Patrícia Koehler-Santos

Caracterização do germoplasma de tangerineiras do Sul do Brasil mediante análises morfológicas e moleculares

Abstract – 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.

BDNF/TrkB Content and Interaction with Gastrin-Releasing Peptide Receptor Blockade in Colorectal Cancer

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.

Prevalence of the BRCA1 founder mutation c.5266dupin Brazilian individuals at-risk for the hereditary breast and ovarian cancer syndrome

About 5-10% of breast and ovarian carcinomas are hereditary and most of these result from germline mutations in the BRCA1 and BRCA2 genes. In women of Ashkenazi Jewish ascendance, up to 30% of breast and ovarian carcinomas may be attributable to mutations in these genes, where 3 founder mutations, c.68_69del (185delAG) and c.5266dup (5382insC) in BRCA1 and c.5946del (6174delT) in BRCA2, are commonly encountered. It has been suggested by some authors that screening for founder mutations should be undertaken in all Brazilian women with breast cancer. Thus, the goal of this study was to determine the prevalence of three founder mutations, commonly identified in Ashkenazi individuals in a sample of non-Ashkenazi cancer-affected Brazilian women with clearly defined risk factors for hereditary breast and ovarian cancer (HBOC) syndrome. Among 137 unrelated Brazilian women from HBOC families, the BRCA1 c.5266dup mutation was identified in seven individuals (5%). This prevalence is similar to that encountered in non-Ashkenazi HBOC families in other populations. However, among patients with bilateral breast cancer, the frequency of c.5266dup was significantly higher when compared to patients with unilateral breast tumors (12.1% vs 1.2%, p = 0.023). The BRCA1 c.68_69del and BRCA2 c.5946del mutations did not occur in this sample. We conclude that screening non-Ashkenazi breast cancer-affected women from the ethnically heterogeneous Brazilian populations for the BRCA1 c.68_69del and BRCA2 c.5946del is not justified, and that screening for BRCA1 c.5266dup should be considered in high risk patients, given its prevalence as a single mutation. In high-risk patients, a negative screening result should always be followed by comprehensive BRCA gene testing. The finding of a significantly higher frequency of BRCA1 c.5266dup in women with bilateral breast cancer, as well as existence of other as yet unidentified founder mutations in this population, should be further assessed in a larger well characterized high-risk cohort.

Prevalence of the TP53 p.R337H mutation in breast cancer patients in Brazil.

Germline TP53 mutations predispose individuals to multiple cancers and are associated with Li-Fraumeni/Li-Fraumeni-Like Syndromes (LFS/LFL). The founder mutation TP53 p.R337H is detected in 0.3% of the general population in southern Brazil. This mutation is associated with an increased risk of childhood adrenal cortical carcinoma (ACC) but is also common in Brazilian LFS/LFL families. Breast Cancer (BC) is one of the most common cancers diagnosed in TP53 mutation carriers. We have assessed the prevalence of p.R337H in two groups: (1) 59 BC affected women with a familial history (FH) suggestive of hereditary cancer syndrome but no LFS/LFL features; (2) 815 BC affected women unselected for cancer FH, diagnosed with BC at or before age 45 or at age 55 or older. Among group 1 and group 2 patients, 2/59 (3.4%, CI95%: 0.4%–11.7%) and 70/815 (8.6%, CI95%: 6.8%–10.7%), respectively, were p.R337H carriers in the germline. The prevalence of p.R337H was higher in women diagnosed with BC at or before age 45 (12.1%, CI95%: 9.1%–15.8%) than at age 55 or older (5.1%, CI95%: 3.2%–7.7%), p<0.001). The Brazilian founder p.R337H haplotype was detected in all carriers analysed. These results suggest that inheritance of p.R337H may significantly contribute to the high incidence of BC in Brazil, in addition to its recently demonstrated impact on the risk of childhood ACC.

Identification of patients at-risk for Lynch syndrome in a hospital-based colorectal surgery clinic.

AIM 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. METHODS 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. RESULTS 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 (P < 0.001). 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. CONCLUSION 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.

Ecological-evolutionary relationships in Passiflora alata from Rio Grande do Sul, Brazil

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 Alegres suburbs. The data presented here, together with the DNA investigations, indicate that P. alata is actively colonizing previously unoccupied areas of this region.

Frequency of the common germline MUTYH mutations p.G396D and p.Y179C in patients diagnosed with colorectal cancer in Southern Brazil

IntroductionMUTYH-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.Materials and methodsA 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.Results and conclusionsBiallelic 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.

Mutação germinativa 1100delC no gene CHEK2: estudo da frequência em famílias brasileiras com câncer de mama e cólon hereditários

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.

Pathogenesis-related proteins in Brazilian wheat genotypes: protein induction and partial gene sequencing

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 (thaumatinlike). 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.

CHEK2 1100DELC germline mutation: a frequency study in hereditary breast and colon cancer Brazilian families

This corrects the article DOI: 10.1590/S0004-28032012000400008.