The High Stability and Selectivity of Electrochemical Sensor Using Low-Cost Diamond Nanoparticles for the Detection of Anti-Cancer Drug Flutamide in Environmental Samples.

In this study, a novel electrochemical sensor was created by fabricating a screen-printed carbon electrode with diamond nanoparticles (DNPs/SPCE). The successful development of the sensor enabled the specific detection of the anti-cancer drug flutamide (FLT). The DNPs/SPCE demonstrated excellent conductivity, remarkable electrocatalytic activity, and swift electron transfer, all of which contribute to the advantageous monitoring of FLT. These qualities are critical for monitoring FLT levels in environmental samples. Various structural and morphological characterization techniques were employed to validate the formation of the DNPs. Remarkably, the electrochemical sensor demonstrated a wide linear response range (0.025 to 606.65 µM). Additionally, it showed a low limit of detection (0.023 µM) and high sensitivity (0.403 µA µM-1 cm-2). Furthermore, the practicability of DNPs/SPCE can be successfully employed in FLT monitoring in water bodies (pond water and river water samples) with satisfactory recoveries.

Heterostructured bismuth oxide/hexagonal-boron nitride nanocomposite: A disposable electrochemical sensor for detection of flutamide.

Aquatic contamination from the accumulation of pharmaceuticals has induced severe toxicological impact to the ecological environment, especially from non-steroidal anti-inflammatory drugs (NSAIDs). Real-time monitoring of flutamide, which is a class of NSAIDs, is very significant in environmental protection. In this work, we have synthesized the hexagonal-h boron nitride decorated on bismuth oxide (Bi2O3/h-BN) based nanocomposite for the effective electrochemical detection of flutamide (FTM). The structural and morphological information of the heterostructured Bi2O3/h-BN nanocomposite was analyzed by using a sequence of characterization methods. Voltammetric techniques were used to evaluate the analytical performance of the Bi2O3/h-BN modified screen-printed carbon electrode (SPCE) for the FTM detection. The Bi2O3/h-BN modified SPCE displays a synergetic catalytic effect for the reduction of FTM due to large surface area, numerous active sites, fast charge transfer and abundant defects. The proposed electrochemical sensing platform demonstrates high selectivity, low detection limit (9.0 nM), good linear ranges (0.04-87 µM) and short response time for the detection of FTM. The feasibility of the electrochemical sensor has been proved by the successful application to determine FTM in environmental samples.

Synthesis of flutamide-conjugates.

In this paper, we designed and extended modification basing on the flutamide structure. A series of flutamide-conjugates were obtained with methyl bromoacetate and ethylenediamine. Through the synthesis of two conjugates with 3,5-bis(dodecyloxy)benzoate derivatives, these flutamide conjugates were tested for anticancer activity. Among the compounds tested, the flutamide-conjugates showed good inhibition activity against cancer cell lines U-251, PC-3 and K-562. The conjugates showed a better inhibitory effect than free flutamide and did not show activity against normal COS-7 monkey kidney fibroblast cells. It was also observed that the flutamide conjugates had an inhibitory effect against human colorectal adenocarcinoma HCT-15.

Degradation of the anticancer drug flutamide by solar photo-Fenton treatment at near-neutral pH: Identification of transformation products and in silico (Q)SAR risk assessment.

Flutamide (FLUT) is a non-steroidal drug mainly used in the treatment of prostate cancer and has been detected in the aquatic environment at ng L-1 levels. The environmental fate and effects of FLUT have not yet been studied. Conventional treatment technologies fail to completely remove pharmaceuticals, so the solar photo-Fenton process (SPF) has been proposed as an alternative. In this study, the degradation of FLUT, at two different initial concentrations in ultra-pure water, was carried out by SPF. The initial SPF conditions were pH0 5, [Fe2+]0 = 5 mg L-1, and [H2O2]0 = 50 mg L-1. Preliminary elimination rates of 53.4% and 73.4%. The kinetics of FLUT degradation could be fitted by a pseudo-first order model and the kobs were 6.57 × 10-3 and 9.13 × 10-3 min-1 t30W and the half-life times were 95.62 and 73.10 min t30W were achieved for [FLUT]0 of 5 mg L-1 and 500 µg L-1, respectively. Analysis using LC-QTOF MS identified thirteen transformation products (TPs) during the FLUT degradation process. The main degradation pathways proposed were hydroxylation, hydrogen abstraction, demethylation, NO2 elimination, cleavage, and aromatic ring opening. Different in silico (quantitative) structure-activity relationship ((Q)SAR) freeware models were used to predict the toxicities and environmental fates of FLUT and the TPs. The in silico predictions indicated that these substances were not biodegradable, while some TPs were classified near the threshold point to be considered as PBT compounds. The in silico (Q)SAR predictions gave positive alerts concerning the mutagenicity and carcinogenicity endpoints. Additionally, the (Q)SAR toolbox software provided structural alerts corresponding to the positive alerts obtained with the different mutagenicity and carcinogenicity models, supporting the positive alerts with more proactive information.

Development of Androgen-Antagonistic Coumarinamides with a Unique Aromatic Folded Pharmacophore.

First-generation nonsteroidal androgen receptor (AR) antagonists, such as flutamide (2a) and bicalutamide (3), are effective for most prostate cancer patients, but resistance often appears after several years due to the mutation of AR. Second-generation AR antagonists are effective against some of these castration-resistant prostate cancers, but their structural variety is still limited. In this study, we designed and synthesized 4-methyl-7-(N-alkyl-arylcarboxamido)coumarins as AR antagonist candidates and evaluated their growth-inhibitory activity toward androgen-dependent SC-3 cells. Coumarinamides with a secondary amide bond did not show inhibitory activity, but their N-methylated derivatives exhibited AR-antagonistic activity. Especially, 19b and 31b were more potent than the lead compound 7b, which was comparable to hydroxyflutamide (2b). Conformational analysis showed that the inactive coumarinamides with a secondary amide bond have an extended structure with a trans-amide bond, while the active N-methylated coumarinamides have a folded structure with a cis-amide bond, in which the two aromatic rings are placed face-to-face. Docking study suggested that this folded structure is important for binding to AR. Selected coumarinamide derivatives showed AR-antagonistic activity toward LNCaP cells with T877A AR, and they had weak progesterone receptor (PR)-antagonistic activity. The folded coumarinamide structure appears to be a unique pharmacophore, different from those of conventional AR antagonists.

Drug-polymer miscibility, interactions, and precipitation inhibition studies for the development of amorphous solid dispersions for the poorly soluble anticancer drug flutamide.

The major goal of this research was to successfully formulate solid dispersion (SD) of the poorly soluble anticancer drug flutamide (FLT) using various hydrophilic polymers. Furthermore, to get more insight into SD, solid-state studies (miscibility and molecular interaction) were correlated with solution study (precipitation inhibition, dissolution). Hydrophilic polymers like PVP K90, HPMC, Eudragit EPO, and PEG 8000 were used at different drug-to-polymer w/w ratios. Solid-state miscibility studies were carried out using modulated differential scanning calorimetry (MDSC). SDs were prepared using solvent-evaporation technique and characterized by powder X-ray diffraction (PXRD) and MDSC. Infrared, Raman spectroscopy and molecular modeling were used to investigate drug-polymer interactions in the dispersions. Precipitation inhibition studies were carried out at various FLT-hydrophilic polymer ratios. Precipitation inhibition studies showed that PEG 8000 has the highest efficiency, followed by PVP K90, while HPMC and EPO showed no effect on precipitation inhibition. In the solid-state, MDSC of the physical mixture (PM) suggested that FLT is miscible to a greater extent with EPO and PEG 8000. Characterization of the amorphous dispersions using MDSC and PXRD concluded that FLT transformed from crystalline to amorphous form in the presence of PVP K90 and PEG 8000. Spectroscopic results confirmed stronger interaction of FLT with PVP K90 and PEG 8000, thereby confirming the in-solution precipitation and molecular modeling binding energy results. Amorphous dispersions formulated with PVP and PEG were stable and showed higher dissolution, an important property necessary to improve the physicochemical properties and drug delivery of poorly soluble anticancer drug FLT.

Combination of 1H nuclear magnetic resonance spectroscopy and principal component analysis to evaluate the lipid fluidity of flutamide-encapsulated lipid nanoemulsions.

The lipid fluidity of various lipid nanoemulsions (LNEs) without and with flutamide (FT) and containing one of two neutral lipids, one of four phosphatidylcholines as a surfactant, and sodium palmitate as a cosurfactant was investigated by the combination of 1H nuclear magnetic resonance (NMR) spectroscopy and principal component analysis (PCA). In the 1H NMR spectra, the peaks from the methylene groups of the neutral lipids and surfactants for all LNE preparations showed downfield shifts with increasing temperature from 20 to 60 °C. PCA was applied to the 1H NMR spectral data obtained for the LNEs. The PCA resulted in a model in which the first two principal components (PCs) extracted 88% of the total spectral variation; the first PC (PC-1) axis and second PC (PC-2) axis accounted for 73 and 15%, respectively, of the total spectral variation. The Score-1 values for PC-1 plotted against temperature revealed the existence of two clusters, which were defined by the neutral lipid of the LNE preparations. Meanwhile, the Score-2 values decreased with rising temperature and reflected the increase in lipid fluidity of each LNE preparation, consistent with fluorescence anisotropy measurements. In addition, the changes of Score-2 values with temperature for LNE preparations with FT were smaller than those for LNE preparations without FT. This indicates that FT encapsulated in LNE particles markedly suppressed the increase in lipid fluidity of LNE particles with rising temperature. Thus, PCA of 1H NMR spectra will become a powerful tool to analyze the lipid fluidity of lipid nanoparticles. Graphical abstract ᅟ.

Flutamide-Loaded Zein Nanocapsule Hydrogel, a Promising Dermal Delivery System for Pilosebaceous Unit Disorders.

Zein is a naturally occurring corn protein having similarity to skin keratin. Owing to its hydrophobicity and biodegradability, zein nanocarriers are promising drug delivery vehicles for hydrophobic dermatological drugs. In this study, zein-based nanocapsules (ZNCs) were exploited for the first time as dermal delivery carriers for flutamide (FLT), an antiandrogen used for the management of pilosebasceous unit disorders. FLT-loaded ZNC of appropriate particle size and negative surface charge were prepared by nanoprecipitation method. The dermal permeation and skin retention of FLT from ZNCs were studied in comparison to corresponding nanoemulsion (NE) and hydroalcoholic drug solution (HA). ZNCs showed a significantly lower permeation flux compared to NE and HA while increasing the skin retention of FLT. Confocal laser scanning microscopy (CLSM) demonstrated the follicular localization of the fluorescently labeled NCs. The incorporation of NCs in chitosan gel or Carbomer® 934 gel was studied. Carbomer® gel increased the skin retention of FLT compared to chitosan gel. Accordingly, Carbomer® hydrogel embedding FLT-loaded ZNCs is a promising inexpensive, biocompatible dermal delivery nanocarrier for localized therapy of PSU disorders suitable for application on oily skin.

Effects of testosterone on mean arterial pressure and aquaporin (AQP)-1, 2, 3, 4, 6 and 7 expressions in the kidney of orchidectomized, adult male Sprague-Dawley rats.

We hypothesized that higher blood pressure in males than females could be due to testosterone effects on aquaporin (AQP) expression in kidneys. METHODS: Orchidectomized adult male Sprague-Dawley (SD) rats received seven days subcutaneous testosterone treatment (125 µg/kg/day or 250 µg/kg/day), with or without flutamide or finasteride. Following completion of treatment, MAP was determined in rats under anaesthesia via carotid artery cannulation. In another cohort of rats, kidneys were removed following sacrifice and AQP-1, 2, 3, 4, 6 and 7 protein and mRNA levels were determined by Western blotting and Real-time PCR respectively. Distribution of AQP subunits' protein in the nephrons were visualized by immunofluorescence. RESULTS: Testosterone caused MAP, AQP-1, 2, 4, 6 and 7 protein and mRNA levels in kidneys to increase while AQP-3 protein and mRNA levels in kidneys to decrease (p < 0.05). AQP-1 and 7 were found to be distributed in the proximal convoluted tubule (PCT) while AQP-2, 3, 4 and 6 were found to be distributed in the collecting ducts (CD). Effects of testosterone were antagonized by flutamide and finasteride. CONCLUSIONS: Elevated expression of AQP-1, 2, 4, 6 and 7 under testosterone influence in kidneys could lead to increase H2O reabsorption which eventually lead to increase in blood pressure.

A New Method To Identify Physically Stable Concentration of Amorphous Solid Dispersions (I): Case of Flutamide + Kollidon VA64.

In this paper, a novel approach to determine stable concentration in API-polymer systems is presented. As a model, binary amorphous mixtures flutamide (FL) drug with a copolymer Kollidon VA64 (PVP/VA) have been used. It is worthwhile to note that finding an effective method to achieve this goal is a matter of great importance because physical stability of the amorphous pharmaceuticals is the key issue that is investigated worldwide. Due to the fact that molecular dynamics was found to be the crucial factor affecting physical stability of disordered pharmaceuticals, we examined it for both neat FL and its PVP/VA mixtures by means of broadband dielectric spectroscopy (BDS). Thorough investigation of the impact of polymeric additive on the molecular mobility of disordered FL reveals unusual, previously unreported behavior. Namely, simultaneously with the beginning of the recrystallization process, we observe some transformation from unstable supersaturated concentration of investigated mixture to the different, unknown concentration of FL-PVP/VA. Observed, during BDS experiment, transformation enables us to determine the limiting, highly physically stable concentration of FL in PVP/VA polymer (saturated solution), which is equivalent to FL + 41% wt. of PVP/VA. The described high physical stability of this unveiled system has been confirmed by means of long-term XRD measurements. According to our knowledge, this is the first time when such a behavior has been observed by means of BDS.

Antiandrogens act as selective androgen receptor modulators at the proteome level in prostate cancer cells.

Current therapies for prostate cancer include antiandrogens, inhibitory ligands of the androgen receptor, which repress androgen-stimulated growth. These include the selective androgen receptor modulators cyproterone acetate and hydroxyflutamide and the complete antagonist bicalutamide. Their activity is partly dictated by the presence of androgen receptor mutations, which are commonly detected in patients who relapse while receiving antiandrogens, i.e. in castrate-resistant prostate cancer. To characterize the early proteomic response to these antiandrogens we used the LNCaP prostate cancer cell line, which harbors the androgen receptor mutation most commonly detected in castrate-resistant tumors (T877A), analyzing alterations in the proteome, and comparing these to the effect of these therapeutics upon androgen receptor activity and cell proliferation. The majority are regulated post-transcriptionally, possibly via nongenomic androgen receptor signaling. Differences detected between the exposure groups demonstrate subtle changes in the biological response to each specific ligand, suggesting a spectrum of agonistic and antagonistic effects dependent on the ligand used. Analysis of the crystal structures of the AR in the presence of cyproterone acetate, hydroxyflutamide, and DHT identified important differences in the orientation of key residues located in the AF-2 and BF-3 protein interaction surfaces. This further implies that although there is commonality in the growth responses between androgens and those antiandrogens that stimulate growth in the presence of a mutation, there may also be influential differences in the growth pathways stimulated by the different ligands. This therefore has implications for prostate cancer treatment because tumors may respond differently dependent upon which mutation is present and which ligand is activating growth, also for the design of selective androgen receptor modulators, which aim to elicit differential proteomic responses dependent upon cellular context.

A Molecular Modeling Study of the Hydroxyflutamide Resistance Mechanism Induced by Androgen Receptor Mutations.

Hydroxyflutamide (HF), an active metabolite of the first generation antiandrogen flutamide, was used in clinic to treat prostate cancer targeting androgen receptor (AR). However, a drug resistance problem appears after about one year's treatment. AR T877A is the first mutation that was found to cause a resistance problem. Then W741C_T877A and F876L_T877A mutations were also reported to cause resistance to HF, while W741C and F876L single mutations cannot. In this study, molecular dynamics (MD) simulations combined with the molecular mechanics generalized Born surface area (MM-GBSA) method have been carried out to analyze the interaction mechanism between HF and wild-type (WT)/mutant ARs. The obtained results indicate that AR helix 12 (H12) plays a pivotal role in the resistance of HF. It can affect the coactivator binding site at the activation function 2 domain (AF2, surrounded by H3, H4, and H12). When H12 closes to the AR ligand-binding domain (LBD) like a lid, the coactivator binding site can be formed to promote transcription. However, once H12 is opened to expose LBD, the coactivator binding site will be distorted, leading to invalid transcription. Moreover, per-residue free energy decomposition analyses indicate that N705, T877, and M895 are vital residues in the agonist/antagonist mechanism of HF.

Histological assessment of follicular delivery of flutamide by solid lipid nanoparticles: potential tool for the treatment of androgenic alopecia.

CONTEXT: Flutamide is a potent anti-androgen with the several unwanted side effects in systemic administration, therefore, it has attracted special interest in the development of topically applied formulations for the treatment of androgenic alopecia. OBJECTIVE: The purpose of this study was to prepare and characterize the solid lipid nanoparticles (SLNs) of Flutamide for follicular targeting in the treatment of the androgenic alopecia. METHODS: Flutamide-loaded SLNs, promising drug carriers for topical application were prepared by hot melt homogenization method. Drug permeation and accumulation in the exercised rat skin and histological study on the male hamsters were performed to assess drug delivery efficiency in vitro and in vivo, respectively. RESULTS: The optimized Flutamide-loaded SLNs (size 198 nm, encapsulation efficiency percentage 65% and loading efficiency percentage 3.27%) exhibited a good stability during the period of at least 2 months. The results of X-ray diffraction showed Flutamide amorphous state confirming uniform drug dispersion in the SLNs structure. Higher skin drug deposition (1.75 times) of SLN formulation compared to Flutamide hydroalcoholic solution represented better localization of the drug in the skin. The in vivo studies showed more new hair follicle growth by utilizing Flutamide-loaded SLNs than Flutamide hydroalcoholic solution which could be due to the higher accumulation of SLNs in the hair follicles as well as slowly and continues release of the Flutamide through the SLNs maximizing hair follicle exposure by antiandrogenic drug. CONCLUSION: It was concluded Flutamide-loaded SLN formulation can be used as a promising colloidal drug carriers for topical administration of Flutamide in the treatment of androgenic alopecia.

19F Nuclear Magnetic Resonance Spectrometric Determination of the Partition Coefficients of Flutamide and Nilutamide (Antiprostate Cancer Drugs) in a Lipid Nano-Emulsion and Prediction of Its Encapsulation Efficiency for the Drugs.

To design a useful lipid drug carrier having a high encapsulation efficiency (EE%) for the antiprostate cancer drugs flutamide (FT) and nilutamide (NT), a lipid nano-emulsion (LNE) was prepared with soybean oil (SO), phosphatidylcholine (PC), and sodium palmitate, and the partition coefficients (K ps) of the drugs for the LNE were determined by 19F nuclear magnetic resonance (NMR) spectrometry. The 19F NMR signal of the trifluoromethyl group of both drugs showed a downfield shift from an internal standard (trifluoroethanol) and broadening according to the increase in the lipid concentration due to their interaction with LNE particles. The difference in the chemical shift (Δδ) of each drug caused by the addition of LNE was measured under different amounts of LNE, and the K p values were calculated from the Δδ values. The results showed that FT has higher lipophilicity than NT. The total lipid concentration (SO + PC) required to encapsulate each drug into LNE with an EE% of more than 95% was calculated from the K p values as 93.3 and 189.9 mmol/L for FT and NT, respectively. For an LNE prepared with the total lipid concentration of 215 mmol/L, the predicted EE% values were 98 and 96% for FT and NT, respectively, while the experimental EE% values determined by a centrifugation method were approximately 99% for both drugs. Thus, the 19F NMR spectrometric method is a useful technique to obtain the K p values of fluorinated drugs and thereby predict the theoretical lipid concentrations and prepare LNEs with high EE% values.

Androgens promote the acquisition of maturation competence in bovine oocytes.

Recent studies in mice suggest that androgens are important for normal follicle development. However, there have been few reports concerning the action of androgens in the growth of oocytes from large animals. The purpose of this study was to determine the roles of androgens in bovine oocyte growth in vitro. Oocyte-granulosa cell complexes (OGCs) collected from 0.4-0.7 mm early antral follicles were cultured for 14 days with 17ß-estradiol (E2) and a non-aromatizable androgen, dihydrotestosterone (DHT). We also examined the ability of an androgen receptor (AR) inhibitor, hydroxyflutamide, to antagonize the effect of androgens on the oocytes. During growth culture, the OGC structures collapsed in the medium with DHT alone, while in the presence of E2, the OGC structures were maintained. In the medium with both androgens and E2, the mean diameter of oocytes was increased from 95 µm to around 120 µm, larger than those grown with E2 alone (115 µm). Also in the maturation culture, oocytes grown with androgens (A4 or DHT) and E2 showed higher percentages of metaphase II oocytes (63% or 69%, respectively) than those grown with E2 alone (32%). Moreover, these maturation rates were decreased by hydroxyflutamide in a dose-dependent manner. Immunostaining showed that ARs were expressed in oocytes and granulosa cells in early antral follicles, and the nuclei of granulosa cells showed intense AR expression. In conclusion, although E2 supports the OGC structure, additional androgens promote oocyte growth and their acquisition of meiotic competence via AR during in vitro growth culture.

[Extraction of flutamide from aqueous solutions].

The objective of the present study was to elucidate peculiarities of flutamide extraction from aqueous solutions using he liquid-liquid extraction technique and spectrophotometry in the visible spectrum. The results of flutamide extraction were shown to depend on the chemical nature of the extractant, pH of the aqueous phase as well as its saturation with water and electrolytes. The study has demonstrated that the optimal conditions for flutamide extraction by ethylacetate are created by using the aqueous phase with the pH values in the range from 1.0 to 12.0. We have calculated extraction factor necessary for the luberation of the desired amount of flutamide from the aqueous solutions with the use of a concrete solvent.

Optimization of Electrochemical Sensitivity in Anticancer Drug Quantification through ZnS@CNS Nanosheets: Synthesis via Accelerated Sonochemical Methodology.

Zinc sulfide/graphitic Carbon Nitride binary nanosheets were synthesized by using a novel sonochemical pathway with high electrocatalytic ability. The as- obtained samples were characterized by various analytical methods such as Transmission Electron Microscopy (TEM), Field emission scanning electron microscopy (FESEM), Energy-dispersive X-ray spectroscopy (EDS), X-ray diffraction analysis (XRD), and X-ray photoelectron spectroscopy (XPS) to evaluate the properties of ZnS@CNS synthesized by this new route. Subsequently, the electrical and electrochemical performance of the proposed electrodes were characterized by using EIS and CV to establish an electroactive ability of the nanocomposites. The complete properties like structural and physical of ZnS@CNS were analyzed. As-prepared binary nanocomposite was applied towards the detection of anticancer drug (flutamide) by various electrochemical methods such as cyclic voltammetry (CV), differential pulse voltammetry (DPV) and amperometry. The glassy carbon electrode modified with a ZnS@CNS composite demonstrates a remarkable electrocatalytic efficiency for detecting flutamide in a pH 7.0 (PBS). The composite modified electrode shows synergistic effect of ZnS and CNS catalyst. The electrochemical sensing performance of the linear range was improved significantly due to high electroactive sites and rapid electron transport pathways. Crucially, the electrochemical method was successfully demonstrated in biological fluids which reveals its potential real-time applicability in the analysis of drug.

Polymeric nanocarriers for magnetic targeted drug delivery: preparation, characterization, and in vitro and in vivo evaluation.

In this paper the preparation of magnetic nanocarriers (MNCs), containing superparamagnetic domains, is reported, useful as potential magnetically targeted drug delivery systems. The preparation of MNCs was performed by using the PHEA-IB-p(BMA) graft copolymer as coating material through the homogenization-solvent evaporation method. Magnetic and nonmagnetic nanocarriers containing flutamide (FLU-MNCs) were prepared. The prepared nanocarriers have been exhaustively characterized by dynamic light scattering (DLS), transmission electron microscopy (TEM), and magnetic measurements. Biological evaluation was performed by in vitro cytotoxicity and cell uptake tests and in vivo biodistribution studies. Magnetic nanocarriers showed dimensions of about 300 nm with a narrow size distribution, an amount of loaded FLU of 20% (w/w), and a superparamagnetic behavior. Cell culture experiments performed on prostate cancer cell line LNCaP demonstrated the cytotoxic effect of FLU-MNCs. In vivo biodistribution studies carried out by the application of an external magnetic field in rats demonstrated the effect of the external magnet on modifying the biodistribution of FLU-MNCs. FLU-MNCs resulted efficiently internalized by tumor cells and susceptible to magnetic targeting by application of an external magnetic field. The proposed nanocarriers can represent a very promising approach to obtain an efficient magnetically targeted anticancer drug delivery system.

Powder crystallography of pharmaceutical materials by combined crystal structure prediction and solid-state 1H NMR spectroscopy.

A protocol for the ab initio crystal structure determination of powdered solids at natural isotopic abundance by combining solid-state NMR spectroscopy, crystal structure prediction, and DFT chemical shift calculations was evaluated to determine the crystal structures of four small drug molecules: cocaine, flutamide, flufenamic acid, and theophylline. For cocaine, flutamide and flufenamic acid, we find that the assigned (1)H isotropic chemical shifts provide sufficient discrimination to determine the correct structures from a set of predicted structures using the root-mean-square deviation (rmsd) between experimentally determined and calculated chemical shifts. In most cases unassigned shifts could not be used to determine the structures. This method requires no prior knowledge of the crystal structure, and was used to determine the correct crystal structure to within an atomic rmsd of less than 0.12 Å with respect to the known reference structure. For theophylline, the NMR spectra are too simple to allow for unambiguous structure selection.

Antitumor agents 290. Design, synthesis, and biological evaluation of new LNCaP and PC-3 cytotoxic curcumin analogs conjugated with anti-androgens.

In our continuing study of curcumin analogs as potential anti-prostate cancer drug candidates, 15 new curcumin analogs were designed, synthesized and evaluated for cytotoxicity against two human prostate cancer cell lines, androgen-dependent LNCaP and androgen-independent PC-3. Twelve analogs (5-12, 15, 16, 19, and 20) are conjugates of curcumin (1) or methyl curcumin (2) with a flutamide- or bicalutamide-like moiety. Two compounds (22 and 23) are C4-mono- and difluoro-substituted analogs of dimethyl curcumin (DMC, 21). Among the newly synthesized conjugates compound 15, a conjugate of 2 with a partial bicalutamide moiety, was more potent than bicalutamide alone and essentially equipotent with 1 and 2 against both prostate tumor cell lines with IC(50) values of 41.8 µM (for LNCaP) and 39.1 µM (for PC-3). A cell morphology study revealed that the cytotoxicity of curcumin analogs or curcumin-anti-androgen conjugates detected from both prostate cancer cell lines might be due to the suppression of pseudopodia formation. A molecular intrinsic fluorescence experiment showed that 1 accumulated mainly in the nuclei, while conjugate 6 was distributed in the cytosol. At the tested conditions, anti-androgens suppressed pseudopodia formation in PC-3 cells, but not in LNCaP cells. The evidence suggests that distinguishable target proteins are involved, resulting in the different outcomes toward pseudopodia suppression.