Arsenic Methyltransferase and Apolipoprotein E Polymorphism in Pregnant Women Exposed to Inorganic Arsenic in Drinking Water in Western Romania.

Previous studies have shown that inorganic arsenic (iAs) exposure may be associated with genotoxic and cytotoxic effects. The aim of this study was to evaluate the relationship between several polymorphisms in AS3MT and APOE genes and urinary As and the relationship between these polymorphisms and pregnancy loss. We determined urinary As concentrations and performed genotyping analysis in 50 cases of spontaneous pregnancy loss and 50 controls, matched to cases on gestational age. The most frequently identified AS3MT polymorphisms in both cases and controls were in rs10748835 (80% cases and 68% controls), rs3740400 (78% cases and 64% controls), rs7085104 (74% cases and 48% controls), and rs1046778 (62% cases and 54% controls). We identified 30 different haplotypes in AS3MT SNPs, with four predominant haplotypes (>8%). Cases with Haplotype 1 had four-fold higher urinary DMA and two-fold higher MMA concentration than those without this haplotype, the MMA levels were lower in cases and controls with Haplotype 4 compared to Haplotype 1, and the DMA levels were significantly lower in cases with Haplotype 4 compared to Haplotype 3. Cases with Haplotype 1 had higher levels of all analyzed biomarkers, suggesting that Haplotype 1 may be associated with greater exposure to iAs and tobacco smoke. Our results suggest the importance of the AS3MT gene in iAs metabolism among pregnant women with low-level drinking water iAs exposure.

Identification of a miRNA Panel with a Potential Determinant Role in Patients Suffering from Periodontitis.

In recent years, the role of microRNA (miRNA) in post-transcriptional gene regulation has advanced and supports strong evidence related to their important role in the regulation of a wide range of fundamental biological processes. Our study focuses on identifying specific alterations of miRNA patterns in periodontitis compared with healthy subjects. In the present study, we mapped the major miRNAs altered in patients with periodontitis (n = 3) compared with healthy subjects (n = 5), using microarray technology followed by a validation step by qRT-PCR and Ingenuity Pathways Analysis. Compared to healthy subjects, 159 differentially expressed miRNAs were identified among periodontitis patients, of which 89 were downregulated, and 70 were upregulated, considering a fold change of ±1.5 as the cut-off value and p ≤ 0.05. Key angiogenic miRNAs (miR-191-3p, miR-221-3p, miR-224-5p, miR-1228-3p) were further validated on a separate cohort of patients with periodontitis versus healthy controls by qRT-PCR, confirming the microarray data. Our findings indicate a periodontitis-specific miRNA expression pattern representing an essential issue for testing new potential diagnostic or prognostic biomarkers for periodontal disease. The identified miRNA profile in periodontal gingival tissue was linked to angiogenesis, with an important molecular mechanism that orchestrates cell fate.

Discovering the Biological Significance and Therapeutic Potential of miR-29b-3p in Triple-Negative Breast Cancer.

The lack of estrogen or progesterone receptors and absence of HER2 amplification/overexpression in triple-negative breast cancer (TNBC) restricts therapeutic options used in clinical management. MicroRNAs (miRNAs) are small, non-coding transcripts which affect important cellular mechanisms by regulating gene expression at the post-transcriptional level. Among this class, attention was focused on miR-29b-3p with a high profile in TNBC and correlated with the overall survival rates, as TCGA data revealed. This study aims to investigate the implication of the miR-29b-3p inhibitor in TNBC cell lines by identifying a potential therapeutic transcript, improving the clinical outcomes of this disease. The experiments were performed on two TNBC cell lines (MDA-MB-231 and BT549) as in vitro models. An established dose of 50 nM was used for all functional assays performed on the miR-29b-3p inhibitor. A decreased level of miR-29b-3p determined a significant reduction in cell proliferation and colony-forming capacity. At the same time, the changes occurring at the molecular and cellular levels were highlighted. We observed that, when inhibiting the expression level of miR-29b-3p, processes such as apoptosis and autophagy were activated. Further, microarray data revealed that the miRNA expression pattern was altered after miR-29b-3p inhibition, pointing out 8 overexpressed and 11 downregulated miRNAs specific for BT549 cells and 33 upregulated and 10 downregulated miRNAs that were specific for MDA-MB-231 cells. As a common signature for both cell lines, three transcripts were observed, two downregulated, miR-29b-3p and miR-29a, and one upregulated, miR-1229-5p. According to DIANA miRPath, the main predicted targets are related to ECM (extracellular matrix) receptor interaction and TP53 signaling. An additional validation step through qRT-PCR was performed, which showed an upregulation of MCL1 and TGFB1. By inhibiting the expression level of miR-29b-3p, it was shown that complex regulatory pathways targeted this transcript in TNBC cells.

Dysregulation of miR-21-5p, miR-93-5p, miR-200c-3p and miR-205-5p in Oral Squamous Cell Carcinoma: A Potential Biomarkers Panel?

Oral squamous cell carcinoma (OSCC) is considered the sixth most common cancer worldwide. To reduce the high mortality of the disease, sensitive and specific diagnostic and prognostic biomarkers are urgently needed. Non-coding RNA, microRNAs (miRNAs), which are short length non-coding transcripts, or long non-coding RNA (lncRNA) seem to be potential biomarkers, considering that they have an important role in regulation of cell fate being involved in a wide range of biological processes. Literature data emphasized the important role of these transcripts as a biomarker for diagnosis and prognosis in oral squamous cell carcinoma. Therefore, we have evaluated the expression levels of a panel of four miRNAs (miR-21-5p, miR-93-5p, miR-200c-3p and miR-205-5p) and H19, MALAT1 by quantitative real-time PCR (qRT-PCR) from 33 fresh frozen tissues and 33 normal adjacent tissues. Our date revealed miR-21-5p and miR-93-5p to be upregulated, while miR-200c-3p and miR-205-5p to be downregulated. Regarding the long non-coding RNAs, H19 and MALAT1, were also downregulated. We also investigated the expression of BCL2, which is another important gene correlated to non-coding RNAs investigated by as, and it was also under-expressed. Additional validation step at protein level was done for KI67, TP53 and BCL2. In our patient cohort no correlation with clinical stage and smoking status was observed. The results of the present study indicated the important role of miR-21-5p, miR-93-5p, miR-200c-3p, miR-205-5p and H19 in OSCC. Differential expression of these transcripts at sub-sites, may serve as a diagnostic marker with further elaboration on a larger sample size. Additional studies should be conducted to confirm the results, particularly the interconnection with coding and non-coding genes.

Alteration of Gene and miRNA Expression in Cervical Intraepithelial Neoplasia and Cervical Cancer.

Background: Cervical cancer is one of the most common malignancies in women in terms of prevalence and mortality. Cervical cancer has some particularities that distinguish it from any other oncologic pathology: first, it is completely preventable by prompt detection of its precursor, cervical intraepithelial neoplasia (CIN); second, the Human Papillomavirus (HPV) infection is a known etiological agent; third, the mean age at diagnosis is much lower than in other oncologic conditions, as a consequence of the sexually-transmitted HPV. Methods: We evaluated the expression level of several long noncoding RNAs and a microRNA in samples from 30 patients with CIN, 9 with cervical cancer and 38 normal samples using qRT-PCR technology. Results: We observed higher expression levels for MEG3, DAPK1, MLH1 and MALAT1 in CIN samples than in normal samples, whereas TIMP3 and SOX1 had lower expression levels. For cancer samples, DAPK1, MLH1 and MALAT1 had higher expression, and MEG3, TIMP3 and SOX1 had lower expression when compared to normal samples. In the case of CIN versus cancer samples, only MEG3 gene showed a statistically significant difference. The expression of miR-205-5p was lower in both CIN and cancer samples compared to normal samples. Conclusion: Decreased MEG3 expression could be considered an alarm signal in the transition from a premalignant cervical lesion to invasive cancer, while altered expression levels of TIMP3, SOX1, MLH1, MALAT1 and miR-205-5p could serve as early biomarkers in the diagnosis of premalignant cervical lesions. Future studies, including a larger number of patients with CIN, will be of particular importance in validating these observations.

Continuous Intravenous Administration of Granulocyte-Colony-Stimulating Factors-A Breakthrough in the Treatment of Cancer Patients with Febrile Neutropenia.

(1) Background: Febrile neutropenia (FN) remains one of the most challenging problems in medical oncology and is a very severe side effect of chemotherapy. Its late consequences, when it is recurrent or of a severe grade, are dose reduction and therapy delays. Current guidelines allow the administration of granulocyte-colony-stimulating factors (G-CSF) for profound FN (except for the case when a pegylated form of G-CSF is administrated with prophylactic intention) in addition to antibiotics and supportive care. (2) Methods: This is a prospective study that included 96 patients with confirmed malignancy, treated with chemotherapy, who developed FN during their oncological therapy, and were hospitalized. They received standard treatment plus a dose of G-CSF of 16 µg/Kg/day IV continuous infusion. (3) Results: The gender distribution was almost symmetrical: Male patients made up 48.96% and 51.04% were female patients, with no significance on recovery from FN (p = 1.00). The patients who received prophylactic G-CSF made up 20.21%, but this was not a predictive or prognostic factor for the recovery time from aplasia (p = 0.34). The median chemotherapy line where patients with FN were included was two and the number of previous chemotherapy cycles before FN was three. The median serological number of neutrophils (PMN) was 450/mm3 and leucocytes (WBC) 1875/mm3 at the time of FN. Ten patients possess PMN less than 100/mm3. The median time to recovery was 25.5 h for 96 included patients, with one failure in which the patient possessed grade 5 FN. Predictive factors for shorter recovery time were lower levels of C reactive protein (p < 0.001) and procalcitonin (p = 0.002) upon hospital admission and higher WBC (p = 0.006) and PMN (p < 0.001) at the time of the provoking cycle of chemotherapy for FN. The best chance for a shorter duration of FN was a short history of chemotherapy regarding the number of cycles) (p < 0.0001). (4) Conclusions: Continuous IV administration of G-CSF could be an alternative salvage treatment for patients with profound febrile neutropenia, with a very fast recovery time for neutrophiles.

New Insights in Gene Expression Alteration as Effect of Paclitaxel Drug Resistance in Triple Negative Breast Cancer Cells.

BACKGROUND/AIMS: Triple negative breast cancer (TNBC) is a highly aggressive form of cancer which lacks targeted therapy options. Thus, TNBC patients have poor outcomes and a decreased survival rate than patients with other types of breast cancers. Due to the lack of surface receptors, TNBC needs a comprehensive investigation to provide more information regarding patient's therapy, as well as to understand the way how to counteract drug resistance mechanisms. Nowadays, chemotherapy remains an unsolved issue which rise a lot of questions in oncology field. METHODS: In this article, we investigated the implication of paclitaxel in TNBC cell lines after a prolong administration, after 12, respectively 24 passages followed by evaluation of morphological alteration, mutational pattern by next generation sequencing and the altered gene expression pattern by microarray technology and validation by qRT-PCR of the resistance to therapy relevant genes. RESULTS: Using functional assays, we showed that paclitaxel exhibits antiproliferative activity on Hs578T/Pax and MDA-MB-231/Pax demonstrating the activation of cell death mechanisms. Confocal microscopy revealed significant modifications which occur in the morphological structure with a disruption of the actin-filaments and also mitotic catastrophe. The presence of these nuclear alterations is due to some modifications at the cellular and molecular levels. Important alterations at the transcriptomic and genomic levels were observed from this a common drug resistance signature (IL-6, CXCL8, VEGFA, EGR1, PTGS2 and TRIB1) for both cell lines at 24 passages was discovered. Also, an important mutation (TP53) linked with drug response was identified. CONCLUSION: These results might be used to furnish novel biomarkers in TNBC, as well as to find a strategy to counteract the resistance to therapy in order to increase survival rate and to enhance the prognosis of patients with TNBC.

New perspectives in triple-negative breast cancer therapy based on treatments with TGFß1 siRNA and doxorubicin.

Triple-negative breast cancer (TNBC), which accounts for 10-20% of all breast cancers, has the worst prognosis. Although chemotherapy treatment is a standard for TNBC, it lacks a specific target. Therefore, new therapeutic strategies are required to be investigated. In this study, a combined doxorubicin (DOX) and small interfering RNA (siRNA) therapy is proposed as therapeutic strategy for targeting TGFß1 gene. Hs578T cell line is used as in vitro model for TNBC, wherein TGFß1siRNA therapy is employed to enhance therapeutic effects. Cell proliferation rate is measured using an MTT test, and morphological alterations are assed using microscopically approached, while gene expression is determined by qRT-PCR analysis. The combined treatment of TGFß1siRNA and DOX reduced levels of cell proliferation and mitochondrial activity and promoted the alteration of cell morphology (dark-field microscopy). DOX treatment caused downregulation of six genes and upregulation of another six genes. The combined effects of DOX and TGFß1siRNA resulted in upregulation of 13 genes and downregulation of four genes. Silencing of TGFß1 resulted in activation of cell death mechanisms in Hs578T cells, to potentiate the effects of DOX, but not in an additive manner, due to the activation of genes involved in resistance to therapy (ABCB1 and IL-6).

Beyond Conventional: The New Horizon of Anti-Angiogenic microRNAs in Non-Small Cell Lung Cancer Therapy.

GLOBOCAN 2018 identified lung cancer as the leading oncological pathology in terms of incidence and mortality rates. Angiogenesis is a key adaptive mechanism of numerous malignancies that promotes metastatic spread in view of the dependency of cancer cells on nutrients and oxygen, favoring invasion. Limitation of the angiogenic process could significantly hamper the disease advancement through starvation of the primary tumor and impairment of metastatic spread. This review explores the basic molecular mechanisms of non-small cell lung cancer (NSCLC) angiogenesis, and discusses the influences of the key proangiogenic factors-the vascular endothelial growth factor-A (VEGF-A), basic fibroblast growth factor (FGF2), several matrix metalloproteinases (MMPs-MMP-2, MMP-7, MMP-9) and hypoxia-and the therapeutic implications of microRNAs (miRNAs, miRs) throughout the entire process, while also providing critical reviews of a number of microRNAs, with a focus on miR-126, miR-182, miR-155, miR-21 and let-7b. Finally, current conventional NSCLC anti-angiogenics-bevacizumab, ramucirumab and nintedanib-are briefly summarized through the lens of evidence-based medicine.

Critical Analysis of Genome-Wide Association Studies: Triple Negative Breast Cancer Quae Exempli Causa.

Genome-wide association studies (GWAS) are useful in assessing and analyzing either differences or variations in DNA sequences across the human genome to detect genetic risk factors of diseases prevalent within a target population under study. The ultimate goal of GWAS is to predict either disease risk or disease progression by identifying genetic risk factors. These risk factors will define the biological basis of disease susceptibility for the purposes of developing innovative, preventative, and therapeutic strategies. As single nucleotide polymorphisms (SNPs) are often used in GWAS, their relevance for triple negative breast cancer (TNBC) will be assessed in this review. Furthermore, as there are different levels and patterns of linkage disequilibrium (LD) present within different human subpopulations, a plausible strategy to evaluate known SNPs associated with incidence of breast cancer in ethnically different patient cohorts will be presented and discussed. Additionally, a description of GWAS for TNBC will be presented, involving various identified SNPs correlated with miRNA sites to determine their efficacies on either prognosis or progression of TNBC in patients. Although GWAS have identified multiple common breast cancer susceptibility variants that individually would result in minor risks, it is their combined effects that would likely result in major risks. Thus, one approach to quantify synergistic effects of such common variants is to utilize polygenic risk scores. Therefore, studies utilizing predictive risk scores (PRSs) based on known breast cancer susceptibility SNPs will be evaluated. Such PRSs are potentially useful in improving stratification for screening, particularly when combining family history, other risk factors, and risk prediction models. In conclusion, although interpretation of the results from GWAS remains a challenge, the use of SNPs associated with TNBC may elucidate and better contextualize these studies.