LncRNA-SNPs in a Brazilian Breast Cancer Cohort: A Case-Control Study.

Long noncoding RNAs (lncRNAs) are a class of non-coding RNAs that contain more than 200 nucleotides and exhibit a versatile regulatory capacity. Genomic alterations in lncRNAs have already been investigated in several complex diseases, including breast cancer (BC). BC is a highly heterogeneous disease and is the most prevalent cancer type among women worldwide. Single nucleotide polymorphisms (SNPs) in lncRNA regions appear to have an important role in BC susceptibility; however, little is known about lncRNA-SNPs in the Brazilian population. This study used Brazilian tumor samples to identify lncRNA-SNPs with a biological role in BC development. We applied a bioinformatic approach intersecting lncRNAs that are differentially expressed in BC tumor samples using The Cancer Genome Atlas (TCGA) cohort data and looked for lncRNAs with SNPs associated with BC in the Genome Wide Association Studies (GWAS) catalog. We highlight four lncRNA-SNPs-rs3803662, rs4415084, rs4784227, and rs7716600-which were genotyped in Brazilian BC samples in a case-control study. The SNPs rs4415084 and rs7716600 were associated with BC development at higher risk. These SNPs were also associated with progesterone status and lymph node status, respectively. The rs3803662/rs4784227 haplotype GT was associated with BC risk. These genomic alterations were also evaluated in light of the lncRNA's secondary structure and gain/loss of miRNA binding sites to better understand its biological functions. We emphasize that our bioinformatics approach could find lncRNA-SNPs with a potential biological role in BC development and that lncRNA-SNPs should be more deeply investigated in a highly heterogeneous disease population.

When guidelines face reality - Lynch syndrome screening in the setting of public health system in a developing country.

Lynch syndrome (LS) is the most common cause of hereditary colorectal cancer (CRC); however, it is still underrecognized and underdiagnosed. While international guidelines gravitate towards universal screening, the underuse of screening methods has been reported in real-world scenarios. This study aims to evaluate screening for LS among patients diagnosed with CRC in a public cancer center in Brazil and evaluate access to genetic counseling and testing for abnormal screens. For that purpose, all patients with CRC registered in our institution from July 2012 to December 2018 had their charts reviewed. Demographic and clinical characteristics were noted, as well as immunohistochemistry and microsatellite instability analysis results, when available. After applying exclusion criteria, a total of 1234 charts were reviewed. Among these, 257 patients were screened for LS, making up a 20.8% screening rate; when considering Jerusalem criteria, screening rate was 24.5%; for Bethesda criteria, it was 35.1%. Almost 80% of patients fulfilling Amsterdam criteria I/II were screened. There were 64 abnormal screens, from which 40 (62.5%) underwent genetic counseling and 12 (18.7%) underwent genetic testing. We concluded that overall screening rates for LS among CRC patients in a public cancer center in Brazil are low, and still very guided by stringent clinical criteria. Referral to genetic counseling and access to testing is limited, calling the whole process into question. Public policies aiming to raise awareness on hereditary cancer and include genetic testing in the public health system could help improve this scenario.

A novel lncRNA derived from an ultraconserved region: lnc-uc.147, a potential biomarker in luminal A breast cancer.

The human genome contains 481 ultraconserved regions (UCRs), which are genomic stretches of over 200 base pairs conserved among human, rat, and mouse. The majority of these regions are transcriptionally active (T-UCRs), and several have been found to be differentially expressed in tumours. Some T-UCRs have been functionally characterized, but of those few have been associated to breast cancer (BC). Using TCGA data, we found 302 T-UCRs related to clinical features in BC: 43% were associated with molecular subtypes, 36% with oestrogen-receptor positivity, 17% with HER2 expression, 12% with stage, and 10% with overall survival. The expression levels of 12 T-UCRs were further analysed in a cohort of 82 Brazilian BC patients using RT-qPCR. We found that uc.147 is high expressed in luminal A and B patients. For luminal A, a subtype usually associated with better prognosis, high uc.147 expression was associated with a poor prognosis and suggested as an independent prognostic factor. The lncRNA from uc.147 (lnc-uc.147) is located in the nucleus. Northern blotting results show that uc.147 is a 2,8 kb monoexonic trancript, and its sequence was confirmed by RACE. The silencing of uc.147 increases apoptosis, arrests cell cycle, and reduces cell viability and colony formation in BC cell lines. Additionally, we identifed 19 proteins that interact with lnc-uc.147 through mass spectrometry and demonstrated a high correlation of lnc-uc.147 with the neighbour gene expression and miR-18 and miR-190b. This is the first study to analyse the expression of all T-UCRs in BC and to functionally assess the lnc-uc.147.

Extracellular vesicles from triple-negative breast cancer cells promote proliferation and drug resistance in non-tumorigenic breast cells.

PURPOSE: Triple-negative breast cancer (TNBC), an aggressive breast cancer subtype, is genetically heterogeneous which challenges the identification of clinically effective molecular makers. Extracellular vesicles (EVs) are key players in the intercellular signaling communication and have been shown to be involved in tumorigenesis. The main goal of this study was to evaluate the role and mechanisms of EVs derived from TNBC cells in modulating proliferation and cytotoxicity to chemotherapeutic agents in non-tumorigenic breast cells (MCF10A). METHODS: EVs were isolated from TNBC cell lines and characterized by nanoparticle tracking analysis, Western blot, and transmission electron microscopy. MCF10A cells were treated with the isolated EVs and evaluated for cell proliferation and cytotoxicity to Docetaxel and Doxorubicin by the MTT and MTS assays, respectively. Gene and miRNA expression profiling was performed in the treated cells to determine expression changes that may be caused by EVs treatment. RESULTS: MCF10A cells treated with HCC1806-EVs (MCF10A/HCC1806-EVs) showed a significant increase in cell proliferation and resistance to the therapeutic agents tested. No significant effects were observed in the MCF10A cells treated with EVs derived from MDA-MB-231 cells. Gene and miRNA expression profiling revealed 138 genes and 70 miRNAs significantly differentially expressed among the MCF10A/HCC1806-EVs and the untreated MCF10A cells, affecting mostly the PI3K/AKT, MAPK, and HIF1A pathways. CONCLUSION: EVs isolated from the HCC1806 TNBC cells are capable of inducing proliferation and drug resistance on the non-tumorigenic MCF10A breast cells, potentially mediated by changes in genes and miRNAs expression associated with cell proliferation, apoptosis, invasion, and migration.

Comparative proteomics of primary breast carcinomas and lymph node metastases outlining markers of tumor invasion.

BACKGROUND: Lymph node metastasis is an important clinicopathological parameter for breast cancer prognostication and treatment. Although the development of metastasis is common in axillary lymph nodes, the mechanisms underlying the locoregional spread are yet poorly understood. In the present study, we outline the involvement of proteins in tumor invasion by comparing the proteome profile of primary breast tumors (PBT) against that of lymph node metastasis (LNM). PATIENTS AND METHODS: The comparative proteome analyses of seven paired samples were performed using two-dimensional gel electrophoresis (2DE) and mass spectrometry (MS). RESULTS: Recurrent proteins were differentially expressed in PBT and LNM across patients. Higher levels of 1433G, 1433T, K2C8, PSME2, SNAA, TPM4, TRFE and VIME were observed in primary tumors compared to the metastatic site. On the other hand, higher levels of ALDH2 and GDIR2 were identified in metastasis related to tumors. These proteins provide a new insight on breast cancer research. CONCLUSION: Our achievements strengthened previous omics-based studies and also support the validation of potential markers of tumor invasion and metastasis.

Allelic variants of XRCC1 and XRCC3 repair genes and susceptibility of oral cancer in Brazilian patients.

BACKGROUND: The capacity for DNA repair is essential in maintaining cellular functions and homeostasis; however, this capacity can be altered based on DNA sequence variations in DNA repair genes, which may contribute to the onset of cancer. Many single-nucleotide polymorphisms (SNPs) in repair genes have been found to be associated with oral cancer. The aim of this study was to investigate the relationship between the presence of allelic variants Arg194Trp (rs:1799782) and Arg399Gln (rs: 25487) of XRCC1 gene and Thr241Met (rs: 861539) of XRCC3 gene and susceptibility to oral cancer. We also attempted to correlate the frequencies obtained for each of the SNPs to histopathological parameters. METHODS: A case-control study was conducted with genomic DNA from 150 patients with oral squamous cell carcinomas and 150 controls. SNPs were genotyped by RFLP-PCR. RESULTS: The presence of the polymorphic variants of the XRCC1 gene within codon 194 (OR 0.82, 95% CI: 0.44-1.51) and codon 399 (OR 0.94, 95% CI: 0.59-1.50) and within the XRCC3 gene (OR 0.72; 95% CI: 0.45-1.16) were not associated with an increased risk of oral cancer. A combinational analysis of SNPs in both genes indicated no association. The presence of the allelic variants of these two genes had no statistically significant effect on tumor differentiation, lymph node invasion or tumor size. CONCLUSIONS: These results suggest that allelic variants of XRCC1 and XRCC3 are not suitable markers for susceptibility to carcinomas of the oral cavity and are also not related to the later stages of such tumors.

Differential loss of heterozygosity profile on chromosome 3p in ductal and lobular breast carcinomas.

The 2 main histologic types of infiltrating breast cancer, invasive lobular and invasive ductal carcinoma, are morphologically and clinically distinct. Studies revealed that different patterns of gene expression and loss of heterozygosity can also distinguish these 2 subtypes. A whole-genome study using single nucleotide polymorphism array found a significantly higher frequency of loss of heterozygosity on 3p in invasive ductal carcinoma when compared with invasive lobular carcinoma. In this study, we performed a loss of heterozygosity analysis of the 3p chromosome region in ductal and lobular breast tumors. Seven microsatellite markers were evaluated in a series of 136 sporadic breast cancer cases (118 invasive ductal carcinoma and 18 invasive lobular carcinoma) and correlated with clinical-histopathologic parameters from the patients. A significantly higher frequency of loss of heterozygosity was observed in invasive ductal carcinoma (65.3%) when compared with invasive lobular carcinoma (38.9%). When the markers were analyzed separately, loss of heterozygosity at 3 of them, D3S1307 in 3p26.3, D3S1286 in 3p24.3, and D3S1300 in 3p14.2, were significantly more frequent in ductal than in lobular tumors. D3S1307 marker showed the highest frequency of loss of heterozygosity in invasive ductal carcinoma (46.2%), and associations between loss of this marker and loss of estrogen and progesterone receptors were found in these samples. Our results confirm the observations that invasive ductal carcinoma has a higher frequency of loss of heterozygosity events across the 3p region than invasive lobular carcinoma and show that specific losses on 3p26.3, 3p24.3, and 3p14.2 regions are more frequent in ductal than in lobular tumors. We discuss our data in relation to the known tumor suppressor genes that are mapped at the 3p loci investigated and their present relevant roles in breast cancer.

Association between the STin2 VNTR polymorphism and smoking behavior in oral cancer patients and healthy individuals.

The serotonergic system may be involved in smoking behavior since the intake of nicotine increases serotonin secretion in the CNS. Moreover, evidence supporting the beneficial effect of selective serotonin reuptake for quitting smoking suggesting that the serotonin transporter (5-HTT) is a plausible target for the understanding and elucidation of smoking behavior. The transcriptional activity of its human gene (SLC6A4) is modulated by a polymorphism described in the second intron, the STin2 VNTR, which thus may interfere with 5-HTT synthesis. In this study was analyzed the polymorphism STin2 VNTR of 60 smokers male patients diagnosed for oral carcinoma, 61 male smokers without cancer and 65 non-smoker healthy blood donors. The STin2. 9 allele carriers were more present in smoker groups (with cancer and without cancer, respectively) than in the non-smoker (OR = 7.11, 95% CI = 0.83-60.91 and OR = 24.73; IC 95% = 3.17-192.66). Conversely, individuals carrying allele 10 were more prevalent in non-smokers compared with smokers (oral cancer patients and individuals without cancer, respectively), showing a protective factor of this allele (OR = 0.56; 95% CI = 0.24-1.33 and OR = 0.46; 95% CI = 0.20-1.07). This is the first report of a study assessing the importance of STin2 VNTR smoking behavior in Brazilian individuals and the association of STin2. 9 allele carriers in nicotine dependence. It is suggested that individuals with low serotonin concentration in the central nervous system, probably due to the presence of the allele for high expression of 5-HTT,especially STin2. 9, were more susceptible to nicotine dependence. Moreover, individuals with the 10 allele might have less risk for nicotine dependence.

Concomitant loss of heterozygosity at the BRCA1 and FHIT genes as a prognostic factor in sporadic breast cancer.

Previous studies have shown loss of heterozygosity (LOH) at the BRCA1 and FHIT genes in sporadic primary breast cancer. The aim of this study was to evaluate concomitant LOH at the BRCA1 and FHIT genes in sporadic breast cancer and investigate its influence on patient survival. Loss of heterozygosity was determined using microsatellite markers. The analysis on the informative cases (n = 72) indicated LOH at both the BRCA1 and FHIT loci in 25 cases (35%), the absence of LOH at both loci in 23 cases (32%), and the presence of LOH at one of the loci in 24 cases (33%). The concomitant LOH was associated with poor prognostic factors, such as large tumors (P = 0.01), axillary nodal involvement (P < 0.01), histologic grade III (P < 0.01), vascular invasion (P = 0.01), and negative hormone receptor (P = 0.02). After a median follow-up period of 48 months, the concomitant LOH group had the shortest survival (P < 0.02 by log-rank test; P < 0.05 by Cox model; hazard ratio of 4.87), compared with patients without LOH. These data suggest that concomitant allelic losses of the BRCA1 and FHIT genes are associated with more aggressive breast tumors.

Meta-analysis and pooled analysis of GSTM1 and CYP1A1 polymorphisms and oral and pharyngeal cancers: a HuGE-GSEC review.

The association of GSTM1 and CYP1A1 polymorphisms and oral and pharyngeal cancers was assessed through a meta-analysis of published case-control studies and a pooled analysis of both published and unpublished case-control studies from the Genetic Susceptibility to Environmental Carcinogens database (http://www.upci.upmc.edu/research/ccps/ccontrol/index.html ). Thirty publications used in the meta-analysis included a total of 7783 subjects (3177 cases and 4606 controls); 21 datasets, 9397 subjects (3130 cases and 6267 controls) were included in the pooled analysis. The GSTM1 deletion was 2-fold more likely to occur in African American and African cases than controls (odds ratio: 1.7, 95% confidence interval: 0.9-3.3), although this was not observed among whites (odds ratio: 1.0, 95% confidence interval: 0.9-1.1). The meta-analysis and pooled analysis showed a significant association between oral and pharyngeal cancer and the CYP1A1 MspI homozygous variant (meta-ORm2/m2: 1.9, 95% confidence interval: 1.4-2.7; Pooled ORm2m2: 2.0, 95% confidence interval: 1.3-3.1; ORm1m2 or [infi]m2m2: 1.3, 95% confidence interval: 1.1-1.6). The association was present for the CYP1A1 (exon 7) polymorphism (ORVal/Val: 2.2, 95% confidence interval: 1.1-4.5) in ever smokers. A joint effect was observed for GSTM1 homozygous deletion and the CYP1A1 m1m2 variant on cancer risk. Our findings suggest that tobacco use and genetic factors play a significant role in oral and pharyngeal cancer.

High frequency of t(12;21)(p13;q22) in children with acute lymphoblastic leukemia and known clinical outcome in southern Brazil.

The presence of the t(12;21)(p13;q22) distinguishes a subset of children with acute lymphoblastic leukemia (ALL) that present a favorable prognosis. This is a cryptic translocation difficult to detect through conventional cytogenetics. In this study, bone marrow samples from 30 children with ALL from southern Brazil were evaluated by fluorescence in situ hybridization (FISH) for the t(12;21), using locus specific probes to detect the TEL/AML1 rearrangement. The selection criteria included: age (0-12 years old); FAB classification (L1 or L2), absence of specific clonal chromosomal aberrations; and adequate cellular integrity to perform FISH analysis. A frequency of 40% of the t(12;21) was observed, in addition to extra copies of the AML1 gene in 7.5% of patients. These findings were analyzed in relation to the patient's clinical parameters and compared with other pediatric populations.