Ventricular rate adaptation: a novel, rapid, cellular-based in-vitro assay to identify proarrhythmic and torsadogenic compounds.

INTRODUCTION: Delayed cardiac repolarization is an established risk factor for proarrhythmia and Torsades-de-Pointes (TdeP) that is typically measured in vitro during slow, regular stimulation. We have developed an alternative, novel, and rapid cellular-based approach for predicting drug-induced proarrhythmia that detects changes in electrical refractoriness based on mechanical responses (measured optically) during increasingly rapid trains of stimulation interspersed with pauses (mimicking the clinically observed short-long-short (SLS) stimulation sequence associated with the TdeP initiation). METHODS: Acutely isolated rabbit ventricular myocytes were superfused and electrically stimulated using an accelerating pacing protocol (APP) consisting of 12 consecutive pacing segments (10 beats per segment) with incrementally faster cycle lengths; trains were separated by pauses to identify loss of stimulus capture as well as to mimic clinically observed SLS sequences. Drug effects were evaluated based on a myocyte's ability to contract during progressively faster pacing segments (rate-adaptation); the earliest rate during which the myocyte fails to respond (longest cycle length with incomplete capture (CLIC)) was used to quantify electrophysiologic effects. RESULTS: Torsadogenic drugs known to delay repolarization during slow stimulation prolonged CLIC and dramatically limited the ability to respond to progressively rapid stimulation. The recognized proarrhythmic compounds E-4031, cisapride, grepafloxacin, and haloperidol rapidly prolonged CLIC at and above therapeutic concentrations in a concentration-dependent manner, while negative controls (captopril, indomethacin, and loratidine) do not affect rate-adaptation. DISCUSSION: Ventricular rate adaptation represents a novel approach for rapidly detecting drugs with torsadogenic risk using rapid rhythms that are typically not employed when evaluating proarrhythmic risk. This method is well suited for detecting and avoiding potential cardiac liabilities early in drug discovery ("frontloading") prior to final selection of candidate drugs.

Noise contribution to the correlation between temperature-induced localized reflectance of diabetic skin and blood glucose.

We used the effect of temperature on the localized reflectance of human skin to assess the role of noise sources on the correlation between temperature-induced fractional change in optical density of human skin (DeltaOD(T)) and blood glucose concentration [BG]. Two temperature-controlled optical probes at 30 degrees C contacted the skin, one was then cooled by -10 degrees C; the other was heated by +10 degrees C. DeltaOD(T) upon cooling or heating was correlated with capillary [BG] of diabetic volunteers over a period of three days. Calibration models in the first two days were used to predict [BG] in the third day. We examined the conditions where the correlation coefficient (R2) for predicting [BG] in a third day ranked higher than R2 values resulting from fitting permutations of randomized [BG] to the same DeltaOD(T) values. It was possible to establish a four-term linear regression correlation between DeltaOD(T) upon cooling and [BG] with a correlation coefficient higher than that of an established noise threshold in diabetic patients that were mostly females with less than 20 years of diabetes duration. The ability to predict [BG] values with a correlation coefficient above biological and body-interface noise varied between the cases of cooling and heating.

Studies on repository compound stability in DMSO under various conditions.

The chemical stability of repository compounds is affected by various environmental conditions during long-term storage. Studies were carried out to evaluate the effects of the following potential causes of instability of compounds in DMSO at a 10-mM concentration: water, oxygen, freeze/thaw cycles, and storage container material. A set of compounds was selected for the study based on structural diversity and functional group representation. Compound concentration was determined with liquid chromatography/ultraviolet spectroscopy/mass spectrometry (LC/UV/MS) analysis relative to an internal standard added to each sample. An accelerated study was conducted, and results demonstrate that most compounds are stable for 15 weeks at 40 degrees C. Water is more important in causing compound loss than oxygen. The freeze/thaw cycle study was done with freezing at -15 degrees C and thawing under nitrogen atmosphere at 25 degrees C. Two methods were used to redissolve compounds after thawing: agitation and repeated aspiration/dispense. The results indicate no significant compound loss after 11 freeze/thaw cycles. Compound recovery was also measured from glass and polypropylene containers for 5 months at room temperature, and no significant difference was found for these 2 types of containers.