Objectives Microsatellite instability (MSI) induction by alkylating agent-based chemotherapy (ACHT) may underlie both tumor resistance to chemotherapy and secondary leukaemias in cancer patients. We investigated if ACHT could induce MSI in tumor-derived plasma-circulating DNA (pfDNA) and in normal peripheral blood mononuclear (PBMN) cells. We also evaluated if amifostine could interfere with this process in an in-vitro model. Methods MSI was determined in pfDNA, PBMN cells and urine cell-free DNA (ufDNA) of 33 breast cancer patients before and after ACHT. MCF-7 cells and PBMN from normal donors were exposed in vitro to melphalan, with or without amifostine. ResultsWe observed at least one MSI event in PBMN cells, pfDNA or ufDNA of 87, 80 and 80% of patients, respectively. In vitro, melphalan induced MSI in both MCF-7 and normal PBMN cells. In PBMN cells, ACHT-induced MSI occurred together with a significant decrease in the expression of the DNA mismatch repair gene hMSH2. Amifostine decreased hMSH2 expression and also prevented MSI induction only in normal PBMN cells. Conclusions ACHT induced MSI in PBMN cells and in tumour-derived pfDNA. Because of its protective effect against ACHT induction of MSI in normal PBMN cells in vitro, amifostine may be a potential agent for preventing secondary leukaemias in patients exposed to ACHT.

Breast cancer (BC) is the second most common cancer worldwide and the first among women. If early diagnosed and treated, this disease has a good prognosis. However, it is believed that 90 % of all patients who have had cancer died due to metastatic disease, which highlights the need for a marker which allows the detection of latent cancer cells spread from the primary tumor. The objective of this study was to investigate the expression of survinin in peripheral blood of patients with breast cancer at diagnosis and during chemotherapy aiming correlation with minimal residual disease, clinical and pathological findings. The study included 40 patients with breast cancer and 12 healthy donors as a comparison group. Survinin expression was verified by real-time PCR. For diagnosis, survinin expression cutoff point was 1.05; considering this cutoff point, we obtained a test sensitivity of 85.3 %, specificity of 75.0 %, positive predictive value of 90.6 %, negative predictive value of 64.3 %, and accuracy of 82.6 %. There was statistical significance between groups (patients × control group), presenting to patients a significantly higher value than the control group (p < 0.001). Patients that presented at the diagnosis a survinin gene expression ≥ 1.05 are 17 times more likely to develop metastatic disease.

BackgroundSeveral studies seek biological markers that give diagnostic and degree of tumor development. The aim of this study was to validate the determination of plasma DNA using nanotechnology (Nanovue™-NV) in samples of 80 patients with prostate cancer.MethodsBlood samples of 80 patients of the Urology Ambulatory of Faculdade de Medicina do ABC with prostate cancer confirmed by anatomical-pathology criteria were analyzed. DNA extraction was performed using a GFX TM kit (Amersham Pharmacia Biotech, Inc, USA) following the adapted protocol. Plasma was subjected to centrifugation.ResultsThere was a big difference between the first and the second value obtained by NanoVue Only two samples had no differences between duplicates. Maximum difference between duplicates was 38 μg/mL. Average variation between 51 samples was 10.29 μg/mL, although 21 samples had differences above this average. No correlation was observed between pDNA obtained by traditional spectrophotometry and by nanotechnology.ConclusionDetermination of plasma DNA by nanotechnology was not reproducible.

<strong><strong>Objectives:</strong> </strong>The study of the involvement of non-neoplastic cells with genomic instability has not been sufficiently investigated. Genomic instability induced by treatment chemotherapy with alkylating agents’ employment has been reported in different biological matrices like PBMN, and fpDNA fuDNA. We investigated the possible use of DNA from oral mucosal cells to observe the presence of genomic instability being a simple protocol, applicable and non - invasive.<br /> <strong>Methods: </strong>Genomic instability was determined in oral mucosal cells of 31 women diagnosed with breast cancer before and after chemotherapy with alkylating agents’ presence. MSI was assessed by a panel with five different microsatellite regions.<br /> <strong>Results:</strong> We observed that 77.41% of patients had any genetic alteration in the oral mucosal cells, with a higher number of MSI events by 32.58% compared to LOH events by 24.97%, mainly in the FMR2 (16.29 %) and BAT 26 (13.04%). The control group did not show genomic instability in oral mucosal cells.<br /> <strong>Conclusions:</strong> The oral mucosal cells are susceptible to genomic instability when exposed to chemotherapy regimens containing alkylating agents which allows us a new approach to chemotherapy regimens and their implications and propose a new biological matrix for assessing such adverse effects.<br /> <br /><strong><br /></strong>

e11505 Background: We have previously shown that alkylating agent based chemotherapy regimens (AQT) could induce MIS in the PBMNF of BC patients in parallel to a decrease in the expression of the protein hMSH2 in these cells (Fonseca et al., 2005, Breast Cancer Res, 7, R28–32). Since plasma DNA derives mainly from tumor cells, we wanted to know if chemotherapy would also produce MIS in tumor DNA and if this phenomenon could be reproduced in vitro. Methods: 33 previously untreated female BC patients with a mean age of 51 years received AQT(16 ACT;3FAC;2 TAC;1FEC;10AC). Samples from 3 additional patients who received Fulvestrant only as neoadjuvant therapy were also included. Blood (pfDNA and PBMNNF) and urine (ufDNA) were evaluated at time 0,3 and 6 months with 6 MIS markers (BAT40,BAT26, MR2,TP53 PCR15.1, APC and ALU). Levels of fpDNA and fuDNA were measured by spectrophotometry. We incubated in vitro cultures of MCF- 7 cells and PBMNF cells with M at a dose of 0.7μg/ml for 30 minutes with and without A at 20% of the M dose and evaluated serially for 48 hours for MIS and hMH2 expression by immunohistochemistry. Results: We observed at least one MIS event in the PBMNF, fpDNA or fuDNA in 87%, 80% and 80% of the patients, respectively, mainly in BAT40 and BAT 26 markers. There was only 14.74% of concordance of MIS alterations between PBMNF and fpDNA and 8. 42% between fpDNA and fuDNA. Patients receiving Hormones also exhibited MIS. Interestingly, fpDNA levels increased significantly in patients with measurable disease who responded to therapy (47.4 ± 13.34 vs 14.37± 5.32; p = 0.021). In vitro, incubating MCF-7 cells and normal PBMNF cells with M ±A, we observed that we could induce MIS in both MCF-7 cells and normal PBMNF cells but A prevented MIS only in normal PBMNF cells. In normal PBMNF cells without A that sustained MIS there was a significantly decreased percentage of cells expressing hMSH2 ( 96% vs 57% p < 0.001). Conclusions: We conclude that Chemotherapy as well as Fulvestran can induce MIS in normal and malignant cells and that in vitro these effects could be reproduced by treatment with M and prevented in normal cells by A. No significant financial relationships to disclose.