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The carcinogenesis in the oral cavity occurs as a multistep process and is often preceded by potentially malignant lesions. The main risk factors for the development of oral cancer are smoking and alcohol intake. The current challenge is to identify patients at greatest risk for the development of oral cancer using noninvasive and effective methods. The aim of this study is to evaluate the microsatellite mutations in the 9p21 locus, the cell proliferative activity, the pattern of epithelial desquamation, and the nucleus/cytoplasm ratio of exfoliated epithelial cells. Cytopathological samples were collected from 131 individuals divided into four groups: control (n = 26), alcohol-smoking (n = 32), leukoplakia (n = 38), and the oral squamous cell carcinoma group (OSCC, n = 35). From the cytological scraping, a slide was silver impregnated for Ag-stained nucleolar organizer region analysis and another slide was stained using the Papanicolaou technique. The remaining cells were used for DNA extraction, followed by PCR amplification and capillary electrophoresis. The cell proliferation velocity rate was higher in the leukoplakia and OSCC groups compared with the control group (P < 0.05). The leukoplakia group showed increased anucleated scales, whereas the nucleated superficial predominated in the control group and the parabasal cells in the OSCC group (P < 0.05). An increased nucleus/cytoplasm ratio was detected only in the OSCC group (P < 0.05). The 9p21 locus mutation frequency was higher in the alcohol-smoking and leukoplakia groups. 9p21 analysis and Ag-stained nucleolar organizer region methods are promising for the screening and monitoring of individuals at higher risk for the development of oral cancer.

Introduction. Studies have shown that T2DM is an inflammatory disease. Thus, the present study was aimed at evaluating whether diacerein could improve the metabolic and inflammatory profile among patients with T2DM under long-term treatment with glucose-lowering agents. Methods. This is a double-blind, parallel, placebo-controlled trial with 72 participants randomly assigned to diacerein 50 mg or placebo for 12 weeks. The primary endpoint was the between-group difference in change in HbA1c. Secondary endpoints included the proportion of patients achieving metabolic control [HbA1c≤7.0% (53 mmol/mol)] and change in inflammatory mediators. Results. Participants in the diacerein group had greater reductions in mean HbA1c level in comparison to placebo (−0.98; 95% CI: −2.02 to 0.05, P=0.06), independently of confounding factors. The difference in HbA1c level was −1.3 (95% CI: −2.3 to −0.4) in favor of diacerein (P=0.007) in those with P=0.9) in those with longer duration. The diacerein group had a 50% increase in the number of participants at the lowest TNF-α level (≤1.46 pg/mL). Conclusions. In patients with long-established T2DM under long-term treatment with glucose-lowering agents, diacerein improves metabolic control as measured by HbA1c level and has a favorable impact on inflammatory profile. Clinical Trial Registry. This trial is registered with Brazilian Clinical Trials Registry (ReBEC) number RBR-29j956.

X-linked adrenoleukodystrophy (X-ALD) is an inherited disease characterized by progressive inflammatory demyelization in the brain, adrenal insufficiency, and an abnormal accumulation of very long chain fatty acids (VLCFA) in tissue and body fluids. Considering that inflammation might be involved in pathophysiology of X-ALD, we aimed to investigate pro- and anti-inflammatory cytokines in plasma from three different male phenotypes (CCER, AMN, and asymptomatic individuals). Our results showed that asymptomatic patients presented increased levels of pro-inflammatory cytokines IL-1β, IL-2, IL-8, and TNF-α and the last one was also higher in AMN phenotype. Besides, asymptomatic patients presented higher levels of anti-inflammatory cytokines IL-4 and IL-10. AMN patients presented higher levels of IL-2, IL-5, and IL-4. We might hypothesize that inflammation in X-ALD is related to plasmatic VLCFA concentration, since there were positive correlations between C26:0 plasmatic levels and pro-inflammatory cytokines in asymptomatic and AMN patients and negative correlation between anti-inflammatory cytokine and C24:0/C22:0 ratio in AMN patients. The present work yields experimental evidence that there is an inflammatory imbalance associated Th1, (IL-2, IL-6, and IFN-γ), Th2 (IL-4 and IL-10), and macrophages response (TNF-α and IL-1β) in the periphery of asymptomatic and AMN patients, and there is correlation between VLCFA plasmatic levels and inflammatory mediators in X-ALD. Furthermore, we might also speculate that the increase of plasmatic cytokines in asymptomatic patients could be considered an early biomarker of brain damage and maybe also a predictor of disease progression.

Germline mutations in the TP53 gene are associated with Li-Fraumeni and Li-Fraumeni-Like Syndromes, characterized by increased predisposition to early-onset cancers. In Brazil, the prevalence of the TP53-p.R337H germline mutation is exceedingly high in the general population and in cancer-affected patients, probably as result of a founder effect. Several genotyping methods are used for the molecular diagnosis of LFS/LFL, however Sanger sequencing is still considered the gold standard. We compared performance, cost and turnaround time of Sanger sequencing, PCR-RFLP, TaqMan-PCR and HRM in the p.R337H genotyping. The performance was determined by analysis of 95 genomic DNA samples and results were 100% concordant for all methods. Sequencing was the most expensive method followed by TaqMan-PCR, PCR-RFLP and HRM. The overall cost of HRM increased with the prevalence of positive samples, since confirmatory sequencing must be performed when a sample shows an abnormal melting profile, but remained lower than all other methods when the mutation prevalence was less than 2.5%. Sequencing had the highest throughput and the longest turnaround time, while TaqMan-PCR showed the lowest turnaround and hands-on times. All methodologies studied are suitable for the detection of p.R337H and the choice will depend on the application and clinical scenario.

CONTEXT: CHEK2 encodes a cell cycle checkpoint kinase that plays an important role in the DNA damage repair pathway, activated mainly by ATM (Ataxia Telangiectasia Mutated) in response to double-stranded DNA breaks. A germline mutation in CHEK2, 1100delC, has been described as a low penetrance allele in a significant number of families with breast and colorectal cancer in certain countries and is also associated with increased risk of contralateral breast cancer in women previously affected by the disease. About 5%-10% of all breast and colorectal cancers are associated with hereditary predisposition and its recognition is of great importance for genetic counseling and cancer risk management. OBJECTIVES: Here, we have assessed the frequency of the CHEK2 1100delC mutation in the germline of 59 unrelated Brazilian individuals with clinical criteria for the hereditary breast and colorectal cancer syndrome. METHODS: A long-range PCR strategy followed by gene sequencing was used. RESULTS: The 1100delC mutation was encountered in the germline of one (1.7%) individual in this high risk cohort. This indicates that the CHEK2 1100delC is not commonly encountered in Brazilian families with multiple diagnoses of breast and colorectal cancer. CONCLUSION: These results should be confirmed in a larger series of families and further testing should be undertaken to investigate the molecular mechanisms underlying the hereditary breast and colorectal cancer phenotype.

Background: The exact contribution of TP53 germline mutations, associated with Li Fraumeni Syndrome — LFS, to the overall burden of cancer is still only partially known. Studies in Southern and Southeastern Brazil have shown that a particular mutant, TP53 p.R337H, has incomplete penetrance and may be present in a significant number of subjects (estimated frequency at the populational level of 1:300 individuals). In an exploratory approach, the aim of this study is to assess the prevalence of the TP53 p.R337H mutation in women with breast cancer diagnosed before 46 and after 55 years of age, unselected for family history of cancer and resident in Southern and Southeastern Brazil. Methods: Formalin-fixed paraffin-embedded (FFPE) non-tumoral tissue of 521 women diagnosed with breast cancer between 2000 and 2010 in two pathology laboratories were obtained retrospectively and consecutively, and analyzed after anonimization. Genomic DNA was isolated with the QIAamp DNA Tissue Kit and genotyping performed by allelic discrimination using a TaqMan assay. Confirmation of all mutation-positive and a sample of mutation-negative cases was done by TP53 exon 10 sequencing. Results: Analysis of the first 299 cases identified the TP53 p.R337H mutation in the germline of 15 (5,0%) cases: 13/142 (9,2%) before 46 years and 2/157 (1,3%) diagnosed after 55 years. The p53 expression pattern assessed by immunohistochemistry in the breast tumors was not different between p.R337H mutation carriers and non-carriers. Conclusion: Preliminary analysis in a sample of women with breast cancer in Southern Brazil indicates that the germline TP53 p.R337H founder mutation is present in a high proportion of cases, especially those diagnosed at a young age. Germline TP53 mutations are considered rare, occurring in about 1:5000 individuals of the general population. The occurrence of this founder mutation at such a high frequency in a particular geographic region has important implications for disease management and cancer risk counseling for these patients and families. This mutation likely contributes to a significant proportion of the health burden associated with breast cancer in Southern Brazil. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-09-07.

MUTYH-associated polyposis (MAP) is an autosomal recessive cancer predisposition syndrome associated with the development of colorectal tumors and colonic polyps at an early age. MAP syndrome is associated to germline biallelic mutations in the MUTYH gene which lead to deficient DNA repair through the base-excision repair system and accumulation of G:C→T:A transversions. Occurrence of such mutations in oncogenes and tumor suppressor genes drives colorectal carcinogenesis and is associated with the development of colonic polyps. Two common mutations, p.Y179C and p.G396D, are present in approximately 70-80% of MAP in European families with identified MUTYH germline mutations. The aim of this study was to assess the frequency of the germline MUTYH mutations p.Y179C and p.G396D in Brazilian patients with MAP and other hereditary colorectal cancer (CRC) phenotypes, as well as in sporadic CRC cases.A total of 75 patients were included. Samples were screened for the MUTYH germline mutations p.Y179C and p.G396D by allelic discrimination assays using allele-specific TaqMan® probes. In all mutation-positive cases, results were confirmed by sequencing.Biallelic germline MUTYH mutations were identified in 4 of 60 (6.6%) patients with a phenotype of hereditary colorectal cancer. Germline MUTYH mutation screening should be considered in the differential diagnosis of hereditary colorectal syndromes, and not only in MAP, but also in familial adenomatous polyposis and Bethesda criteria-positive families. Additional mutation screening studies of the MUTYH gene in a larger number of Brazilian patients will be necessary to confirm these results and determine the validity and applicability of MUTYH mutation screening in our population.

Background: Previous studies in Brazilian patients diagnosed with the core-cancers of Li-Fraumeni syndrome (LFS) and its variants (LFL) indicate that a particular germline mutation, TP53 p.R337H, may be highly prevalent and strongly associated with pediatric adrenocortical tumors in Southern Brazil. However, other tumours types, including breast cancer, have been observed in carriers, raising the possibility that the TP53 p.R337H mutation may also contribute to breast tumorigenesis. Breast phyllodes tumour, a very rare and aggressive breast neoplasia that accounts for less than 1% of all breast cancer cases, may be associated with LFS, and has suggested by some to be particularly frequent in TP53 mutation carriers. Methods: Formalin fixed paraffin-embedded (FFPE) tissue from disease-free lymph nodes of an unselected consecutive series of women diagnosed with benign and malignant phyllodes tumours between 2000 and 2010, was obtained from 7 Pathology Laboratories in Southern Brazil. Genomic DNA was isolated from FFPE with the QIAamp DNA Tissue Kit. Genotyping was performed by qPCR using TaqMan technology and direct bi-directional sequencing for confirmation in mutation-positive cases. Results: Analysis of the first 48 cases included in the study (9 malignant, 3 borderline and 36 benign phyllodes tumors) resulted in identification of the TP53 p.R337H mutation in the germline of a 61 year-old patient with malignant phyllodes and a 38 year-old patient with a benign phyllodes tumor of the breast. Conclusion: Preliminary analysis of a case series of breast phyllodes tumors indicates that germline the founder mutation TP53 p.R337H may be particularly common in women with these tumors in Southern Brazil. Further analysis of the entire TP53 coding region and evaluation of the founder haplotype in the mutation-positive cases is ongoing. The identification of germline TP53 mutations in patients diagnosed with phyllodes tumors of the breast has important implications for disease management and in cancer risk counseling for these patients and families. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P3-06-02.

Objective: Neurotrophin and neuropeptide pathways are emerging targets in cancer. Here we show that brain-derived neurotrophic factor (BDNF) and its receptor, TrkB, are present in colorectal cancer and that BDNF levels are increased in tumors compared to nontumor tissue. In addition, we investigate the role of BDNF in influencing the response of colorectal cancer cells to inhibition of gastrin-releasing peptide receptors (GRPR). Methods: Fresh-frozen sporadic colorectal adenocarcinoma specimens and adjacent nonneoplastic tissue from 30 patients, as well as paraffin-embedded colorectal cancer samples from 21 patients, were used in this study. Cell proliferation and mRNA and protein levels were examined in HT-29 or SW620 cells treated with a GRPR antagonist, human recombinant BDNF (hrBDNF), a Trk antagonist K252a, or cetuximab. Results: Expression of BDNF and TrkB was detected in tumor samples and cell lines. BDNF levels were higher in tumor samples compared to nonneoplastic tissue. BDNF expression and secretion were increased by GRPR blockade in HT-29 cells through a mechanism dependent on epidermal growth factor receptors. Treatment with hrBDNF prevented the effect of GRPR blockade on cell proliferation, whereas a Trk inhibitor reduced proliferation. Conclusions: BDNF and TrkB are present in colorectal cancer and might contribute to resistance to GRPR antagonists.

Lynch syndrome is an autosomal dominant cancer predisposition syndrome caused by germline mutations in one of the mismatch repair (MMR) genes: MLH1, MSH2, MSH6 and PMS2. Clinically, Lynch syndrome is characterized by early onset (45 years) of colorectal cancer (CRC), as well as extra-colonic cancer. Male and female carriers of Lynch syndrome-associated mutations have different lifetime risks for CRC and in women endometrial cancer (EC) may be the most common tumor. Whenever Amsterdam criteria are not fulfilled, the currently recommended laboratory screening strategies involve microsatellite instability testing and immunohistochemistry staining of the tumor for the major MMR proteins. The aim of this study was to estimate the frequency of MMR deficiencies in women diagnosed with EC who are at-risk for Lynch syndrome. Thirty women diagnosed with EC under the age of 50 years and/or women with EC and a first degree relative diagnosed with a Lynch syndrome-associated tumor were included. To assess MMR deficiencies four methods were used: multiplex PCR, Single Strand Conformation Polymorphism, Immunohistochemistry and Methylation Specific-Multiplex Ligation-dependent Probe Amplification. Twelve (40%) patients with EC fulfilling one of the inclusion criteria had results indicative of MMR deficiency. The identification of 5 women with clear evidence of MMR deficiency and absence of either Amsterdam or Bethesda criteria among 10 diagnosed with EC under the age of 50 years reinforces previous suggestions by some authors that these women should be considered at risk and always screened for Lynch syndrome after informed consent.

A total of 85 wild plants were collected in different areas of southern and south-eastern Brazil, and studied for nrDNA internal transcribed spacers (ITS). Thirty-two of them were also investigated for two other nuclear (G3pdh and LEAFY), as well as eight plastid systems. Sixty-one ITS sequence types were identified, in a region of 412 bp, whereas 42 G3pdh haplotypes were found in a region of 1010 bp, and 23 LEAFY haplotypes in a region of 565 bp. GC content was higher in ITS (65%) than in the two other nuclear regions (40–42%), whereas nucleotide diversity was markedly higher in G3pdh than in the other two. No variability was found in the plastid systems. Most of the nuclear molecular diversity (56–86%) occurs at the intrapopulation level, and there is clear evidence of population expansion and little geographical structure. Historical information and field studies indicate that Passiflora alata is actively colonizing previously unoccupied areas in southern Brazil. High indices of intrapopulation variability were found in the region that is being colonized, similar to those found in areas where P. alata is native. Multiple introductions, as well the species’ genetic structure and human interference, may explain these findings. © 2006 The Linnean Society of London, Biological Journal of the Linnean Society, 2006, 88, 611–630.

– The objective of this work was to characterize mandarin (Citrus spp.) germplasm fromSouthern Brazil by morphological and molecular analyses. Thirty seven cultivars from 34 distinctmandarin varieties were evaluated by morphological and agronomic traits of leaves, flowers and fruits,and by microsatellite markers. The morphological and agronomic characteristics suggested that almostall varieties can be produced for commercial use, and some, as the Satsuma variety, are recommended forbreeding programs. Pooled DNA samples from 1-5 plants belonging to each cultivar were tested. Eightof the nine primers detected polymorphisms. Specific markers were found for some accessions. Thedendrogram constructed with the morphological results divided the 37 cultivars into four groups, whilethat obtained with the microsatellites clustered 35 of the 37 cultivars into three groups only. Generally,intervarietal differences are not high, and this lack of agreement in the two multifactorial analysesindicates that diverse evolutionary factors are acting at these two levels of investigation.Index terms: citrus, genetic variation, microsatellites, breeding methods.Caracterizacao do germoplasma de tangerineiras do Sul do Brasilmediante analises morfologicas e molecularesResumo – O objetivo deste trabalho foi caracterizar o germoplasma de tangerineiras ( Citrus spp.) pormeio de analises morfologicas e moleculares. Trinta e sete cultivares, pertencentes a 34 variedadesdistintas de tangerineiras, foram avaliadas pelas caracteristicas morfologicas e agronomicas de folhas,flores e frutos, bem como por analises de microssatelites. As caracteristicas morfologicas e agronomicassugerem que praticamente todas as variedades podem ser usadas comercialmente e algumas delas, comoa variedade Satsuma, sao recomendadas para uso em cruzamentos dirigidos. O DNA foi obtido deamostras extraidas de uma a cinco plantas de cada cultivar. Oito de nove primers detectaram polimorfismos.Foram encontrados marcadores especificos para alguns acessos. O dendrograma construido com osresultados morfologicos dividiu as 37 cultivares em quatro grupos, enquanto o obtido com osmicrossatelites agrupou 35 das 37 cultivares em tres grupos. As diferencas intervarietais nao sao gran-des, e esta falta de concordância nas duas analises multivariadas indica que fatores evolutivos diversosestao agindo nesses dois niveis de investigacao.Termos para indexacao: citros, variacao genetica, microsatelites, metodo de melhoramento.

Leaves from 14 Brazilian genotypes of Triticum aestivum L. were treated with salicylic acid to induce pathogenesis-related (PR) proteins. Inter and intracellular extracts were then obtained and investigated through polyacrilamide gel electrophoresis. Seven bands were observed. Material related to two of them (of 40 and 24 kDa) occurred in intracellular spaces only. DNA from these same genotypes was then amplified through PCR using primers developed from three sequences encoding PR proteins, and compared with previously described sequences. The fragments presented homologies to PR groups 1, 3 (chitinases), and 5 (thaumatin-like). The PR3-like sequence also showed a site characteristic of PRs induced by ethylene and a portion without homology with previous sequences. No variation among genotypes were observed, either for protein extracts or DNA sequences.

To determine the prevalence of a family history suggestive of Lynch syndrome (LS) among patients with colorectal cancer (CRC) followed in a coloproctology outpatient clinic in Southern Brazil.A consecutive sample of patients with CRC were interviewed regarding personal and family histories of cancer. Clinical data and pathology features of the tumor were obtained from chart review.Of the 212 CRC patients recruited, 61 (29%) reported a family history of CRC, 45 (21.2%) were diagnosed under age 50 years and 11 (5.2%) had more than one primary CRC. Family histories consistent with Amsterdam and revised Bethesda criteria for LS were identified in 22 (10.4%) and 100 (47.2%) patients, respectively. Twenty percent of the colorectal tumors had features of the high microsatellite instability phenotype, which was associated with younger age at CRC diagnosis and with Bethesda criteria (P0.001). Only 5.3% of the patients above age 50 years had been previously submitted for CRC screening and only 4% of patients with suspected LS were referred for genetic risk assessment.A significant proportion of patients with CRC were at high risk for LS. Education and training of health care professionals are essential to ensure proper management.

The geographical distribution, ecological characteristics, flowering and fruiting times, and pollinating agents of Passiflora alata are considered and related to molecular genetic data gathered simultaneously. The first report on this species in Rio Grande do Sul was made in 1934, only in cultivated gardens. Approximately 20 years later, however, the species was already classified as efferata (wild) in Porto Alegre's suburbs. The data presented here, together with the DNA investigations, indicate that P. alata is actively colonizing previously unoccupied areas of this region. A distribuição geográfica, as características ecológicas, as épocas de florescimento e frutificação, e os agentes polinizadores de Passiflora alata são considerados e relacionados a estudos genético-moleculares desenvolvidos simultaneamente. O primeiro registro da espécie no Rio Grande do Sul foi feito em 1934, apenas em área cultivada. Cerca de 20 anos depois, no entanto, a espécie já era classificada como efferata (selvagem) nos subúrbios de Porto Alegre. Os dados aqui apresentados, junto com as investigações de DNA, indicam que P. alata está colonizando ativamente áreas previamente não ocupadas desta região.

e12546 Background: The clinical diagnosis of Lynch syndrome is usually established when a family fulfills the Amsterdam Criteria, and confirmed by identification of MMR deficiency and/or germline mutations in one of the MMR genes. In a subset of families with Amsterdam criteria, no evidence of DNA mismatch repair deficiency can be identified and this clinical entity has been designated “Familial Colorectal Cancer Type X (FCCTX)”. Methods: The prevalence of MMR deficiency was assessed in a group of 25 individuals with colorectal or endometrial cancer from 20 families fulfilling Amsterdam criteria in Southern Brazil. MMR deficiency was assessed by IHC testing using a panel of antibodies against MSH2, MLH1, MSH6, and PMS2. In cases showing loss of nuclear expression of MLH1, presence of the BRAF p.V600E mutation and microsatellite instability were assessed in the tumor. Results: Fourteen of the 20 families studied fulfilled Amsterdam II criteria. MMR deficiency was identified in the tumor of 11 index cases and in the remaining 9, nuclear expression of all four MMR proteins was normal, suggesting the diagnosis of FCCTX. This FCCTX phenotype was observed in both Amsterdam I and Amsterdam II families. CRC with MMR-deficiency was diagnosed at an earlier age (41.2 years) than CRC showing MMR proficiency (47.2 years), although the difference did not reach significance. Conclusions: FCCTX is a frequent differential diagnosis in hereditary colorectal cancer in patients presenting to high risk cancer genetics clinics in Southern Brazil and further molecular characterization of hereditary colorectal cancer families with these features is warranted.

e22154 Background: Recent studies in North American and British series have suggested an association between germline TP53 mutations and early onset HER2-positive breast cancer (BC). Mutations in the TP53 gene are estimated to occur in 1:2,000-1:5,000 individuals of the general population. Among women with BC who are unselected for family history they occur in up to 0.25%. In women with early-onset BC (A; p.R337H) has been encountered in 0.3% of the general population, in 13.3% of Li-Fraumeni-like families and in 0.5-2.4% of BC-affected women in Brazil. In an exploratory approach, the aim of this study was to investigate, in our series, if there is an association between human epidermal growth factor receptor 2 (HER2) amplification and the germline TP53 p.R337H mutation in a series of BC-affected women. Methods: A series of 718 Brazilian BC-affected women was genotyped in our previous study and 64 (8.9%) were mutation carriers. These cases were recruited from 3 centers and HER2 analysis was performed by immunohistochemistry (IHC) in each center according to validated protocols. Statistical analysis was done using SPSS software. Results: In 47 of 64 mutation carriers (73.5%), BC showed HER expression (either 1+, 2+ or 3+) while among 654 non-carriers, this immunophenotype was observed in 320 (48.9%) (p

Introduction Lynch syndrome (LS) is an autosomal dominant cancer predisposition syndrome caused by germline mutations in one of the genes involved in mismatch repair (MMR): MLH1, MSH2, MSH6 and PMS2. It is the most common hereditary colorectal cancer (CRC) syndrome accounting for approximately 3-5% of all CRC diagnoses. Clinically, LS is characterized by early onset CRC (average age 45 years) as well as extra-colonic cancers, including endometrial, ovarian, gastric, small bowel, pancreas, hepatobiliary and brain tumors. The clinical diagnosis of LS is made when a family fulfills the Amsterdam criteria. Less strict criteria of Bethesda have been used to identify individuals at risk due to suggestive family history, early age at CRC onset or characteristic tumor histologies. MLH1 and MSH2 germline mutations account for the majority of known mutations in LS families (50-90%). To date, over 513 different DNA alterations have been reported principally in Western populations. One of the most interesting characteristics of the Brazilian population is its heterogeneity: apart from native populations, Brazil, especially Southern Brazil, has received immigrants from different countries, including an important contribution from Southern European (mainly Portuguese colonizers in the 16th century). In 1999, a novel mutation, consisting in a single base substitution of nucleotide 2152 C>T at exon 13 in MHS2, resulting in a glutamine amino acid change to a stop codon (Q718X), was describe in Portuguese Lynch families. The aim of this study is to assess the frequency of the c.2152 C>T mutation in MSH2 in Brazilian Lynch syndrome families. Methods Peripheral blood samples were obtained from 49 Lynch syndrome families after informed consent. DNA was extracted from leukocytes using the Ilustra blood genomicPrep Mini Spin kit (GE Healthcare) as described by the manufacturer. Exon 13 of the MSH2 gene was amplified by PCR and the resulting amplicons were separated on a ABI 3500 (Life Technologies) equipment. Results were analysed by the CLC Main WorkBench V6.1.1 software. Results The 2152 C>T - MHS2 mutation was identified in three (6.12%) of the 49 unrelated index cases, all of whom refer a Portuguese origin. All three families fulfilled Amsterdam criteria, and the mean of age at CRC diagnosis was 31 years. Conclusion In families meeting Amsterdam criteria, who very likely carry a germline MMR mutation, the initial genetic testing approach is often sequence analysis of the most commonly affected genes (MLH1 and MSH2), or targeted sequencing of the gene(s) whose product(s) is (are) not expressed in the colorectal tumor. The occurrence of the germline 2152 C>T - MHS2 mutation in Brazilian families at a considerable frequency, suggests that it may be a hotspot mutation in this region. Although the findings of this study should be confirmed in a larger series of patients, screening of Brazilian LS families for this mutation should be considered as an initial approach to diagnosis.

1522 Background: The exact contribution of TP53 germline mutations, associated with Li Fraumeni Syndrome, to the overall burden of cancer is still only partially known. Studies in Southern and Southeastern Brazil have shown that a specific germline mutation at codon 337 (c.1010G>A; p.R337H), has incomplete penetrance and may be present in a significant number of subjects (estimated frequency at the populational level of 1:300 individuals). In an exploratory approach, the aim of the study is to assess the frequency of the p.R337H mutation in women from different Brazilian regions, diagnosed with breast cancer (BC) before 46 and after 55 years of age, and unselected for family history of cancer. Methods: Formalin-fixed paraffin-embedded (FFPE) non-tumoral tissue (lymph nodes or normal breast) of women diagnosed with BC between 2000 and 2010 in 3 pathology laboratories from the Brazilian cities of Porto Alegre, São Paulo and Barretos were obtained retrospectively and analyzed after anonimization. Genomic DNA was isolated with standard methods and genotyping performed in duplicates by qPCR (TaqMan assay). Confirmation of all mutation-positive and a sample of mutation-negative cases were done by TP53 exon 10 sequencing or by a second independent qPCR analysis. Results: Analysis of 515 BC-affected women identified the p.R337H mutation in the germline of 70 (8,6%) cases: 49/403 (12,1%) diagnosed before 46 years and 21/412 (5,1%) diagnosed after 55 years. BC occurred earlier in p.R337H mutation carriers than in non-carriers (p=0.001). Conclusions: Preliminary analysis in a sample of women with BC indicates that p.R337H founder mutation is present in a high proportion of cases, especially those diagnosed at a young age. Regional genetic background and recruitment strategies may account for the different mutation frequencies observed in the centers of study. The occurrence of this mutation at such a high frequency in a particular geographic region has important implications for disease management and cancer risk counseling for these patients and families. This mutation likely contributes to a significant proportion of the health burden associated with BC in Brazil. Financial support: FIPE-HCPA, CAPES, FAPERGS and Glaxo Smith Kline.