Renzo Barbazza

Prevalence of Different Subtypes of Serrated Polyps and Risk of Synchronous Advanced Colorectal Neoplasia in Average-Risk Population Undergoing First-Time Colonoscopy

OBJECTIVES:A growing body of evidence indicates that patients with sessile serrated adenoma/polyp (SSA/P) and traditional serrated adenoma (TSA) are at risk for subsequent malignancy. Despite increasing knowledge on histological categorization of serrated polyps (SPs) data are lacking on the actual prevalence and the association of each SP subtype with advanced colorectal neoplasia.METHODS:We prospectively determined the prevalence of different SP subtypes and evaluate the association with synchronous advanced neoplasia in asymptomatic average-risk subjects undergoing first-time colonoscopy. All retrieved polyps were examined by two independent pathologists. Serrated lesions were classified into hyperplastic polyps (HP), SSA/P (without and with cytological dysplasia, SSA/P/DIS), and TSA, and were screened for BRAF and K-ras mutations.RESULTS:Among 258 polyps detected in 985 subjects, the proportion of SSA/P and TSA was 8.9% and 1.9% with an overall prevalence of 2.3% and 0.6%, respectively. SSA/Ps were small without significant difference in their location between proximal and distal colon; TSA were predominantly left-sided. BRAF mutation was common in SSA/Ps and K-ras mutation was present in all TSA. Independent predictors of advanced neoplasia were male sex (odds ratio (OR)=2.0, 95% confidence interval (CI) 1.0–4.0), increasing age (OR=4.5, 95% CI 1.5–13.4 for 50–69 years and OR=9.9, 95% CI 3.1–31.5 for >70 years), current smoking (OR=2.0, 95% CI 1.3–6.8), >3 tubular adenoma (OR=3.6, 95% CI 1.9–6.4), and SSA/P (OR=6.0, 95% CI 1.9–19.5).CONCLUSIONS:The substantial prevalence of BRAF-mutated SSA/P and the independent association with synchronous advanced colorectal neoplasia in asymptomatic average-risk subjects support the overall impact of the serrated pathway on colorectal cancer (CRC) risk in general population. The endoscopic characteristics of SSA/P emphasize the need of high-quality colonoscopy as a key factor for an effective CRC screening program.

Immunoglobulin heavy chain Diversity genes rearrangement pattern indicates that MALT-type gastric lymphoma B cells have undergone an antigen selection process

Gastric MALT lymphoma usually develops from chronic gastritis, the vast majority of which (>90%) is associated with Helicobacter pylori infection. We sequenced the third complementarity determining region (CDR3) of immunoglobulin heavy chain genes in 19 gastric MALT lymphoma clones to determine the pattern of variable (V), diversity (D) and joining (J) gene utilization during immunoglobulin gene rearrangement.

Management of stage II colon cancer - the use of molecular biomarkers for adjuvant therapy decision

BackgroundThere is uncertainty on the benefit of adjuvant chemotherapy in patients with stage II colorectal cancers. The aim of this study is to investigate the combined role of clinical, pathological and molecular parameters to identify those stage II patients who better benefit from adjuvant therapy.MethodsWe examined 120 stage II colon cancer patients. Of these, 60 patients received adjuvant 5-FU chemotherapy after surgery and the other 60 did not receive therapy. Immunohistochemical (IHC) analyses were performed to evaluate the expressions of Thymidylate synthetase (TYMS), TP53 (p53), β-catenin (CTNNB1) and CD8. For TYMS, its mRNA expression levels were also investigated by real time qRT-PCR. The entire case study was characterized by the presence of a defect in the MMR (mismatch repair) system, the presence of the CpG island methylator phenotype (CIMP or CIMP-High) and for the V600E mutation in the BRAF gene. At the histo-pathological level, the depth of tumour invasion, lymphovascular invasion, invasion of large veins, host lymphocytic response and tumour border configuration were recorded.ResultsThe presence of the V600E mutation in the BRAF gene was a poor prognostic factor for disease free and overall survival (DFS; hazard ratio [HR], 2.57; 95% CI: 1.03 -6.37; p = 0.04 and OS; HR, 3.68; 95% CI: 1.43-9.47; p < 0.01 respectively), independently of 5-FU treatment. Adjuvant therapy significantly improved survival in patients with high TYMS levels (p = 0.04), while patients with low TYMS had a better outcome if treated by surgery alone (DFS; HR, 6.07; 95% CI, 0.82 to 44.89; p = 0.04). In patients with a defect in the MMR system (dMMR), 5-FU therapy was associated to reduced survival (DFS; HR, 37.98; 95% CI, 1.04 to 1381.31; p = 0.04), while it was beneficial for CIMP-High associated tumours (DFS; HR, 0.17; 95% CI, 0.02 to 1.13; p = 0.05).ConclusionsPatients’ characterization according to MMR status, CIMP phenotype and TYMS mRNA expression may provide a more tailored approach for adjuvant therapy in stage II colon cancer.

The proliferation marker Ki67, but not neuroendocrine expression, is an independent factor in the prediction of prognosis of primary prostate cancer patients

Abstract Background Neuroendocrine markers, which could indicate for aggressive variants of prostate cancer and Ki67 (a well-known marker in oncology for defining tumor proliferation), have already been associated with clinical outcome in prostate cancer. The aim of this study was to investigate the prognostic value of those markers in primary prostate cancer patients. Patients and methods NSE (neuron specific enolase), ChrA (chromogranin A), Syp (Synaptophysin) and Ki67 staining were performed by immunohistochemistry. Then, the prognostic impact of their expression on overall survival was investigated in 166 primary prostate cancer patients by univariate and multivariate analyses. Results NSE, ChrA, Syp and Ki67 were positive in 50, 45, 54 and 146 out of 166 patients, respectively. In Kaplan-Meier analysis only diffuse NSE staining (negative vs diffuse, p = 0.004) and Ki67 (≤ 10% vs > 10%, p < 0.0001) were significantly associated with overall survival. Ki67 expression, but not NSE, resulted as an independent prognostic factor for overall survival in multivariate analysis. Conclusions A prognostic model incorporating Ki67 expression with clinical-pathological covariates could provide additional prognostic information. Ki67 may thus improve prediction of prostate cancer outcome based on standard clinical-pathological parameters improving prognosis and management of prostate cancer patients.

Is Human Papillomavirus Associated with Prostate Cancer Survival

The role of human papillomavirus (HPV) in prostate carcinogenesis is highly controversial: some studies suggest a positive association between HPV infection and an increased risk of prostate cancer (PCa), whereas others do not reveal any correlation. In this study, we investigated the prognostic impact of HPV infection on survival in 150 primary PCa patients. One hundred twelve (74.67%) patients had positive expression of HPV E7 protein, which was evaluated in tumour tissue by immunohistochemistry. DNA analysis on a subset of cases confirmed HPV infection and revealed the presence of genotype 16. In Kaplan-Meier analysis, HPV-positive cancer patients showed worse overall survival (OS) (median 4.59 years) compared to HPV-negative (median 8.24 years, P = 0.0381). In multivariate analysis age (P < 0.001), Gleason score (P < 0.001), nuclear grading (P = 0.002), and HPV status (P = 0.034) were independent prognostic factors for OS. In our cohort, we observed high prevalence of HPV nuclear E7 oncoprotein and an association between HPV infection and PCa survival. In the debate about the oncogenic activity of HPV in PCa, our results further confirm the need for additional studies to clarify the possible role of HPV in prostate carcinogenesis.

Evidence of multiple infectious agents in mycosis fungoides lesions

The etiology of mycosis fungoides (MF) remains to be determined. Several studies have proposed a viral etiology with controversial results. In this case-control study we investigated the presence of Epstein-Barr virus (EBV) and the debated presence of Human T-cell lymphotrophic virus I (HTLV-I) sequences, by polymerase chain reaction on nucleic acid extracts from formalin-fixed paraffin-embedded skin biopsies. Moreover, by a multivariate approach we analyzed in the same case-control study also the contribution of two previously examined pathogens: Hepatitis C virus (HCV) and Borrelia burgdorferi (Bb). Significant differences in the frequency of infectious agents in cases and controls were detected for Bb, HTLV-I and EBV. In MF patients we found the concurrent presence of two or three of these pathogen sequences in 21 out of 83 cases, but only in 1 out of 83 healthy controls. Our results suggest that the persistence of multiple infectious agents may cause a long-term antigenic stimulation contributing to the malignant transformation of T lymphocytes, especially when associated with HTLV-I like sequences. However, these infectious agents do not seem to have effects on disease progression.

Are Breast Cancer Molecular Classes Predictive of Survival in Patients with Long Follow-Up?

In this study we investigate the clinical outcomes of 305 breast cancer (BC) patients, aged 55 years or younger, with long follow-up and according to intrinsic subtypes. The cohort included 151 lymph node negative (LN−) and 154 lymph node positive (LN+) patients. Luminal A tumors were mainly LN−, well differentiated, and of stage I; among them AR was an indicator of good prognosis. Luminal B and HER2 positive nonluminal cancers showed higher tumor grade and nodal metastases as well as higher proliferation status and stage. Among luminal tumors, those PR positive and vimentin negative showed a longer survival. HER2-positive nonluminal and TN patients showed a poorer outcome, with BC-specific death mostly occurring within 5 and 10 years. Only luminal tumor patients underwent BC death over 10 years. When patients were divided in to LN− and LN+ no differences in survival were observed in the luminal subgroups. LN− patients have good survival even after 20 years of follow-up (about 75%), while for LN+ patients survival at 20 years (around 40%) was comparable to HER2-positive nonluminal and TN groups. In conclusion, in our experience ER-positive breast tumors are better divided by classical clinical stage than molecular classification, and they need longer clinical follow-up especially in cases with lymph node involvement.

Reduced expression of α-L-Fucosidase-1 (FUCA-1) predicts recurrence and shorter cancer specific survival in luminal B LN+ breast cancer patients

Background The lysosomal enzyme α-L-Fucosidase-1 (FUCA-1) catalyzes the hydrolytic cleavage of terminal fucose residues. FUCA-1 gene is down-regulated in highly aggressive and metastatic human tumors as its inactivation perturbs the fucosylation of proteins involved in cell adhesion, migration and metastases. Results Negativity to FUCA-1 was significantly related to the development of later recurrences in breast cancer patients with lymph node involvement at diagnosis. Cancer specific survival of luminal B LN+ patients was influenced by FUCA-1 expression as luminal B LN+ patients with positive expression had a longer cancer specific survival. FUCA-1 mRNA expression was inversely related to cancer stage and lymph node involvement. WB and qPCR analysis of FUCA-1 expression in breast cancer-derived cell lines confirmed an inverse relationship with tumor aggressiveness. Conclusions This study shows that, within LN+ breast cancer patients, FUCA-1 is able to identify a sub-set of non recurrent patients characterized by the positive expression of FUCA-1 and that, within luminal B LN+ patients, the expression of FUCA-1 predicts longer cancer specific survival. Methods We have analyzed FUCA-1 in 305 breast cancer patients by Immunohistochemistry (IHC), and by qPCR in breast cancer patients and in breast cancer cell lines.

Standards for Equipment and Reagents

The proper maintenance of laboratory equipment and the need for high-quality reagents were earlier recognized as core elements of Good Laboratory Practice (GLP) from international regulatory agencies. Most of the equipment and of the reagents used in molecular pathology laboratories were designed for research and successively adapted for clinical use. For these reasons, the instrumentation requires rigorous surveillance of its performance and prompt technical support; chemicals need proper storage and periodical monitoring of their quality. This chapter focuses on a collection of GLP-compliant suggestions for proper management of the basic laboratory equipment and reagents. Although relevant for an assay performance, these items are often neglected and researchers are faced with unexpected assay results or unexplainable test failures: laboratory compliance to GLP principles is indubitably demanding but allows getting rid of many confounder elements. The standards described in this chapter are based partly on applicable parts of existing guidance elements (in particular, the Laboratory Requirements as described by 42 CFR part 493 of CLIA) and partly on sound lab practices followed in the IMPACTS institutions.

Internal Quality Control (IQC) Organization

According to the OECD definition (1998), Good Laboratory Practice (GLP) generally refers to a quality system concerned with the organizational process and the conditions under which studies are planned, performed, monitored, recorded, archived, and reported. Acceptance of and compliance to this system is a prerequisite for the reliability of results and for their mutual acceptance among laboratories from all countries. Particular criticity is held by the results coming from translational research, because of their relevance in transferring knowledge into practice, from bench to bedside applications. This chapter lists a set of procedures that should be undertaken by the staff of a laboratory for continuous monitoring of the workflow and of the produced results (Internal Quality Controls, ICQ), in keeping with the most relevant international GLP guidelines. Two main branches of the IQC procedures are recognized: those related to the examined samples and those referred to the entire laboratory.