Methodological developments vs. regulatory requirements

This is a discussion of the following three papers appearing in this special issue on adaptive designs: 'FDA's critical path initiative: A perspective on contributions of biostatistics' by Robert T. O'Neill, 'A regulatory view on adaptive/flexible clinical trial design' by H. M. James Hung, Robert T. O'Neill, Sue-Jane Wang and John Lawrence; and 'Confirmatory clinical trials with an adaptive design' by Armin Koch.     

Opening the adaptive toolbox

This is a discussion of the following three papers appearing in this special issue on adaptive designs: 'A regulatory view on adaptive/flexible clinical trial design' by H. M. James Hung, Robert T. O'Neill, Sue-Jane Wang and John Lawrence; 'Confirmatory clinical trials with an adaptive design' by Armin Koch; and 'FDA's critical path initiative: A perspective on contributions of biostatistics' by Robert T. O'Neill.     

Increased expression of TGF-β1 reduces tumor growth of human U-87 Glioblastoma Cells in vivo

The role that transforming growth factor β1 (TGF-β1) plays in influencing growth of glioma cells is somewhat controversial. To further understand the potential growth-regulatory effects of TGF-β1,we constructed an animal astroglial tumor model by injecting either wild-type or virally transduced human U-87 glioblastoma cells into nude rat brains. Wild type U-87 cells produced very low amounts of TGF-β1 and were highly tumorigenic. In contrast, U-87 cells transduced to express high levels of TGF-β1 showed reduced tumor size in vivo, in a dose-dependent manner. This reduction in tumor size was not due to either decreased vascularity or increased apoptosis. To test whether TGF-β1 overproduction inhibited tumor growth through an autocrine mechanism, the highest TGF-β1 producing cells were then double transduced with a vector expressing the kinase-truncated type II TGF-β receptor. Cells expressing high levels of truncated TGF-β receptor were less sensitive to TGF-β1 mediated growth inhibition in vitro and produced more aggressive tumors in vivo. The data suggest that the degree of tumorigenicity of the U-87 high-grade glioblastoma cell line may be associated with correspondingly low level of production of TGF-β1. These results also would tend to support the possibility that TGF-β1 may be useful in treating some high-grade gliomas.     

Reduction of infarct size with D-myo-inositol trisphosphate: role of PI3-kinase and mitochondrial K(ATP) channels

Prophylactic treatment with D-myo-inositol 1,4,5-trisphosphate hexasodium [D-myo-Ins(1,4,5)P3], the sodium salt of the endogenous second messenger Ins(1,4,5)P3, triggers a reduction of infarct size comparable in magnitude to that seen with ischemic preconditioning (PC). However, the mechanisms underlying D-myo-Ins(1,4,5)P3-induced protection are unknown. Accordingly, our aim was to investigate the role of four archetypal mediators implicated in PC and other cardioprotective strategies (i.e., PKC, PI3-kinase/Akt, and mitochondrial and/or sarcolemmal K(ATP) channels) in the infarct-sparing effect of D-myo-Ins(1,4,5)P3. Fifteen groups of isolated buffer-perfused rabbit hearts [5 treated with D-myo-Ins(1,4,5)P3, 5 treated with PC, and 5 control cohorts] underwent 30 min of coronary artery occlusion and 2 h of reflow. One set of control, D-myo-Ins(1,4,5)P3, and PC groups received no additional treatment, whereas the remaining sets were infused with chelerythrine, LY-294002, 5-hydroxydecanoate (5-HD), or HMR-1098 [inhibitors of PKC, PI3-kinase, and mitochondrial and sarcolemmal ATP-sensitive K+ (K(ATP)) channels, respectively]. Infarct size (delineated by tetrazolium staining) was, as expected, significantly reduced in both D-myo-Ins(1,4,5)P3- and PC-treated hearts versus controls. D-myo-Ins(1,4,5)P3-induced cardioprotection was blocked by 5-HD but not HMR-1098, thereby implicating the involvement of mitochondrial, but not sarcolemmal, K(ATP) channels. Moreover, the benefits of D-myo-Ins(1,4,5)P3 were abrogated by LY-294002, whereas, in contrast, chelerythrine had no effect. These latter pharmacological data were corroborated by immunoblotting: D-myo-Ins(1,4,5)P3 evoked a significant increase in expression of phospho-Akt but had no effect on the activation/translocation of the cardioprotective epsilon-isoform of PKC. Thus PI3-kinase/Akt signaling and mitochondrial K(ATP) channels participate in the reduction of infarct size afforded by prophylactic administration of D-myo-Ins(1,4,5)P3.     

Deglutitive upper esophageal sphincter relaxation: a study of 75 volunteer subjects using solid-state high-resolution manometry

This study aimed to use a novel high-resolution manometry (HRM) system to establish normative values for deglutitive upper esophageal sphincter (UES) relaxation. Seventy-five asymptomatic controls were studied. A solid-state HRM assembly with 36 circumferential sensors spaced 1 cm apart was positioned to record from the hypopharynx to the stomach. Subjects performed ten 5-ml water swallows and one each of 1-, 10-, and 20-ml volume swallows. Pressure profiles across the UES were analyzed using customized computational algorithms that measured 1) the relaxation interval (RI), 2) the median intrabolus pressure (mIBP) during the RI, and 3) the deglutitive sphincter resistance (DSR) defined as mIBP/RI. The automated analysis succeeded in confirming bolus volume modulation of both the RI and the mIBP with the mean RI ranging from 0.32 to 0.50 s and mIBP ranging from 5.93 to 13.80 mmHg for 1- and 20-ml swallows, respectively. DSR was relatively independent of bolus volume. Peak pharyngeal contraction during the return to the resting state postswallow was almost 300 mmHg, again independent of bolus volume. We performed a detailed analysis of deglutitive UES relaxation with a novel HRM system and customized software. The enhanced spatial resolution of HRM allows for the accurate, automated assessment of UES relaxation and intrabolus pressure characteristics, in both cases confirming the volume-dependent effects and absolute values of these parameters previously demonstrated by detailed analysis of concurrent manometry/fluoroscopy data. Normative values were established to aid in future clinical and investigative studies.     

Kidney plays a major role in ammonia homeostasis after portasystemic shunting in patients with cirrhosis

The kidney plays an important role in ammonia metabolism. In this study the hypothesis was tested that the kidney can acutely diminish ammonia release after portacaval shunting. Thirteen patients with cirrhosis (6 female/7 male, age 54.4 +/- 3.3 yr) were studied. Blood was sampled prior to and 1 h after transjugular intrahepatic stent-shunt (TIPSS) insertion from the portal vein, a hepatic vein, the right renal vein, and the femoral vein, and renal and liver plasma flow were measured. Prior to TIPSS, renal ammonia release was significantly higher than ammonia release from the splanchnic region, which was not significantly different from zero. TIPSS insertion did not change arterial ammonia concentration or ammonia release from the splanchnic region but reduced renal ammonia release into the circulation (P < 0.05) to values that were not different from zero. TIPSS resulted in a tendency toward increased venous-arterial ammonia concentration differences across leg muscle. Post-TIPSS ammonia efflux via portasystemic shunts was estimated to be seven times higher than renal efflux. Kidneys have the ability to acutely diminish systemic ammonia release after portacaval shunting. Diminished renal ammonia release and enhanced muscle ammonia uptake are important mechanisms by which the cirrhotic patient maintains ammonia homeostasis after portasystemic shunting.     

TREK-1, a K+ channel involved in polymodal pain perception

The TREK-1 channel is a temperature-sensitive, osmosensitive and mechano-gated K+ channel with a regulation by Gs and Gq coupled receptors. This paper demonstrates that TREK-1 qualifies as one of the molecular sensors involved in pain perception. TREK-1 is highly expressed in small sensory neurons, is present in both peptidergic and nonpeptidergic neurons and is extensively colocalized with TRPV1, the capsaicin-activated nonselective ion channel. Mice with a disrupted TREK-1 gene are more sensitive to painful heat sensations near the threshold between anoxious warmth and painful heat. This phenotype is associated with the primary sensory neuron, as polymodal C-fibers were found to be more sensitive to heat in single fiber experiments. Knockout animals are more sensitive to low threshold mechanical stimuli and display an increased thermal and mechanical hyperalgesia in conditions of inflammation. They display a largely decreased pain response induced by osmotic changes particularly in prostaglandin E2-sensitized animals. TREK-1 appears as an important ion channel for polymodal pain perception and as an attractive target for the development of new analgesics.