Incorporation of lipolysis in monolayer permeability studies of lipid-based oral drug delivery systems.

Lipid-based drug delivery systems, a well-tolerated class of formulations, have been evaluated extensively to enhance the bioavailability of poorly soluble drugs. However, it has been difficult to predict the in vivo performance of lipid dosage forms based on conventional in vitro techniques such as cell monolayer permeability studies because of the complexity of the gastrointestinal processing of lipid formulations. In the current study, we explored the feasibility of coupling Caco-2 and Madin-Darby canine kidney monolayer permeability studies with lipolysis, a promising in vitro technique to evaluate lipid systems. A self-emulsifying lipid delivery system was formulated using a blend of oil (castor oil), surfactant (Labrasol® or PL497), and co-surfactant (lecithin). Formulations demonstrating high drug solubility and rapid self-emulsification were selected to study the effect of lipolysis on in vitro cell permeability. Lipolysis of the formulations was carried out using pancreatin as the digestive enzyme. All the digested formulations compromised monolayer integrity as indicated by lowered trans-epithelial electrical resistance (TEER) and enhanced Lucifer yellow (LY) permeability. Further, the changes in TEER value and LY permeability were attributable to the digestion products of the formulation rather than the individual lipid excipients, drug, digestion enzyme, or the digestion buffer. The digested formulations were fractionated into pellet, oily phase, and aqueous phase, and the effect of each of these on cell viability was examined. Interestingly, the aqueous phase, which is considered important for in vivo drug absorption, was responsible for cytotoxicity. Because lipid digestion products lead to disruption of cell monolayer, it may not be appropriate to combine lipolysis with cell monolayer permeability studies. Additional in vivo studies are needed to determine any potential side effects of the lipolysis products on the intestinal permeability barrier, which could determine the suitability of lipid-based systems for oral drug delivery.

Highly Eribulin-resistant KBV20C Oral Cancer Cells Can Be Sensitized by Co-treatment with the Third-generation P-Glycoprotein Inhibitor, Elacridar, at a Low Dose.

BACKGROUND/AIM: Eribulin mesylate, also called Halaven® (HAL), was recently developed as a microtubule-targeting drug and is used in the clinic for resistant or metastatic cancer. Previously, we showed that P-glycoprotein (P-gp)-overexpressing KBV20C oral cancer cells are highly resistant to HAL compared to sensitive KB cells. This qualitative study was designed to identify specific P-gp inhibitors that increase the sensitivity of highly resistant cancer cells to HAL. MATERIALS AND METHODS: In order to identify functional P-gp inhibitors, HAL-treated KBV20C cells were co-treated with P-gp inhibitors, verapamil, elacridar, cyclosporine A, mitotane, piperine, fumagillin, curcumin, indomethacin, probenecid, sulindac, tesmilifene, and C-4. We then evaluated which P-gp inhibitors required a low dose to sensitize KBV20C cells to HAL. We also determined whether a low dose of a P-gp inhibitor could inhibit P-gp efflux pumping. RESULTS: We found that cyclosporine A sensitized HAL-treated KBV20C cells at a low dose, whereas verapamil, another first-generation P-gp inhibitor, required a dose that was nearly 10-fold higher. We also found that the natural products, piperine and mitotane, sensitized KBV20C cells to HAL co-treatment. Interestingly, we found that elacridar, a third-generation P-gp inhibitor, sensitized HAL-treated cells at a low dose. Elacridar required approximately a 500-fold lower dose than that of verapamil to exert a similar effect. All inhibitors showed P-gp inhibitory activity that correlated with sensitivity to HAL. CONCLUSION: These results suggest that highly HAL-resistant cancer cells can be sensitized with cyclosporine A or elacridar, specific P-gp inhibitors that exert their effects at a low dose. These findings provide important information regarding the sensitization of highly HAL-resistant cells with selective P-gp inhibitors and indicate that elacridar may be used to treat such highly HAL-resistant cancer cells.

[Therapy and Rehabilitation of Patients with Pulmonary Tuberculosis and Different Treatment Adherence].

The pulmonary tuberculosis process as dependent on the disease form and the therapy efficacy with the use of Cycloferon in the treatment scheme were investigated. The study had two stages. At the first stage the data concerning 358 patients with primary pulmonary tuberculosis and infiltration (93 patients) or degradation (89 patients) and 176 patients with pulmonary fibrocavernous tuberculosis were analysed. At the second stage the efficacy of the treatment schemes applied to the patients with pulmonary fibrocavernous tuberculosis was compared. The etiotropic therapy intensive phase was applied to all the patients. Moreover, 56 patients (group 1) under the therapy and rehabilitatinon were treated with Cycloferon in a dose of 0.25 administered intramuscularly twice a week (not less than 16 injections for the course), 60 patients (group 2) were treated with Omega 3, 30 patients (group 3) were given the standard complex (vitamins and tonics), 30 patients (group 4) were under the etiotropic therapy alone. The following additional factors promoting progression and aggravation of the tuberculosis process were confirmed: degradation at the time of the disease diagnosis, high resistance of the pathogen to antituberculosis drugs, low adherence to the treatment, social desadaptation and especially psychofunctional state of the patients. The use of Cycloferon in the schemes of the intensive phase treatment of the primary fibrocavernous tuberculosis resulted in reduction of the intoxication signs, bacteria isolation, positive dynamics of the cavity healing, lower lung infiltration and consequently high frequency of the treatment positive outcomes (94.1 ± 3.33%).

[Syndrome "sickly child"].

Scientific review is devoted to the urgent problem of child health care--Syndrome "sickly child" described in detail the category of "frequently ill children" repeated respiratory diseases, the pathogens that cause diseases of the respiratory tract. Paying attention to factors contributing to re-respiratory morbidity, including genetically determined causes (disturbances in the state of health of the mother), is represented by the genetic determinism of repeated and recurrent diseases related to blood groups. The development of an immune imbalance is manifested by changes in the cellular, humoral immune response and nonspecific resistance factors characterizing the changes of local immunity in this category of children that shape the development of chronic disease. We describe the effect of an allergic component to the severity of respiratory disease and the relationship with the mechanisms of neuroendocrine and immune systems. Correction of the immune resistance of sickly children was conducted cycloferon contributing to reduce the incidence and duration of repeated episodes of acute respiratory infections per year.

[Influence of immunotropic preparation cycloferon and phytopreparations of Cynara scolimus L. on blood cytokines profile of the patients with chronic viral hepatitis C in medical rehabilitation period].

The effect of the immunotropic drug cycloferon and herbal medicine resources on the basis of Cynara scolimus L. on the blood cytokine profile in the patients with chronic viral hepatitis C (CVHC) in medical rehabilitation (MR) period. Established that prior to MR period in the patients with CVHC was noted significantly increased levels of proinflammatory cytokines (CK) at the blood serum, and the level of antiinflammatory CK changed significantly. The use of cycloferon and herbal medicine resources on the basis of Cynara scolimus L. in the MR complex provided to normalize the studied CK concentration in the serum of the patients with CVHC.

[Apoptosis of the hypothalamus neurosecretory cells in stress and ageing: the role of immune modulators].

There is assumption about active role of immune modulators in cell death process. The involvement of interferon-alpha and cycloferon in apoptosis regulation of hypothalamic neurons of mice during stress and aging was studied. We determined the expression of apoptosis markers (Bcl-2, Mcl-1, Bax) in comparison with apoptosis level. We have found that immune modulators suppress activity of nonapeptidergic neurons. Thus, interferon-alpha treatment reduces synthesis of Bcl-2; cycloferon treatment inhibits expression of Bax and Bcl-2. So the role of immune modulators in neuron apoptosis depends on the stage of ontogenesis and type of immune modulator. Cycloferon is able to reduce the level of age-dependent apoptosis of neurons in aging, but under stress condition both interferon-alpha and cycloferon act as protectors of cell death.

[Monitoring of side effects and estimation of cycloferon efficacy in treatment of children with frequent and prolonged diseases].

Two hundred fifty patients, including 100 children with frequent and prolonged diseases at the age of 4 to 7 years, 76 children at the age of 7 to 18 years and 74 subjects at the age of 22 to 57 years were observed. The patients were treated with cycloferon in two courses with a 2-week interval according to the standard scheme. The tonsil surface microflora and its susceptibility to antibiotics were determined. Cycloferon lowered the Staphylococcus aureus titre and increased the culture susceptibility to benzylpenicillin, oxacillin, rifampicin, and erythromycin, reducing the variety of the fauces nonpathogenic microflora. The use of cycloferon induced no adverse (pathologic) reactions in 94.8% of the cases. In 4.4% of the children under school age the adverse reactions were transitory and did not require discontinuation of the drug use. Unforeseen reactions were recorded in 0.8% of the children and the use of the drug in them was discontinued. The use of cycloferon in two courses with a 2-week interval according to the standard scheme is recommended for prophylaxis of acute respiratory diseases in the group of children with frequent and prolonged diseases during epidemiologically unfavourable periods and for complex therapy of rhinopharinx infections as an agent increasing efficacy of other antibacterials.

The effects of a telomere destabilizing agent on cancer cell-cycle dynamics--integrated modelling and experiments.

The pentacyclic acridinium salt RHPS4 displays anti-tumour properties in vitro as well as in vivo and is potentially cell-cycle specific. We have collected experimental data and formulated a compartmental model using ordinary differential equations to investigate how the compound affects cells in each stage of the cell cycle. In addition to a control case in which no drug was used, we treated colorectal cancer cells with three different concentrations of the drug and fitted simulations from our models to experimental observations. We found that RHPS4 caused a concentration-dependent, marked cell death in treated cells, which is best modelled by allowing the rate parameters corresponding to cell death to be sigmoidal functions of time. We have shown that the model is "identifiable", meaning that, at least in principle, the parameter values can be determined from observable quantities. We find that at low concentrations RHPS4 primarily affects the cells in the G(2)/M phase, and that the drug has a delayed effect with the delay decreasing at larger doses. Since the drug diffuses into the nucleus, the observed delayed effect of the compound is unexpected and is a novel finding of our research into this compound.

[Characteristics of immune response to etiotropic and pathogenetic therapy in children with chronic hepatitis C].

Features of the immune response of children with chronic hepatitis C to the antiviral and pathogenetic therapy have been studied. It is shown that the antiviral therapy is accompanied by stimulation of the immune response as manifested by the synthesis of cytokines (IL-4, IFN-alpha, IFN-gamma) with retention of increased production of IFN-a for more than two years after the end of the course of treatment. In children that previously received interferon inductor (cycloferon) for 12 months, high level of IFN-a production is retained, which ensures antiviral protection. Phytotherapy did not influence the production of cytokines.

Investigation of cytotoxic activity on human cancer cell lines of arborinine and furanoacridones isolated from Ruta graveolens.

The cytotoxic effects of a series of furanoacridones isolated from Ruta graveolens L. (Rutaceae) and of two further acridone alkaloids (arborinine and evoxanthine) were investigated by means of the MTT assay, using the human cell lines HeLa, MCF7 and A431. Arborinine proved best in inhibiting the proliferation of all three cell lines. The cytotoxic potency of the furacridone alkaloids was a function of their lipid solubility, which was determined by means of PAMPA. The capacity of the most effective furanoacridones to induce apoptosis was demonstrated by flow cytometric cell cycle analysis and by staining with ethidium bromide and acridine orange. This finding was reinforced by determining the apoptosis-regulating factors Bcl-2 and Bax, which were revealed by means of RT-PCR to change dose-dependently. The data presented here indicate that naturally occurring furanoacridones can be regarded as excellent starting structures for the potential development of new anticancer agents.

Cytochrome C release induces apoptosis of nasopharyngeal carcinoma (NPC) by antitumor glyfoline.

The mechanism of action, leakage of cytochrome c from mitochondria into cytosol, for the antineoplastic compound glyfoline was examined. Additionally, our current studies revealed that glyfoline induced apoptotic changes and arrested cell cycle procession at the G2/M phase in nasopharyngeal carcinoma (NPC). A reverse transcriptase polymerase chain reaction (RT-PCR) showed no specific changes of apoptosis-related gene expression (i. e., bax, ICE-alpha,beta, bcl-2, and c-myc). However, no similar changes were detected in fibroblasts and peripheral lymphocytes after glyfoline treatment suggesting that glyfoline has a higher affinity for tumor cells than for normal cells.

[Use of cycloferon in a combined treatment of chlamydial conjunctivitis]. / Primenenie preparata tsikloferona pri kompleksnom lechenii khlamidiinogo kon"iunktivita.

A high therapeutic efficiency of cycloferon, an interferon inductor, was demonstrated when it is combined with antibiotics in the treatment of chlamidial conjuctivitis. The use of the mentioned drug for acute chlamidial conjuctivitis prevents relapses.

[A comparative evaluation of treatment methods for a genital papillomavirus infection in women with different viral genotypes]. / Sravnitel'naia otsenka nekotorykh sposobov lecheniia genital'noi papillomavirusnoi infektsii u zhenshchin s razlichnymi genotipami virusa.

Such genotypes of human papilloma virus (HPV) as "malignant" (180), "benign" (49) and indeterminate (127) were identified in 356 females by PCR and cytological procedures. The following therapeutic techniques were compared: cryodestruction of the uterine cervix (60), antiviral medication (AVM) (19), combination of AVM and cryodestruction (224), and combination of AVM and laser destruction (54). In the "malignant" HPV group, AVM in conjunction with cryo- or laser destruction was significantly more effective than cryodestruction or AVM alone. Both cryodestruction alone and in combination with AVM were more effective in treating "benign" genotypes. Repeat courses showed the same tendency.

Parasitological evaluation of curative and subcurative doses of 9-acridanone-hydrazone drugs against Schistosoma mansoni in baboons, and observations on changes in serum levels of anti-egg antibodies detected by ELISA.

Derivatives in the class of 9-acridanone-hydrazones were found to be highly active against Schistosoma mansoni in baboons. Single doses of 25 mg/kg were fully effective. Data are presented showing changes detected by ELISA in antibody levels against schistosome eggs which correlated positively with the effect of chemotherapy. This approach may help to evaluate the effects of treatment of human schistosomiasis where the detection of low egg counts is problematic.

Anti-neoplastic activity of C205: a new topical anti-cancer formulation.

C205, a topical ointment currently in clinical trial, was tested by a method developed to evaluate the topical activity of new drugs. Subcutaneous B-16 melanoma was used as the primary tumor model. Tumor size of treated animals as percentage of tumor size of control animals (T/C ratio) was found to be 14, with 50% of the animals tested free of viable and palpable tumor at 60 days post treatment. The anti-neoplastic activity of C205 was superior to 5-fluorouracil using the same method of evaluation. Only minimal loss of animal body weight occurred after C205. C205 also showed anti-tumor activity against subcutaneously implanted Lewis lung carcinoma.

[Chemotherapeutic activity of benzylpenicillin in combination with a perhydroacridine derivative studied in experimental staphylococcal infection]. / Izuchenie khimioterapevticheskoi aktivnosti benzilpenitsillina v sochetanii s proizvodnym pergidroakridina pri éksperimental'noi stafilokokkovoi infektsii.

Chemotherapeutic activity of various benzylpenicillin combinations with 10-nitro-so-trans-anti-cys-perhydroacridine (MT-2) was studied on 2 experimental staphylococcal infections caused by pathogenic penicillin resistant strains of staphylococci. It was found that the combined use of the antibiotic and MT-2 in doses of 25-100 and 12.5 mg/kg respectively administered subcutaneously increased the therapeutic effect of benzylpenicillin as compared to the antibiotic used alone. The oral use of MT-2 in a dose of 400 mg/kg also increased the therapeutic action of benzylpenicillin used in a dose of 100 mg/kg subcutaneously simultaneously with MT-2. A favourable effect of ointments containing MT-2 in a concentration of 10-20 per cent and benzylpenicillin in a dose of 50 000 or 100 000 units per 1 g of the ointment was noted in treatment of mice with localized staphylococcal infection.

Studies on vomitory effect of some acridine compounds.

Vomitory effect of 16 chosen acridine derivatives was observed on pigeons. The expected effect of the preparations as well as their interaction with, causing vomiting lobeline were considered. Two groups were distinguished: 1) preparations which alone caused vomitory reaction (C-283, C-410, C-541, C-609, C-684, C-702, C-829, C-835, C-846, C-1006 and C-1020). 2) Preparations which caused this effect while combined with a subthreshold dose of lobeline (C-429, C-516, C-666, C-857 and C-1005). Dependence of the effects on chemical structure and pharmacological activity of the compounds is discussed.

Antimalarial activity of Floxacrine (HOE 991) I. Studies on blood schizontocidal action of Floxacrine against Plasmodium berghei, P. vinckei and P. cynomolgi.

Floxacrine (HOE 991), 7-chloro-10-hydroxy-3-(4-trifluoromethylphenyl)3,4-dihydroacridine-1,9-(2H, 10H) dion, shows a high level of antimalarial action against blood-induced infection of drug-sensitive and drug-resistant lines of Plasmodium berghei in mice, rats and Syrian hamsters. The drug is also a potent blood schizontocide against drug-sensitive P. vinckei strains in rodents and P. cynomolgi in rhesus monkeys. The CD50/CD90 values against the drug-sensitive P. berghei strain ascertained in the '28-day test' in mice were 4.3/6.7 mg/kg after the oral route and 1.7/3.6 mg/kg after the subcutaneous (sc) route. In the 'two- and four-day test' the ED50 against sensitive P. vinckei was 0.7 mg/kg in both mice and rats. A moderate prophylactic effect could be demonstrated after the sc route probably due to a 'depot effect' of the water-insoluble active principle. Floxacrine was also highly active against P. berghei-lines which were resistant to chloroquine, mepacrine, dihydrofolate reductase inhibitors, sulfadoxine and dapsone. Resistance to HOE 991 could be developed in P. berghei and P. cynomolgi when the compound was used alone and administered repeatedly in subcurative doses. The antimalarial activity of the compound was not influenced by p-aminobenzoic acid or folic acid supplements in diets. Structural changes induced by floxacrine on pigment cytoplasm and nucleus in erythrocytic stages of P. berghei differed in some aspects from those of mepacrine and chloroquine. It is therefore assumed that the mode of action of floxacrine differs from that of the known antimalarial drugs. The general tolerance of the compound in rodents and rhesus monkeys is good and there is a wide range between the effective and maximum tolerated doses. Floxacrine was also effective at 100 ppm against pathogen Eimeria species in chickens, at 1000 mg/kg orally against Fasciola hepatica in rats and at 300-800 mg/kg orally against Heterakis spumosa in rats.

Oral absorption and selective tissue localization of 4'-(9-acridinylamino)-methanesulfon-m-anisidide.

The disposition of 4'-(9-acridinylamino)-methanesulfon-m-anisidide (AMSA), a new antitumor agent presently undergoing clinical evaluation, was studied in mice and rats following oral administration and compared to that observed following intravenous administration. The metabolic fate of AMSA was the same with either intravenous or oral administration; however, the tissue distribution of AMSA differed significantly between the two routes of administration. Following absorption from the GI tract, AMSA was rapidly cleared from plasma by the liver and excreted in the bile as metabolites. Concentrations of AMSA in the liver were relatively high after oral administration and were sufficient to exert a cytotoxic effect on L1210 cells implanted at the site. The results indicate the use of AMSA orally to attain selective localization in the liver with decreased systemic exposure, which may prove useful against tumor metastases to the liver or primary hepatocellular carcinoma.