population-based case–control study in Sweden included women with BOT between 1999 and 2020 and 10 matched controls. Data from nationwide registers were analyzed using conditional logistic regression, adjusting for age, residential district, educational level and parity. Among 4782 cases and 45,167 controls, 2.0% of cases and 1.3% of controls had a history of PID. Previous PID was associated with an increased risk of BOT overall (aOR, 1.48; 95% CI, 1.19–1.85). Significant association was observed with serous tumors (aOR, 1.76; 95% CI, 1.36–2.29), while not with mucinous tumors (aOR, 0.95; 95% CI, 0.60–1.49). A dose–response relationship between number of PID episodes and serous BOT risk was noted (Ptrend <.001). This study demonstrates that PID is associated with increased risk of serous BOT, with a dose response relationship. The study highlights the potential serious implications of upper reproductive tract infections and inflammation. This underscores the need for further investigation of biological mechanisms and possible impact of PID on serous BOT development.

Endometriosis, affecting 10% of women, is defined as implantation, survival, and growth of endometrium-like/endometriotic tissue outside the uterine cavity, causing inflammation, infertility, pain, and susceptibility to ovarian cancer. Despite extensive studies, its etiology and pathogenesis are poorly understood and largely unknown. The prevailing view is that the immune system of endometriosis patients fails to clear ectopically disseminated endometrium from retrograde menstruation. Exosomes are small extracellular vesicles that exhibit immunomodulatory properties. We studied the role of endometriotic tissue-secreted exosomes in the pathophysiology of endometriosis. Two exosome-mediated mechanisms known to impair the immune response were investigated: 1) downregulation of NKG2D-mediated cytotoxicity and 2) FasL- and TRAIL-induced apoptosis of activated immune cells. We showed that secreted endometriotic exosomes isolated from supernatants of short-term explant cultures carry the NKG2D ligands MICA/B and ULBP1-3 and the proapoptotic molecules FasL and TRAIL on their surface, i.e., signature molecules of exosome-mediated immune suppression. Acting as decoys, these exosomes downregulate the NKG2D receptor, impair NKG2D-mediated cytotoxicity, and induce apoptosis of activated PBMCs and Jurkat cells through the FasL- and TRAIL pathway. The secreted endometriotic exosomes create an immunosuppressive gradient at the ectopic site, forming a “protective shield” around the endometriotic lesions. This gradient guards the endometriotic lesions against clearance by a cytotoxic attack and creates immunologic privilege by induction of apoptosis in activated immune cells. Taken together, our results provide a plausible, exosome-based mechanistic explanation for the immune dysfunction and the compromised immune surveillance in endometriosis and contribute novel insights into the pathogenesis of this enigmatic disease.

Endometriosis, affecting 10% of women, is defined as implantation, survival, and growth of endometriumlike/endometriotic tissue outside the uterine cavity, causing inflammation, infertility, pain andsusceptibility to ovarian cancer. Despite extensive studies, its etiology and pathogenesis are poorlyunderstood and largely unknown. The prevailing view is that the immune system of endometriosispatients fails to clear ectopically disseminated endometrium from retrograde menstruation. Exosomes aresmall extracellular vesicles that exhibit immunomodulatory properties. We studied the role ofendometriotic tissue-secreted exosomes in the pathophysiology of endometriosis. Two exosome-mediatedmechanisms known to impair the immune response were investigated: 1) downregulation of NKG2Dmediatedcytotoxicity and 2) FasL- and TRAIL-induced apoptosis of activated immune cells. We showedthat secreted endometriotic exosomes isolated from supernatants of short-term explant cultures carry theNKG2D ligands MICA/B and ULBP1-3; and the proapoptotic molecules FasL and TRAIL on theirsurface, i.e. signature molecules of exosome-mediated immune suppression. Acting as decoys, theseexosomes downregulate the NKG2D receptor, impair NKG2D-mediated cytotoxicity and induce apoptosisof activated PBMC and Jurkat cells through the FasL- and TRAIL pathway. The secreted endometrioticexosomes create an immunosuppressive gradient at the ectopic site, forming a “protective shield” aroundthe endometriotic lesions. This gradient guards the endometriotic lesions against clearance by a cytotoxicattack and creates immunologic privilege by induction of apoptosis in activated immune cells. Takentogether, our results provide a plausible, exosome-based mechanistic explanation for the immunedysfunction and the compromised immune surveillance in endometriosis and contribute with novelinsights into the pathogenesis of this enigmatic disease.

Background: Epithelial ovarian cancer is an insidious disease, and women are often diagnosed when the disease is beyond curative treatment. Accordingly, identifying modifiable risk factors is of paramount importance. Inflammation predisposes an individual to cancer in various organs, but whether pelvic inflammatory disease is associated with an increased risk of epithelial ovarian cancer has not been fully determined.

Objective: This study aimed to investigate a possible association between clinically verified pelvic inflammatory disease and the risk of epithelial ovarian cancer.

Study Design: In this national population-based case-control study, all women in Sweden diagnosed with epithelial ovarian cancer between 1999 and 2020 and 10 controls for each were identified, matched for age and residential district. Using several Swedish nationwide registers, data on previous pelvic inflammatory disease and potential confounding factors (age, parity, educational level, and previous gynecologic surgery) were retrieved. Adjusted odds ratios and 95% confidence intervals were estimated using conditional logistic regression. Histotype-specific analyses were performed for the subgroup of women diagnosed with epithelial ovarian cancer between 2015 and 2020. Moreover, hormonal contraceptives and menopausal hormone therapy were adjusted in addition to the aforementioned confounders.

Results: This study included 15,072 women with epithelial ovarian cancer and 141,322 controls. Most women (9102 [60.4%]) had serous carcinoma. In a subgroup of cases diagnosed between 2015 and 2020, high-grade serous carcinoma (2319 [60.0%]) was identified. A total of 168 cases (1.1%) and 1270 controls (0.9%) were diagnosed with pelvic inflammatory disease. Previous pelvic inflammatory disease was associated with an increased risk of epithelial ovarian cancer (adjusted odds ratio, 1.39; 95% confidence interval, 1.17–1.66) and serous carcinoma (adjusted odds ratio, 1.46; 95% confidence interval, 1.18–1.80) for the entire study population. For the subgroup of women diagnosed in 2015–2020, pelvic inflammatory disease was associated with high-grade serous carcinoma (adjusted odds ratio, 1.43; 95% confidence interval, 1.01–2.04). The odds ratios of the other histotypes were as follows: endometrioid (adjusted odds ratio, 0.13; 95% confidence interval, 0.02–1.06), mucinous (adjusted odds ratio, 1.55; 95% confidence interval, 0.56–4.29), and clear cell carcinoma (adjusted odds ratio, 2.30; 95% confidence interval, 0.90–5.86). A dose-response relationship was observed between the number of pelvic inflammatory disease episodes and the risk of epithelial ovarian cancer (P_trend<.001).

Conclusion: A history of pelvic inflammatory disease is associated with an increased risk of epithelial ovarian cancer and a dose-response relationship is evident. Histotype-specific analyses show an association with increased risk of serous epithelial ovarian cancer and high-grade serous carcinoma and potentially also with clear cell carcinoma, but there is no significant association with other histotypes. Infection and inflammation of the upper reproductive tract might have serious long-term consequences, including epithelial ovarian cancer.

Problem: Tumors compromise the patients’ immune system to promote their own survival. We have previously reported that HGSC exosomes play a central role, downregulating NKG2D cytotoxicity. Primary surgery's effect on tumor exosomes and NKG2D cytotoxicity in HGSC patients has not been studied before. The overall objective of this study was to explore the effect of surgery on the exosome-induced impairment of NKG2D cytotoxicity in HGSC.

Method of study: Paired pre- and post-operative blood samples were subjected to cell and exosome analyses regarding the NKG2D receptor and ligands, and NKG2D-mediated cytotoxicity. Lymphocytes were phenotyped by immunoflow cytometry. Exosomes, isolated by ultracentrifugation, and characterized by nanoparticle tracking analysis, transmission and immune electron microscopy and western blot were used in functional cytotoxic experiments. HGSC explant culture-derived exosomes, previously studied by us, were used for comparison.

Results: HGSC exosomes from patients’ sera downregulated NKG2D-mediated cytotoxicity in NK cells of healthy donors. In a subgroup of subjects, NKG2D expression on CTLs and NK cells was upregulated after surgery, correlating to a decrease in the concentration of exosomes in postoperative sera. An overall significantly improved NKG2D-mediated cytotoxic response of the HGSC patients’ own NK cells in postoperative compared to preoperative samples was noted.

Conclusions: Surgical removal of the primary tumor has a beneficial effect, relieving the exosome-mediated suppression of NKG2D cytotoxicity in HGSC patients, thus boostering their ability to combat cancer.

Background: CA125 is the best available yet insufficiently sensitive biomarker for early detection of ovarian cancer. There is a need to identify novel biomarkers, which individually or in combination with CA125 can achieve adequate sensitivity and specificity for the detection of earlier-stage ovarian cancer.

Methods: In the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort, we measured serum levels of 92 preselected proteins for 91 women who had blood sampled ≤18 months prior to ovarian cancer diagnosis, and 182 matched controls. We evaluated the discriminatory performance of the proteins as potential early diagnostic biomarkers of ovarian cancer.

Results: Nine of the 92 markers; CA125, HE4, FOLR1, KLK11, WISP1, MDK, CXCL13, MSLN and ADAM8 showed an area under the ROC curve (AUC) of ≥0.70 for discriminating between women diagnosed with ovarian cancer and women who remained cancer-free. All, except ADAM8, had shown at least equal discrimination in previous case-control comparisons. The discrimination of the biomarkers, however, was low for the lag-time of >9–18 months and paired combinations of CA125 with any of the 8 markers did not improve discrimination compared to CA125 alone.

Conclusion: Using pre-diagnostic serum samples, this study identified markers with good discrimination for the lag-time of 0–9 months. However, the discrimination was low in blood samples collected more than 9 months prior to diagnosis, and none of the markers showed major improvement in discrimination when added to CA125.

INTRODUCTION: Endometrioid endometrial carcinoma is a cancer type with generally excellent prognosis when diagnosed at an early stage, but there is a subset of patients with relapsing disease in spite of early diagnosis and surgical treatment. There is a need to find prognostic markers to identify these patients with increased risk of relapse. Depth of myometrial invasion, histological grade, and presence of lymphovascular invasion are known risk factors. DNA content (ploidy) and proliferation measured as S-phase fraction (SPF) have been discussed as prognostic markers but need additional evaluation.

MATERIAL AND METHODS: We evaluated relapse-free survival (RFS) with respect to ploidy and SPF, which was analyzed by flow cytometry on fresh tumor tissue, in a cohort of 1001 women treated for stage I endometrioid endometrial carcinoma in northern Sweden during the period of 1993-2010, with a median follow up time of 12.0 years. Data were obtained from historical records.

RESULTS: In simple analysis, both aneuploidy and high SPF were associated to increased risk of relapse with hazard ratios (HR) 2.37 (95% CI 1.52-3.70) and 1.94 (95% CI 1.24-3.02), respectively. Our data also confirmed stage, tumor grade, and ploidy as independent prognostic markers in an age adjusted cox regression multivariable analysis but we did not find SPF to contribute to prognosis. However, the combination of aneuploidy and high SPF identified a group of patients with increased risk of relapse, HR 2.02 (95% CI 1.19-3.44).

CONCLUSION: In this study, which is the largest study of ploidy and SPF in stage I endometrioid endometrial carcinoma using fresh frozen tissue, aneuploidy was shown to be an independent prognostic marker. Furthermore, the combination of aneuploidy and high SPF could be used to identify patients with increased risk of relapse.