Browsing by Subject "Pharmaceutics"
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Item Active transport and CNS delivery of novel tyrosine kinase inhibitors.(2009-03) Chen, YingTyrosine kinase inhibitors (TKIs) are promising agents for specific inhibition of malignant cell growth and metastasis formation. They directly interfere with TK enzymes that are activated in tumor cells and are critical to tumor growth. The success of the first generation TKI, imatinib, in the treatment of CML has led to the broader examination of its application in the treatment of other tumors, such as glioma. However, early studies showed imatinib has difficulty penetrating blood-brain barrier (BBB). One component of the BBB that may limit the delivery of imatinib into the CNS is the drug efflux transporters, such as ABCB1 (p-glycoprotein) and ABCG2 (breast cancer resistance protein). Dasatinib is a second-generation TKI developed to overcome the molecular resistance to imatinib and may be very promising in the treatment of brain tumor. Thus far little information is known about the CNS delivery of dasatinib, including the action of relevant BBB transporters in modulating this delivery. The objectives of this work were to assess the influence of various drug efflux transporters, such as ABCB1 and ABCG2, on the specific delivery of imatinib and dasatinib to CNS and the possibility of improving CNS delivery of imatinib and dasatinib by effective pharmacological inhibition. In in vitro studies, we demonstrated that imatinib is a substrate of Abcg2 by using cellular accumulation and permeability methods. In in vivo studies, we further explored that ABCB1and ABCG2 together play an important role in limiting the CNS delivery of imatinib. Saturation or inhibition of ABCB1 and ABCG2 could effectively improve CNS delivery of imatinib. In vitro evidence pointed that dasatinib is a substrate of ABCB1 and Abcg2. In vivo results revealed that the CNS delivery of dasatinib was low. ABCB1 and ABCG2 could be a factor limiting the CNS delivery of dasatinib. ABCB1 plays a more important role than ABCG2 in effecting the CNS delivery of dasatinib. The use of potent inhibitors for both ABCB1 and ABCG2 can improve dasatinib CNS delivery. These findings provide significant insight into current and new clinical strategies to more effectively use the TKIs for CNS disease treatment and prevention.Item Formulation and delivery of polymeric nanoparticle-assisted vaccine against melanoma(2015-04) Niu, LinPoly (lactide-co-glycolide) (PLGA) nanoparticle (NP) is a widely used biodegradable carrier for drug and vaccine delivery. This thesis focused on the formulation, delivery and efficacy of PLGA NP for its potential application in melanoma immunotherapy. To enable reliable PLGA NP formulation for clinical use such as vaccination, lyophilization is the method of choice to manufacture dry NP dosage form. A major risk of the lyophilization product development for NPs is the irreversible NP aggregation due to freezing and drying stress. Based on real-time imaging, freezing stress could be attributed to freeze-concentration of NPs. Cryo-scanning electron microscopy (cryo-SEM) revealed individual NP separately embedded in the freeze-concentrate interstitial space of the sucrose formulation, leading to corroborative support for the "particle isolation" hypothesis of cryo-protection. Various sphere packing models were investigated to guide the rational design of cryo-/lyo-protectant containing NP formulations. To facilitate precise intradermal delivery of NP formulation for vaccination, microneedle array-mediated administration was utilized to deliver large volume of NPs into the skin. The majority of the infused PLGA NPs were retained locally. A PLGA NP vaccine formulation delivered intradermally elicited robust humoral and cellular immunity. Antigen-loaded NP formulation triggered quicker and stronger high affinity antibody responses compared to the soluble antigen formulation. Vemurafenib, a selective inhibitor of BRAF V600E, induces apoptotic melanoma cell death and remarkable tumor burden reduction. However, drug resistance invariably occurs. Novel TLR7/8 agonists were encapsulated in PLGA particulate formulation as immunostimulatory nanoparticles (ISNP) to boost immune response against drug-resistant melanoma. NP-mediated intracellular delivery contributed to enhanced dendritic cell activation in vitro and antigen-specific CD8+ T cell proliferation in vivo. The prophylactic vaccination using NP-assisted whole tumor cell formulation prolonged the survival of mice challenged with melanoma. To take advantage of the clearance of melanoma antigens by immune system in the context of BRAF inhibition, an ISNP-assisted in situ whole tumor cell vaccination strategy was investigated using BRAF V600E positive mouse SM1 melanoma cells. Despite the suppressed tumor growth, no survival benefit was observed in this therapeutic vaccination model.Item Hygroscopicity of pharmaceutical crystals.(2009-01) Chen, DabingThe active pharmaceutical ingredients (APIs) as well as excipients in a solid dosage form can take up water vapor both during manufacture and subsequent storage of the product. Uptake of unacceptable amount of water can cause adverse effects on physical and chemical stability of APIs and functionality of excipients. It is prudent to select drug candidates with low hygroscopicity to minimize the development risk and time. The objectives of this study are: (i) to investigate the risk in predicting long-term water uptake from short-term water sorption studies, (ii) to understand the thermodynamic and kinetic factors that affect water uptake by pharmaceutical crystals. Automated sorption microbalance (ASM) is often used to determine the hygroscopicity, in which the small sample size and gas purge are believed to accelerate the water sorption process so that equilibrium could be attained in a short time period. However, caution must be exercised when the rates of water vapor diffusion or heat transfer at the solid-vapor interface are not the limiting factors. Four cases are discussed in this thesis, in which ASM failed to predict long-term water uptake. 1) Water vapor was believed to diffuse into the lattice of a metastable crystalline form and induced a polymorphic transformation. The crystallization of the stable form led to a decrease in water content. 2) Adsorbed water formed a surface solution and enhanced the mobility of surface molecules. Nucleation rate of hydrate could be the rate-limiting step. 3) Water sorption induced a crystal to liquid crystal transformation in a surface-active compound, where the latter retains orientational but lacks positional order of molecular packing. 4) The formation of a metastable liquid crystalline phase was kinetically favored for amorphous materials formed in surface-active compounds. The metastable liquid crystalline phase was stable for 3 months when stored under ambient conditions.Item Immunological benefits of a novel polycaprolactone-polyorthoester-based therapeutic vaccine in a mouse model of glioma(2014-08) Grinnen, Karen LynnCancer immunotherapy has led to significant improvement in the survival of patients with previously untreatable malignancies. The use of therapeutic vaccines is a promising form of immunotherapy, but their efficacy remains ambiguous. Much of the difficulty in identifying the optimal formulation and delivery is related to the complicated nature of the immune response, where it is uncertain which aspects would be most effective in destroying cancer cells. In this thesis, a novel polymeric delivery system, involving poly (caprolactone)-co-poly (ortho ester) [PCL-POE], was used to deliver tumor antigens and adjuvants in a controlled manner. We hypothesized that persistent release of tumor antigens from the biodegradable polymer would result in an increase in the number and persistence of anti-tumor lymphocytes in the effector state. To test this hypothesis, vaccines were administered to mice and the time dependent immunological response was evaluated. The polymeric delivery system resulted in an in vitro release profile characterized by a burst release of both antigen and adjuvant followed, in both cases, by a much slower phase of release. We also observed that the slow release provided by the PCL-POE polymer stimulated prolonged maturation of dendritic cells, activation and persistence of anti-OVA antibodies and antigen-specific T cells following a single vaccination. The vaccine system was also tested in a mouse model of glioblastoma multiforme (GBM). We observed a significant, potentially translatable increase in overall survival.Item Improving delivery of molecularly targeted agents to glioma.(2011-06) Agarwal, Sagar SureshTreatment of glioblastoma multiforme is at a crossroads. Promising new molecularly-targeted agents have failed to show any significant clinical benefit. Treatment is particularly challenging since the tumor resides in a tissue that is considered to be a pharmacological and immunological sanctuary due to the presence of the blood-brain barrier. Protective mechanisms at the blood-brain barrier (BBB), such as the endothelial tight junctions and drug efflux transporters, restrict the passage of most large and small molecules into the brain. Limited drug delivery to the tumor is a plausible explanation for the failure of molecularly-targeted therapy in glioma. If therapeutic agents do not reach their target, regardless of their potency, they cannot be effective. The objective of this work was to show that active efflux transporters at BBB restrict delivery of potent molecularly-targeted agents to their targets. More importantly, the aim was to demonstrate that the targets in question are in invasive tumor cells that are left behind after surgery and remain shielded behind an intact blood-brain barrier. The ultimate goal of this endeavor is to improve delivery of molecularly-targeted therapy to the tumor and show that this can translate to enhanced efficacy against this lethal disease. We show that brain distribution of the tyrosine kinase inhibitors, gefitinib, erlotinib and sorafenib, is restricted due to active efflux mediated by p-glycoprotein (P-gp) and the breast cancer resistance protein (BCRP). We further demonstrate that delivery of these drugs to the brain increases dramatically when the two transporters are genetically absent or pharmacologically inhibited. Using a rat xenograft model and a spontaneous mouse model of glioma, we show that the BBB is heterogeneously disrupted in the brain. The blood-brain barrier is disrupted in the tumor core resulting in high tumoral concentrations of erlotinib and dasatinib. However, it is intact in areas immediately adjacent to the tumor, and therefore restricts drug delivery to these sites. Thus, clinical assessment of drug delivery when using drug concentrations in tumor core (the resected tissue) as a guide for the adequacy of drug delivery can be misleading. Furthermore, we show that increasing drug delivery to these areas, by genetic deletion or pharmacological inhibition of P-gp and BCRP, results in a remarkable enhancement in efficacy of the tyrosine kinase inhibitor, dasatinib. Finally, we show that efficacy of dasatinib increases dramatically in tumor bearing transgenic mice, that are deficient in P-gp and BCRP, and consequently, these mice survive for a significantly longer time compared to the wild-type mice. These observations underline that restricted delivery of molecularly-targeted agents to their targets can be a significant determinant of drug efficacy against glioma. In an invasive tumor, such as glioblastoma, it is important to realize that the target resides within the invasive glioma cells, that remain shielded by an intact blood-brain barrier, and evade chemotherapy. Overall, this work highlights the need to develop strategies to improve drug delivery to the invasive tumor in glioma and translate these strategies to the clinic.Item Intranasal targeting of neuropeptides to the central nervous system: evaluation of pharmacokinetics, pharmacodynamics, and a novel vasoconstrictor formulation.(2009-02) Dhuria, Shyeilla VDelivery of therapeutics to the central nervous system (CNS) for the treatment of neurological and psychiatric diseases and disorders is a formidable challenge due to the presence of the blood-brain barrier (BBB). The intranasal route of administration is a noninvasive method to bypass the BBB and rapidly target therapeutics to the CNS by utilizing the olfactory and trigeminal neural pathways connecting the nasal mucosa to the brain and spinal cord. Despite the enormous potential of the intranasal method, it is limited by the low efficiency of delivery to the CNS. The overall objective of this research was to assess targeting of neuropeptides to the CNS following intranasal administration by evaluating pharmacokinetics, pharmacodynamics, and a novel vasoconstrictor formulation. To assess if intranasal administration targeted the neuropeptide, hypocretin-1 (HC), to the CNS, pharmacokinetics and targeting were compared over a two hour period following intranasal and intravenous administration of 10 nmol of 125I-HC to anesthetized rats. Results indicated that intranasal administration resulted in less exposure to the blood and peripheral tissues (~10-fold), similar brain concentrations, increased tissue-to-blood concentration ratios (trigeminal nerve, 14-fold; olfactory bulbs, 9-fold), and greater drug targeting efficiency to the brain (5- to 8-fold) compared to an equivalent intravenous dose. Approximately 80% of the brain exposure after intranasal administration was due to direct transport pathways from the nasal passages. Results from these studies indicated that intranasal administration targets HC to the CNS within 30 minutes of dosing, along direct pathways involving the trigeminal and olfactory nerves. To determine if intranasal administration resulted in pharmacodynamic effects in the CNS, food consumption, water intake, and wheel running activity were monitored following intranasal administration of 100 nmol of HC. Further, HC signaling pathways were investigated to understand the molecular mechanisms underlying the behavioral effects of intranasal HC. Intranasal administration of HC increased food consumption and wheel running activity over the first four hours following dosing, but had no effect on water intake. Intranasal HC activated HC signaling pathways in the diencephalon and in the brainstem, which are brain areas involved in the regulation of appetite and locomotor activity. These findings indicate that intranasal HC reaches the CNS in its biologically active form and at concentrations sufficient to affect HC-mediated behaviors and to activate signaling pathways. To determine if a vasoconstrictor nasal formulation could enhance targeting to the CNS, drug targeting was compared 30 minutes following intranasal administration of 125I-labeled neuropeptides (HC; L-Tyr-D-Arg, D-KTP) in the presence and absence of a vasoconstrictor (phenylephrine, PHE). Results showed that intranasal administration of HC or D-KTP with 1% PHE reduced absorption into the blood, increased deposition in the olfactory epithelium, and increased delivery to the olfactory bulbs. Concentrations in the remaining brain regions and in the trigeminal nerve were reduced in the presence of the vasoconstrictor. The dramatic reduction in the blood concentrations contributed to tissue-to-blood ratios that were increased for HC throughout the brain. For D-KTP, ratios were increased in the olfactory bulbs with 1% PHE, and throughout the brain using a higher concentration of 5% PHE. Use of vasoconstrictor formulations could be used with CNS therapeutics having adverse side effects, where systemic exposure would be limited. The key findings of this research are that intranasal administration targets neuropeptides to the CNS compared to intravenous administration, with brain concentrations that are sufficient to affect CNS-mediated behaviors and signaling pathways. Intranasal administration of HC and other neuropeptides has potential for treating CNS diseases involving the hypocretinergic system, including narcolepsy, Alzheimer's disease, and appetite disorders. In addition, use of a vasoconstrictor nasal formulation could improve intranasal treatments by reducing systemic exposure and enhancing delivery to rostral brain areas, which could be important for CNS therapeutics having adverse systemic effects.Item Mechanisms and analysis of the CNS distribution of cediranib, a molecularly-targeted anti-angiogenic agent.(2011-08) Wang, TianliThe role of vascular endothelial growth factor (VEGF) in brain tumor angiogenesis via stimulation of its receptor (VEGFR) is well established, indicating that the tumor endothelium may be a potential target for brain tumor treatment. However, the current angiogenesis inhibitors used in clinical trials so far have shown limited effects on tumor growth and improvement in survival. Cediranib is an orally available small-molecule kinase inhibitor of all three VEGFR isoforms, with additional activity against PDGFRß and c-KIT. Its broad activities against critical targets, especially the anti-angiogenic activity, make cediranib an attractive option for therapy in central nervous system (CNS) tumors. Cediranib has shown promising anti-angiogenic efficacy in early clinical trials in glioblastoma (GBM), but its anti-tumor mechanisms and its effect on the efficacy of concurrent chemotherapies remain unclear. ATP-binding cassette transporters p-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) have been shown to work in concert to restrict brain penetration of several tyrosine kinase inhibitors. This study investigated the influence of P-gp and BCRP at the blood-brain barrier (BBB) on the CNS penetration of cediranib. Although in vitro studies suggest that both P-gp and BCRP significantly affect intracellular delivery of cediranib, in vivo data indicated that P-gp is a dominant efflux transporter for cediranib and BCRP plays a minor role in limiting transport of cediranib across the BBB. The interaction of cediranib with these brain efflux transporters could limit the antiangiogenic and anti-tumor action in cells in the brain parenchyma and might lead to poor iv outcomes in clinical trials. A more thorough understanding of the mechanisms controlling the delivery of cediranib to its targets will allow more efficacious use of this drug in GBM. To better understand the brain distributional kinetics, simulation strategies were employed to explore the appropriate ways to compare the true brain partitioning among different transporter deficient transgenic mouse groups, which, in general, helps in exploring the contribution of each drug transporter to the brain drug delivery. There are two partial-areas analyses utilized to determine the exit rate constant from the brain, the performances of which were evaluated and compared in the current study. The requirement for accurate determination of the brain-to-plasma ratio of the area under the concentration-time curve (AUC) also warrants the investigation of a Bayesian approach to estimate the variability around the ∞ o AUC and the tissue-to-plasma ∞ o AUC ratio obtained by destructive sampling Since anti-angiogenic agents are commonly used in combination therapy for GBM, the influence of anti-angiogenic therapy on tumor delivery of traditional chemotherapy and molecularly-targeted agents was examined using a xenograft GBM model. It has been shown that restoration of the BBB integrity by cediranib and bevacizumab could decrease the tumor site delivery of both temozolomide and erlotinib, and even the delivery of cediranib itself, which could also be one of the reasons for the limited efficacy of cediranib in clinical trials.Item The metabolism of 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone [NNK] and the enantiomers of 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol [NNAL] in the isolated perfused rat lung system.(2010-08) Maertens, Laura A.4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is a potent carcinogen found specifically in tobacco products. It has been shown to be a lung-specific carcinogen in rodents, and may play a critical role in the formation of lung cancer in smokers. One of the enantiomers of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), a metabolite of NNK, may be important to the selective pulmonary carcinogenicity of NNK. The objective of the current research was to better characterize the pulmonary metabolism of NNK, (S)-NNAL, and (R)-NNAL using the isolated perfused rat lung (IPRL) system to elucidate the mechanisms behind the lung-specific nature of NNK. This research examined metabolite formation, distribution of the metabolites between the perfusate and tissue, the formation of individual DNA adducts in the tissue, and the effects of concentration and the chemopreventive agent PEITC. The results showed that NNK was readily metabolized and DNA adducts were detected in the tissue at the end of the 180 min perfusions. Both an increase in NNK concentration and the co-administration of PEITC were shown to inhibit NNK metabolism. PEITC was also shown to significantly reduce the formation of DNA adducts. The results obtained for the NNK perfusions were in agreement with previously published results. (S)-NNAL and (R)-NNAL were not metabolized as extensively by the lung as NNK. The metabolism of the two enantiomers was similar, which was in contrast to previous in vitro and in vivo results. The only observed difference between the two enantiomers was the formation of low levels of a pyridyloxobutyl (POB)-DNA adduct in the (S)-NNAL perfusions, which indicated reoxidation to NNK. The unexpected results for the NNAL enantiomers may be a result of diffusional barriers to the preformed metabolites that do not exist when the enantiomers are formed from NNK in the tissue. This work showed that the IPRL system was a valid system for examining the pulmonary metabolism of NNK and the formation of DNA adducts, but it may have some limitations for more polar compounds that cannot penetrate the diffusional barriers of the lung and the cells to gain access to the enzymatic sites responsible for metabolism.Item Pharmacokinetics and Pharmacodynamics of Strategically Substituted Agmatines(2022-10) Clements, BenjaminChronic pain remains a major issue affecting patients, with current pharmacotherapy limited to drugs with low efficacy, negative side effects, and/or social stigma. Thus, there is a critical need to develop novel pharmacotherapies that are effective in reducing chronic pain while being safe for long-term use. Agmatine has been extensively shown to reduce chronic pain behaviors in animal models with no cardiac, motor, or neurological adverse effects1. This reduction is due to antagonism of the N-methyl-D-aspartate (NMDA) receptor, specifically at the GluN2B subunit2-4. This specificity allows agmatine to modulate the biological changes underlying chronic pain without the negative side effects observed with complete inhibition of the NMDA receptor. However, agmatine is a polar small molecule and does not efficiently diffuse passively across biological membranes. It has been shown to cross the intestinal epithelium and blood-brain barrier (BBB)5,6, but these distributions are limited, requiring high doses to generating quantifiable disposition. Additionally, the barrier in drug appearance in the CNS necessitates high doses (30-300 mg/kg in animal models) to achieve pharmacological effects. Furthermore, although agmatine has a long half-life in the CNS, its short elimination half-life in plasma restricts efficacy as a systemic therapeutic7. Therefore, our team and collaborators have designed and synthesized a series of agmatine-based prodrugs, the Strategically Substituted Agmatines (SSAs), which are designed to have increased lipophilicity, prolonged plasma half-lives, and equivalent pharmacological activity to agmatine. The central hypothesis of this thesis research is that the SSAs reduce pain behaviors in preclinical models of chronic pain in a manner comparable to agmatine while exhibiting improved pharmacokinetic parameters in rat. From this hypothesis, my objective has been to answer several questions: How can these compounds be measured in plasma and the central nervous system (CNS)? Do strategic substitutions improve pharmacokinetic parameters over agmatine? Do the SSAs distribute more readily to the CNS? Are the SSAs metabolized to agmatine within the CNS? And do the SSAs show improved potency over agmatine following oral administration? To answer these questions, my goals have been to develop techniques to quantify agmatine and the SSAs in complex tissues, determine the pharmacokinetic parameters of agmatine and the SSAs after IV and oral administration, investigate the prodrug activity of the SSAs, and explore the efficacy on chronic pain of these compounds as oral therapeutics. I accomplished these goals by completing the following specific aims: Specific Aim 1: Develop and Validate Analytical Methods to Accurately Quantify Agmatine and the SSAs in Complex MatricesUsing HPLC-MS/MS, a series of tissue preparation methods and chromatographic techniques were developed and validated according to FDA guidance8 to determine the concentration of agmatine, SSA1, SSA2, SSA3, and SSA4 in rat plasma, and quantify agmatine and SSA3 in rat brain and spinal cord. Specific Aim 2: Determine the Plasma Pharmacokinetic Parameters of Agmatine and the SSAs in Rat Following Intravenous and Oral DeliveryI hypothesized that the lipophilic substitutions of the SSAs would increase plasma half-life and volume of distribution over agmatine, as well as improve the oral bioavailability over agmatine. Sprague-Dawley rats with surgically implanted catheters received IV or oral agmatine, SSA1, SSA2, SSA3, or SSA4. Serial blood samples were collected, and plasma was analyzed using HPLC-MS/MS. These plasma concentrations were used to generate individual pharmacokinetic profiles of each drug in individual rats. Specific Aim 3: Assess the Tissue Distribution Profiles of Agmatine and SSA3 in RatI hypothesized that SSA3 would show increased CNS distribution across all tissues compared to agmatine. Furthermore, I hypothesized that SSA3 would be metabolized to agmatine within the CNS. Following IV administration of agmatine or SSA3 via tail-vein in Sprague-Dawley rat, multiple tissues from within the CNS were collected and heat-treated, along with plasma. Agmatine and SSA3 content were assessed using HPLC-MS/MS in each tissue, from which I determined distribution and pharmacokinetic profiles of each compound within individual tissues, such as spinal cord, cortex, ventral tegmental area (VTA), and nucleus accumbens (NAcc). Agmatine content was also assessed in CNS tissues following IV SSA3 and SSA4 to estimate prodrug activity. Specific Aim 4: Evaluate the Efficacy of Agmatine and the SSAs on NMDA-Evoked Responses and Chronic Pain Following Systemic Administration Intrathecal NMDA evokes characteristic behaviors in mice, including biting/scratching and tail-flick hyperalgesia. I hypothesized that the SSAs, due to their increased lipophilicity, would have increased systemic activity at lower doses than agmatine in the NMDA-evoked behavioral model. Additionally, I hypothesized that increased activity at lower doses would be observed in reversal of tactile hypersensitivity in inflammatory pain. I assessed changes in NMDA-evoked behaviors following subcutaneous and oral administration of agmatine and the SSAs to determine the CNS effects of these compounds from non-central delivery. Furthermore, these compounds were tested after oral administration for reduction of tactile hypersensitivity following inflammatory injury via CFA administration.Item Structure-property relationships of solids in pharmaceutical processing(2012-11) Chattoraj, SayantanPharmaceutical development and manufacturing of solid dosage forms is witnessing a seismic shift in the recent years. In contrast to the earlier days when drug development was empirical, now there is a significant emphasis on a more scientific and structured development process, primarily driven by the Quality-by-Design (QbD) initiatives of US Food and Drug Administration (US-FDA). Central to such an approach is the enhanced understanding of solid materials using the concept of Materials Science Tetrahedron (MST) that probes the interplay between four elements, viz., the structure, properties, processing, and performance of materials. In this thesis work, we have investigated the relationships between the structure and those properties of pharmaceutical solids that influence their processing behavior. In all cases, we have used material-sparing approaches to facilitate property assessment using very small sample size of materials, which is a pre-requisite in the early stages of drug development when the availability of materials, drugs in particular, is limited. The influence of solid structure, either at the molecular or bulk powder levels, on crystal plasticity and powder compaction, powder flow, and solid-state amorphization during milling, has been investigated in this study. Through such a systematic evaluation, we have captured the involvement of structure-property correlations within a wide spectrum of relevant processing behaviors of pharmaceutical solids. Such a holistic analysis will be beneficial for addressing both regulatory and scientific issues in drug development.Item Targeted magnetic hyperthermia for lung cancer(2013-01) Sadhukha, TanmoyLung cancer (specifically, non-small cell lung cancer; NSCLC) is the leading cause of cancer-related deaths in the United States. Poor response rates and survival with current treatments clearly indicate the urgent need to develop an effective means to treat NSCLC. Magnetic hyperthermia is a novel non-invasive approach for ablation of lung tumors, and is based on heat generation by magnetic materials, such as superparamagnetic iron oxide (SPIO) nanoparticles, when subjected to an alternating magnetic field. However, inadequate delivery of magnetic nanoparticles to tumor cells can result in sub-lethal temperature change and induce resistance. Additionally, non-targeted delivery of these particles to the healthy tissues can result in toxicity. To overcome these problems, we used aerosol-based, tumor-targeted SPIO nanoparticles to induce highly selective hyperthermia for the treatment of lung cancer.Mechanistic study on the mode of cell kill by magnetic hyperthermia revealed that the extent and mechanism of MH-induced cell kill is dramatically altered with aggregation of SPIO nanoparticles. While well-dispersed SPIO nanoparticles induced apoptosis similar to that observed with conventional hyperthermia, sub-micron size aggregates, induced temperature-dependent autophagy through generation of oxidative stress. Micron size aggregates caused rapid membrane damage and acute cell kill, likely due to physical motion of the aggregates in alternating magnetic field. Overall, this work highlighted the potential for developing highly effective anticancer therapeutics through designed aggregation of SPIO nanoparticles. Cancer stem cells (CSCs) are a sub-population of stem-like cells that are thought to be responsible for tumor drug resistance and relapse. We determined the susceptibility of CSCs to magnetic hyperthermia. Multiple assays for CSCs, including side population phenotype, aldehyde dehydrogenase expression, mammosphere formation and in vivo xenotransplantation, indicated that magnetic hyperthermia reduced or, in some cases, eliminated the CSC sub-population in treated cells. Magnetic hyperthermia demonstrated pleiotropic effects, inducing acute necrosis in some cells while stimulating reactive oxygen species generation and slower cell kill in others. These results suggest the potential for lower rates of tumor recurrence after magnetic hyperthermia compared to conventional cancer therapies. We then studied the effectiveness of inhalation delivery of tumor targeted SPIO nanoparticles for magnetic hyperthermia treatment of lung cancer. We developed EGFR-targeted, inhalable SPIO nanoparticles for magnetic hyperthermia of NSCLC. EGFR targeting resulted in 50% higher concentration of iron oxide in the lungs 1 week post inhalation, when compared to non-targeted SPIO nanoparticles. Magnetic hyperthermia using targeted SPIO nanoparticles resulted in significant inhibition of in vivo tumor growth over a period of one month. Overall, this work demonstrates the potential for developing an effective anticancer treatment modality for the treatment of NSCLC, using targeted magnetic hyperthermia.Item Thermo-activated drug release.(2010-09) Zeng, PengyunInhalation is an effective means of drug administration for treatment of respiratory diseases. Development of a respirable, stimuli-responsive aerosol formulation would further enhance the drug delivery efficiency. In this thesis, it is postulated that a magnetite/lipid formulation stimulated by alternating magnetic fields can be adapted for use as a thermal-activated delivery system to achieve the desired dose and temporal control of drug release. To test this hypothesis, the following specific aims were carried out: (1) Determine the thermal response of superparamagnetic nanoparticles (SPNs) to alternating magnetic fields, (2) Evaluate the release of solute from temperature sensitive aerosol particles, (3) Assess magnetic-activated release of drug from a lipid matrix, and (4) Study the feasibility of magnetic-activated release of solutes with varying polarity from lipid particles. SPNs heat production was found to be quantitatively consistent with theory, and incorporation of SPNs into solid lipid matrices allowed magnetic heating. For the second aim, thermal activation was shown to be necessary and sufficient for the release of encapsulated solute using naturally occurring lipids. For the third aim, stimuli sensitive release of a test solute was demonstrated, which coincided with melting of the matrix. As such, "on-off" drug release was shown to be controlled by a magnetic field. The release was diffusion controlled, such that existing transport theory can be used to guide the development of delivery systems with appropriate release characteristics. Finally, solid lipid particles containing test compounds were characterized and assessed in vitro for thermal and magnetic stimuli release. Surface release and particle erosion mechanism were suggested for nanoparticles containing a hydrophobic compound. For the release from microparticles, magnetic activation was observed in microscopic images. Magnetic activated release was detected for core-lipid shell particles containing a hydrophilic solute, which may be a consequence of physical rotation of the SPNs. A quantitative framework was established to judge the feasibility of developing a magnetic-sensitive drug delivery system that is also respirable. In light of this analysis, significant practical challenges were revealed that make this approach impractical with currently available technology.