A novel areneisonitrile Tc complex inhibits the transport activity of MDR P-glycoprotein.

P-glycoprotein (Pgp), the product of the multidrug resistance (MDR1) gene, has been an important cancer target for development of MDR modulators that act to inhibit Pgp efflux transport activity. From a series of novel substituted areneisonitrile analogues of Tc-sestamibi, a known Pgp transport substrate, emerged the hexakis(3,4,5-trimethoxyphenylisonitrile)Tc(I) complex (Tc-TMPI) as a potential modulator of Pgp. Tracer 99mTc-TMPI showed net cellular accumulation in inverse proportion to expression of Pgp and enhancement upon addition of classic MDR modulators. At pharmacological concentrations, the carrier-added 94Tc-TMPI complex showed potent inhibition of Pgp-mediated 99mTc-sestamibi transport (EC50, 1.1 +/- 0.2 microM) and displacement of a Pgp-specific photolabel in a concentration-dependent manner. We conclude that 99Tc-TMPI directly inhibited Pgp transport activity and serves as a convenient template for development of nonradioactive Re(I) analogues as novel MDR modulators.

Optimization of the synthesis of peptide combinatorial libraries using a one-pot method.

Conditions for the synthesis of synthetic peptide combinatorial libraries (SPCLs) from mixtures of amino acids were explored. In a one-pot synthesis, the effect of the starting concentrations of amino acids on the resulting library composition was studied, and the optimum balance of amino acids was determined. Protein sequencing, MALDI-TOF, and amino acid analysis were used for the evaluation of the libraries, and their relative merits-are discussed. The effects of continuous-flow automated synthesis instrumentation in conjunction with polyethylene glycol-polystyrene (PEG-PS) graft supports and various cleavage cocktails on the successful synthesis of SPCLs were examined.

Stereochemical probes for the estrogen receptor: synthesis and receptor binding of (17 alpha,20E/Z)-21-phenyl-19-norpregna-1,3,5(10), 20-tetraene-3,17 beta-diols.

Previous studies from our laboratory using 17 alpha-E- and 17 alpha-Z-halovinyl and phenylthiovinyl estradiols demonstrated a marked preference for the Z stereochemistry and a significant steric tolerance for the Z-vinyl substituent. To further explore the extent of that stereochemical preference and steric tolerance we have prepared stereoselectively the 17 alpha-E- and 17 alpha-Z-phenylvinyl estradiols (E- and Z-styrylestradiols). The results, in addition to demonstrating a facile preparation of the target compounds, supported the previously observed stereochemical and steric effects. The relative binding affinities for the Z isomer were 3-4 fold greater than the E isomer at both 4 degrees C and 25 degrees C, and only one-half to one-fourth those of estradiol under similar conditions. The developing model for ligand-accessible space within the estrogen receptor suggests that Z-phenylvinyl estradiols may provide interesting and useful probes for mapping the receptor.

Synthesis and estrogen receptor binding of (17 alpha, 20E)- and (17 alpha, 20Z)-21-phenylthio- and 21-phenylseleno-19-norpregna-1,3,5(10),20-tetraene-3,17 beta-diols.

Previous studies from our laboratory using 17 alpha-E- and 17 alpha-Z-halovinyl estradiols demonstrated a marked enhancement of receptor binding by the Z-isomers. This suggested tolerance at the 17 alpha-position was not previously observed by investigations using 16 alpha and 17 alpha-substituted estradiols. Because of the synthetic access provided by vinyl tin chemistry, we prepared the 17 alpha-E and Z-phenylthiovinyl and phenylselenovinyl estradiols and compared their binding characteristics to those of the previously reported 16 alpha/17 alpha-phenylseleno and methylseleno estradiols. The results, in addition to demonstrating a facile preparation of the target compounds, indicated that significant receptor affinity was retained by these compounds (relative binding affinity = 24.5-117). The highest affinity was demonstrated by the 17 alpha-Z-phenylthiovinyl estradiol 5a, which, by molecular modeling, exhibited a significantly different molecular conformation from the corresponding 17 alpha-E-phenylthiovinyl isomer or the 17 alpha-phenyl-thioethynyl analog. The current series possessed better binding characteristics than the phenylseleno and methylseleno estradiols but somewhat poorer binding than the 17 alpha-E/Z-halovinyl series. The observations suggest that some steric limitations exist in a portion of the 17 alpha-region, and that the region is better accessed by compounds possessing Z-vinyl stereochemistry.

Novel hexakis(areneisonitrile)technetium(I) complexes as radioligands targeted to the multidrug resistance P-glycoprotein.

Transport substrates and modulators of the human multidrug resistance (MDR1) P-glycoprotein (Pgp) are generally lipophilic cationic compounds, many with substituted aryl moieties. We sought to synthesize aromatic technetium-isonitrile complexes to enable functional detection in vivo of Pgp expression in tissues. A series of substituted aromatic isonitrile analogs were synthesized from their corresponding amines by reaction with dichlorocarbene under phase transfer-catalyzed conditions, and the non-carrier-added hexakis(areneisonitrile)Tc-99m(I) complexes were produced by reaction with pertechnetate in the presence of sodium dithionite. Cellular accumulation in vitro, whole body biodistribution, and the imaging properties of these lipophilic, monocationic organometallic complexes were determined in Chinese hamster lung fibroblasts expressing MDR Pgp, in normal rats, and in rabbits, respectively. For this initial series, verapamil (50 microM), the classical Pgp modulator, significantly enhanced cellular accumulation or displaced binding of Tc complexes of 1b, 1d, 1h, 2a, 2d, 3a, and 3b, indicative of targeted interactions with Pgp. Most complexes, despite their modestly high lipophilicity, were excluded by the blood/brain barrier, and several complexes displayed simultaneously high hepatobiliary and renal excretion in vivo, consistent with the physiological expression pattern of Pgp in these tissues. Selected Tc- and Re-areneisonitrile complexes of this class have potential applicability to the functional imaging and modulation, respectively, of MDR Pgp in human tissues.

Contrast material-carrying liposomes: biodistribution, clearance, and imaging characteristics.

PURPOSE: To assess the biodistribution, clearance, and computed tomographic (CT) imaging characteristics of interdigitation-fusion (IF) liposomes that carry iotrolan in their aqueous phases. MATERIALS AND METHODS: Biodistribution and clearance of liposomes containing iotrolan produced with the IF method (IF vesicles) were assessed in rats. CT scans of rats and dogs were obtained after injection of IF vesicles at 100 and 250 mg of iodine per kilogram of body weight. RESULTS: A high initial uptake (63%-96% of the injected dose) was found in the liver and spleen. Liver elimination showed half-lives to be 12.9 days at 250 mg of iodine per kilogram, 10.9 days at 100 mg, and 8.7 days at 25 mg. At 250 mg of iodine per kilogram, the rats had an average of 96 HU of hepatic and 321 HU of splenic enhancement. The dogs had 116 HU of hepatic and 65 HU of splenic enhancement. CONCLUSION: IF liposomes have favorable biodistribution, clearance, and imaging characteristics as hepatosplenic contrast agents.

The use of pentafluorophenyl derivatives for the 18F labelling of proteins.

A simple, one-step method for the radiolabelling of proteins with 18F is described. A series of pentafluorophenyl derivatives were synthesized and tested for 18F exchange using tetrabutylammonium-[18F]fluoride in DMSO, with microwave heating. A number of the compounds examined incorporated 18F quickly and in high yield. Two compounds, pentafluorobenzaldehyde and 2,3,5,6-tetrafluorophenylpentafluorobenzoate, were used to label HSA in good yield. The methods produce low specific activity labelled proteins, but are fast. The yields are reasonable and the reagents do not require a separate modification or activation step for protein labelling.

Structure-localization relationships of 11C-labeled phentermine derivatives: effect of aromatic substitution.

A series of phentermine analogs, including the unsubstituted, the para-F, -Cl, -Br and -I, and the meta-CF3 derivatives, were labeled by [11C]methylation and evaluated in rats to determine the structure-localization relationships for this class of regional cerebral blood flow imaging agents. All the phentermines were well-localized in the brain; however, only the para-substituted agents were well-retained. Localization in the nontarget tissue was affected by the lipophilicity of the substituent. Comparison with the radioiodinated analogs showed virtually identical results, which suggests that the compounds were not significantly metabolized. The agent with the best biodistribution characteristics was the N-[11C]methyl-p-iodophentermine, with the p-bromo analog almost equivalent.

Synthesis and evaluation of 1-[11C]methyl-4-aryl-piperazinium salts as myocardial imaging agents.

Our previous study with radioiodinated 1-methyl-1-(2-hydroxyethyl)-4- phenylpiperazinium derivatives indicated that these compounds localize selectively in the rat myocardium. Based upon these results, 1-[11C]methyl-1-alkyl-4-phenylpiperazinium cations were synthesized and evaluated for their potential as myocardial imaging agents. Biodistribution studies were performed in rats and imaging studies in normal dogs. The results indicate high concentrations of 3-4% ID/g in the rat heart at 5 min, comparable to those obtained with the radioiodinated analogs. Sequential imaging of the canine heart showed a similar pattern of uptake. The level of activity reached at 20 min was maintained over the 1-h time period. These results suggest that the phenylpiperazinium agents may have potential application as myocardial imaging agents.

Comparative effects of neutral dipolar compounds and lipophilic anions on technetium 99m-hexakis (2-methoxyisobutyl isonitrile) accumulation in cultured chick ventricular myocytes.

RATIONALE AND OBJECTIVES: Non-flow-dependent myocellular accumulation and uptake kinetics of the myocardial perfusion and viability imaging agent, hexakis (2-methoxyisobutyl isonitrile) technetium 99m(I) (Tc-SESTAMIBI), are thermodynamically driven by large negative sarcolemmal and mitochondrial membrane potentials, and can be enhanced by addition of the lipophilic anion, tetraphenylborate (TPB). To further understand the general properties required of a co-administered compound for increasing the kinetic response of Tc-SESTAMIBI to membrane potential, a systematic appraisal of additional candidate lipid-soluble anions and neutral dipolar compounds was undertaken. METHODS: Each compound was biologically tested for its ability to enhance Tc-SESTAMIBI accumulation in a cultured heart cell model, and electronic dipole moments were evaluated using semi-empirical molecular orbital calculations. RESULTS: Of this series, phloretin (100 microM), TPB (10 microM), and to a lesser degree, 8-anilino-1-naphthalene sulfonate (100 microM) enhanced myocellular accumulation of Tc-SESTAMIBI. Phloretin enhancement was pH-dependent, showing maximal effect at pH 7.4, and was not additive to the augmentation induced by TPB. A series of additional lipid soluble anions and structural analogues of phloretin were without effect. CONCLUSION: Although selected compounds enhanced Tc-SESTAMIBI accumulation, overall, no direct relationship of dipole moment to biologic enhancement was demonstrated.