Cystinosin-deficient rats recapitulate the phenotype of nephropathic cystinosis.

The lysosomal storage disease cystinosis is caused by mutations in CTNS, encoding the cystine transporter cystinosin, and in its severest form leads to proximal tubule dysfunction followed by kidney failure. Patients receive the drug-based therapy cysteamine from diagnosis. However, despite long-term treatment, cysteamine only slows the progression of end-stage renal disease. Preclinical testing in cystinotic rodents is required to evaluate new therapies; however, the current models are suboptimal. To solve this problem, we generated a new cystinotic rat model using CRISPR/Cas9-mediated gene editing to disrupt exon 3 of Ctns and measured various parameters over a 12-mo time course. Ctns-/- rats display hallmarks of cystinosis by 3-6 mo of age, as demonstrated by a failure to thrive, excessive thirst and urination, cystine accumulation in tissues, corneal cystine crystals, loss of LDL receptor-related protein 2 in proximal tubules, and immune cell infiltration. High levels of glucose, calcium, albumin, and protein were excreted at 6 mo of age, consistent with the onset of Fanconi syndrome, with a progressive diminution of urine urea and creatinine from 9 mo of age, indicative of chronic kidney disease. Kidney histology and immunohistochemistry showed proximal tubule atrophy and glomerular damage as well as classic "swan neck" lesions. Overall, Ctns-/- rats show a disease progression that more faithfully recapitulates nephropathic cystinosis than existing rodent models. The Ctns-/- rat provides an excellent new rodent model of nephropathic cystinosis that is ideally suited for conducting preclinical drug testing and is a powerful tool to advance cystinosis research.NEW & NOTEWORTHY Animal models of disease are essential to perform preclinical testing of new therapies before they can progress to clinical trials. The cystinosis field has been hampered by a lack of suitable animal models that fully recapitulate the disease. Here, we generated a rat model of cystinosis that closely models the human condition in a timeframe that makes them an excellent model for preclinical drug testing as well as being a powerful tool to advance research.

A non-opioid analgesic implant for sustained post-operative intraperitoneal delivery of lidocaine, characterized using an ovine model.

Appropriate management of post-operative pain is an ongoing challenge in surgical practice. At present, systemic opioid administration is routinely used for analgesia in the post-operative setting. However, due to significant adverse effects and potential for misuse, there is a perceived need for the development of alternative, opioid-sparing treatment modalities. Continuous infusion of local anesthetic into the peritoneum after major abdominal surgery reduces pain and opioid consumption, and enhances recovery from surgery. Here we describe a non-opioid, poly(ethylene-co-vinyl-acetate) intraperitoneal implant for the sustained delivery of local anesthetic following major abdominal surgery. A radio-opaque core had the required mechanical strength to facilitate placement and removal procedures. This core was enclosed by an outer shell containing an evenly dispersed local anesthetic, lidocaine. Sustained release of lidocaine was observed in an ovine model over days and the movement modelled between peritoneal fluid and circulating plasma. While desirably high levels of lidocaine were achieved in the peritoneal space these were several orders of magnitude higher than blood levels, which remained well below toxic levels. A pharmacokinetic model is presented that incorporates in vitro release data to describe lidocaine concentrations in both peritoneal and plasma compartments, predicting similar release to that suggested by lidocaine concentrations remaining in the device after 3 and 7 days in situ. Histological analysis revealed similar inflammatory responses following implantation of the co-extruded implant and a commercially used silicone drain after three days. This non-opioid analgesic implant provides sustained release of lidocaine in an ovine model and is suitable for moving onto first in human trials.

Preformulation studies of l-glutathione: physicochemical properties, degradation kinetics, and in vitro cytotoxicity investigations.

Objectives: l-Glutathione (GSH) is an endogenous tripeptide with super antioxidant properties. In this study, preformulation parameters of GSH and its degradation products were fully investigated.Significance: To date, no experimental preformulation data is available for GSH. Therefore, to the author's knowledge, this is the first study to experimentally determine the preformulation parameters of GSH, which can be considered more reliable for further studies.Methods: An HPLC method for GSH was optimized and validated to accurately quantify the GSH amount in solution, used to investigate GSH's solubility and Log P. Differential Scanning Calorimeter and Thermogravimetric Analyzer were used to evaluate the thermal properties of GSH. Polarized microscope and Fourier-transform Infrared Spectroscopy were used to determine GSH's crystal habits and functional groups, respectively. Forced degradation kinetics and the degradation products were investigated and identified by LC-MS, respectively. GSH's cellular cytotoxicity on fibroblasts was investigated by MTT assay.Results: It was determined that GSH has high aqueous solubility (252.7 mg/mL), low Log P (-3.1), a melting endotherm of 195 °C and decomposition at 210°C, negligible moisture content, and a rectangular/cylindrical-shaped crystalline form. Seven degradation products were identified; one of the major degradation products of GSH under different conditions is first order kinetic oxidation into glutathione disulfide. No cytotoxicity was observed when fibroblasts were treated with GSH (0.005-10.000 mg/mL).Conclusions: Precise preformulation parameters of GSH were obtained, and these are imperative for the development and optimization of advanced GSH formulations.

Population pharmacokinetics of intravenous acetaminophen and its metabolites in major surgical patients.

Intravenous acetaminophen is a commonly used analgesic following surgery. The aims of this study were to determine the population pharmacokinetic profile of intravenous acetaminophen and its metabolites in adult surgical patients and to identify patient characteristics associated with acetaminophen metabolism in the postoperative period. 53 patients were included in the dataset; 28 were men, median age (range) 60 years (33-87), median weight (range) 74 kg (54-129). Patients received 1, 1.5 or 2 g of intravenous acetaminophen every 4-6 h. Plasma and urine samples were collected at various intervals for up to 6 days after surgery. Simultaneous modelling of parent acetaminophen and its metabolites was conducted in Phoenix(®) NLME™ to estimate pharmacokinetic parameters. The population mean estimate (CV%) for central (plasma) volume of distribution of parent acetaminophen (VC) was 13.9 (4.41) L, peripheral (tissue) volume of distribution (VT) was 50.9 (2.96) L, and intercompartmental clearance (Q) was 77.5 (9.29) L/h. The population mean (CV%) metabolic clearances for glucuronidation (CLPG) was 8.92 (3.25) L/h, sulfation (CLPS) was 0.903 (3.47) L/h, and oxidation (CLPO) was 0.533 (7.90) L/h. The population mean (CV%) urinary clearances of parent acetaminophen (CLRP) was 0.137 (5.46) L/h, acetaminophen glucuronide (CLRG) was 3.81 (6.71) L/h, acetaminophen sulfate (CLRS) was 3.13 (4.32) L/h, and acetaminophen cysteine + mercapturate (CLRO) was 3.51 (9.98) L/h. Age was found to be a significant covariate on the formation of acetaminophen glucuronide, and renal function (estimated as creatinine clearance) on the urinary excretion of acetaminophen glucuronide.