N-Trifluoromethyl Amines and Azoles: An Underexplored Functional Group in the Medicinal Chemist's Toolbox.

Introducing trifluoromethyl groups is a common strategy to improve the properties of biologically active compounds. However, N-trifluoromethyl moieties on amines and azoles are very rarely used. To evaluate their suitability in drug design, we synthesized a series of N-trifluoromethyl amines and azoles, determined their stability in aqueous media, and investigated their properties. We show that N-trifluoromethyl amines are prone to hydrolysis, whereas N-trifluoromethyl azoles have excellent aqueous stability. Compared to their N-methyl analogues, N-trifluoromethyl azoles have a higher lipophilicity and can show increased metabolic stability and Caco-2 permeability. Furthermore, N-trifluoromethyl azoles can serve as bioisosteres of N-iso-propyl and N-tert-butyl azoles. Consequently, we suggest that N-trifluoromethyl azoles are valuable substructures to be considered in medicinal chemistry.

The impact of azole antifungal drugs on imatinib metabolism in human liver microsomes.

1. Imatinib is widely used for the treatment of hematologic malignancies. It is common that imatinib is clinically co-prescribed with azole antifungal agents since these patients are more prone to invasive antifungal infection. The present study was to investigate the effects of azole antifungal drugs, including ketoconazole, fluconazole, voriconazole, itraconazole and posaconazole on imatinib metabolism. 2. The main metabolites, 1-OH midazolam and N-desmethyl imatinib, were determined in the absence and in the presence of various levels of ketoconazole, fluconazole, voriconazole, itraconazole and posaconazole. The relevant assay was also performed to screen mechanism-based inhibitors (MBI). 3. The inhibition ability of 1-OH midazolam formation from midazolam based on IC50 values was ketoconazole (0.09 µM)>itraconazole (0.31 µM)> posaconazole (0.68 µM)>voriconazole (2.10 µM) > fluconazole (8.90 µM). Similarly, the rank order of inhibitory effects on formation of N-desmethyl imatinib from imatinib was ketoconazole (4.58 µM)>itraconazole (17.45 µM)> posaconazole (31.02 µM)> voriconazole (367.9 µM) >fluconazole (1.11 mM). Posaconazole and itraconazole displayed evidence of MBI. Additionally, imatinib was also shown as a MBI of CYP3A with IC50 value of 5.40 µM against the midazolam. 4. The significant difference in IC50 values of midazolam and imatinib inhibited by azole antifungal agents was observed. The role of CYP2C8 in imatinib metabolism and imatinib autoinhibits CYP3A activity may explain this difference. Our findings suggest that the azole antifungal agents might have limited impacts on imatinib exposure by CYP3A activity.

18F-Flortanidazole Hypoxia PET Holds Promise as a Prognostic and Predictive Imaging Biomarker in a Lung Cancer Xenograft Model Treated with Metformin and Radiotherapy.

Metformin may improve tumor oxygenation and thus radiotherapy response, but imaging biomarkers for selection of suitable patients are still under investigation. First, we assessed the effect of acute metformin administration on non-small cell lung cancer xenograft tumor hypoxia using PET imaging with the hypoxia tracer 18F-flortanidazole. Second, we verified the effect of a single dose of metformin before radiotherapy on long-term treatment outcome. Third, we examined the potential of baseline 18F-flortanidazole as a prognostic or predictive biomarker for treatment response. Methods: A549 tumor-bearing mice underwent a 18F-flortanidazole PET/CT scan to determine baseline tumor hypoxia. The next day, mice received a 100 mg/kg intravenous injection of metformin. 18F-flortanidazole was administered intravenously 30 min later, and a second PET/CT scan was performed to assess changes in tumor hypoxia. Two days later, the mice were divided into 3 therapy groups: controls (group 1), radiotherapy (group 2), and metformin + radiotherapy (group 3). Animals received saline (groups 1-2) or metformin (100 mg/kg; group 3) intravenously, followed by a single radiotherapy dose of 10 Gy (groups 2-3) or sham irradiation (group 1) 30 min later. Tumor growth was monitored triweekly by caliper measurement, and tumor volume relative to baseline was calculated. The tumor doubling time (TDT), that is, the time to reach twice the preirradiation tumor volume, was defined as the endpoint. Results: Thirty minutes after metformin treatment, 18F-flortanidazole demonstrated a significant change in tumor hypoxia, with a mean intratumoral reduction in 18F-flortanidazole tumor-to-background ratio (TBR) from 3.21 ± 0.13 to 2.87 ± 0.13 (P = 0.0001). Overall, relative tumor volume over time differed across treatment groups (P < 0.0001). Similarly, the median TDT was 19, 34, and 52 d in controls, the radiotherapy group, and the metformin + radiotherapy group, respectively (log-rank P < 0.0001). Both baseline 18F-flortanidazole TBR (hazard ratio, 2.0; P = 0.0004) and change from baseline TBR (hazard ratio, 0.39; P = 0.04) were prognostic biomarkers for TDT irrespective of treatment, and baseline TBR predicted metformin-specific treatment effects that were dependent on baseline tumor hypoxia. Conclusion: Using 18F-flortanidazole PET imaging in a non-small cell lung cancer xenograft model, we showed that metformin may act as a radiosensitizer by increasing tumor oxygenation and that baseline 18F-flortanidazole shows promise as an imaging biomarker.

Isavuconazole: A new broad-spectrum azole. Part 2: pharmacokinetics and clinical activity.

Isavuconazole, the active moiety of its prodrug isavuconazonium, is a new extended-spectrum triazole whose activity against yeasts, molds, including Aspergillus and mucorales, and dimorphic fungi has been shown in vitro and in preclinical models. The most relevant pharmacokinetics features are water-solubility of the prodrug, rapid cleavage of the prodrug into active moiety and cleavage product by plasmatic esterases, high oral bioavailability of isavuconazole with an extensive penetration into most tissues and a good safety profile even in case of renal impairment. The results of two main clinical studies have led to an approval by FDA and EMA in the treatment of invasive aspergillosis and invasive mucormycosis. Isavuconazole is non-inferior to voriconazole in terms of response and survival in invasive aspergillosis and has shown improved safety and tolerability. Importantly, less hepatobiliary, skin and eye disorders have been reported in isavuconazole-treated patients. Isavuconazole has therefore been granted a grade A-I recommendation by the European Conference on Infections in Leukemia (ECIL) for the treatment of invasive aspergillosis. Efficacy has also been demonstrated in mucormycosis in an open-label study. Survival was similar to the survival of matched patients from the international Fungiscope registry and treated with an amphotericin B formulation. Isavuconazole failed to show non-inferiority to caspofungin in a large double-blind candidemia trial. The aim of this review is to give the reader an overview of the data available so far to support inclusion of isavuconazole in the anti-mold therapeutic arsenal.

Pharmacokinetics of Azole Antifungals in Cystic Fibrosis.

Defects in mucociliary clearance predispose cystic fibrosis (CF) patients to airway colonization and infection by various fungi, especially Aspergillus fumigatus. Although the clinical significance of airway fungal colonization is not clear, several studies have suggested its association with worsening lung function and increased risk of CF exacerbations. Antifungal triazole agents have been used in CF patients with airway fungal colonization or infections with varying results. Limited pharmacokinetic studies to date have demonstrated high inter-subject variability of triazole levels among CF patients. This review discusses the basic principles of pharmacokinetics, the pharmacokinetic changes associated with CF and the effect of CF on the pharmacokinetic principles of azole antifungals. The inconsistent azole serum levels in CF patients may be associated with sub-therapeutic (thus risk of therapeutic failure and/or emergence of azole-resistant fungi) or supratherapeutic exposures (thus potential risk of azole toxicity), suggesting that therapeutic dose monitoring is necessary in CF patients.

Antifungal pharmacodynamics: Latin America's perspective

Abstract The current increment of invasive fungal infections and the availability of new broad-spectrum antifungal agents has increased the use of these agents by non-expert practitioners, without an impact on mortality. To improve efficacy while minimizing prescription errors and to reduce the high monetary cost to the health systems, the principles of pharmacokinetics (PK) and pharmacodynamics (PD) are necessary. A systematic review of the PD of antifungals agents was performed aiming at the practicing physician without expertise in this field. The initial section of this review focuses on the general concepts of antimicrobial PD. In vitro studies, fungal susceptibility and antifungal serum concentrations are related with different doses and dosing schedules, determining the PD indices and the magnitude required to obtain a specific outcome. Herein the PD of the most used antifungal drug classes in Latin America (polyenes, azoles, and echinocandins) is discussed.

Azole drug import into the pathogenic fungus Aspergillus fumigatus.

The fungal pathogen Aspergillus fumigatus causes serious illness and often death when it invades tissues, especially in immunocompromised individuals. The azole class of drugs is the most commonly prescribed treatment for many fungal infections and acts on the ergosterol biosynthesis pathway. One common mechanism of acquired azole drug resistance in fungi is the prevention of drug accumulation to toxic levels in the cell. While drug efflux is a well-known resistance strategy, reduced azole import would be another strategy to maintain low intracellular azole levels. Recently, azole uptake in Candida albicans and other yeasts was analyzed using [(3)H]fluconazole. Defective drug import was suggested to be a potential mechanism of drug resistance in several pathogenic fungi, including Cryptococcus neoformans, Candida krusei, and Saccharomyces cerevisiae. We have adapted and developed an assay to measure azole accumulation in A. fumigatus using radioactively labeled azole drugs, based on previous work done with C. albicans. We used this assay to study the differences in azole uptake in A. fumigatus isolates under a variety of drug treatment conditions, with different morphologies and with a select mutant strain with deficiencies in the sterol uptake and biosynthesis pathway. We conclude that azole drugs are specifically selected and imported into the fungal cell by a pH- and ATP-independent facilitated diffusion mechanism, not by passive diffusion. This method of drug transport is likely to be conserved across most fungal species.

Design of a drug-drug interaction study of vincristine with azole antifungals in pediatric cancer patients using clinical trial simulation.

BACKGROUND: The aim of the current work was to perform a clinical trial simulation (CTS) analysis to optimize a drug-drug interaction (DDI) study of vincristine in children who also received azole antifungals, taking into account challenges of conducting clinical trials in this population, and, to provide a motivating example of the application of CTS in the design of pediatric oncology clinical trials. PROCEDURE: A pharmacokinetic (PK) model for vincristine in children was used to simulate concentration-time profiles. A continuous model for body surface area versus age was defined based on pediatric growth curves. Informative sampling time windows were derived using D-optimal design. The CTS framework was used to different magnitudes of clearance inhibition (10%, 25%, or 40%), sample size (30-500), the impact of missing samples or sampling occasions, and the age distribution, on the power to detect a significant inhibition effect, and in addition, the relative estimation error (REE) of the interaction effect. RESULTS: A minimum group specific sample size of 38 patients with a total sample size of 150 patients was required to detect a clearance inhibition effect of 40% with 80% power, while in the case of a lower effect of clearance inhibition, a substantially larger sample size was required. However, for the majority of re-estimated drug effects, the inhibition effect could be estimated precisely (REE < 25%) in even smaller sample sizes and with lower effect sizes. CONCLUSION: This work demonstrated the utility of CTS for the evaluation of PK clinical trial designs in the pediatric oncology population.

PXR antagonists and implication in drug metabolism.

Adopted orphan nuclear receptor (NR), pregnane X receptor (PXR), plays a central role in the regulation of xeno- and endobiotic metabolism. Since the discovery of the functional role of PXR in 1998, there is evolving evidence for the role of PXR agonists in abrogating metabolic pathophysiology (e.g., cholestasis, hypercholesterolemia, and inflammation). However, more recently, it is clear that PXR is also an important mediator of adverse xeno- (e.g., enhances acetaminophen toxicity) and endobiotic (e.g., hepatic steatosis) metabolic phenotypes. Moreover, in cancer therapeutics, PXR activation can induce drug resistance, and there is growing evidence for tissue-specific enhancement of the malignant phenotype. Thus, in these instances, there may be a role for PXR antagonists. However, as opposed to the discovery efforts for PXR agonists, there are only a few antagonists described. The mode of action of these antagonists (e.g., sulforaphane) remains less clear. Our laboratory efforts have focused on this question. Since the original discovery of azoles analogs as PXR antagonists, we have preliminarily defined an important PXR antagonist pharmacophore and developed less-toxic PXR antagonists. In this review, we describe our published and unpublished findings on recent structure-function studies involving the azole chemical scaffold. Further work in the future is needed to fully define potent, more-selective PXR antagonists that may be useful in clinical application.

Novel 1,3,4-heterodiazole analogues: synthesis and in-vitro antitumor activity.

The synthesis of some new heterodiazole and their annulated imidazo[2,1-b]1,3,4-oxa/thiadiazolone 6a-d, 7a-d; 1,3,4-oxa or thiadiazole[3,2-a]pyrimidine diamine 8a-d and 1,3,4-oxa or thiadiazole-3-piperidino-1-propamide 11a,b derivatives have been described. The obtained compounds were evaluated for their in-vitro antitumor activity. A single dose (10 µM) of the test compounds were used in the full National Cancer Institute (NCI) 60 cell lines panel assay. Compounds 6c and 6d displayed appreciable anticancer activity against leukemia, non-small cell lung, CNS and showed moderate activity against colon, melanoma, and breast cancer cells lines. Compound 6c possessed remarkable broad-spectrum antitumor activity which almost 4 fold more active than the known drug 5-FU with GI(50), TGI, and LC(50) values of 6.0, 17.4, and 55.1 µM, respectively.

Use of external metabolizing systems when testing for endocrine disruption in the T-screen assay.

Although, it is well-established that information on the metabolism of a substance is important in the evaluation of its toxic potential, there is limited experience with incorporating metabolic aspects into in vitro tests for endocrine disrupters. The aim of the current study was a) to study different in vitro systems for biotransformation of ten known endocrine disrupting chemicals (EDs): five azole fungicides, three parabens and 2 phthalates, b) to determine possible changes in the ability of the EDs to bind and activate the thyroid receptor (TR) in the in vitro T-screen assay after biotransformation and c) to investigate the endogenous metabolic capacity of the GH3 cells, the cell line used in the T-screen assay, which is a proliferation assay used for the in vitro detection of agonistic and antagonistic properties of compounds at the level of the TR. The two in vitro metabolizing systems tested the human liver S9 mix and the PCB-induced rat microsomes gave an almost complete metabolic transformation of the tested parabens and phthalates. No marked difference the effects in the T-screen assay was observed between the parent compounds and the effects of the tested metabolic extracts. The GH3 cells themselves significantly metabolized the two tested phthalates dimethyl phthalate (DMP) and diethyl phthalate (DEP). Overall the results and qualitative data from the current study show that an in vitro metabolizing system using liver S9 or microsomes could be a convenient method for the incorporation of metabolic and toxicokinetic aspects into in vitro testing for endocrine disrupting effects.

Pharmacological considerations for azole antifungal drug management in cystic fibrosis lung transplant patients.

This paper aims to present our experience in the pharmacological approach of the management of azole antifungal drugs in cystic fibrosis lung transplant patients. Cystic fibrosis (CF) lung transplantation is associated with multi-factorial care management, because of immunosuppressive requirements, risk of infections, frequency of gastro-oesophageal reflux disease, hepatic alterations and CF pharmacokinetics (PK) specificities that result in important PK variability. CF is associated with frequent colonization of the airways by filamentous fungi, especially by Aspergillus species. Today the antifungal therapeutic arsenal offers several possibilities for long-term oral therapy including azole drugs (itraconazole, voriconazole and posaconazole). Therefore, nephrotoxic amphotericin B should be avoided. The liver is important in the pharmacological profile of azole drugs, due to metabolic elimination, hepatotoxicity and PK drug-drug interaction (DDI) involving CYP3A4 metabolic inhibition. Targets for such DDI are numerous, but immunosuppressive drugs are of major concern, justifying combined therapeutic drug monitoring (TDM) of both azoles (inhibitors) and immunosuppressants (targets) on an individualized patient basis to adjust the coprescription quantitatively. The risk of long under-dosed periods, frequently addressed in this population, could justify, on a PK basis, the need for combination with an exclusive parenteral antifungal while waiting for azole relevant drug level. High PK variability, the risk of low exposure, therapeutic issues and DDI management in this complex underlying disease justify close monitoring with systematic combined TDM of azole and immunosuppressants, in case of coprescription.

Azole-based inhibitors of AKT/PKB for the treatment of cancer.

Through a combination of screening and structure-based rational design, we have discovered a series of N(1)-(5-(heterocyclyl)-thiazol-2-yl)-3-(4-trifluoromethylphenyl)-1,2-propanediamines that were developed into potent ATP competitive inhibitors of AKT. Studies of linker strand-binding adenine isosteres identified SAR trends in potency and selectivity that were consistent with binding interactions observed in structures of the inhibitors bound to AKT1 and to the counter-screening target PKA. One compound was shown to have acceptable pharmacokinetic properties and to be a potent inhibitor of AKT signaling and of in vivo xenograft tumor growth in a preclinical model of glioblastoma.

Design, synthesis, biological evaluation and pharmacokinetics of bis(hydroxyphenyl) substituted azoles, thiophenes, benzenes, and aza-benzenes as potent and selective nonsteroidal inhibitors of 17beta-hydroxysteroid dehydrogenase type 1 (17beta-HSD1).

17beta-Estradiol (E2), the most potent female sex hormone, stimulates the growth of mammary tumors and endometriosis via activation of the estrogen receptor alpha (ERalpha). 17beta-Hydroxysteroid dehydrogenase type 1 (17beta-HSD1), which is responsible for the catalytic reduction of the weakly active estrogen estrone (E1) into E2, is therefore discussed as a novel drug target. Recently, we have discovered a 2,5-bis(hydroxyphenyl) oxazole to be a potent inhibitor of 17beta-HSD1. In this paper, further structural optimizations were performed: 39 bis(hydroxyphenyl) azoles, thiophenes, benzenes, and aza-benzenes were synthesized and their biological properties were evaluated. The most promising compounds of this study show enhanced IC 50 values in the low nanomolar range, a high selectivity toward 17beta-HSD2, a low binding affinity to ERalpha, a good metabolic stability in rat liver microsomes, and a reasonable pharmacokinetic profile after peroral application. Calculation of the molecular electrostatic potentials revealed a correlation between 17beta-HSD1 inhibition and the electron density distribution.

ortho-Substituted azoles as selective and dual inhibitors of VEGF receptors 1 and 2.

We have developed a series of novel potent ortho-substituted azole derivatives active against kinases VEGFR-1 and VEGFR-2. Both specific and dual ATP-competitive inhibitors of VEGFR-2 were identified. Kinase activity and selectivity could be controlled by varying the arylamido substituents at the azole ring. The most specific molecule (17) displayed > 10-fold selectivity for VEGFR-2 over VEGFR-1. Compound activities in enzymatic and cell-based assays were in the range of activities for reported clinical and development candidates (IC50 < 100 nM), including Novartis' PTK787 (Vatalanib). High permeability of active compounds across the Caco-2 cell monolayer (> 30x10(-5) cm/min) is indicative of their potential for intestinal absorption upon oral administration.

Small-molecule screening identifies the selanazal drug ebselen as a potent inhibitor of DMT1-mediated iron uptake.

HEK293T cells overexpressing divalent metal transporter-1 (DMT1) were established to screen for small-molecule inhibitors of iron uptake. Using a fluorescence-based assay, we tested 2000 known bioactive compounds to find 3 small molecules that potently block ferrous iron uptake. One of the inhibitors, ebselen, is a seleno compound used in clinical trials as a protective agent against ischemic stroke. Ebselen inhibited Fe(II) uptake (IC(50) of approximately 0.22 microM), but did not influence Fe(III) transport or DMT1-mediated manganese uptake. An unrelated antioxidant, pyrrolidine dithiobarbamate (PDTC), also inhibited DMT1 activity (IC(50) of approximately 1.54 microM). Both ebselen and PDTC increased cellular levels of reduced glutathione. These observations indicate that Fe(II) transport by DMT1 can be modulated by cellular redox status and suggest that ebselen may act therapeutically to limit iron-catalyzed damage due to transport inhibition.

Novel C-17-heteroaryl steroidal CYP17 inhibitors/antiandrogens: synthesis, in vitro biological activity, pharmacokinetics, and antitumor activity in the LAPC4 human prostate cancer xenograft model.

New chemical entities, steroidal C-17 benzoazoles (5, 6, 9 and 10) and pyrazines (14 and 15) were rationally designed and synthesized. The key reaction for synthesis of the benzoazoles involved the nucleophilic vinylic "addition-elimination" substitution reaction of 3beta-acetoxy-17-chloro-16-formylandrosta-5,16-diene (2) and benzoazole nucleophiles, while that for synthesis of pyrazines involved palladium-catalyzed cross-coupling reaction of 17-iodoandrosta-5,16-dien-3beta-ol (13) with tributylstannyl diazines. Some of the compounds were shown to be potent inhibitors of human CYP17 enzyme as well as potent antagonist of both wild type and mutant androgen receptors (AR). The most potent CYP17 inhibitors were 3beta-hydroxy-17-(1H-benzimidazole-1-yl)androsta-5,16-diene (5, code named VN/124-1), 3beta-hydroxy-17-(5(1)-pyrimidyl)androsta-5,16-diene (15) and 17-(1H-benzimidazole-1-yl)androsta-4,16-dien-3-one (6), with IC(50) values of 300, 500 and 915 nM, respectively. Compounds 5, 6, 14 and 15 were effective at preventing binding of (3)H-R1881 (methyltrienolone, a stable synthetic androgen) to both the mutant LNCaP AR and the wild-type AR, but with a 2.2- to 5-fold higher binding efficiency to the latter. Compounds 5 and 6 were also shown to be potent pure AR antagonists. The cell growth studies showed that 5 and 6 inhibit the growth of DHT-stimulated LNCaP and LAPC4 prostate cancer cells with IC(50) values in the low micromolar range (i.e., <10 microM). Their inhibitory potencies were comparable to that of casodex but remarkably superior to that of flutamide. The pharmacokinetics of compounds 5 and 6 in mice were investigated. Following s.c. administration of 50 mg/kg of 5 and 6, peak plasma levels of 16.82 and 5.15 ng/mL, respectively, occurred after 30 to 60 min, both compounds were cleared rapidly from plasma (terminal half-lives of 44.17 and 39.93 min, respectively), and neither was detectable at 8 h. Remarkably, compound 5 was rapidly converted into a metabolite tentatively identified as 17-(1H-benzimidazol-1-yl)androsta-3-one. When tested in vivo, 5 proved to be very effective at inhibiting the growth of androgen-dependent LAPC4 human prostate tumor xenograft, while 6 was ineffective. Compound 5 (50 mg/kg/twice daily) resulted in a 93.8% reduction (P = 0.00065) in the mean final tumor volume compared with controls, and it was also significantly more effective than castration. To our knowledge, this is the first example of an antihormonal agent (an inhibitor of androgen synthesis (CYP17 inhibitor)/antiandrogen) that is significantly more effective than castration in suppression of androgen-dependent prostate tumor growth. In view of these impressive anticancer properties, compound 5 is a strong candidate for development for the treatment of human prostate cancer.

A new era of antifungal therapy.

Invasive fungal infections pose major management problems for clinicians caring for hematopoietic cell transplant patients. Two major fungal genera, Candida and Aspergillus, account for most fungal infections. Rates of systemic Candida infection range from 15% to 25%, mostly in the pre-engraftment period. Prophylaxis by fluconazole has dramatically reduced the frequency of early Candida infections. Caspofungin has recently been shown to offer an excellent alternative to amphotericin B (with less toxicity) or fluconazole (with a broader spectrum) for therapy of systemic Candida infections. Aspergillus infections occur in 15% to 20% of allogeneic hematopoietic cell transplant patients, most frequently in the post-engraftment period; they are associated with a severe diminution of cell-mediated immune responses by graft-versus-host disease and prolonged corticosteroid use. Voriconazole, a recently introduced broad-spectrum azole, has excellent activity against Aspergillus and is generally well tolerated. Voriconazole currently offers the best prospect for success and tolerance as a first-line treatment for aspergillosis. Second-line therapies include lipid formulations of amphotericin B, caspofungin, or intravenous itraconazole. Unfortunately, early initiation of therapy for aspergillosis is frequently not possible because of inaccurate diagnostics. One new diagnostic, the galactomannan assay, has recently been approved, and others are in development; these offer promise for earlier diagnosis without the need for invasive procedures. It is hoped that these new therapies and new diagnostics will usher in a new era of antifungal therapy.

Modeling of the anticancer action for radical derivatives of nitroazoles: quantitative structure-activity relationship (QSAR) study.

A QSAR analysis of the anti-tumor, anti-metastasis and anti-colony formation (for metastatic colonies) activities of eighteen nitroazoles (including metronidazole and hypoxic radiosensitizers RP-170, KU-2285 and sanazole (drug AK-2123)) and their nitro and nitroso anion radical derivatives against melanoma B16 in mice has been performed. The QSAR models were built with the use of the frontal polygon method. This approach has features of different 3D QSAR methodologies and belongs to the group of "indirect" methods. The procedure allows to build robust models with high predictive ability even in series of diverse and conformationally flexible compounds. Key atomic characteristics accompany the geometrical requirements in the analysis of local 3D molecular similarity. By variation of weight coefficients for hydrophobicity, refraction increments, and partial charge it is possible to achieve better quality of QSAR and evaluate the importance of each characteristic for biological activity under consideration. It was found that hydrophobicity, molar refraction and charge characteristics of nitro anion radical derivatives are more significant for interaction with molecular targets than those of the parent compounds and of the nitroso anion radical derivatives. Size and hydrophobic properties of substituents in nitro anion radicals play significant role for ligand-target interaction in the processes of inhibition of metastatic spreading and growth of metastatic colonies by nitroazoles. A scheme of competitive interaction of parent nitroazoles and their anion radicals with a target in organism is suggested. It can be concluded that the step of one-electron reduction of nitroazoles can be important for anticancer activity of these drugs.

Ebselen-binding equilibria between plasma and target proteins.

The antiinflammatory drug ebselen (2-phenyl-1,2-benzisoselenazo-3(2H)-one) is known to bind covalently to thiols to form seleno disulfides that, directly or indirectly, are responsible for its pharmacological effects. Due to its reactive thiol group and high plasma concentration, albumin is a preferred target of ebselen, which it binds covalently. Ebselen should not, then, be available for intracellular actions at other target proteins. We have addressed this question, and show by difference spectroscopy that the interaction of ebselen with albumin occurs stoichiometrically under ring opening, but is readily reversible in the presence of glutathione. With intact human polymorphonuclear leukocytes (PMN), a similar stoichiometric reaction with distinct spectral features was observed with ebselen that was completely abolished by pretreatment of PMN with N-ethylmaleimide, but not by selective depletion of cellular glutathione. Human platelets, again, exhibited different spectral changes upon addition of ebselen. In agreement with results reported in the literature, we show that 14C-ebselen is in dynamic equilibrium with all accessible thiol groups and, hence, despite mostly being bound covalently to albumin, it will exchange rapidly with other target proteins in PMN or platelets.