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Development of Predictive Degradation Models and Determination of Bioequivalence of Pharmaceutical Preparations Contained in the Medical Kits on Board the International Space Station (First Award Fellowship)

Principal Investigator:
Lei Wu, Ph.D.

University of Houston

The environmental factors of space flight may cause accelerated physical-chemical degradation or alterations in the integrity of formulations stored on board space missions for extended periods such as on the International Space Station (ISS) and exploration missions. The degradation of the active pharmaceutical ingredient (API) and adjuvants in addition to alterations of the chemical matrix of the formulation can decrease potency and bioavailability, and increase the risk of toxicity of degraded medications. Therefore, characterizing degradation profiles and developing predictive models of degradation using results from ground-based accelerated stability, bioavailability and toxicity studies is important to ensure safe and effective pharmacotherapeutics for astronauts during long duration space missions.

The overall goal of the proposed research is to develop and validate predictive degradation models for select pharmaceutical preparations contained in the ISS medical kits. A related objective of our research is to examine whether or not integrity and performance of degraded formulations is compromised as indicated by bioequivalence in a ground-based animal model.

The following specific aims will be pursued to accomplish our research goal:

1) Establish a validated HPLC-MS/MS assay method that can be used for the identification and quantifications of API and degradation products of the selected formulations.

2) Characterize degradation profiles of medications received from ISS and compare with existing data from ISS payload and irradiation studies.

3) Conduct ground-based accelerated stability studies with a matching set of medications selected for the above aims to develop predictive degradation models.

4) Estimate bioequivalence between earth-based control and SpaceX returned formulations in a small animal model (rat) to examine integrity and performance of medications stored on board the ISS.

Our proposed study is relevant to the identified “Risk of ineffective and toxic medications due to long term storage” on the Human Research Roadmap and Gap Pharm02: We do not know how long medications may be safe and effective beyond their expiration dates.

NASA Taskbook Entry

Technical Summary

Therapeutic efficacy and safety of pharmaceuticals remain a critical issue for successful NASA medical operations of long-term space missions of International Space Station (ISS) and for future Exploration Medical Capabilities (ExMC). It is known that many pharmaceuticals on Earth are subjected to accelerated degradation when exposed to heat, light and humidity. It is evident that the space flight environment has unique variables, including radiation, excessive vibration, microgravity, and enrichment of CO2, as well as humidity and temperature variations, that affect chemical degradation profiles of medications. Pharmaceutical instability may modify efficacy, safety or patient acceptability. While pharmaceutical product stability is a function of all of its components which include the integrity of the active drug compound, excipients, packaging and product label claims; most stability studies concentrate on quantifying loss of active drug compound from the formulation to estimate loss of potency and shelf life. (Chen XQ, 2006).

The need for a customized protocol and models to evaluate and predict drug degradation in space is obvious from results of a recent investigation led by the mentor (Putcha) on the stability of some of the medications stored on the Space Shuttle and ISS that included antibiotics, pain medications, sleep aids, and others (Du et al., The AAPS Journal, 2011, 55, 1197). Results indicated that 73% of the solid formulations failed to meet the acceptance criteria for active pharmaceutical ingredient (API) and in-flight degradation rates for most drugs were consistently higher than those from matching-lot ground controls. These differences are attributable to the exposure to unique conditions of space environment and therefore, warrant the need for the development of specific methods and protocols for ground-based testing to ensure the stability of the drugs to be used and stored on space missions.

Technologies like BilcareOptimaTM (Ajith Nair, “Time to show your work and that includes packaging," Pharmaceutical Processing, Aug. 2012) developed by Bilcare Research are used by pharmaceutical companies to understand the factors causing the degradation of the drug products (primarily solid dosages) in the prevailing Earth conditions and use the data to model an optimum packaging. BilcareOptimaTM offers a standardized testing program with three distinctive aspects: (1) a forced degradation protocol to quantitatively understand the effect of humidity, light and temperature on the physico-chemical degradation of pharmaceutical solid dosages and determine its critical parameters and moisture threshold values; (2) an algorithm to quantify the sensitivity of each products on a scale of 0 to 10 using results obtained from forced degradation protocol; and (3) a mathematical model to determine optimum moisture and light barrier properties required in primary packing material to protect the product to its shelf life period. However the focus of this model is humidity and light related degradation at different temperatures as those are major environmental factors causing degradation to the drug product on Earth. Hence, there is a need for a modified model which is applicable for the space conditions.

Pharmaceutical stability analysis consists of multiple tests that are described in the United States Pharmacopeia (USP) manual. All methods used for the analysis of pharmaceuticals in this study are implemented strictly and followed from the USP manual. Standard methods for pharmaceutical analysis to determine API content use high/ultra- performance liquid chromatography (HPLC). In an earlier stability study of vitamin B after storage on ISS, chromatograms of different brand of vitamin B were obtained, and the degradation of compounds were indicated by the chromatograms showing a split peak appearing as a notch at the peak tip (Putcha, 2011). LC-MS/MS analysis is an efficient method with high sensitivities for pharmaceutical analysis to determine API content and mass spectral analysis can be used to determine the structure of degradation products for each formulation. Both reference control pharmaceuticals and selected medications obtained from the ISS medical kit will be quantified and compared. Animal models are frequently used in bioavailability and bioequivalence studies of pharmaceuticals. The therapeutic and toxicological potential can be assessed by animal testing. From the previous several reviews on interspecies differences in drug pharmacokinetic, small animals like rats, mice and rabbits are most suitable for determining the mechanism of drug absorption and bioavailability values (Martinez M, 2009; Martignoni M, 2006). In the proposed research project, healthy Sprague-Dawley (SD) rats will be used for the assessment of bioequivalence of reference ground-based control and space flight samples.

Sample preparation and transportation

In the proposed ground-based studies, we will include 4 representative medications obtained from the ISS medical kit packs, including promethazine injection and tablets (for space motion sickness; a racemic mixture of enantiomers), ciprofloxacin tablets (antibiotic) and ibuprofen tablets (Nonsteroidal anti-inflammatory).

A request to transfer all remaining units of selected medications obtained from the ISS medical kit packs that will be returned on all SpaceX missions in the future from the JSC Pharmacy to Pharmacotherapeutics stability research lab in Building 37 will be submitted for approval by Division management. Once received, the medication lots will be stored in the controlled environmental chambers along with reference control pharmaceuticals until pharmaceutical stability analyses are completed. The reference control pharmaceutical preparations from the same manufacturer as those returned from SpaceX will be procured from the contracting pharmacy vendor of JSC pharmacy.

Stability analysis

Chromatographic analysis will be performed using an Agilent 1200 series high/ultra- performance liquid chromatography (HPLC) system and mass spectral analysis will use an Waters LC-MS/MS of 3200 QTRAP® system, which is a hybrid triple quadrupole linear ion trap equipped with a TurboIonSpray ion source. Optimal multiple reaction monitoring (MRM) was used to detect transition ions from a specific precursor ion to product ion [M+H] +. Concentrations of active pharmaceutical ingredient (API) and any detectable degradation products will be measured for each formulation.

Physical and chemical stability indicating parameters that include API concentration will be determined. Pharmaceutical analysis methods described in the Standard Operating Procedures (SOPs) existing in the laboratory (BCI data base) will be implemented. For formulations without standard SOP in the lab, Physical and chemical stability will be determined using methods described in the current United States Pharmacopoeia (USP) and in accordance with the FDA Guidance, Q1A (R2).

Data will be collated and results analyzed for assessment of differences between reference ground and ISS sample lots.  A reference database on degradation characteristics and stability profiles of ISS medications will be compiled.  Where possible, a repository of all samples will be maintained to facilitate repeat analyses in conjunction with SpaceX payloads returned in the future.  Additionally, adequate reference control samples will be also stored for repeat analyses as needed.

In addition to the sample analysis and compilation of results from physical and chemical analyses, predictive stability/shelf-life models will be compiled and validated using commercial off-the-shelf FDA approved shelf-life assessment software (Novatek/Bilcare) using all available data.

In vitro release study

Different groups of formulations will be tested for in vitro release.  The dialysis bag (MWCO: 6,000-8,000) will be used to load selected medications, certain PH, temperature condition with USP diluting solution will be tested and selected to conduct the drug release study. Different dissolution medium, such as 0.1 M HCL and deionized water will be tested and selected as media. The concentrations from the aliquots will be quantified by the LC-MS/MS assay. The cumulative percent of release versus time plot will be obtained.

Bioequivalence studies in rats

Healthy Sprague-Dawley (SD) rats will be used and selected medications from ground-based reference control and ISS groups will be administered to the animals. Animals will be kept in standard animal facilities with free access to food and water, in a temperature and humidity-controlled room with 12 hour on-off light cycle. All experiments would be conducted in accordance with NIH Guidelines for the Care and Use of Animals and with an approved animal protocol from the University of Houston Institutional Animal Care and Use Committee (IACUC).

Pharmacokinetic studies will be conducted. The optimization of dosage of selected medications will be selected for this animal study taking into the consideration of the body surface area and toxicity. The administration routes are oral gavage and tail vein intravenous injection base on the formulations of selected space medications.

The animals will be divided into eight groups (n=5 each), four groups of animals received promethazine injection and tablets, ciprofloxacin tablets and ibuprofen tablets at selected doses, respectively, and the other four groups  will receive medications from ground-based reference control as a reference to calculate the relevant bioavailability. Two hundred μl of the blood samples were withdrawn via tail vein at 15 and 30 minutes and 1, 2, 4, 6, 8, 10, 12 and 24 hours for the oral groups, and at the same time points up to 12 hours for the i.v. group. The blood samples were centrifuged at 16,000 g for 15 minutes immediately to collect the plasma samples to be stored at -80 C until analysis.

Blood samples will be collected and analyzed by validated LC-MS/MS methods. The pharmacokinetics of selected medications in reference ground and ISS sample lots will be characterized and bioavailability will be calculated by using Phoenix PK Modeling software.

Earth Applications

Results of this study will provide important information on identification of degradation profiles for formulations; formulations will be selected based on results from shuttle and ISS operational and payload studies results. Results from the proposed analysis will also serve as preliminary data for the NRA proposal on the development of finger print degradation profiles for predictive model construction using commercial stability modeling programs.  Mass spectra of ISS medications generated can serve as gold standard finger prints for comparison and validation of nondestructive IR and Raman spectral analyses in the future.  These data will also be useful in understanding whether or not degradation patterns of medications onboard space missions is different from those on the ground. The success of development of predictive degradation models will also be useful in identifying alternate methods of preparation and storage of these specific drug products to minimize loss of potency during space missions. The in vivo study of selected drug products will give more information about the impact of drug degradation while the percentage cumulative drug release profiles and relative bioavailability results can provide more information on selection of drug dosage forms.