The aim of this study is to analyse the association between vaginal microbiota and the histological finding of CIN. From July 2016 until June 2017, we included 110 consecutive patients with abnormal cervical cytology results referred for colposcopy to Riga East Clinical University Hospital Outpatient department in the study group. 118 women without cervical pathology were chosen as controls. Certified colposcopists performed interviews, gynaecological examinations and colposcopies for all participants. Material from the upper vaginal fornix was taken for pH measurement and wet-mount microscopy. Cervical biopsy samples were taken from all subjects in the study group and in case of a visual suspicion for CIN in the control group. Cervical pathology was more often associated with smoking (34.6% vs. 11.0%, p < 0.0001), low education level (47.2% vs. 25.5%, p = 0.001), increased vaginal pH (48.2% vs. 25.4%, p < 0.0001), abnormal vaginal microbiota (50% vs. 31.4%, p = 0.004) and moderate to severe aerobic vaginitis (msAV) (13.6% vs. 5.9%, p = 0.049) compared to controls. The most important independent risk factors associated with CIN2+ were smoking (OR 3.04 (95% CI 1.37–6.76), p = 0.006) and msAV (OR 3.18 (95% CI 1.13–8.93), p = 0.028). Bacterial vaginosis (BV) was found more often in CIN1 patients (8/31, 25.8%, p = 0.009) compared with healthy controls (8/118, 6.8%), or CIN2+ cases (8/79, 10.1%). In the current study msAV and smoking were the most significant factors in the development of CIN in HPV-infected women, especially high grade CIN. We suggest that AV changes are probably more important than the presence of BV in the pathogenesis of CIN and progression to cervix cancer and should not be ignored during the evaluation of the vaginal microbiota.

We aimed to determine the diagnostic value of anti-parietal cell antibodies (anti-PCA), anti-intrinsic factor antibodies (anti-IFA), pepsinogen ratio (PGI/II), and gastrin-17 (G-17) in corpus-restricted atrophic gastritis (CRAG) detected by ELISA (Inova, Biohit). Our study compared 29 CRAG cases against 58 age- and sex-matched controls with mild or no atrophy. Anti-PCA and anti-IFA positive cutoff values were ≥25 units for both. PGI/II value 5 pg/L and >30 EIU. Anti-PCA was positive in 65.5% For CRAG cases and 13.8% of the controls (p < 0.0001), anti-IFA was positive in 13.8% and 0% (p = 0.01), respectively. Decreased pepsinogen levels were present in 79.3% of CRAG cases and 10.3% of the controls (p < 0.0001). PGI/II ratio was the best single biomarker, with sensitivity = 79%, specificity = 90%, and AUC 0.90. The combined use of PGI/II and anti-PCA resulted in AUC 0.93 for detecting CRAG. Our study suggests that the best combination of non-invasive biomarkers for detecting CRAG is PGI/II with anti-PCA. The addition of G-17 and anti-IFA is of little utility in clinical application.

Introduction––Serum pepsinogen tests for gastric cancer screening have been debated for decades. We assessed the performance of two pepsinogen assays with or without gastrin-17 for the detection of different precancerous lesions alone or as a composite endpoint in a Latvian cohort. Methods––Within the intervention arm of the GISTAR population-based study, participants with abnormal pepsinogen values by ELISA or latex-agglutination tests, or abnormal gastrin-17 by ELISA and a subset of subjects with all normal biomarker values were referred for upper endoscopy with biopsies. Performance of biomarkers, corrected by verification bias, to detect five composite outcomes based on atrophy, intestinal metaplasia, dysplasia or cancer was explored. Results––Data from 1045 subjects were analysed, of those 273 with normal biomarker results. Both pepsinogen assays showed high specificity (>93%) but poor sensitivity (range: 18.4–31.1%) that slightly improved when lesions were restricted to corpus location (40.5%) but decreased when dysplasia and prevalent cancer cases were included (23.8%). Adding gastrin-17 detection, sensitivity reached 33–45% while specificity decreased (range: 61.1–62%) and referral rate for upper endoscopy increased to 38.6%. Conclusions––Low sensitivity of pepsinogen assays is a limiting factor for their use in population-based primary gastric cancer screening, however their high specificity could be useful for triage.

Over the past decade, extracellular vesicles (EVs) have emerged as a promising source of cancer-derived RNAs for liquid biopsies. However, blood contains a pool of heterogeneous EVs released by a variety of cell types, making the identification of cancer RNA biomarkers challenging. Here, we performed deep sequencing of plasma EV RNA cargo in 32 patients with locally advanced breast cancer (BC) at diagnosis and 7 days after breast surgery and in 30 cancer-free healthy controls (HCs). To identify BC-derived RNA biomarkers, we searched for RNAs that had higher levels in BC EVs at the time of diagnosis compared with HCs and decreased after surgery. Data analysis showed that the fractions of miRNAs, snRNAs, snoRNAs, and tRFs were increased, but the fraction of lncRNAs was decreased in BC EVs as compared to HCs. BC-derived biomarker candidates were identified across various RNA biotypes. Considered individually, they had very high specificity but moderate sensitivity for the detection of BC, whereas a biomarker model composed of eight RNAs: SNORD3H, SNORD1C, SNORA74D, miR-224-5p, piR-32949, lnc-IFT-122-2, lnc-C9orf50-4, and lnc-FAM122C-3 was able to distinguish BC from HC EVs with an AUC of 0.902 (95% CI = 0.872–0.931, p = 3.4 × 10⁻⁹) in leave-one-out cross-validation. Furthermore, a number of RNA biomarkers were correlated with the ER and HER2 expression and additional biomarker models were created to predict hormone receptor and HER2 status. Overall, this study demonstrated that the RNA composition of plasma EVs is altered in BC patients and that they contain cancer-derived RNA biomarkers that can be used for BC detection and monitoring using liquid biopsies.

Extracellular vesicles (EVs) are g7aining increased attention as carriers of cancer-derived molecules for liquid biopsies. Here, we studied the dynamics of EV levels in the plasma of breast cancer (BC) patients undergoing neoadjuvant chemotherapy (NAC) and explored the relevance of their RNA cargo for the prediction of patients’ response to the therapy. EVs were isolated from serial blood samples collected at the time of diagnosis, at the end of NAC, and 7 days, 6, and 12 months after the surgery from 32 patients with locally advanced BC, and 30 cancer-free healthy controls (HCs) and quantified by nanoparticle tracking analysis. The pre-treatment levels of EVs in BC patients were higher than in HCs, significantly increased during the NAC and surgery, and decreased to the levels found in HCs 6 months after surgery, thus showing that a substantial fraction of plasma EVs in BC patients are produced due to the disease processes and treatment. RNA sequencing analysis revealed that the changes in the EV levels were associated with the alterations in the proportions of various RNA biotypes in EVs. To search for RNA biomarkers that predict response to the NAC, patients were dichotomized as responders and non-responders based on Miller-Payne grades and differential expression analyses were carried out between responders and non-responders, and HCs. This resulted in the identification of 6 miRNAs, 4 lncRNAs, and 1 snoRNA that had significantly higher levels in EVs from non-responders than responders at the time of diagnosis and throughout the NAC, and significantly lower levels in HCs, thus representing biomarkers for the prediction of response to NAC at the time of diagnosis. In addition, we found 14 RNAs representing piRNA, miRNA, lncRNA, snoRNA, and snRNA biotypes that were induced by NAC in non-responders and 2 snoRNAs and 1 piRNA that were induced by NAC in patients with early disease progression, thus warranting further functional studies on their role in chemoresistance and metastasis.

Stromal tumour infiltrating lymphocytes (sTILs) are a strong prognostic marker in triple negative breast cancer (TNBC). Consistency scoring sTILs is good and was excellent when an internet-based scoring aid developed by the TIL-WG was used to score cases in a reproducibility study. This study aimed to evaluate the reproducibility of sTILs assessment using this scoring aid in cases from routine practice and to explore the potential of the tool to overcome variability in scoring. Twenty-three breast pathologists scored sTILs in digitized slides of 49 TNBC biopsies using the scoring aid. Subsequently, fields of view (FOV) from each case were selected by one pathologist and scored by the group using the tool. Inter-observer agreement was good for absolute sTILs (ICC 0.634, 95% CI 0.539–0.735, p < 0.001) but was poor to fair using binary cutpoints. sTILs heterogeneity was the main contributor to disagreement. When pathologists scored the same FOV from each case, inter-observer agreement was excellent for absolute sTILs (ICC 0.798, 95% CI 0.727–0.864, p < 0.001) and good for the 20% (ICC 0.657, 95% CI 0.561–0.756, p < 0.001) and 40% (ICC 0.644, 95% CI 0.546–0.745, p < 0.001) cutpoints. However, there was a wide range of scores for many cases. Reproducibility scoring sTILs is good when the scoring aid is used. Heterogeneity is the main contributor to variance and will need to be overcome for analytic validity to be achieved.