Novel symmetric bis-benzimidazoles: Synthesis, DNA/RNA binding and antitrypanosomal activity.

The novel benzimidazol-2-yl-fur-5-yl-(1,2,3)-triazolyl dimeric series with aliphatic and aromatic central linkers was successfully prepared with the aim of assessing binding affinity to DNA/RNA and antitrypanosomal activity. UV-Visible spectroscopy, thermal denaturation showed interaction of heterocyclic bis-amidines with ctDNA. Circular dichroism studies indicated uniform orientation of heterocyclic bis-amidines along the chiral double helix axis, revealing minor groove binding as the dominant binding mode. The amidino fragment and 1,4-bis(oxymethylene)phenyl spacer were the main determinants of activity against Trypanosoma brucei. The bis-benzimidazole imidazoline 15c, which had antitrypanosomal potency in the submicromolar range and DNA interacting properties, emerged as a candidate for further structural optimization to obtain more effective agents to combat trypanosome infections.

Mechanistic investigation of charge-remote and charge-driven fragmentation processes in 2,5-diphenyl-3,4-ethylenedioxythiophene diamidines.

RATIONALE: Diphenylfuran diamidines represent an important class of DNA minor groove binders of high therapeutic interest as antitumor and antibacterial agents. This study aimed to investigate fragmentation patterns in mass spectra of four diamidine derivatives with significant antitumor activity, in order to gain more insight into the structures and stability of their putative biological metabolites. METHODS: Compounds were investigated by electrospray ionization tandem mass spectrometry (ESI-MS/MS) using low-energy collision-induced dissociation (CID). Density functional theory calculations were performed to confirm the main fragmentation paths. RESULTS: The most abundant ion present in mass spectra is the doubly protonated molecule, whereas singly protonated molecules are present to a lesser extent. In the simplest compound, 2,5-bis(4-amidinophenyl)-3,4-ethylenedioxythiophene, the main fragmentation path was loss of ammonia, followed by loss of HCN where possible. The fragmentation of the N-alkyl derivatives (N-isopropyl-, N-isobutyl-, N-cyclopentyl-) includes competition between loss of alkene and the corresponding amine, followed by loss of another alkene and formation of fragment ions present in the pathway of the parent compound. CONCLUSIONS: The primary sites of fragmentations of investigated compounds are amidine groups, while breaking the core 3,4-ethylenedioxythiophene ring system does not take place. Fragmentation of the singly protonated molecule [M + H](+) occurs primarily on the charged side of the molecule, but a charge-remote process is energetically viable. The fragmentation mechanism of the alkyl derivatives revealed that singly and doubly protonated molecules cleave to the singly and doubly protonated molecules of the parent compound. Once formed, they are gradually transformed into nitrile. Copyright © 2016 John Wiley & Sons, Ltd.

An Aromatic Diamidine That Targets Kinetoplast DNA, Impairs the Cell Cycle in Trypanosoma cruzi, and Diminishes Trypomastigote Release from Infected Mammalian Host Cells

Trypanosoma cruzi is the etiological agent of Chagas disease, affecting approximately 10 million people in the Americas and with some 40 million people at risk. The objective of this study was to evaluate the anti-T. cruzi activity of three new diamidines that have a 3,4-ethylenedioxy extension of the thiophene core, designated MB17, MB19, and MB38. All three diamidines exhibited dose-dependent inhibition of epimastigote replication. The mechanisms of action of these diamidines were investigated. Unlike MB17 and MB19, MB38 exhibited a significant increase in the number of annexin-propidium iodide double-labeled cells compared to levels in control parasites. As MB17 had shown a lower 50% inhibitory concentration (IC50) against epimastigote growth, the mechanism of action of this drug was studied in more detail. MB17 triggered a decrease in the intracellular ATP levels. As a consequence, MB17 affected the genomic DNA and kinetoplast DNA (kDNA) and impaired the parasite cell cycle. Moreover, MB17 caused DNA fragmentation, with a more severe effect on kDNA than on nuclear DNA, resulting in dyskinetoplastic cells. MB17 was tested for toxicity and effectiveness for the treatment of infected CHO-K-1 cells, exhibiting a 50% cytotoxic concentration (CC50) of 13.47 +/- 0.37 mu Mand an IC50 of 0.14 +/- 0.12 mu Magainst trypomastigote release. MB17 also diminished the infection index by 60% at 0.5 mu M. In conclusion, despite belonging to the same family, these diamidines have different efficiencies. To summarize, MB17 was the most potent of these diamidines against epimastigotes, producing DNA damage preferentially in kDNA, impairing the parasite cell cycle, and decreasing the infection index and trypomastigote release from infected mammalian host cells, with a high selectivity index (SI) (< 90). These data suggest that MB17 could be an interesting lead compound against T. cruzi

[Significance of blood transfusion in the development of cytotoxic antibodies in patients on hemodialysis]. / Znacaj transfuzija krvi za razvoj citotoksicnih antitela kod bolesnika na hemodijalizi.

The aim was to evaluate the influence of red blood cells (RBC) transfusion on the development of cytotoxic antibodies (C-Ab) in patients subjected to hemodialyses (HD) and planned for the kidney transplantation. The group of 71 HD patients, of mean age 42 years (19-65), 48 males and 23 females, planned for the kidney information was examined. Out of 71 HD patients, only 42 (59.19%) HD patients (group I) received subcutaneously recombinant human erythropoietin--rhuEPO (Eprex--epoetin-alpha or Recormon SE--epoetin-beta in dosage of 4,000 IU during every HD; i.e. one to three times a week) and they were not treated by RBC transfusion. The other 29 (40.85%) HD patients (group II) received RBC transfusion: 18 (62.07%) HD patients received < 10 units 18 of RBC, 8 (27.59%) HD patients received 10-20 units of RBC; 3 (10.35%) HD patients received > 20 units of RBC. Testing of C-Ab was done in all patients every three months by standard lymphocytotoxicity test on the panel from 20 different lymphocyte donors with definite class I phenotype of antigen HLA. C-Ab was not found in HD patients who were not treated by RBC transfusion. Out of 18 HD patients who received < 10 units of RBC only 3 (16.67%) HD patients developed C-Ab; out of 8 HD patients who received 10-20 units of RBC, in 4 (50%) patients was proved C-Ab; and C-Ab was proved in all 3 HD patients who received > 20 units of RBC. RhuEPO administration is very important for the transfusiologic treatment of HD patients; especially those who are planned for the kidney transplantation. Development of C-Ab is in direct correlation with the number of transfunded units of RBC. HD patients who received 10 or more units of RBC were at great risk to develop C-Ab.

Histocompatibility antigens in patients with hepatocellular carcinoma.

The distribution of class I (A, B, C) and class II (DR antigens) histocompatibility antigens (HLA) was examined in 82 patients with hepatocellular carcinoma (HCC) and in 147 patients with chronic liver disease as controls. The diagnosis of HCC was confirmed by histological examination of liver tissue. HLA-B15 antigen was found more frequently in the subgroup of HCC patients who were positive for HBsAG (13/36, 36.1%) compared to the control group (8/147; 5.4%) [p < 0.001, Pc < 0.05, RR = 9.8] and a HBsAg positive control subgroup (1/25, 4%) [p < 0.001, Pc < 0.05, RR = 13.6]. No other statistically significant difference was found for any of the HLA antigens examined either in HCC patients as a whole group or in the subgroups according to sex, course of illness, AFP status, alcohol consumption, liver cirrhosis or blood groups. These data are further evidence that there may be a link between hepatitis B viral (HBV) infection and HLA antigens. The association of HLA-B15 antigen and HbsAg supports the idea of some genetic control of HBV infection in the patients with HCC.

[Kidney transplantation in high-risk patients]. / Transplantacija bubrega u bolesnika sa visokim rizikom.

Immunosuppression with Cyclosporine A in kidney transplantation, triple therapy (CyA + Imuran + corticosteroids) and plasmapheresis before and after kidney transplantation in high risk recipients (positive cytotoxic antibody, MLC at the level of non related persons), also in high risk patients (juvenile diabetes, patients over 50 years old). In 1988 we had done in our Centre, kidney transplantation in 52.8% (28: 53) in high and increased risk patients. Triple therapy with plasmapheresis before and after kidney transplantation (if the level of cytotoxic antibodies is over 15%) allows successful kidney transplantation in high risk kidney recipients. Patients with juvenile diabetes are also available kidney recipients with therapy and permanent regulation of blood sugar. The patients of the age group between 50-60 years should be considered as suitable for kidney transplantation.

[Correlation between the frequency of HL-A antigens and antigens of the P system in the population of Serbia]. / Korelacija izmedu zastupljenosti HLA antigena i antigena p sistema eritrocitnih krvnih grupa u populaciji SR Srbije.

We studied the correlation between the antigens of the P system and the antigens and haplotypes of the HLA system in 334 nonrelated persons and 46 chosen families consisting of 166 members. On our material we found a distinct linear correlation between the antigens of the segregant serie A and especially of the segregant serie B and the P antigen of the blood groups. The correlation between the association of the antigens of series A and B and the antigens of the P system same as between the most frequent HLA haplotypes and the antigens of the P system, is also distinct. We described two cases of crossing over between the P and HLA system; the frequency of crossing over of 0,0435. We have confirmed the linkage between HLA and P and the likelihood that the genes are located on the same chromosome.

[Occurrence of cytotoxic anti-HL-A antibodies in female patients sensitized to Rh antigens]. / Pojava citotoksicnih anti-HL-A antitela u pacijentkinja senzibilisanih na antigene Rhesus-sistema

The frequency of sentization to HL-A system antigens in the categories of Rh-sentized patients was studied, considering the observation that in this category the precentage of senzitization to the system of transplantation antigens is much higher than in other groups. A total of 211 samples of sera of Rh-sensitized patients were examined of which 127 were sensitized to the HL-A system at the same time, while the remaining 74 showed absence of HL-A antibodies. For work the technique of Indirect antihuman globulin test was used (for Rh-sentization) while for HL-A system sensitization examination, the Two-stage microlymphocytotoxicyty method of Terasaki, modified by Dausset, was used. In this study the possibility of the favourable influence of sensitization to the erythrocyte antigen Rh-system and sensitization to transplantation antigens during pregnancy are discussed.

[Correlation between the presence of cytotoxic anti-HL-A antibodies in the clinical course of hemolytic disease of the newborn infant due to anti-Rh (D) or immune anti-A/B erythrocyte antibodies]. / Korelacija izmedu prisustva citotoksicnih anti-HL-A antitela i klinickog toka hemoliticke bolesti novorodencadi uzrokovane anti-Rh-(D) ili imunim anti-A/Beritrocitnim antitelima

In a retrospective study the simultaneous influence of catotoxic HL-A antibodies on the clinical course of 60 cases of infants affected by Haemolytic disease on the Newborn due to anti Rh (D) or immune anti A/B antibodies, is shown. In all cases the treatment was by exange transfusion. In the group of infants in whose cord blood anti HL-A antibodies were found Exange transfusion had a weak efect so that it had to be repeated in 96 per cent of cases. In the group of infants in whose cord blood anti HL-S antibodies were not found, Exange transfusion was repeated only in one case, that is 2,7 per cent. In a group of Rh isommunised mothers whose children were affected by Haemolytic disease of the Newborn, antt HL-A antibodies were found in 61,7 per cent while 80,5 per cnet of the antibodies passed through the placente. In a group of ABO isoimmunised mothers cytotoxic HL-A antibodies were found in 42,3 per cent, while 45,4 per cent passed through the placente. A significant difference in the number of leucocytes, limphocytes, platelets, term of birth, level of bilirubin, amount of haemoglobin and Apgar Score was not found between the group of newborn who in their cord blood had, besides the already present isoimmunhaemagllutinines, cytotoxic HL-A antibodies and the group of infants with no cytotoxic anti HL-A antibodies, present. Cytotoxic HL-A antibodies in a way, react with the "unmasked" erythrocyte membrane, increasing haemolysis, so that the therapeutic effect of Exange transfusion was discriminated.