Synthesis and Preliminary Evaluation of an ASGPr-Targeted Polycationic -Cyclodextrin Carrier for Nucleosides and Nucleotides.

Most antiviral and anticancer nucleosides are prodrugs that require stepwise phosphorylation to their triphosphate nucleotide form for biological activity. Monophosphorylation may be rate-limiting, and the nucleotides may be unstable and poorly internalized by target cells. Effective targeting and delivery systems for nucleoside drugs, including oligonucleotides used in molecular therapeutics, could augment their efficacy.

Influence of 2-Nitroimidazoles in the Response of FaDu Cells to Ionizing Radiation and Hypoxia/Reoxygenation Stress.

Cellular adaptations to hypoxia promote resistance to ionizing radiation (IR). This presents a challenge for treatment of head and neck cancer (HNC) that relies heavily on radiotherapy. Standard radiosensitizers often fail to reach diffusion-restricted hypoxic cells, whereas nitroimidazoles (NIs) [such as iodoazomycin arabinofuranoside (IAZA) and fluoroazomycin arabinofuranoside (FAZA)] can preferentially accumulate in hypoxic tumours.

Radiation Dosimetry of Theragnostic Pairs for Isotopes of Iodine in IAZA.

Theragnostic pairs of isotopes are used to infer radiation dosimetry for a therapeutic radiopharmaceutical from a diagnostic imaging study with the same tracer molecule labelled with an isotope better suited for the imaging task. We describe the transfer of radiation dosimetry from the diagnostic radioiodine isotope I, labelled for the hypoxia tracer molecule iodoazomycin arabinoside ([I]IAZA), to isotopes I (therapeutic) and I (PET imaging). Uncertainties introduced by the dissimilar isotope half-lives are discussed in detail.

Synthesis of [F]F-γ-T-3, a Redox-Silent γ-Tocotrienol (γ-T-3) Vitamin E Analogue for Image-Based In Vivo Studies of Vitamin E Biodistribution and Dynamics.

Vitamin E, a natural antioxidant, is of interest to scientists, health care pundits and faddists; its nutritional and biomedical attributes may be validated, anecdotal or fantasy. Vitamin E is a mixture of tocopherols (TPs) and tocotrienols (T-3s), each class having four substitutional isomers (α-, β-, γ-, δ-). Vitamin E analogues attain only low concentrations in most tissues, necessitating exacting invasive techniques for analytical research.

A Simple Computational Tool for Accurate, Quantitative Prediction of One-Electron Reduction Potentials of Hypoxia-Activated Tirapazamine Analogues.

The reduction potentials of bioreductively-activated drugs represent an important design parameter to be accommodated in the course of creating lead compounds and improving the efficacy of older generation drugs.  Reduction potentials are traditionally reported as single-electron reduction potentials, E(1), measured against reference electrodes under strictly defined experimental conditions.  More recently, computational chemists have described redox properties in terms of a molecule's highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO), in electron volts (eV).

Putative electron-affinic radiosensitizers and markers of hypoxic tissue: Synthesis and preliminary in vitro biological characterization of C3-amino-substituted benzotriazine dioxides (BTDOs).

Introduction: The redox characteristics of 1,2,4-benzotriazine-1,4-dioxides (BTDOs) make them potential radiosensitizing agents for hypoxic cells in solid human cancers. Tirapazamine (TPZ) is the most clinically tested BTDO radiosensitizer, despite its toxicity at effective doses. To date, no BTDOs have been developed as diagnostic markers of tissue hypoxia.

Development of [I]I-EOE-TPZ and [I]I-EOE-TPZMO: Novel Tirapazamine (TPZ)-Based Radioiodinated Pharmaceuticals for Application in Theranostic Management of Hypoxia.

: Benzotriazine-1,4-dioxides (BTDOs) such as tirapazamine (TPZ) and its derivatives act as radiosensitizers of hypoxic tissues. The benzotriazine-1-monoxide (BTMO) metabolite (SR 4317, TPZMO) of TPZ also has radiosensitizing properties, and via unknown mechanisms, is a potent enhancer of the radiosensitizing effects of TPZ. Unlike their 2-nitroimidazole radiosensitizer counterparts, radiolabeled benzotriazine oxides have not been used as radiopharmaceuticals for diagnostic imaging or molecular radiotherapy (MRT) of hypoxia.

Synthesis of [F]FAZA Using Nosyl and Iodo Precursors for Nucleophilic Radiofluorination.

Background: 1-α-D-(5-Deoxy-5-[18F]fluoroarabinofuranosyl)-2-nitroimidazole ([18F]FAZA) is manufactured by nucleophilic radiofluorination of 1-α-D-(2',3'-di-O-acetyl-5'-O-toluenesulfonylarabinofuranosyl)- 2-nitroimidazole (DiAcTosAZA) and alkaline deprotection to afford [18F]FAZA. High yields (>60%) under optimized conditions frequently revert to low yields (<20%) in large scale, automated syntheses. Competing side reactions and concomitant complex reaction mixtures contribute to substantial loss of product during HPLC clean-up.

Positron Emission Tomography (PET) and Pharmacokinetics: Classical Blood Sampling Versus Image-Derived Analysis of [18F]FAZA and [18F]FDG in a Murine Tumor Bearing Model.

Purpose: Pharmacokinetic (PK) data are generally derived from blood samples withdrawn serially over a defined period after dosing. In small animals, blood sampling after dosing presents technical difficulties, particularly when short time intervals and frequent sampling are required. Positron emission tomography (PET) is a non-invasive functional imaging technique that can provide semi-quantitative temporal data for defined volume regions of interest (vROI), to support kinetic analyses in blood and other tissues.

Pharmacokinetics and Scintigraphic Imaging of the Hypoxia-Imaging Agent [I]IAZA in Healthy Adults Following Exercise-Based Cardiac Stress †.

The objective of this work is to evaluate the potential effect of cardiac stress exercise on the accumulation of [I]IAZA, a radiopharmaceutical used to image focal tissue hypoxia, in otherwise normal myocardium in healthy volunteers, and to determine the impact of exercise on [I]IAZA pharmacokinetics. The underlying goal is to establish a rational basis and a baseline for studies of focal myocardial hypoxia in cardiac patients using [I]IAZA. Three healthy male volunteers ran the 'Bruce' treadmill protocol, a clinically-accepted protocol designed to expose myocardial ischemia in patients.

β -[F]Fluoro Azomycin Arabinoside (β -[F]FAZA): Synthesis, Radiofluorination and Preliminary PET Imaging of Murine A431 Tumors.

Background: 1-α-D-(5-Deoxy-5-[18F]fluoroarabinofuranosyl)-2-nitroimidazole([18F] FAZA) is a PET radiotracer that demonstrates excellent potential in imaging regional hypoxia, and is clinically used in diagnosing a wide range of solid tumors in cancer patients. [18F]FAZA, however, is radiofluorinated in only moderate recovered radiochemical yield (rRCY, ~12%). It is postulated that the relative stability of the C1' β-anomeric bond at C5' will make 1-β-D-(5-fluoro-5-deoxyarabinofuranosyl)-2-nitroimidazole (β-FAZA), the β-conformer of FAZA, an attractive candidate for clinical hypoxia imaging.

Synthesis and Biological Evaluation of Iodoglucoazomycin (I-GAZ), an Azomycin-Glucose Adduct with Putative Applications in Diagnostic Imaging and Radiotherapy of Hypoxic Tumors.

Iodoglucoazomycin (I-GAZ; N-(2-iodo-3-(6-O-glucosyl)propyl)-2-nitroimidazole), a non-glycosidic nitroimidazole-6-O-glucose adduct, was synthesized, radioiodinated, and evaluated as a substrate of glucose transporter 1 (GLUT1) for radiotheranostic (therapy+diagnostic) management of hypoxic tumors. Nucleophilic iodination of the nosylate synthon of I-GAZ followed by deprotection afforded I-GAZ in 74 % overall yield. I-GAZ was radioiodinated via 'exchange' labeling using [(123/131) I]iodide (50-70 % RCY) and then purified by Sep-Pak™ (>96 % RCP).

Detecting functional changes with [(18)F]FAZA in a renal cell carcinoma mouse model following sunitinib therapy.

Background: The multitargeting tyrosine kinase inhibitor (TKI) sunitinib is currently the first-line drug therapy for metastasizing renal cell carcinoma (RCC). TKIs have profound effects on tumor angiogenesis, leading to modifications of the tumor microenvironment. The goal of this study was to determine whether these treatment-induced changes can be detected with [(18)F]FAZA.

The chemistry and radiochemistry of hypoxia-specific, radiohalogenated nitroaromatic imaging probes.

Hypoxia is prevalent in many solid tumors. Hypoxic tumors tend to exhibit rapid growth and aberrant vasculature, which lead to oxygen (O2) depletion and impaired drug delivery. The reductive environment in hypoxic tumors alters cellular metabolism, which can trigger transcriptional responses; induce genetic alterations; promote invasion, metastasis, resistance to radiotherapy and chemotherapy, tumor progression, and recurrence; and leads to poor local control and reduced survival rates.

Microwave-assisted radiosynthesis of the hypoxia marker 1-α-D-(5- deoxy-5-[18F]fluoroarabinofuranosyl)-2-nitroimidazole ([18F]FAZA).

Unlabelled: The routine manufacture of most short-lived positron-emitting radiopharmaceuticals (PERs) involves conventional heating to accelerate the radiolabeling process. Nucleophilic radiofluorination reactions are generally slow at lower temperatures, and are accompanied by thermal decomposition of both precursor and product at higher temperatures. This necessitates HPLC purification and results in lower recovered radiochemical yields (rRCYs).

Pharmacokinetics of 3'-O-retinoyl-5-fluoro-2'-deoxyuridine (RFUdR), a dual acting mutually masking prodrug, and its metabolites in tumor bearing mice.

3'-O-Retinoyl-5-fluoro-2'-deoxyuridine (RFUdR) is a putative dual-acting, mutually-masking (DAMM) prodrug for the treatment of cancer. As part of the proof of principle for the DAMM concept, the concentrations of RFUdR and its post-hydrolysis active metabolites, 5-fluoro-2'-deoxyuridine (FUdR) and all-trans-retinoic acid (RA), were determined in plasma and selected tissues following either bolus intravenous (i.v.

[131I]IAZA as a molecular radiotherapeutic (MRT) drug: wash-out with cold IAZA accelerates clearance in a murine tumor model.

Based on animal model studies, [131I]IAZA may be useful as an adjunct radiotherapeutic (MRT) drug for the treatment of tumor hypoxia. However, radioactivity in the blood of patients and healthy volunteers dosed with [123I]IAZA has a protracted terminal elimination phase in which clearance is influenced by free [123I]IAZA and possibly by unidentified metabolites. The current work reports that about 40% of the radioactivity in human serum is associated with the serum protein fraction, and that the free:bound ratio is constant at about 60:40 for at least the first 135 min after injection, as determined by radio-HPLC analyses.

In vitro and in vivo evaluation of [(18)F]F-GAZ, a novel oxygen-mimetic azomycin-glucose conjugate, for imaging hypoxic tumor.

Several F-18-labeled 2-nitroimidazole (azomycin) derivatives have been proposed for imaging hypoxia using positron emission tomography (PET). Their cell penetration is based on passive diffusion, which limits their intracellular concentration maxima. The purpose of this study was to investigate the uptake of N-(2-[(18)F]fluoro-3-(6-O-glucosyl)propyl-azomycin ([(18)F]F-GAZ), a new azomycin-glucose conjugate, in vitro and in vivo.

Design, synthesis, and preliminary biological evaluation of 6-O-glucose-azomycin adducts for diagnosis and therapy of hypoxic tumors.

Several 2-nitroimidazole-based molecules (NIs) are used as clinical hypoxic tumor radiodiagnostics, but they are not effective as radiosensitizers/radiochemotherapeutics. These NIs permeate tumor cells nonselectively via diffusion, and in therapy, where high doses are required, their dose limiting toxicities preclude success. The synthesis and preliminary in vitro evaluations of three glucoazomycins, members of a novel class of C6-O-glucose-linked-azomycin conjugates that are putative substrates of glucose transport proteins (GLUTs) and possess hypoxia-selective radiosensitization features, are now reported.

Biochemistry and biology of 2'-Fluoro-2'-deoxythymidine (FT), a putative highly selective substrate for thymidine kinase type 2 (TK2).

2'-Deoxy-2'-fluorothymidine (FT) is a bioisostere of both thymidine (TdR), in which F replaces H at C-2' in the ribosyl configuration, and methyluridine, in which F replaces OH at C-2' in the ribosyl configuration. Fluorine is bioisosteric with H with respect to atomic radius and is bioisosteric with OH with respect to polarity and H-bonding as an H acceptor. The consequences of this C-2' F for H substitution on cytotoxicity, nucleoside transporter affinity, phosphorylation by thymidine kinases (TK1, TK2), cell uptake and biodistribution of FT in a murine tumor model are now reported.

Synthesis of hypoxia imaging agent 1-(5-deoxy-5-fluoro-α-D-arabinofuranosyl)-2-nitroimidazole using microfluidic technology.

Introduction: Microfluidic technology allows fast reactions in a simple experimental setup, while using very low volumes and amounts of starting material. Consequently, microfluidic technology is an ideal tool for radiolabeling reactions involving short-lived positron emitters. Optimization of the complex array of different reaction conditions requires knowledge of the different reaction parameters linked to the microfluidic system as well as their influence on the radiochemical yields.

Confocal laser scanning microscopy (CLSM) based evidence for cell permeation by mono-4-(N-6-deoxy-6-amino-β-cyclodextrin)-7-nitrobenzofuran (NBD-β-CyD).

Beta-cyclodextrin (β-CyD), amantadine and glucose were fluorescently tagged with 4-chloro-7-nitrobenz-2-oxa-1,3-diazole (NBD chloride) to afford NBD-β-CyD, NBD-amantadine and NBD-glucose, respectively. NBD-β-CyD/amantadine and β-CyD/NBD-amantadine inclusion complexes were prepared. Fluorescence emission maxima (λ(max) 544nm) and relative fluorescence intensities for NBD-β-CyD and NBD-β-CyD/amantadine were virtually identical, precluding the use of emission spectrum shifts for distinguishing free NBD-β-CyD from the complex.

Biodistribution and imaging of 1-(2-deoxy-beta-d-ribofuranosyl)-2,4-difluoro-5-[123/125I]iodobenzene (dRF[(123/125)I]IB), a nonpolar thymidine-mimetic nucleoside, in rats and tumor-bearing mice.

1-(2-Deoxy-beta-D-ribofuranosyl)-2,4-difluoro-5-iodobenzene (dRFIB) is a putative bioisostere of iododeoxyuridine (IUdR). The advantages of dRFIB over IUdR for in vivo studies include resistance to both phosphorolytic cleavage of the nucleoside bond and de-iodination. dRFIB was radioiodinated (dRF(123/125)IB) by copper-catalyzed exchange using commercial sodium [(123/125)I]iodide.

Synthesis, radiofluorination, and hypoxia-selective studies of FRAZ: A configurational and positional analogue of the clinical hypoxia marker, [18F]-FAZA.

The current work evaluates 1-alpha-d-(2-deoxy-2-fluororibofuranosyl)-2-nitroimidazole (FRAZ), a novel azomycin nucleoside that is a potential radiosensitizer of tumor hypoxia. FRAZ is a ribose analogue of 1-alpha-d-(2-deoxy-2-fluoroarabinofuranosyl)-2-nitroimidazole ([(18)F]-FAZA), a clinically used hypoxia marker. Preliminary assessment of the cytotoxicity and hypoxia-specific in vitro binding in HCT-110 colorectal cancer cells indicate that the radiosensitization properties of FRAZ are similar to that of FAZA, with a sensitizer enhancement ratio (SER) of approximately 1.