MicroRNAs as possible indicators of drug sensitivity in breast cancer cell lines.

MicroRNAs (miRNAs) regulate gene expression post-transcriptionally. In this way they might influence whether a cell is sensitive or resistant to a certain drug. So far, only a limited number of relatively small scale studies comprising few cell lines and/or drugs have been performed. To obtain a broader view on miRNAs and their association with drug response, we investigated the expression levels of 411 miRNAs in relation to drug sensitivity in 36 breast cancer cell lines. For this purpose IC50 values of a drug screen involving 34 drugs were associated with miRNA expression data of the same breast cancer cell lines. Since molecular subtype of the breast cancer cell lines is considered a confounding factor in drug association studies, multivariate analysis taking subtype into account was performed on significant miRNA-drug associations which retained 13 associations. These associations consisted of 11 different miRNAs and eight different drugs (among which Paclitaxel, Docetaxel and Veliparib). The taxanes, Paclitaxel and Docetaxel, were the only drugs having miRNAs in common: hsa-miR-187-5p and hsa-miR-106a-3p indicative of drug resistance while Paclitaxel sensitivity alone associated with hsa-miR-556-5p. Tivantinib was associated with hsa-let-7d-5p and hsa-miR-18a-5p for sensitivity and hsa-miR-637 for resistance. Drug sensitivity was associated with hsa-let-7a-5p for Bortezomib, hsa-miR-135a-3p for JNJ-707 and hsa-miR-185-3p for Panobinostat. Drug resistance was associated with hsa-miR-182-5p for Veliparib and hsa-miR-629-5p for Tipifarnib. Pathway analysis for significant miRNAs was performed to reveal biological roles, aiding to find a potential mechanistic link for the observed associations with drug response. By doing so hsa-miR-187-5p was linked to the cell cycle G2-M checkpoint in line with this checkpoint being the target of taxanes. In conclusion, our study shows that miRNAs could potentially serve as biomarkers for intrinsic drug resistance and that pathway analyses can provide additional information in this context.

Antioxidant activity of CAPE (caffeic acid phenethyl ester) in vitro can protect human sperm deoxyribonucleic acid from oxidative damage.

PURPOSE: Sperm processing (e.g., centrifugation) used in preparation for assisted reproduction can result in excessive generation of reactive oxygen species (ROS) and potential sperm damage. The use of antioxidants during sperm processing has been shown to prevent iatrogenic sperm damage, including DNA damage. In this study, we evaluated the effect of caffeic acid phenethyl ester (CAPE) on oxidative stress mediated sperm dysfunction and DNA damage. METHODS: Semen samples were obtained to liquefy at room temperature. After centrifugation and washing protocols, spermatozoa were incubated in a single step supplemented medium with either of 10, 50 or 100 µmol/L CAPE for 2 hours at 36 °C. After incubation period, MDA levels of seminal plasma were measured. The fragmentation in sperm DNA was detected by light microscopy via use of an aniline blue assay, while ultrastructural morphology was analyzed by transmission electron microscopy. RESULTS: Significant increase has been observed in percent chromatin condensation (assessed by aniline blue staining) and Malondialdehyde (Mmol/L) in oligoasthenoteratozoospermia group before the centrifugation (0.57 ±â€¯0.15). Incubation of samples with 100 µmol/L CAPE after centrifugation resulted in a significantly lower percent chromatin condensation compared to samples incubated without CAPE (0.42 ±â€¯0.12) (P < 0.0033). Incubation of all samples with CAPE (10 µmol/L, 50 µmol/L, 100 µmol/L.) after centrifugation resulted in a significantly lower percentage of Malondialdehyde levels. CONCLUSIONS: The data suggests that preincubation of spermatozoa with the antioxidant CAPE offers protection against oxidative DNA damage in vitro.

Understanding drugs in breast cancer through drug sensitivity screening.

With substantial numbers of breast tumors showing or acquiring treatment resistance, it is of utmost importance to develop new agents for the treatment of the disease, to know their effectiveness against breast cancer and to understand their relationships with other drugs to best assign the right drug to the right patient. To achieve this goal drug screenings on breast cancer cell lines are a promising approach. In this study a large-scale drug screening of 37 compounds was performed on a panel of 42 breast cancer cell lines representing the main breast cancer subtypes. Clustering, correlation and pathway analyses were used for data analysis. We found that compounds with a related mechanism of action had correlated IC50 values and thus grouped together when the cell lines were hierarchically clustered based on IC50 values. In total we found six clusters of drugs of which five consisted of drugs with related mode of action and one cluster with two drugs not previously connected. In total, 25 correlated and four anti-correlated drug sensitivities were revealed of which only one drug, Sirolimus, showed significantly lower IC50 values in the luminal/ERBB2 breast cancer subtype. We found expected interactions but also discovered new relationships between drugs which might have implications for cancer treatment regimens.

Effects of aluminum chloride on some trace elements and erythrocyte osmotic fragility in rats.

Aluminum (Al) is a nonessential, toxic element to which humans are constantly exposed as a result of an increase in industrialization and improving technology practices. The aim of the study was to investigate the effects of different durations and doses of Al exposure on serum and tissue element levels and erythrocyte osmotic fragility in rats. A total of 40 male Wistar Albino rats were divided into five groups: control, group I (3 weeks, 8 mg/kg), group II (6 weeks, 8 mg/kg), group III (3 weeks, 16 mg/kg), and group IV (6 weeks, 16 mg/kg). Al chloride (AlCl3) was injected intraperitoneally (i.p.) five times a week. At the end of the experimental period, levels of Al, iron (Fe), copper (Cu), and zinc (Zn) in serum, liver, and kidney tissues were measured using inductively coupled plasma optical emission spectrometry. Osmotic fragility was determined using a spectrophotometer. The results of the experiment indicate that Al induced a statistically significant increase in Al and Fe concentrations in liver and serum as well as in Cu in the kidney. The Fe concentration in serum and kidney tissues was significantly lower in all the groups. As a result of our study, it may be concluded that tissue Al accumulation may lead to an increase in osmotic fragility of erythrocytes and abnormal trace element levels.

CHEK2*1100delC homozygosity in the Netherlands--prevalence and risk of breast and lung cancer.

The 1100delC mutation in the CHEK2 gene has a carrier frequency of up to 1.5% in individuals from North-West Europe. Women heterozygous for 1100delC have an increased breast cancer risk (odds ratio 2.7). To explore the prevalence and clinical consequences of 1100delC homozygosity in the Netherlands, we genotyped a sporadic breast cancer hospital-based cohort, a group of non-BRCA1/2 breast cancer families, and breast tumors from a tumor tissue bank. Three 1100delC homozygous patients were found in the cohort of 1434 sporadic breast cancer patients, suggesting an increased breast cancer risk for 1100delC homozygotes (odds ratio 3.4, 95% confidence interval 0.4-32.6, P=0.3). Another 1100delC homozygote was found in 592 individuals from 108 non-BRCA1/2 breast cancer families, and two more were found after testing 1706 breast tumors and confirming homozygosity on their wild-type DNA. Follow-up data was available for five homozygous patients, and remarkably, three of them had developed contralateral breast cancer. A possible relationship between 1100delC and lung cancer risk was investigated in 457 unrelated lung cancer patients but could not be confirmed. Due to the small number of 1100delC homozygotes identified, the breast cancer risk estimate associated with this genotype had limited accuracy but is probably higher than the risk in heterozygous females. Screening for CHEK2 1100delC could be beneficial in countries with a relatively high allele frequency.

miRNA expression profiling of 51 human breast cancer cell lines reveals subtype and driver mutation-specific miRNAs.

INTRODUCTION: Breast cancer is a genetically and phenotypically complex disease. To understand the role of miRNAs in this molecular complexity, we performed miRNA expression analysis in a cohort of molecularly well-characterized human breast cancer cell lines to identify miRNAs associated with the most common molecular subtypes and the most frequent genetic aberrations. METHODS: Using a microarray carrying LNA™ modified oligonucleotide capture probes), expression levels of 725 human miRNAs were measured in 51 breast cancer cell lines. Differential miRNA expression was explored by unsupervised cluster analysis and was then associated with the molecular subtypes and genetic aberrations commonly present in breast cancer. RESULTS: Unsupervised cluster analysis using the most variably expressed miRNAs divided the 51 breast cancer cell lines into a major and a minor cluster predominantly mirroring the luminal and basal intrinsic subdivision of breast cancer cell lines. One hundred and thirteen miRNAs were differentially expressed between these two main clusters. Forty miRNAs were differentially expressed between basal-like and normal-like/claudin-low cell lines. Within the luminal-group, 39 miRNAs were associated with ERBB2 overexpression and 24 with E-cadherin gene mutations, which are frequent in this subtype of breast cancer cell lines. In contrast, 31 miRNAs were associated with E-cadherin promoter hypermethylation, which, contrary to E-cadherin mutation, is exclusively observed in breast cancer cell lines that are not of luminal origin. Thirty miRNAs were associated with p16INK4 status while only a few miRNAs were associated with BRCA1, PIK3CA/PTEN and TP53 mutation status. Twelve miRNAs were associated with DNA copy number variation of the respective locus. CONCLUSION: Luminal-basal and epithelial-mesenchymal associated miRNAs determine the subdivision of miRNA transcriptome of breast cancer cell lines. Specific sets of miRNAs were associated with ERBB2 overexpression, p16INK4a or E-cadherin mutation or E-cadherin methylation status, which implies that these miRNAs may contribute to the driver role of these genetic aberrations. Additionally, miRNAs, which are located in a genomic region showing recurrent genetic aberrations, may themselves play a driver role in breast carcinogenesis or contribute to a driver gene in their vicinity. In short, our study provides detailed molecular miRNA portraits of breast cancer cell lines, which can be exploited for functional studies of clinically important miRNAs.