HIV treatments: a history of scientific advance.
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Anti-HIV Agents↗
Biomedical subjects
R A Smith
Publications and source records attributed to R A Smith.
HIV prevention for the third decade of AIDS. The need for a renewed commitment.
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HIV Infections↗
Early lung cancer detection: current and ongoing challenges.
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Humans↗
Selective targeting of a redox-active ubiquinone to mitochondria within cells: antioxidant and antiapoptotic properties.
With the recognition of the central role of mitochondria in apoptosis, there is a need to develop specific tools to manipulate mitochondrial function within cells. Here we report on the development of a novel antioxidant that selectively blocks mitochondrial oxidative damage, enabling the roles of mitochondrial oxidative stress in different types of cell death to be inferred. This antioxidant, named mitoQ, is a ubiquinone derivative targeted to mitochondria by covalent attachment to a lipophilic triphenylphosphonium cation through an aliphatic carbon chain. Due to the large mitochondrial membrane potential, the cation was accumulated within mitochondria inside cells, where the ubiquinone moiety inserted into the lipid bilayer and was reduced by the respiratory chain. The ubiquinol derivative thus formed was an effective antioxidant that prevented lipid peroxidation and protected mitochondria from oxidative damage. After detoxifying a reactive oxygen species, the ubiquinol moiety was regenerated by the respiratory chain enabling its antioxidant activity to be recycled. In cell culture studies, the mitochondrially localized antioxidant protected mammalian cells from hydrogen peroxide-induced apoptosis but not from apoptosis induced by staurosporine or tumor necrosis factor-alpha. This was compared with untargeted ubiquinone analogs, which were ineffective in preventing apoptosis. These results suggest that mitochondrial oxidative stress may be a critical step in apoptosis induced by hydrogen peroxide but not for apoptosis induced by staurosporine or tumor necrosis factor-alpha. We have shown that selectively manipulating mitochondrial antioxidant status with targeted and recyclable antioxidants is a feasible approach to investigate the role of mitochondrial oxidative damage in apoptotic cell death. This approach will have further applications in investigating mitochondrial dysfunction in a range of experimental models.
Animals↗
Discovery of a new class of p38 kinase inhibitors.
The MAP kinase p38 has been implicated in cytokine signaling, and its inhibitors are potentially useful for the treatment of arthritis and osteoporosis. Novel small-molecule inhibitors of p38 kinase were derived from a combinatorial chemistry effort and exhibit activity in the nanomolar range. Very steep structure-activity relationships are observed within this class.
Antirheumatic Agents↗
31P and 1H NMR studies of the effect of the counteracting osmolyte trimethylamine-N-oxide on interactions of urea with ribonuclease A.
31P NMR spectroscopy has been used to show that the activity of RNase A, which is lowered in the presence of urea, can be recovered with trimethylamine-N-oxide (TMAO). A 1:1 ratio of TMAO:urea was sufficient to recover the enzyme activity. (1)H nuclear Overhauser effect spectroscopy NMR studies with RNase A have shown that even at relatively low effective concentrations of TMAO, some modification of the three-dimensional structure of the biomolecule is apparent.
Animals↗
Mitochondrially targeted antioxidants and thiol reagents.
Mitochondrial oxidative damage and dysfunction contributes to a number of cell pathologies. To investigate how this damage affects cell function we have developed mitochondrially targeted antioxidants and thiol reagents by covalently linking them to lipophilic cations. The cation drives the selective accumulation of these reagents into mitochondria within cells where the antioxidants decrease oxidative damage and the thiol reagents enable measurement of the redox status of thiol proteins. In conjunction with cell and animal models of apoptosis, oxidative damage, and nitric oxide signaling, these molecules may provide new insights into the roles of mitochondria in human pathologies.
Animals↗
Structure-directed discovery of potent non-peptidic inhibitors of human urokinase that access a novel binding subsite.
BACKGROUND: Human urokinase-type plasminogen activator has been implicated in the regulation and control of basement membrane and interstitial protein degradation. Because of its role in tissue remodeling, urokinase is a central player in the disease progression of cancer, making it an attractive target for design of an anticancer clinical agent: Few urokinase inhibitors have been described, which suggests that discovery of such a compound is in the early stages. Towards integrating structural data into this process, a new human urokinase crystal form amenable to structure-based drug design has been used to discover potent urokinase inhibitors. RESULTS: On the basis of crystallographic data, 2-naphthamidine was chosen as the lead scaffold for structure-directed optimization. This co-crystal structure shows the compound binding at the primary specificity pocket of the trypsin-like protease and at a novel binding subsite that is accessible from the 8-position of 2-napthamidine. This novel subsite was characterized and used to design two compounds with very different 8-substituents that inhibit urokinase with K(i) values of 30-40 nM. CONCLUSIONS: Utilization of a novel subsite yielded two potent urokinase inhibitors even though this site has not been widely used in inhibitor optimization with other trypsin-like proteases, such as those reported for thrombin or factor Xa. The extensive binding pockets present at the substrate-binding groove of these other proteins are blocked by unique insertion loops in urokinase, thus necessitating the utilization of additional binding subsites. Successful implementation of this strategy and characterization of the novel site provides a significant step towards the discovery of an anticancer clinical agent.
Binding Sites↗
Drug delivery to mitochondria: the key to mitochondrial medicine.
The major function of mitochondria in human cells is to provide ATP by oxidative phosphorylation. However, mitochondria have many other roles including the modulation of intracellular calcium concentration and the regulation of apoptotic cell death. Furthermore, the mitochondrial respiratory chain is a major source of damaging free radicals. Consequently, mitochondrial dysfunction contributes to a number of human diseases, ranging from neurodegenerative diseases and ischaemia-reperfusion injury to obesity and diabetes. In addition, mutations to nuclear or mitochondrial DNA cause a number of human diseases. Therefore, strategies to prevent mitochondrial damage or to manipulate mitochondrial function in clinically useful ways may provide new therapies for a range of human disorders. Here we outline why mitochondria are a potentially important target for drug delivery and discuss how to deliver bioactive molecules selectively to mitochondria within cells.
Animals↗
Detecting and reporting medical errors: why the dilemma?
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Communication↗
Macrophage cytokine response to particles and lipopolysaccharide in vitro.
Several investigators have suggested that biologic molecules adsorbed onto particles may play a key role in determining macrophage response. Adsorbed endotoxins (bacterial debris) may be of particular importance since they are widely present exogenously and endogenously and adhere strongly to many materials. Murine-transformed peritoneal macrophages (IC-21) were used in this in vitro study. Secretions of IL-1 beta, TNF alpha, and IL-6 were used as a measure of macrophage response to micron-range particles of high-density polyethylene and Co-Cr-Mo alloy, with and without adsorbed lipopolysaccharide (LPS) endotoxin. Little cytokine secretion was measured in response to particles (and to polypropylene experimental chambers) cleaned with ethanol and saline and not exposed to LPS. The lack of macrophage response to cleaned particles has been reported by others and may help reconcile conflicting reports in the literature. Cytokine secretion levels were high in all cases if the chambers (with or without particles) were exposed to LPS (and rinsed to minimize nonbound LPS). Secretion patterns were different with particles present and for polymer versus metal particles. Overall, these results suggest that (1) adsorbed molecules on material surfaces strongly affect macrophage response and (2) particle surface chemistry and microstructure affect the concentration and configuration of adsorbed molecules, further influencing particle interaction with macrophage surface receptors.
Animals↗
A novel method for prediction of long-term outcome of women with T1a, T1b, and 10-14 mm invasive breast cancers: a prospective study.
BACKGROUND: Women with small mammographically detected breast cancers generally have good long-term outcomes, but a few with T1a (1-5 mm) and T1b (6-10 mm) tumours will eventually die from breast cancer. We investigated whether women at high risk of breast-cancer death could be identified with mammographic criteria and differentiated from women with small cancers of the breast and good outcomes. METHODS: We prospectively applied mammographic classifications of tumour type to a consecutive series of 343 mammograms of invasive breast cancers of size 1-14 mm. Classifications were: stellate (spiculated) mass with no calcifications; circular or oval lesions with no calcifications; spiculated or circular lesions with non-casting-type calcifications; and casting-type calcifications. FINDINGS: 20-year survival for women with 1-14 mm invasive tumours with casting-type calcifications was 55%. 14% of 138 women with 1-9 mm tumours had casting-type calcifications on mammography, which accounted for 73% of all breast-cancer deaths (p<0.001). T1a, T1b, and 10-14 mm tumours with casting-type calcifications behaved as if they were larger lesions, since the rate of death was similar to that for women with advanced high-grade tumours. Most women who died were node-negative. The long-term survival of women who had tumours of 1-9 mm with no casting-type calcifications was about 95%. INTERPRETATION: Mammographic classification seemed to reliably predict good and bad long-term outcomes for survival in tumours of 14 mm or smaller, and especially for those smaller than 10 mm. The implications for therapy are substantial.
Breast Neoplasms↗
Site-specific incorporation of nucleoside analogs by HIV-1 reverse transcriptase and the template grip mutant P157S. Template interactions influence substrate recognition at the polymerase active site.
Studies of drug-resistant reverse transcriptases (RTs) reveal the roles of specific structural elements and amino acids in polymerase function. To characterize better the effects of RT/template interactions on dNTP substrate recognition, we examined the sensitivity of human immunodeficiency virus type 1 (HIV-1) RT containing a new mutation in a "template grip" residue (P157S) to the 5'-triphosphates of (-)-beta-2',3'-dideoxy-3'-thiacytidine (3TC), (-)-beta-2',3'-dideoxy-5-fluoro-3'-thiacytidine (FTC), and 3'-azido-3'-deoxythymidine (AZT). A primer extension assay was used to monitor quantitatively drug monophosphate incorporation opposite each of multiple target sites. Wild-type and P157S RTs had similar catalytic activities and processivities on heteropolymeric RNA and DNA templates. When averaged over multiple template sites, P157S RT was 2-7-fold resistant to the 5'-triphosphates of 3TC, FTC, and AZT. Each drug triphosphate inhibited polymerization more efficiently on the DNA template compared with an RNA template of identical sequence. Moreover, chain termination by 3TC and FTC was strongly influenced by template sequence context. Incorporation of FTC and 3TC monophosphate varied up to 10-fold opposite 7 different G residues in the DNA template, and the P157S mutation altered this site specificity. In summary, these data identify Pro(157) as an important residue affecting nucleoside analog resistance and suggest that interactions between RT and the template strand influence dNTP substrate recognition at the RT active site. Our findings are discussed within the context of the HIV-1 RT structure.
Catalytic Domain↗
Linear estimates of disease progression predict survival in patients with amyotrophic lateral sclerosis.
We have shown that linear estimates of rates of disease progression (LEP), derived from isometric myometry [grip or foot dorsiflexion (FD) strength] and forced vital capacity (FVC%), are clinically and statistically significant predictors of survival of patients with amyotrophic lateral sclerosis (ALS) from date of disease onset and, except those based on grip strength, of survival from the date of measurement. We tested these results in 2 additional groups of patients: 1) those who participated in a previously reported Protropin (GH) study; and 2) those enrolled in two other clinical trials (group 2). The LEP were derived and tested as predictors of survival. In a Cox proportional hazards model, LEP based on all measures predicted survival from disease onset in both groups of patients. Using cutoff points determined within the original group to stratify patients in the validation groups into faster and slower progressing subgroups resulted in statistically significant separation of survival curves from disease onset in group 2 for all LEP and in group 1 (the GH group) for LEP derived from FD strength; and, for survival from date of measurement in group 2, when stratified by LEP based on FD strength or FVC%. LEP based on data generated by myometry or pulmonary function studies have now been shown to predict survival in 3 unrelated groups of patients with ALS entering clinical trials. Their precise use in clinical trial design needs to be explored further.
Adult↗
Epidemiology of lung cancer.
There are few relationships in the epidemiology of cancer between an exposure and disease that are as consistent as those observed between tobacco and lung cancer. The health consequences of tobacco use are not limited to lung cancer; the 1990 Surgeon General's Report described that the use of cigarettes was the leading cause of avoidable mortality in the United States, with about 434,000 preventable deaths per year. Although a majority of the adults in the United States are current or former smokers, smoking cessation and prevention efforts have been successful, although currently about one in four adults still smoke cigarettes. The decline in ever-smoking has reached a plateau in the past few years, however, and rates of teenage smoking have begun to increase. In 1997, smoking rates among high school students in the United States were 32% higher compared with 1991. As noted by Cinciripini et al, adult smokers who quit or die are being replaced by children who smoke. Until recently, lung cancer control efforts primarily have focused on smoking prevention in youth and cessation among adults, with little obvious potential for reducing deaths through early detection. With the recent publication of early results from the Early Lung Cancer Action Project showing remarkably more favorable screening performance compared with chest radiography, the potential to detect lung cancer early and save lives is being revisited. Ultimately, the preferred disease control strategy is the prevention of lung cancer through the elimination of tobacco use altogether, but for the foreseeable future a legacy of decades of tobacco use in a significant proportion of the US population means we are still challenged to develop public health strategies to reduce deaths and suffering from those destined to develop lung cancer.
Aged↗
The Swedish Two-County Trial twenty years later. Updated mortality results and new insights from long-term follow-up.
The benefit of invitation to mammographic screening observed in this trial is maintained as a highly significant 32% reduction in breast cancer mortality. Mammographic screening for breast cancer continues to save lives after up to 20 years. Screening derives this benefit by improving the distribution of tumors diagnosed with respect to prognostic categories based on node status, size, and histology of tumors. There is potential for modern screening programs with shorter interscreening intervals to achieve even greater improvements in prognostic category and greater reductions in breast cancer mortality. Mammography can discriminate a subpopulation of high-risk cases, those displaying casting-type calcifications on the mammogram, among very small tumors, with fundamental implications for diagnosis and treatment.
Adult↗
Adhesion of microvascular endothelial cells to metallic implant surfaces.
The objective of this study was to explore the molecular mechanisms of adhesion of endothelial cells (ECs) to implant grades of titanium alloy (Ti) and stainless steel (SS), compared to tissue culture polystyrene (PS). The idea is that promotion of EC adhesion to implant surfaces during the initial stages of healing may be critical in the formation of a capillary bed intimately associated with the implant surface. Ultimately this could be expected in turn to promote bone formation close to the surface and a more stable implant/bone interface. Surfaces were coated with either peak 1 fibrinogen gammaAgammaA, fibrinogen Fr I-9, fibrinogen fragment D1, fibronectin, vitronectin, or fetal calf serum and then post-coated with bovine serum albumin (BSA) to block non-specific cell adhesion. Surfaces with BSA alone and no other protein coating were also evaluated. Fibronectin coating maximized cell adhesion on all three surfaces, and adhesion was highest on PS. BSA blocked cell adhesion to PS (and most adhesion to SS) much better than to Ti. These results provide evidence that BSA adsorption on the metal surface is unable to effectively block the adhesion of the cells to the Ti. These data may provide a basis for understanding in vivo observations that soft tissue becomes attached to a Ti surface more rapidly and with more bone formation than to SS. Evidence is also presented that alphavbeta3 plays an important role in adhesion of ECs to the Ti surface. These experiments also provide preliminary data which may reflect some of the features of initial EC adhesion to metal implants.
Journal Article↗
Effects of the early diagnosis of amyotrophic lateral sclerosis on the patient: disadvantages.
While the early diagnosis of disease is generally desirable, this is not always so. There is reason to be concerned about the reliability of clinical diagnosis of amyotrophic lateral sclerosis (ALS), based on currently available methods, early in the disease process. Applying the standard of cost-effectiveness it is possible to estimate the effort required to extend the lifespan of persons who are detected through a screening programme. Until it is possible to alter the treatment outcome, patients have little to gain by the early detection of ALS.
Cost of Illness↗