[Development and validation of predictive models for esophageal squamous cell carcinoma and its precancerous lesions using terminal motif analysis in circulating cell-free DNA].

Objectives: To develop and validate predictive models for esophageal squamous cell carcinoma (ESCC) using circulating cell-free DNA (cfDNA) terminal motif analysis. The goal was to improve the non-invasive detection of early-stage ESCC and its precancerous lesions. Methods: Between August 2021 and November 2022, we prospectively collected plasma samples from 448 individuals at the Department of Endoscopy, Cancer Hospital, Chinese Academy of Medical Sciences for cfDNA extraction, library construction, and sequencing. We analyzed 201 cases of ESCC, 46 high-grade intraepithelial neoplasia (HGIN), 46 low-grade intraepithelial neoplasia (LGIN), 176 benign esophageal lesions, and 29 healthy controls. Participants, including ESCC patients and control subjects, were randomly assigned to a training set (n=284) and a validation set (n=122). The training cohort underwent z-score normalization of cfDNA terminal motif matrices and a selection of distinctive features differentiated ESCC cases from controls. The random forest classifier, Motif-1 (M1), was then developed through principal component analysis, ten-fold cross-validation, and recursive feature elimination. M1's efficacy was then validated in the validation and precancerous lesion sets. Subsequently, individuals with precancerous lesions were included in the dataset and participants were randomly allocated to newly formed training (n=243), validation (n=105), and test (n=150) cohorts. Using the same procedure as M1, we trained the Motif-2 (M2) random forest model with the training cohort. The M2 model's accuracy was then confirmed in the validation cohort to establish the optimal threshold and further tested by performing validation in the test cohort. Results: We developed two cfDNA terminal motif-based predictive models for ESCC and associated precancerous conditions. The first model, M1, achieved a sensitivity of 90.0%, a specificity of 77.4%, and an area under the curve (AUC) of 0.884 in the validation cohort. For LGIN, HGIN, and T1aN0 stage ESCC, M1's sensitivities were 76.1%, 80.4%, and 91.2% respectively. Notably, the sensitivity for jointly predicting HGIN and T1aN0 ESCC reached 85.0%. Both the predictive accuracy and sensitivity increased in line with the cancer's progression (P＜0.001). The second model, M2, exhibited a sensitivity of 87.5%, a specificity of 77.4%, and an AUC of 0.857 in the test cohort. M2's sensitivities for detecting precancerous lesions and ESCC were 80.0% and 89.7%, respectively, and it showed a combined sensitivity of 89.4% for HGIN and T1aN0 stage ESCC. Conclusions: Two predictive models based on cfDNA terminal motif analysis for ESCC and its precancerous lesions are developed. They both show high sensitivity and specificity in identifying ESCC and its precancerous stages, indicating its potential for early ESCC detection.

Platelet to Lymphocyte Ratio Correlates With Carcinoma Progression in Pancreatic Intra Epithelial Neoplasia.

BACKGROUND/AIM: Pancreatic intraepithelial neoplasia (PanIN) is the most common precursor lesion to pancreatic adenocarcinoma (PDAC). Yet no criteria to quantify patients at risk for progression to PDAC with PanIN exist. Platelet to lymphocyte ratio is an inflammatory marker that has been associated with overall survival in patients with invasive malignancies including pancreatic cancer. Preoperative sarcopenia has been linked to more aggressive diseases in pancreatic neoplasms. We aimed to assess a relation between PLR and sarcopenia as predictors for tumor progression in patients undergoing pancreatic resection for IPMN. PATIENTS AND METHODS: We retrospectively reviewed 102 patients (46 females, 56 males) who underwent pancreatic resection for PanIn. PLR was calculated and quantified using a cutoff of 110, sarcopenia was quantified using the skeletal muscle index (SMI) on preoperative abdominal imaging. Both were co-evaluated with additional demographic, clinical, pathological, and imaging data for possible correlation with PanIN associated PDAC. RESULTS: PLR was significantly elevated in patients with PanIN - associated PDAC (p=0.006). In the multivariate analysis, invasive carcinomas were significantly more prevalent in patients with PLR above 110 (OR=4.06, 95%CI=3.91-4.12, p=0.04). Patients with elevated PLR had a two-times higher risk to die in the postoperative period (HR=2.26, 95%CI=1.04-2.21, p=0.001). Patients with elevated PLR, preoperative jaundice and sarcopenia were the most likely to have PanIN-associated PDAC (OR=3.48, 95%CI=2.98-8.41, p=0.02). CONCLUSION: PLR is an independent predictive marker for the presence of PanIN associated invasive carcinoma.

Thyroid Hormones and Morphological Features of Primary Breast Cancer.

BACKGROUND/AIM: Many experimental studies have suggested the importance of thyroid hormones in breast cancer (BC) morphogenesis. The aim of this study was to evaluate the association of thyroid hormone levels in serum of patients with primary BC with morphological presentations of the disease in pathological specimens and prognosis. PATIENTS AND METHODS: We measured the serum levels of thyroid-stimulating hormone (TSH), free triiodothyronine (FT3), and free thyroxine (FT4), along with serum thymidine kinase 1 activity and examined their relation to pathological features and prognosis of 158 patients with primary BC. RESULTS: We found a significant positive association of serum FT3 level with the presence of carcinoma in situ component (CIS) (p=0.032) and its size (p=0.047), with the presence (p=0.022) and the number of multifocal/multicentric tumors (MMTs) (p=0.002), as well as with increased proliferative activity in terms of serum thymidine kinase 1 (p=0.002). Moreover, we report that each 1.0 unit rise of FT3/FT4 ratio×10 was associated with an odds ratio of 1.77 (95% confidence interval=1.17-3.30, p=0.007), 1.97 (95% confidence interval=1.17-2.67, p=0.010) and 1.56 (95% confidence interval=1.02-2.37, p=0.039) for the detection of patients with CIS, MMTs and lymphovascular invasion, respectively, after adjusting for age. We did not find statistically significant associations of serum TSH level with breast cancer`s parameters. A Cox regression survival analysis identified serum FT3 level >5.95 pmol/l as a risk factor for BC recurrence (relative risk=2.65, p=0.017), a finding that retained significance in a multivariate model (relative risk=2.52, p=0.027). CONCLUSION: The FT3/FT4 ratio is a valuable parameter predicting the presence of CIS, MMTs and lymphovascular invasion in pathological specimens. An elevated serum FT3 level is associated with the presence of CIS, MMTs, increased proliferative activity and poor prognosis.

Human papillomavirus 16 L1 gene methylation as a potential biomarker for predicting anal intraepithelial neoplasia in men who have sex with men (MSM).

The human papillomavirus (HPV) 16 early promoter and L1 gene methylation were quantitatively measured using pyrosequencing assay in anal cells collected from men who have sex with men (MSM) to determine potential biomarkers for HPV-related anal cancer. The methylation patterns of HPV16 genes, including the early promoter (CpG 31, 37, 43, 52, and 58) and L1 genes (CpG 5600, 5606, 5609, 5615, 7136, and 7145), were analyzed in 178 anal samples. The samples were diagnosed as normal, anal intraepithelial neoplasia (AIN) 1, AIN2, and AIN3. Low methylation levels of the early promoter (< 10%) and L1 genes (< 20%) were found in all detected normal anal cells. In comparison, medium to high methylation (≥ 20-60%) in the early promoter was found in 1.5% (1/67) and 5% (2/40) of AIN1 and AIN2-3 samples, respectively. Interestingly, slightly increased L1 gene methylation levels (≥ 20-60%), especially at the HPV16 5'L1 regions CpGs 5600 and 5609, were demonstrated in AIN2-3 specimen. Moreover, a negative correlation between high HPV16 L1 gene methylation at CpGs 5600, 5609, 5615, and 7145 and a percentual CD4 count was found in AIN3 HIV positive cases. When comparing the methylation status of AIN2-3 to that of normal/AIN1 lesions, the results indicated the potential of using HPV16 L1 gene methylation as a biomarker for HPV-related cancer screening.

Early Detection of Pancreatic Intraepithelial Neoplasias (PanINs) in Transgenic Mouse Model by Hyperpolarized 13C Metabolic Magnetic Resonance Spectroscopy.

While pancreatic cancer (PC) survival rates have recently shown modest improvement, the disease remains largely incurable. Early detection of pancreatic cancer may result in improved outcomes and therefore, methods for early detection of cancer, even premalignant lesions, may provide more favorable outcomes. Pancreatic intraepithelial neoplasias (PanINs) have been identified as premalignant precursor lesions to pancreatic cancer. However, conventional imaging methods used for screening high-risk populations do not have the sensitivity to detect PanINs. Here, we have employed hyperpolarized metabolic imaging in vivo and nuclear magnetic resonance (1H-NMR) metabolomics ex vivo to identify and understand metabolic changes, towards enabling detection of early PanINs and progression to advanced PanINs lesions that precede pancreatic cancer formation. Progression of disease from tissue containing predominantly low-grade PanINs to tissue with high-grade PanINs showed a decreasing alanine/lactate ratio from high-resolution NMR metabolomics ex vivo. Hyperpolarized magnetic resonance spectroscopy (HP-MRS) allows over 10,000-fold sensitivity enhancement relative to conventional magnetic resonance. Real-time HP-MRS was employed to measure non-invasively changes of alanine and lactate metabolites with disease progression and in control mice in vivo, following injection of hyperpolarized [1-13C] pyruvate. The alanine-to-lactate signal intensity ratio was found to decrease as the disease progressed from low-grade PanINs to high-grade PanINs. The biochemical changes of alanine transaminase (ALT) and lactate dehydrogenase (LDH) enzyme activity were assessed. These results demonstrate that there are significant alterations of ALT and LDH activities during the transformation from early to advanced PanINs lesions. Furthermore, we demonstrate that real-time conversion kinetic rate constants (kPA and kPL) can be used as metabolic imaging biomarkers of pancreatic premalignant lesions. Findings from this emerging HP-MRS technique can be translated to the clinic for detection of pancreatic premalignant lesion in high-risk populations.

Unexpected metastasis of intraductal papillary neoplasm of the bile duct without an invasive component to the brain and lungs: A case report.

BACKGROUND: Despite an expanding number of studies on intraductal papillary neoplasm of the bile duct (IPNB), distant metastasis remains unexplained especially in cases of carcinoma in situ. In the present study, we report a rare and interesting case of IPNB without invasive components that later metastasized to lungs and brain. CASE SUMMARY: A 69-year-old male was referred to our hospital due to suspected cholangiocarcinoma. Laboratory tests on admission reported a mild elevation of alkaline phosphatase, γ-glutamyl transpeptidase, and total bilirubin in serum. Endoscopic retrograde cholangiography revealed a filling defect in the common bile duct (CBD) extending to the left hepatic duct. Peroral cholangioscopy delineated a tumor in the CBD that had a papillary pattern. Multidetector computed tomography and magnetic resonance cholangiopancreatography detected partial blockage ot interlude in the CBD leading to cholestasis without evidence of metastasis. Therefore, a diagnosis of IPNB cT1N0M0 was established. Left hepatectomy with bile duct reconstruction was performed. Pathological examination confirmed an intraepithelial neoplasia pattern without an invasive component and an R0 resection achievement. The patient was monitored carefully by regular examinations. However, at 32 mo after the operation, a 26 mm tumor in the lungs and a 12 mm lesion in the brain were detected following a suspicious elevated CA 19-9 level. Video-assisted thoracoscopic surgery of left upper lobectomy and stereotactic radiotherapy are indicated. In addition to histopathological results, a genomic profiling analysis using whole exome sequencing subsequently confirmed lung metastasis originating from bile duct cancer. CONCLUSION: This case highlights the important role of genomic profiling analysis using whole exome sequencing in identifying the origin of metastasis in patients with IPNB.

Decreased NK cell activity after partial vulvectomy in pregnant patient with vulvar intraepithelial neoplasia 3 (VIN3).

INTRODUCTION AND OBJECTIVE: The study presents the problem of immune disturbances in pregnant women with vulvar carcinoma in situ (VIN3). MATERIAL AND METHODS: NK cell and T reg activity in the study patient were analysed using flow cytometry. RESULTS: Decreased NK cell activity and but increased T reg activity were observed after vulvectomy, with subsequent successful pregnancy outcome. CONCLUSIONS: Although vulvar cancer may influence immune cell activity, this issue merits further study.

Can germ cell neoplasia in situ be diagnosed by measuring serum levels of microRNA371a-3p?

PURPOSE: Diagnosing germ cell neoplasia in situ (GCNis) can detect germ cell tumours (GCTs) at the pre-invasive stage. To date, testicular biopsy with the potential of surgical complications is the only way of safely diagnosing GCNis. Recently, microRNAs (miRs) 371-3, and miR 367 were shown to be valuable serum biomarkers of GCTs. We explored the usefulness of these candidate miRs as a marker for GCNis. METHODS: 27 patients with GCNis and no concomitant GCT were enrolled. All patients underwent measuring serum levels of miR-371a-3p and miR-367-3p before treatment, 11 had repeat measurement after treatment, 2 also had testicular vein blood examinations. Serum levels were measured by quantitative PCR. In addition, four orchiectomy specimens of patients with GCT were examined immunohistochemically and by in situ hybridization (ISH) with a probe specific for miR-371a-3p to look for the presence of this miR in GCNis cells. RESULTS: The median serum level of miR-371a-3p was significantly higher in patients with GCNis than in controls, miR-367 levels were not elevated. Overall, 14 patients (51.9%) had elevated serum levels of miR-371a-3p. The highest levels were found in patients with bilateral GCNis. Levels in testicular vein serum were elevated in both of the cases. After treatment, all elevated levels dropped to normal. In two orchiectomy specimens, miR-371a-3p was detected by ISH in GCNis cells. CONCLUSIONS: Measuring miR-371a-3p serum levels can replace control biopsies after treatment of GCNis. In addition, the test can guide clinical decision making regarding the need of testicular biopsy in cases suspicious of GCNis.

A Noninvasive Blood-based Combinatorial Proteomic Biomarker Assay to Detect Breast Cancer in Women Under the Age of 50 Years.

BACKGROUND: Despite significant advances in breast imaging, the ability to detect breast cancer (BC) remains a challenge. To address the unmet needs of the current BC detection paradigm, 2 prospective clinical trials were conducted to develop a blood-based combinatorial proteomic biomarker assay (Videssa Breast) to accurately detect BC and reduce false positives (FPs) from suspicious imaging findings. PATIENTS AND METHODS: Provista-001 and Provista-002 (cohort one) enrolled Breast Imaging Reporting and Data System 3 or 4 women aged under 50 years. Serum was evaluated for 11 serum protein biomarkers and 33 tumor-associated autoantibodies. Individual biomarker expression, demographics, and clinical characteristics data from Provista-001 were combined to develop a logistic regression model to detect BC. The performance was tested using Provista-002 cohort one (validation set). RESULTS: The training model had a sensitivity and specificity of 92.3% and 85.3% (BC prevalence, 7.7%), respectively. In the validation set (BC prevalence, 2.9%), the sensitivity and specificity were 66.7% and 81.5%, respectively. The negative predictive value was high in both sets (99.3% and 98.8%, respectively). Videssa Breast performance in the combined training and validation set was 99.1% negative predictive value, 87.5% sensitivity, 83.8% specificity, and 25.2% positive predictive value (BC prevalence, 5.87%). Overall, imaging resulted in 341 participants receiving follow-up procedures to detect 30 cancers (90.6% FP rate). Videssa Breast would have recommended 111 participants for follow-up, a 67% reduction in FPs (P < .00001). CONCLUSIONS: Videssa Breast can effectively detect BC when used in conjunction with imaging and can substantially reduce unnecessary medical procedures, as well as provide assurance to women that they likely do not have BC.

Normalizers for microRNA quantification in plasma of patients with vulvar intraepithelial neoplasia lesions and vulvar carcinoma.

The role of circulating microRNAs as a promising tool for diagnosing cancer and monitoring anticancer therapies has been widely studied in the past decades. To date, no suitable reference microRNAs for normalizing quantitative real-time polymerase chain reaction assays has been identified in vulvar intraepithelial neoplasia lesions and vulvar squamous cell carcinoma. The purpose of this study was to select appropriate references for gene expression studies in plasma of patients with these lesions. Expression levels of six microRNAs-hsa-miR-425-5p, hsa-miR-191-5p, hsa-miR-93-5p, hsa-miR-423-5p, hsa-miR-103a-3p, and hsa-miR-16-5p-were analyzed by quantitative reverse transcription polymerase chain reaction in plasma samples obtained from 17 patients with vulvar intraepithelial neoplasia lesion and 27 patients with vulvar squamous cell carcinoma. The expression stability of these candidate normalizers was assayed using geNorm algorithm. hsa-miR-93-5p was revealed as the most stably expressed reference in plasma samples of both vulvar intraepithelial neoplasia lesion and vulvar squamous cell carcinoma patients. The results pointed at hsa-miR-93-5p and hsa-miR-425-5p as microRNAs that retained the greatest robustness in plasma of vulvar intraepithelial neoplasia lesion and vulvar squamous cell carcinoma patients, respectively. Our work is the first report on reference microRNA selection for quantitative real-time polymerase chain reaction applications in vulvar intraepithelial neoplasia lesion and vulvar squamous cell carcinoma. The candidate microRNA stability values for the two types of lesions are provided and might serve for normalization of the future novel microRNA biomarkers in these rare entities.

Seroprevalence of Cutaneous Human Papillomaviruses and the Risk of External Genital Lesions in Men: A Nested Case-Control Study.

BACKGROUND: A variety of cutaneous human papillomaviruses (HPV) are detectable in genital epithelial lesions in men and non-melanoma skin cancer patients. It remains unclear whether these viruses are associated causally with skin lesions. To date, no study has prospectively examined the association between cutaneous HPV seropositivity and development of external genital lesions (EGLs) in men. OBJECTIVES: To examine the association between seropositivity to cutaneous HPV types and the risk of subsequent development of EGLs. METHODS: A nested case-control study including 163 incident EGL cases and 352 EGL-free controls in the HPV Infection in Men (HIM) Study cohort was conducted. Cases were ascertained at each of up to 10 biannual clinical visits and verified through biopsy and pathological diagnoses. EGLs were categorized as condyloma, suggestive of condyloma, penile intraepithelial neoplasia (PeIN), and other EGLs. Archived serum specimens collected at baseline were tested for antibodies against 14 cutaneous HPV types (ß types (5, 8, 12, 14, 17, 22, 23, 24, 38, and 47), α type 27, γ type 4, µ type 1, and ν type 41) using a GST L1-based multiplex serology assay. Socio-demographic and sexual behavior data were collected through a questionnaire. Using logistic regression, adjusted odds ratios (AOR) and 95% confidence intervals (CI) were estimated. RESULTS: Overall, seropositivity to ≥1 cutaneous HPV type (any-HPV) and ≥1 ß types (any-ß) was 58.3% and 37.5% among other EGL cases, 71.6% and 46.8% among condyloma, 66.8% and 50.0% among PeIN, and 71.9% and 38.4% among controls, respectively. Type-specific seropositivity was most common for ɤ-HPV 4, µ-HPV 1, and ß-HPV 8. No statistically significant association was observed between any-HPV, any-ß, and type-specific HPV seropositivity and subsequent development of EGLs across all pathological diagnoses. CONCLUSIONS: Overall, seropositivity to cutaneous HPV was common among men; however, it appears that cutaneous HPV is not associated with the development of genital lesions in men.

Metabonomic changes from pancreatic intraepithelial neoplasia to pancreatic ductal adenocarcinoma in tissues from rats.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most malignant tumors and is difficult to diagnose in the early phase. This study was aimed at obtaining the metabolic profiles and characteristic metabolites of pancreatic intraepithelial neoplasia (PanIN) and PDAC tissues from Sprague-Dawley (SD) rats to establish metabonomic methods used in the early diagnosis of PDAC. In the present study, the animal models were established by embedding 7,12-dimethylbenzanthracene (DMBA) in the pancreas of SD rats to obtain PanIN and PDAC tissues. After the preprocessing of tissues, (1) H nuclear magnetic resonance (NMR) spectroscopy combined with multivariate and univariate statistical analysis was applied to identify the potential metabolic signatures and the corresponding metabolic pathways. Pattern recognition models were successfully established and differential metabolites, including glucose, amino acids, carboxylic acids and coenzymes, were screened out. Compared with the control, the trends in the variation of several metabolites were similar in both PanIN and PDAC. Kynurenate and methionine levels were elevated in PanIN but decreased in PDAC, thus, could served as biomarkers to distinguish PanIN from PDAC. Our results suggest that NMR-based techniques combined with multivariate statistical analysis can distinguish the metabolic differences among PanIN, PDAC and normal tissues, and, therefore, present a promising approach for physiopathologic metabolism investigations and early diagnoses of PDAC.

Assessment of Cervicovaginal Cancer Antigen 125 Levels: A Preliminary Study for Endometrial Cancer Screening.

AIM: We primarily aimed to compare the levels of serum and cervicovaginal cancer antigen 125 (CA 125) in women with and without endometrial carcinoma in order to reveal whether cervicovaginal CA 125 could be used as a non-invasive method. METHODS: A preliminary case-control study was designed. The study group consisted of patients who were operated for endometrial adenocarcinoma or endometrial intraepithelial neoplasia. The control group consisted of patients who underwent surgery for benign gynecological diseases. Serum and cervicovaginal secretions were immediately collected before surgery to compare levels of CA 125. RESULTS: The mean cervicovaginal CA 125 levels in patients with endometrial cancer and controls were 1,598.1 ± 1,691.1 versus 947.0 ± 1,282.7 U/ml, respectively (p = 0.016). Whereas area under receiver operating characteristic curve was 0.62 for serum CA 125, it was 0.68 for cervicovaginal CA 125. The optimal threshold of CA 125 in cervicovaginal secretion was calculated to be 575 U/ml, which detected endometrial precancer or cancer with sensitivity of 78% and specificity of 57%. The positive and negative predictive values for this threshold were 38.7 and 88.2%, respectively. CONCLUSION: In conclusion, detection of CA 125 in cervicovaginal secretion has a potential role for the non-invasive screening of endometrial precancers and cancers.

Polychlorinated biphenyls and their association with survival following breast cancer.

BACKGROUND: Polychlorinated biphenyls (PCBs) are hypothesised to influence breast carcinogenesis due to their persistence and potential to induce oestrogenic and anti-oestrogenic effects. Whether PCBs influence survival following breast cancer is unknown. METHODS: A population-based cohort of women diagnosed with first primary invasive or in situ breast cancer in 1996-1997 and with blood-measured PCBs (n=627) collected shortly after diagnosis was followed for vital status through 2011. After 5 and 15 years, we identified 54 and 187 deaths, respectively, of which 36 and 74 were breast cancer related. Using Cox regression, we estimated hazard ratios (HRs) and 95% confidence intervals (CIs) for mortality for baseline PCB concentrations, individually and as oestrogenic (ΣGroup 1B: PCB101, PCB174, PCB177, PCB187, and PCB199), anti-oestrogenic (ΣGroup 2A: PCB66, PCB74, PCB105, and PCB118; ΣGroup 2B: PCB138 and PCB170), and cytochrome P450 enzyme-inducing (ΣGroup 3: PCB99, PCB153, PCB180, PCB183, and PCB203) groups. RESULTS: The highest PCB174 tertile was associated with an increase in all-cause (HR=2.22, 95% CI: 1.14-4.30) and breast cancer-specific (HR=3.15, 95% CI: 1.23-8.09) mortalities within 5 years of diagnosis and remained associated with breast cancer-specific mortality (HR=1.88, 95% CI: 1.05-3.36) at 15 years. At 5 years, the highest tertile of PCB177 was positively associated with all-cause mortality (HR=2.12, 95% CI: 1.05-4.30). At 15 years, the highest tertiles of ΣGroup 2A congeners and PCB118 were inversely associated with all-cause mortality (HR=0.60, 95% CI: 0.39-0.83; HR=0.63, 95% CI: 0.43-0.92, respectively). CONCLUSIONS: In this first US study of PCBs and breast cancer survival, PCBs were associated with mortality in biologically plausible directions. The investigation of other, structurally similar, chemicals may be warranted.

ProEx™ C is a useful ancillary study for grading anal intraepithelial neoplasia alone and in combination with other biomarkers.

Anal intraepithelial neoplasia (AIN) is a precursor to invasive anal squamous cell carcinoma. Histologic evaluation is hampered by intra- and interobserver variability. Various biomarkers have been investigated to improve the accuracy and reproducibility of diagnosis and grading, but interpretation can be challenging. ProEx™ C is an antibody cocktail for proteins upregulated in cervical intraepithelial neoplasia. This study investigated ProEx™ C's role alone and with p16 and Ki-67 in the diagnosis and grading of AIN. Sixty-seven anal tissue samples (22 AIN I, 25 AIN II/III, and 20 non-dysplastic) were stained for ProEx™ C, Ki-67, and p16. Staining patterns were recorded and correlated with morphologic diagnoses. Considering AIN II/III vs I, full-thickness ProEx™ C staining was more frequent in AIN II/III (p = 0.0373), and showed the highest sensitivity of the biomarkers. In combination with Ki-67, sensitivity was lower, but specificity for AIN II/III rose to 83%. For differentiating non-dysplasia from AIN I, negative ProEx™ C staining correlated with non-dysplasia (p < 0.0001) and had the highest sensitivity (90%). In combination with Ki-67, sensitivity dropped to 80%, but specificity was high (96%). ProEx™ C is useful for diagnosing and grading AIN, performing as well or better than other markers at identifying AIN II/III and non-dysplastic epithelium.

Pancreatic preneoplastic lesions plasma signatures and biomarkers based on proteome profiling of mouse models.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies with a mortality that is almost identical to incidence. Because early detected PDAC is potentially curable, blood-based biomarkers that could detect currently developing neoplasia would improve patient survival and management. PDAC develops from pancreatic intraepithelial neoplasia (PanIN) lesions, graded from low grade (PanIN1) to high grade (PanIN3). We made the hypothesis that specific proteomic signatures from each precancerous stage exist and are detectable in plasma. METHODS: We explored the peptide profiles of microdissected PanIN cells and of plasma samples corresponding to the different PanIN grade from genetically engineered mouse models of PDAC using capillary electrophoresis coupled to mass spectrometry (CE-MS) and Chip-MS/MS. RESULTS: We successfully characterised differential peptides profiles from PanIN microdissected cells. We found that plasma from tumor-bearing mice and age-matched controls exhibit discriminative peptide signatures. We also determined plasma peptide signatures corresponding to low- and high-grade precancerous step present in the mice pancreas using the two mass spectrometry technologies. Importantly, we identified biomarkers specific of PanIN3. CONCLUSIONS: We demonstrate that benign and advanced PanIN lesions display distinct plasma peptide patterns. This strongly supports the perspectives of developing a non-invasive screening test for prediction and early detection of PDAC.

The existence of Th22, pure Th17 and Th1 cells in CIN and Cervical Cancer along with their frequency variation in different stages of cervical cancer.

BACKGROUND: Recently, it is found that T-helper (Th) 22 cells are involved in different types of autoimmune and tumor diseases. But, till now, no study has been carried out to understand the involvement of these cells in cervical cancer (CC). METHODS: Flow cytometry was used to determine the expression of interferon gamma (IFN-γ), Interleukin-22 (IL-22), IL-17 in the peripheral blood of healthy controls (HC), CIN and cervical cancer patients. From peripheral blood mononuclear cells (PBMCs), mRNA expression levels of Aryl hydrocarbon receptor (AHR), RAR-related orphan receptor C (RORC), TNF-α and IL-6 were respectively determined. Using the method of ELISA, plasma concentrations of IL-22, IL-17 and TNF-α were examined. RESULTS: Th22 and Th17 cells were elevated in CC and CIN patients. Th1 cells and the plasma concentrations of IL-22 in CC patients were significantly increased compared with HC. In CC patients, an increased prevalence of Th22 cells was associated with lymph node metastases. There was a positive correlation between Th22 and Th17 cells, but an approximately negative correlation between Th22 and Th1 cells in CC patients. The mRNA expression of RORC, TNF-α and IL-6 was significantly high in CC patients. CONCLUSIONS: Our results indicate that there is a higher circulatory frequency of Th22, Th17 and Th1 cells in CC which may conjointly participate in the pathogenesis and growth of CC.

Natural killer (NK) cell profiles in blood and tumour in women with large and locally advanced breast cancer (LLABC) and their contribution to a pathological complete response (PCR) in the tumour following neoadjuvant chemotherapy (NAC): differential restoration of blood profiles by NAC and surgery.

BACKGROUND: NK cells contribute to tumour surveillance, inhibition of growth and dissemination by cytotoxicity, secretion of cytokines and interaction with immune cells. Their precise role in human breast cancer is unclear and the effect of therapy poorly studied. The purpose of our study was to characterise NK cells in women with large (≥3 cm) and locally advanced (T3-4, N1-2, M0) breast cancers (LLABCs) undergoing neoadjuvant chemotherapy (NAC) and surgery, and to ascertain their possible contribution to a pathological complete response (pCR). METHODS: Women with LLABCs (n = 25) and healthy female donors [HFDs (n = 10)] were studied. Pathological responses in the breast were assessed using established criteria. Blood samples were collected pre and post NAC and surgery. Flow cytometry and labelled monoclonal antibodies established absolute numbers (AbNs) and percentages (%) of NK cells, and expressing granzyme B/perforin and NKG2D. In vitro NK cytotoxicity was assessed and NK cells and cytokines (IL-2, INF-γ, TGF-ß) documented in tumours using immunohistochemical techniques. Data was analysed by SPSS. RESULTS: Women with LLABCs had significantly reduced AbNs (160.00 ± 40.00 cells/µl) but not % of NK cells, compared with HFDs (NK: 266.78 ± 55.00 cells/µl; p = 0.020). NAC enhanced the AbN (p = 0.001) and % (p = 0.006) of NK cells in patients with good pathological responses. Granzyme B(+)/perforin(+) cells were significantly reduced (43.41 ± 4.00%), compared with HFDs (60.26 ± 7.00%; p = 0.003). NAC increased the % in good (p = 0.006) and poor (p = 0.005) pathological responders. Pretreatment NK cytotoxicity was significantly reduced in good (37.80 ± 8.05%) and poor (22.80 ± 7.97%) responders (p = 0.001) but remained unchanged following NAC. NK-NKG2D(+) cells were unaltered and unaffected by NAC; NKG2D expression was increased in patients with a pCR (p = 0.001). Surgery following NAC was not beneficial, except in those with a pCR. Tumour-infiltrating NK cells were infrequent but increased peritumourally (p = 0.005) showing a significant correlation (p = 0.004) between CD56(+) cells and grade of response. Tumour cytokines had no effect. CONCLUSION: Women with LLABCs have inhibited blood innate immunity, variably reversed by NAC (especially with tumour pCRs), which returned to pretreatment levels following surgery. These and in situ tumour findings suggest a role for NK cells in NAC-induced breast pCR.

Circulating prolactin and in situ breast cancer risk in the European EPIC cohort: a case-control study.

INTRODUCTION: The relationship between circulating prolactin and invasive breast cancer has been investigated previously, but the association between prolactin levels and in situ breast cancer risk has received less attention. METHODS: We analysed the relationship between pre-diagnostic prolactin levels and the risk of in situ breast cancer overall, and by menopausal status and use of postmenopausal hormone therapy (HT) at blood donation. Conditional logistic regression was used to assess this association in a case-control study nested within the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort, including 307 in situ breast cancer cases and their matched control subjects. RESULTS: We found a significant positive association between higher circulating prolactin levels and risk of in situ breast cancer among all women [pre-and postmenopausal combined, ORlog2=1.35 (95% CI 1.04-1.76), Ptrend=0.03]. No statistically significant heterogeneity was found between prolactin levels and in situ cancer risk by menopausal status (Phet=0.98) or baseline HT use (Phet=0.20), although the observed association was more pronounced among postmenopausal women using HT compared to non-users (Ptrend=0.06 vs Ptrend=0.35). In subgroup analyses, the observed positive association was strongest in women diagnosed with in situ breast tumors<4 years compared to ≥4 years after blood donation (Ptrend=0.01 vs Ptrend=0.63; Phet=0.04) and among nulliparous women compared to parous women (Ptrend=0.03 vs Ptrend=0.15; Phet=0.07). CONCLUSIONS: Our data extends prior research linking prolactin and invasive breast cancer to the outcome of in situ breast tumours and shows that higher circulating prolactin is associated with increased risk of in situ breast cancer.

Sports-induced blood sugar utilization prevents development of pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a malignant tumor of extremely high lethality in humans. Pancreatic intraepithelial neoplasia (PanIN) is the predominant precancerous lesion for PDAC and is frequently detected in the normal and inflamed pancreas. However, only a few of PanIN eventually progress into PDAC. Thus, understanding of the regulation of PanIN-to-PDAC conversion appears to be critical for prevention of the occurrence of PDAC. Here, we evaluated the effect of sports on the progression of PanIN into PDAC in an established mouse PDAC model (Ptf1a-Cre; K-ras fx/fx). We found that swimming (3 min twice per day) since 12 weeks of age significantly decreased the incidence of the development of PDAC in these PanIN-baring mice at 24 weeks of age. Moreover, swimming significantly decreased fasting blood sugar and improved glucose response in these mice, compared to the control. Furthermore, implantation of insulin pellets into the mice not only reduced fasting blood sugar and improved glucose response, but also significantly reduced the incidence of development of PDAC, which mimicked the effect of swimming. Taken together, our study suggests that sports-induced blood sugar utilization may prevent development of PDAC.