2013 UMKC Dissertations - Freely Available Online
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This community contains the collections of dissertations submitted electronically to the School of Graduate Studies by doctoral degree candidates at the University of Missouri-Kansas City. The items in this collection are dissertations that are available to the general public.
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Item Development of Pentablock Copolymer Based Formulations for the Sustained Delivery of Protein Therapeutics in the Treatment of Posterior Segment Ocular Diseases(2013) Patel, Sulabh P.; Mitra, Ashim K., 1954-We have successfully synthesized pentablock (PB) copolymers comprised of various FDA approved polymer blocks such as polyethylene glycol (PEG), polycaprolactone (PCL), polylactic acid (PLA) and polyglycolic acid (PGA). PB copolymers with different composition, molecular weights and block arrangements were utilized to develop protein-embedded thermosensitive gels or nanoparticles (NPs) for sustained delivery in the treatment of posterior segment ocular diseases. In order to eliminate the burst release effect, we have studied PB composite formulation comprised of protein-encapsulated PB NPs dispersed in PB thermosensitive gel. The composite formulation eliminated burst release effect and exhibited nearly zero-order protein release for significantly longer durations. In this research work, we have utilized various model proteins (lysozyme, IgG-Fab, IgG, BSA, and catalase) and therapeutic proteins (octreotide, insulin and bevacizumab) to optimize the formulation. We have synthesized various triblock (TB) (PCL-PEG-PCL, B-A-B) and PB (PLA-PCL-PEG-PCL-PLA (C-B-A-B-C) and PEG-PCL-PLA-PCL-PEG (A-B-C-B-A)) copolymers based thermosensitive gelling polymers. We have observed distinct effect of block arrangement and molecular weights of block copolymers on the sol-gel transition and on the kinematic viscosity of aqueous solutions. PB copolymers with A-B-C-B-A block arrangement exhibited significantly lower viscosity relative to TB copolymers or other types of PB copolymers (C-B-A-B-C). The difference in viscosity and sol-gel transition behavior has been explained by two different processes of micellization for A-B-C-B-A and B-A-B, or C-B-A-B-C types of copolymers. Moreover, a PB copolymer based formulation sustained the release of IgG up to ~20 days, which is significantly longer relative to TB copolymers based formulations. In order to sustain release for longer duration, we have synthesized various PB copolymers (PLA-PCL-PEG-PCL-PLA and PGA-PCL-PEG-PCL-PGA) with high molecular weight and utilized them for the fabrication of protein-encapsulated NPs. We observed a significant effect of the presence of PLA or PGA on entrapment efficiency (EE), drug loading (DL) and in vitro release behavior. This may be due to the fact that PB copolymers exhibited significantly reduced crystallinity relative to TB copolymers. In addition, we have successfully optimized NP preparation methods to achieve maximum possible DL. This achievement allowed the loading of a large amount of drug which can last for ~6 month in a limited injection volume (100 μL). The optimized methods were successfully utilized to encapsulate a wide variety of peptides and proteins with molecular weights ranging from 1 - 237 kDa in PB NPs. PB NPs alone exhibited significant burst release in the first few days of release study. However, a composite formulation comprised of protein-encapsulated PB-NPs prepared with optimized method and optimized PB copolymers (PB copolymers for NPs and thermosensitive gel) exhibited protein release for significantly longer duration of time (~6 months) with nearly zero-order release rate. We have evaluated the structural integrity of released protein at different time intervals by CD spectroscopy. Moreover, biological activity of bevacizumab was evaluated by cell proliferation and cell migration assays. Enzymatic activity of lysozyme and catalase were confirmed with their respective enzymatic assays. Our results indicated that proteins retained their structural integrity and bioactivity during the preparation of formulation and also during the release process. In vitro cell culture studies such as cell viability, cytotoxicity and biocompatibility studies performed on various ocular cell lines confirmed the safety of PB copolymers for ocular applications. Further, we have performed in vivo ocular tolerability studies with optimized PB formulations which demonstrated no inflammation, retinal toxicity, change in intraocular pressure or cataract even after 16 week of exposure. Moreover, in vivo studies further revealed that PB copolymers based formulations were slowly degraded and dissolved in vitreous humor confirming biodegradability of polymers. Our studies indicated that PB copolymer based composite formulation can serve as a platform technology for the development of sustained release therapy in the treatment of posterior segment ocular diseases such as wet age-related macular degeneration (wet-AMD), diabetic macular edema (DME) and diabetic retinopathy (DR). This technology has a scope beyond ocular treatments and can also be used for the treatment of other chronic diseases.Item Novel Transporter Targeted Lipid Ester Prodrugs of Cidofovir and Sustained Release Formulations for Cytomegalovirus Infection(2013) Gokulgandhi, Mitan R.; Mitra, Ashim K., 1954-Cidofovir (CDF) has shown potential in vitro and in vivo antiviral activity against cytomegalovirus (CMV) retinitis. However, low CDF permeability due to its hydrophilic nature limits its effectiveness. Furthermore, physicochemical properties such as high water solubility (170 mg/mL) and log p (-3.9) of CDF are unfavorable for passive transcellular absorption. Therefore, we have tested novel transporter targeted lipid prodrugs of CDF and their nanoparticle formulations to achieve sustain drug delivery for the treatment of CMV retinitis. These prodrugs contains lipid linker of different carbon chain length (e.g. C2, C6, and C12) carrying minimum two functional groups to link one end with CDF and another end with ligand (biotin) targeting sodium dependent multivitamin transporter (SMVT). We have successfully synthesized three biotinylated (B) derivatives (B-C2-cCDF, B-C6-cCDF, and B-C12-cCDF) and two lipid ester derivatives (C6-cCDF and C12-cCDF) of cCDF. Structure and purity of all the produgs were confirmed using ³¹P NMR and LC-MS/MS analysis. All synthesized prodrugs were characterized in terms of its physicochemical properties, in vitro cellular accumulation, cytotoxicity, transporter affinity, ocular tissue stability and in vitro antiviral potency against various virus strains. Furthermore, prodrug loaded PLGA nanoparticle formulations were prepared and characterized. Overall, improvement in physicochemical properties, lipophilicity, interaction with SMVT transporter, higher tissue-buffer partition coefficient and rapid conversion into parent molecule (in retina–choroid) were observed with higher lipid chain length (B-C12-cCDF > B-C6-cCDF > B-C2-cCDF). In vitro antiviral activity shows B-C12-cCDF as the potential prodrug candidate for future in vivo evaluation. Sustained release (~3 weeks) formulation of B-C12-cCDF loaded PLGA nanoparticles may serve as a viable intravitreal delivery approach for the treatment of CMV retinitis. Based on the above finding we hypothesized that lipid raft facilitates enhanced prodrug interaction with cell membrane and thereby assist docking of targeted ligand into the binding domain of transporter. Therefore, current novel approach combines both lipids mediated facilitate diffusion and transporter targeted delivery to generate synergistic drug effect. Overall, these findings indicate that transporter-targeted lipid analogue of CDF could be a viable strategy for the treatment of CMV retinitis.Item Provider Perceptions of a Mature School-Based Health Center's Effect on Adolescent Health and Wellness: A Case Study(2015-06-02) Blacksin, Beth Amy; Kelly, Patricia J. (Patricia Jane)School-based health centers (SBHCs) have emerged as a model of care that enhances utilization of health care services for U.S. adolescents. Staffed by interdisciplinary teams, physicians play a largely supervisory role in these centers, where nurses and Nurse Practitioners play a critical role. Yet the school-based health center model is unfamiliar to many nurses. The passage of comprehensive national health care reform included dedicated funding for school-based health centers, so this model is expected to expand in the near future. The aims of this study were: (1) To explore how a mature SBHC functions to impact the health and well-being of adolescents; and (2) To explore how adolescent health care providers in a mature SBHC perceive the effect of its utilization on the health and well-being of adolescent users. The research question was: How does a SBHC affect the health and well-being of its adolescent users, as perceived by providers in a mature school health center? The study used a single case study methodology and employed multiple methods of data collection, including interviews with the providers, staff and historians of the center, document review, descriptive epidemiologic data, and limited observation. An adaptation of Bronfenbrenner’s ecological model of multiple influencers as the theoretical framework guided interviews and data analysis. Findings included the themes of: easy access, providers as connectors, and care of the whole adolescent. This SBHC was able to construct a robust nexus of care that fit the needs of adolescent users, creating a complex network of internal and external partnerships with the school and local community.Item Transporter-Targeted Prodrug Delivery to Improve Oral Bioavailability of Saquinavir(2015-05-22) Wang, Zhiying; Mitra, Ashim K., 1954-Saquinavir (SQV) is the first antiretroviral protease inhibitor approved by Food and Drug Administration in the United States for the treatment of HIV infection due to its potent anti-HIV activity. However, some unfavorable properties including low aqueous solubility, low intestinal absorption and fast biotransformation lead to its poor oral bioavailability and limited therapeutic efficacy. The objective of this dissertation project is to investigate whether stereoisomerized peptide prodrugs targeting influx transporter system could improve intestinal absorption and oral bioavailability of SQV. Four stereoisomeric dipeptide prodrugs of SQV including L-valine-L-valine-SQV (LLS), L-valine-D-valine-SQV (LDS), D-valine-L-valine-SQV (DLS) and D-valine-Dvaline- SQV (DDS) and two amino acid prodrugs including L-valine-SQV (LS) and Dvaline- SQV (DS) were synthesized and investigated using in vitro cell culture models. All dipeptide prodrugs exhibit improved aqueous solubility, lowered cytotoxicity, and reduced P-gp/MRP2-mediated efflux activities regardless of stereochemistry in promoieties. SQV attached with L-isomers shows higher affinity for peptide transporters but lower stability and higher toxicity, whereas conjugation with D-isomers can enhance stability and reduce toxicity, but not be recognized by peptide transporters. Subsequently, results of metabolism studies performed in rat liver microsomes indicate that elimination half life of SQV was prolonged dramatically by 7- to 40-fold due to prodrug modification with the rank order of DDS > DLS > LDS > LLS > DS > LS > SQV. In comparison with D-configuration, L-configuration favors the interaction between prodrugs and rat hepatic CYP3A enzymes, resulting in a relatively rapid clearance by CYP3A. Stereoselectivity is also observed in protein binding of prodrugs in rat plasma. Lower protein binding was obtained by LS, LLS and LDS but higher by DS, DDS and DLS as compared with SQV. Pharmacokinetics studies performed in rats provide further evidences that oral bioavailability of SQV is drastically enhanced by conjugation with dipeptide L-valine-Dvaline. In conclusion, stereoisomeric dipeptide prodrug modification targeting specific influx transporters could be a successful strategy to improve bioavailability of poorly absorbed drug SQV. Additionally, influence of exogenous human efflux transporters (P-gp, MRP2 and BCRP) on functional activities of endogenous peptide transporters (PepT) was also delineated in MDR-transfected MDCK cell lines in this project. Results demonstrate that overexpression of MDR genes reduces PepT function probably due to the phenomenon of transporter-compensation resulting in down-regulation of endogenous genes. It may provide some mechanistic insight into possible reasons for underestimation in drug screening using these cell models.Item The relationship between African American high school students' desire to attend college, their perceived likelihood to attend college and actual college entrollment(2013) King, Makini Lateefah, 1981-; Barber, Carolyn E.African American students continue to be an underrepresented population in institutions of higher education. This study uses Mickelson’s Attitude-Achievement Paradox to explain the effect of individual and contextual SES, students’ sense of belonging, achievement and engagement on student’s desire to attend college and perceived likelihood of attending college and eventual college enrollment. Using waves I and III of the Adolescent Health dataset, the researcher explores how SES and individual high school experiences impact the desire and perceived likelihood of college enrollment and actual college enrollment for 1775 African American 9-11th grade students. Multi-level model analysis showed that individual and contextual SES and students’ high school experiences were more strongly related to perceived likelihood of attending college than to desire and that these variables also predicted actual college attendance. Perceived likelihood partially mediated the relationship between desire, SES, belonging, achievement and actual college attendance. These results suggest the importance of encouraging positive high school experiences for African American students, assisting students with locating financial support for college and increasing students’ perception that they are likely to attend college.