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As the primary screening test, E6/E7 mRNA has shown similar sensitivity for CIN3+ and lower positivity rate than the HPV DNA test. Nevertheless, the overall mRNA positivity is too high for immediate colposcopy, making a triage test necessary. The aim was to estimate the mRNA performance as a primary test with different triage strategies. All HPV DNA-positives were tested for mRNA, cytology and p16/ki67. A sample of HPV DNA-negatives was also tested for mRNA to estimate test specificity. We included all CIN3+ histologically diagnosed within 24 months since recruitment. Of the 41 127 participants, 7.7% were HPV DNA-positive, of which 66.4% were mRNA-positive. Among the HPV DNA-negatives, 10/1108 (0.9%) were mRNA-positive. Overall, 97 CIN3+ were found. If mRNA was used as the primary test, it would miss about 3% of all CIN3+ with a 22% reduction of positivity compared with HPV DNA. The weighted specificity estimate forCIN2 was 94.5% (95% CI = 93.9%-94.9%) and sensitivity for CIN3+ was 96.9% (95% CI = 91.3%-99.1%). If all the weighted estimated 6.0% mRNA-positive women had been referred to colposcopy, PPV for CIN3+ would have been 4.2%. Cytology or p16/ki67 triage would decrease immediate referral to 1.7% and 2.0%, increasing PPV to 11.2% and 11.7%, respectively; total colposcopy referral would be 4.0% and 3.9%, respectively. As the primary screening test, the mRNA assay showed a positivity rate lower than that of HPV DNA, with a small number of CIN3+ missed. Triage with cytology or p16/ki67 would only marginally decrease overall colposcopy referral.

Background p16/Ki-67 dual staining is a candidate biomarker for triaging human papillomavirus (HPV)-positive women. Reproducibility is needed for adopting a test for screening. This study assessed interlaboratory reproducibility in HPV-positive women. Methods All women positive for HPV from the Italian New Technologies for Cervical Cancer 2 study, were included in this study. ThinPrep slides were immunostained for p16/Ki-67 in 4 laboratories and were interpreted in 7 laboratories. Each slide had 3 reports from different laboratories. Slides were classified as valuable or inadequate, and valuable slides were classified as positive (at least 1 double-stained cell) or negative. Interlaboratory reproducibility was evaluated with κ values. Results Overall, we obtained 9300 reports for 3100 cases; 905 reports (9.7%) were inadequate. The overall adequacy concordance was poor (κ = 0.224; 95% confidence interval [CI], 0.183-0.263). The overall positivity concordance was moderate (κ = 0.583; 95% CI, 0.556-0.610). Of the 176 cervical intraepithelial neoplasia 2+ (CIN-2+) lesions found in HPV DNA-positive women, 158 had a valid result: 107 were positive in all 3 reports (sensitivity for CIN-2+, 67.7%; 95% CI, 59.8%-74.9%), 23 were positive in 2 reports (sensitivity of the majority report, 82.3%; 95% CI, 75.4%-87.9%), and 15 were positive in 1 report (sensitivity of at least 1 positive result, 91.8%; 95% CI, 86.3%-95.5%). Thirteen CIN-2+ cases were negative in all 3 reports. The overall positivity concordance in CIN-2+ samples was κ = 0.487 (95% CI, 0.429-0.534), whereas in the non-CIN-2+ samples, it was κ = 0.558 (95% CI, 0.528-0.588). Conclusions The p16/Ki-67 assay showed poor reproducibility for adequacy and good reproducibility for positivity comparable to that of cervical cytology. Nevertheless, the low reproducibility does not affect the sensitivity for CIN-2+.

Objective To evaluate performance of the first round of HPV-based screening in Tuscany region and compare it with the prior round of Pap-based screening Setting Tuscany region of Italy, where HPV-based cervical cancer screening started in 2013, with a strong level of centralization screening tests at the Regional Laboratory for Cancer Prevention (ISPRO). Methods The transition from Pap- to HPV-based screening was initiated for older women and at 3 out of 12 Tuscany Local Health Units (LHUs). Data from the Florence and Grosseto LHUs (about 300,000 women) were analysed and performance screening indicators estimated. Results HPV-based indicators recorded good performance, with increased compliance vs. the Pap-based programme. We registered a substantial decrease in waiting times from sampling to test reporting, probably related to the centralization strategy. Since the screening protocol was the same and conducted at a single laboratory, we could hypothesize that the difference in HPV positivity (6.8% in Florence vs. 8.4% in Grosseto) was due to a real difference in HPV prevalence among women of the two LHUs. The transition to HPV-based screening led to a significant increase both in colposcopy referral rate (4.3% vs. 1.2%) and CIN2+ detection rate (8.3‰ vs. 3.4‰). Conclusions HPV-based is more effective in detecting high-grade precancerous and cancerous lesions than Pap-based screening and is characterized by an “anticipatory effect” in the detection of CIN2+ lesions. The transition from Pap-based to HPV-based screening programme should include increased resources dedicated to colposcopy services. Centralization in a laboratory with long experience in this field promotes efficiency of the screening process.

Objective To present performance indicators from the second round of the Vallecamonica-Sebino HPV screening programme in women who had tested negative about four years earlier (mean 45 months). Methods From 2010 to 2012, the target female population (aged 25–64) was invited to the first HPV screening round. In 2013–2017, women were rescreened for the second round. HPV-negative women at the first round were initially rescreened after three years. The interval was gradually increased to five years. HPV-positive women underwent cytology triage: positives were referred to colposcopy and negatives to repeat testing after one year. If HPV was persistently positive, women were referred to colposcopy, if negative, to normal interval rescreening. Results In the second round, of 13,824 previously HPV-negative women, 598 were HPV-positive (4.3%), of whom 297 were positive at cytology triage. Of those referred to one-year HPV test, 291 complied (98.0%), 133 (50.2%) of whom were persistently positive. Total referral was 3.1% compared with 6.6% in the first round (age-adjusted relative referral 0.59, 95% CI: 0.53–0.65). There were 24 cervical intraepithelial neoplasia 2+ (three cervical intraepithelial neoplasia 3+). Detection was 0.17%, compared with 0.9% in the first round. Age-adjusted relative detections were 0.25 (95% CI: 0.16–0.39) and 0.18 (95% CI: 0.05–0.61) for cervical intraepithelial neoplasia 2+ and cervical intraepithelial neoplasia 3+, respectively. Positive predictive value was 5.7%, compared with 14.6% in the first round. Conclusions At second round, referral was half that at first round, while cervical intraepithelial neoplasia 2+ detection decreased nine-fold. Consequently, positive predictive value decreased dramatically. Rescreening four years after an HPV-negative test makes the process inefficient due to the low prevalence of lesions.

Background A large free-of-charge quadrivalent HPV (qHPV) vaccination program, covering four cohorts annually (women 11, 14, 17 and 24 years), has been implemented in Basilicata since 2007. This study evaluated vaccine and non-vaccine HPV prevalence 5-7 years post-vaccination program implementation in vaccinated and unvaccinated women. Methods This population-based, cross-sectional study was conducted in the public screening centers of the Local Health Unit in Matera between 2012 and 2014. Cervical samples were obtained for Pap and HPV testing (HC2, LiPA Extra® assay) and participants completed a sociodemographic and behavioral questionnaire. Detailed HPV vaccination status was retrieved from the official HPV vaccine registry. HPV prevalence was described overall, by type and vaccination status. The association between HPV type-detection and risk/protective factors was studied. Direct vaccine protection (qHPV vaccine effectiveness [VE]), cross-protection, and type-replacement were evaluated in cohorts eligible for vaccination, by analyzing HPV prevalence of vaccine and non-vaccine types according to vaccination status. Results Overall, 2793 women (18-50 years) were included, 1314 of them having been in birth cohorts eligible for the HPV vaccination program (18- to 30-year-old women at enrolment). Among the latter, qHPV vaccine uptake was 59% (at least one dose), with 94% completing the schedule; standardized qHPV type prevalence was 0.6% in vaccinated versus 5.5% in unvaccinated women (P

In Italy, the cohorts of women who were offered Human papillomavirus (HPV) vaccination in 2007/08 will reach the age (25years) for cervical cancer (CC) screening from 2017. The simultaneous shift from cytology-based screening to HPV test-based screening gives the opportunity for unprecedented reorganisation of CC prevention. The ONS (National Screening Monitoring Centre) Directive and the GISCi (Italian Group for Cervical Screening) identified the consensus conference as the most suitable method for addressing this topic. A summary of consensus recommendations is reported here. The main objective was to define the best screening methods in girls vaccinated against HPV and the knowledge required for defining evidence-based screening strategies. A Jury made recommendations about questions and proposals formulated by a panel of experts representative of Italian scientific societies involved in CC prevention and based on systematic reviews of literature and evidence. The Jury considered changing the screening protocols for girls vaccinated in their twelfth year as appropriate. Tailored screening protocols based on vaccination status could be replaced by "one size fits all" protocols only when a herd immunity effect has been reached. Vaccinated women should start screening at age 30, instead of 25, with HPV test. Furthermore, there is a strong rationale for applying longer intervals for re-screening HPV negative women than the currently recommended 5years, but research is needed to determine the optimal screening time points. For non-vaccinated women and for women vaccinated in their fifteenth year or later, the current protocol should be kept.

BACKGROUND The accumulation of cyclin-dependent kinase inhibitor 2A (p16ink4a) protein in a cell is associated with neoplastic progression in precancerous cervical lesions. Dual staining for p16ink4a and Ki-67 has been proposed as a triage test in cervical cancer screening for women who test positive for human papillomavirus DNA. In this study, interobserver reproducibility of the interpretation of this test was assessed. METHODS Forty-two immunostained, liquid-based cytology slides were divided into 2 sets and were interpreted by 17 to 21 readers from 9 different laboratories, yielding a total of 816 reports. Immunostaining results were classified as positive, negative, inconclusive, or inadequate. After evaluation of the first set of slides and before circulation of the second set, the results were discussed in a plenary meeting. The 10 slides with the most discordant results were evaluated again by selected expert cytopathologists. RESULTS The overall κ value was 0.612 (95% confidence interval [CI], 0.523-0.701), it was higher for the positive and negative categories (κ = 0.692 and κ = 0.641, respectively), and it was almost null for the inconclusive category (κ = 0.058). Considering only readers from laboratories with documented experience, the κ value was higher (κ = 0.747; 95% CI, 0.643-0.839) compared with nonexperienced centers (κ = 0.498; 95% CI, 0.388-0.616). The results were similar in both sets of slides (κ = 0.505 [95% CI, 0.358-0.642] and κ = 0.521 [95% CI, 0.240-0.698] for the first and second sets, respectively). Reinterpretation of the slides with the most discordant results did not provide any improvement (first evaluation, κ = 0.616 [95% CI, 0.384-0.866]; second evaluation, κ = 0.403 [95% CI, 0.182-0.643]). CONCLUSIONS Dual staining for p16 ink4a and Ki-67 demonstrated good reproducibility, confirming its robustness, which is a necessary prerequisite for its adoption as a triage test in cervical cancer screening programs that use human papillomavirus DNA as a primary test. Cancer Cytopathol 2017;125:212–220. © 2016 American Cancer Society.

Background The study presents cross-sectional accuracy of E6 and E7 (E6/E7) mRNA detection and p16/ki67 dual staining, alone or in combination with cytology and human papillomavirus (HPV)16/18 genotyping, as a triage test in HPV DNA-positive women and their impact on cervical intraepithelial neoplasia (CIN2+) overdiagnosis. Methods Women aged 25-64 years were recruited. HPV DNA-positive women were triaged with cytology and tested for E6/E7 mRNA and p16/ki67. Cytology positive women were referred to colposcopy, and negatives were randomly assigned to immediate colposcopy or to 1-year HPV retesting. Lesions found within 24 months since recruitment were included. All P values were 2-sided. Results 40 509 women were recruited, and 3147 (7.8%) tested HPV DNA positive; 174 CIN2+ were found: sensitivity was 61.0% (95% confidence interval [CI] = 53.6 to 68.0), 94.4% (95% CI = 89.1 to 97.3), and 75.2% (95% CI = 68.1 to 81.6) for cytology, E6/E7 mRNA, and p16/ki67, respectively. Immediate referral was 25.6%, 66.8%, and 28.3%, respectively. Overall referral was 65.3%, 78.3%, and 63.3%, respectively. Cytology or p16/ki67, when combined with HPV16/18 typing, reached higher sensitivity with a small impact on referral. Among the 2306 HPV DNA-positive and cytology-negative women, relative CIN2+ detection in those randomly assigned at 1-year retesting vs immediate colposcopy suggests a -28% CIN2+ regression (95% CI = -57% to +20%); regression was higher in E6/E7 mRNA-negatives (Pinteraction = .29). HPV clearance at 1 year in E6/E7 mRNA and in p16/ki67 negative women was about 2 times higher than in positive women (Pinteraction < .001 for both). Conclusions p16/ki67 showed good performance as a triage test. E6/E7 mRNA showed the highest sensitivity, at the price of too high a positivity rate to be efficient for triage. However, when negative, it showed a good prognostic value for clearance and CIN2+ regression.

Background and objectives A randomized trial was conducted in Tuscany, Italy, to evaluate the effectiveness of HPV vaccination for 25 year old (yo) women who attend at the first time cervical cancer screening. The trial also evaluated immune response after vaccination, reductions of cytological abnormalities and the impact of vaccination on screening activity. Study design During 2010–2011, all 25 yo women who were invited to the Florence cervical cancer screening programme were also asked to participate in the trial. Enrolled women were randomized into study and control groups. Those in the study group were offered HPV vaccination after the usual Pap test. The cytology distribution and prevalence for any high risk (hr) HPV type were compared at the subsequent screening round in an intention-to-treat analysis. The impact of HPV vaccination was evaluated per protocol comparing vaccinated women with the control group. Results Our results showed a reduction in HPV prevalence at recall for any hr-HPV type but it was not statistically significant, being 17.1% vs 21.4%, p = 0.20 in the study and control groups, respectively. If we restricted the analysis to vaccinated women, strong reductions of the HPV 16,18,31,33,45 and HPV 31,33,45 infections were observed, being 5.3% vs 12.8%, p < 0.01 and 2.1% vs 6.5%, p = 0.02, respectively. Significant reductions for any hr-HPV infection and for HPV 16 infection were also observed in women HPV 16/18 negative at enrolment, being 12% vs 21.4%, p < 0.01 and 0.6% vs 6.7%, p-value < 0.01, respectively. In women hr-HPV negative at enrolment no infections due to HPV 16 or HPV 18 were observed and there was a big reduction for any hr-HPV infection (7.1% vs 21.4% p < 0.01). A strong antibody response was observed not only for HPV 16 & 18 but also for their related types. Conclusions Our findings suggest that HPV vaccination at the age 25 is beneficial if it is offered to hr-HPV negative women. Our data will assist in developing a cost effectiveness model for choosing the best strategy to integrate screening and vaccination for the coming years. Clinical trial registration number is NCT02296255. (Less)

Background Measuring viral DNA methylation in human papillomavirus (HPV) infected women showed promise for accurate detection of high-grade cervical lesions and cancer. Methylation status has been widely investigated for HPV16, sporadically for other HPV types. Methods Objective of this methodological study was to set up molecular methods to test the methylation levels in the twelve oncogenic HPV types by pyrosequencing, minimizing the number of HPV type-specific PCR protocols. Target CpGs were selected on the HPV L1 (two regions, L1 I and L1 II) and L2 genes. Study samples included DNA stored at Turin, Italy, purified by cervical cells collected in Standard Transport Medium or PreservCyt from women who participated in two studies (N = 126 and 140) nested within the regional organized screening programme. PCR consensus primers were designed by PyroMark Assay Design software to be suitable for amplification of many different oncogenic HPV types. Results Generation of consensus primers was successful for L1 I and II regions, unsuccessful for L2 region, for which HPV type-specific primers remained necessary. The difference between replicated tests on the same sample was ≤4% in 88%, 77% and 91% of cases when targeting the L1 I, L1 II and L2 regions, respectively. The corresponding intra-class correlation coefficients (ICC) were 0.94, 0.87 and 0.97 respectively. When comparing methylation measures based on consensus and type-specific primers, ICC was 0.97 for the L1 I region and 0.99 the for L1 II region. Conclusions The proposed protocols, applying consensus primers suitable to amplify the oncogenic HPV types and minimize the number of PCR reactions, represent a promising tool to quantify viral methylation in women positive for any high risk HPV type. Impact Potential application of these methylation protocols in screening settings can be explored to identify women with high probability of progression to high grade lesions.

Objective We present the results of the first screening round and the first year of the second round of the Valcamonica Human Papillomavirus (HPV) pilot screening project. Setting From 2010 to 2012, the entire target female population (aged 25–64) was invited to the first HPV screening round in an area where Pap test screening had been active since 2002. Methods For HPV-negative women, the interval was three years. For HPV-positive women, a cytological smear was stained and interpreted. Positive cytologies were referred to colposcopy; negatives were referred to repeat HPV after one year. If HPV was persistently positive, women were referred to colposcopy; if negative, to normal screening. Results In 2010–12 18728 women were screened, slightly higher participation than with Pap test (18233 64.7%); 1633 were HPV-positive (8.7%); 843 were positive at cytology triage (referral rate at baseline 4.5%). Of those referred at the one year HPV test, 84% complied (660/780); 356 were persistently positive (1.9%). The total referral rate was 6.4% compared with 3.7% for the Pap test. The detection rate was 9.2/1000 compared with 5.0% for the Pap test. The HPV positivity rate during the second round in women previously negative was 3.9% and the detection rate in HPV-positive cytology-positive women was 0.8/1000. Conclusions HPV-based screening increases colposcopies at the first round, but also strongly increases the detection rate. At the second round, HPV prevalence was much lower and the detection rate also fell, corroborating the need for longer screening intervals in HPV-negative women.

With the introduction of Human papillomavirus (HPV) vaccination in young girls in 2007, it is important to monitor HPV infections and epidemiological changes in this target population. The present study has evaluated the detection of human papillomavirus DNA in paired cervical and urine samples to understand if HPV testing in urine could be used as non-invasive method to monitor HPV status in young women. The study enrolled 216 twenty five-year-old women, resident in Florence and invited for the first time to the cervical cancer Screening Program within a project evaluating the impact of HPV vaccination. HPV genotyping was performed on 216 paired urine and cervical samples. The overall concordance between cervix and urine samples, investigated by HPV genotyping (INNO-LiPA HPV Genotyping Extra), was: 85.6% (184/215), 84.6% (182/215), 80% (172/215) when the same HPV, at least the same HR HPV and all HR HPV, respectively, were detected. HPV type specific concordance in paired urine and cervical samples was observed in 85.8% (175/204) of women with normal cytology and in seven out of nine women with abnormal cytology. Urine seems to be a suitable and reliable biological material for HPV DNA detection as evidenced by the high concordance with HPV detected in cervical samples. These results suggest that urine could be a good noninvasive tool to monitor HPV infection in vaccinated women.

The accumulation of cyclin-dependent kinase inhibitor 2A (p16Forty-two immunostained, liquid-based cytology slides were divided into 2 sets and were interpreted by 17 to 21 readers from 9 different laboratories, yielding a total of 816 reports. Immunostaining results were classified as positive, negative, inconclusive, or inadequate. After evaluation of the first set of slides and before circulation of the second set, the results were discussed in a plenary meeting. The 10 slides with the most discordant results were evaluated again by selected expert cytopathologists.The overall κ value was 0.612 (95% confidence interval [CI], 0.523-0.701), it was higher for the positive and negative categories (κ = 0.692 and κ = 0.641, respectively), and it was almost null for the inconclusive category (κ = 0.058). Considering only readers from laboratories with documented experience, the κ value was higher (κ = 0.747; 95% CI, 0.643-0.839) compared with nonexperienced centers (κ = 0.498; 95% CI, 0.388-0.616). The results were similar in both sets of slides (κ = 0.505 [95% CI, 0.358-0.642] and κ = 0.521 [95% CI, 0.240-0.698] for the first and second sets, respectively). Reinterpretation of the slides with the most discordant results did not provide any improvement (first evaluation, κ = 0.616 [95% CI, 0.384-0.866]; second evaluation, κ = 0.403 [95% CI, 0.182-0.643]).Dual staining for p16

Objective An important internal quality control system used in the Cancer Prevention and Research Institute cytopathology laboratory in Florence is the peer review procedure, based on the review of all abnormal cytological smears which routinely emerge. Peer review is an important training opportunity for all cytologists, especially for those with less experience. This article shows the results of the peer review procedure. Methods Of the 63 754 Papanicolaou (Pap) smears screened in 2011, 1086 were considered to be abnormal [at least atypical squamous cells of undetermined significance (ASC-US+)] on primary screening (selected by a single cytologist) and were subjected to the peer review procedure. The overall performance of the laboratory's cytologists was evaluated using a multiple rater analysis and the comparison of each cytologist with the final diagnosis. Further, the agreement was assessed by means of Cohen's kappa and weighted kappa statistics. Results In general, a moderate/substantial level of agreement between the ten cytologists and the final diagnoses was evident. Kappa values for each reader compared with the final diagnosis ranged from 0.54 to 0.69. The overall kappa value was 0.62 [95% confidence interval (CI), 0.58–0.66] and overall weighted kappa value was 0.76 (95% CI, 0.74–0.79). The category-specific agreement showed the lowest values for atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion (ASC-H). Conclusion In summary, peer review represents an important internal quality control in the evaluation and improvement of inter-observer agreement and of the functioning of the laboratory as a whole. Multi-head microscope sessions may improve particularly the reproducibility of borderline diagnoses and, above all, can be an important training contribution for cytologists.

G. Tinacci, A. Biggeri, A. Pellegrini, M.P. Cariaggi, M.L. Schiboni and M. Confortini The use of digital images to evaluate the interobserver agreement on cervical smear readings in Italian cervical cancer screening Objective: The aim of this study was to measure interobserver agreement among cytologists on using a set of digital images. Methods: A set of 90 selected Papanicolaou-stained cervical smears were digitalized and the digital images circulated among 117 readers, from laboratories spread across almost all Italian regions. Three representative fields of each smear were displayed at 20× and 40× magnification (overall six images for each case). The diagnoses made by the cytologists who provided the images were taken as target diagnoses. Results: The κ values were: very low for the categories atypical squamous cells of undetermined significance (ASC-US), and atypical squamous cells – cannot exclude high-grade squamous intraepithelial lesion (ASC-H); poor for the categories atypical glandular cells (AGC), high-grade squamous intraepithelial lesion (HSIL) and invasive cancer; and fair to good for the categories negative and low-grade squamous intraepithelial lesion (LSIL). However, we found a cluster of 42 best concordant readers. The overall κ value and overall weighted κ with the target diagnosis for these 42 readers were 0.45 and 0.66, respectively. This finding is in contrast with the overall κ value and overall weighted κ for the other readers of 0.39 and 0.59, respectively. Conclusions: As this finding is an estimate of the accuracy of the readers, we can infer that it will be very important to reach this level of concordance for all the participating readers. Future effort will facilitate common experiences in order to improve the reproducibility of diagnostic criteria. Digital images could be the key to reach this aim.

Background: In Italy, cervical cancer screening programmes actively invite women aged 25–64 years. Programmes are hindered by low participation. Methods: A sample of non-responder women aged 35–64 years, belonging to three different programmes (in Rome, Florence and Teramo), was randomly split into four arms: two control groups received standard recall letters to perform either Pap-test (first group) or human papillomavirus (HPV) test (second group) at the clinic. A third arm was sent letters offering a self-sampler for HPV testing, to be requested by phone, whereas a fourth group was directly sent the self-samplers home. Results: Compliance with standard recall was 13.9% (N619). Offering HPV test at the clinic had a nonsignificant effect on compliance (N616, relative risk (RR)=1.08; 95% CI=0.82–1.41). Self-sampler at request had the poorest performance, 8.7% (N622, RR=0.62; 95% CI=0.45–0.86), whereas direct mailing of the self-sampler registered the highest compliance: 19.6% (N616, RR=1.41; 95% CI=1.10–1.82). This effect on compliance was observed only in urban areas, Florence and Rome (N438, RR=1.69; 95% CI=1.24–2.30), but not in Abruzzo (N178, RR=0.95; 95% CI=0.61–1.50), a prevalently rural area. Conclusions: Mailing self-samplers to non-responders may increase compliance as compared with delivering standard recall letters. Nevertheless, effectiveness is context specific and the strategy costs should be carefully considered.

Please cite this paper as: Castle P, Bulten J, Confortini M, Klinkhamer P, Pellegrini A, Siebers A, Ronco G, Arbyn M. Age-specific patterns of unsatisfactory results for conventional Pap smears and liquid-based cytology: data from two randomised clinical trials. BJOG 2010;117:1067–1073. Objective To investigate the rate of unsatisfactory cervical cell samples in liquid-based cytology (LBC) versus conventional cytology (CC) by age. Design Randomised clinical trials. Setting Population-based cervical cancer screening in the Netherlands and Italy. Population Asymptomatic women invited for screening enrolled in two randomised trials: Netherlands ThinPrep® versus conventional cytology (NETHCON; 39 010 CC, 46 064 LBC) and New Technologies in Cervical Cancer Screening (NTCC; 22 771 CC, 22 403 LBC). Methods Comparison of categorical variables using Pearson’s chi-square test, logistic regression and trend tests. Main outcome measures Proportion of unsatisfactory samples, ratio of LBC versus CC, and variation by 5-year group. Results In NETHCON, a lower percentage of LBC samples were judged to be unsatisfactory compared with CC samples (0.33 versus 1.11%). There was no significant trend in unsatisfactory results by age group for conventional cytology (Ptrend = 0.54), but there was a trend towards an increasing percentage of unsatisfactory results with increasing age for LBC (Ptrend

Background Faecal occult blood testing (FOBT) in population screening has proved to be effective in reducing mortality from colorectal cancer. In Italy a latex agglutination FOBT has been adopted for a single-sample screening programme. The aim of this study was to examine the performance of FOBTs in the Florence screening programme over several seasons to evaluate the impact of variations in ambient temperature on the performance of the screening test. Methods Measured haemoglobin (Hb) concentrations were aggregated into seasons with their average ambient temperature (AAT). Using logistic regression, the AAT over the period preceding the test measurement was analysed. This period included the time between faecal sampling and return of the test sample (mean 7 days) and the time in the laboratory refrigerator before analysis (mean 4 days). The AAT from days 5–11 before analysis of the test sample was considered a determinant of test positivity. The Kruskal–Wallis rank test was used to evaluate the significance of seasonal and/or AAT-related differences in Hb concentration. A logistic regression model adjusted for sex, age, season and screening episode (first or repeated examination) was constructed. Results 199 654 FOBT results were examined. Mean FOBT seasonal Hb concentrations (ng/ml) were: spring 27.6 (95% CI 26.2 to 29.1); summer 25.2 (95% CI 23.1 to 27.3); autumn 29.2 (95% CI 27.7 to 30.6); winter 29.5 (95% CI 27.9 to 31.1). Logistic regression showed that there was a 17% lower probability of the FOBT being positive in summer than in winter. The results of the logistic regression showed that an increase in temperature of 1°C produced a 0.7% reduction in probability of a FOBT being positive. In the summer the probability of detecting a cancer or an advanced adenoma was about 13% lower than in the winter. Conclusions This study showed that there is a significant fall in Hb concentration at higher ambient temperatures. These results will have important implications for the organisation of immunochemical FOBT-based screening programmes, particularly in countries with high ambient temperatures.

BACKGROUND: In the New Technologies for Cervical Cancer Screening (NTCC) randomized controlled trial, no significant increase in the sensitivity of liquid-based cytology (LBC) was observed compared with conventional cytology. Both were interpreted by cytologists who had limited previous LBC experience. The objective of the current study was to assess whether different results could be expected with experienced LBC interpreters. METHODS: A stratified, random sample of 818 LBC slides from the NTCC study was obtained. These slides were reviewed blindly and independently by 3 international experts who did not participate in the NTCC. The sensitivity and specificity of external experts were estimated for cervical intraepithelial neoplasia grade 2 or greater (CIN2+) and for CIN3+ histology, and the differences were compared with the sensitivity and specificity of the original cytologic interpretation using cutoffs of atypical squamous cells of undetermined significance (ASCUS) and low-grade squamous intraepithelial lesion (LSIL). RESULTS: With the endpoint of CIN2+ histology, the difference in sensitivity between external experts and the original interpretation was −5.3 (95% confidence interval [CI], −16.0 to 5.4) with ASCUS as the cutoff and 3.8 (95% CI, −8.2 to 15.8) with LSIL as the cutoff. External experts had slightly lower specificity using ASCUS as the cutoff (−3.4; 95% CI, −3.9 to −2.9) and LSIL as the cutoff (−0.7; 95% CI, −1.0 to −0.4). CONCLUSIONS: The accuracy of the external experts' interpretation was similar to that of the original interpretation. Therefore, the current results indicated that LBC is not expected to increase sensitivity even if it is used by interpreters who have extensive experience with this technique. Cancer (Cancer Cytopathol) 2010. © 2010 American Cancer Society.

Objective We report performance indicators and costs of the first round of a cervical cancer screening programme based on the human papillomavirus (HPV)-DNA test. Methods We implemented a demonstration study using HPV as the primary test in Guidonia, Italy (90,000 inhabitants). All women aged 25-64 were invited to undergo a Hybrid Capture II high-risk HPV test. Two cervical samplings, smear and liquid, were taken. The smear was dyed and interpreted only for HPV-positive (HPV+) women. Women with a non-negative Pap smear were referred for colposcopy, women HPV+/cytology negative were referred to one-year follow-up with HPV. A cost-analysis indicated the price at which the HPV-based and cytological screening would cost the same per screened woman and per lesion found. Results Of 24,000 women invited, 7639 accepted and 427 (5.6%) were HPV+; 141 (34%) of these had a non-negative Pap test, and 20 cervical intraepithelial neoplasia (CIN) 2 or higher were found (positive predictive value 15%). Compliance to one-year follow-up was 58% (166/286); 90 (54%) were HPV-positive and five additional lesions were found (positive predictive value 9%; overall detection rate 3.4/1000). The cost analysis showed that at a price of 8.3 euros per HPV DNA test, the strategy using HPV as primary test followed by cytological triage would cost the same per screened woman, while at a price of 12.7 euros it would have the same cost per CIN2+ found. Conclusion The workload for management of positive women was similar to cytological screening. Low compliance to one-year follow-up was the main barrier to effectiveness. The price of HPV test should be about 9 euros to maintain the same screening budget, and can go as high as 13 euros per lesion found.

Summary Background Human papillomavirus (HPV) testing is known to be more sensitive, but less specific than cytology for detecting cervical intraepithelial neoplasia (CIN). We assessed the efficacy of cervical-cancer screening policies that are based on HPV testing. Methods Between March, 2004, and December, 2004, in two separate recruitment phases, women aged 25–60 years were randomly assigned to conventional cytology or to HPV testing in combination with liquid-based cytology (first phase) or alone (second phase). Randomisation was done by computer in two screening centres and by sequential opening of numbered sealed envelopes in the remaining seven centres. During phase one, women who were HPV-positive and aged 35–60 years were referred to colposcopy, whereas women aged 25–34 years were referred to colposcopy only if cytology was also abnormal or HPV testing was persistently positive. During phase two, women in the HPV group were referred for colposcopy if the HPV test was positive. Two rounds of screening occurred in each phase, and all women had cytology testing only at the second round. The primary endpoint was the detection of grade 2 and 3 CIN, and of invasive cervical cancers during the first and second screening rounds. Analysis was done by intention to screen. This trial is registered, number ISRCTN81678807. Findings In total for both phases, 47 001 women were randomly assigned to the cytology group and 47 369 to HPV testing. 33 851 women from the cytology group and 32 998 from the HPV-testing group had a second round of screening. We also retrieved the histological diagnoses from screening done elsewhere. The detection of invasive cervical cancers was similar for the two groups in the first round of screening (nine in the cytology group vs seven in the HPV group, p=0·62); no cases were detected in the HPV group during round two, compared with nine in the cytology group (p=0·004). Overall, in the two rounds of screening, 18 invasive cancers were detected in the cytology group versus seven in the HPV group (p=0·028). Among women aged 35–60 years, at round one the relative detection (HPV vs cytology) was 2·00 (95% CI 1·44–2·77) for CIN2, 2·08 (1·47–2·95) for CIN3, and 2·03 (1·60–2·57) for CIN2 and 3 together. At round two the relative detection was 0·54 (0·23–1·28) for CIN2, 0·48 (0·21–1·11) for CIN3, and 0·51 (0·28–0·93) for CIN2 and 3 together. Among women aged 25–34 years, there was significant heterogeneity between phases in the relative detection of CIN3. At round one the relative detection was 0·93 (0·52–1·64) in phase one and 3·91 (2·02–7·57) in phase two. At round two the relative detection was 1·34 (0·46–3·84) in phase one and 0·20 (0·04–0·93) in phase two. Pooling both phases, the detection ratio of CIN2 for women aged 25–34 years was 4·09 (2·24–7·48) at round one and 0·64 (0·23–1·27) at round two. Interpretation HPV-based screening is more effective than cytology in preventing invasive cervical cancer, by detecting persistent high-grade lesions earlier and providing a longer low-risk period. However, in younger women, HPV screening leads to over-diagnosis of regressive CIN2. Funding European Union, Italian Ministry of Health, Regional Health Administrations of Piemonte, Tuscany, Veneto and Emilia-Romagna, and Public Health Agency of Lazio.

Immunochemical faecal occult blood tests have shown a greater sensitivity than guaiac test in colorectal cancer screening, but optimal number of samples and cutoff have still to be defined. The aim of this multicentric study was to evaluate the performance of immunochemical-based screening strategies according to different positivity thresholds (80, 100, 120 ng ml(-1)) and single vs double sampling (one, at least one, or both positive samples) using 1-day sample with cutoff at 100 ng ml(-1) as the reference strategy. A total of 20 596 subjects aged 50-69 years were enrolled from Italian population-based screening programmes. Positivity rate was 4.5% for reference strategy and 8.0 and 2.0% for the most sensitive and the most specific strategy, respectively. Cancer detection rate of reference strategy was 2.8 per thousand, and ranged between 2.1 and 3.4 per thousand in other strategies; reference strategy detected 15.6 per thousand advanced adenomas (range=10.0-22.5 per thousand). The number needed to scope to find a cancer or an advanced adenoma was lower than 2 (1.5-1.7) for the most specific strategies, whereas it was 2.4-2.7, according to different thresholds, for the most sensitive ones. Different strategies seem to have a greater impact on adenomas rather than on cancer detection rate. The study provides information when deciding screening protocols and to adapt them to local resources.

Summary Background Human-papillomavirus (HPV) testing is more sensitive, but less specific, than conventional cytology for detecting high-grade cervical intraepithelial neoplasia (CIN). We assessed whether HPV testing with triage by p16-INK4A overexpression can increase specificity while maintaining sensitivity. Methods HPV-positive women were enrolled between June 10, 2003, and Dec 31, 2004 in a multicentre randomised controlled trial, which compared stand-alone HPV testing by Hybrid Capture 2 (experimental group) with conventional cytology, were referred for colposcopy. In seven of nine centres, cytospin preparations from these women were tested for p16-INK4A overexpression by immunostaining. The sensitivity and specificity for CIN grade 2 or more, determined at blind review of histology, were calculated for these women. We also estimated the relative sensitivity and relative referral to colposcopy that would have been obtained by HPV testing with p16-INK4A triage compared with conventional cytology. This trial is registered as a Standard Randomised Controlled Trial, number ISRCTN81678807. Findings 24 661 women were randomly assigned to the experimental group. 1137 women (74% of those undergoing colposcopy in relevant centres), including 50 with CIN2 and 42 with CIN3 or cancer, had valid p16-INK4A immunostaining. For the endpoint of CIN2+, sensitivity and specificity of p16-INK4A (deemed positive with any number of stained cells—except endocervical, metaplastic, and atrophic cells if morphologically normal) in HPV-positive women of any age were 88% (81 of 92; 95% CI 80–94) and 61% (633 of 1045; 57–64), respectively. In the 35–60-year age group, the relative sensitivity of HPV testing and p16-INK4A triage versus conventional cytology for CIN2+ was 1·53 (95% CI 1·15–2·02) and relative referral was 1·08 (0·96–1·21). In the 25–34-year age group, relative sensitivity was 3·01 (1·82–5·17) and relative referral was 1·15 (0·96–1·37). In the latter age group, when 5% or more stained cells were deemed positive, the corresponding values were 2·06 (1·20–3·68) and 0·58 (0·46–0·73), respectively. Interpretation HPV testing with p16-INK4A triage produces a significant increase in sensitivity compared with conventional cytology, with no substantial increase in referral to colposcopy. Funding European Union, Italian Ministry of Health, Regional Health Administrations of Piemonte, Tuscany, Veneto, and Emilia-Romagna, and Public Health Agency of Lazio Region.

OBJECTIVE To compare conventional cervical testing (CCT) and liquid-based cytology (LBC) within a randomized trial performed during 2001-2002 in the Abruzzo Region of Italy, including a cost-outcome comparative analysis. STUDY DESIGN Study subjects were recruited in the framework of a controlled, randomized study organized in the Abruzzo Region. Women aged 2 6-64 years were randomized to an active arm (LBC) or control arm (CC1). The particip ating laboratories had no previous ex perience with LBC. RESULTS The inadequacy rate was 4.3% in CCT and 1.3% in the LBC arm (D < 0.001). Atypical squamous cells of undetermined sign ifi cance and atypical glands of undetermined significance reports were more frequent at CCT vs. LBC. A small, insignificant excess of low grade squamous intraepithelial lesions or high grade squamous epithelial lesions+ reports was observed in the LBC arm. The cervical intraepithelial neoplasia 2+ (CIN2+) detection rate was not statistically different in the 2 arms (CCT=0.54%, LBC= 0.66%, p = 0.28). In the overall series positive predictive value was slightly but not significantly higher in the LBC arm. LBC increased costs by 4.2% per both screened women and CIN2+ detected. CONCLUSION The study reflects the introductory phase of LBC in laboratories without prior LBC experience. In this setting LBC reduced the inadequacy rate and decreased reading and was at least as sensitive as and more specific than CCT. Utilization of LBC in organized screening programs will be based on local feasibility, considering that the high cost of LBC is only partially compensated for by other benefits, such as residual cellular material, available for molecular testing, including human papillomavirus testing.

The (32)P-post-labelling assay has emerged as a major tool for detecting bulky DNA adducts in subjects exposed to carcinogens, especially aromatic compounds. However, the (32)P-post-labelling protocol still requires the use of high amounts of radioactivity, i.e. 25-50 muCi per sample, an obstacle that limits its use in large studies. The characterization of the DNA adducts measured is also limited. Methodological improvements and increased DNA adduct characterization are necessary to make this assay capable of achieving higher throughput. A new protocol was tested to ensure efficient hydrolysis to reduce the use of radioactive material and to obtain higher chromatography resolution. Different chromatography systems based on high-urea or ammonium hydroxide systems were also employed to characterize the adducts being measured. Improvements were tested by re-analysing DNA adducts in a group of police officers and urban residents in Genoa, Italy. The analysis of carcinogen-modified DNA standards was also included in the study for qualitative and quantitative comparison. An efficient DNA digestion was obtained using a method involving hydrolysis by micrococcal nuclease and a mixture of two spleen phosphodiesterases at fixed concentrations. A 72% reduction of the amount of radioactivity used for labelling was achieved in respect to the non-modified protocol without loss of DNA adduct sensitivity. An improved chromatography resolution was obtained by reducing the volume of sample to be spotted on the chromatogram. Lower volume of spotting sample can decrease sample diffusion and the formation of unresolved spots on the thin-layer chromatography plate. The amount of output produced using a single batch of carrier-free [gamma-(32)P]ATP was increased by about 3.5-fold. A complex pattern of DNA adducts was observed in leukocytes using both high-urea or isopropanol-ammonium hydroxide systems, two techniques effective in the detection of aromatic DNA adducts. The above observations indicate that DNA adducts being measured are likely to have been induced by aromatic compounds.

We assessed the correlation between quantitative results of immunological faecal occult blood testing (I-FOBT) and colonic lesions (191 colorectal cancers, 890 adenomas) detected at colonoscopy in 2597 FOBT+ (cutoff 100 ng ml(-1) Hb) subjects. At univariate analysis, a higher average faecal Hb content was significantly associated with male gender (P=0.003), age (P=0.02), and colonoscopy findings (P=0.000). Among adenomas, higher faecal Hb content was significantly associated with size (P=0.0000), presence of severe dysplasia (P=0.0001), presence of villous component (P=0.0002), and location in the left colon (P=0.003). At multivariate analysis adjusting for potential confounders, age (P=0.03), size (P=0.0000), and location in the left colon (P=0.0005) were confirmed as having an independent association with higher faecal Hb content. Immunological FOBT is confirmed to be a specific screening test to detect cancer and adenoma, with a low positivity rate (3.7%) and a high positive predictive value (41.5%). Faecal Hb content is significantly higher for those lesions (cancer and high-risk adenomas) screening is aimed at detecting.

Within a multicentre controlled trial framework, an external quality control (EQC) was scheduled to evaluate the interlaboratory reproducibility of liquid-based cytology. In particular, this EQC intended to evaluate the reproducibility of the ASCUS diagnosis. A selected set of 30 slides (4 within normal limit cases, 16 atypical squamous cells of undetermined significance; 4 low-grade squamous intraepithelial lesions and 6 high-grade squamous intraepithelial lesions) circulated among the 13 laboratories involved in the trial. Kappa values were obtained from the comparison between individual laboratory diagnoses and majority diagnoses with target diagnoses. Specific kappa values resulted moderate to high for HSIL and low to moderate for LSIL and WNL. Meanwhile, the specific kappa for ASCUS was below 0.4 in 12 of 13 participating laboratories. The lack of reproducibility for ASCUS was not a result of the introduction of this new technology but rather to the low reproducibility of the ASCUS category itself stemming from intrinsic uncertainties in the reporting criteria. Diagn. Cytopathol. 2007;35:541-544. © 2007 Wiley-Liss, Inc.

Compared to spontaneous screening, an organized screening programme is characterized by the presence of protocols and recommendations for all stages including follow-up. Despite the availability of well-functioning screening programmes throughout the country, the follow-up protocol after an abnormal Pap test and negative colposcopy is not clearly defined in Italy, and there is no uniformity of indications. HPV testing for oncogenic human papillomavirus (hr-HPV) has a high negative predictive value (NPV) and high positive predictive value (PPV) for CIN2+ and its employment can reduce follow-up assessments. In order to provide indications about the management of women with ASC-US+ and the follow-up of women with cytological abnormalities and negative colposcopy, a literature analysis was carried out, taking into consideration European and American guidelines and good practice recommendations from the most important scientific associations and regulatory agencies. GISCi (Italian Group for Cervical Screening) drafted recommendations for the management of women with ASC-US, L-SIL, ASC-H, AGC, and H-SIL until their return to the routine screening interval. This protocol can be applied not only in the management of abnormal Pap smears in cytology-based programmes, but also in the management of abnormal Pap test triage after HPV positive test when HPV is the primary screening test. The protocols approved within the screening programmes must have an extensive consensus among all involved professionals, including any that women might meet outside the programme.

The aim of this work was to evaluate the role of human papillomavirus (HPV) testing and anal cytology, considering a population of HIV-positive and negative men who have sex with men (MSM), at high risk of sexually transmitted diseases (STD), in order to ascertain which of the methods examined is the best screening strategy for the prevention of anal cancer.In the period 06/2013-07/2014 at the "MTS Centre" of the University of Florence, 87 male patients, homo/bi-sexual, of which 46 HIV-negative and 41 HIV-positive, were recruited for anal Pap smear and HPV testing. All patients with an "abnormal" cytological result underwent anoscopy with possible biopsy.HPV testing was positive in 73 patients (83.6%). Cytology was negative in 50 patients (57.5%), inconclusive in 14 patients (16.1%), abnormal in 23 patients (26.4%): 14 ASC-US (19.2%), 4 ASC-H (5.5%), 5 L-SIL (6.8%), 0 H-SIL. Anoscopy with biopsy led to diagnosis of AIN I in 10 cases, of which 6 ASC-US+ and 4L-SIL+, AIN II in only 1 case, LS-IL+.Anal HPV testing, when used in primary screening, lead to a high number of "false positives", given the too high prevalence of HPV infection in MSM, the highest risk population targeted for screening. So we propose a screening program with anal cytology which has a high sensitivity for detection of AIN while is a poor predictor of the severity of these lesions; therefore, all patients with abnormal anal Pap smear should undergo anoscopy with biopsy.

Background Harmonization of fecal immunochemical tests for hemoglobin (FIT-Hb) is crucial to compare clinical outcomes in screening programs. The lack of reference materials and standard procedures does not allow the use of usual protocols to compare methods. We propose 2 protocols, based on artificial biological samples (ABS), to discriminate preanalytical and analytical variation and investigate clinical performances. The protocols were used to compare 2 FIT systems available on European markets: the OC-Sensor Diana (Eiken, Tokyo, Japan) and HM-JACKarc (Kyowa-Medex, Tokyo, Japan). Methods ABS were obtained adding Hb to Hb-free feces. In the first procedure, 35 ABS were collected for each collection device and analyzed on both systems. In the second, 188 ABS (106 positive and 82 negative) were ­collected and tested on the specific systems. Passing-Bablock (PB), Pearson's correlation coefficients (R) and Bland-Altman difference analysis were used to compare data. Results PB, R and mean standard errors for Bland-Altman analysis (Diana vs. Arc) results were 0.93x-0.56: R = 0.97 and 19%; and 1.09x + 5.60: R = 0.96 and −18%; for Diana and Arc devices, respectively. No correlations and no difference in positive/negative assessment were observed with the second protocol. Conclusions A good correlation was observed in comparing data generated using collection devices on the 2 systems. Manufacturers have developed different sample collection procedures for feces: therefore, data from different systems cannot easily be compared. Adoption of protocols to discriminate preanalytical and analytical variation would be a significant contribution to harmonization of FIT, facilitating data comparison and information acquisition for sample collection strategy and effect of buffers on systems.

Background: Human papillomavirus (HPV)–based screening needs triage. In most randomized controlled trials (RCTs) on HPV testing with cytological triage, cytology interpretation has been blind to HPV status. Methods: Women age 25 to 60 years enrolled in the New Technology in Cervical Cancer (NTCC) RCT comparing HPV testing with cytology were referred to colposcopy if HPV positive and, if no cervical intraepithelial neoplasia (CIN) was detected, followed up until HPV negativity. Cytological slides taken at the first colposcopy were retrieved and independently interpreted by an external laboratory, which was only aware of patients’ HPV positivity. Sensitivity, specificity, and positive (PPV) and negative (NPV) predictive values were computed for histologically proven CIN2+ with HPV status–informed cytology for women with a determination of atypical squamous cells of undetermined significance (ASCUS) or more severe. All statistical tests were two-sided. Results: Among HPV-positive women, informed cytology had cross-sectional sensitivity, specificity, PPV and 1-NPV for CIN2+ of 85.6% (95% confidence interval [CI] = 76.6 to 92.1), 65.9% (95% CI = 63.1 to 68.6), 16.2% (95% CI = 13.0 to 19.8), and 1.7 (95% CI = 0.9 to 2.8), respectively. Cytology was also associated with subsequent risk of newly diagnosed CIN2+ and CIN3+. The cross-sectional relative sensitivity for CIN2+ vs blind cytology obtained by referring to colposcopy and following up only HPV positive women who had HPV status–informed cytology greater than or equal to ASCUS was 1.58 (95% CI = 1.22 to 2.01), while the corresponding relative referral to colposcopy was 0.95 (95% CI = 0.86 to 1.04). Conclusions: Cytology informed of HPV positivity is more sensitive than blind cytology and could allow longer intervals before retesting HPV-positive, cytology-negative women.

Objective: The aim of this study was to evaluate the inter-laboratory reproducibility for atypical glandular cells (AGC) (The Bethesda System (TBS) 2001) of the laboratories involved in the screening programmes in Italy. Methods: A set of 35 selected slides were circulated among 167 laboratories involved in local population-based cervical screening programmes. Each laboratory provided one single diagnosis per smear. The smears were read blind to the original diagnosis and to the diagnoses provided by other laboratories. A ‘majority’ diagnosis was defined for each case and assumed as the reference standard. The diagnosis provided from each laboratory was compared with the majority diagnosis. Results: According to the majority report the 35 slides in the set were classified as negative in nine cases, AGC in eight, adenocarcinoma in eight, and squamous lesion or squamous + glandular lesion in 10. The crude agreement between all pairs of laboratories was 49.43%. K-values were 0.46, 0.21, 0.34, 0.36 and 0.32 for negative, AGC/AIS (adenocarcinoma in situ of endocervix), AdenoCa, Sq/Sq + Gl and all reporting categories respectively. Concordance according to overall K was moderate to substantial in 77% of the participating laboratories. Conclusions: The present study shows that the AGC category is not easily reproducible. The data confirmed the importance, in a screening scenario, of AGC/AIS diagnoses, but also presented difficulties in differentiating between the two diagnoses. In addition to the results obtained from the circulation of the slides, laboratories which had annually a low number of cervical smears were able to gain experience focused on particular morphological pictures.

The purpose of this study was to determine the prognostic significance of the expression of p53 and Ki-67 in non-small cell lung cancer (NSCLC) using immunocytochemical detection. All consecutive NSCLC cases were selected for study, and, after surgery, a part of each tumor sample was frozen at -20 degrees C and stored for immunocytochemical studies. Overexpression of p53 was associated significantly with worse patient outcome in stage I disease, whereas no excess risk was evident in stage II and III cases. The same pattern was observed for Ki-67 expression. The excess risk in stage I cases with p53 and Ki-67 overexpression was observed only in adenocarcinoma. These findings are in agreement with other retrospective studies and support the hypothesis that p53 alteration may have different roles in adenocarcinoma and in squamous cell carcinoma, such as a carcinogenic factor for both cellular types but progression only for adenocarcinoma.

The free-to-total prostate-specific antigen ratio (F/T PSA) is associated with the presence of prostate cancer and is thus used as an indicator for suspicion of prostate cancer and as a determinant for biopsy. We reviewed a recent retrospective series of 966 consecutive prostate biopsies where F/T PSA was blindly determined and did not influence biopsy indication. We simulated the association of F/T PSA with biopsy outcome and its impact as a biopsy determinant. When adopting an F/T PSA cutoff of 10%, 13%, 16% or 20% among random sextant biopsies in the 4–10 ng/mL total PSA range, the sensitivity was 15%, 37%, 55% and 72% and the specificity 89%, 80%, 64% and 44%, respectively. Using F/T PSA as a biopsy determinant, from 1.7 to 2.6 cancer biopsies would have been delayed to avoid 10 benign biopsies. As this balance is not acceptable, F/T PSA has no role as a biopsy indicator and its clinical use is questionable.

Within a large Italian randomized trial on new technologies for cervical cancer screening involving 7 laboratories with different levels of experience, an intralaboratory and interlaboratory quality control program for human papillomavirus (HPV) DNA testing by Hybrid Capture 2 (HC2; Digene, Gaithersburg, MD) was implemented. To monitor the hybridization and detection steps, target samples containing purified, concentration-defined, HPV DNA were introduced in each test run. Only 3 of 1,024 showed a mistake in a positive vs negative classification with a 1 relative light unit (RLU)/positive control specimen (PC) ratio cutoff. To monitor the preanalytic steps (particularly denaturation), blinded specimens (33 collected in PreservCyt [Cytyc, Boxborough, MA] and 36 in Specimen Transport Medium [STM, Digene]) were centrally prepared, divided into aliquots, and sent to each laboratory. The multiple-rater κ scores for negative (

The CellSlide Processor (Menarini Diagnostics) is an automatic computer-controlled instrument, designed to prepare standardized, thin-layer cytological preparations using a filtration process. In this report, this new processor has been tested in a multicentric investigation, designed to evaluate cell recovery, cellular morphology and possible time saved in reading the slide. The results of this study show that the CellSlide processing system is suitable for use in the preparation of liquid-based gynaecology samples. Furthermore, the high-quality smear produced by the CellSlide decreases the reading time by approximately 20-25%.

BACKGROUND The current study was conducted to evaluate the cost-effectiveness of triaging for colposcopy using human papillomavirus (HPV) testing. METHODS HPV tests were performed in a consecutive series of women who were referred for colposcopy for persistent atypical squamous cells of undetermined significance (ASCUS)-favor reactive (n = 35 women), ASCUS-favor squamous epithelial lesion (n = 164 women), atypical glandular cells of undetermined significance (n = 74 women), low-grade squamous epithelial lesion (n = 161 women), or high-grade squamous epithelial lesion (n = 78 women). The cost effectiveness of triaging women with ASCUS results using HPV testing was determined compared with the current protocol. RESULTS The sensitivity of HPV testing for cervical intraepithelial neoplasia > Grade 2 was very high. Cost analysis showed a moderate increase in cost with the addition of HPV triage. CONCLUSIONS Because HPV testing is highly sensitive, it may be useful as an alternative to the current policy of 6-month repeat cytology for women with ASCUS. Cancer (Cancer Cytopathol) 2005. © 2004 American Cancer Society.

The study was aimed at assessing interlaboratory reproducibility in the reporting of cervical smears in the atypical squamous cells of undetermined significance (ASCUS) category. A set of 50 selected slides circulated among 89 laboratories, currently involved in population-based screening programmes for cervical cancer, which provided a diagnostic report according to four main reporting categories based on the 1991 Bethesda system. Interlaboratory agreement was determined according to kappa (K) statistics: overall and weighted K values were determined for each laboratory and for single reporting categories. The results showed a very low reproducibility for the ASCUS category. This finding supports the Bethesda system 1991 recommendation to limit the use of this reporting category and suggests that the clinical response to ASCUS reports should be decided locally, based on the observed positive predictive value for cervical intraepithelial neoplasia 2 or more severe lesions.

The cost-effectiveness of qualifying ASCUS cases into two different subcategories, favoring a reactive (ASCUS-R) or dysplastic process (ASCUS-S), was evaluated at the Centro per lo Studio e la Prevenzione Oncologica of Florence in a prospective study. The study determined the positive predictive value (PPV) for histologically confirmed cervical intraepithelial neoplasia grade 2 (CIN2) or more (CIN>) severe lesion of the two ASCUS subgroups. ASCUS-S had a PPV (10.78%) comparable to low-grade squamous intraepithelial lesions (LSIL) (11.40%). For ASCUS-R cases, the recommendation of 6-mo repeat cytology prompting colposcopy in cases of persistent ASCUS or more severe cytology was also effective, as it selected a subgroup with a relatively high PPV (10.34%). The cost-effectiveness of a protocol based on ASCUS qualification was compared with two other possible options for nonqualified ASCUS cases: immediate colposcopy and colposcopy in persistent ASCUS at 6-mo repeat cytology.. The detection rate of CIN2> was substantially higher using ASCUS qualification (35.9 vs 14.8 or 17.1). The cost per ASCUS subject was €24.99, 27.11, or 25.14 and that per CIN2> detected was €697, 1,831 or 1,470 for the three options, respectively. The evidence that ASCUS detection option implies a higher detection rate of CIN2> and subsequently a lower cost per CIN2> detection must be considered with caution and deserves confirmation by other comparative studies. Diagn. Cytopathol. 2003;29:4–7. © 2003 Wiley-Liss, Inc.

The study objective was assessing whether circulation of smears and discussion of those with differing interpretation can increase reproducibility between laboratories. The study included: the blind interpretation of a first set of 194 smears among seven laboratories, the discussion of smears with discrepant diagnoses during the previous phase and the blind interpretation of a second set of smears of same size and characteristics. After discussions, the overall weighted kappa increased in five laboratories (substantially in three : +50%, +27% and +20%). However, no change was observed in one laboratory and a slight decrease (-4%) in another. The latter interpreted the second set of smears at a longer time interval from discussions. Agreement improved for all diagnostic classes except low grade intraepithelial neoplasia (LSIL). Overall, the intervention increased diagnostic agreement, but its effect varied with laboratory and by diagnostic class and could be transient. Continued programmes of smear exchange and discussion appear to be advisable.

Interlaboratory reproducibility of the immunocytochemical assessment of oestrogen and progesterone receptors and proliferative activity in fine needle aspiration of breast cancer The purpose of this study was to establish the interlaboratory reproducibility of immunocytochemical analysis of oestrogen (ER) and progesterone (PR) expression and Mib1 growth fraction on fine needle aspiration (FNA) smears. A set of 44 immunostained slides for ER, PR and Mib1 were randomly selected from the archives of the Center for the Study and Prevention of Cancer (CSPO) of Florence, Italy, and submitted for reading to 6 Italian laboratories. The generalized kappa statistic was used as an indicator of agreement among the six laboratories. A good correlation for ER and PR was evident. For Mib1 the results showed some discrepancies. In addition to adequate standardization of procedures, these data confirm that the reliability of the immunocytochemistry is strictly linked to accurate analysis of the results.

With the introduction of Human papillomavirus (HPV) vaccination in young girls in 2007, it is important to monitor HPV infections and epidemiological changes in this target population. The present study has evaluated the detection of human papillomavirus DNA in paired cervical and urine samples to understand if HPV testing in urine could be used as non-invasive method to monitor HPV status in young women. The study enrolled 216 twenty five-year-old women, resident in Florence and invited for the first time to the cervical cancer Screening Program within a project evaluating the impact of HPV vaccination. HPV genotyping was performed on 216 paired urine and cervical samples. The overall concordance between cervix and urine samples, investigated by HPV genotyping (INNO-LiPA HPV Genotyping Extra), was: 85.6% (184/215), 84.6% (182/215), 80% (172/215) when the same HPV, at least the same HR HPV and all HR HPV, respectively, were detected. HPV type specific concordance in paired urine and cervical samples was observed in 85.8% (175/204) of women with normal cytology and in seven out of nine women with abnormal cytology. Urine seems to be a suitable and reliable biological material for HPV DNA detection as evidenced by the high concordance with HPV detected in cervical samples. These results suggest that urine could be a good noninvasive tool to monitor HPV infection in vaccinated women.

In this case-cohort study, we examined the association between bulky DNA adducts and the risk of lung cancer within the European Prospective Investigation into Cancer and Nutrition (EPIC) Spanish cohort with an average 7-year follow-up, including 98 cases of primary lung cancer and 296 subjects randomly selected from the cohort. Aromatic adducts were measured using 32 P-postlabeling in leukocyte DNA from blood samples collected at enrollment. The association between DNA adducts and the risk of lung cancer was estimated using a Cox proportional hazards model with a modified partial likelihood. There was an overall significant increased risk for developing lung cancer when DNA adduct concentrations were doubled, with relative risk (RR) adjusting for all relevant confounders of 1.36 with 95% confi dence interval (CI) 1.18–157. There was a significant increased risk for developing lung cancer when DNA adduct concentrations were doubled for current smokers and among subjects exposed to PAH at work; there was also a slightly higher increase among males than females. However, no statistically significant differ ences were observed for the effect of adduct levels across smoking status, sex or occupational exposure to PAH. A meta-analysis combined four prospective studies, including this study, resulting in a significant association among current smokers, with an over all estimate of 34% increase in the risk of lung cancer when doubling the level of aromatic DNA adducts in leukocytes.

Human papillomavirus (HPV) infection is highly prevalent in women migrating from countries where cervical screening is not implemented. The variety of HPV genotypes, their prevalence and the association with cervical abnormalities has been investigated by several groups in women moving mainly from Eastern Europe, Africa, and Southern Asia to Italy. All studies are concordant on the elevated rate of HPV infection among immigrants, which is four times higher than that observed among age-matched Italian women. The HPV prevalence among short-term migrants and characterization of viral variants showed that the high prevalence of HPV reflects either individual lifestyle or high prevalence of HPV in the country of origin. The high burden of HPV infection correlates very well with the high incidence of cervical cancer in migrant women. In fact, during the years 2000–2004 the cervical cancer incidence in women from Central and Eastern Europe and living in Central Italy was 38.3 per 100,000, which is statistically significant higher than that of native Italian women (6 per 100,000). In this study, we pooled together the results of three independent studies originally designed to assess the distribution and the prevalence of HPV genotypes among 499 immigrant women living in Southern Italy. A total of 39 mucosal HPV genotypes were identified. The 12 genotypes (HPV16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, and 59) classified as carcinogenic to humans (group 1) accounted for >80% of all infections. HPV16 was the most common viral type in all groups with frequency rates ranging from 15.4% in Africa to 51.1% in Eastern and Southern European HPV-positive women. The high prevalence of oncogenic HPVs and cervical cancer risk among migrant women, together with the lower participation in screening programs, demands for an urgent implementation of preventive strategies to increase screening and vaccine coverage and viral monitoring of uncommon HPV genotypes potential spreading in settled population.

Background Although among women a decreasing prevalence of human papillomavirus (HPV) infection with increasing age has been consistently observed in high-resource countries, different age profiles have been reported elsewhere. Methods We compared the age profile of high-risk (HR)-HPV prevalence in nine different areas of Northern and Central Italy by studying the women recruited in the intervention arm of the New Technologies in Cervical Cancer study and tested by Hybrid Capture 2. Differences in the age-distribution of HPV infection were investigated in each centre by the joinpoint approach in a logistic model. 46,900 women aged 25 to 60 years were included in the analysis. Results The HR-HPV age-standardised (on Italian population) prevalence ranged from 5.7% (Trento) to 10.3% (Ravenna). HR-HPV prevalence decreased as a logistic function of increasing age in 6 of 9 centres (Trento, Verona, Florence, Bologna, Imola, and Viterbo). The effect of age on HR-HPV prevalence slopes did not differ significantly among these 6 centres, whereas significant heterogeneity in intercepts (p