Mutant IDH inhibits HNF-4α to block hepatocyte differentiation and promote biliary cancer.

Mutations in isocitrate dehydrogenase 1 (IDH1) and IDH2 are among the most common genetic alterations in intrahepatic cholangiocarcinoma (IHCC), a deadly liver cancer. Mutant IDH proteins in IHCC and other malignancies acquire an abnormal enzymatic activity allowing them to convert α-ketoglutarate (αKG) to 2-hydroxyglutarate (2HG), which inhibits the activity of multiple αKG-dependent dioxygenases, and results in alterations in cell differentiation, survival, and extracellular matrix maturation. However, the molecular pathways by which IDH mutations lead to tumour formation remain unclear. Here we show that mutant IDH blocks liver progenitor cells from undergoing hepatocyte differentiation through the production of 2HG and suppression of HNF-4α, a master regulator of hepatocyte identity and quiescence. Correspondingly, genetically engineered mouse models expressing mutant IDH in the adult liver show an aberrant response to hepatic injury, characterized by HNF-4α silencing, impaired hepatocyte differentiation, and markedly elevated levels of cell proliferation. Moreover, IDH and Kras mutations, genetic alterations that co-exist in a subset of human IHCCs, cooperate to drive the expansion of liver progenitor cells, development of premalignant biliary lesions, and progression to metastatic IHCC. These studies provide a functional link between IDH mutations, hepatic cell fate, and IHCC pathogenesis, and present a novel genetically engineered mouse model of IDH-driven malignancy.

Are two consecutive negative RT-PCR results enough to rule out COVID-19?

The coronavirus disease 2019 (COVID-19) pandemic is redefining the world we live in, and scientists are struggling to find the best severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnostic tool. Routine testing is currently performed using real-time reverse transcription PCR (RT-PCR) of upper or lower respiratory tract secretions. We sought to demonstrate the importance of conducting RT-PCR using deep sampling when initial upper respiratory testing is negative in cases of high index of suspicion for COVID-19. We present the case of a 47-year-old man admitted for fever and bilateral pneumonia diagnosed via chest computed tomographic scan amidst the early peak of the COVID-19 pandemic, suggesting a SARS-CoV-2 infection. Two RT-PCR results from nasopharyngeal swab samples were negative. A bronchoscopy was then performed, and RT-PCR testing on bronchoalveolar lavage samples yielded positive results, confirming the diagnosis of COVID-19 pneumonia. RT-PCR samples of the lower respiratory tract likely contain a higher virus load and thus retain a higher sensitivity for SARS-CoV-2 detection.

Posterolateral corner injuries: Epidemiology, anatomy, biomechanics and diagnosis.

Increased internal and external rotational laxity of the knee may result from a wide range of pathologies in or around the knee. However, the principal cause of increased external rotational laxity is damage to the posterolateral corner (PLC). The aim of the review is to discuss the epidemiology, anatomy, biomechanics and diagnosis of PLC injuries.

Isocitrate Dehydrogenase Mutations Confer Dasatinib Hypersensitivity and SRC Dependence in Intrahepatic Cholangiocarcinoma.

UNLABELLED: Intrahepatic cholangiocarcinoma (ICC) is an aggressive liver bile duct malignancy exhibiting frequent isocitrate dehydrogenase (IDH1/IDH2) mutations. Through a high-throughput drug screen of a large panel of cancer cell lines, including 17 biliary tract cancers, we found that IDH mutant (IDHm) ICC cells demonstrate a striking response to the multikinase inhibitor dasatinib, with the highest sensitivity among 682 solid tumor cell lines. Using unbiased proteomics to capture the activated kinome and CRISPR/Cas9-based genome editing to introduce dasatinib-resistant "gatekeeper" mutant kinases, we identified SRC as a critical dasatinib target in IDHm ICC. Importantly, dasatinib-treated IDHm xenografts exhibited pronounced apoptosis and tumor regression. Our results show that IDHm ICC cells have a unique dependency on SRC and suggest that dasatinib may have therapeutic benefit against IDHm ICC. Moreover, these proteomic and genome-editing strategies provide a systematic and broadly applicable approach to define targets of kinase inhibitors underlying drug responsiveness. SIGNIFICANCE: IDH mutations define a distinct subtype of ICC, a malignancy that is largely refractory to current therapies. Our work demonstrates that IDHm ICC cells are hypersensitive to dasatinib and critically dependent on SRC activity for survival and proliferation, pointing to new therapeutic strategies against these cancers. Cancer Discov; 6(7); 727-39. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 681.

Preparation of ethyl cellulose microcapsules using the complex emulsion method.

In the present work, an investigation was designed to encapsulate a drug with ethyl cellulose using the complex emulsion method. The technique involved the dispersion of the drug in chloroformic solution of ethyl cellulose, followed by emulsification of the polymer solution in water using sodium lauryl sulfate. After continuous stirring for 5 h at room temperature, the solvent evaporates and a rigid polymer film was formed. The optimum surfactant concentration was determined and the effect of different polymer concentrations and core/coat ratios on the quality of the microcapsules produced was investigated. Drug dissolution was retarded as a result of microencapsulation and was controlled by changing core/coat ratios. The dissolution pattern was indicative of the suitability of this technique for the preparation of prolonged action medication.

Preparation of microcapsules using the n-butyl half-ester of PVM/MA coacervate system.

The n-butyl half-ester of the polymer polyvinyl methyl ether maleic anhydride was used for the preparation of microcapsules applying a simple encapsulation process which is less difficult to control than other methods of phase separation encapsulation. The data demonstrated that the release of the drug from the microcapsules can be controlled by changing the ratio of the amount of the drug to shell substance.

Development of implants for sustained release of 5-fluorouracil using low molecular weight biodegradable polymers.

Anticancer drugs have poor efficacy especially against solid tumors that hinder drug penetration into the tumor. Thus, the dose has to be increased, but toxicity is a limiting factor. Local administration of a polymeric biodegradable poly-L-lactic acid (PLA) and poly(L-lactic acid-co-glycolic acid) copolymer (PLGA) implant containing an anticancer drug may be an acceptable method of concentrating the drug near the tumor site. This work sought to synthesize low molecular weight PLA and PLGA by polycondensation to yield polymers with good physical properties to make them suitable for use in implantable therapy. The synthesized polymers were characterized by determining their molecular weight, melting point, and percentage crystallinity using DSC. Fourier transformationinfra red spectrum (FT-IR), nuclear magnetic resonance (NMR) and specific optical rotation measurement were also used to characterize the synthesized polymers. Morphological characteristics were assessed using scanning electron microscopy (SEM). Implants were manufactured using compression (C) and injection molding (IM) and were loaded with 12 mg 5-fluorouracil (5-FU) per 120 mg implant. In vitro release patterns of all implants were assessed in phosphate buffered saline pH 7.4 (PBS 7.4) at 37°C. Factors affecting the release of 5-FU from implants were the polymer species, manufacturing technique, drug particle size, drug concentration, implant dimensions, and coating of the implant. Implants prepared with PLGA had significantly faster release of 5-FU than those prepared with PLA. Those manufactured using compression had significantly faster drug release than those prepared by injection molding. A PLA implant that contained 12 mg 5-FU/120 mg with a diameter of 0.3 cm and that was loaded with a drug particle size smaller than 150 µm and prepared by injection molding and then subsequently coated with PLA had the longest release period of 45 days.

Mycoflora associated with Hyoscyamus muticus growing under an extremely arid desert environment (Aswan region, Egypt).

Hyoscyamus muticus L. (Egyptian henbane) is one of the desert medicinal plants of family Solanaceae. The plant produces pharmaceutically important compounds (tropane alkaloids) as secondary metabolites. In the present study, we describe mycoflora of H. muticus grown in four different locations in Egyptian southern desert (Aswan region): Aswan university campus, Wadi Allaqi down stream part, Aswan airport road, and Sahari city. Eighty-one species and two varieties belonging to 31 genera were isolated from soils surrounding H. muticus plants, the surface of the plants, and inside the plants as endophytic fungi. Aspergillus was the most common genus in all study areas. The highest number of genera and species of fungi were recorded in Aswan university campus followed by Aswan airport road. Fungal diversity analysis revealed that these two locations have higher fungal diversity compared to other two locations. A higher number of fungal species were isolated from rhizosphere soil and rhizoplane than from non-rhizosphere and other plant organs. Endophytic fungi were isolated from all plant parts of H. muticus. Communications between H. muticus plants and fungi under desert conditions both in rhizosphere and inside the plant are deduced.

Preparation of microcapsules from complex coacervation of Gantrez-gelatin. III. Bioavailability of nitrofurantoin microcapsules.

The bioavailability of marketed nitrofurantoin capsules was compared to capsules filled with Gantrez-gelatin nitrofurantoin microcapsules of a core:coat ratio of 1:2. The collective results obtained from the analysis of urine samples of five volunteers indicated that nitrofurantoin microcapsules provided a prolonged release compared with that of the control formulation. Analysis of variance showed statistically significant differences between the control formulation and the capsule filled with the microcapsule. Non-significant intersubject variation was noticed.

Preparation and release kinetics of hydrochlorothiazide from butyl half-ester of PVM/MA microcapsules.

This study describes the principle of a simple rapid method for encapsulating hydrochlorothiazide in butyl half-ester of polyvinyl methyl ether-maleic anhydride copolymer to produce a controlled release dosage form. Unplasticized microcapsules and microcapsules plasticized with Tween 80 and castor oil were prepared. Particle size distribution, flow properties and drug content of microcapsules suggested the suitability of the method for encapsulating a wide variety of materials. The in-vitro release rate was studied as a function of core: coat ratio, type and concentration of plasticizer. Various release mechanisms were considered but no single mechanism can explain all the data completely.

Preparation and evaluation of slow-release pan-coated indomethacin granules.

Granules containing indomethacin crystals are coated with Eudragit solutions of different RL/RS ratios using a pan coating technique. The process is reproducible with regard to drug content, inexpensive and the formed granules were directly compressed into tablets. In vitro release of indomethacin from coated granules, tablets and capsules was studied as a function of different ratios of Eudragit RL/RS in the coating solution. The release of the drug was significantly reduced by the coating process in comparison with a formulation made from uncoated granules, prepared using 10 per cent gelatin solution as a binder. Release data were found to follow a diffusion-controlled model.

Preparation of microcapsules from complex coacervation of Gantrez-gelatin. I. Development of the technique.

Trials to induce complex coacervation between two grades Gantrez-AN polymer (G-AN), and Type A gelatin were made. Physical parameters influencing the coacervation process were studied. Maximum coacervation was attained when the pH of the gelatin solution was at 6.8. Increasing the molecular weight of Gantrez decreased the ratio of combination of both polymers. The ratio for optimum coacervation was 1:4 for Gantrez-AN 119-gelatin system and 2:3 for Gantrez-AN 149-gelatin system with total colloid concentration of 2.5 g per cent w/v in both cases. High stirring speed gave almost spherical uniform coacervates. Recovery of the product as water-insoluble discrete units required the use of formaldehyde and isopropanol for coacervate denaturation and flocculation, respectively.

Preparation of microcapsules from complex coacervation of Gantrez-gelatin. II. In vitro dissolution of nitrofurantoin microcapsules.

Nitrofurantoin crystals were encapsulated in a Gantrez-gelatin complex coacervation system. The encapsulation process was reproducible and inexpensive and the microcapsules were free flowing and directly compressible into tablets. In vitro release of nitrofurantoin from Gantrez-gelatin microcapsules was studied as a function of the core:coat ratio, the molecular weight of Gantrez and the particle size of the microcapsules. The release of the drug was significantly reduced using G149-gelatin microcapsules of core:coat ratio of 1:2. Release data were examined kinetically and were found to follow a diffusion-controlled model. In vitro release of the drug from the microcapsules filled in capsules and compressed into tablets confirmed the efficiency of the encapsulation process for preparing prolonged release formulations.

Preparation and in vitro evaluation of slow release ketoprofen microcapsules formulated into tablets and capsules.

Ketoprofen powder was encapsulated with Eudragit RL/RS polymer solutions in isopropanol-acetone 1:1, using a simple and rapid method. Microcapsules were prepared using Eudragit solutions with different RL/RS ratios. The encapsulation process produces free-flowing microcapsules with good drug content and marked decrease in dissolution rate. The retardation in release profile of ketoprofen from microcapsules was a function of the polymer ratio employed in the encapsulation process. In vitro release of ketoprofen from microcapsules either filled in gelatin capsules or compressed into tablets, using calcium sulphate as diluent, confirmed the efficiency of the encapsulation process for preparing prolonged release medication. A capsule formulation with optimum sustained-release profile was suggested.