A Physiologically-Based, Pharmacokinetic Model for the Clearance of PEGylated Nanomedicines

Physiologically-based pharmacokinetic (PBPK) models are valuable for conducting large-scale in silico experiments to assist in various stages of drug development, including determining optimal dosing strategies, modifying these strategies for special populations, or testing different applications for therapeutics. In doing so, PBPK serves as an alternative for time-consuming, costly, and potentially unethical in vivo experiments or clinical trials: it can be used to predict and optimize the success of proposed treatments using known (well-determined physiologically) or learned (through experiments or data-driven analyses) kinetics of the system mimicking complex human physiology. In turn, PBPK modeling can enable more efficient design and optimization of in vivo experiments, thus accelerating pre-clinical screening and development of therapeutics. In this thesis, I discuss the application of PBPK modeling to an important problem in the medical community – the accelerated clearance of PEGylated drugs in the presence of anti-PEG antibodies (APA). For example, Krystexxa, a PEGylated uricase used to treat severe chronic gout, is ineffective in approximately half of patients, and 5% of the patients experience severe hypersensitivity, including anaphylaxis, as a result of APA. Here I present a multi-compartment PBPK model to accurately predict the biodistribution and clearance behavior of PEGylated liposomal drug carriers, and validate the model predictions against drug biodistribution data obtained via PET/CT technology. I then analyze the model to identify mechanistic behaviors underlying APA-mediated accelerated clearance using parameter estimation and optimization techniques, namely Latin Hypercube Sampling. Finally, I demonstrate that pre-injection with a high molecular weight free PEG effectively restores plasma Krystexxa levels to a naïve-like biodistribution in mice with clinical levels of APA, suggesting a potential strategy for mitigating the accelerated blood clearance (ABC) effect in the clinic. In considering such a translation, PBPK modeling is a powerful tool that holds the potential to suggest further optimization of our strategy to mitigate ABC.Doctor of Philosoph

Dermal Lymphatic Capillaries Do Not Obey Murray's Law

Lymphatic vessels serve as a major conduit for the transport of interstitial fluid, immune cells, lipids and drugs. Therefore, increased knowledge about their development and function is relevant to clinical issues ranging from chronic inflammation and edema, to cancer metastasis to targeted drug delivery. Murray's Law is a widely-applied branching rule upheld in diverse circulatory systems including leaf venation, sponge canals, and various human organs for optimal fluid transport. Considering the unique and diverse functions of lymphatic fluid transport, we specifically address the branching of developing lymphatic capillaries, and the flow of lymph through these vessels. Using an empirically-generated dataset from wild type and genetic lymphatic insufficiency mouse models we confirmed that branching blood capillaries consistently follow Murray's Law. However surprisingly, we found that the optimization law for lymphatic vessels follows a different pattern, namely a Murray's Law exponent of ~1.45. In this case, the daughter vessels are smaller relative to the parent than would be predicted by the hypothesized radius-cubed law for impermeable vessels. By implementing a computational fluid dynamics model, we further examined the extent to which the assumptions of Murray's Law were violated. We found that the flow profiles were predominantly parabolic and reasonably followed the assumptions of Murray's Law. These data suggest an alternate hypothesis for optimization of the branching structure of the lymphatic system, which may have bearing on the unique physiological functions of lymphatics compared to the blood vascular system. Thus, it may be the case that the lymphatic branching structure is optimized to enhance lymph mixing, particle exchange, or immune cell transport, which are particularly germane to the use of lymphatics as drug delivery routes

Changes in cholesterol homeostasis and acute phase response link pulmonary exposure to multi-walled carbon nanotubes to risk of cardiovascular disease

AbstractAdverse lung effects following pulmonary exposure to multi-walled carbon nanotubes (MWCNTs) are well documented in rodents. However, systemic effects are less understood. Epidemiological studies have shown increased cardiovascular disease risk after pulmonary exposure to airborne particles, which has led to concerns that inhalation exposure to MWCNTs might pose similar risks.We analyzed parameters related to cardiovascular disease, including plasma acute phase response (APR) proteins and plasma lipids, in female C57BL/6 mice exposed to a single intratracheal instillation of 0, 18, 54 or 162μg/mouse of small, entangled (CNTSmall, 0.8±0.1μm long) or large, thick MWCNTs (CNTLarge, 4±0.4μm long). Liver tissues and plasma were harvested 1, 3 and 28days post-exposure. In addition, global hepatic gene expression, hepatic cholesterol content and liver histology were used to assess hepatic effects.The two MWCNTs induced similar systemic responses despite their different physicochemical properties. APR proteins SAA3 and haptoglobin, plasma total cholesterol and low-density/very low-density lipoprotein were significantly increased following exposure to either MWCNTs. Plasma SAA3 levels correlated strongly with pulmonary Saa3 levels. Analysis of global gene expression revealed perturbation of the same biological processes and pathways in liver, including the HMG-CoA reductase pathway. Both MWCNTs induced similar histological hepatic changes, with a tendency towards greater response following CNTLarge exposure.Overall, we show that pulmonary exposure to two different MWCNTs induces similar systemic and hepatic responses, including changes in plasma APR, lipid composition, hepatic gene expression and liver morphology. The results link pulmonary exposure to MWCNTs with risk of cardiovascular disease

MWCNTs of different physicochemical properties cause similar inflammatory responses, but differences in transcriptional and histological markers of fibrosis in mouse lungs

Multi-walled carbon nanotubes (MWCNTs) are extensively produced and used in composite materials and electronic applications, thus increasing risk of worker and consumer exposure. MWCNTs are an inhomogeneous group of nanomaterials that come in various lengths, shapes and with different metal contaminations, which makes hazard evaluation difficult. However, several studies suggest that length plays an important role in the toxicity induced by MWCNTs. How the length influences toxicity at the molecular level is yet to be characterized. Female C57BL/6 mice were exposed by single intratracheal instillation to 18, 54 or 162 µg/mouse of a short MWCNT (NRCWE-026, 847±102 nm in length) or long MWCNT (NM-401, 4048±366 nm in length). The two MWCNTs were extensively characterized. Lung tissues were harvested 24 h, 3 d and 28 d after exposure. We employed DNA microarrays, bronchoalveolar lavage fluid analysis, comet assay and dichlorodihydrofluorescein assay in order to profile the pulmonary responses. Bioinformatics tools were then applied to compare and contrast the expression profiles and to build a length dependent property-response matrix for gene-by-gene comparison. The toxicogenomic analysis of the global mRNA changes after exposure to the short, entangled NRCWE-026 or the longer, stiffer NM-401 showed high degree of similarities. The toxicity of both MWCNTs was driven by strong inflammatory and acute phase responses, which peaked at day 3 and was observed both in bronchoalveolar lavage cell influx and in gene expression profiles. The inflammatory response was sustained at post-exposure day 28. Also, at the sub-chronic level, we identified a sub-set of 14 fibrosis related genes that were uniquely differentially regulated after exposure to NM-401. Acellular ROS production occurred almost exclusively with NRCWE-026, however the longer NM-401 induced in vivo DNA strand breaks and differential regulation of genes involved in free radical scavenging more readily than NRCWE-026. Our results indicate that the global mRNA response after exposure to MWCNTs is length independent at the acute time points, but that fibrosis may be length dependent sub-chronic end point.JRC.H.6-Digital Earth and Reference Dat

Transcriptomic Analysis Reveals Novel Mechanistic Insight into Murine Biological Responses to Multi-Walled Carbon Nanotubes in Lungs and Cultured Lung Epithelial Cells

There is great interest in substituting animal work with in vitro experimentation in human health risk assessment; however, there are only few comparisons of in vitro and in vivo biological responses to engineered nanomaterials. We used high-content genomics tools to compare in vivo pulmonary responses of multiwalled carbon nanotubes (MWCNT) to those in vitro in cultured lung epithelial cells (FE1) at the global transcriptomic level. Primary size, surface area and other properties of MWCNT- XNRI -7 (Mitsui7) were characterized using DLS, SEM and TEM. Mice were exposed via a single intratracheal instillation to 18, 54, or 162 μg of Mitsui7/mouse. FE1 cells were incubated with 12.5, 25 and 100 μg/ml of Mitsui7. Tissue and cell samples were collected at 24 hours post-exposure. DNA microarrays were employed to establish mechanistic differences and similarities between the two models. Microarray results were confirmed using gene-specific RT-qPCR. Bronchoalveolar lavage (BAL) fluid was assessed for indications of inflammation in vivo. A strong dose-dependent activation of acute phase and inflammation response was observed in mouse lungs reflective mainly of an inflammatory response as observed in BAL. In vitro, a wide variety of core cellular functions were affected including transcription, cell cycle, and cellular growth and proliferation. Oxidative stress, fibrosis and inflammation processes were altered in both models. Although there were similarities observed between the two models at the pathway-level, the specific genes altered under these pathways were different, suggesting that the underlying mechanisms of responses are different in cells in culture and the lung tissue. Our results suggest that careful consideration should be given in selecting relevant endpoints when substituting animal with in vitro testing

Modeling the Dynamics of Cancerous Tumors in Vivo

A key area of interest in cancer research is understanding tumor dynamics, so that tumor growth can be predicted more easily and treated more reliably. We explored in vivo tumor dynamics starting from simple exponential and logistic growth and progressing to more general models. We then applied these models to analyze the growth of breast cancer, liver cancer, and two types of neurological cancer using five data sets. We found that exponential growth gave the best fit to breast and liver cancers, while surface growth (a 2/3's power law) gave the best fit to neurological cancers.Honors thesi