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Gusella M.,Laboratory of Pharmacology and Molecular Biology
Pharmacogenomics Journal | Year: 2016

Candidate genes involved in DNA repair, 5-fluorouracil metabolism and drug detoxification were genotyped in 124 patients receiving neoadjuvant chemoradiation treatment for locally advanced esophageal cancer and their predictive role for long-term relapse-free survival (RFS) and cancer-specific survival (CSS) were evaluated. A panel including MTHFR 677TT, MDR1 2677GT, GSTP1 114CC, XPC 499CC and XPC 939AC+CC, defined as high-risk genotypes, discriminated subgroups with significantly different outcomes. When the panel was combined with histology, patients split into two subsets with 5-year RFS and CSS rates of 65% vs 27% (hazard ratio (HR) 3.0, P<0.0001) and 69% vs 31% (HR 2.9, P<0.0001), respectively. Combining the 5-single-nucleotide polymorphism (5-SNP) panel with pathological response defined two major informative risk classes with 5-year PFS and CSS rates of 79.4% vs 17.7% (HR 6.71, P<0.0001) and 79.3% vs 26.3% (HR 6.25, P<0.0001), respectively. This classification achieved a sensitivity of 79%, a specificity of 85.4% and an accuracy of 81.8%.The Pharmacogenomics Journal advance online publication, 1 March 2016; doi:10.1038/tpj.2016.9. © 2016 Macmillan Publishers Limited Source

Danese E.,University of Verona | Minicozzi A.M.,Queen Mary, University of London | Benati M.,University of Verona | Montagnana M.,University of Verona | And 7 more authors.
PLoS ONE | Year: 2015

Background: Although recent advances in circulating DNA analysis allow the prediction of tumor genomes by noninvasive means, some challenges remain, which limit the widespread introduction of cfDNA in cancer diagnostics. We analyzed the status of the two best characterized colorectal cancer (CRC) genetic and epigenetic alterations in a cohort of CRC patients, and then compared the degree to which the two patterns move from tissue to plasma in order to improve our understanding of biology modulating the concordance between tissues and plasma methylation and mutation profiles. Methods: Plasma and tumor tissues were collected from 85 patients (69±14 years, 56 males). KRAS and SEPT9 status was assessed by allele refractory mutation system quantitative PCR and quantitative methylation-specific PCR, respectively. Six of the most common point mutations at codon 12 and 13 were investigated for KRAS analysis. Results: KRAS mutations and SEPT9 promoter methylation were present in 34% (29/85) and in 82% (70/85) of primary tumor tissue samples. Both genetic and epigenetic analyses of cfDNA revealed a high overall concordance and specificity compared with tumor-tissue analyses. Patients presenting with both genetic and epigenetic alterations in tissue specimens (31.8%, 27/85) were considered for further analyses. The median methylation rates in tumour tissues and plasma samples were 64.5%(12.2-99.8%) and 14.5%(0-45.5%), respectively. The median KRAS mutation load (for matched mutations) was 33.6% (1.8-86.3%) in tissues and 2.9% (0-17.3) in plasma samples. The plasma/tissue (p/t) ratio of SEPT9 methylation rate was significantly higher than the p/t ratio of KRAS mutation load, especially in early stage cancers (p=0.0108). Conclusion: The results of this study show a discrepant rate of epigenetic vs. genetic alterations moving from tissue to plasma. Many factors could affect mutation cfDNA analysis, including both presence of tumor clonal heterogeneity and strict compartmentalization of KRAS mutation profile. The present study highlights the importance of considering the nature of the alteration when analyzing tumor-derived cfDNA. © 2015 Danese et al. Source

Nicoletto M.O.,Istituto Oncologico Veneto | Palma M.D.,Istituto Oncologico Veneto | Donach M.E.,New York University | Gusella M.,Laboratory of Pharmacology and Molecular Biology | And 11 more authors.
Tumori | Year: 2010

Aims and background. To assess feasibility and toxicity of intraperitoneal administration of cisplatin and paclitaxel, followed by intravenous chemotherapy in pretreated patients with suboptimal ovarian cancer (residuum >1 cm) or primary peritoneal tumor, and suffering from ascites and/or intestinal obstruction. Methods. Fourteen relapsed ovarian cancer patients, 5 of whom were platinum sensitive (platinum-free interval >6 mo), 7 platinum-resistant (platinum-free interval >6 mo), and 2 platinum-refractory, received one cycle of intraperitoneal cisplatin, 100 mg/m2 on day 1, and two cycles of intraperitoneal paclitaxel, 120 mg/m2 on days 8 and 14. Intravenous chemotherapy was administrated 4 weeks following the last intraperitoneal paclitaxel instillation. Blood and peritoneal fluid samples were harvested at 0, 1, 4 and 24 h after ending paclitaxel delivery to guarantee proper tumor exposure and patient safety. Results. Intraperitoneal cisplatin determined 6 cases of vomiting grade 1-2 (40% of the morbidity). Intraperitoneal paclitaxel was associated with 6 events of grade 1-2 abdominal pain; the only grade 4 toxicity was one case of neutropenia and one ofmucositis. Ascites decreased in 11 patients: the median time to first need for paracentesis was 5 months, compared to a median baseline paracentesis of 4 weeks. Three intestinal normalizations were obtained. Themedian overall survival was 10 months for our cohort of patients. Intraperitoneal paclitaxel clearance was significantly higher in patients with suboptimal tumor and symptomatic disease than in patients with smaller residual masses and without ascites (P = 0.004). Conclusions. Intraperitoneal treatment was feasible, and enhanced response to the following intravenous chemotherapy was seen in these patients. Free full text available at www.tumorionline.it. Source

Nisi A.,University of Padua | Panfili M.,University of Padua | De Rosa G.,University of Padua | Boffa G.,University of Padua | And 3 more authors.
Journal of Clinical Pharmacology | Year: 2013

Pentoxifylline (PTX) is extensively metabolized in the body, and all its 3 plasma metabolites (M1, M4, M5) are pharmacologically active. The authors evaluated the pharmacokinetics of PTX and its metabolites in 20 patients with chronic heart failure (CHF). Eleven had moderate and 9 severe CHF. The time courses of PTX, M1, M4, and M5 plasma levels were determined after oral administration of a sustained-release 600-mg tablet of PTX, and for each compound, AUC, maximal plasma concentration (Cmax), and time to Cmax (T peak ) were calculated. Compared with patients with moderate CHF, those with severe CHF showed a significant delay in T peak of PTX (3.9 vs 1.6 hours) and M5 (5.6 vs 3.6 hours), a 59% significant increase in M5 AUC, and a 56% nonsignificant increase in PTX AUC. In the whole population, the AUCs of PTX, M4, and M5 were inversely correlated with markers of liver function, whereas the AUCs of M4 and M5 were inversely correlated with the creatinine clearance. In view of the kinetic features of slow-release formulations (flip-flop phenomenon), the delay in T peak of PTX in patients with severe CHF compared with moderate CHF should be ascribed to a reduced elimination rate. © The Author(s) 2012. Source

Danese E.,University of Verona | Minicozzi A.M.,University of Verona | Benati M.,University of Verona | Montagnana M.,University of Verona | And 6 more authors.
British Journal of Cancer | Year: 2013

Background: Tumour-released DNA in blood represents a promising biomarker for cancer detection. Although epigenetic alterations such as aberrant promoter methylation represent an appealing perspective, the discordance existing between frequencies of alterations found in DNA extracted from tumour tissue and cell-free DNA (cfDNA) has challenged their practical clinical application. With the aim to explain this bias of agreement, we investigated whether protocadherin 10 (PCDH10) promoter methylation in tissue was associated with methylation pattern in matched cfDNA isolated from plasma of patients with colorectal cancer (CRC), and whether the strength of concordance may depend on levels of cfDNA, integrity index, as well as on different clinical-pathological features. Methods: A quantitative methylation-specific PCR was used to analyse a selected CpG site in the PCDH10 promoter of 67 tumour tissues, paired normal mucosae, and matched plasma samples. The cfDNA integrity index and cfDNA concentration were assessed using a real-time PCR assay.Results:The PCDH10 promoter methylation was detected in 63 out of 67 (94.0%) surgically resected colorectal tumours and in 42 out of 67 (62.7%) plasma samples. The median methylation rate in tumour tissues and plasma samples was 43.5% (6.3-97.8%) and 5.9% (0-80.9%), respectively. There was a significant correlation between PCDH10 methylation in cfDNA and tumour tissue in patients with early CRC (P<0.0001). The ratio between plasma and tissue methylation rate increases with increasing cfDNA integrity index in early-stage cancers (P=0.0299) and with absolute cfDNA concentration in advanced cancers (P=0.0234). Conclusion: Our findings provide new insight into biological aspects modulating the concordance between tissues and plasma methylation profiles. © 2013 Cancer Research UK. All rights reserved. Source

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