Three-Dimensional NMR Spectroscopy of Fluorinated Pharmaceutical Solids under Ultrafast Magic Angle Spinning.

High-resolution solid-state analysis of multicomponent molecular systems, e.g., pharmaceutical formulations, is a great challenge. Solid-state nuclear magnetic resonance (ssNMR) spectroscopy plays a critical role in the characterization of solid dosage forms due to its capabilities of chemical identification, quantification, and structural elucidation at a molecular level. However, the low NMR sensitivity as well as the high spectral complexity and low drug loading of multicomponent products hinder an in-depth investigation of the active pharmaceutical ingredient (API) at the natural isotopic abundance. Herein, we developed two new three-dimensional (3D) ssNMR methods, including 1H-19F-1H and 19F-19F-1H correlations and successfully applied them to characterize a fluorinated drug molecule, aprepitant, and its commercial nanoparticulate formulation EMEND (Merck & Co, Inc., Kenilworth, NJ, USA). These 1H-detection methods utilize the significantly enhanced sensitivity and resolution of 1H and 19F afforded by 60 kHz ultrafast magic angle spinning (MAS) and enable the analysis of milligram samples. The 3D techniques simultaneously provide homonuclear 1H-1H and 19F-19F, and heteronuclear 1H-19F correlations of the crystalline aprepitant without interferences from other pharmaceutical components in the drug product. Moreover, our results demonstrate that 19F is a highly sensitive spin for probing molecular details of fluorinated drug substances in solid formulations, due to its high isotopic abundance, large gyromagnetic ratio, and absence of signal interference from pharmaceutical excipients, as well as for characterizing structural properties of a broad range of fluorine-containing materials.

Audiogenic seizure activity following HSV-1 GAD65 sense or antisense injection into inferior colliculus of Long-Evans rat.

Herpes virus technology involving manipulation of GAD65 was used to study effects on audiogenic seizures (AGS). Audiogenic seizure behaviors were examined following injections of replication-defective herpes simplex virus (HSV-1) vectors incorporating sense or antisense toward GAD65 along with 10% lac-Z into the central nucleus of inferior colliculus (CNIC) of Long-Evans rats. In seizure-sensitive animals developmentally primed by intense sound exposure, injection of GAD65 in the sense orientation increased wild running latencies and reduced incidence of clonus compared with lac-Z only, unoperated, and vehicle seizure groups. In contrast, infection of CNIC with GAD65 antisense virus resulted in 100% incidence of wild running and clonus behaviors in AGS animals. Unprimed animals not operated continued to show uniform absence of seizure activity. Administration of GAD65 antisense virus into CNIC produced novel wild running and clonus behaviors in some unprimed animals. Staining for ß-galactosidase in all vector animals revealed no differences in pattern or numbers of immunoreactive cells at injection sites. Qualitatively, typical small and medium multipolar/stellate and medium fusiform neurons appeared in the CNIC of vector animals. These results demonstrate that HSV-1 vector constructs implanted into the CNIC can predictably influence incidence and severity of AGS and suggest that viral vectors can be useful in studying GABA mechanisms with potential for therapeutic application in epilepsy. This article is part of a Special Issue entitled "Genetic and Reflex Epilepsies, Audiogenic Seizures and Strains: From Experimental Models to the Clinic".

Nanomedicines in the treatment of emesis during chemotherapy: focus on aprepitant.

Aprepitant, a selective high-affinity antagonist of human substance P/neurokinin 1 (NK1) receptors, is the active ingredient of EMEND which has recently been approved by the FDA for the prevention of chemotherapy-induced nausea and vomiting (CINV). Aprepitant undergoes extensive metabolism, primarily via CYP3A4 mediated oxidation. It is eliminated primarily by metabolism and is not renally excreted. The apparent terminal half-life in humans ranged from 9 to 13 hours. Early development studies led to the development of a nanoparticle formulation to enhance exposure and minimize food effects. Two large randomized trials accruing 1099 patients studied the effect in patients receiving cisplatin of adding aprepitant to ondansetron and dexamethasone on day 1 then to dexamethasone on days 2 and 3 to control delayed emesis. The complete response of no vomiting and no rescue medication overall from days 1 to 5 improved from 48% to 68% (p<0.001), a 13% improvement in acute emesis but a 21% improvement in delayed emesis with the improvement from 51% to 72% (p<0.001). Similarly, 866 patients treated with cyclophosphamide plus either doxorubicin or epirubicin, received either ondansetron, dexamethasone, and aprepitant on day 1 followed by aprepitant on days 2 and 3 or ondansetron and dexamethasone on day 1 and dexamethasone on days 2 and 3. The overall complete response rate over 5 days was better for the aprepitant group 50.8% vs 42.5% (p=0.015). Complete responses were reported in more patients taking aprepitant in both the acute (76% vs 69%, p=0.034) and delayed (55% vs 49%, p=0.064) phases of vomiting. There were no clinically relevant differences in toxicity by adding aprepitant and improvements in the quality of life of patients on chemotherapy were recorded.

Characterization of crystalline drug nanoparticles using atomic force microscopy and complementary techniques.

PURPOSE: The purpose of this work was to image crystalline drug nanoparticles from a liquid dispersion and in a solid dosage form for the determination of size, shape, and distribution. METHODS: Crystalline drug nanoparticles were adsorbed from a colloidal dispersion on glass for atomic force microscopy (AFM) imaging. Nanoparticles that were spray coated onto a host bead were exposed by ultramicrotomy for scanning electron microscopy and AFM examination. RESULTS: The adsorbed drug nanoparticles were measured by AFM to have a mean diameter of 95 nm and an average aspect ratio of 1.3. Nanoparticles observed in the solid dosage form had a size and shape similar to drug nanoparticles in the dispersion. Particle size distribution from AFM measurement agreed well with data from field emission scanning electron microscopy, static light scattering, and X-ray powder diffraction. CONCLUSION: AFM is demonstrated to be a valuable tool in visualization and quantification of drug nanoparticle crystals in formulations. In addition to accurate size measurement, AFM readily provides shape and structural information of nanoparticles, which cannot be obtained by light scattering. Ultramicrotomy is a good sample preparation method to expose the interior of solid dosage forms with minimal structural alteration for microscopic examination.

Characterization of an extemporaneous liquid formulation of lisinopril.

The stability of lisinopril in an extemporaneously prepared suspension stored at or below 25 degrees C for 28 days under ambient light exposure was studied. A formulation of 1-mg/mL oral suspension was prepared from commercially available 20-mg lisinopril tablets, using Bicitra and Ora-Sweet SF as the compounding vehicles to make a final volume of 200 mL. Individual samples, stored in 8-oz amber polyethylene terephthalate bottles, were used for each test performed. All samples were stored at 25 degrees C. Appropriateness of the extemporaneous preparation method was performed by shaking three lots of each suspension for 30, 60, and 90 seconds. To test the robustness and reproducibility of the method, two chemists prepared the suspensions from the same three lots of lisinopril tablets. Chemical and physical stability were established by analyzing duplicate samples at time zero and after one, two, four, and six weeks. The solubility of lisinopril was tested from suspensions stored for four weeks. In-use stability was also examined over four weeks. Photochemical stability was examined by exposing three batches of the suspension to maximum light stress in accordance with the International Conference on Harmonization. Antimicrobial-effectiveness testing was also conducted with freshly prepared suspensions and suspensions stored for six weeks. The preparation method used was appropriate and effective. Lisinopril is fully dissolved in the suspension matrix. Satisfactory chemical, physical, and microbiological results were obtained after the suspensions were stored for six weeks at 25 degrees C and 35% relative humidity. Lisinopril suspensions extemporaneously prepared from tablets are stable for at least four weeks when stored at or below 25 degrees C under ambient light exposure.