Modeling the Prognostic Impact of Circulating Tumor Cells Enumeration in Metastatic Breast Cancer for Clinical Trial Design Simulation.

Despite the strong prognostic stratification of circulating tumor cells (CTCs) enumeration in metastatic breast cancer (MBC), current clinical trials usually do not include a baseline CTCs in their design. This study aimed to generate a classifier for CTCs prognostic simulation in existing datasets for hypothesis generation in patients with MBC. A K-nearest neighbor machine learning algorithm was trained on a pooled dataset comprising 2436 individual MBC patients from the European Pooled Analysis Consortium and the MD Anderson Cancer Center to identify patients likely to have CTCs ≥ 5/7 mL blood (StageIVaggressive vs StageIVindolent). The model had a 65.1% accuracy and its prognostic impact resulted in a hazard ratio (HR) of 1.89 (Simulatedaggressive vs SimulatedindolentP < .001), similar to patients with actual CTCs enumeration (HR 2.76; P < .001). The classifier's performance was then tested on an independent retrospective database comprising 446 consecutive hormone receptor (HR)-positive HER2-negative MBC patients. The model further stratified clinical subgroups usually considered prognostically homogeneous such as patients with bone-only or liver metastases. Bone-only disease classified as Simulatedaggressive had a significantly worse overall survival (OS; P < .0001), while patients with liver metastases classified as Simulatedindolent had a significantly better prognosis (P < .0001). Consistent results were observed for patients who had undergone CTCs enumeration in the pooled population. The differential prognostic impact of endocrine- (ET) and chemotherapy (CT) was explored across the simulated subgroups. No significant differences were observed between ET and CT in the overall population, both in terms of progression-free survival (PFS) and OS. In contrast, a statistically significant difference, favoring CT over ET was observed among Simulatedaggressive patients (HR: 0.62; P = .030 and HR: 0.60; P = .037, respectively, for PFS and OS).

Evaluation of the cardiovascular risk in patients undergoing major non-cardiac surgery: role of cardiac-specific biomarkers.

Major adverse cardiovascular events are frequently observed in patients undergoing major non-cardiac surgery during the peri-operative period. At this time, the possibility to predict cardiovascular events remains limited, despite the introduction of several algorithms to calculate the risk of adverse events, mainly death and major adverse cardiovascular events (MACE) based on the clinical history, risk factors (sex, age, lipid profile, serum creatinine) and non-invasive cardiac exams (electrocardiogram, echocardiogram, stress tests). The cardiac-specific biomarkers natriuretic peptides (NPs) and cardiac troponins (cTn) have been proposed as additional tools for risk prediction in the peri-operative period, particularly for the identification of myocardial injury in patients undergoing major non-cardiac surgery. The prognostic information from the measurement of BNP/NT-proBNP and hs-cTn is independent and complementary to other important indicators of risk, also including ECG and imaging techniques. Elevated levels of cardiac-specific biomarkers before surgery are associated with a markedly higher risk of MACE during the peri-operative period. BNP/NT-proBNP and hs-cTn should be measured in all patients during the clinical evaluation before surgery, particularly during intermediate- or high-risk surgery, in patients aged >65 years and/or with comorbidities. Several questions remain to be assessed in dedicated clinical studies, such as how to optimize the management of patients with raised cardiac specific biomarkers before surgery, and whether a strategy based on biomarker measurement improves patient outcomes and is cost-effective.

Anyplex II HPV test in detection and follow-up after surgical treatment of CIN2+ lesions.

Human papillomavirus (HPV) tests differ for technology, targets, and information on the genotype and viral load. In this study, we evaluated the performance of the Seegene Anyplex II HPV HR (Anyplex) assay in the detection of cervical intraepithelial lesions (CIN) and as a test-of-cure in the follow-up after surgical treatment. One hundred and sixty-seven women referred to the European Institute of Oncology, Milan, for surgical treatment of CIN2+ were enrolled. A cervical sample was taken before treatment and at the first follow-up visit: on these samples, Qiagen Hybrid Capture 2 (HC2), Roche Linear Array HPV Test (Linear Array), cytology and histology were performed at baseline, HC2, and cytology at follow-up. Anyplex genotyping HPV test was performed on a post aliquot from liquid-based cytology specimens when available. The concordance between Anyplex and HC2 was 93.6% at baseline and 76.7% at follow-up (3-9 months after treatment), respectively. The concordance between Anyplex and Linear Array was evaluable only at baseline (92.9%). No recurrence occurred in women without the persistence of the same genotype at follow-up. Seven women relapsed: six had persistence of the same genotypes (five HPV16, one HPV33, and one HPV39), while one tested negative not only with Anyplex but also with HC2 for the persistence of low-risk genotype infection (HPV73 only detected by Linear Array). Anyplex test represents a valid option for HPV detection and genotyping in order to stratify women at risk of high-grade lesions at baseline and to monitor patients treated for CIN2+ lesions during follow-up.

High-sensitivity cardiac troponin I and T methods for the early detection of myocardial injury in patients on chemotherapy.

Important advances achieved in pharmacological cancer treatment have led progressively to a reduction in mortality from many forms of cancer, and increasing numbers of previously incurable patients can now hope to become cancer-free. Yet, to achieve these improved outcomes a high price has been paid in terms of untoward side effects associated with treatment, cardio-toxicity in particular. Several recent studies have reported that cardiac troponin assay using high-sensitivity methods (hs-cTn) can enable the early detection of myocardial injury related to chemotherapy or abuse of drugs that are potentially cardiotoxic. Several authors have recently suggested that changes in hs-cTn values enable the early diagnosis of cardiac injury from chemotherapy, thus potentially benefitting cancer patients with increased troponin values by initiating early cardioprotective therapy. However, large randomised clinical trials are needed in order to evaluate the cost/benefit ratio of standardised protocols for the early detection of cardiotoxicity using the hs-cTn assay in patients treated with chemotherapy.

Onclarity Performance in Human Papillomavirus DNA Detection in Formalin-Fixed Paraffin-Embedded Cervical Samples.

OBJECTIVES: Diagnosis of HPV infection is usually performed from cervical liquid-based cytology specimens (LBC), but these often contain a large amount of human papillomavirus (HPV) genotypes, most of which might cause transient infections. The aim of the study was to evaluate the performance of BD Onclarity HPV test genotyping method on formalin-fixed, paraffin-embedded (FFPE) cervical specimens compared with genotyping results from LBC. MATERIALS AND METHODS: Formalin-fixed, paraffin-embedded specimens from women surgically treated for cervical intraepithelial lesions (CINs) at the European Institute of Oncology, Milan, from September 2012 to June 2013 were retrieved from the archives of the Department of Pathology of the European Institute of Oncology. The FFPE and LBC specimens were genotyped using the same extended genotyping Onclarity assay. RESULTS: We collected 99 samples (26 CIN 1, 30 CIN 2, and 43 CIN 3+), but 15 were excluded from the analysis: these 84 samples show an overall agreement of 89% for HPV status between FFPE Onclarity samples versus LBC samples. The FFPE and LBC samples showed identical genotype in 75% samples, compatible genotype (at least 1 of the genotypes detected in LBC sample was found in the tissue sample) in 14% specimens, and discrepant genotype in 11% samples. CONCLUSIONS: Our data demonstrate a very good concordance between HPV genotypes found in cytological and tissue samples, suggesting that the Onclarity method could also be used to detect HPV in tissue samples and that the HPV genotype detected in FFPE samples is one of the HPV detected in cytological samples, supporting the thesis that one lesion is caused by one HPV genotype.

Human Papillomavirus Same Genotype Persistence and Risk: A Systematic Review.

OBJECTIVE: The aim of the study was to examine whether high-grade cervical intraepithelial neoplasia (CIN) was more closely associated with human papillomavirus (HPV) same-genotype persistence (SGTP) versus clearance of prior infection with a subsequent infection by a new genotype (genotype switch [GS]), clearance of HPV infection, or acquisition of a new HPV infection after a negative infection status, during a follow-up testing subsequent to abnormal screening results. MATERIALS AND METHODS: MEDLINE, Cochrane Library, Health Technology Assessment, and clinicaltrials.gov were searched from January 2000 to July 2019 for prospective controlled trials and observational studies of women and retrospective studies using HPV assays with extended- or full-genotype reporting. The primary outcome was high-grade CIN after at least 2 rounds of testing. Overall quality of evidence for the risk estimate outcomes was assessed. Of the 830 identified abstracts, 66 full-text articles were reviewed, and 7 studies were included in the synthesis. The study protocol was registered with the PROSPERO International Prospective Register of Systematic Reviews (CRD42018091093). RESULTS: Continued HPV-positive women falls in 2 equally large groups: SGTP and GS. Sensitivity, positive predictive value, and positive likelihood ratio of SGTP were significantly higher than for GS. Human papillomavirus genotypes may be ranked into 3 tiers (immediate colposcopy, follow-up testing, return to routine screening), according to associated risk of persistence for high-grade CIN and to prevailing clinical action thresholds. CONCLUSIONS: There is moderately high-quality evidence to support the clinical utility of SGTP to improve risk discrimination for high-grade CIN compared with qualitative HPV testing without genotype-specific information.

Clinical performance of Xpert Bladder Cancer (BC) Monitor, a mRNA-based urine test, in active surveillance (AS) patients with recurrent non-muscle-invasive bladder cancer (NMIBC): results from the Bladder Cancer Italian Active Surveillance (BIAS) project.

PURPOSE: To investigate the clinical performance of a new mRNA-based urine test, aiming to avoid unnecessary follow-up cystoscopy in patients under active surveillance (AS) for recurrent NMIBC. METHODS: This is a prospective cohort study enrolling patients with history of low-grade (LG) NMIBC, who developed a recurrence during the follow-up and underwent AS. Their urinary samples were analyzed by Xpert BC Monitor (Cepheid, Sunnyvale, CA, USA). The primary endpoint was to investigate if Xpert BC Monitor could avoid unnecessary cystoscopy during the follow-up period. Its sensitivity, specificity, PPVs and NPVs were calculated. A cutoff of 0.4 "linear discriminant analysis" (LDA) was optimized for the AS setting. RESULTS: The cohort consisted of 106 patients with a mean age of 72 ± 9.52 and a median follow-up from AS start of 8.8 (range 0-56.5) months. No statistically significant difference was found for the mean age, smoker status, lesion size, and number of lesions with a cutoff of 0.4. Of 106 patients, 22 (20.8%) were deemed to require treatment because of cystoscopic changes in size and/or number of lesions during the follow-up period. Using a cutoff value of < 0.4, 34 (33.7%) cystoscopies could be avoided due to low LDA value, missing 2/22 (9%) failures, none with high-grade (HG) NMIBC. Further research on larger population remains mandatory before its clinical use. CONCLUSION: Xpert BC Monitor seems to be a reliable assay, which might avoid unnecessary cystoscopies without missing HG NMIBC when its cutoff is optimized for the AS setting.

Association between cardiac troponin I and mortality in patients with COVID-19.

BACKGROUND: Severe pneumonia is pathological manifestation of Coronavirus Disease 2019 (COVID-19), however complications have been reported in COVID-19 patients with a worst prognosis. Aim of this study was to evaluate the role of high sensitivity cardiac troponin I (hs-TnI) in patients with SARS-CoV-2 infection. METHODS: we retrospectively analysed hs-TnI values measured in 523 patients (median age 64 years, 68% men) admitted to a university hospital in Milan, Italy, and diagnosed COVID-19. RESULTS: A significant difference in hs-TnI concentrations was found between deceased patients (98 patients) vs discharged (425 patients) [36.05 ng/L IQR 16.5-94.9 vs 6.3 ng/L IQR 2.6-13.9, p < 0.001 respectively]. Hs-TnI measurements were independent predictors of mortality at multivariate analysis adjusted for confounding parameters such as age (HR 1.004 for each 10 point of troponin, 95% CI 1.002-1.006, p < 0.001). The survival rate, after one week, in patients with hs-TnI values under 6 ng/L was 97.94%, between 6 ng/L and the normal value was 90.87%, between the normal value and 40 ng/L was 86.98, and 59.27% over 40 ng/L. CONCLUSION: Increase of hs-TnI associated with elevated mortality in patients with COVID-19. Troponin shows to be a useful biomarker of disease progression and worse prognosis in COVID-19 patients.

Clinical Utility of Human Papillomavirus Genotyping in Cervical Cancer Screening: A Systematic Review.

OBJECTIVE: Thirteen human papillomavirus (HPV) genotypes are associated with the highest risk of cervical disease/cancer; however, the risk of disease progression and cancer is genotype dependent. The objective of this systematic review was to examine evidence for high-grade cervical intraepithelial neoplasia (≥CIN 3) risk discrimination using HPV genotyping. MATERIALS AND METHODS: A systematic review of English and non-English articles through MEDLINE, Cochrane, clinicaltrials.gov, and abstracts presented at relevant professional society conferences were searched from 2000 to 2019. Search terms included: cervical cancer screening, HPV genotyping, CIN, HPV persistence, humans, and colposcopy; prospective, controlled trials, observational studies, and retrospective studies of residual specimens; evidence included HPV genotyping (beyond genotypes 16/18/45) results. Data were obtained independently by authors using predefined fields. Risk of bias was evaluated with a modified Newcastle-Ottawa Scale. The Grading of Recommendations, Assessment, Development and Evaluation methodology facilitated overall quality of evidence evaluation for risk estimation. The study protocol was registered with the PROSPERO International Prospective Register of Systematic Reviews (CRD42018091093). The primary outcome was CIN 3 or worse risk both at baseline and at different follow-up periods. RESULTS: Of 236 identified sources, 60 full texts were retrieved and 16 articles/sources were included. Risk of bias was deemed low; the overall quality of evidence for CIN 3 or worse risk with negative for intraepithelial lesions or malignancies or low-grade squamous intraepithelial cytology was assessed as moderate; that with atypical squamous cells-undetermined significance and "all cytology" was assessed as high. Clinical and methodological heterogeneity precluded meta-analysis. Human papillomavirus genotyping discriminated risk of CIN 3 or worse to a clinically significant degree, regardless of cytology result. CONCLUSIONS: The evidence supports a clinical utility for HPV genotyping in risk discrimination during cervical cancer screening.

Chest CT in patients with a moderate or high pretest probability of COVID-19 and negative swab.

OBJECTIVES: We aimed to assess the diagnostic performance of CT in patients with a negative first RT-PCR testing and to identify typical features of COVID-19 pneumonia that can guide diagnosis in this case. METHODS: Patients suspected of COVID-19 with a negative first RT-PCR testing were retrospectively revalued after undergoing CT. CT was reviewed by two radiologists and classified as suspected COVID-19 pneumonia, non-COVID-19 pneumonia or negative. The performance of both first RT-PCR result and CT was evaluated by using sensitivity (SE), specificity (SP), positive predictive value (PPV), negative predictive value (NPV) and area under the curve (AUC) and by using the second RT-PCR test as the reference standard. CT findings for confirmed COVID-19 positive or negative were compared by using the Pearson chi-squared test (P values < 0.05) RESULTS: Totally, 337 patients suspected of COVID-19 underwent CT and nasopharyngeal swabs in March 2020. Eighty-seven out of 337 patients had a negative first RT-PCR result; of these, 68 repeated RT-PCR testing and were included in the study. The first RT-PCR test showed SE 0, SP = 100%, PPV = NaN, NPV = 70%, AUC = 50%, and CT showed SE = 70% SP = 79%, PPV = 86%, NPV = 76%, AUC = 75%. The most relevant CT variables were ground glass opacity more than 50% and peripheral and/or perihilar distribution. DISCUSSION: Negative RT-PCR test but positive CT features should be highly suggestive of COVID-19 in a cluster or community transmission scenarios, and the second RT-PCR test should be promptly requested to confirm the final diagnosis.

Neutrophil, Platelets, and Eosinophil to Lymphocyte Ratios Predict Gleason Score Upgrading in Low-Risk Prostate Cancer Patients.

BACKGROUND: Several biochemical and clinical markers have been proposed for selecting patients for active surveillance (AS). However, some of these are expensive and not easily accessible. Moreover, currently about 30% of patients on AS harbor aggressive disease. Hence, there is an urgent need for other tools to accurately identify patients with low-risk prostate cancer (PCa). PATIENTS: We retrospectively reviewed the medical records of 260 patients who underwent radical prostatectomy and were eligible for AS according to the following criteria: clinical stage T2a or less, prostate-specific antigen level < 10 ng/mL, 2 or fewer cores involved with cancer, Gleason score (GS) ≤6 grade, and prostate-specific antigen density < 0.2 ng/mL/cc. METHODS: Univariate and multivariate analyses were performed to evaluate the association of patient and tumor characteristics with reclassification, defined as upstaged (pathological stage >pT2) and upgraded (GS ≥7) disease. A base model (age, prostate-specific antigen, prostate volume, and clinical stage) was compared with models considering neutrophil to lymphocyte ratio (NLR) or platelets to lymphocyte ratio (PLR), monocyte to lymphocyte (MLR), and eosinophil to lymphocyte ratio (ELR). OR and 95% CI were calculated. Finally, a decision curve analysis was performed. RESULTS: Univariate and multivariate analyses showed that NLR, PLR, and ELR upgrading were significantly associated with upgrading (ORs ranging from 2.13 to 4.13), but not with upstaging except for MLR in multivariate analysis, showing a protective effect. CONCLUSION: Our results showed that NLR, PLR, and ELR are predictors of Gleason upgrading. Therefore, these inexpensive and easily available tests might be useful in the assessment of low-risk PCa, when considering patients for AS.

Distribution of High-Risk Human Papillomavirus Genotypes and Multiple Infections in Preneoplastic and Neoplastic Cervical Lesions of Unvaccinated Women: A Cross-sectional Study.

OBJECTIVE: The aim of the study was to investigate the distribution of high-risk (HR) human papillomavirus (HPV) genotypes and the role of multiple infection in preneoplastic and neoplastic cervical lesions, according to histology, age, and the number of genotypes per infection. MATERIALS AND METHODS: Nine hundred eighty-eight women affected by known HPV-related cervical lesions and attending the European Institute of Oncology, Milan, Italy, from December 2006 to December 2014, were selected for a cross-sectional study. Prevalence of HPV genotypes was calculated by histology and the number of genotypes per infection. Univariate and multivariable cervical intraepithelial neoplasia (CIN) 2-3 versus CIN 1 risks were estimated by logistic regression models. RESULTS: Overall, HPV 16 (53.1%), HPV 31 (15.1%), and HPV 58 (6.4%) were the most frequent genotypes in precancerous lesions. At multivariable analysis, HPV 16 (p = .02), 18 (p = .013), and 56 (p = .01) were significantly associated to worsen histology, whereas HPV 39 (p = .03) and 45 (p = .03) were statistically correlated only to the increasing number of genotypes per infections. Human papillomavirus 33 was the only genotype significantly related to both the number of genotypes per infection (p = .005) and age (p = .03). Infections by HR-HPV (odds ratio [OR] = 9.48, 95% CI = 3.77-23.8, p < .001), HPV genotypes covered by current vaccines (OR = 6.28, 95% CI = 4.05-9.75, p < .001), single HPV genotype (OR = 8.13, 95% CI = 4.12-16.0, p < .001), as well as age (OR = 1.13, 95% CI = 1.07-1.19, p < .001) were significantly associated to higher risk of CIN 2-3. CONCLUSIONS: The most of CIN 2+ lesions are sustained by HR-HPV genotypes, especially the ones covered by 9-valent vaccine; therefore, the widespread use of prophylactic HPV vaccines could significantly reduce the incidence of preneoplastic and neoplastic cervical lesions.

Onclarity Human Papillomavirus Extended Genotyping in the Management of Cervical Intraepithelial Neoplasia 2+ Lesions.

OBJECTIVE: Many methods are available today for human papillomavirus (HPV) testing; they differ for technology, targets, and information on the genotypes detected. In this study, we evaluated the performance of the Onclarity HPV assay in detection and follow-up of cervical preneoplastic lesions. MATERIALS AND METHODS: One hundred sixty-seven women referred to the European Institute of Oncology, Milan, for treatment of cervical lesions were enrolled. We investigated the utility of Onclarity extended genotyping HPV test in the management of cervical intraepithelial neoplasia (CIN) 2+ preneoplastic lesion. RESULTS: At baseline, the concordance was 92% (150/163) between Onclarity and Hybrid Capture 2 (HC2) and 93% (142/152) between Onclarity and linear array, respectively. At follow-up, the concordance between Onclarity and HC2 was 80%. Seven women relapsed: 6 had persistence of the same genotypes and 1 patient tested negative not only with Onclarity but also with HC2 for the presence of a low-risk genotype in the sample. CONCLUSIONS: This study showed that the evaluation of the HPV genotype persistence may represent a valid option to monitor patients treated for CIN 2+ lesions, because relapses were detected only in patients with persistence of the same genotype detected at baseline.

Diagnostic and Prognostic Utility of Circulating Cytochrome c in Acute Myocardial Infarction.

RATIONALE: In contrast to cardiomyocyte necrosis, which can be quantified by cardiac troponin, functional cardiomyocyte impairment, including mitochondrial dysfunction, has escaped clinical recognition in acute myocardial infarction (AMI) patients. OBJECTIVE: To investigate the diagnostic accuracy for AMI and prognostic prediction of in-hospital mortality of cytochrome c. METHODS AND RESULTS: We prospectively assessed cytochrome c serum levels at hospital presentation in 2 cohorts: a diagnostic cohort of patients presenting with suspected AMI and a prognostic cohort of definite AMI patients. Diagnostic accuracy for AMI was the primary diagnostic end point, and prognostic prediction of in-hospital mortality was the primary prognostic end point. Serum cytochrome c had no diagnostic utility for AMI (area under the receiver-operating characteristics curve 0.51; 95% confidence intervals 0.44-0.58; P=0.76). Among 753 AMI patients in the prognostic cohort, cytochrome c was detectable in 280 (37%) patients. These patients had higher in-hospital mortality than patients with nondetectable cytochrome c (6% versus 1%; P<0.001). This result was mainly driven by the high mortality rate observed in ST-segment-elevation AMI patients with detectable cytochrome c, as compared with those with nondetectable cytochrome c (11% versus 1%; P<0.001). At multivariable analysis, cytochrome c remained a significant independent predictor of in-hospital mortality (odds ratio 3.0; 95% confidence interval 1.9-5.7; P<0.001), even after adjustment for major clinical confounders (odds ratio 4.01; 95% confidence interval 1.20-13.38; P=0.02). CONCLUSIONS: Cytochrome c serum concentrations do not have diagnostic but substantial prognostic utility in AMI.

Prevalence and Risk Factors of Human Papillomavirus Infection in 18-Year-Old Women: Baseline Report of a Prospective Study on Human Papillomavirus Vaccine.

OBJECTIVES: Little is known about the epidemiology of human papillomavirus (HPV) in Italy before the age of 25. At the European Institute of Oncology, a prospective observational study on cervical HPV infection in 18-year-old women undergoing quadrivalent HPV vaccination is ongoing. METHODS: At the first visit before vaccination, all the young women answered an epidemiological questionnaire, and then, the presence of high-risk HPV (hrHPV) was tested. Samples positive for hrHPV were genotyped. Liquid-based cytology was done only to women declaring not to be virgins. Any positivity at cytology or HPV testing was completed with colposcopy and eventually biopsies. RESULTS: Seven hundred and thirty women were enrolled. Two hundred sixty-six women were virgins; 7 (2.6%) of these resulted positive to hrHPV: 1 had HPV16 and CP6108, whereas the other 6 resulted negative at genotyping. Of the 464 nonvirgins, 61 (13.1%) were HPV positive: 19 had HPV16, 4 were positive to HPV18 with other hrHPVs, 25 to other hrHPVs, 7 to low-risk HPV, whereas 13 resulted negative at genotyping. HPV positivity was significantly associated to both smoking and having more than 3 partners. Cervical cytology was negative in 433 cases (93.3%), ASC-US in 10 cases (2.2%), low-grade squamous intraepithelial lesion in 20 cases (4.3%), and ASC-H in 1 case (0.2%). No CIN2+ was identified. CONCLUSIONS: Overall, we found a low positivity to HPV in this population; however, the rate of HPV positivity was significantly related to smoking and sexual life. The cytology result low-grade squamous intraepithelial lesion was more frequent than in the screening population, whereas no CIN2+ was identified, confirming the indication to avoid screening at this age.

The BD Onclarity HPV Assay on Samples Collected in SurePath Medium Meets the International Guidelines for Human Papillomavirus Test Requirements for Cervical Screening.

This study describes a validation of the BD Onclarity HPV (Onclarity) assay using the international guidelines for HPV test requirements for cervical cancer screening of women 30 years old and older using Danish SurePath screening samples. The clinical specificity (0.90, 95% confidence interval [CI] = 0.88 to 0.91) and sensitivity (0.97, 95% CI = 0.87 to 1.0) of the Onclarity assay were shown to be not inferior to the reference assay (specificity, 0.90 [95% CI = 0.88 to 0.92]; sensitivity, 0.98 [95% CI = 0.91 to 1.0]). The intralaboratory reproducibility of Onclarity was 97%, with a lower confidence bound of 96% (kappa value, 0.93). The interlaboratory agreement was 97%, with a lower confidence bound of 95% (kappa value, 0.92). The BD Onclarity HPV assay fulfills all the international guidelines for a new HPV test to be used in primarily screening. This is the first clinical validation of a new HPV assay using SurePath screening samples, and thus the Onclarity HPV assay is the first HPV assay to hold an international validation for both SurePath and ThinPrep.

Optimization and Standardization of Circulating MicroRNA Detection for Clinical Application: The miR-Test Case.

BACKGROUND: The identification of circulating microRNAs (miRNAs) in the blood has been recently exploited for the development of minimally invasive tests for the early detection of cancer. Nevertheless, the clinical transferability of such tests is uncertain due to still-insufficient standardization and optimization of methods to detect circulating miRNAs in the clinical setting. METHODS: We performed a series of tests to optimize the quantification of serum miRNAs that compose the miR-Test, a signature for lung cancer early detection, and systematically analyzed variables that could affect the performance of the test. We took advantage of a large-scale (>1000 samples) validation study of the miR-Test that we recently published, to evaluate, in clinical samples, the effects of analytical and preanalytical variables on the quantification of circulating miRNAs and the clinical output of the signature (risk score). RESULTS: We developed a streamlined and standardized pipeline for the processing of clinical serum samples that allows the isolation and analysis of circulating miRNAs by quantitative reverse-transcription PCR, with a throughput compatible with screening trials. The major source of analytical variation came from RNA isolation from serum, which could be corrected by use of external (spike-in) or endogenous miRNAs as a reference for normalization. We also introduced standard operating procedures and QC steps to check for unspecific fluctuations that arise from the lack of standardized criteria in the collection or handling of the samples (preanalytical factors). CONCLUSIONS: We propose our methodology as a reference for the development of clinical-grade blood tests on the basis of miRNA detection.

Serum HER2 extracellular domain levels and HER2 circulating tumor cell status in patients with metastatic breast cancer.

AIM: To shed light on the clinical role of HER2 status in serum as extracellular domain (ECD) and corresponding circulating tumor cells (CTCs) in metastatic breast cancer patients. METHODS: 68 patients were analyzed. Serum HER2 was determined by ADVIA Centaur(®) Serum HER2 test. CellSearch System was performed for CTC quantification. RESULTS: HER2 was overexpressed in 21 primary tumors. In total, 19 patients had ECD >15 ng/ml (the cut-off used), 48 patients had at least one CTC. ECD positivity was associated with CTC number (p = 0.01), HER2-positive CTC (p = 0.01) and the ratio HER2-positive CTC/total CTC (p = 0.02). ECD was not associated with survival. CONCLUSION: ECD in combination with HER2 CTC status would deserve further investigation in larger series for addressing its putative prognostic relevance.

Chlamydia trachomatis infection and HPV/Chlamydia trachomatis co-infection among HPV-vaccinated young women at the beginning of their sexual activity.

PURPOSE: This study investigated the prevalence of Chlamydia trachomatis infection, co-infection with Human Papillomavirus (HPV) and associated risk factors in a cohort of sexually active young women enrolled in an ongoing trial on HPV vaccination at the European Institute of Oncology (IEO, Milan, Italy). METHODS: Cervical samples were collected from 591 girls (median age 18.8 years) at the beginning of their sexual activity. At the time of sample collection, 354 women had not yet been vaccinated, and 237 women had been vaccinated for at least 12 months. All samples were analyzed through a molecular assay for the detection of C. trachomatis infection. Demographic, behavioral risk factors and high-risk HPV (HR-HPV) status were investigated. RESULTS: The prevalence of C. trachomatis infection was 4.9 % and HPV/C. trachomatis co-infection rate was 1.5 %. The exact analysis has not underlined statistical significance for the variables considered, except for the infection with HR-HPV (p < 0.001). The prevalence of C. trachomatis infection among women who had not been immunized and those already vaccinated was similar (5.6 vs 3.8 %). However, the rate of HPV/C. trachomatis co-infection was twice as high in unvaccinated women (2 %) compared to vaccinated women (0.8 %). CONCLUSIONS: Over 16 % of young women had at least one of the two STIs investigated. The risk of C. trachomatis infection was higher in HR-HPV infected compared to HR-HPV uninfected young women. The rate of co-infection was halved in HPV-vaccinated compared to unvaccinated women. This study underlines that HPV vaccination can confer benefits also in terms of co-infections prevention, leading to a decreased risk of developing cervical malignancies.

Portal vein-circulating tumor cells predict liver metastases in patients with resectable pancreatic cancer.

Pancreatic cancer patients underwent surgical resection often present distant metastases early after surgery. Detection of circulating tumor cells (CTCs) has been correlated to a worse oncological outcome in patients with advanced pancreatic cancer. The objective of this pilot study is to investigate the possible prognostic role of CTCs in patients undergoing surgery for pancreatic cancer. In 20 patients undergoing pancreatic resection, 10 mL blood sample was collected intraoperatively from both systemic circulation (SC) and portal vein (PV). Blood sample was analyzed for CTCs with CellSearch® system. All patients underwent an oncologic follow-up for at least 3 years, quarterly. CTCs were detected in nine (45%) patients: five patients had CTCs in PV only, three patients in both SC and PV, and one patient in SC only. CTC-positive and CTC-negative patients were similar for demographics and cancer stage pattern. No significant differences were found in both overall and disease-free survival between CTC-positive and CTC-negative patients. At 3-year follow-up, portal vein CTC-positive patients presented a higher rate of liver metastases than CTC-negative patients (53 vs. 8%, p = 0.038). CTCs were found in 45% of the patients. No correlation between CTCs and survival was found. The presence of CTCs in portal vein has been associated to higher rate of liver metastases after surgery.