Purity profile of the indoloquinone anticancer agent EO-9 and chemical stability of EO-9 freeze dried with 2-hydroxypropyl-beta-cyclodextrin.

Two new bladder instillations of the investigational anticancer agent EO-9 containing 2-hydroxypropyl-beta-cyclodextrin (HP beta CD) and the alkalizers sodium bicarbonate (NaHCO3) and tri(hydroxymethyl)aminomethane (Tris) were developed. During the stability study of these freeze-dried products, formation of new degradation products was seen. We have characterized these products by high performance liquid chromatography in combination with photodiode array detection and mass spectrometry. In total, five new degradation products were identified of which three were detected in both freeze-dried products and two only in the freeze-dried product composed of EO-9/HP beta CD/NaHCO3. Furthermore, the purity profile of two lots of EO-9 drug substance was investigated. Five, probably synthetic intermediates were found. However, the amount of total impurities was very small for both lots of drug substance and below acceptable international limits for pharmaceutical use.

Complexation study of the anticancer agent EO-9 with 2-hydroxypropyl-beta-cyclodextrin.

For the development of a bladder instillation of the indoloquinone agent EO-9, use of the complexing agent 2-hydroxypropyl-beta-cyclodextrin (HPbetaCD) was considered. Therefore, a complexation study of EO-9 with HPbetaCD was performed. Complexation was studied in aqueous solution and in solid freeze-dried products. A phase solubility study, UV-visible spectroscopy (UV/VIS), and analysis of the effect of HPbetaD on the stability of EO-9 were performed. With the phase solubility study, a complexation constant (K1:1) of 32.9, a complexation efficiency (CE) of 0.0457, and a utility number (UCD) of 38.3 were calculated. These K1:1 and CE values indicate a weak complex, but the UCD shows that HPbetaCD can be very useful as solubilizer in the desired formulation. Furthermore, a positive effect of HPbetaCD on the chemical stability of EO-9 in solution was seen. Subsequently, complexation in the freeze-dried products was studied more thoroughly using Fourier transform infrared (FTIR), differential scanning calorimetry (DSC), X-ray diffraction (XRD), and scanning electron microscopy (SEM) analyses. HPbetaCD was found to be an excellent pharmaceutical complexing agent for application in formulations for EO-9 bladder instillations. Reconstitution before use of the developed freeze-dried products can be simply accomplished with water for injection.

Assessment of performance of manufacturing procedures in a unit for production of investigational anticancer agents, using a mixed effects analysis.

PURPOSE: To identify the magnitude and sources of variability of a generic, aseptic manufacturing process for experimental anticancer agents employed at our facility, and to estimate the effects on product quality. MATERIALS AND METHODS: In-process and quality control data of all products manufactured according to this generic process (composed of weighing, dissolution, filtration, filling, semi-stoppering and lyophilization) over a 3-year period were retrospectively analyzed using mixed-effects analysis. RESULTS: Variability in the filling process was shown to be marginal and of minor importance for product quality in terms of content and content uniformity. An overall content of 101% was found with batch-to-batch and vial-to-vial variability up to 4.21% and 2.57%, respectively. Estimation of the overall batch failure revealed that structural bias in content and a high batch-to-batch variability in content were the most prominent factors determining batch failure. Furthermore, content and not content uniformity was shown to be most important parameter influencing batch failure. Calculated Process Capability Indices (CpKs) calculated for each product showed that the process is capable of manufacturing products which will routinely comply with the specification of 90-110% for content. However, the CpK values decreased dramatically using the specification of 95-105% as required for approved drug products. CONCLUSION: These results indicate that at the early stage of product development less tight specification limits must be applied to prevent unnecessary batch rejection of investigational agents.

EO-9 bladder instillations: formulation selection based on stability characteristics and in vitro simulation studies.

A bladder instillation of EO-9 (EOquin) is currently used in phase II clinical trials for the treatment of superficial bladder cancer. Three alternative formulations were developed to improve its pharmaceutical properties and clinical acceptability. Freeze-dried products composed of EO-9, 2-hydroxypropyl-beta-cyclodextrin (HPbetaCD), tri(hydroxymethyl) aminomethane (Tris), and sodium bicarbonate (NaHCO(3)) were tested. Selection of one formulation for further development was based on stability studies. These studies comprised stability of the freeze-dried products, stability after reconstitution and dilution and stability during bladder instillation in an experimental set-up. The stability study of the freeze-dried products showed that the formulation composed of EO-9/HPbetaCD/Tris (4/600/1mg/vial) was most stable. After reconstitution and dilution all products were stable for at least 8h. The product composed of EO9/HPbetaCD/NaHCO(3) (4/600/20mg/vial) was the least stable product both as freeze-dried formulation and after reconstitution and dilution. The bladder instillation simulation experiment showed that all products were stable when mixed with urine of pH 8 and unstable in urine of pH 4 and 6. The degradation products formed in urine were EO-5a and EO-9-Cl. Based on these results, the product composed of EO-9/HPbetaCD/Tris (4/600/1mg/vial) was selected for further pharmaceutical development.

Pharmaceutical development, quality control, stability and compatibility of a parenteral lyophilized formulation of the investigational polymer-conjugated platinum antineoplastic agent AP5346.

AP5346 is a low molecular weight polymer-conjugated platinum antineoplastic agent. The lyophilized drug product has completed a phase I clinical trial. In order to guarantee a constant quality of AP5346 pharmaceutical products, quality control and analysis of the drug substance and final product were performed. The identity of AP5346 was confirmed using 1H NMR, 195Pt NMR and IR spectroscopy. Furthermore, the free platinum content, platinum release characteristics, molecular size and size distribution were established. With the selected analytical techniques, AP5346 could be distinguished very well from its polymeric analogues, such as AP5280 and AP5279. Stability experiments revealed that AP5346 final product is stable for 12 months at 5 degrees C, in the dark. For administration to patients, AP5346 final product is reconstituted with 5% w/v dextrose and diluted in infusion containers. To investigate the influence of container materials, the stability of AP5346 after reconstitution and dilution in infusion containers was determined. The infusion containers investigated were composed of glass, polyvinyl chloride (PVC, intraflex) and low density polyethylene (LD-PE, Ecoflac). AP5346 was shown to be stable after reconstitution and dilution with 5% w/v dextrose in these infusion containers for at least 96 h at 2-8 degrees C in the dark and at room temperature with ambient light conditions.