Improving health behaviors in patients with peripheral arterial disease - A pilot study of supported self-management.

PURPOSE OF THE RESEARCH: This pilot study reports the feasibility of a future randomized controlled trial (RCT) investigating the effect of supported self-management through low-intensity psychological intervention in patients with peripheral arterial disease (PAD) resulting in claudication. The study protocol, measurement instrument, data collection, and analysis were evaluated. Clinical outcome measures include depression and anxiety scores, smoking cessation, activity (step count), weight, and quality of life. Both Quantitative and Qualitative data were collected to evaluate participant experience and the clinical impact of a supported self-management intervention delivered in a routine clinical setting. METHODS: Participants received an initial one to one assessment with a health psychologist. Demographic data and baseline clinical outcome measures were recorded. These included Hospital Anxiety and Depression Scale score (HADS), health-related quality of life questionnaire (EQ-5D-3 L), number of cigarettes smoked daily, weight/BMI, and daily step count. Participants each received an activity tracker to record daily step count and were followed up weekly to provide psychological input, including goal setting, overcoming barriers, and preventing relapse. Quantitative data collection was scheduled at baseline, 3 and 6 months (final follow-up). At the final follow-up, participants provided qualitative feedback reflecting upon their experience of the intervention and its impact. Descriptive statistical analysis and simple paired samples t-test were employed in data evaluation. RESULTS: The sample size was small (n = 30). Twenty-three participants were followed up to 6 months. Depression scores improved with statistical significance from baseline to 6-month follow-up. Eight participants stopped smoking (47% reduction); a further 9 greatly reduced their intake. A borderline statistically significant increase of daily step count was achieved between baseline and 6-month follow-up. In addition, positive weight loss trends were observed in a predominantly obese or overweight cohort. The qualitative feedback highlighted participants understood and embraced the information delivered regarding the importance of health behavior change. Participants were able to link tailored action plans with what mattered to them with the help of the Health Psychologist. CONCLUSION: This study embraces the contemporary ideology of enabling self-management of long-term conditions to improve clinical outcomes. As a pilot study, we have shown that an expanded, randomized controlled trial is both safe and feasible. A positive trend in clinical outcomes suggests this patient group may benefit from supported self-management through low-intensity psychological intervention, where other forms of early intervention have historically faltered.

Sex and race differences of cerebrospinal fluid metabolites in healthy individuals.

INTRODUCTION: Analyses of cerebrospinal fluid (CSF) metabolites in large, healthy samples have been limited and potential demographic moderators of brain metabolism are largely unknown. OBJECTIVE: Our objective in this study was to examine sex and race differences in 33 CSF metabolites within a sample of 129 healthy individuals (37 African American women, 29 white women, 38 African American men, and 25 white men). METHODS: CSF metabolites were measured with a targeted electrochemistry-based metabolomics platform. Sex and race differences were quantified with both univariate and multivariate analyses. Type I error was controlled for by using a Bonferroni adjustment (0.05/33 = .0015). RESULTS: Multivariate Canonical Variate Analysis (CVA) of the 33 metabolites showed correct classification of sex at an average rate of 80.6% and correct classification of race at an average rate of 88.4%. Univariate analyses revealed that men had significantly higher concentrations of cysteine (p < 0.0001), uric acid (p < 0.0001), and N-acetylserotonin (p = 0.049), while women had significantly higher concentrations of 5-hydroxyindoleacetic acid (5-HIAA) (p = 0.001). African American participants had significantly higher concentrations of 3-hydroxykynurenine (p = 0.018), while white participants had significantly higher concentrations of kynurenine (p < 0.0001), indoleacetic acid (p < 0.0001), xanthine (p = 0.001), alpha-tocopherol (p = 0.007), cysteine (p = 0.029), melatonin (p = 0.036), and 7-methylxanthine (p = 0.037). After the Bonferroni adjustment, the effects for cysteine, uric acid, and 5-HIAA were still significant from the analysis of sex differences and kynurenine and indoleacetic acid were still significant from the analysis of race differences. CONCLUSION: Several of the metabolites assayed in this study have been associated with mental health disorders and neurological diseases. Our data provide some novel information regarding normal variations by sex and race in CSF metabolite levels within the tryptophan, tyrosine and purine pathways, which may help to enhance our understanding of mechanisms underlying sex and race differences and potentially prove useful in the future treatment of disease.

HTRF analysis of soluble huntingtin in PHAROS PBMCs.

OBJECTIVE: We measured the levels of mutant huntingtin (mtHtt) and total huntingtin (tHtt) in blood leukocytes from Prospective Huntington At-Risk Observational Study (PHAROS) subjects at 50% risk of carrying the Huntington disease mutation using a homogeneous time-resolved fluorescence (HTRF) assay to assess its potential as a biomarker. METHODS: Peripheral blood mononuclear cells from consenting PHAROS subjects were analyzed by HTRF using antibodies that simultaneously measured mtHtt and tHtt. mtHtt levels were normalized to tHtt, double-stranded DNA, or protein and analyzed according to cytosine-adenine-guanine repeat length (CAGn), demographics, predicted time to clinical onset or known time since clinical onset, and available clinical measures. RESULTS: From 363 assayed samples, 342 met quality control standards. Levels of mtHtt and mt/tHtt were higher in 114 subjects with expanded CAG repeats (CAG ≥ 37) compared with 228 subjects with nonexpanded CAG repeats (CAG <37) (p < 0.0001). Analysis of relationships to predicted time to onset or to phenoconversion suggested that the HTRF signal could mark changes during the Huntington disease prodrome or after clinical onset. CONCLUSIONS: The HTRF assay can effectively measure mtHtt in multicenter sample sets and may be useful in trials of therapies targeting huntingtin.

Pharmacometabolomics of response to sertraline and to placebo in major depressive disorder - possible role for methoxyindole pathway.

Therapeutic response to selective serotonin (5-HT) reuptake inhibitors in Major Depressive Disorder (MDD) varies considerably among patients, and the onset of antidepressant therapeutic action is delayed until after 2 to 4 weeks of treatment. The objective of this study was to analyze changes within methoxyindole and kynurenine (KYN) branches of tryptophan pathway to determine whether differential regulation within these branches may contribute to mechanism of variation in response to treatment. Metabolomics approach was used to characterize early biochemical changes in tryptophan pathway and correlated biochemical changes with treatment outcome. Outpatients with MDD were randomly assigned to sertraline (n = 35) or placebo (n = 40) in a double-blind 4-week trial; response to treatment was measured using the 17-item Hamilton Rating Scale for Depression (HAMD17). Targeted electrochemistry based metabolomic platform (LCECA) was used to profile serum samples from MDD patients. The response rate was slightly higher for sertraline than for placebo (21/35 [60%] vs. 20/40 [50%], respectively, χ(2)(1)  = 0.75, p = 0.39). Patients showing a good response to sertraline had higher pretreatment levels of 5-methoxytryptamine (5-MTPM), greater reduction in 5-MTPM levels after treatment, an increase in 5-Methoxytryptophol (5-MTPOL) and Melatonin (MEL) levels, and decreases in the (KYN)/MEL and 3-Hydroxykynurenine (3-OHKY)/MEL ratios post-treatment compared to pretreatment. These changes were not seen in the patients showing poor response to sertraline. In the placebo group, more favorable treatment outcome was associated with increases in 5-MTPOL and MEL levels and significant decreases in the KYN/MEL and 3-OHKY/MEL; changes in 5-MTPM levels were not associated with the 4-week response. These results suggest that recovery from a depressed state due to treatment with drug or with placebo could be associated with preferential utilization of serotonin for production of melatonin and 5-MTPOL.

A novel method for detecting 7-methyl guanine reveals aberrant methylation levels in Huntington disease.

Guanine methylation is a ubiquitous process affecting DNA and various RNA species. N-7 guanine methylation (7-MG), although relatively less studied, could have a significant role in normal transcriptional regulation as well as in the onset and development of pathological conditions. The lack of a sensitive method to accurately quantify trace amounts of altered bases such as 7-MG has been a major deterrent in delineating its biological function(s). Here we report the development of methods to detect trace amounts of 7-MG in biological samples using electrochemical detection combined with high-performance liquid chromatography (HPLC) separation of compounds. We further sought to assess global alterations in DNA methylation in Huntington disease (HD), where transcriptional dysregulation is a major factor in pathogenesis. The developed method was used to study guanine methylation in cytoplasmic and nuclear nucleic acids from human and transgenic mouse HD brain and controls. Significant differences were observed in the guanine methylation levels in mouse and human samples, consistent with the known transcriptional pathology of HD. The sensitivity of the method makes it capable of detecting subtle aberrations. Identification of changes in methylation pattern will provide insights into the molecular mechanism changes that translate into onset and/or development of symptoms in diseases such as HD.

A high-throughput screen to identify inhibitors of SOD1 transcription.

Amyotrophic lateral sclerosis (ALS) is a fatal degenerative motor neuron disease. Approximately 20 percent of familial ALS cases are caused by mutations in the Cu/Zn superoxide dismutase (SOD1) gene. Rodents expressing mutant SOD1 transgenes develop progressive, fatal motor neuron disease and disease onset and progression is dependent on the level of SOD1. We investigated the possibility that a reduction in SOD1 protein may be of therapeutic benefit in ALS and screened 30,000 compounds for inhibition of SOD1 transcription. The most effective inhibitor identified was N-{4-[4-(4-methylbenzoyl)-1-piperazinyl]phenyl}-2-thiophenecarboxamide (Compound ID 7687685), which in PC12 cells showed an EC50 of 10.6 microM for inhibition of SOD1 expression and an LD50 more than 30 microM. This compound was subsequently shown to reduce endogenous SOD1 levels in HeLa cells and to exhibit a modest reduction of SOD1 protein levels in mouse spinal cord tissue. These data suggest that the efficacy of compound 7687685 as an inhibitor of SOD1 gene expression is not likely to be clinically useful, although the strategy reported could be applied broadly to screening for small molecule inhibitors of gene expression.

Reduced creatine kinase as a central and peripheral biomarker in Huntington's disease.

A major goal of current clinical research in Huntington's disease (HD) has been to identify preclinical and manifest disease biomarkers, as these may improve both diagnosis and the power for therapeutic trials. Although the underlying biochemical alterations and the mechanisms of neuronal degeneration remain unknown, energy metabolism defects in HD have been chronicled for many years. We report that the brain isoenzyme of creatine kinase (CK-BB), an enzyme important in buffering energy stores, was significantly reduced in presymptomatic and manifest disease in brain and blood buffy coat specimens in HD mice and HD patients. Brain CK-BB levels were significantly reduced in R6/2 mice by approximately 18% to approximately 68% from 21 to 91 days of age, while blood CK-BB levels were decreased by approximately 14% to approximately 44% during the same disease duration. Similar findings in CK-BB levels were observed in the 140 CAG mice from 4 to 12 months of age, but not at the earliest time point, 2 months of age. Consistent with the HD mice, there was a grade-dependent loss of brain CK-BB that worsened with disease severity in HD patients from approximately 28% to approximately 63%, as compared to non-diseased control patients. In addition, CK-BB blood buffy coat levels were significantly reduced in both premanifest and symptomatic HD patients by approximately 23% and approximately 39%, respectively. The correlation of CK-BB as a disease biomarker in both CNS and peripheral tissues from HD mice and HD patients may provide a powerful means to assess disease progression and to predict the potential magnitude of therapeutic benefit in this disorder.

Phase 2 study of sodium phenylbutyrate in ALS.

The objective of the study was to establish the safety and pharmacodynamics of escalating dosages of sodium phenylbutyrate (NaPB) in participants with ALS. Transcription dysregulation may play a role in the pathogenesis of ALS. Sodium phenylbutyrate, a histone deacetylase inhibitor, improves transcription and post-transcriptional pathways, promoting cell survival in a mouse model of motor neuron disease. Forty research participants at eight sites enrolled in an open-label study. Study medication was increased from 9 to 21 g/day. The primary outcome measure was tolerability. Secondary outcome measures included adverse events, blood histone acetylation levels, and NaPB blood levels at each dosage. Twenty-six participants completed the 20-week treatment phase. NaPB was safe and tolerable. No study deaths or clinically relevant laboratory changes occurred with NaPB treatment. Histone acetylation was decreased by approximately 50% in blood buffy-coat specimens at screening and was significantly increased after NaPB administration. Blood levels of NaPB and the primary metabolite, phenylacetate, increased with dosage. While the majority of subjects tolerated higher dosages of NaPB, the lowest dose (9 g/day), was therapeutically efficient in improving histone acetylation levels.

Therapeutic attenuation of mitochondrial dysfunction and oxidative stress in neurotoxin models of Parkinson's disease.

Parkinson's disease (PD) is a progressive neurodegenerative disorder for which there is no current therapy preventing cumulative neuronal loss. There is substantial evidence that mitochondrial dysfunction, oxidative stress, and associated caspase activity underlie the neurodegeneration observed. One potential drug therapy is the potent free radical scavenger and antioxidant cystamine, which has demonstrated significant clinical potential in models of neurodegenerative disorders and human neurological disease. This study examined the oral efficacy of cystamine in the MPTP and 6-hydroxydopamine neurotoxin models of PD. The neuroprotective effects of cystamine treatment significantly ameliorated nigral neuronal loss, preserved striatal dopaminergic projections, and improved striatal dopamine and metabolite levels, as compared to MPTP alone. Cystamine normalized striatal 8-hydroxy-2'-deoxyguanosine levels and ATP concentrations, consistent with reduced oxidative stress and improved mitochondrial function. Cystamine also protected against MPTP-induced mitochondrial loss, as identified by mitochondrial heat shock protein 70 and superoxide dismutase 2, with concomitant reductions in cytochrome c and caspase-3 activities. The neuroprotective value of cystamine was confirmed in the 6-hydroxydopamine model. Together these findings show cystamine's therapeutic benefit to reduce neuronal loss through attenuation of oxidative stress and mitochondrial dysfunction, providing the rationale for human clinical trials in PD patients.

Modulation of nucleosome dynamics in Huntington's disease.

Transcriptional dysregulation and aberrant chromatin remodeling are central features in the pathology of Huntington's disease (HD). In order to more fully characterize these pathogenic events, an assessment of histone profiles and associated gene changes were performed in transgenic N171-82Q (82Q) and R6/2 HD mice. Analyses revealed significant chromatin modification, resulting in reduced histone acetylation with concomitant increased histone methylation, consistent with findings observed in HD patients. While there are no known interventions that ameliorate or arrest HD progression, DNA/RNA-binding anthracyclines may provide significant therapeutic potential by correcting pathological nucleosome changes and realigning transcription. Two such anthracyclines, chromomycin and mithramycin, improved altered nucleosome homeostasis in HD mice, normalizing the chromatin pattern. There was a significant shift in the balance between methylation and acetylation in treated HD mice to that found in wild-type mice, resulting in greater acetylation of histone H3 at lysine 9 and promoting gene transcription. Gene expression profiling in anthracycline-treated HD mice showed molecular changes that correlate with disease correction, such that a subset of downregulated genes were upregulated with anthracycline treatment. Improved nucleosomal dynamics were concurrent with a significant improvement in the behavioral and neuropathological phenotype observed in HD mice. These data show the ability of anthracycline compounds to rebalance epigenetic histone modification and, as such, may provide the rationale for the design of human clinical trials in HD patients.

Dose ranging and efficacy study of high-dose coenzyme Q10 formulations in Huntington's disease mice.

There is substantial evidence that a bioenergetic defect may play a role in the pathogenesis of Huntington's Disease (HD). A potential therapy for remediating defective energy metabolism is the mitochondrial cofactor, coenzyme Q10 (CoQ10). We have reported that CoQ10 is neuroprotective in the R6/2 transgenic mouse model of HD. Based upon the encouraging results of the CARE-HD trial and recent evidence that high-dose CoQ10 slows the progressive functional decline in Parkinson's disease, we performed a dose ranging study administering high levels of CoQ10 from two commercial sources in R6/2 mice to determine enhanced efficacy. High dose CoQ10 significantly extended survival in R6/2 mice, the degree of which was dose- and source-dependent. CoQ10 resulted in a marked improvement in motor performance and grip strength, with a reduction in weight loss, brain atrophy, and huntingtin inclusions in treated R6/2 mice. Brain levels of CoQ10 and CoQ9 were significantly lower in R6/2 mice, in comparison to wild type littermate control mice. Oral administration of CoQ10 elevated CoQ10 plasma levels and significantly increased brain levels of CoQ9, CoQ10, and ATP in R6/2 mice, while reducing 8-hydroxy-2-deoxyguanosine concentrations, a marker of oxidative damage. We demonstrate that high-dose administration of CoQ10 exerts a greater therapeutic benefit in a dose dependent manner in R6/2 mice than previously reported and suggest that clinical trials using high dose CoQ10 in HD patients are warranted.

Therapeutic effects of cystamine in a murine model of Huntington's disease.

The precise cause of neuronal death in Huntington's disease (HD) is unknown. Proteolytic products of the huntingtin protein can contribute to toxic cellular aggregates that may be formed in part by tissue transglutaminase (Tgase). Tgase activity is increased in HD brain. Treatment in R6/2 transgenic HD mice, using the transglutaminase inhibitor cystamine, significantly extended survival, improved body weight and motor performance, and delayed the neuropathological sequela. Tgase activity and N(Sigma)-(gamma-L-glutamyl)-L-lysine (GGEL) levels were significantly altered in HD mice. Free GGEL, a specific biochemical marker of Tgase activity, was markedly elevated in the neocortex and caudate nucleus in HD patients. Both Tgase and GGEL immunoreactivities colocalized to huntingtin aggregates. Cystamine treatment normalized transglutaminase and GGEL levels in R6/2 mice. These findings are consistent with the hypothesis that transglutaminase activity may play a role in the pathogenesis of HD, and they identify cystamine as a potential therapeutic strategy for treating HD patients.

Immunologic quantitation of the carcinoma specific human carcinoma antigen in clinical samples.

BACKGROUND: Based on the cross-reactivity of the human carcinoma antigen, HCA, with epiglycanin, a mouse mammary carcinoma cell surface glycoprotein, HCA has been detected in the tissue and blood of patients with every type of epithelium-derived cancer tested. METHODS: Competitive binding assays utilized the following antiepiglycanin antibodies: a polyclonal rabbit antiserum (immunoglobulin [Ig] G and IgM) in a radioimmunoassay; mouse monoclonal antibodies (Ab-1, IgM) on immunoplates; anti-idiotypic (Ab-2) and anti-anti-idiotypic (Ab-3) monoclonal antibodies (both IgG) from spleen cells of C57BL mice immunized, respectively, with Ab-1 and Ab-2, and utilized on immunoplates. IgG and IgM antibodies were evaluated for their ability to detect HCA and to distinguish between the blood of patients with, or without, carcinomas. RESULTS: Assays with the rabbit antiserum distinguished plasmas of metastatic breast carcinoma patients from those of patients with benign breast disease with a sensitivity of approximately 93% (specificity 90%). Antiepiglycanin IgM monoclonal antibodies (i.e., AE3) showed high specificity and sensitivity (> 90%) with sera from advanced carcinoma patients when compared with normal sera. The IgG anti-anti-idiotypic (Ab-3) monoclonal antibodies (i.e., AF2), which bind the same epitope as Ab-1, appear to possess less nonspecific binding capacity, however, than the Ab-1 (IgM) antibodies. Anti-Ab-1 (i.e., C8F2) anti-idiotypic monoclonal antibodies, which bear an idiotope equivalent to the epitope present in epiglycanin and the HCA, demonstrated greater consistency as a standard calibrator and for coating wells than epiglycanin. CONCLUSIONS: The concentration of the HCA in the body fluids of patients with carcinomas may be accurately determined by competitive binding assays. It is suggested that the use of anti-idiotypic antibodies (IgG), rather than epiglycanin/HCA, and Ab-3 anti-anti-idiotypic antibodies (IgG), rather than Ab-1 (IgM), will improve the consistency, as well as the sensitivity and specificity, of the assay.