Fragment based drug discovery using fluorescence correlation: spectroscopy techniques: challenges and solutions.

Novel starting points for medicinal chemistry programmes can be effectively identified by screening libraries of fragment molecules in biochemical assays at high concentration. The key to success with this approach is the combination of a high quality fragment library with sensitive biochemical screening methods. There are an increasing number of literature reports where weakly active fragment molecules have been identified by high concentration biochemical assays. We have successfully demonstrated the use of high concentration screening of fragments, using a portfolio of single-molecule Fluorescence Correlation Spectroscopy (FCS+plus) detection techniques to ensure the highest reproducibility and sensitivity, and have determined the binding mode of active fragments to target proteins by X-ray crystallography. Further biophysical detection methods are reviewed for their applicability to studies of fragment binding.

Saving clinicians' time by delegating routine aspects of therapy to a computer: a randomized controlled trial in phobia/panic disorder.

BACKGROUND: The demand for time-consuming psychotherapy of phobia/panic exceeds the supply of trained therapists. Delegating routine therapy aspects to a computer might ease this problem. METHOD: Ninety-three out-patients with phobia or panic disorder were randomized in a 2: 2 : 1 ratio to have self-exposure therapy guided either mainly by a stand-alone computer system (FearFighter) or entirely face-to-face by a clinician, or to have mainly computer-guided self-relaxation as a placebo. Both computer groups (FearFighter and relaxation) had brief back-up advice from a clinician. Primary outcome measures were self- and blind-assessor ratings of Main Problem and Goals, and Global Phobia. RESULTS: Drop-outs occurred significantly more often in the two self-exposure groups (43% if mainly computer-guided, 24% if entirely clinician-guided) than with self-relaxation (6%); the difference between the two self-exposure groups was not significant. Even with all drop-outs included, the mainly computer-guided exposure group and the relaxation group had 73% less clinician time per patient than did the entirely clinician-guided exposure group. The two self-exposure groups had comparable improvement and satisfaction at post-treatment and at 1-month follow-up, while relaxation was ineffective. Mean improvement on the primary outcome measures (self- and assessor-rated) was 46% computer, 49% clinician, 9% relaxation at post-treatment (week 10) and 58% computer, 53% clinician and -4% relaxation at 1-month follow-up (week 14). Mean effect sizes on the primary outcome measures were 2.9 computer, 3.5 clinician and 0.5 relaxation at post-treatment; and 3.7 computer, 3.5 clinician and 0.5 relaxation at 1-month follow-up. The assessor did not rate patients at follow-up. CONCLUSIONS: Despite its (non-significantly) higher dropout rate, self-exposure therapy for panic/ phobia cut clinician time per patient by 73% without losing efficacy when guided mainly by a computer rather than entirely by a clinician. The finding needs confirmation at a follow-up that is longer and includes a blind assessor. Self-relaxation had the highest rate of completers but was ineffective.

13-week oral toxicity study with stanol esters in rats.

Plant sterols and their saturated derivatives, known as stanols, reduce serum cholesterol when consumed in amounts of approximately 2 g per day. Stanol fatty acid esters have been developed as a highly fat-soluble form that may lower cholesterol more effectively than stanols. Stanol esters occur naturally in human diets, but at levels far below those known to lower cholesterol. The present study was conducted to assess the safety of stanol esters upon subchronic ingestion at levels comparable to or exceeding those recommended for lowering cholesterol. Two stanol fatty acid ester preparations, wood-derived stanol esters and vegetable oil-derived stanol esters, were fed to groups of 20 male and 20 female Wistar rats for 13 weeks, at dietary concentrations of 0, 0.2, 1, and 5% total stanols (equivalent to 0, 0.34, 1.68, and 8.39% wood-derived stanol esters and 0, 0.36, 1.78, and 8.91% vegetable oil-derived stanol esters). Both preparations were well tolerated as evidenced by the absence of clinical changes or major abnormalities in growth, food and water consumption, ophthalmoscopic findings, routine hematological and clinical chemistry values, renal concentrating ability, composition of the urine, appearance of the feces, estrus cycle length, organ weights, gross necropsy findings, and histopathological findings. Plasma cholesterol and phospholipids were slightly decreased in males fed the stanol esters. In both sexes, plasma levels of plant sterols were decreased whereas those of stanols tended to increase. Fecal excretion of sterols, including cholesterol, and stanols was markedly increased in the stanol ester groups. Compared to controls, male rats fed stanol esters showed somewhat lower liver weights and more pronounced glycogen depletion. These hepatic changes were considered to reflect an altered nutritional condition and not a pathological condition. Plasma levels of vitamin E, vitamin K1, and, to a lesser extent, vitamin D were decreased in males and females fed the high-dose diets. Hepatic levels of vitamins E and D showed similar changes (vitamin K1 in the liver was not determined). For both preparations, the mid-dose level (1% total stanols in the diet) was a no-observed-adverse-effect level. This dietary level provided approximately 0.5 g total stanols/kg body wt/day.

Developmental toxicity study of D-tagatose in rats.

D-tagatose is a low-calorie sweetener that tastes like sucrose. The developmental toxicity of D-tagatose was investigated in Crl:CD(SD)BR rats administered D-tagatose at three dose levels (4000, 12,000, and 20,000 mg/kg body wt/day) via gastric intubation on days 6-15 of gestation. No compound-related toxicity was seen among any of the maternal groups. No treatment-related clinical effects were seen in the maternal animals at the 4000 mg/kg/day dose level. At the mid- and high-dose levels, most maternal animals had unformed or watery stools; this effect was most prominent early in the treatment period (Gestation Days 6-8). This effect was attributed to the osmotic effect of the large amount of D-tagatose given to the animals at these doses. Since D-tagatose is not digested or absorbed to a large extent, most of the sugar passes into the colon where it absorbs water and is fermented by colonic bacteria. Mean weight gain for the low- and mid-dose animals was comparable to the control; however, the high-dose group experienced a mean weight loss over the Gestation Day 6-9 interval. Over the entire treatment interval, however, mean weight gain for the high-dose animals was comparable to control. The decreased weight gain in the high-dose animals during the Gestation Day 6-9 interval was considered to be a direct result of laxation. In addition to the effect of laxation on body weight, reduced food consumption also contributed to the decreased weight gain. In the low-dose animals, no effect on food consumption was seen; however, both mid- and high-dose animals had food consumption values that were statistically significantly lower than the control. Food consumption was lowest during the Gestation Day 6-9 interval, the period when laxation was most prominent. Food consumption rebounded and was statistically significantly higher than the control for the mid- and high-dose animals during the posttreatment interval. Maternal liver weight for the low-dose animals was comparable to the control. However, a statistically significant increase in mean maternal liver weight was noted for the mid-and high-dose animals. Based on a lack of any corresponding histopathology, the increased liver weights were not considered toxicologically significant. There were no adverse effects on reproductive performance noted in any treatment group. No adverse treatment-related fetal effects on fetal weight, sex distribution, liver weight, or external, skeletal, or visceral malformations were noted at any dose level.

Multicenter evaluation of electrical stimulation systems for walking.

OBJECTIVE: To test the long-term benefits of several noninvasive systems for functional electrical stimulation (FES) during walking. DESIGN: Forty subjects (average years since injury, 5.4) were studied in four centers for an average time of 1 year. Gait parameters were tested for all subjects with and without FES. Thus, subjects served as their own controls, since the specific effect of using FES could be separated from improvements resulting from other factors (e.g., training). SETTING: Subjects used the devices in the community, but were tested in a university or hospital setting. PATIENTS: Subjects with spinal cord injury (n = 31) were compared to subjects with cerebral damage (n = 9). MAIN OUTCOME MEASURES: Gait parameters (speed, cycle time, stride length). Acceptance was studied by means of a questionnaire. RESULTS: Some initial improvement in walking speed (average increase of >20%) occurred, and continuing gains were seen (average total improvement, 45%). The largest relative gains were seen in the slowest walkers (speeds of <0.3 m/sec). Acceptance of the FES systems was good and improved systems have been developed using feedback from the subjects. CONCLUSIONS: Based on the improvements in speed and the acceptance of these FES systems, a greatly increased role for FES in treating gait disorders is suggested.

The fate of heavy oil wastes in soil microcosms. II: A performance assessment of source correlation indices.

Chemical fingerprinting is commonly undertaken to assist in the resolution of multi-party liability disputes, particularly when contaminants have migrated beyond property boundaries, in litigation-driven environmental assessments related to oil spills, and in assessing potential environmental impacts following releases of petroleum products into the environment. In this paper, we present data relating to the performance of source correlation indices for selected heavy oils over the course of a 9-month microcosm study. The results obtained in this study demonstrated that hopane pair indices varied little in magnitude, and may therefore be considered reliable source correlation indices. Over the course of the 9-month microcosm study [17 alpha(H)21 beta(H)-norhopane: 17 alpha(H)21 beta(H)-hopane] exhibited mean values of 0.7 +/- 0.1 for a heavy ballast oil, and mean values that varied between 0.6 and 0.7 (+/- 0.05) for a crude oil. Similarly [(17 alpha(H)21 beta(H)-homohopane (22S): 17 alpha(H)21 beta(H)-homohopane (22R)] gave a mean value of 1.3 (precisions less than 0.05) and [17 alpha(H)21 beta(H)-bishomohopane: 17 alpha(H)21 beta(H)-methylhopane] varied between 1.3 and 1.6 (precision up to 0.1) for the same crude oil. These source correlation indices may be used to support a correlation between fresh and weathered oil samples for source identification purposes involving heavy and crude oil contamination of the terrestrial environment.

Combinatorial chemistry as a tool for drug discovery.

The question 'will combinatorial chemistry deliver real medicines' has been posed [96]. First it is important to realise that the chemical part of the drug discovery process cannot stand alone; the integration of synthesis and biological assays is fundamental to the combinatorial approach. The results presented in Tables 3.1 to 3.8 suggest that so far smaller directed combinatorial libraries have obtained equivalent results to those obtained previously from traditional medicinal chemistry analogue programs. Unfortunately, because of the long time it takes to develop pharmaceutical drugs there are no examples yet of marketed drugs discovered by combinatorial methods. There are interesting examples where active leads have been discovered from the screening of the same library against multiple targets (e.g. libraries 13, 39, 43, 66, 71 and 76). It is now possible to handle much larger libraries of non-oligomeric structures and the chemistry required for such applications is becoming available. Whether combinatorial approaches can also be adapted to deal with all the other requirements of a successful pharmaceutical (lack of toxicity, bioavailability etc.) is open to question but there are already examples such as cassette dosing [235-237]. However we can still be optimistic about the possibility of larger libraries producing avenues of investigation for the medicinal chemist to develop into real drugs. Combinatorial chemistry is an important tool for the medicinal chemist.