Ovarian cancer (OC) is the most lethal of all gynecological cancers. Due to vague symptoms, OC is mostly detected at advanced stages, with a 5-year survival rate (SR) of only 30%; diagnosis at stage ...I increases the 5-year SR to 90%, suggesting that early diagnosis is essential to cure OC. Currently, the clinical need for an early, reliable diagnostic test for OC screening remains unmet; indeed, screening is not even recommended for healthy women with no familial history of OC for fear of post-screening adverse events. Salivary diagnostics is considered a major resource for diagnostics of the future. In this work, we searched for OC biomarkers (BMs) by comparing saliva samples of patients with various stages of OC, breast cancer (BC) patients, and healthy subjects using an unbiased, high-throughput proteomics approach. We analyzed the results using both logistic regression (LR) and machine learning (ML) for pattern analysis and variable selection to highlight molecular signatures for OC and BC diagnosis and possibly re-classification. Here, we show that saliva is an informative test fluid for an unbiased proteomic search of candidate BMs for identifying OC patients. Although we were not able to fully exploit the potential of ML methods due to the small sample size of our study, LR and ML provided patterns of candidate BMs that are now available for further validation analysis in the relevant population and for biochemical identification.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Facing the SARS-CoV-2 epidemic requires intensive testing on the population to early identify and isolate infected subjects. During the first emergency phase of the epidemic, RT-qPCR on ...nasopharyngeal (NP) swabs, which is the most reliable technique to detect ongoing infections, exhibited limitations due to availability of reagents and budget constraints. This stressed the need to develop screening procedures that require fewer resources and are suitable to be extended to larger portions of the population. RT-qPCR on pooled samples from individual NP swabs seems to be a promising technique to improve surveillance. We performed preliminary experimental analyses aimed to investigate the performance of pool testing on samples with low viral load and we evaluated through Monte Carlo (MC) simulations alternative screening protocols based on sample pooling, tailored to contexts characterized by different infection prevalence. We focused on the role of pool size and the opportunity to develop strategies that take advantage of natural clustering structures in the population, e.g. families, school classes, hospital rooms. Despite the use of a limited number of specimens, our results suggest that, while high viral load samples seem to be detectable even in a pool with 29 negative samples, positive specimens with low viral load may be masked by the negative samples, unless smaller pools are used. The results of MC simulations confirm that pool testing is useful in contexts where the infection prevalence is low. The gain of pool testing in saving resources can be very high, and can be optimized by selecting appropriate group sizes. Exploiting natural groups makes the definition of larger pools convenient and potentially overcomes the issue of low viral load samples by increasing the probability of identifying more than one positive in the same pool.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Background
The objective of this study was to evaluate the performance of a combined approach of liquid-based anal cytology and human papillomavirus (HPV) testing in predicting patients who should ...undergo high-resolution anoscopy for the early detection of anal cancer and anal intraepithelial neoplasia (AIN)-2+.
Methods
We conducted a prospective single-center quality improvement study. We consecutively enrolled men who had sex with men (MSM) attending our sexually transmitted disease clinic to undergo anal Papanicolaou (Pap) and HPV tests. All patients with an abnormal anal Pap test result and/or positive HPV test result underwent high-resolution anoscopy.
Results
We enrolled 217 MSM, 80 HIV-positive patients, and 137 HIV-negative patients. Cytology showed a sensitivity of 100%, a specificity of 64.1%, an accuracy of 66.7%, a positive predictive value (PPV) of 15.7%, and a negative predictive value (NPV) of 100% for the detection of AIN-2+. The high-risk (HR)-HPV test showed sensitivity, specificity, accuracy, PPV, and NPV of 100%, 36.4%, 40%, 9.4%, and 100%, respectively. The combination of abnormal cytology with identification of infection by at least 1 HR-HPV strain on the HPV test had a sensitivity of 100%, a specificity of 73%, an accuracy of 74.6%, a PPV of 19.1%, and an NPV of 100%.
Conclusion
Anal HR-HPV testing, complementary to cytology, improves the diagnostic accuracy of screening for anal cancer.
The role of total plasma cell-free DNA (cfDNA) in lung cancer (LC) screening with low-dose computed tomography (LDCT) is uncertain. We hypothesized that cfDNA could support differentiation between ...malignant and benign nodules observed in LDCT. The baseline cfDNA was measured in 137 subjects of the ITALUNG trial, including 29 subjects with screen-detected LC (17 prevalent and 12 incident) and 108 subjects with benign nodules. The predictive capability of baseline cfDNA to differentiate malignant and benign nodules was compared to that of Lung-RADS classification and Brock score at initial LDCT (iLDCT). Subjects with prevalent LC showed both well-discriminating radiological characteristics of the malignant nodule (16 of 17 were classified as Lung-RADS 4) and markedly increased cfDNA (mean 18.8 ng/mL). The mean diameters and Brock scores of malignant nodules at iLDCT in subjects who were diagnosed with incident LC were not different from those of benign nodules. However, 75% (9/12) of subjects with incident LC showed a baseline cfDNA ≥ 3.15 ng/mL, compared to 34% (37/108) of subjects with benign nodules (p = 0.006). Moreover, baseline cfDNA was correlated (p = 0.001) with tumor growth, measured with volume doubling time. In conclusion, increased baseline cfDNA may help to differentiate subjects with malignant and benign nodules at LDCT.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Clinical radiosensitivity varies considerably among patients, and radiation-induced side effects developing in normal tissue can be therapy limiting. Some single nucleotide polymorphisms (SNPs) have ...been shown to correlate with hypersensitivity to radiotherapy. We conducted a prospective study of 87 female patients with breast cancer who received radiotherapy after breast surgery. We evaluated the association between acute skin reaction following radiotherapy and 11 genetic polymorphisms in DNA repair genes: XRCC1 (Arg399Gln and Arg194Trp), XRCC3 (Thr241Met), XPD (Asp312Asn and Lys751Gln), MSH2 (gIVS12-6T>C), MLH1 (Ile219Val), MSH3 (Ala1045Thr), MGMT (Leu84Phe), and in damage-detoxification GSTM1 and GSTT1 genes (allele deletion).
Individual genetic polymorphisms were determined by polymerase chain reaction and single nucleotide primer extension for single nucleotide polymorphisms or by a multiplex polymerase chain reaction assay for deletion polymorphisms. The development of severe acute skin reaction (moist desquamation or interruption of radiotherapy due to toxicity) associated with genetic polymorphisms was modeled using Cox proportional hazards, accounting for cumulative biologically effective radiation dose.
Radiosensitivity developed in eight patients and was increased in carriers of variants XRCC3-241Met allele (hazard ratio HR unquantifiably high), MSH2 gIVS12-6nt-C allele (HR=53.36; 95% confidence intervals 95% CI, 3.56-798.98), and MSH3-1045Ala allele (HR unquantifiably high). Carriers of XRCC1-Arg194Trp variant allele in combination with XRCC1-Arg399Gln wild-type allele had a significant risk of radiosensitivity (HR=38.26; 95% CI, 1.19-1232.52).
To our knowledge, this is the first report to find an association between MSH2 and MSH3 genetic variants and the development of radiosensitivity in breast cancer patients. Our findings suggest the hypothesis that mismatch repair mechanisms may be involved in cellular response to radiotherapy. Genetic polymorphisms may be promising candidates for predicting acute radiosensitivity, but further studies are necessary to confirm our findings.
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GEOZS, IJS, NUK, OILJ, UL, UM, UPUK
The ITALUNG trial started in 2004 and compared lung cancer (LC) and other-causes mortality in 55-69 years-aged smokers and ex-smokers who were randomized to four annual chest low-dose CT (LDCT) or ...usual care. ITALUNG showed a lower LC and cardiovascular mortality in the screened subjects after 13 years of follow-up, especially in women, and produced many ancillary studies. They included recruitment results of a population-based mimicking approach, development of software for computer-aided diagnosis (CAD) and lung nodules volumetry, LDCT assessment of pulmonary emphysema and coronary artery calcifications (CAC) and their relevance to long-term mortality, results of a smoking-cessation intervention, assessment of the radiations dose associated with screening LDCT, and the results of biomarkers assays. Moreover, ITALUNG data indicated that screen-detected LCs are mostly already present at baseline LDCT, can present as lung cancer associated with cystic airspaces, and can be multiple. However, several issues of LC screening are still unaddressed. They include the annual vs. biennial pace of LDCT, choice between opportunistic or population-based recruitment. and between uni or multi-centre screening, implementation of CAD-assisted reading, containment of false positive and negative LDCT results, incorporation of emphysema. and CAC quantification in models of personalized LC and mortality risk, validation of ultra-LDCT acquisitions, optimization of the smoking-cessation intervention. and prospective validation of the biomarkers.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
This multicenter study describes the type-specific prevalence of HPV infection in the general population from central and southern Italy, comparing the data with previously published Italian studies.
...Women aged from 25 to 65 who attended cervical cancer screening in five different Italian regions were tested for HPV infection with Hybrid Capture II (HCII) low and high risk probes. Women repeating Pap-test upon unsatisfactory or positive results, or as a post-treatment and post-colposcopy follow-up analysis, were excluded from our study. High risk (HR) HPV positive samples were typed using GP5+/GP6+ primed PCR, followed by Reverse Line Blot for 18 high/intermediate risk HPV types, while low risk (LR) HPV positive samples were tested with type specific primers for HPV6 and HPV11.
3817 women had a valid HCII test: 350 of them (9.2%) were positive for HR probes, 160 (4.2%) for LR probes, while 57 women were positive for both. Multiple infections were detected in 97 HR HPV positive women. The most common types were HPV 16 (3%), 31 (1.2%), 51 (1%). HPV6 ranked fifth (0.6%), HPV18 ranked tenth (0.5%) and HPV11 sixteenth (0.3%).In Sardinia the prevalence of high-risk infection was 13%, significantly higher than the mean value (p < 0.00005).The distribution of the most frequent types did not significantly differ by centre (p = 0.187) and age (p = 0.085).
Because cervical cancer incidence and Pap test coverage is lower in southern than in northern Italy, a lower prevalence of high-risk infections in the general population was expected in the south. However, prevalence detected in this study for the south of the country is slightly but significantly higher than the rest of Italy. The consequence may be an epidemic of cervical cancer in the next decades if adequate screening programs are not implemented there.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Abstract
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.
HPV type distribution by cytological status represents useful information to predict the impact of mass vaccination on screening programs.
women aged from 25 to 64 who attended cervical cancer ...screening in five different Italian regions were tested for HPV infection with Hybrid Capture II (HCII) low and high risk probes. Women repeating Pap-test upon unsatisfactory or positive results, or as a post-treatment and post-colposcopy follow-up analysis, were excluded from our study. High risk (HR) HPV positive samples were typed using GP5+/GP6+ primed PCR, followed by Reverse Line Blot for 18 high/intermediate risk HPV types, while low risk (LR) HPV positive samples were tested with type specific primers for HPV6 and HPV11.
3410 women had a valid HCII and Pap-test. The prevalence of HR and LR infections was 7.0% and 3.6%, 29.1% and 13.7%, 68.1% and 31.9%, 60.0% and 0.0%, 65.0% and 12.0%, for negative, ASC-US, L-SIL, ASC-H and H-SIL cytology, respectively. The fraction of ASC-US+ cytology due to HPV 16 and 18 ranged from 11.2 (HPV 16/18 alone) to 15.4% (including HPV 16/18 in co-infection with other virus strains), and that due to HPV 6 and 11 ranged from 0.2% (HPV 6/11 alone) to 0.7% (including HPV 6/11 in co-infection with other LR virus strains).
mass vaccination with bivalent or quadrivalent HPV vaccine would modestly impact on prevalence of abnormal Pap-test in screening.
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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 for <CIN2 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.
What's new?
Human papillomavirus (HPV) DNA‐based assays are now widely recommended for cervical cancer screening, but they lack specificity. As a primary screening test, E6/E7 mRNA testing has shown a similar sensitivity for CIN3+ but a lower positivity rate than HPV DNA testing. This trial study shows that primary E6/E7 mRNA screening would miss about 3% of CIN3+ cases. Overall positivity would be 22% lower than that of HPV DNA. Triage with cytology or p16/ki67 dual staining would only marginally decrease overall colposcopy referral if retesting of E6/E7 mRNA‐positive/triage‐negative women was performed with HPV DNA assays after 12 months.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
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