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Charles Wilson

Publications and source records attributed to Charles Wilson.

18 recordsLinked to original sources
PubMed2006

Building oligonucleotide therapeutics using non-natural chemistries.

Modified nucleotides are increasingly being utilized in all categories of therapeutic oligonucleotides to increase nuclease-resistance, target affinity and specificity. The extent to which these substitutions are tolerated varies with the different modes of action exploited by various modalities, but fully modified oligonucleotides have now been discovered for most types of therapeutic oligonucleotide. Fully phosphorothioate-substituted antisense oligonucleotides have been used for several years. The first fully modified siRNA was reported in 2006 with a 2'-O-methyl sense strand and a phosphorothioate antisense strand. The first fully modified aptamer (2'-O-methyl) was reported in 2005. It is expected that future candidate therapeutic oligonucleotides will have even more drug-like characteristics as a result of the inclusion of modified nucleotides.

Oligonucleotides↗
PubMed2006

Cross-calibration and minimum precision standards for dual-energy X-ray absorptiometry: the 2005 ISCD Official Positions.

The International Society for Clinical Densitometry (ISCD) Committee on Standards of Bone Measurement (CSBM) consists of experts in technical aspects of bone densitometry. The CSBM recently reviewed the scientific literature on cross-calibration and precision assessment. A report with recommendations was presented at the 2005 ISCD Position Development Conference (PDC). Based on a thorough review of the data by the ISCD Expert Panel during the conference, the ISCD adopted Official Positions with respect to (1) cross-calibration when changing or replacing hardware; (2) the approach to cross-calibration when an entire system is changed to one made by either the same or a different manufacturer; (3) when no cross-calibration study or bone mineral density (BMD) comparison is done between facilities; and (4) the minimum acceptable precision for an individual technologist. We present here the ISCD Official Positions on these topics that were established as a result of the 2005 PDC, together with the associated rationales and supportive evidence.

Absorptiometry, Photon↗
PubMed2006

2'-Deoxy purine, 2'-O-methyl pyrimidine (dRmY) aptamers as candidate therapeutics.

Aptamers are short oligonucleotides that fold into well-defined three-dimensional architectures thereby enabling specific binding to molecular targets such as proteins. To be successful as a novel therapeutic modality, it is important for aptamers to not only bind their targets with high specificity and affinity, but also to exhibit favorable properties with respect to in vivo stability, cost-effective synthesis, and tolerability (i.e., safety). We describe methods for generating aptamers comprising 2 - deoxy purines and 2 -O-methyl pyrimidines (dRmY) that broadly satisfy many of these additional constraints. Conditions under which dRmY transcripts can be efficiently synthesized using mutant T7 RNA polymerases have been identified and used to generate large libraries from which dRmY aptamers to multiple target proteins, including interleukin (IL)-23 and thrombin, have been successfully discovered using the SELEX process. dRmY aptamers are shown to be highly nuclease-resistant, long-lived in vivo, efficiently synthesized, and capable of binding protein targets in a manner that inhibits their biologic activity with K(D) values in the low nM range. We believe that dRmY aptamers have considerable potential as a new class of therapeutic aptamers.

Animals↗
PubMed2005

Direct in vitro selection of a 2'-O-methyl aptamer to VEGF.

Aptamers (protein binding oligonucleotides) have potential as a new class of targeted therapeutics. For applications requiring chronic systemic administration, aptamers must achieve high-affinity target binding while simultaneously retaining high in vivo stability, tolerability, and ease of chemical synthesis. To this end, we describe a method for generating aptamers composed entirely of 2'-O-methyl nucleotides (mRmY). We present conditions under which 2'-O-methyl transcripts can be generated directly and use these conditions to select a fully 2'-O-methyl aptamer from a library of 3 x 10(15) unique 2'-O-methyl transcripts. This aptamer, ARC245, is 23 nucleotides in length, binds to vascular endothelial growth factor (VEGF) with a Kd of 2 nM, and inhibits VEGF activity in cellular assays. Notably, ARC245 is so stable that degradation cannot be detected after 96 hr in plasma at 37 degrees C or after autoclaving at 125 degrees C. We believe ARC245 has considerable potential as an antiangiogenesis therapeutic.

Angiogenesis Inhibitors↗
PubMed2005

Conjugation to polyethylene glycol polymer promotes aptamer biodistribution to healthy and inflamed tissues.

Here, we examine biodistribution of radiolabeled aptamers and assess the relative ability of different stabilized aptamer compositions (mixed 2'-F/2'-O-Me; fully 2'-O-Me modified) to access inflamed tissues in a murine inflammation model. Biodistribution of 3H-labeled aptamers, including pegylated and unpegylated compositions, was assessed 3 hours postadministration using quantitative whole body autoradiography (QWBA). Aptamer penetration of cells in kidney and liver was also examined at a qualitative level by microautoradiography. To evaluate aptamer distribution to diseased tissues, inflammation was induced locally in animal hind limbs by treatment with carrageenan just prior to aptamer dosing. Aptamer compositions examined exhibited significant variation in distribution levels among organs and tissues. Highest concentrations of radioactivity in whole body tissues for all animals were observed in the kidney and urinary bladder contents. Relatively little radioactivity was associated with brain, spinal cord, and adipose tissue. Overall, the total level of radioactivity in whole body tissues was significantly higher for a 20-kDa PEG conjugate than for other aptamers. Comparatively high levels of the 20-kDa conjugate were seen in well-perfused organs and tissues, including liver, lungs, spleen, bone marrow, and myocardium. A fully 2'-O-Me composition aptamer had the lowest level of radioactivity in whole body tissues but distributed to higher concentrations in the gastrointestinal tract contents relative to other aptamers. Interestingly, the 20-kDa PEG-conjugated aptamer showed significantly higher levels of distribution to inflamed paw tissues than did either unconjugated or fully 2'-O-Me-modified aptamers.

Animals↗
PubMed2004

Conventional adjuvant chemotherapy versus single-cycle, autograft-supported, high-dose, late-intensification chemotherapy in high-risk breast cancer patients: a randomized trial.

BACKGROUND: Breast cancer patients with four or more positive axillary lymph nodes who are treated with conventional adjuvant therapy have a poor prognosis. In uncontrolled studies, high-dose chemotherapy produced much better results than conventional therapy. We compared the benefits of a single cycle of high-dose chemotherapy and the benefits of conventional chemotherapy in patients with high-risk breast cancer in a prospective, unblinded, randomized trial. METHODS: Between February 23, 1995, and June 29, 1999, 605 patients with breast cancer who had four or more positive lymph nodes were randomly assigned to treatment (307 to high-dose therapy and 298 to conventional therapy). The conventional chemotherapy regimen was four cycles of doxorubicin (75 mg/m2) followed by eight cycles of CMF (cyclophosphamide [600 mg/m2], methotrexate [50 mg/m2], and 5-fluorouracil [600 mg/m2]), all given intravenously on day 1 of a 21-day cycle. The high-dose regimen was four cycles of doxorubicin (75 mg/m2), followed by a single cycle of intermediate-dose cyclophosphamide (4000 mg/m2) supported by filgrastim (300 microg/day) for up to 10 days followed by high-dose cyclophosphamide (6000 mg/m2) and thiotepa (800 mg/m2). Peripheral blood progenitor cells were harvested by leukapheresis after treatment with cyclophosphamide and filgrastim and then re-infused after the high-dose cycle. Log-rank tests were used to compare survival rates. All statistical analyses were two-sided. RESULTS: At a median follow-up of 6 years, no statistically significant differences were detected between the arms in 5-year relapse-free survival (high-dose arm = 57%, 95% confidence interval [CI] = 51% to 63%; conventional-dose arm = 54%, 95% CI = 48% to 61% (P =.73) or in 5-year overall survival (high-dose arm = 62%, 95% CI = 56% to 68%; conventional-dose arm = 64%, 95% CI = 57% to 70%) (P =.38). CONCLUSION: Autograft-supported, high-dose therapy is not superior to conventional chemotherapy in patients with breast cancer who have multiple involved lymph nodes. This conclusion should be viewed in the context of improving the success of conventional chemotherapy.

Adult↗
PubMed2004

A novel strategy for selection of allosteric ribozymes yields RiboReporter sensors for caffeine and aspartame.

We have utilized in vitro selection technology to develop allosteric ribozyme sensors that are specific for the small molecule analytes caffeine or aspartame. Caffeine- or aspartame-responsive ribozymes were converted into fluorescence-based RiboReporter trade mark sensor systems that were able to detect caffeine or aspartame in solution over a concentration range from 0.5 to 5 mM. With read-times as short as 5 min, these caffeine- or aspartame-dependent ribozymes function as highly specific and facile molecular sensors. Interestingly, successful isolation of allosteric ribozymes for the analytes described here was enabled by a novel selection strategy that incorporated elements of both modular design and activity-based selection methods typically used for generation of catalytic nucleic acids.

Allosteric Regulation↗
PubMed2004

Recognition of planar and nonplanar ligands in the malachite green-RNA aptamer complex.

Ribonucleic acids are an attractive drug target owing to their central role in many pathological processes. Notwithstanding this potential, RNA has only rarely been successfully targeted with novel drugs. The difficulty of targeting RNA is at least in part due to the unusual mode of binding found in most small-molecule-RNA complexes: the ligand binding pocket of the RNA is largely unstructured in the absence of ligand and forms a defined structure only with the ligand acting as scaffold for folding. Moreover, electrostatic interactions between RNA and ligand can also induce significant changes in the ligand structure due to the polyanionic nature of the RNA. Aptamers are ideal model systems to study these kinds of interactions owing to their small size and the ease with which they can be evolved to recognize a large variety of different ligands. Here we present the solution structure of an RNA aptamer that binds triphenyl dyes in complex with malachite green and compare it with a previously determined crystal structure of a complex formed with tetramethylrosamine. The structures illustrate how the same RNA binding pocket can adapt to accommodate both planar and nonplanar ligands. Binding studies with single- and double-substitution mutant aptamers are used to correlate three-dimensional structure with complex stability. The two RNA-ligand complex structures allow a discussion of structural changes that have been observed in the ligand in the context of the overall complex structure. Base pairing and stacking interactions within the RNA fold the phosphate backbone into a structure that results in an asymmetric charge distribution within the binding pocket that forces the ligand to adapt through a redistribution of the positive partial charge.

Coloring Agents↗
PubMed2004

Pharmacokinetics and biodistribution of novel aptamer compositions.

PURPOSE: Aptamers are highly selective nucleic acid-based drugs that are currently being developed for numerous therapeutic indications. Here, we determine plasma pharmacokinetics and tissue distribution in rat of several novel aptamer compositions, including fully 2'-O-methylated oligonucleotides and conjugates bearing high-molecular weight polyethylene glycol (PEG) polymers, cell-permeating peptides, and cholesterol. METHODS: Levels of aptamer conjugates in biological samples were quantified radiometrically and by a hybridization-based dual probe capture assay with enzyme-linked fluorescent readout. Intact aptamer in urine was detected by capillary gel electrophoresis and matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF). RESULTS: Aptamer compositions examined exhibited a wide range of mean residence times in circulation (0.6-16 h) and significant variation in distribution levels among organs and tissues. Among the conjugates tested, in vivo properties of aptamers were altered most profoundly by conjugation with PEG groups. Complexation with a 20 kDa PEG polymer proved nearly as effective as a 40 kDa PEG polymer in preventing renal clearance of aptamers. Conjugation with 20 kDa PEG prolonged aptamer circulatory half-life, while reducing both the extent of aptamer distribution to the kidneys and the rate of urinary elimination. In contrast, the fully 2'-O-Me aptamer composition showed rapid clearance from circulation, and elimination with intact aptamer detectable in urine at 48 h post-administration. CONCLUSIONS: We find that conjugation and chemical composition can alter fundamental aspects of aptamer residence in circulation and distribution to tissues. Though the primary effect of PEGylation was on aptamer clearance, the prolonged systemic exposure afforded by presence of the 20 kDa moiety appeared to facilitate distribution of aptamer to tissues, particularly those of highly perfused organs.

Animals↗
PubMed2004

ADP-specific sensors enable universal assay of protein kinase activity.

Two molecular sensors that specifically recognize ADP in a background of over 100-fold molar excess of ATP are described. These sensors are nucleic-acid based and comprise a general method for monitoring protein kinase activity. The ADP-aptamer scintillation proximity assay is configured in a single-step, homogeneous format while the allosteric ribozyme (RiboReporter) sensor generates a fluorescent signal upon ADP-dependent ribozyme self-cleavage. Both systems perform well when configured for high-throughput screening and have been used to rediscover a known protein kinase inhibitor in a high-throughput screening format.

Adenosine Diphosphate↗
PubMed2003

Advances in understanding the process of epileptogenesis based on patient material: what can the patient tell us?

Many different types of epileptic seizures and epileptic syndromes exist. The process of epileptogenesis and the progressive nature of epilepsy, however, can most easily be investigated in the acquired epilepsies, in which a brain insult presumably gives rise to changes in neuronal systems that ultimately become capable of generating spontaneous ictal events. Invasive in vivo and in vitro research can be carried out in patients with acquired epileptogenic lesions in the course of epilepsy surgery; however, such studies are possible only for those epileptic conditions that can be treated surgically, and can be used only to examine an end stage of the epileptogenic process. Consequently, experimental animal models of human epileptic conditions are still required to study mechanisms by which specific cerebral insults initiate the epileptogenic process and the progression of an epileptic disturbance. Most current parallel human/animal invasive research has been focused on temporal lobe epilepsy, and particularly that form associated with hippocampal sclerosis, the most common human epileptogenic lesion. Studies indicate that epileptogenesis in this condition is initiated by specific types of cell loss and neuronal reorganization, which results not only in enhanced excitation, but also in enhanced inhibition, predisposing to hypersynchronization. Even within this single, well-studied epileptic disorder, evidence is found for more than one type of ictal onset, and individual seizures can demonstrate a transition from one ictal mechanism to another. Recent in vivo and in vitro parallel, reiterative investigations in patients with mesial temporal lobe epilepsy, and in rats with intrahippocampal kainate-induced hippocampal seizures, have revealed the presence of interictal epileptiform events, termed "fast ripples," which appear to be unique in tissue capable of generating spontaneous seizures. Pursuit of the fundamental mechanisms underlying these abnormalities should elucidate the neurobiologic basis of epileptogenicity in this disorder. Furthermore, if these events are markers for epileptogenicity, they may have clinical value for diagnosis and pharmacologic, as well as surgical, treatment. Further research is needed to determine if these observations are relevant to other types of epilepsies.

Animals↗
PubMed2003

Muscarinic and nicotinic cholinergic mechanisms in the mesostriatal dopamine systems.

The striatum and its dense dopaminergic innervation originating in the midbrain, primarily from the substantia nigra pars compacta and the ventral tegmental area, compose the mesostriatal dopamine (DA) systems. The nigrostriatal system is involved mainly in motor coordination and in disorders such as Tourette's syndrome, Huntington's disease, and Parkinson's disease. The dopaminergic projections from the ventral tegmental area to the striatum participate more in the processes that shape behaviors leading to reward, and addictive drugs act upon this mesolimbic system. The midbrain DA areas receive cholinergic innervation from the pedunculopontine tegmentum and the laterodorsal pontine tegmentum, whereas the striatum receives dense cholinergic innervation from local interneurons. The various neurons of the mesostriatal systems express multiple types of muscarinic and nicotinic acetylcholine receptors as well as DA receptors. Especially in the striatum, the dense mingling of dopaminergic and cholinergic constituents enables potent interactions. Evidence indicates that cholinergic and dopaminergic systems work together to produce the coordinated functioning of the striatum. Loss of that cooperative activity contributes to the dysfunction underlying Parkinson's disease.

Acetylcholine↗
PubMed2003

Endovascular treatment for dural arteriovenous fistulae of the superior petrosal sinus.

OBJECTIVE: To determine the usefulness of endovascular embolization for treatment of dural arteriovenous fistulae of the superior petrosal sinus. METHODS: We performed a retrospective review of 18 patients treated during a 16-year period. Transarterial and/or transvenous embolizations were performed as a preoperative adjunct or definitive therapy. Clinical follow-up status was supplemented by telephone interviews to determine Glasgow Outcome Scale scores. RESULTS: Fourteen patients (78%) were treated with a combination of endovascular therapy and open surgery, and 4 were treated by embolization alone (22%). Angiographic cure was achieved in all patients (100%). Thirty-day morbidity and mortality were 11 and 0%, respectively. The mean follow-up period was 5.4 years. At the latest follow-up examination, all patients had returned to independent clinical status (Glasgow Outcome Scale scores of 1 or 2). CONCLUSION: Endovascular treatment of dural arteriovenous fistulae of the superior petrosal sinus can result in cure when access to the site of the fistula can be achieved. Preoperative embolization is a safe and effective adjunct to minimize bleeding during open neurosurgery.

Adult↗
PubMed2002

Epileptogenesis after self-sustaining status epilepticus.

PURPOSE: To describe the natural history of chronic epilepsy after experimental self-sustaining status epilepticus (SSSE) and to correlate patterns of SSSE with ictal, interictal, and plastic changes that characterize chronic epilepsy. METHODS: SSSE was induced in adult Wistar rats by 30-min intermittent electrical stimulation of the perforant path. In some animals, SSSE was treated by short-term administration of antiepileptic drugs (AEDs). After SSSE, EEG and animal behavior were monitored for </=1 year. Some animals were killed to study mossy fiber sprouting in the dentate gyrus. RESULTS: Despite the high reproducibility of the electrographic and behavioral manifestations of SSSE, patterns of chronic epilepsy varied considerably among animals in terms of seizure frequency, initial seizure pattern at the onset of chronic epilepsy, and frequency of interictal spikes. Statistically significant correlations were found between spike frequency during SSSE and interictal spike frequency, as well as between the frequency of spontaneous seizures and degree of mossy fiber sprouting. Early treatment of SSSE prevented the occurrence of spontaneous seizures and significantly decreased frequency of interictal spikes. Late treatment of SSSE did not prevent spontaneous seizures, but significantly decreased their frequency, and eventually may lead to remission of epilepsy. CONCLUSIONS: SSSE leads after a "silent" period to chronic epilepsy, which is maintained for > or =1 year in the rat. The silence is only behavioral, because EEG paroxysmal activity is seen in every animal. In this model of SSSE, the timing of treatment is a major determinant of outcome. Early treatment reduces the incidence of chronic epilepsy, whereas late treatment only reduces its severity. The possibility that this reduction of the severity of epilepsy may led to spontaneous remissions merits further study.

Animals↗