Anil Mishra

Polylysine-Gd-DTPAn and polylysine-Gd-DOTAn coupled to anti-CEA F(ab')2 fragments as potential immunocontrast agents. Relaxometry, biodistribution, and magnetic resonance imaging in nude mice grafted with human colorectal carcinoma.

RATIONALE AND OBJECTIVESnImmunocontrast agents used for magnetic resonance imaging require antibodies that preserve the immunoreactivity while containing a high number of chelated paramagnetic ions.nnnMETHODSnAnti-CEA F(ab)2 fragments were coupled to polylysine-Gd-DOTA and polylysine-Gd-DTPA. A paramagnetic load as high as n = 24 to 28 metal ions per antibody was reached.nnnRESULTSnThe immunoreactivity of the gadolinium (Gd)-labeled anti-CEA F(ab)2 immunoconjugates was 80% to 85%. Compared with that of commercial chelates, the relaxivity (R1) increase is as follows: Gd-DTPA < Gd-DOTA < Gd-H2O < PL-Gd-DTPA24-28 < PL-Gd-DTPA24-28 F(ab)2 < PL-Gd-DOTA24-28 < PL-Gd-DOTA24-28 F(ab)2. 1H nuclear magnetic relaxation dispersion data of immunoconjugates showed that the high relaxivity enhancement was the result of a reduction of the molecular tumbling rate. Twenty-four hours after intravenous injection of 50 micrograms (1 mumol Gd/kg) of Gd-labeled immunoconjugates to nude mice grafted with human colorectal carcinoma LS 174T, the tumor uptake was 10% to 15%, resulting in an increase of R1 of up to 15% to 20% versus noninjected mice. No difference was found between PL-Gd-DTPA24-28 F(ab)2 and PL-Gd-DOTA24-28 F(ab)2 immunoconjugates for tumor, liver, and kidney uptake. A high signal intensity of tumor was observed in 50% of the tested mice.

Mucoadhesive chitosan microspheres of carvedilol for nasal administration

The aim of the present study was to develop and characterize chitosan mucoadhesive microspheres of carvedilol (CRV) for nasal delivery to improve bioavailability for treatment of hypertension and angina pectoris. The microspheres were prepared by emulsification-cross-linking method and evaluated for size, shape, entrapment efficiency (EE), in vitro mucoadhesion, in vitro drug release, differential scanning calorimetry (DSC) and X-ray diffraction (XRD). The mucoadhesive properties were also evaluated by Freundlich and Langmuir adsorption isotherms. In vivo tests were carried out in rabbits. The microspheres were spherical with size of 20–50u2009µm, which is favorable for intranasal absorption. The EE was observed from 42% to 68% while percentage mucoadhesion was from 74% to 88%. A strong interaction between mucin and chitosan microspheres was detected explaining adsorption with electrostatic interaction. The microspheres released around 75% of drug in 8u2009h. DSC and XRD studies revealed that CRV was molecularly dispersed. The absorption rate was rapid and the absolute bioavailability was high, 72.29%. The gamma scintigraphy indicated that the microspheres cleared slowly from the nasal cavity. It was concluded that chitosan microspheres could be used to deliver CRV following nasal administration for improving the bioavailability.

Design and development of nanoemulsion drug delivery system of amlodipine besilate for improvement of oral bioavailability

Nanoemulsion (NE) of amlodipine besilate (AB) was developed by spontaneous emulsification method with the aim to enhance the solubility and oral bioavailability of AB and to achieve localized delivery of drug at target site. Pseudoternary phase diagrams were constructed to identify the NE region. The selected formulations from NE region were subjected to droplet size analysis, partitioning study and in vitro drug release. The partition coefficient was calculated and correlated with percent dissolution efficiency as a tool to predict in vitro drug release from NEs. The release of drug from NEs was significantly higher (pu2009<u20090.01) than the marketed tablet formulation. The optimal formulation contained 15% Labrafil M, 35% [Tween 80: ethanol (2:1)], and 50% by weight aqueous phase (NE3) was characterized by transmission electron microscopy (TEM) and for thermodynamic stability. The pharmacokinetics and biodistribution studies of the optimized radiolabeled formulation (99mTc-labeled) in mice (p.o.) demonstrated a relative bioavailability of 475% against AB suspension. In almost all the tested organs, the uptake of AB from NE was significantly higher (pu2009<u20090.05) than AB suspension especially in heart with a drug targeting index of 44.1%, also confirming the efficacy of nanosized formulation at therapeutic site. A three times increase in the overall residence time of NE further signifies the advantage of NEs as drug carriers for enhancing bioavailability of AB.

A convenient, novel approach for the synthesis of polyaza macrocyclic bifunctional chelating agents

Abstract The convenient, synthetically useful bifunctional chelating agents, (10-p-aminobenzyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triacetate (p-ABz-DO3A) 3 and (11-p-aminobenzyl)-1,4,8,11-tetraazacyclotetradecane-1,4,8-triacetate (p-ABz-TE3A) 6 , were obtained by reaction of an ethyl bromoacetate with 1,4,7,10-tetraazacyclododecane and 1,4,8,11-tetraazacyclotetradecane followed by reaction with nitrobenzyl bromide. This method proved more efficient than any described in the literature since the overall yield in a two-step synthesis sequence starting from tetraazamacrocycles was 68%.

Radiolabeling, pharmacoscintigraphic evaluation and antiretroviral efficacy of stavudine loaded 99mTc labeled galactosylated liposomes

The study was aimed to optimize radiolabeling with 99mTc, to determine the antiretroviral activity and to study the biodistribution of 99mTc labeled galactosylated liposomes loaded with stavudine. Liposomes were prepared using reverse-phase evaporation method followed by extrusion through 200nm polycarbonate membranes. The galactosylated liposomes were assessed for in vitro ligand-specific activity and the aggregation of galactosylated liposomes was found to increase as lectin concentration was increased from 5microg/ml to 30microg/ml. Free stavudine and stavudine loaded plain and galactosylated liposomes were radiolabeled with 99mTc by direct labeling method using stannous chloride as a reducing agent. Labeling method was optimized for stannous chloride quantity to achieve maximum labeling efficiency >95%. Antiretroviral activity was determined using human immunodeficiency virus-1 (HIV) infected MT2 cell line. A dose-dependent inhibition of p24 production was observed upon treatment of HIV-1 infected MT2 cells with stavudine loaded liposomes and galactosylated liposomes. Scintigraphic imaging and quantitative biodistribution of 99mTc labeled drug and liposomes showed that liposomal formulations were better taken up by the liver and spleen. Free drug solution was cleared from the blood. Further, a significantly higher (P<0.05) liver and spleen retention was observed over a period of 24h in case of galactosylated liposomes as compared to free drug and plain liposomes. Reduced uptake of the galactosylated liposomes in bone and higher and prolonged accumulation in mononuclear phagocyte system (MPS)-rich organs indicates the excellent potential of this formulation in the treatment of HIV infection.

Intranasal Cabergoline: Pharmacokinetic and Pharmacodynamic Studies

Aims of this investigation were to prepare and characterize cabergoline intranasal microemulsion formulations, determine brain drug delivery through biodistribution using technetium-99m (99mTc) as a tracer, and assess its performance pharmacodynamically in weight control. Cabergoline microemulsions of different compositions were prepared by water titration method and characterized for globule size and zeta potential. Microemulsion with maximum drug solubilization and stability was considered optimal and taken for further studies with or without addition of mucoadhesive agent. Pharmacokinetics of optimized 99mTc-labeled cabergoline formulations and 99mTc-labeled drug solution were studied by estimating radioactivity in brain and blood of albino rats post intranasal, intravenous, and oral administrations. To confirm localization of drug in brain following intranasal, intravenous, and oral administrations, gamma scintigraphy imaging was also performed. To assess weight control performance of formulations, body weight, white adipose tissue mass, serum lipids, leptin, and prolactin were determined before and after 40xa0days of intranasal administrations of these formulations to Wistar rats. Microemulsions were found to be stable both physically and chemically when stored at various stress conditions. Brain/blood uptake ratios, drug targeting efficiency, and direct drug transport were found to be highest for drug mucoadhesive microemulsion followed by drug microemulsion and drug solution post-intranasal administration compared to intravenous drug microemulsion. Significant (pu2009<u20090.05) reduction in assessed pharmacodynamic parameters was observed after intranasal administration of mucoadhesive microemulsion against control group. The results of the studies conclusively demonstrate that intranasal microemulsion formulations developed in this investigation are stable and can deliver cabergoline selectively and in higher amounts to the brain compared to both drug administrations as a solution intranasally or microemulsion intravenously. The results also demonstrate reduction in weight, adipose tissue mass, serum lipids, and serum prolactin after intranasal administration of drug microemulsion. Hence, long-term studies in at least two more animal models followed by extensive clinical evaluation can safely result into a product for clinical use.

Pharmacokinetics and biodistribution of samarium-153-labelled OC125 antibody coupled to CITCDTPA in a xenograft model of ovarian cancer

The use of samarium-153 in the context of radioimmunotherapy of cancers has been limited by the instability of antibody labelling, which produces high uptake concentrations in liver and bone. This study compares the pharmacokinetics and biodistribution of153Sm-labelled OC125 monoclonal antibody, in whole or F(ab′)2 fragment form and with diethylene triamine penta-acetic acid (DTPA) or 6-p-isothiocyanatobenzyl diethylene triamine penta-acetic acid (CITCDTPA) coupling, in nude mice grafted subcutaneously with an ovarian adenocarcinoma line (SHIN-3) expressing CA125 antigen. The specific activity of the immunoconjugates was 18.5–55.5 MBq/mg, and their immunoreactivity exceeded 65%. With153Sm-DTPA-OC125F(ab′)2, the stability study in serum indicated that 50% of the metal remained bound to the antibody. The pharmacokinetic study showed a retention half-life of 25.1 h and blood clearance of 0.72 ml/h. The biodistribution study indicated tumour uptake of 4.53%±9% of injected activity per gram (%ID/g) at 24 h and tumour-to-liver and tumour-to-bone ratios of 0.23±0.02 and 1.54±0.49 respectively at 24 h. With153Sm-CITCDTPA-OC125F(ab′)2, serum stability was greater (87% of the metal remaining bound to the antibody), retention half-life was 22.25 h and blood clearance was 2.23 ml/h. Tumour was better targeted (8.30%±3.56%ID/g at 24 h), and tumour-to-liver and tumour-to-bone ratios were 1.17±0.36 and 7.08±3.09 respectively at 24 h. However, renal retention remained elevated (29.76%±9.41%ID/g at 24 h). With intact IgG, renal uptake decreased (1.41%±0.49%ID/g at 24 h), but tumour uptake was lower than with fragments (1.46%±0.58%ID/g at 24 h). Liver uptake was higher (tumour-to-liver ratio 0.10±0.05), and blood clearance was slower. The stability and distribution of153Sm-CITCDTPA were more favourable than those of153Sm-DTPA for application in radioimmunotherapy. Quantitative analysis performed using digitized images obtained by conventional autoradiography and the imaging plate system indicated that the latter system is suitable for bio-distribution studies of immunoconjugates.

In Vitro Assessment of Acyclovir Permeation Across Cell Monolayers in the Presence of Absorption Enhancers

The aim of the investigation was to establish transepithelial permeation of acyclovir across Caco-2 and Madin-Darby canine kidney (MDCK) cell monolayers and attempt to improve its permeation by employing absorption enhancers (dimethyl β cyclodextrin, chitosan hydrochloride and sodium lauryl sulfate) and combinations thereof. Caco-2 and MDCK cell monolayers have been widely employed in studying drug transport, mechanisms of drug transport, and screening of absorption enhancers and excipients. Transepithelial electrical resistance and permeation of 99mTc-mannitol were employed as control parameters to assess the tight junction and paracellular integrity. Permeation of acyclovir in the presence of absorption enhancers was found to be significantly higher compared with drug permeation in their absence when assessed as apparent permeability coefficients (Papp). Synergistic improvements in Papp values of acyclovir were obtained in case-selected combinations of absorption enhancers; dimethyl β cyclodextrin–chitosan hydrochloride, chitosan hydrochloride–sodium lauryl sulfate, and dimethyl β cyclodextrin–sodium lauryl sulfate, were used. Recovery and viability assessment studies of both cell monolayers suggested reestablishment of paracellular integrity and no damage to cell membranes. Significantly improved permeation of acyclovir in the presence of selected combinations of absorption enhancers may be used as a viable approach in overcoming the problem of limited oral bioavailability of acyclovir.

In vivo evaluation of alginate microspheres of carvedilol for nasal delivery.

Mucoadhesive alginate microspheres of carvedilol (CRV) for nasal administration intended to avoid first pass metabolism and to improve bioavailability were prepared and evaluated. The microspheres were prepared by emulsification cross-linking method. Radiolabeling of CRV and its microspheres was performed by direct labeling with reduced technetium-99m ((99m) Tc). In vivo studies were performed on New Zealand white rabbits by administering the microspheres intranasally using monodose nasal insufflator. The radioactivity was measured in a well-type gamma scintillation counter. The noncompartmental pharmacokinetic analysis was performed. The pattern of deposition and clearance of the microspheres were evaluated using a radioactive tracer and the noninvasive technique of gamma scintigraphy. The clearance of alginate microsphere was compared with that of control lactose. The microspheres were nonaggregated, free flowing powders with spherical shape, and smooth surface. Pharmacokinetics study displayed an increase in area under the curve and hence in relative bioavailability when compared with intravenous administration of drug. The nasal bioavailability was 67.87% which indicates that nasal administration results in improved absorption of CRV. The results of gamma scintigraphy showed that the alginate microspheres had significantly reduced rates of clearance from the rabbit nasal cavity when compared with the control lactose.

Rhenium-188-labeled anti-neural cell adhesion molecule antibodies with 2-iminothiolane modification for targeting small-cell lung cancer

We have evaluated the potential of188Re-labeled monoclonal antibodies (MAbs) modified with 2-iminothiolane (2IT) for targeting small-cell lung cancer (SCLC). Radiolabeled MAbs NK1NBL1 and C218 recognizing neural cell adhesion molecule were injected i.v. into athymic mice inoculated with human SCLC tumors, and the biodistribution was examined. NK1NBL1 localized in the tumors better than C218.188Re-labeled MAbs cleared from the blood faster than125I-labeled counterparts, resulting in higher tumor-to-blood ratios. In conclusion, the188Re-labeled MAbs are attractive candidates for imaging and therapy of SCLC.