64Cu-DOTATATE PET/CT for Imaging Patients with Known or Suspected Somatostatin Receptor-Positive Neuroendocrine Tumors: Results of the First U.S. Prospective, Reader-Masked Clinical Trial.

Studies demonstrate that the investigational 64Cu-DOTATATE radiopharmaceutical may provide diagnostic and logistical benefits over available imaging agents for patients with somatostatin receptor (SSTR)-positive neuroendocrine tumors (NETs). Accordingly, we aimed to prospectively determine the lowest dose of 64Cu-DOTATATE that facilitates diagnostic-quality scans and evaluated the diagnostic performance and safety in a phase III study of patients with SSTR-expressing NETs. Methods: A dose-ranging study was conducted on 12 patients divided into 3 dose groups (111 MBq [3.0 mCi], 148 MBq [4.0 mCi], and 185 MBq [5.0 mCi] ± 10%) to determine the lowest dose of 64Cu-DOTATATE that produced diagnostic-quality PET/CT images. Using the 64Cu-DOTATATE dose identified in the dose-ranging study, 3 independent nuclear medicine physicians who were masked to all clinical information read PET/CT scans from 21 healthy volunteers and 42 NET-positive patients to determine those with disease or no disease, as well as those with localized versus metastatic status. Masked-reader evaluations were compared with a patient-specific standard of truth, which was established by an independent oncologist who used all previously available pathology, clinical, and conventional imaging data. Diagnostic performance calculated for 64Cu-DOTATATE included sensitivity, specificity, negative predictive value, positive predictive value, and accuracy. Inter- and intrareader reliability, as well as ability to differentiate between localized and metastatic disease, was also determined. Adverse events were recorded from 64Cu-DOTATATE injection through 48 h after injection. Results: The dose-ranging study identified 148 MBq (4.0 mCi) as the optimal dose to obtain diagnostic-quality PET/CT images. After database lock, diagnostic performance from an initial majority read of the 3 independent readers showed a significant 90.9% sensitivity (P = 0.0042) and 96.6% specificity (P < 0.0001) for detecting NETs, which translated to a 100.0% sensitivity and 96.8% specificity after correcting for an initial standard-of-truth misread. Excellent inter- and intrareader reliability, as well as ability to distinguish between localized and metastatic disease, was also noted. No adverse events were related to 64Cu-DOTATATE, and no serious adverse events were observed. Conclusion:64Cu-DOTATATE PET/CT is a safe imaging technique that provides high-quality and accurate images at a dose of 148 MBq (4.0 mCi) for the detection of somatostatin-expressing NETs.

Evaluation of DOTA-chelated neurotensin analogs with spacer-enhanced biological performance for neurotensin-receptor-1-positive tumor targeting.

INTRODUCTION: Neurotensin receptor 1 (NTR1) is overexpressed in many cancer types. Neurotensin (NT), a 13 amino acid peptide, is the native ligand for NTR1 and exhibits high (nM) affinity to the receptor. Many laboratories have been investigating the development of diagnostic and therapeutic radiopharmaceuticals for NTR1-positive cancers based on the NT peptide. To improve the biological performance for targeting NTR1, we proposed NT analogs with a 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelation system and different lengths of spacers. METHODS: We synthesized four NTR1-targeted conjugates with spacer lengths from 0 to 9 atoms (null (N0), ß-Ala-OH (N1), 5-Ava-OH (N2), and 8-Aoc-OH (N3)) between the DOTA and the pharmacophore. In vitro competitive binding, internalization and efflux studies were performed on all four NT analogs. Based on these findings, metabolism studies were carried out on our best performing conjugate, (177)Lu-N1. Lastly, in vivo biodistribution and SPECT/CT imaging studies were performed using (177)Lu-N1 in an HT-29 xenograft mouse model. RESULTS: As shown in the competitive binding assays, the NT analogs with different spacers (N1, N2 and N3) exhibited lower IC50 values than the NT analog without a spacer (N0). Furthermore, N1 revealed higher retention in HT-29 cells with more rapid internalization and slower efflux than the other NT analogs. In vivo biodistribution and SPECT/CT imaging studies of (177)Lu-N1 demonstrated excellent accumulation (3.1 ± 0.4%ID/g) in the NTR1-positive tumors at 4h post-administration. CONCLUSIONS: The DOTA chelation system demonstrated some modest steric inhibition of the pharmacophore. However, the insertion of a 4-atom hydrocarbon spacer group restored optimal binding affinity of the analog. The in vivo assays indicated that (177)Lu-N1 could be used for imaging and radiotherapy of NTR1-positive tumors.

Innovative approach for urease inhibition by Ficus carica extract-fabricated silver nanoparticles: An in vitro study.

In the present study, a rapid, low-cost, and ecofriendly method of stable silver nanoparticles (AgNPs) synthesis using leaves extract of Ficus carica (F. carica), a plant with diverse metabolic consortium, is reported for the first time. An absorption peak at 422 nm in UV-Vis spectroscopy, a spherical shape with an average size of 21 nm in transmission electron microscopy, and crystalline nature in X-ray powder diffraction studies were observed for the synthesized AgNPs. Fourier transform infrared analysis indicated that proteins of F. carica might have a vital role in AgNP synthesis and stabilization. AgNPs were found to inhibit urease, a key enzyme responsible for the survival and pathogenesis of the bacterium, Helicobacter pylori. Inhibition of urease by AgNPs was monitored spectrophotometrically by the evaluation of ammonia release. The urease inhibition potential of AgNPs can be explored in the treatment of H. pylori by preparing novel combinations of standard drugs with AgNPs- or AgNPs-encapsulated drug molecules.

The influence of linker length on the properties of cathepsin S cleavable (177)Lu-labeled HPMA copolymers for pancreatic cancer imaging.

N-(2-hydroxypropyl)-methacrylamide (HPMA) copolymers have shown promise for application in the detection and staging of cancer. However, non-target accumulation, particularly in the liver and spleen, hinders the detection of resident or nearby metastatic lesions thereby decreasing diagnostic effectiveness. Our laboratory has pursued the development of cathepsin S susceptible linkers (CSLs) to reduce the non-target accumulation of diagnostic/radiotherapeutic HPMA copolymers. In this study, we ascertain if the length of the linking group impacts the cleavage and clearance kinetics, relative to each other and a non-cleavable control, due to a reduction in steric inhibition. Three different CSLs with linking groups of various lengths (0, 6 and 13 atoms) were conjugated to HPMA copolymers. In vitro cleavage studies revealed that the longest linking group (13 atoms) led to more rapid cleavage when challenged with cathepsin S. The CSL incorporated HPMA copolymers demonstrated significantly higher levels of excretion and a significant decrease in long-term hepatic and splenic retention relative to the non-cleavable control. Contrary to in vitro observations, the length of the linking group did not substantially impact the non-target in vivo clearance. In the case of HPAC tumor retention, the CSL with the null (0 atom) linker demonstrated significantly higher levels of retention relative to the other CSLs. Given these results, we find that the length of the linking group of the CSLs did not substantially impact non-target clearance, but did influence tumor retention. Overall, these results demonstrate that the CSLs can substantially improve the non-target clearance of HPMA copolymers thereby enhancing clinical potential.

Synthesis and in vitro and in vivo evaluation of hypoxia-enhanced 111In-bombesin conjugates for prostate cancer imaging.

UNLABELLED: Receptor-targeted agents, such as gastrin-releasing peptide receptor (BB2r)-targeted peptides, have been investigated extensively in preclinical and clinical studies. In an attempt to increase the effectiveness of diagnostic or radiotherapeutic agents, we have begun to explore the incorporation of the hypoxia-selective prodrug 2-nitroimidazole into receptor-targeted peptides. Hypoxia is a well-known characteristic of many solid tumors, including breast, prostate, and pancreatic cancers. The aim of this approach is to use the hypoxia-trapping capability of 2-nitroimidazoles to increase the retention of the agent in hypoxic, BB2r-positive tumors. We have demonstrated that incorporation of one or more 2-nitroimidazoles into the BB2r-targeted peptide significantly increases the in vitro retention of the agent in hypoxic prostate cancer cells. The study described herein represents our first investigation of the in vivo properties of these hypoxia-enhanced BB2r-targeted agents in a PC-3 xenograft mouse model. METHODS: Four (111)In-labeled BB2r-targeted conjugates--(111) IN-1, (111) IN-2, (111) IN-3, and (111) IN-4, composed of 2-nitroimidazole moieties of 0, 1, 2, and 3, respectively--were synthesized, labeled, and purified. The BB2r binding affinities, externalization, and protein-association properties of these radioconjugates were assessed using the BB2r-positive PC-3 human prostate cancer cell line under hypoxic and normoxic environments. The in vivo biodistribution and micro-SPECT/CT imaging of the (111) IN-1, (111) IN-2, and (111) IN-4 radioconjugates were investigated in PC-3 tumor-bearing severely combined immunodeficient mice. RESULTS: All conjugates and (nat)In-conjugates demonstrated nanomolar binding affinities. (111) IN-1, (111) IN-2, (111) IN-3, and (111) IN-4 demonstrated 41.4%, 60.7%, 69.1%, and 69.4% retention, correspondingly, of internalized radioactivity under hypoxic conditions relative to 34.8%, 35.3%, 33.2%, and 29.7% retention, respectively, under normoxic conditions. Protein-association studies showed significantly higher levels of association under hypoxic conditions for 2-nitroimidazole-containing BB2r-targeted radioconjugates than for controls. On the basis of the initial 1-h uptake in the PC-3 tumors, (111) IN-1, (111) IN-2, and (111) IN-4 demonstrated tumor retentions of 1.5%, 6.7%, and 21.0%, respectively, by 72 h after injection. Micro-SPECT/CT imaging studies of (111) IN-1, (111) IN-2, and (111) IN-4 radioconjugates resulted in clear delineation of the tumors. CONCLUSION: On the basis of the in vitro and in vivo studies, the BB2r-targeted agents that incorporated 2-nitroimidazole moieties demonstrated improved retention. These results indicate that further exploration into the potential of hypoxia-selective trapping agents for BB2r-targeted agents, as well as other targeted compounds, is warranted.

177Lu-labeled HPMA copolymers utilizing cathepsin B and S cleavable linkers: synthesis, characterization and preliminary in vivo investigation in a pancreatic cancer model.

INTRODUCTION: A major barrier to the advancement of therapeutic nanomedicines has been the non-target toxicity caused by the accumulation of the drug delivery systems in organs associated with the reticuloendothelial system, particularly the liver and spleen. Herein, we report the development of peptide based metabolically active linkers (MALs) that are enzymatically cleaved by cysteine cathepsin B and S, two proteases highly expressed in the liver and spleen. The overall goal of this approach is to utilize the MALs to lower the non-target retention and toxicity of radiolabeled drug delivery systems, thus resulting in higher diagnostic and radiotherapeutic efficacy. METHODS: In this study three MALs (MAL0, MAL1 and MAL2) were investigated. MAL1 and MAL2 are composed of known substrates of cathepsin B and S, respectively, while MAL0 is a non-cleavable control. Both MAL1 and MAL2 were shown to undergo enzymatic cleavage with the appropriate cathepsin protease. Subsequent to conjugation to the HPMA copolymer and radiolabeling with (177)Lu, the peptide-polymer conjugates were renamed (177)Lu-metabolically active copolymers ((177)Lu-MACs) with the corresponding designations: (177)Lu-MAC0, (177)Lu-MAC1 and (177)Lu-MAC2. RESULTS: In vivo evaluation of the (177)Lu-MACs was performed in an HPAC human pancreatic cancer xenograft mouse model. (177)Lu-MAC1 and (177)Lu-MAC2 demonstrated 3.1 and 2.1 fold lower liver retention, respectively, compared to control ((177)Lu-MAC0) at 72h post-injection. With regard to spleen retention, (177)Lu-MAC1 and (177)Lu-MAC2 each exhibited a nearly fourfold lower retention, relative to control, at the 72h time point. However, the tumor accumulation of the (177)Lu-MAC0 was two to three times greater than (177)Lu-MAC1 and (177)Lu-MAC2 at the same time point. The MAL approach demonstrated the capability of substantially reducing the non-target retention of the (177)Lu-labeled HPMA copolymers. CONCLUSIONS: While further studies are needed to optimize the pharmacokinetics of the (177)Lu-MACs design, the ability of the MAL to significantly decrease non-target retention establishes the potential this avenue of research may have for the improvement of diagnostic and radiotherapeutic drug delivery systems.

Cycloart-23-ene-3ß, 25-diol stimulates GLP-1 (7-36) amide secretion in streptozotocin-nicotinamide induced diabetic Sprague Dawley rats: a mechanistic approach.

In previous study, we have reported cycloart-23-ene-3ß, 25-diol is an active antidiabetic constituent isolated from stem bark of Pongamia pinnata (Linn.) Pierre. The objective of the present investigation was to evaluate cycloart-23-ene-3ß, 25-diol stimulates glucagon like peptide-1 (GLP-1) (7-36) amide secretion in streptozotocin-nicotinamide induced diabetic Sprague Dawley rats. Molecular docking studies were performed to elucidate the molecular basis for GLP-1 receptor agonistic activity. Type 2 diabetes was induced in overnight fasted Sprague Dawley rats pre-treated with nicotinamide (100mg/kg, i.p.) followed by administration of streptozotocin (55 mg/kg, i.p.) 20 min after. The rats were divided into following groups; I- non-diabetic, II- diabetic control, III- sitagliptin (5mg/kg, p.o.), IV- cycloart-23-ene-3ß, 25-diol (1mg/kg, p.o.). The cycloart-23-ene-3ß, 25-diol and sitagliptin treatment was 8 week. Plasma glucose was estimated every week (week 0 to week 8). Body weight, food and water intake were recorded daily. Glycosylated haemoglobin, lipid profile, plasma and colonic active (GLP-1) (7-36) amide, mRNA expression of proglucagnon GLP-1, plasma and pancreatic insulin, histology of pancreata as well as biomarkers of oxidative stress (superoxidase dismutase, reduced glutathione, malondialdehyde, glutathione peroxidase, glutathione S transferase) were measured after 8th week treatment. In acute study, active GLP-1 (7-36) amide release, plasma glucose and insulin were measured during oral glucose tolerance test. The docking data clearly indicated cycloart-23-ene-3ß, 25-diol bind to the GLP-1 receptor. It decreased plasma glucose level, increased plasma and pancreatic insulin level as well as increased plasma and colonic active GLP-1 (7-36) amide secretion in streptozotocin-nicotinamide induced diabetic Sprague Dawley rats.

Development of hypoxia enhanced 111In-labeled Bombesin conjugates: design, synthesis, and in vitro evaluation in PC-3 human prostate cancer.

The gastrin-releasing peptide receptor (BB2r) has shown great promise for tumor targeting due to the increase of the receptor expression in a variety of human cancers including prostate, breast, small-cell lung, and pancreatic cancer. From clinical investigations, prostate cancer has been shown to be among the most hypoxic of the cancers investigated. Many solid tumors contain regions of hypoxia due to poor organization and efficiency of the vasculature. However, hypoxia is typically not present in normal tissue. Nitroimidazoles, a thoroughly investigated class of hypoxia selective drugs, have been shown to be highly retained in hypoxic tissues. The purpose of this study is to determine if the incorporation of hypoxia trapping moieties into the structural paradigm of BB2r-targeted peptides will increase the retention time of the agents in prostate cancer tumors. The present work involves the design, syntheses, purification, and in vitro investigation of hypoxia enhanced (111)In-BB2r-targeted radioconjugates. A total of four BB2r-targeted conjugates (1-4) were synthesized and coupled with increasing numbers of 2-nitroimidazoles, a hypoxia trapping moiety. Conjugates were radiolabeled with (111)In and purified by HPLC prior to in vitro studies. Receptor saturation assays under both normoxic and hypoxic conditions showed that the BB2r receptor expression on the PC-3 human prostate cancer cell line was not significantly affected by oxygen levels. Competitive binding assays revealed that incorporation of 2-nitroimidazoles had a detrimental effect to BB2r binding when adequate spacer groups, between the hypoxia trapping agent and the pharmacophore, were not employed. All of the 2-nitroimidazole containing BB2r-targeted agents exhibited significantly higher longitudinal retention in PC-3 cells under hypoxic conditions compared to the analogous normoxic studies. Protein association analysis revealed a 3-fold increase in binding of a 2-nitroimidazole containing BB2r-targeted agent under hypoxic relative to normoxic conditions. The positive nature of these results indicate that further exploration into the potential of hypoxia selective trapping agents for BB2r-targeted agents, as well as other targeted compounds, is warranted.

Evaluation of anti-inflammatory and analgesic activity of a new class of biphenyl analogs in animal models of inflammation.

A new class of 4'-methylbiphenyl-2-(substituted phenyl)carboxamide derivatives had been previously evaluated in vivo for their anti-inflammatory activities in animal models of inflammation. In the present study, the most active compound of that series, compound 4e (4'-methylbiphenyl-2-(4-carboxy phenyl)carboxamide), was investigated in detail for its anti-inflammatory, analgesic and ulcerogenic potential. Pretreatment of rats with 4e (100 mg/kg) reduced carrageenan induced rat paw edema at 3 h compared to control group. Dose dependent percent inhibition of granuloma formation, exudate volume, total leukocyte count was observed in 4e (25, 50 and 100 mg/kg) and celecoxib (CAS 169590-42-5; 5 mg/kg) treated groups in the cotton pellet granuloma and granuloma pouch technique, respectively, in rats. C-reactive proteins were absent in the 4e treated group. Compound 4e inhibited acetic acid induced writhing dose dependently (10, 20 and 30 mg/kg). Compound 4e was inactive in the hot plate test. Gastric toxicity screening experiments showed that compound 4e, both after single and repeated oral administration, is devoid of any gastric irritation in rats. The LD50 was found to be more than 2000 mg/kg.

The influence of workplace environment on lung function of flour mill workers in Jalgaon urban center.

The workplace environment affects the health of workers. Unhygienic conditions are observed in the workplace environment of flour mills as fine organic flour dust gets airborne in the indoor environment of the flour mills. The present work was undertaken to study the health problems related to the workplace environment of flour mill workers. The results show that flour mill workers are receiving a heavy dose (average exposure concentration, 624 microg/m3) of flour dust. To determine the impact of flour dust on the lung function of the workers spirometric analysis was conducted. Significant declines in forced vital capacity (FVC), peak expiratory flow rate (PEFR) and forced expiratory volume in one second (FEV1) were observed in the flour mill workers as compared to expected values. This study reveals reduced lung efficiency of flour mill workers due to excessive exposure to fine organic dust prevalent in the workplace environment. The impairment in lung efficiency was increased with duration of exposure in the flour mill workers. The analysis of questionnaires used to generate information on self-reported problems reveals that most of the workers were suffering from asthma and respiratory problems. Furthermore, the data shows that 42% of the flour mill workers were having shortness of breath problems, 34% of workers were having frequent coughing, and 19% workers were having respiratory tract irritation. We recommend the compulsory use of personal protective equipment (nose mask) by flour mill workers during working hours. This would help to protect the workers health from the flour dust prevalent in the workplace environment. A regular periodic examination is necessary to measure the impact of particulate matter on the health of the flour mill workers.

3D-QSAR of histone deacetylase inhibitors: hydroxamate analogues.

The histone deacetylase enzyme has increasingly become an attractive target for developing novel anticancer drugs. Hydroxamates are a new class of anticancer agents reported to act by selective inhibition of the histone deacetylase (HDAC) enzyme. Comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were employed to study three-dimensional quantitative structure-activity relationships (3D-QSARs). QSAR models were derived from a training set of 40 molecules. An external test set consisting of 17 molecules was used to validate the CoMFA and CoMSIA models. All molecules were superimposed on the template structure by atom-based, multifit and the SYBYL QSAR rigid body field fit alignments. The statistical quality of the QSAR models was assessed using the parameters r(2)(conv), r(2)(cv) and r(2)(pred). In addition to steric and electronic fields, ClogP was also taken as descriptor to account for lipophilicity. The resulting models exhibited a good conventional r(2)(conv) and cross-validated r(2)(cv) values up to 0.910 and 0.502 for CoMFA and 0.987 and 0.534 for CoMSIA. Robust cross-validation by 2 groups was performed 25 times to eliminate chance correlation. The CoMFA models exhibited good external predictivity as compared to that of CoMSIA models. These 3D-QSAR models are very useful for design of novel HDAC inhibitors.

3D-QSAR of histone deacetylase inhibitors as anticancer agents by genetic function approximation.

Histone deacetylases (HDACs) play a critical role in gene transcription and are implicated in cancer therapy and other diseases. Inhibitors of HDACs induce cell differentiation and suppress cell proliferation in the tumor cells. Although many such inhibitors have been designed and synthesized, but selective inhibitors for HDAC isoforms are lacking. Various hydroxamic acid analogues have been reported as HDAC inhibitors. Here, we report a three-dimensional quantitative structure-activity relationship (3D-QSAR) study performed using genetic function approximation (GFA) for this class of molecules. QSAR models were generated using a training set of 39 molecules and the predictive ability of final model was assessed using a test set of 17 molecules. The internal consistency of the final QSAR model was 0.712 and showed good external predictivity of 0.585. The results of the present QSAR study indicated that molecular shape analysis (MSA). thermodynamic and structural descriptors are important for inhibition of HDACs.

Exposure to vehicular pollution and respiratory impairment of traffic policemen in Jalgaon City, India.

The ambient air quality monitoring was carried during the May 2003 to April 2004 along the (NH-6) passing through Jalgaon city. The average concentration of SOx 64 microg/m3, NOx 58 microg/ m3, particulates (> 10 micro) 515 microg/m3 and respirable dust particulates 224 microg/m3 was reported at Prabhat during the study period (May 2003-April 2004). This location represents the major highway crossings (four) in the study area. The present investigations are on the survey of health status and lung function of traffic policemen exposed to the inferior air quality as observed on the highway crossings. The spirometric analysis of traffic policemen shows significant variation in Peak Expiratory Flow Rate (PEFR), Forced Expiratory Volume in one second (FEV1) and Forced Vital Capacity (FVC). The parameters were significantly affected in the traffic policemen as against the control group of population. It reveals significant respiratory impairment in the traffic policemen due to exposure to vehicular pollution. The study suggest the compulsory use of personal protective equipment (nose mask) by the traffic policemen during duty hours will help for the protection from vehicular pollution. The regular periodic health checkup is required to understand the impact of vehicular pollution on the health of traffic policemen.

Ecofriendly approach of urban solid waste management: a case study of Jalgaon city, Maharashtra.

In India, the traditional methods are used for urban solid waste management. These practices are associated with degradation of the urban environment Hence, an environmentally sound garbage management system is required for the urban waste management. The concept of waste minimization needs to be adopted in such systems. It is possible to implement certain corrective measures at collection, storage, transport and disposal of urban solid waste to minimize the adverse impacts on the environment The present investigations are emphasized on generation, characterzation and ecofriendly disposal of solid waste generated in Jalgaon city, Maharashtra. In the present study the waste generation at source was studied in the various income groups of the city. The per capita waste generation was studied by sampling the waste directly from the source of generation. The study reveals that the waste generation is large in the high-income group followed by middle and low-income group public. Further the physicochemical characterization of the waste was also studied. The data generated will help to develop environmentally sound and economically feasible solid waste management system for Ihe city.