The Alzheimer's Disease Assessment Scale: evaluation of psychometric properties and patterns of cognitive decline in multicenter clinical trials of mild to moderate Alzheimer's disease.

PURPOSE: To evaluate the psychometric properties and patterns of decline on the total score and item scores of the cognitive subscale of the Alzheimer's Disease Assessment Scale (ADAS-Cog) in patients with Alzheimer Disease (AD). METHODS: We analyzed data from 536 AD outpatients randomized to the placebo group in two identical 26-week multicenter drug trials. RESULTS: Mean deterioration at week 26 on the ADAS-Cog total score for subjects with moderate dementia was 84% greater than that for those with milder severity ( p < 0.001). After adjusting for this effect, age ( p = 0.015) and educational level ( p = 0.01) also predicted cognitive decline. In a model, absolute change for most individual ADAS items was less than 10% of the possible change, and it was generally smaller than one-third of the standard deviation of the measure. Measurement error variability was greatest for word recognition and the "placebo" effect was greatest for word recall. Variability increased with trial duration in a model. CONCLUSIONS: There is a relationship between baseline severity and magnitude of cognitive decline. In 6-month trials, measurement error makes a substantial contribution to the variance in ADAS-Cog change scores. Sensitivity to intervention effects will therefore depend both on the variability and magnitude of change. Such data must be considered when designing future trials to minimize measurement error variability and increase sensitivity for specific populations.

The Bayer-Activities of Daily Living Scale (B-ADL): results from a validation study in three European countries.

The Bayer-Activities of Daily Living Scale (B-ADL) is a 25-item, informant-rated questionnaire which was developed as a brief and internationally applicable instrument for assessing functional disabilities. The scale's target group are elderly patients suffering from mild to moderate dementia or cognitive impairment. To investigate the reliability and validity of different language versions, the B-ADL was administered in the UK, Germany, and Spain to a total of 1,433 subjects with a wide range of cognitive decline. The results from the three country samples were very similar, with internal consistency being above 0.98 (Cronbach alpha). A factor analysis revealed that a one-factor solution accounted for most of the variance. The B-ADL total score significantly increased between adjacent Global Deterioration Scale (GDS) stages 1 to 5. A second factor analysis entering additional variables (GDS stage, Mini-Mental State Examination or MMSE subscores, age, years of education, gender, and country) revealed that all B-ADL items loaded on the same factor, "dementia severity", and that they were not related to age, education, gender, or country. In the identification of subjects with clinically manifest dementia symptoms (GDS stages 4 and 5), the B-ADL proved to be as efficient as the MMSE in the UK and German samples and superior to the MMSE in the Spanish sample.

Interpretation of complex forensic DNA mixtures.

Forensic evidentiary samples routinely contain DNA from multiple contributors. The interpretation of these mixtures can be a challenging task for the DNA scientist. Several approaches are discussed (no calculation- qualitative statement; probability of exclusion; likelihood ratio estimates; presumptive genotype assignment based on peak heights), which have been employed to assess the significance of an inclusion/match when DNA mixtures have been detected in casework samples. These statistical approaches are discussed in light of technical challenges that can arise when evaluating evidentiary samples.

Fluorescence in situ hybridization (FISH) for rapid detection of aneuploidy: experience in 911 prenatal cases.

Fluorescence in situ hybridization (FISH) was performed with probes specific for chromosomes 13, 18, 21, X and Y on 911 of 11123 (8.2%) amniotic fluid samples submitted to the present authors' laboratory for cytogenetic analysis over an 8-year period. Altogether 3516 hybridizations were performed with an interpretable FISH result on all chromosomes requested in 884/911 (97%) of cases. An uninformative FISH result occurred in 44 hybridizations among 27 cases (3%). Of a total of 89 karyotypically proven cases with aneuploidy that might have been detected by FISH, the overall detection rate was 84%. An inconclusive or incomplete FISH result occurred in 9/89 (10%) of these proven aneuploid cases. In the remaining 80 informative proven aneuploid cases, correct detection of aneuploidy was accomplished in 75/80 (94%) of samples. A false-negative result occurred in the remaining 5/80 (6%) of such informative cases. Eighteen cases had karyotypically proven abnormalities that could not have been detected by the targeted FISH. Aside from these 18 cases, FISH allowed correct detection of normal disomy in 785/804 (98%) of such cases. An incomplete FISH result occurred in 18 normal disomic cases. There was a single possible 'false-positive' FISH result for chromosome 21. Interphase FISH analysis of uncultured amniotic fluid cells has been shown to be a useful laboratory tool for rapid fetal aneuploidy screening during pregnancy. As with all clinical laboratory diagnostic tests, incomplete or inconclusive results (or even interpretive errors) occur in a small percentage of cases. Nevertheless, FISH results accompanied by other data and by appropriate counseling provide clinicians and patients with valuable information for clinical decision-making surrounding family planning and pregnancy management.

Overview of human identity testing and forensic genetics.

This unit discusses the development of the field of forensic genetics, touching on the quality control and legal issues that are central to this area. It closes with a discussion of some caveats for interpreting forensic DNA results.

Collecting and handling samples for parentage and forensics DNA-based genetic testing.

The unit covers Variable Numbers of Tandem Repeats (VNTR) based paternity analysis as well as the newer methods relying on PCR to analyze sequence specific polymorphisms and microsatellite regions. The discussion of data analysis and probability calculations has been expanded to address a number of special circumstances, such as the lack of sample from an alleged father, motherless cases, and more.

Isolation of DNA from forensic evidence.

This unit covers the many and varied methods for extracting DNA from such diverse specimens as blood, tissue, stamps and envelopes, and cigarette butts, among others. Modifications to the methods that allow the DNA to be used for either PCR or Southern blotbased analyses are also included.

RFLP analysis of forensic DNA samples with single-locus VNTR genetic markers.

This unit covers the many and varied methods for extracting DNA from such diverse specimens as blood, tissue, stamps and envelopes, and cigarette butts, among others. Modifications to the methods that allow the DNA to be used for either PCR or Southern blotbased analyses are also included.

Manual methods for PCR-based forensic DNA analysis.

This unit provides validated PCR-based methods to test for sequence polymorphisms and length polymorphisms. It includes a description of the reverse dot blot method for detecting sequence polymorphisms. The forensic PCR systems used to detect length polymorphisms are based on detection of different-sized PCR products following electrophoresis in either native or denaturing polyacrylamide gels. The unit offers a description of the amplification and detection of the variable number of tandem repeat (VNTR) length polymorphism at the D1S80 locus. It also describes the multiplex amplification of three separate autosomal short tandem repeats (STRs) and the X- and Y-linked amelogenin alleles. These PCR products are electrophoresed on a denaturing polyacrylamide gel. Support protocols for creating permanent records of silver-stained gels, checking the quality of reagents, and interpreting PCR-based tests are provided.

Randomized, double-blind, placebo-controlled, multicenter study to evaluate the safety and tolerability of metrifonate in patients with probable Alzheimer disease. The Metrifonate Study Group.

A randomized, double-blind, placebo-controlled, parallel-group study was undertaken to evaluate the safety and tolerability of a once-daily oral administration of metrifonate in patients with probable mild to moderate Alzheimer disease. Metrifonate was given as a loading dose of 125-225 mg based on weight (2.5 mg/kg) for 2 weeks, followed by a maintenance dose of 50-90 mg based on weight (1.0 mg/kg) for 4 weeks. Twenty-nine patients received metrifonate, and 10 patients received placebo. Metrifonate produced a mean erythrocyte acetylcholinesterase inhibition at the end of treatment of 86.3%. The proportion of patients who experienced at least one adverse event was comparable between the metrifonate (76%) and placebo (80%) groups. Selected adverse events in disfavor of metrifonate (defined as those for which the incidence in the metrifonate and placebo groups differed by at least 10%) were diarrhea, nausea, leg cramps, and accidental injury. Adverse events were predominantly mild in intensity and transient. No severe adverse events were experienced by any patient. The most notable hemodynamic change observed during metrifonate treatment was a clinically insignificant mean decrease in the heart rate (by electrocardiogram) of approximately 9 beats/min, compared with an approximate 3-beats/min decrease for the placebo group. No muscle weakness was observed in this study. No clinically relevant laboratory abnormalities, such as liver toxicity, or changes in exercise tolerance or pulmonary function tests were found with metrifonate treatment. This metrifonate dose provided a high level of acetylcholinesterase inhibition, which was associated in these patients with a favorable safety and tolerability profile. Indeed, the magnitude of the peripheral acetylcholinesterase inhibition is the highest tolerable inhibition level yet observed.

Detection of mosaicism in amniotic fluid cultures: a CYTO2000 collaborative study.

PURPOSE: To evaluate the assumptions on which the American College of Medical Genetics (ACMG) Standards and Guidelines for detecting mosaicism in amniotic fluid cultures are based. METHODS: Data from 653 cases of amniotic fluid mosaicism were collected from 26 laboratories. A chi-square goodness-of-fit test was used to compare the observed number of mosaic cases with the expected number based on binomial distribution theory. RESULTS: Comparison of observed data from the in situ colony cases with the expected distribution of cases detected based on the binomial distribution did not reveal a significant difference (P = 0.525). CONCLUSIONS: The empirical data fit the binomial distribution. Therefore, binomial theory can be used as an initial discussion point for determining whether ACMG Standards and Guidelines are adequate for detecting mosaicism.

Twins, placentas, and genetics: acardiac twinning in a dichorionic, diamniotic, monozygotic twin gestation.

We describe a human acardiac twin with associated vascular anastomoses in a dichorionic diamniotic fused twin placenta. A 22-year-old woman delivered a healthy 3,554 g male infant and a fused diamniotic dichorionic twin placenta with a 230 g umbilical cord-attached, skin-covered, ovoid mass, consistent with acardiac amorphus. By gross and histological examination, the placental dividing membranes comprised four leaves, one amnion from each placenta, and two centrally fused chorions, diagnostic of dichorionicity. Placental barium injection of the normal twin's umbilical vein showed an anastomosis with the acardiac twin which traversed the dividing membranes, then supplied major vessels of the acardiac mass via its 5.5 cm umbilical cord. DNA-typing studies of the normal twin's placenta and of the acardiac twin's tissues revealed identical alleles at 11 distinct genetic polymorphic loci, consistent with monozygosity. Our findings demonstrate that vascular anastomoses can occur in dichorionic twin placentas, and that human acardiac twinning is not, as heretofore believed, restricted to monochorionic placentas.

Metrifonate treatment of the cognitive deficits of Alzheimer's disease. Metrifonate Study Group.

The efficacy and safety of metrifonate, an acetylcholinesterase inhibitor, was evaluated clinically in patients diagnosed with mild to moderate Alzheimer's disease (AD). This was a prospective, 30-week, multicenter, double-blind, randomized, parallel group, dose-finding study, which included a 2-week screening period, a 12-week treatment period, and follow-up visits at 8 and 16 weeks post-treatment. Patients received placebo or metrifonate once daily. Metrifonate-treated patients received a loading dose of 0.5 mg/kg (25 to 45 mg), 0.9 mg/kg (45 to 80 mg), or 2.0 mg/kg (100 to 180 mg) for 2 weeks, followed by a maintenance dose of 0.2 mg/kg (10 to 20 mg), 0.3 mg/kg (15 to 25 mg), or 0.65 mg/kg (30 to 60 mg) for 10 weeks. Four hundred eighty patients were enrolled. Percentages of patients completing double-blind treatment were 96% in the placebo group and 89 to 94% in the metrifonate group. Metrifonate significantly improved cognitive ability, as assessed by the Alzheimer's Disease Assessment Scale-Cognitive Subscale (ADAS-Cog), and enhanced global function, as assessed the Clinicians's Interview-Based Impression of Change with Caregiver Input (CIBIC-Plus). At 3 months, in the intent-to-treat patients, the treatment difference for the change in ADAS-Cog score in favor of metrifonate was 2.94 points (95% CI, 1.61 to 4.27; p = 0.0001). These patients also exhibited a 0.35-point improvement on the CIBIC-Plus relative to the placebo patients (95% CI, 0.15 to 0.54; p = 0.0007). Patients receiving lower drug doses had scores intermediate to those of the placebo and the 0.65 mg/kg metrifonate groups on both performance scales. The drug was well tolerated; side effects were predominantly gastrointestinal in nature, and no hepatic toxicity was observed. Therefore, in this study, metrifonate safely improved the cognitive deficits and benefited the global function of AD patients.

Metrifonate benefits cognitive, behavioral, and global function in patients with Alzheimer's disease.

OBJECTIVE: To evaluate the efficacy and safety of metrifonate, an acetylcholinesterase inhibitor, in patients clinically diagnosed with probable Alzheimer's disease (AD) of mild to moderate severity. METHODS: A prospective, 36-week, multicenter, double-blind, randomized, parallel group study of metrifonate in probable AD patients, including a 2-week screening period, a 26-week double-blind treatment period, and a follow-up visit at 8 weeks post-treatment. A total of 24 ambulatory clinics in the United States in a variety of settings, including contract research organizations, public health facilities, and universities. Patients met diagnostic criteria for probable AD as defined by the work group of the National Institute for Neurological and Communicative Diseases and Stroke and the Alzheimer's Disease and Related Disorders Association. Patients had Mini-Mental State Examination (MMSE) scores of 10 to 26 and Ischemic Scores (Rosen Modification) of <4. A total of 408 patients were enrolled. Percentages of patients completing double-blind treatment were 88% and 79% in the placebo and metrifonate groups, respectively. Rates of discontinuation due to adverse events were 4% in the placebo group and 12% in the metrifonate group. Placebo or metrifonate was administered once daily. Metrifonate-treated patients received a loading dose of 100 to 180 mg based on weight (2.0 mg/kg) for 2 weeks, followed by a maintenance dose of 30 to 60 mg based on weight (0.65 mg/kg) for 24 weeks. Primary efficacy variables were the Alzheimer's Disease Assessment Scale-Cognitive Subscale (ADAS-Cog) and the Clinician's Interview-Based Impression of Change with Caregiver Input (CIBIC-plus). Secondary efficacy variables included the Neuropsychiatric Inventory (NPI), the Disability Assessment in Dementia, the Global Deterioration Scale (GDS), the ADAS-Noncognitive subscale (ADAS-Noncog), the MMSE, and the Clinician's Interview-Based Impression of Severity with Caregiver Input (CIBIS-plus). Outcome measures reflected changes from baseline at week 26 for all variables. Safety was assessed with incidences of premature termination, treatment-emergent events and mortality, and routine safety evaluations. RESULTS: After 26 weeks of metrifonate therapy, a 2.86-point treatment difference (p = 0.0001) was observed in the ADAS-Cog scores of the intent-to-treat AD patients. The treatment difference in the mean CIBIC-plus score at this time was 0.28 points (p = 0.0071). At week 26, treatment differences also were observed in the mean NPI total score (p = 0.0161). Analysis of the remaining secondary efficacy variables showed treatment differences that favored metrifonate but did not reach statistical significance. Metrifonate adverse events were predominantly mild in intensity. No hepatotoxicity was observed. CONCLUSIONS: Metrifonate was safe and well-tolerated. It enhanced not only the cognitive and global function, but also the behavioral function of patients diagnosed with mild to moderate AD. Therefore, metrifonate appears to be useful in the symptomatic treatment of AD.

Safety and tolerability of metrifonate in patients with Alzheimer's disease: results of a maximum tolerated dose study.

Metrifonate, a pro-drug that is transformed non-enzymatically into a potent inhibitor of acetylcholinesterase (AChE), has been used in the tropics for over 30 years for the treatment of schistosomiasis. A pilot study, and Phase I and Phase II studies of metrifonate in Alzheimer's disease (AD) patients conducted prior to the current study showed benign, dose-dependent adverse event profiles consisting primarily of gastrointestinal events, optimal daily dosing with a loading phase (in the absence of a loading dose phase, 6-8 weeks were required to attain steady-state AChE inhibition levels), and an improvement in Alzheimer's Disease Assessment Scale (ADAS) scores. The current open-label study was designed to evaluate the safety and tolerability of relatively high loading doses, followed by lower maintenance doses of metrifonate in the same patient population, and to determine the maximum tolerated dose (MTD) of metrifonate. Accordingly, the first cohort of 8 probable AD patients (per National Institute of Neurological and Communicative Disorders and Stroke-Alzheimer's Disease and Related Disorders Association [NINCDS-ADRDA] criteria) were administered once-daily loading doses of 2.5 mg/kg (125-225 mg) for 14 days, followed by 4.0 mg/kg (200-360 mg) for an additional 3 days. These patients were maintained on once-daily doses of 2.0 mg/kg (100-180 mg) for 14 days. AChE inhibition for this cohort ranged from 88% to 94%. On Day 28 of 31, this cohort was discontinued due to moderate to severe side effects in 6 patients; consequently, the second cohort of 8 probable AD patients received a once-daily loading dose of 2.5 mg/kg (125-225 mg) for 14 days followed by a once-daily maintenance dose of 1.5 mg/kg (75-135 mg) for 35 days. This maintenance dose yielded an AChE inhibition level ranging from 89% to 91%. In spite of an AChE inhibition level comparable to that achieved with the higher dose, the reduced dose was associated with a more favorable adverse event profile which was mainly gastrointestinal and musculoskeletal in nature. The maximum tolerated dose was established at 1.5 mg/kg/day (75-135 mg/day) for maintenance dosing in AD patients.

Pharmacokinetics, pharmacodynamics, and safety of metrifonate in patients with Alzheimer's disease.

Metrifonate is converted nonenzymatically to 2.2, dimethyl dichlorovinyl phosphate (DDVP), an inhibitor of acetylcholinesterase (AChE). This 21-day, randomized, double-blind, placebo-controlled trial of metrifonate in patients with Alzheimer's disease (n = 27) evaluated four doses, each administered orally once daily. All patients received a loading dose (LD) for 6 days followed by a maintenance dose (MD) for 15 days. The treatment groups were: panel 1, LD = 1.5 mg/kg (75-135 mg), MD = 0.25 mg/kg (12.5-25 mg); panel 2, LD = 2.5 mg/kg (125-225 mg), MD = 0.40 mg/kg (20-35 mg); panel 3, LD = 4.0 mg/kg (200-335 mg), MD = 0.65 mg/kg (30-60 mg); and panel 4, LD = 4.0 mg/kg (200-335 mg), MD = 1.0 mg/kg (50-90 mg). All metrifonate doses were well tolerated. Most adverse events were mild to moderate in intensity, gastrointestinal in nature, and transient. Mean area under the concentration-time curve (AUC) and maximum concentration (Cmax) for both metrifonate and DDVP increased in relation to dose. Metrifonate and DDVP had similar, largely dose-independent mean values for time to Cmax (tmax) and half-life (t1/2). There was little or no accumulation of either metrifonate or DDVP with long-term administration. After 21 days of treatment, mean percent erythrocyte AChE inhibition was 14%, 35%, 66%, 77%, and 82% for placebo and panels 1 through 4, respectively. Cognitive improvement was observed with the two highest metrifonate doses. These results reflect favorable safety and pharmacokinetic profiles for the use of metrifonate in the treatment of Alzheimer's disease.

The Alzheimer's Disease Assessment Scale: patterns and predictors of baseline cognitive performance in multicenter Alzheimer's disease trials.

The cognitive subscale of the Alzheimer's Disease Assessment Scale (ADAS-Cog) is used as an efficacy measure in clinical drug trials of Alzheimer's disease (AD). We used data from 1,648 AD participants in two identical 26-week multicenter drug trials to examine the distribution of baseline ADAS-Cog scores in relation to selected demographic and clinical variables, Mini-Mental State Exam (MMSE), Global Deterioration Scale (GDS), and Geriatric Evaluation by Relative's Rating Instrument (GERRI) scores. At baseline, the mean (+/-SD) MMSE score was 18 +/- 4, the ADAS-Cog score was 28 +/- 11, and most subjects were in GDS stage 4 or 5. The ADAS-Cog score was statistically significantly correlated with MMSE (R = -0.76, p < 0.0001) and GERRI (R = 0.40, p < 0.0001) total scores. Correlations among the ADAS-Cog items ranged from 0.19 to 0.59 and all were statistically significant (p < 0.0001). In a multiple regression model, younger age, male gender, older age at onset of dementia, use of concurrent estrogen, and use of concurrent anti-inflammatory agents were statistically significantly associated with superior cognitive performance. We also present data on the distribution of ADAS-Cog scores in relation to subjects' age, level of education, MMSE score, and GDS stage. Because age, MMSE score, and GDS stage (and not the ADAS-Cog) are commonly used to select subjects for AD clinical trials, our data should improve the ability of sponsors to predict ADAS-Cog scores of the subjects in their trials on the basis of the inclusion criteria used. Our data also suggest that age, gender, age at onset of dementia, level of education, and use of estrogen (in women) or anti-inflammatory drugs are related to cognitive abilities in AD. Further studies are needed to assess how and when cognitive differences related to these variables arise.

Memory, language, and praxis in Alzheimer's disease: norms for outpatient clinical trial populations.

The cognitive subscale of the Alzheimer's Disease Assessment Scale (ADAS-Cog) is designed specifically to assess memory, language, and praxis dysfunctions characteristic of Alzheimer's disease (AD). In this report, we use data from 1,648 AD participants in two identical 26-week multicenter drug trials to derive clinical trial population-based norms for ADAS-Cog subtest scores. All 11 ADAS-Cog item scores were sensitive to differences in dementia severity judged either by baseline Mini-Mental State Exam (MMSE) scores or by Global Deterioration Scale (GDS) stage of dementia. Using ADAS-Cog subtest scores alone, 85 percent of GDS 4 subjects and 69 percent of GDS 5 subjects could be classified correctly. In a stepwise discriminant analysis, orientation was the item with the greatest discriminatory power between subjects with GDS 4 and GDS 5 stages of dementia. ADAS-Cog subtest scores also varied by age, gender, educational level, and concurrent use of anti-inflammatory drugs or estrogen (in women). Such normative data may facilitate the selection of appropriate outcome measures to investigate differential treatment effects on specific cognitive domains or in specific AD subpopulations.

An ancient conserved gene expressed in the human inner ear: identification, expression analysis, and chromosomal mapping of human and mouse antiquitin (ATQ1).

We constructed and screened a human fetal cochlear cDNA library to identify genes involved in hearing and deafness. From this library we isolated a cDNA corresponding to the highly conserved ancient gene antiquitin (ATQ1). The plant homolog of ATQ1 is thought to be involved in regulating turgor pressure, a function that also would be essential for cells of the mammalian cochlea. Northern blots of 13 human fetal tissues show antiquitin to be highly expressed in cochlea, ovary, eye, heart, and kidney. Using RT-PCR of rat cochlear hair cell-specific cDNA libraries, we detect antiquitin expression in outer hair cells, but not in inner or vestibular type 1 hair cells, suggesting that antiquitin is not expressed ubiquitously in the cochlea. Human ATQ1 was mapped to human chromosome region 5q31 using fluorescence in situ hybridization, and mouse ATQ1 was mapped to mouse chromosome 18 by single-strand conformation polymorphism mapping of interspecific backcross progeny DNAs. Four human antiquitin-like sequences, possibly pseudogenes, were also identified and mapped.

Mapping and characterization of a novel cochlear gene in human and in mouse: a positional candidate gene for a deafness disorder, DFNA9.

Previously we identified a partial human cDNA for a novel cochlear transcript, hCoch-5B2 (HGMW-approved symbol D14S564E), using subtractive hybridization techniques. Herein we report isolation and characterization of both human and mouse (D12H14S564E) cDNAs for Coch-5B2. Full-length Coch5B2 deduced amino acid sequences reveal a very high degree of conservation in the coding region (89% nucleotide and 94% amino acid identity and a potential signal peptide and two regions of extensive homology to the collagen-binding type A domains of von Willebrand factor, also present in other secreted proteins, including extracellular matrix components. High levels of hCoch-5B2 expression are seen only in human fetal inner ear structures, cochlea, and vestibule, among a large panel of human fetal and adult tissues. Coch-5B2 expression in the mouse is more widespread than in the human, with message detected in mouse adult spleen, cerebrum, cerebellum/medulla, and thymus. In both species very low level expression is detected in total eye. More specifically, mouse retina shows a higher level of mCoch-5B2 message than sclera and choroid. We have mapped hCoch-5B2 to human 14q11.2-q13 by somatic cell hybrid analysis and FISH and, more precisely, using radiation hybrids to a region of markers linked to DFNA9, a nonsyndromic autosomal dominant sensorineural hearing loss with vestibular defects. Furthermore, we detect hCoch-5B2 on three overlapping YACs, two of which also contain one of the markers linked to DFNA9. mCoch-5B2 was genetically mapped in the mouse to chromosome 12, in a region of homologous synteny with human 14q11.2-q13, which contains the asp1 (audiogenic seizure prone) locus in the mouse.