Correlating Basal Gene Expression across Chemical Sensitivity Data to Screen for Novel Synergistic Interactors of HDAC Inhibitors in Pancreatic Carcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal malignancies. Development of the chemoresistance in the PDAC is one of the key contributors to the poor survival outcomes and the major reason for urgent development of novel pharmacological approaches in a treatment of PDAC. Systematically tailored combination therapy holds the promise for advancing the treatment of PDAC. However, the number of possible combinations of pharmacological agents is too large to be explored experimentally. In respect to the many epigenetic alterations in PDAC, epigenetic drugs including histone deacetylase inhibitors (HDACi) could be seen as the game changers especially in combined therapy settings. In this work, we explored a possibility of using drug-sensitivity data together with the basal gene expression of pancreatic cell lines to predict combinatorial options available for HDACi. Developed bioinformatics screening protocol for predictions of synergistic drug combinations in PDAC identified the sphingolipid signaling pathway with associated downstream effectors as a promising novel targets for future development of multi-target therapeutics or combined therapy with HDACi. Through the experimental validation, we have characterized novel synergism between HDACi and a Rho-associated protein kinase (ROCK) inhibitor RKI-1447, and between HDACi and a sphingosine 1-phosphate (S1P) receptor agonist fingolimod.

Synthesis, characterization, and in vivo evaluation of the anticancer activity of a series of 5- and 6-(halomethyl)-2,2'-bipyridine rhenium tricarbonyl complexes.

We report the synthesis, characterization, and in vivo evaluation of the anticancer activity of a series of 5- and 6-(halomethyl)-2,2'-bipyridine rhenium tricarbonyl complexes. The study was promoted in order to understand if the presence and position of a reactive halomethyl substituent on the diimine ligand system of fac-[Re(CO)3]+ species may be a key molecular feature for the design of active and non-toxic anticancer agents. Only compounds potentially able to undergo ligand-based alkylating reactions show significant antiproliferative activity against colorectal and pancreatic cell lines. Of the new species presented in this study, one compound (5-(chloromethyl)-2,2'-bipyridine derivative) shows significant inhibition of pancreatic tumour growth in vivo in zebrafish-Panc-1 xenografts. The complex is noticeably effective at 8 µM concentration, lower than its in vitro IC50 values, being also capable of inhibiting in vivo cancer cells dissemination.

Methylenetetrahydrofolate reductase polymorphic variants C677T and A1298C in rectal cancer in Slavic population: significance for cancer risk and response to chemoradiotherapy.

Background: Methylenetetrahydrofolate reductase (MTHFR) single nucleotide polymorphisms (SNPs) have been suggested as risk, prognostic, and predictive factors for colorectal cancer in various populations, but have not been validated so far. The aim of this study was to examine the association of MTHFR C677T (rs1801133) and A1298C (rs1801131) single nucleotide polymorphisms with the risk of rectal cancer as well as the response to neoadjuvant chemoradiotherapy (nCRT) based on 5-Fluorouracil (5-FU)/leucovorin (LV) in the locally advanced setting. Patients and methods: This case-control study included 119 healthy controls and 97 patients with locally advanced rectal cancer (LARC). For MTHFR genotyping, restriction fragment length polymorphism analysis (PCR-RFLP) was employed. Results: In silico analysis highlighted that SNPs C677T and A1298T correlate with MTHFR gene expression, and that gene expression profile correlates with cancer risk and stage. Using dominant and recessive models, it was found that the MTHFR 677CC vs. 677CT+677TT have increased risk of cancer development (odds ratio (OR): 2.27; 95% confidence interval (CI): 1.30-3.95, p = 0.002) as well as 677CC+677CT compared to 677TT (OR: 4.18, 95% CI: 1.16-14.99, p = 0.014). MTHFR 1298AA also shown increased risk for cancer development compared to 1298AC+1298CC (OR:2.0, 95% CI: 1.20-3.59, p = 0.035) Statistical analysis of combined genotypes highlighted the protective role of CT/AC combined genotype (OR: 3.15 95% CI: 1.576-6.279, p = 0.002) while the CC/AA genotype showed an increased risk for rectal cancer development (OR: 2.499, 95% CI: 1.246-5.081, p = 0.016) The carriers of the 677C/1298A haplotype had the highest risk for developing rectal cancer (OR: 1.74; 95% CI: 1.198-2.530, p = 0.002) while the 677T/1298C haplotype seems to provide a protective effect. (OR: 0.44; 95%CI 0.248-0.795, p = 0.003). No significant association with response to chemoradiotherapy was found. Conclusion: Our data point to MTHFR 667C allele and 1298A alleles as low-penetrance risk factors for rectal cancer in our population. To the best of our knowledge, this is the first study of this type performed on the Slavic population in the Western Balkan, as various population-based factors might also be significant our findings can be used for future meta-analyses and the construction of genetic cancer risk prediction panels.

Discovery of 1-Benzhydryl-Piperazine-Based HDAC Inhibitors with Anti-Breast Cancer Activity: Synthesis, Molecular Modeling, In Vitro and In Vivo Biological Evaluation.

Isoform-selective histone deacetylase (HDAC) inhibition is promoted as a rational strategy to develop safer anti-cancer drugs compared to non-selective HDAC inhibitors. Despite this presumed benefit, considerably more non-selective HDAC inhibitors have undergone clinical trials. In this report, we detail the design and discovery of potent HDAC inhibitors, with 1-benzhydryl piperazine as a surface recognition group, that differ in hydrocarbon linker. In vitro HDAC screening identified two selective HDAC6 inhibitors with nanomolar IC50 values, as well as two non-selective nanomolar HDAC inhibitors. Structure-based molecular modeling was employed to study the influence of linker chemistry of synthesized inhibitors on HDAC6 potency. The breast cancer cell lines (MDA-MB-231 and MCF-7) were used to evaluate compound-mediated in vitro anti-cancer, anti-migratory, and anti-invasive activities. Experiments on the zebrafish MDA-MB-231 xenograft model revealed that a novel non-selective HDAC inhibitor with a seven-carbon-atom linker exhibits potent anti-tumor, anti-metastatic, and anti-angiogenic effects when tested at low micromolar concentrations.

Synthesis, characterization, antitumor potential, and investigation of mechanism of action of copper(ii) complexes with acylpyruvates as ligands: interactions with biomolecules and kinetic study.

Considering the urgency of finding a cure for vicious diseases such as tumors, we have synthesized and characterized a small series of new copper(ii) complexes with biologically important ligands such as acylpyruvate. In addition to this, we used another four copper(ii) complexes, with ligands of the same type to examine the antitumor potential. The antitumor potential of the copper(ii) complexes was examined on three tumor cell lines and one normal human cell line using the MTT assay. All seven tested complexes showed very good cytotoxic effects. Two copper complexes that showed the best antitumor potential were selected for further testing that showed the best potential for potential application in the future. The mechanism of activity of these complexes was examined in detail using tests such as cell cycle, ROS level, oxidative DNA damage, and proteins related to hypoxia analysis. In addition, we examined the binding abilities of these complexes with biomolecules (Guo, Ino, 5'-GMP, BSA, and DNA). The results showed that the tested compounds bind strongly to DNA molecules through intercalation. Also, it has been shown that the tested compounds adequately bind to the BSA molecule, which indicates an even greater potential for some future application of these compounds in clinical practice.

Association Between Oxidative Stress and Melanoma Progression.

BACKGROUND: Overproduction of free radicals accompanied with their insufficient removal/neutralization by antioxidative defense system impairs redox hemostasis in living organisms. Oxidative stress has been shown to be involved in all the stages of carcinogenesis and malignant melanocyte transformation. The aim of this study was to examine association between oxidative stress development and different stages of melanoma. METHODS: The measured oxidative stress parameters included: superoxide anion radical, total and manganese superoxide dismutase, catalase and malondialdehyde. Oxidative stress parameters were measured spectrophotometrically in serum samples from melanoma patients (n=72) and healthy control subjects (n=30). Patients were classified according to AJCC clinical stage. RESULTS: Average superoxide anion and malondialdehyde concentrations were significantly higher in melanoma patients than in control group, with the highest value of superoxide anion in stage III, while malondialdehyde highest value was in stage IV. The activity of total and manganese superoxide dismutase was insignificantly higher in melanoma patients than in control group, while catalase activity was significantly higher. The highest activity of total activity of manganese superoxide dismutase was in stage IV. Catalase activity was increasing with the disease progression achieving the maximum in stage III. CONCLUSION: Results of our study suggest that melanoma is oxidative stress associated disease, as well as deteriorated cell functioning at mitochondrial level.

In vitro antioxidant activity of thiazolidinone derivatives of 1,3-thiazole and 1,3,4-thiadiazole.

The initial steps in preclinical drug developing research concern the synthesis of new compounds for specific therapeutic use which needs to be confirmed by in vitro and then in vivo testing. Nine thiazolidinone derivatives (numerically labeled 1-9) classified as follows: 1,3-thiazole-based compounds (1 and 2); 1,3,4-thiadiazole based compounds (3 and 4); substituted 5-benzylideno-2-adamantylthiazol[3,2-b][1,2,4]triazol-6(5H)ones (5-8); and an ethylaminothiazole-based chalcone (9), were tested for antioxidant activity (AOA) by using three in vitro assays: DPPH (1,1-diphenyl-2-picrylhydrazyl scavenging capacity test); FRAP (ferric reducing antioxidant power test); and TBARS (thiobarbituric acid reactive substances test). Compounds 1-4 and 9 in particular are newly synthesized compounds. Also, traditional antioxidants Vitamins E and C and α-lipoic acid (α-LA) were tested. The results of DPPH testing: Vitamin C 94.35%, Vitamin E 2.99% and α-LA 1.57%; compounds: 4 33.98%; 2 18.73%; 1 15.62%; 5 6.59%; 3 4.99%; 6-9 demonstrated almost no AOA. The results of TBARS testing (% of LPO inhibition): Vitamin C 62.32%; Vitamin E 36.29%; α-LA 51.36%; compounds: 1 62.11%; 5 66.71%; 9 60.93%; 4, 6 and 7 demonstrated ∼50%; 3 and 8 displayed ∼38%; 2 23.51%. By FRAP method, Vitamins E and C showed equal AOA, ∼100%, unlike α-LA (no AOA), and AOA of the tested compounds (expressed as a fraction of the AOA of Vitamin C) were: 2 and 4-75%; 8, 3 and 1-45%; 5-7 and 9-27%. Different red-ox reaction principles between these assays dictate different AOA outcomes for a single compound. Vitamin C appeared to be the superior antioxidant out of the traditional antioxidants; and compound 4 was superior to other tested thiazolidinone derivatives. Vitamin C appeared to be the superior antioxidant out of the traditional antioxidants; and compound 4 was superior to other tested thiazolidinone derivatives. Phenyl-functionalized benzylidene, amino-carbonyl functional domains and chelating ligand properties of the thiazolidinone derivatives correlated with AOA.

Subacute alcohol and/or disulfiram intake affects bioelements and redox status in rat testes.

The aim of the study was to investigate if alcohol and disulfiram (DSF) individually and in combination affect bioelements' and red-ox homeostasis in testes of the exposed rats. The animals were divided into groups according to the duration of treatments (21 and/or 42 days): C21/C42 groups (controls); OL21 and OL22-42 groups (0.5 mL olive oil intake); A1-21 groups (3 mL 20% ethanol intake); DSF1-21 groups (178.5 mg DSF/kg/day intake); and A21+DSF22-42 groups (the DSF ingestion followed previous 21 days' treatment with alcohol). The measured parameters in testes included metals: zinc (Zn), copper (Cu), iron (Fe), magnesium (Mg) and selenium (Se); as well as oxidative stress (OS) parameters: superoxide anion radical (O2•-), glutathione reduced (GSH) and oxidized (GSSG), malondialdehyde (MDA), hydrogen peroxide (H2O2) decomposition and activities of total superoxide dismutase (tSOD), glutathione-S-transferase (GST) and glutathione reductase (GR). Metal status was changed in all experimental groups (Fe rose, Zn fell, while Cu increased in A21+DSF24-32 groups). Development of OS was demonstrated in A1-21 groups, but not in DSF1-21 groups. In A21+DSF22-42 groups, OS was partially reduced compared to A groups (A1-21>MDA>C; A1-21

Disulfiram moderately restores impaired hepatic redox status of rats subchronically exposed to cadmium.

Examination of cadmium (Cd) toxicity and disulfiram (DSF) effect on liver was focused on oxidative stress (OS), bioelements status, morphological and functional changes. Male Wistar rats were intraperitoneally treated with 1 mg CdCl2/kg BW/day; orally with 178.5 mg DSF/kg BW/day for 1, 3, 10 and 21 days; and co-exposed from 22nd to 42nd day. The co-exposure nearly restored previously suppressed total superoxide dismutase (SOD), catalase (CAT) and increased glutathione peroxidase (GPx) activities; increased previously reduced glutathione reductase (GR) and total glutathione-S-transferase (GST) activities; reduced previously increased superoxide anion radical (O2·-) and malondialdehyde (MDA) levels; increased zinc (Zn) and iron (Fe), and decreased copper (Cu) (yet above control value), while magnesium (Mg) was not affected; and decreased serum alanine aminotransferases (ALT) levels. Histopathological examination showed signs of inflammation process as previously demonstrated by exposure to Cd. Overall, we ascertained partial liver redox status improvement, compared with the formerly Cd-induced impact.

Antioxidant activity and protective role on protein glycation of synthetic aminocoumarins

Background: Synthesized aminocoumarins are heterocyclic compounds possessing potential for the treatment of insulin-dependent diabetes mellitus with unexplored anti-glycative action. Results: In this study 4-aminocoumarin derivatives (4-ACDs) were evaluated in vitro for antiglycation (AG) activities by using the human serum albumin (HSA)/glucose system, for 8 weeks of incubation. The glycation and conformational alteration of HSA in the presence of the tested compounds were evaluated by Congo red assay, fluorescence and circular dichroism spectroscopy. The antioxidant (AO) capacity were also tested by four different assays including: DPPH (2,2'-diphenyl-1-picrylhydrazyl radical), ABTS (2,2-azinobis (3-ethylbenzothiazoline-6-sulphonate) diammonium salt), FRAP (ferric reducing antioxidant power) and β-carotene-linoleic acid assay. The tested compounds showed AG and AO effects. The intensity of the accomplished AO potential is related to the type of the used assay. Significant alterations in the secondary (monitored by CD spectropolarimetry) and tertiary structure (assessed by spectrofluorimetry) of HSA upon glycation were mitigated by the 4-ACDs, suggesting their suppressive role in the late stage (post-Amadori) of the HSA glycation. Conclusions: By the analogues, in vitro ascertained AO and AG properties of 4-ACD may be recognized as rationale for their protective role against oxidative changes of proteins, thereby precluding diabetic complications in humans.

Transcranial theta-burst stimulation alters GLT-1 and vGluT1 expression in rat cerebellar cortex.

Repetitive transcranial magnetic stimulation (rTMS) induces changes in expression of proteins engaged in activity of excitatory and inhibitory systems as well as redox homeostasis. Our aim was to investigate the effect of single (SS) and repeated session (RS) of intermittent and continuous theta-burst stimulation (iTBS; cTBS) on the expression of vesicular and plasmatic glutamate transporters 1 (vGluT1 and GLT-1), glial fibrillary acidic protein (GFAP) and influence on oxidative status in rats cerebellar tissue and plasma. Redox state parameters in cerebellar tissue and plasma were assessed 24 h after single and 48 h after the last TBS session. Molecular changes were examined by immunofluorescence. Stimulation significantly increased thiol groups (SH) in tissue of SS iTBS group, and decreased in iTBS RS. Activity of glucose-6-phosphate-dehydrogenase (G6PD) was increased markedly in cTBS RS. Immunoreactivity of vGluT1 in cTBS RS decreased, while GLT-1 increased in cTBS SS and cTBS RS, compared to control. Present study gives insight in molecular and biochemical mechanisms by which iTBS and cTBS exerts its effects on rats cerebellar cortex.

Oxidative stress, bioelements and androgen status in testes of rats subacutely exposed to cadmium.

The objective of our study was to examine testicular toxicity of cadmium (Cd), focusing on oxidative stress (OS), essential metals and androgenic status and morphological changes. Male Wistar rats [controls and four Cd-subgroups (n = 6) organized according to the exposure (1, 3, 10 and 21 days)] were intraperitoneally (i.p.) treated with 1 mg CdCl2/kg/day. Testicular Cd deposition was noticed from the 1st day. After 10 and 21 days, copper (Cu) and iron (Fe) increased by 60-109% and 43-67%, respectively, while zinc (Zn) decreased by 24-33%. During 1-21 days of the exposure, decrease in testicular total superoxide dismutase (SOD) and total glutathione-s-transferase (GST) activities occurred gradually by 30-78% and 15-84%, respectively, while superoxide anion radical (O2(-)) increased gradually by 114-271%. After 10-21 days, decrease in testicular catalase (CAT) activity appeared by 13-31%. After 21 days, malondialdehyde (MDA) decreased by 44% and the ratio of oxidized glutathione/reduced glutathione (GSSG/GSH) increased by 130% in testes of the rats exposed to Cd. Additionally, decreased testicular testosterone level and the relative testes mass, along with induced microscopic and macroscopic changes were occured, what can be explained as the consequence of instantly developed OS, impaired essential metals status and Cd testicular deposition.

Robust fault detection of turbofan engines subject to adaptive controllers via a Total Measurable Fault Information Residual (ToMFIR) technique.

This paper provides a new design of robust fault detection for turbofan engines with adaptive controllers. The critical issue is that the adaptive controllers can depress the faulty effects such that the actual system outputs remain the pre-specified values, making it difficult to detect faults/failures. To solve this problem, a Total Measurable Fault Information Residual (ToMFIR) technique with the aid of system transformation is adopted to detect faults in turbofan engines with adaptive controllers. This design is a ToMFIR-redundancy-based robust fault detection. The ToMFIR is first introduced and existing results are also summarized. The Detailed design process of the ToMFIRs is presented and a turbofan engine model is simulated to verify the effectiveness of the proposed ToMFIR-based fault-detection strategy.