Publications by authors named "Dirce Maria Carraro"

84 Publications

The mutational repertoire of uterine sarcomas and carcinosarcomas in a Brazilian cohort: A preliminary study.

Clinics (Sao Paulo) 2021 20;76:e2324. Epub 2021 Jan 20.

Laboratorio de Ginecologia Estrutural e Molecular, Disciplina de Ginecologia, Hospital das Clinicas (HCFMUSP), Faculdade de Medicina, Universidade de Sao Paulo, Sao Paulo, SP, BR.

Objectives: The present study aimed to contribute to the catalog of genetic mutations involved in the carcinogenic processes of uterine sarcomas (USs) and carcinosarcomas (UCSs), which may assist in the accurate diagnosis of, and selection of treatment regimens for, these conditions.

Methods: We performed gene-targeted next-generation sequencing (NGS) of 409 cancer-related genes in 15 US (7 uterine leiomyosarcoma [ULMS], 7 endometrial stromal sarcoma [ESS], 1 adenosarcoma [ADS]), 5 UCS, and 3 uterine leiomyoma (ULM) samples. Quality, frequency, and functional filters were applied to select putative somatic variants.

Results: Among the 23 samples evaluated in this study, 42 loss-of-function (LOF) mutations and 111 missense mutations were detected, with a total of 153 mutations. Among them, 66 mutations were observed in the Catalogue of Somatic Mutations in Cancer (COSMIC) database. TP53 (48%), ATM (22%), and PIK3CA (17%) were the most frequently mutated genes. With respect to specific tumor subtypes, ESS showed mutations in the PDE4DIP, IGTA10, and DST genes, UCS exhibited mutations in ERBB4, and ULMS showed exclusive alterations in NOTCH2 and HER2. Mutations in the KMT2A gene were observed exclusively in ULM and ULMS. In silico pathway analyses demonstrated that many genes mutated in ULMS and ESS have functions associated with the cellular response to hypoxia and cellular response to peptide hormone stimulus. In UCS and ADS, genes with most alterations have functions associated with phosphatidylinositol kinase activity and glycerophospholipid metabolic process.

Conclusion: This preliminary study observed pathogenic mutations in US and UCS samples. Further studies with a larger cohort and functional analyses will foster the development of a precision medicine-based approach for the treatment of US and UCS.
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http://dx.doi.org/10.6061/clinics/2021/e2324DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7798418PMC
January 2021

Deep Learning Predicts Underlying Features on Pathology Images with Therapeutic Relevance for Breast and Gastric Cancer.

Cancers (Basel) 2020 Dec 9;12(12). Epub 2020 Dec 9.

Laboratory of Computational Biology Bioinformatics, CIPE/A.C. Camargo Cancer Center, São Paulo 01508-010, Brazil.

DNA repair deficiency (DRD) is an important driver of carcinogenesis and an efficient target for anti-tumor therapies to improve patient survival. Thus, detection of DRD in tumors is paramount. Currently, determination of DRD in tumors is dependent on wet-lab assays. Here we describe an efficient machine learning algorithm which can predict DRD from histopathological images. The utility of this algorithm is demonstrated with data obtained from 1445 cancer patients. Our method performs rather well when trained on breast cancer specimens with homologous recombination deficiency (HRD), AUC (area under curve) = 0.80. Results for an independent breast cancer cohort achieved an AUC = 0.70. The utility of our method was further shown by considering the detection of mismatch repair deficiency (MMRD) in gastric cancer, yielding an AUC = 0.81. Our results demonstrate the capacity of our learning-base system as a low-cost tool for DRD detection.
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http://dx.doi.org/10.3390/cancers12123687DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7763049PMC
December 2020

Hepatoblastomas exhibit marked downregulation driven by promoter DNA hypermethylation.

Tumour Biol 2020 Dec;42(12):1010428320977124

Human Genome and Stem Cell Research Center, Department of Genetics and Evolutionary Biology, Institute of Biosciences, University of São Paulo, São Paulo, Brazil.

Hepatoblastomas exhibit the lowest mutational burden among pediatric tumors. We previously showed that epigenetic disruption is crucial for hepatoblastoma carcinogenesis. Our data revealed hypermethylation of nicotinamide N-methyltransferase, a highly expressed gene in adipocytes and hepatocytes. The expression pattern and the role of nicotinamide N-methyltransferase in pediatric liver tumors have not yet been explored, and this study aimed to evaluate the effect of nicotinamide N-methyltransferase hypermethylation in hepatoblastomas. We evaluated 45 hepatoblastomas and 26 non-tumoral liver samples. We examined in hepatoblastomas if the observed nicotinamide N-methyltransferase promoter hypermethylation could lead to dysregulation of expression by measuring mRNA and protein levels by real-time quantitative polymerase chain reaction, immunohistochemistry, and Western blot assays. The potential impact of nicotinamide N-methyltransferase changes was evaluated on the metabolic profile by high-resolution magic angle spinning nuclear magnetic resonance spectroscopy. Significant nicotinamide N-methyltransferase downregulation was revealed in hepatoblastomas, with two orders of magnitude lower nicotinamide N-methyltransferase expression in tumor samples and hepatoblastoma cell lines than in hepatocellular carcinoma cell lines. A specific TSS1500 CpG site (cg02094283) of nicotinamide N-methyltransferase was hypermethylated in tumors, with an inverse correlation between its methylation level and nicotinamide N-methyltransferase expression. A marked global reduction of the nicotinamide N-methyltransferase protein was validated in tumors, with strong correlation between gene and protein expression. Of note, higher nicotinamide N-methyltransferase expression was statistically associated with late hepatoblastoma diagnosis, a known clinical variable of worse prognosis. In addition, untargeted metabolomics analysis detected aberrant lipid metabolism in hepatoblastomas. Data presented here showed the first evidence that nicotinamide N-methyltransferase reduction occurs in hepatoblastomas, providing further support that the nicotinamide N-methyltransferase downregulation is a wide phenomenon in liver cancer. Furthermore, this study unraveled the role of DNA methylation in the regulation of nicotinamide N-methyltransferase expression in hepatoblastomas, in addition to evaluate the potential effect of nicotinamide N-methyltransferase reduction in the metabolism of these tumors. These preliminary findings also suggested that nicotinamide N-methyltransferase level may be a potential prognostic biomarker for hepatoblastoma.
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http://dx.doi.org/10.1177/1010428320977124DOI Listing
December 2020

Impact of BRCA1/2 Mutations on the Efficacy of Secondary Cytoreductive Surgery.

Ann Surg Oncol 2020 Nov 21. Epub 2020 Nov 21.

Department of Medical Oncology, A.C. Camargo Cancer Center, São Paulo, SP, Brazil.

Background: Phase III trials evaluating the role of secondary cytoreductive surgery (SCS) in recurrent ovarian cancer have pointed to the importance of patient selection. Two studies showed conflicting results regarding the benefit of SCS in BRCA1/2 mutation carriers. Our aim was to evaluate the impact of SCS on recurrent ovarian cancer according to BRCA1/2 status.

Methods: All patients with ovarian carcinoma with platinum-sensitive recurrent disease and tested for BRCA1/2 germline mutations were included. Cox regression and log rank test were used to evaluate the impact of SCS on progression-free survival (PFS) and the influence of BRCA1/2 mutations on the effect of SCS.

Results: 127 patients were included, 45.6% were treated with SCS and chemotherapy and 54.3% treated with chemotherapy only. Patients treated with SCS were younger, presented better performance status, had lower CA125, and had a longer platinum-free interval. In multivariate analysis SCS was associated with longer PFS (HR 0.42, 95% CI 0.25-0.72, p = 0.002). BRCA1/2 mutations were found in 35 patients (27.5%), and 11.8% of patients were treated with PARP inhibitors. Although not statistically significant, both BRCA1/2 wild type patients (PFS: 21.6 vs 18.4 months; p = 0.114) and BRCA1/2 mutation carriers (PFS: 23.1 vs 18.2 months, p = 0.193) appeared to derive benefit from SCS.

Discussion: The present study suggests a benefit of SCS irrespective of BRCA1/2 status among patients mostly not treated with PARP inhibitor. Further data on post hoc analysis from the phase III trials are warranted to confirm whether BRCA1/2 mutated patients should be selected for SCS.
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http://dx.doi.org/10.1245/s10434-020-09366-wDOI Listing
November 2020

Using Co-segregation and Loss of Heterozygosity Analysis to Define the Pathogenicity of Unclassified Variants in Hereditary Breast Cancer Patients.

Front Oncol 2020 2;10:571330. Epub 2020 Oct 2.

Molecular Oncology Research Center, Barretos Cancer Hospital, São Paulo, Brazil.

The use of gene panels introduces a new dilemma in the genetics field due to the high frequency of variants of uncertain significance (VUS). The objective of this study was to provide evidence that may help in the classification of these germline variants in terms of their clinical impact and association with the disease in question. A total of 52 unrelated women at-risk for HBOC and negative for pathogenic variants were evaluated through a gene panel comprising 14 breast and/or ovarian cancer susceptibility genes. Of the 453 germline variants identified, 15 variants (classes 3, 4, and 5) in the , and genes were evaluated via databases, co-segregation studies and loss of heterozygosity in the tumor. The co-segregation analysis allowed the establishment of an association with the presence of variants and the risk of cancer for variant c.316C>T in the gene. Four variants of uncertain significance showed loss of heterozygosity in the tumor ( c.4709T>C; c.1036C>T; c.1001A>G, and c.281T>C), which is an indication of pathogenicity. Thus, the present study provides novel evidence that favors the association of variants in moderate-risk genes with the development of hereditary breast cancer.
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http://dx.doi.org/10.3389/fonc.2020.571330DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7566163PMC
October 2020

Mutational Portrait of Lung Adenocarcinoma in Brazilian Patients: Past, Present, and Future of Molecular Profiling in the Clinic.

Front Oncol 2020 2;10:1068. Epub 2020 Jul 2.

Genomics and Molecular Biology Group, International Research Center, A.C. Camargo Cancer Center, São Paulo, Brazil.

Approximately 60% of lung adenocarcinomas (LAs) carry mutations that can guide treatment with tyrosine-kinase inhibitors (TKI) and other targeted therapies. Data on activating mutations in and other tyrosine-kinase receptor (TKR) genes in highly admixed populations, such as that of Brazil, are scarce. In this study, we comprehensively analyzed the actionable alteration profile of LA in Brazilian patients. driver mutation data were collected from a large Brazilian LA cohort covering an 8-year period of molecular testing in a single institution. Tests were performed using three distinct methods, and demographic and histopathological data were analyzed. For a subset of patients, driver mutations in , and and gene fusions involving TKR genes (before TKI treatment) and T790M (after TKI treatment) were assessed. mutations were detected in 25% of 1,316 LAs evaluated, with exon 19 deletions and exon 21 L858R TKI sensitizing mutations representing 72.5% of all mutations. Mutation rates were higher in women and non-smokers ( < 0.001). Next-generation sequencing was very sensitive, with a lower rate of inconclusive results compared with Sanger sequencing and pyrosequencing. hotspot gene panels were applied in 495 LA cases and detected oncogenic mutations in 51.3% of samples, most frequently in (22.4%) and (26.9%). In subgroups of 36 and 35 patients, gene fusions were detected in 11.1% of tumors and T790M resistance mutations were detected in 59% of plasma samples from patients previously treated with TKI, respectively. This report provides the first comprehensive actionable alteration portrait of LA in Brazil. The high rate of actionable alterations in and other driver genes in LA reinforces the need to incorporate TKI guided by molecular diagnostics into clinical routines for patients in both public and private healthcare systems.
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http://dx.doi.org/10.3389/fonc.2020.01068DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7343968PMC
July 2020

Clinical and Molecular Assessment of Patients with Lynch Syndrome and Sarcomas Underpinning the Association with Germline Pathogenic Variants.

Cancers (Basel) 2020 Jul 9;12(7). Epub 2020 Jul 9.

Genomics and Molecular Biology Group, International Research Center/CIPE, A.C.Camargo Cancer Center, São Paulo 01508-010, Brazil.

Lynch syndrome (LS) is a hereditary cancer-predisposing syndrome associated most frequently with epithelial tumors, particularly colorectal (CRC) and endometrial carcinomas (EC). The aim of this study was to investigate the relationship between sarcomas and LS by performing clinical and molecular characterization of patients presenting co-occurrence of sarcomas and tumors from the LS spectrum. We identified 27 patients diagnosed with CRC, EC, and other LS-associated tumors who had sarcomas in the same individuals or families. Germline genetic testing, mismatch repair (MMR) protein immunohistochemistry, microsatellite instability (MSI), and other molecular analyses were performed. Five LS patients presenting personal or family history of sarcomas were identified (3 carriers and 2 ), with 2 having Muir-Torre phenotypes. For two carriers we confirmed the etiology of the sarcomas (one liposarcoma and two osteosarcomas) as LS-related, since the tumors were MSH2/MSH6-deficient, MSI-high, or presented a truncated MSH2 transcript. Additionally, we reviewed 43 previous reports of sarcomas in patients with LS, which revealed a high frequency (58%) of alterations. In summary, sarcomas represent a rare clinical manifestation in patients with LS, especially in carriers, and the analysis of tumor biological characteristics can be useful for definition of tumor etiology and novel therapeutic options.
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http://dx.doi.org/10.3390/cancers12071848DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7408879PMC
July 2020

Three photobiomodulation protocols in the prevention/treatment of radiotherapy-induced oral mucositis.

Photodiagnosis Photodyn Ther 2020 Sep 30;31:101906. Epub 2020 Jun 30.

Stomatology Department- A.C. Camargo Cancer Center, Sao Paulo, Brazil; Stomatology Department School of Dentistry, Sao Paulo University, Sao Paulo, Brazil. Electronic address:

Purpose: To compare three Photobiomodulation protocols to prevent/treat oral mucositis associated to radiotherapy.

Methods: Seventy-three patients with cancer in oral cavity, oropharynx, and nasopharynx, who underwent RT with dose in facial fields equal or higher than 6000 cGy were randomized into three groups (mean RT dose = 66 cGy ±4.9). Protocols of Group 1 was 660 nm, 15 mW, 3.8 J/cm2, Group 2 660 nm, 25 mW, 6.3 J/cm2 both starting on the first day of radiotherapy, and group 3 660 nm, 15 mW, 3.8 J/cm2 for therapeutic purpose. The patients of group 1 and 2 were irradiated at 40 points daily covering non-keratinizing oral mucosa. The spot size (probe's tip surface size) was 0.040 cm2 for all groups. Oral mucositis was evaluated according to both WHO and NCI scales, and pain related to oral mucositis was scored using the VAS.

Results: Patients from group 1 presented with grade II oral mucositis later than groups 2 and 3 (p < 0.001). Moreover, groups 2 and 3 also presented with a mean higher of oral mucositis grade than group 1, p < 0.001. Pain scores were lower in group 1 (p = 0.002).

Conclusions: The Photobiomodulation used in Group 1 was more effective than the protocols used in groups 2 and 3 in controlling the grade II oral mucositis intensity, and mean pain scores.
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http://dx.doi.org/10.1016/j.pdpdt.2020.101906DOI Listing
September 2020

Assessment of somatic mutations in urine and plasma of Wilms tumor patients.

Cancer Med 2020 08 26;9(16):5948-5959. Epub 2020 Jun 26.

Laboratory of Genomics and Molecular Biology, International Research Center/CIPE, A. C. Camargo Cancer Center, São Paulo, Brazil.

Tumor DNA has been detected in body fluids of cancer patients. Somatic tumor mutations are being used as biomarkers in body fluids to monitor chemotherapy response as a minimally invasive tool. In this study, we evaluated the potential of tracking somatic mutations in free DNA of plasma and urine collected from Wilms tumor (WT) patients for monitoring treatment response. Wilms tumor is a pediatric renal tumor resulting from cell differentiation errors during nephrogenesis. Its mutational repertoire is not completely defined. Thus, for identifying somatic mutations from tumor tissue DNA, we screened matched tumor/leukocyte DNAs using either a panel containing 16 WT-associated genes or whole-exome sequencing (WES). The identified somatic tumor mutations were tracked in urine and plasma DNA collected before, during and after treatment. At least one somatic mutation was identified in five out of six WT tissue samples analyzed. Somatic mutations were detected in body fluids before treatment in all five patients (three patients in urine, three in plasma, and one in both body fluids). In all patients, a decrease of the variant allele fraction of somatic mutations was observed in body fluids during neoadjuvant chemotherapy. Interestingly, the persistence of somatic mutations in body fluids was in accordance with clinical parameters. For one patient who progressed to death, it persisted in high levels in serial body fluid samples during treatment. For three patients without disease progression, somatic mutations were not consistently detected in samples throughout monitoring. For one patient with bilateral disease, a somatic mutation was detected at low levels with no support of clinical manifestation. Our results demonstrated the potential of tracking somatic mutations in urine and plasma DNA as a minimally invasive tool for monitoring WT patients. Additional investigation is needed to check the clinical value of insistent somatic mutations in body fluids.
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http://dx.doi.org/10.1002/cam4.3236DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7433816PMC
August 2020

Insights Into the Somatic Mutation Burden of Hepatoblastomas From Brazilian Patients.

Front Oncol 2020 5;10:556. Epub 2020 May 5.

Department of Genetics and Evolutionary Biology, Human Genome and Stem-Cell Research Center, Institute of Biosciences, University of São Paulo, São Paulo, Brazil.

Hepatoblastoma is a very rare embryonal liver cancer supposed to arise from the impairment of hepatocyte differentiation during embryogenesis. In this study, we investigated by exome sequencing the burden of somatic mutations in a cohort of 10 hepatoblastomas, including a congenital case. Our data disclosed a low mutational background and pointed out to a novel set of candidate genes for hepatoblastoma biology, which were shown to impact gene expression levels. Only three recurrently mutated genes were detected: and two novel candidates, and . A relevant finding was the identification of a recurrent mutation (A235G) in two hepatoblastomas at the gene; evaluation of RNA and protein expression revealed upregulation of in tumors. The analysis was replicated in two independents cohorts, substantiating that an activation of the pathway occurs in hepatoblastomas. In inflammatory regions of hepatoblastomas, CX3CL1/CX3CR1 were not detected in the infiltrated lymphocytes, in which they should be expressed in normal conditions, whereas necrotic regions exhibited negative labeling in tumor cells, but strongly positive infiltrated lymphocytes. Altogether, these data suggested that upregulation may be a common feature of hepatoblastomas, potentially related to chemotherapy response and progression. In addition, three mutational signatures were identified in hepatoblastomas, two of them with predominance of either the COSMIC signatures 1 and 6, found in all cancer types, or the COSMIC signature 29, mostly related to tobacco chewing habit; a third novel mutational signature presented an unspecific pattern with an increase of C>A mutations. Overall, we present here novel candidate genes for hepatoblastoma, with evidence that chemokine signaling pathway is likely involved with progression, besides reporting specific mutational signatures.
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http://dx.doi.org/10.3389/fonc.2020.00556DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7214543PMC
May 2020

MLH1 intronic variants mapping to + 5 position of splice donor sites lead to deleterious effects on RNA splicing.

Fam Cancer 2020 10;19(4):323-336

Instituto de Medicina Traslacional e Ingeniería Biomédica (IMTIB) [HIBA-IUHI-CONICET], C1199ABH, Buenos Aires, Argentina.

Germline pathogenic variants in the DNA mismatch repair genes (MMR): MLH1, MSH2, MSH6, and PMS2, are causative of Lynch syndrome (LS). However, many of the variants mapping outside the invariant splice site positions (IVS ± 1, IVS ± 2) are classified as variants of unknown significance (VUS). Three such variants (MLH1 c.588+5G>C, c.588+5G>T and c.677+5G>A) were identified in 8 unrelated LS families from Argentina, Brazil and Chile. Herein, we collected clinical information on these families and performed segregation analysis and RNA splicing studies to assess the implication of these VUS in LS etiology. Pedigrees showed a clear pattern of variant co-segregation with colorectal cancer and/or other LS-associated malignancies. Tumors presented deficient expression of MLH1-PMS2 proteins in 7/7 of the LS families, and MSI-high status in 3/3 cases. Moreover, RNA analyses revealed that c.588+5G>C and c.588+5G>T induce skipping of exon 7 whereas c.677+5G>A causes skipping of exon 8. In sum, we report that the combined clinical findings in the families and the molecular studies provided the evidences needed to demonstrate that the three MLH1 variants are causative of LS and to classify c.588+5G>C and c.677+5G>A as class 5 (pathogenic), and c.588+5G>T as class 4 (likely-pathogenic). Our findings underline the importance of performing clinical and family analyses, as well as RNA splicing assays in order to determine the clinical significance of intronic variants, and contribute to the genetic counseling and clinical management of patients and their relatives.
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http://dx.doi.org/10.1007/s10689-020-00182-5DOI Listing
October 2020

Germline variants in DNA repair genes associated with hereditary breast and ovarian cancer syndrome: analysis of a 21 gene panel in the Brazilian population.

BMC Med Genomics 2020 02 10;13(1):21. Epub 2020 Feb 10.

Department of Genetics, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, SP, Brazil.

Background: The Hereditary Breast and Ovarian Cancer Syndrome (HBOC) occurs in families with a history of breast/ovarian cancer, presenting an autosomal dominant inheritance pattern. BRCA1 and BRCA2 are high penetrance genes associated with an increased risk of up to 20-fold for breast and ovarian cancer. However, only 20-30% of HBOC cases present pathogenic variants in those genes, and other DNA repair genes have emerged as increasing the risk for HBOC. In Brazil, variants in ATM, ATR, CHEK2, MLH1, MSH2, MSH6, POLQ, PTEN, and TP53 genes have been reported in up to 7.35% of the studied cases. Here we screened and characterized variants in 21 DNA repair genes in HBOC patients.

Methods: We systematically analyzed 708 amplicons encompassing the coding and flanking regions of 21 genes related to DNA repair pathways (ABRAXAS1, ATM, ATR, BARD1, BRCA1, BRCA2, BRIP1, CDH1, CHEK2, MLH1, MRE11, MSH2, MSH6, NBN, PALB2, PMS2, PTEN, RAD50, RAD51, TP53 and UIMC1). A total of 95 individuals with HBOC syndrome clinical suspicion in Southeast Brazil were sequenced, and 25 samples were evaluated for insertions/deletions in BRCA1/BRCA2 genes. Identified variants were assessed in terms of population allele frequency and their functional effects were predicted through in silico algorithms.

Results: We identified 80 variants in 19 genes. About 23.4% of the patients presented pathogenic variants in BRCA1, BRCA2 and TP53, a frequency higher than that identified among previous studies in Brazil. We identified a novel variant in ATR, which was predicted as pathogenic by in silico tools. The association analysis revealed 13 missense variants in ABRAXAS1, BARD1, BRCA2, CHEK2, CDH1, MLH1, PALB2, and PMS2 genes, as significantly associated with increased risk to HBOC, and the patients carrying those variants did not present large insertions or deletions in BRCA1/BRCA2 genes.

Conclusions: This study embodies the third report of a multi-gene analysis in the Brazilian population, and addresses the first report of many germline variants associated with HBOC in Brazil. Although further functional analyses are necessary to better characterize the contribution of those variants to the phenotype, these findings would improve the risk estimation and clinical follow-up of patients with HBOC clinical suspicion.
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http://dx.doi.org/10.1186/s12920-019-0652-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7011249PMC
February 2020

BAP1 tumor predisposition syndrome case report: pathological and clinical aspects of BAP1-inactivated melanocytic tumors (BIMTs), including dermoscopy and confocal microscopy.

BMC Cancer 2019 Nov 9;19(1):1077. Epub 2019 Nov 9.

Laboratory of Genomics and Molecular Biology, A.C. Camargo Cancer Center, Rua Taguá, 440, São Paulo, SP, CEP: 0508-010, Brazil.

Background: BRCA1 associated-protein 1 (BAP1) tumor predisposition syndrome is associated with an increased risk for malignant mesotheliomas, uveal and cutaneous melanomas, renal cell carcinomas, and singular cutaneous lesions. The latter are referred to as BAP1-inactivated melanocytic tumors (BIMTs). When multiple BIMTs manifest, they are considered potential markers of germline BAP1 mutations.

Case Presentation: Here, we report a novel pathogenic BAP1 germline variant in a family with a history of BIMTs, cutaneous melanomas, and mesotheliomas. We also describe singular pathological aspects of the patient's BIMT lesions and their correlation with dermoscopic and reflectance confocal microscopy findings.

Conclusions: This knowledge is crucial for the recognition of BIMTs by dermatologists and pathologists, allowing the determination of appropriate management for high-risk patients, such as genetic investigations and screening for potentially aggressive tumors.
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http://dx.doi.org/10.1186/s12885-019-6226-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6842488PMC
November 2019

Influence of BRCA1 Germline Mutations in the Somatic Mutational Burden of Triple-Negative Breast Cancer.

Transl Oncol 2019 Nov 13;12(11):1453-1460. Epub 2019 Aug 13.

CIPE, International Research Center, A.C. Camargo Cancer Center, São Paulo, SP, Brazil; National Institute of Science and Technology in Oncogenomics and Therapeutic Innovation, São Paulo, SP, Brazil. Electronic address:

The majority of the hereditary triple-negative breast cancers (TNBCs) are associated with BRCA1 germline mutations. Nevertheless, the understanding of the role of BRCA1 deficiency in the TNBC tumorigenesis is poor. In this sense, we performed whole-exome sequencing of triplet samples (leucocyte, tumor, and normal-adjacent breast tissue) for 10 cases of early-onset TNBC, including 5 hereditary (with BRCA1 germline pathogenic mutation) and 5 sporadic (with no BRCA1 or BRCA2 germline pathogenic mutations), for assessing the somatic mutation repertoire. Protein-affecting somatic mutations were identified for both mammary tissues, and Ingenuity Pathway Analysis was used to investigate gene interactions. BRCA1 and RAD51C somatic promoter methylation in tumor samples was also investigated by bisulfite sequencing. Sporadic tumors had higher proportion of driver mutations (≥25% allele frequency) than BRCA1 hereditary tumors, whereas no difference was detected in the normal breast samples. Distinct gene networks were obtained from the driver genes in each group. The Cancer Genome Atlas data analysis of TNBC classified as hereditary and sporadic reinforced our findings. The data presented here indicate that in the absence of BRCA1 germline mutations, a higher number of driver mutations are required for tumor development and that different defective processes are operating in the tumorigenesis of hereditary and sporadic TNBC in young women.
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http://dx.doi.org/10.1016/j.tranon.2019.07.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6706625PMC
November 2019

TET Upregulation Leads to 5-Hydroxymethylation Enrichment in Hepatoblastoma.

Front Genet 2019 12;10:553. Epub 2019 Jun 12.

Human Genome and Stem-Cell Research Center, Department of Genetics and Evolutionary Biology, Institute of Biosciences, University of São Paulo, São Paulo, Brazil.

Hepatoblastoma is an embryonal liver tumor carrying few genetic alterations. We previously disclosed in hepatoblastomas a genome-wide methylation dysfunction, characterized by hypermethylation at specific CpG islands, in addition to a low-level hypomethylation pattern in non-repetitive intergenic sequences, in comparison to non-tumoral liver tissues, shedding light into a crucial role for epigenetic dysregulation in this type of cancer. To explore the underlying mechanisms possibly related to aberrant epigenetic modifications, we evaluated the expression profile of a set of genes engaged in the epigenetic machinery related to DNA methylation (, , , , , , , and ), as well as the 5-hydroxymethylcytosine (5hmC) global level. We observed in hepatoblastomas a general disrupted expression of these genes from the epigenetic machinery, mainly , , and upregulation, in association with an enrichment of 5hmC content. Our findings support a model of active demethylation by in hepatoblastoma, probably during early stages of liver development, which in combination with overexpression would lead to DNA hypomethylation and an increase in overall 5hmC content. Furthermore, our data suggest that decreased 5hmC content might be associated with poor survival rate, highlighting a pivotal role of epigenetics in hepatoblastoma development and progression.
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http://dx.doi.org/10.3389/fgene.2019.00553DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6582250PMC
June 2019

Association of Folate and Vitamins Involved in the 1-Carbon Cycle with Polymorphisms in the Methylenetetrahydrofolate Reductase Gene (MTHFR) and Global DNA Methylation in Patients with Colorectal Cancer.

Nutrients 2019 Jun 18;11(6). Epub 2019 Jun 18.

Department of Pelvic Surgery, A.C. Camargo Cancer Center, Rua Professor Antônio Prudente, 211, Liberdade, São Paulo, São Paulo CEP 01509-010, Brazil.

Folate, vitamin B2, vitamin B6, vitamin B12, choline, and betaine are nutrients involved in the 1-carbon cycle that can alter the levels of DNA methylation and influence genesis and/or tumor progression. Thus, the objective of this study was to evaluate the association of folate and vitamins involved in the 1-carbon cycle and MTHFR polymorphisms in global DNA methylation in patients with colorectal cancer gene. The study included 189 patients with colorectal adenocarcinoma answering a clinical evaluation questionnaire and the Food Frequency Questionnaire (FFQ) validated for patients with colon and rectal cancer. Blood samples were collected for evaluation of MTHFR gene polymorphisms in global DNA methylation in blood and in tumor. The values for serum folate were positively correlated with the equivalent total dietary folate (total DFE) (rho = 0.51, = 0.03) and global DNA methylation (rho = 0.20, = 0.03). Individuals aged over 61 years ( = 0.01) in clinicopathological staging III and IV ( = 0.01) and with + heterozygous mutated homozygous genotypes for the MTHFR A1298C gene had higher levels of global DNA methylation ( = 0.04). The association between dietary intake of folate, serum folate, and tumor stage were predictive of global DNA methylation in patients' blood. The levels of serum folate, the dietary folate and the status of DNA methylation can influence clinicopathological staging.
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http://dx.doi.org/10.3390/nu11061368DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6627304PMC
June 2019

Enhanced type I interferon gene signature in primary antiphospholipid syndrome: Association with earlier disease onset and preeclampsia.

Autoimmun Rev 2019 Apr 14;18(4):393-398. Epub 2019 Feb 14.

Rheumatology Division, Faculdade de Medicina da Universidade de São Paulo (USP), Brazil.

Objective: Recently, two studies demonstrated that a relevant percentage of primary antiphospholipid syndrome (PAPS) patients had an upregulation of interferon (IFN) genes. However, 20%-28% of these patients had anti-dsDNA, a highly specific systemic lupus erythematosus (SLE) autoantibody. This study aimed to determine the prevalence of the type I IFN signature in the peripheral blood mononuclear cells of PAPS patients without specific SLE autoantibodies and search for its clinical associations.

Methods: Fifty-three PAPS patients (Sydney's criteria) were consecutively selected and age-matched with 50 healthy controls. A third group of nonimmune-mediated thrombophilia patients was also included. The expression of 41 IFN-induced genes was analyzed using real time quantitative PCR. A principal component analysis determined which genes composed the IFN signature, and the z-score was calculated. An ROC curve defined the signature cut-off.

Results: Six genes remained in the IFN signature DNAJA1, IFIT5, IFI27, MX1, IFI6, and TYK2. The ROC cutoff was 3.9-fold (AUC = 0.706, S = 0.49, E = 0.86, PPV = 0.79, NPV = 0.61). The type I IFN signature was present in 49% of the patients with PAPS compared with 14.0% of the healthy controls and 17% of the nonimmune-mediated thrombophilia patients (p < .0001). The IFN signature was associated with a younger age at the first antiphospholipid syndrome event (p = .023) and with preeclampsia (p = .032).

Conclusion: Our results indicate that PAPS patients without lupus-specific antibodies have an enhanced type I IFN gene signature that is not observed in nonimmune-mediated thrombophilia. Also, this overexpression of type I IFN-regulated genes associated with an earlier onset of antiphospholipid syndrome event and preeclampsia.
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http://dx.doi.org/10.1016/j.autrev.2018.11.004DOI Listing
April 2019

Prevalence of BRCA1 and BRCA2 pathogenic and likely pathogenic variants in non-selected ovarian carcinoma patients in Brazil.

BMC Cancer 2019 Jan 3;19(1). Epub 2019 Jan 3.

Department of Medical Oncology, AC Camargo Cancer Center, Rua Professor Antonio Prudente 211, São Paulo, CEP: 01509-900, Brazil.

Background: BRCA1/2 pathogenic (P) and likely pathogenic (LP) germline variants are frequent among patients with ovarian carcinoma. However, these variants have not been extensively characterized in patients with ovarian cancer in Brazil.

Methods: In this retrospective study we evaluated clinical characteristics and BRCA1/2 genetic test results from patients with ovarian carcinoma who underwent genetic counseling at A.C.Camargo Cancer Center (Brazil) between 2015 and 2017 and had performed germline genetic testing of BRCA1/2 genes.

Results: Among 158 patients, 33 P and LP variants and were found (20.8%), 27 in BRCA1 and six in BRCA2, and six variants of unknown clinical significance (VUS). Thirteen percent of the patients did not have Multiplex Ligation-dependent Probe Amplification (MLPA) results. Three P variants in BRCA1 were found in more than one patient: c.5266dupC (p.Gln1756Profs*74), c.3331_3334delCAAG (p.Gln1111Asnfs5*), and c.211A > G (p.Arg71Gly). One LP variant in BRCA1 had not been previously described, c.4153_4154delCT (p.Leu1385Ilefs*5). Patients with previous diagnosis of breast cancer were carriers of P or LP variant in 8 of 12 cases (66.7%), and patients with a family history of ovarian or breast cancer in first- or second-degree relatives were carriers of P or LP variant in 26.7% of cases compared to 16.9% for patients without family history (p = 0.166).

Conclusion: Prevalence of BRCA1/2 germline P and LP variants is slightly higher than previously described by the largest occidental studies, with a high prevalence of variant c.5266dupC (p.Gln1756Profs*74) in BRCA1 observed. Moreover, we identified a new LP variant.
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http://dx.doi.org/10.1186/s12885-018-5235-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6319008PMC
January 2019

From colorectal cancer pattern to the characterization of individuals at risk: Picture for genetic research in Latin America.

Int J Cancer 2019 07 5;145(2):318-326. Epub 2018 Dec 5.

Department of Tumor Biology, Institute for Cancer Research, Oslo University Hospital, Oslo, Norway.

Colorectal cancer (CRC) is one of the most common cancers in Latin America and the Caribbean, with the highest rates reported for Uruguay, Brazil and Argentina. We provide a global snapshot of the CRC patterns, how screening is performed, and compared/contrasted to the genetic profile of Lynch syndrome (LS) in the region. From the literature, we find that only nine (20%) of the Latin America and the Caribbean countries have developed guidelines for early detection of CRC, and also with a low adherence. We describe a genetic profile of LS, including a total of 2,685 suspected families, where confirmed LS ranged from 8% in Uruguay and Argentina to 60% in Peru. Among confirmed LS, path_MLH1 variants were most commonly identified in Peru (82%), Mexico (80%), Chile (60%), and path_MSH2/EPCAM variants were most frequently identified in Colombia (80%) and Argentina (47%). Path_MSH6 and path_PMS2 variants were less common, but they showed important presence in Brazil (15%) and Chile (10%), respectively. Important differences exist at identifying LS families in Latin American countries, where the spectrum of path_MLH1 and path_MSH2 variants are those most frequently identified. Our findings have an impact on the evaluation of the patients and their relatives at risk for LS, derived from the gene affected. Although the awareness of hereditary cancer and genetic testing has improved in the last decade, it is remains deficient, with 39%-80% of the families not being identified for LS among those who actually met both the clinical criteria for LS and showed MMR deficiency.
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http://dx.doi.org/10.1002/ijc.31920DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6587543PMC
July 2019

Expanding morphological and clinical aspects of hereditary leiomyomatosis and renal cell carcinoma (HLRCC): a case report in a patient with unusual morphology and clinical presentation.

Virchows Arch 2018 Dec 31;473(6):775-779. Epub 2018 Aug 31.

Department of Pathology, AC Camargo Cancer Center, Rua Professor Antônio Prudente 211, São Paulo, 01509-900, Brazil.

Renal cell carcinoma (RCC) accounts for 2-3% of all malignant disease in adults. Hereditary RCC represents 5 to 8% of kidney tumors. Hereditary leiomyomatosis and renal cell carcinoma (HLRCC) represents an autosomal dominant syndrome that results from a germline mutation in fumarate hydratase gene (FH). HLRCC patients typically present with skin or uterine leiomyomas and renal neoplasms. HLRCC was recently recognized as a distinct renal tumor subtype by the WHO 2016 classification. Many morphological patterns such as papillary, solid, tubular, and cystic had been described as part of morphological aspects of HLRCC. In this study, we describe a case of a patient that had a history of persistence of ductus arteriosus (PDA) and cryptorchidism. In addition, the renal tumor showed a very unusual hystiocytoid morphological aspect. We confirmed the presence of a FH germline mutation both in the patient and his mother.
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http://dx.doi.org/10.1007/s00428-018-2420-3DOI Listing
December 2018

The germline mutational landscape of BRCA1 and BRCA2 in Brazil.

Sci Rep 2018 06 15;8(1):9188. Epub 2018 Jun 15.

Programa de Pós-Graduação em Genética e Biologia Molecular, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil.

The detection of germline mutations in BRCA1 and BRCA2 is essential to the formulation of clinical management strategies, and in Brazil, there is limited access to these services, mainly due to the costs/availability of genetic testing. Aiming at the identification of recurrent mutations that could be included in a low-cost mutation panel, used as a first screening approach, we compiled the testing reports of 649 probands with pathogenic/likely pathogenic variants referred to 28 public and private health care centers distributed across 11 Brazilian States. Overall, 126 and 103 distinct mutations were identified in BRCA1 and BRCA2, respectively. Twenty-six novel variants were reported from both genes, and BRCA2 showed higher mutational heterogeneity. Some recurrent mutations were reported exclusively in certain geographic regions, suggesting a founder effect. Our findings confirm that there is significant molecular heterogeneity in these genes among Brazilian carriers, while also suggesting that this heterogeneity precludes the use of screening protocols that include recurrent mutation testing only. This is the first study to show that profiles of recurrent mutations may be unique to different Brazilian regions. These data should be explored in larger regional cohorts to determine if screening with a panel of recurrent mutations would be effective.
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http://dx.doi.org/10.1038/s41598-018-27315-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6003960PMC
June 2018

DNA methylation landscape of hepatoblastomas reveals arrest at early stages of liver differentiation and cancer-related alterations.

Oncotarget 2017 Nov 25;8(58):97871-97889. Epub 2016 Dec 25.

Department of Genetics and Evolutionary Biology, Institute of Biosciences, University of São Paulo, São Paulo, Brazil.

Hepatoblastomas are uncommon embryonal liver tumors accounting for approximately 80% of childhood hepatic cancer. We hypothesized that epigenetic changes, including DNA methylation, could be relevant to hepatoblastoma onset. The methylomes of eight matched hepatoblastomas and non-tumoral liver tissues were characterized, and data were validated in an independent group (11 hepatoblastomas). In comparison to differentiated livers, hepatoblastomas exhibited a widespread and non-stochastic pattern of global low-level hypomethylation. The analysis revealed 1,359 differentially methylated CpG sites (DMSs) between hepatoblastomas and control livers, which are associated with 765 genes. Hypomethylation was detected in hepatoblastomas for ~58% of the DMSs with enrichment at intergenic sites, and most of the hypermethylated CpGs were located in CpG islands. Functional analyses revealed enrichment in signaling pathways involved in metabolism, negative regulation of cell differentiation, liver development, cancer, and Wnt signaling pathway. Strikingly, an important overlap was observed between the 1,359 DMSs and the CpG sites reported to exhibit methylation changes through liver development (p<0.0001), with similar patterns of methylation in both hepatoblastomas and fetal livers compared to adult livers. Overall, our results suggest an arrest at early stages of liver cell differentiation, in line with the hypothesis that hepatoblastoma ontogeny involves the disruption of liver development. This genome-wide methylation dysfunction, taken together with a relatively small number of driver genetic mutations reported for both adult and pediatric liver cancers, shed light on the relevance of epigenetic mechanisms for hepatic tumorigenesis.
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http://dx.doi.org/10.18632/oncotarget.14208DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5716698PMC
November 2017

BRCA1 deficiency is a recurrent event in early-onset triple-negative breast cancer: a comprehensive analysis of germline mutations and somatic promoter methylation.

Breast Cancer Res Treat 2018 02 7;167(3):803-814. Epub 2017 Nov 7.

Laboratory of Genomics and Molecular Biology, CIPE - A. C. Camargo Cancer Center, São Paulo, Brazil.

Purpose: BRCA1 germline mutation is closely associated with triple-negative breast cancer. BRCA deficiency leads to impaired DNA repair and tumor development, and understanding this deficiency, in both hereditary and sporadic scenarios, is of great clinical and biological interest. Here, we investigated germline or somatic events that might lead to BRCA1 impairment in triple-negative breast cancer. We also analyzed the clinical implications associated with BRCA deficiency.

Methods: Next-generation sequencing for the BRCA1/2 genes and multiplex ligation-dependent probe amplification (MLPA) for the BRCA1 gene were performed for mutation screening. A customized bisulfite next-generation sequencing approach was used for assessing BRCA1 promoter methylation status in tumor tissue.

Results: A total of 131 triple-negative cases were assessed, and germline pathogenic variants were detected in 13.0% of all cases and in 26% of cases diagnosed in young women. Most germline pathogenic variants (88.2%) occurred in the BRCA1 gene. BRCA1 promoter hypermethylation was detected in 20.6% of tumors; none of these tumors were in BRCA1/2 pathogenic variant carriers. BRCA1 impairment by either germline or somatic events was significantly more frequent in young women (55% in those ≤ 40 years; 33% in those 41-50 years; 22% in those > 50 years of age) and associated with better overall and disease-free survival rates in this group of patients.

Conclusions: BRCA1 deficiency was recurrent in early-onset triple-negative breast cancer in Brazilian patients and associated with improved survival. With the new treatment modalities being investigated, including poly (ADP-ribose)-polymerase (PARP) inhibitor therapy, our results suggest that a significant proportion of young women with this subtype of tumor might benefit from PARP inhibitor treatment, which warrants further investigation.
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http://dx.doi.org/10.1007/s10549-017-4552-6DOI Listing
February 2018

Pitfalls in genetic testing: a case of a SNP in primer-annealing region leading to allele dropout in .

Mol Genet Genomic Med 2017 Jul 11;5(4):443-447. Epub 2017 May 11.

Laboratory of Genomics and Molecular BiologyInternational Research CenterAC Camargo Cancer CenterSão PauloBrazil.

Background: Hereditary breast and ovarian cancer is characterized by mutations in or genes and PCR-based screening techniques, such as capillary sequencing and next-generation sequencing (NGS), are considered gold standard methods for detection of pathogenic mutations in these genes. Single-nucleotide polymorphisms (SNPs) constitute a vast source of variation in the human genome and represent a risk for misdiagnosis in genetic testing, since the presence of a SNP in primer-annealing sites may cause false negative results due to allele dropout. However, few reports are available and the frequency of this phenomenon in diagnostic assays remains unknown.

Methods And Results: In this article, we investigated the causes of a false negative capillary sequencing result in involving a mother-daughter dyad. Using several molecular strategies, including different DNA polymerases, primer redesign, allele-specific PCR and NGS, we established that the initial misdiagnosis was caused by a SNP located in the primer-annealing region, leading to allele dropout of the mutated allele.

Conclusion: Assuming that this problem can also occur in any PCR-based method that are widely used in diagnostic settings, the clinical report presented here draws attention for one of the limitations of genetic testing in general, for which medical and laboratory communities need to be aware.
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http://dx.doi.org/10.1002/mgg3.295DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5511807PMC
July 2017

DNA repair genes PAXIP1 and TP53BP1 expression is associated with breast cancer prognosis.

Cancer Biol Ther 2017 06 5;18(6):439-449. Epub 2017 May 5.

a Programa de Pesquisa Clínica , Coordenação de Pesquisa, Instituto Nacional de Câncer , Rio de Janeiro , RJ , Brazil.

Despite remarkable advances in diagnosis, prognosis and treatment, advanced or recurrent breast tumors have limited therapeutic approaches. Many treatment strategies try to explore the limitations of DNA damage response (DDR) in tumor cells to selectively eliminate them. BRCT (BRCA1 C-terminal) domains are present in a superfamily of proteins involved in cell cycle checkpoints and the DDR. Tandem BRCT domains (tBRCT) represent a distinct class of these domains. We investigated the expression profile of 7 tBRCT genes (BARD1, BRCA1, LIG4, ECT2, MDC1, PAXIP1/PTIP and TP53BP1) in breast cancer specimens and observed a high correlation between PAXIP1 and TP53BP1 gene expression in tumor samples. Tumors with worse prognosis (tumor grade 3 and triple negative) showed reduced expression of tBRCT genes, notably, PAXIP1 and TP53BP1. Survival analyses data indicated that tumor status of both genes may impact prognosis. PAXIP1 and 53BP1 protein levels followed gene expression results, i.e., are intrinsically correlated, and also reduced in more advanced tumors. Evaluation of both genes in triple negative breast tumor samples which were characterized for their BRCA1 status showed that PAXIP1 is overexpressed in BRCA1 mutant tumors. Taken together our findings indicate that PAXIP1 status correlates with breast cancer staging, in a manner similar to what has been characterized for TP53BP1.
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http://dx.doi.org/10.1080/15384047.2017.1323590DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5536937PMC
June 2017

Biobanking and cytopathology: Challenges and opportunities from a Brazilian perspective.

Cancer Cytopathol 2017 06 29;125(6):373-377. Epub 2017 Mar 29.

Department of Anatomic Pathology, A.C. Camargo Cancer Center, Sao Paulo, Brazil.

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http://dx.doi.org/10.1002/cncy.21836DOI Listing
June 2017

Genetic and epigenetic characterization of the BRCA1 gene in Brazilian women at-risk for hereditary breast cancer.

Oncotarget 2017 Jan;8(2):2850-2862

Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, SP, Brazil.

This study aimed to characterize women at-risk for hereditary BC regarding their clinical and molecular characteristics (mutation and methylation in the BRCA1 gene) and correlate the gene expression levels with histopathological, clinical and family history information. BRCA1 real time qPCR was performed to evaluate methylation status and gene expression. The study included 88 women grouped according to the BRCA1 mutational status: 23 BRCA1 mutated, 22 with a Variant of Unknown Significance (VUS) in BRCA1 and 43 BRCA1 WT. Most BRCA1 mutated tumors were triple negative (69.6%) and had histologic grade III (61.0%). Patients with VUS/WT BRCA1 were predominantly of luminal B subtype with histological grades I and II. Regarding the methylation profile, BRCA1 hypermethylation was observed in only two patients (both WT) and none had association with pathogenic BRCA1 mutation. In one patient methylation was present in both, tumor and normal tissues. Hypermethylated tumors had ductal histology, negativity for ER and occurred in < 50 years patients. Gene expression profile showed in all groups lower BRCA1 mRNA levels in tumor tissue compared to the adjacent breast tissue, thereby indicating the loss/decrease of gene function. No association was found between the levels of BRCA1 gene expression and family history of cancer. In summary, our findings suggested that methylation at the BRCA1 gene is not the "second" event in the development of BC in patients with germline mutations in BRCA1 and, although rare, BRCA1 epimutations can constitute an explanation for a fraction of HBOC families.
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http://dx.doi.org/10.18632/oncotarget.13750DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5356847PMC
January 2017

Germline mutations in BRCA1 and BRCA2 in epithelial ovarian cancer patients in Brazil.

BMC Cancer 2016 12 3;16(1):934. Epub 2016 Dec 3.

Departamento de Radiologia e Oncologia, Centro de Investigação Translacional em Oncologia, Instituto do Câncer do Estado de São Paulo, Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo, São Paulo, SP, Brazil.

Background: Approximately 8-15% epithelial ovarian cancer patients are BRCA1 or BRCA2 germline mutation carriers. Brazilian inhabitants may have peculiar genetic characteristics associated with ethnic diversity, and studies focusing on the entire BRCA1/BRCA2 gene sequencing in Brazilian ovarian cancer patients are still lacking. The aim of this study was to evaluate BRCA1/2 mutations, through entire gene sequencing, in a Brazilian population of women with epithelial ovarian cancer.

Methods: In a cross sectional study performed in one reference centre for cancer treatment in São Paulo, Brazil, 100 patients diagnosed with epithelial ovarian cancer unselected for family history of breast and/or ovarian cancer were included. The complete coding sequence of BRCA1/2 genes was evaluated through Next-Generation or capillary sequencing. Large deletions were investigated through Multiplex Ligation-dependent Probe Amplification (MLPA).

Results: Nineteen pathogenic mutations (BRCA1: n = 17 and BRCA2: n = 2) featuring 14 different mutations, including two large deletions in BRCA1 (exon 1-2 deleted and exon 5-7 deleted) were identified. Three mutations were detected more than once (c.3331_3334delCAAG, c.5266dupC and c.4484G > T). Two novel frameshift mutations were identified, one in BRCA1 (c.961_962delTG) and one in BRCA2 (c.1963_1963delC). BRCA1/2 mutations were seen in 35.5% of the patients with first and/or second-degree relatives with breast and/or ovarian cancer. Nineteen variants of uncertain significance (VUS) were detected (BRCA1: n = 2 and BRCA2: n = 17), including five distinct missense variants (BRCA1: c.5348 T > C; BRCA2: c.2350A > G, c.3515C > T, c.7534C > T, and c.8351G > A).

Conclusions: Among epithelial ovarian cancer patients unselected for family history of cancer, 19% were BRCA1/2 germline mutation carriers. Almost ¾ of the BRCA mutations, including two large deletions, were detected only once. Our work emphasizes the need of entire gene sequencing and MLPA screening in Brazil.
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http://dx.doi.org/10.1186/s12885-016-2966-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5135756PMC
December 2016

Infections with multiple high-risk HPV types are associated with high-grade and persistent low-grade intraepithelial lesions of the cervix.

Cancer Cytopathol 2017 Feb 21;125(2):138-143. Epub 2016 Nov 21.

Department of Pathology, A.C. Camargo Cancer Center, Sao Paulo, Brazil.

Background: Infections with multiple human papillomavirus (HPV) types (mHPV) in Papanicolaou tests have been reported but the histologic correlation and clinical meaning remains debatable.

Methods: The authors prospectively tested 37 HPV types using the Linear Array HPV Genotyping Test and correlated the results to cytology and histology findings in 260 women evaluated from June 2009 to October 2011 and followed for up to 60 months.

Results: HPV was detected in 148 of 235 samples (63%) and high-risk HPV was detected in 132 samples (56%). mHPV infection was found to be twice as common as single HPV (sHPV) infection and was detected more frequently in low-grade squamous intraepithelial lesion (LSIL) (48 of 83 samples [58%]) and high-grade squamous intraepithelial lesion or invasive carcinoma (HSIL + (26 of 47 samples [55%]) compared with other categories (P<.001). Of 34 LSIL/cervical intraepithelial neoplasia 1 (CIN1) index cases, 13 of 21 patients with mHPV (61.9%) persisted on CIN1, whereas no histologic abnormality was detected during follow-up in all 12 patients with sHPV infection (high risk or low risk) (P<.001). Eighteen of 20 patients with HSIL/cervical intraepithelial neoplasia 2 (CIN2) (90%) and high-risk mHPV persisted on HSIL+/CIN2 + whereas 6 of 11 patients with sHPV infection did not demonstrate HSIL+/CIN2 + on follow-up (54.5%) (P = .066). Approximately 40% of women with HSIL were infected by high-risk HPV types other than types 16 or 18.

Conclusions: High-risk mHPV infection identified patients with persistent LSIL/CIN1 and may to help identify patients at higher risk of disease progression to HSIL+/CIN2+. Longer follow-up will clarify the role of mHPV testing in patient care. Cancer Cytopathol 2017;125:138-143. © 2016 American Cancer Society.
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http://dx.doi.org/10.1002/cncy.21789DOI Listing
February 2017

Epidermal growth factor receptor as an adverse survival predictor in squamous cell carcinoma of the penis.

Hum Pathol 2017 03 15;61:97-104. Epub 2016 Nov 15.

Department of Anatomic Pathology, A.C. Camargo Cancer Center, 01508-010, São Paulo, SP, Brazil; General Pathology, Faculty of Dentistry, University of São Paulo, 05508-000, São Paulo, SP, Brazil. Electronic address:

Penile carcinoma (PC) is more frequent in underdeveloped countries, generally is diagnosed at an advanced stage when therapeutic options are restricted, and thus is associated with high morbidity/mortality rates. Recent studies have demonstrated clinical benefits with epidermal growth factor receptor (EGFR)-targeted therapy in patients with PC, although there is no test that provides accurate patient selection. The aim of the present study was to evaluate the prognostic value of EGFR gene and protein status in tumor samples from patients with primary penile squamous cell carcinoma. We assessed the expression of wild-type and 2 mutant EGFR isoforms (delA746-E750 and mL858R) by immunohistochemistry in 139 samples, of which 49 were also evaluated for EGFR copy number by fluorescence in situ hybridization (FISH). Positive immunohistochemical staining of wild-type and mutant EGFR was evidenced by complete and strong membranous staining. For FISH analysis, cases were considered unaltered, polysomic, or amplified, as determined by signals of the EGFR gene and chromosome 7. An independent cohort of 107 PC samples was evaluated for mutations in EGFR, KRAS, and BRAF. Protein overexpression was noted in nearly half of the cases and was associated with cancer recurrence (P=.004) and perineural invasion (P=.005). Expression of the 2 mutated EGFR isoforms was not observed. The FISH status was not associated with protein expression. Altered FISH (polysomy and gene amplification) was an independent risk factor for dying of cancer. Only 1 patient of 107 presented KRAS mutations, and no mutations of EGFR or BRAF were observed.
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http://dx.doi.org/10.1016/j.humpath.2016.07.041DOI Listing
March 2017