Small molecule therapy has been widely used for the treatment of a variety of diseases. Small molecule drugs can be easily administered to patients, and are advantageous in that they can cross the ...blood-brain barrier, do not cause autoimmune responses, and have lower manufacturing costs. In this chapter, we focus on different strategies and methods for small molecule drug development as it applies to Fabry disease. The steps involved in developing an appropriate high throughput screen to identify activators and inhibitors of alpha galactosidase A are outlined. Assay development includes optimization of the assay pH, time course, enzyme and substrate concentration and the amount of sodium taurocholate used. The assay must then be validated and confirmed using additional screens. The optimized screens can be used to identify novel lead compounds that can serve as new starting points for drug development for Fabry disease.
A uniquely attenuated disruption of cholesterol homeostasis has been characterized in certain Niemann-Pick, type C (NP-C) fibroblasts. Uptake of LDL-cholesterol by cultured fibroblasts derived from ...two clinically affected brothers with this variant biochemical phenotype led to less intracellular accumulation of unesterified cholesterol than found in other typical cell lines. This limited cholesterol lipidosis in the variant NP-C cells reflected cholesterol processing errors that differed from the cellular lesions in classical NP-C cells in the following ways: (1) a more limited intracellular distribution of the excessive unesterified cholesterol; (2) shorter and more transient delays in the induction of cholesterol-mediated homeostatic responses; and (3) more efficient intracellular transport of exogenously derived cholesterol to the plasma membrane and the endoplasmic reticulum. Activation of acyl-CoA cholesterol acyltransferase (ACAT) was greater than 100-fold in both control and NP-C fibroblasts when cell cultures were preconditioned with 25-hydroxycholesterol, but the subsequent esterification of exogenous non-lipoprotein 3Hcholesterol remained deficient in all NP-C cells. In the variant NP-C cells conditioned with the oxysterol, this esterification of exogenous 3Hcholesterol was less affected than in classical NP-C cultures. The NP-C mutation affects a broad spectrum of metabolic responses related to the processing of exogenously derived cholesterol. Among this pleiotropic array of deficient responses, retarded intracellular cholesterol transport appears most closely linked to the primary mutation. This conclusion is supported by two current observations: (1) the degree to which sterol transport is affected in mutant cells in turn reflects the extent to which cholesterol-homeostatic responses are compromised; and (2) sterol transport remains deficient despite concurrent normal activation of other cellular responses, such as cholesterol esterification.
NPC disease is an autosomal recessive neurovisceral storage disorder. A pleiotropic array of secondary enzymatic and storage abnormalities has in the past obscured a cohesive understanding of the ...underlying metabolic basis of this disorder. Recent findings, reviewed in this report, demonstrate that NPC disease is a cholesterol lipidosis resulting from defective intracellular cholesterol transport. The sequence of cellular events characteristic of NPC is 1) deficient intracellular transport of exogenously derived cholesterol resulting in retarded induction of cellular cholesterol homeostatic regulation; 2) accumulation of cholesterol in lysosomes; and 3) secondary cellular effects. Retarded esterification of exogenous cholesterol and accumulation of unesterified cholesterol in lysosomes is tightly coupled to the primary defect and serves as the basis for biochemical diagnosis of NPC.
Colorectal cancer is a leading cause of death. Colonoscopy is the criterion standard for detection and removal of precancerous lesions and has been shown to reduce mortality. The polyp miss rate ...during colonoscopies is 22% to 28%. DEEP DEtection of Elusive Polyps (DEEP2) is a new polyp detection system based on deep learning that alerts the operator in real time to the presence and location of polyps. The primary outcome was the performance of DEEP2 on the detection of elusive polyps.
The DEEP2 system was trained on 3611 hours of colonoscopy videos derived from 2 sources and was validated on a set comprising 1393 hours from a third unrelated source. Ground truth labeling was provided by offline gastroenterologist annotators who were able to watch the video in slow motion and pause and rewind as required. To assess applicability, stability, and user experience and to obtain some preliminary data on performance in a real-life scenario, a preliminary prospective clinical validation study was performed comprising 100 procedures.
DEEP2 achieved a sensitivity of 97.1% at 4.6 false alarms per video for all polyps and of 88.5% and 84.9% for polyps in the field of view for less than 5 and 2 seconds, respectively. DEEP2 was able to detect polyps not seen by live real-time endoscopists or offline annotators in an average of .22 polyps per sequence. In the clinical validation study, the system detected an average of .89 additional polyps per procedure. No adverse events occurred.
DEEP2 has a high sensitivity for polyp detection and was effective in increasing the detection of polyps both in colonoscopy videos and in real procedures with a low number of false alarms. (Clinical trial registration number: NCT04693078.)
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Clinical studies have shown that negative symptoms of schizophrenia unresponsive to antipsychotic given alone can improve after augmentation with SSRI antidepressant. Laboratory investigations into ...the mechanism of this synergism showed that co-administration of SSRI and antipsychotic produces changes in GABAA receptor and related systems, which differ from the effects of each drug alone. To examine the clinical relevance of these findings, the current study examined the effects of SSRI augmentation treatment on GABAA receptor and related systems in schizophrenia patients. Schizophrenia patients with high levels of negative symptoms unresponsive to antipsychotic treatment received add-on fluvoxamine (100 mg/d). Blood was taken before and 1, 3 and 6 wk after adding fluvoxamine and peripheral mononuclear cells (PMC) isolated. RNA encoding for GABAAβ3, 5-HT2A, and 5-HT7 receptors, PKCβ2, and brain-derived neurotrophic factor (BDNF) was assayed with real-time RT-PCR. Plasma BDNF protein was assayed using ELISA. Clinical symptoms were assessed with validated rating scales. We found significant increase in mRNA encoding for GABAAβ3 and 5-HT2A, 5-HT7 receptors and BDNF and a reduction in PKCβ2 mRNA. Plasma BDNF protein concentrations were increased. There were significant correlations among the genes. Clinical symptoms improved significantly. mRNA expression of PKCβ2, 5-HT2A and 5-HT7 showed significant associations with clinical symptoms. Combined SSRI+antipsychotic treatment is associated with changes in GABAA receptor and in related signalling systems in patients. These changes may be part of the mechanism of clinically effective drug action and may prove to be biomarkers of pharmacological response.
In this study, we evaluated the safety and efficacy of a personalized mode of treatment for Crohn's disease (CD) by oral administration of Alequel an extract of autologous colonic proteins.
...Thirty-one patients with moderate to severe CD were enrolled in a 27-wk randomized, double-blind, placebo-controlled trial. Patients were randomized to receive either a placebo or the study drug prepared from autologous colonic extract.
Oral administration of autologous colonic proteins resulted in clinical remission (58% vs 29%; 46.6% vs 26.6%, using an intention to treat analysis, p= NS), clinical response (67% vs 43%; 53.3% vs 40%, using an intention to treat analysis, p= NS) and improved quality of life (Inflammatory Bowel Disease Questionnaire score improvement 43%vs 12%) in the drug study group, compared to placebo group. No treatment-related adverse events were noted. Only in the study-drug-treated cohort who achieved clinical remission (DR), there was a decreased number of subject-specific, antigen-directed, IFNgamma spot-forming colonies. DR subjects had a lower initial C-reactive protein level than DNOR or placebo subjects, an increased percentage of peripheral blood nature killer T cells, and an increased CD4+/CD8+ T-cell ratio throughout the period of drug administration.
Oral administration of Alequel is a safe method for treatment of patients with moderate to severe CD, and its efficacy needs to be proven. Several markers may be applicable as surrogate markers for the clinical effect.
The phenomenon of Levantine nanism in the Mediterranean Sea has so far been described in invertebrates and fish. We explored the possibility that it would also apply to marine mammals. To that end, ...we compared total body length (TBL) and skull condylobasal length (CBL) of adult common bottlenose dolphinsTursiops truncatuscollected along the Israeli coastline (representing the Levantine subpopulation) to those of specimens collected along the shores of western Mediterranean seas. Significant differences were found between mean (±SD) CBL values of 40 skulls from Israel and a pooled sample of 40 skulls from the Adriatic, Tyrrhenian, Ligurian and Balearic Seas (49.70 ± 1.87 and 52.18 ± 1.47 cm, respectively, p < 0.001). The mean (±SD) TBL of 26 Israeli animals were significantly smaller than those of 28 animals from the Spanish Mediterranean coast and 36 animals from the French Mediterranean coast (272 ± 18.0, 317.3 ± 16.1 and 313.4 ± 14.8 cm, respectively, p < 0.001). The results clearly demonstrate that animals of the Levantine subpopulation are significantly smaller than those residing in the west. A difference of ~16% for TBL between populations fits the range of within-species dwarf morphs in cetaceans and, when translated into volume and mass, also fits the definition of Levantine nanism.
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