Thesis (M.S.)--West Virginia University, 2001. / Title from document title page. Document formatted into pages; contains vi, 96 p. : ill. (some col.). Vita. Includes abstract. Includes bibliographical references (p. 95).

Thesis (Ph. D.)--University of California, San Diego and San Diego State University, 2002. / Vita. Includes bibliographical references (leaves 84-93).

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BACKGROUND:Nephropathic cystinosis is an autosomal recessive disease resulting from intracellular accumulation of cystine leading to multiple organ failure.CASE REPORT:We describe the clinical course of a patient managed from the age of six until his death at the age of 33 years. He underwent multiple surgery, including two renal transplants, developed transplant renal artery stenosis that was managed medically, and progressive heart failure at the age of 33 years. His death from a ruptured pseudoaneurysm associated with a restrictive cardiomyopathy is noteworthy. A limited cardiac autopsy revealed the presence of cystine crystals in interstitial cardiac histiocytes and one myocardial cell, along with 1000-fold higher tissue cystine content of the left ventricular myocardium compared to patients without cystinosis, suggesting the possibility of direct cystine mediated metabolic injury.

Most marketed formulations for treating anterior ocular conditions are topical, with conventional eyedrops representing the most utilized modality. However, due to the natural clearance mechanisms of the eye, less than 5% of an applied dose remains bioavailable following administration. To overcome the shortcomings associated with conventional eyedrops, a series of enzymatically degradable, mucoadhesive thermogels were developed. Thermogels can be applied as a solution, like a conventional eyedrops, but gel against the heat of eye. To avoid obstructing vision, these thermogels were designed to be instilled within the inferior fornix of the eye. In these studies, the base thermogelling polymer (pNAM) was crosslinked with the natural polymer chitosan. Not only does crosslinking strengthen the typically weak thermogels, but chitosan can be enzymatically degraded by lysozyme, the highest concentration protein found in tear fluid. Therefore, the developed thermogels can be applied to the inferior fornix and degrade over multiple days. A limitation of applying materials to the inferior fornix is they tend to be poorly retained. To anchor the developed thermogels within the inferior fornix, the mucoadhesive properties were tailored based on the chitosan utilized as well as the inclusion of a disulfide monomer capable of covalently bonding with the natural mucosal layer covering the surface of the eye. The disulfide bridging monomer could be further conjugated with therapeutic components which were released as a function of mucosal interaction. Conjugates investigated included cysteamine for treating cystinosis, n-acetyl cysteine for treating dry eye, the adhesion peptide RGDC as a model peptide/protein, and polyethylene glycol for modulating material properties. The release of the drugs Ketotifen Fumarate, for treating allergic conjunctivitis, and atropine, for treating myopia, were also investigated. The safety of the developed thermogels were studied both in vivo and extensively in vitro utilizing both rat and rabbit models. / Dissertation / Doctor of Philosophy (PhD) / Topical eyedrops are the most utilized treatment option for the vast majority of ocular diseases. However, eyedrops are largely ineffective with less than 5% of an applied dose reaching the desired cite of action. Therefore, eyedrops need to be frequently reapplied. To overcome these limitations, an eyedrop was developed which can be applied as a liquid but gels against the heat of the eye. This gel allows for prolonged drug release over multiple days, greatly increasing drug efficacy as well as patient comfort and compliance. To prevent obstructing vision, these gels can be applied under the lower eyelid. To keep these gels retained under the lower eyelid, they were designed to anchor to the natural mucus layer which covers the surface of eye. The developed eyedrops represent a significant advancement in ocular care; bettering the convenience, comfort, and effectiveness for patients of a topical formulation compared to traditional eyedrops.

Cystinosis is a metabolic disorder characterised by the abnormal accumulation of the amino acid cystine in cells leading to a slow destruction of all major organs. If patients diagnosed with cystinosis are untreated, death due to kidney failure ensues in the second decade of life. A number of studies have shown the ability of the drug cysteamine (Cystagon®) to lower cystine accumulation within cells resulting in reduced organ and tissue damage. Cysteamine therapy however, is associated with a number of side effects involving the gastrointestinal tract and the central nervous system. Most of these arise due to the large amount of cysteamine present in the stomach and gut following administration. In addition, cysteamine possesses an unpleasant taste and smell, resulting in poor patient compliance. In an attempt to overcome these problems, a number of pro-drug derivatives of cysteamine and cystamine, the disulfide analogue of cysteamine, have been synthesised and evaluated. Pro-drugs were synthesised using a route established in our laboratories. Briefly, cystamine dihydrochloride was basified and allowed to react with a number of cyclic anhydrides under basic conditions. The resulting di-acids were reacted with carbonyldiimidazole and monoBoc-cystamine to yield the desired pro-drugs. Removal of the tBoc-protecting group was achieved in a facile manner by use of trifluoroacetic acid to yield product. The efficacy of the synthesised pro-drugs was determined by incubation of 50μM compound in a suspension of cultured cystinotic fibroblasts, with 50μM cysteamine as control. Cell growth was measured at 72 h and the level of thiol determined. All except one of the pro-drugs tested were significantly more effective than the control at lowering the cystine burden of the cells. Further work will concentrate on repeating these studies and evaluating a more robust Structure Activity Relationship for these compounds. The overall aim of all this work remains the production of an odourless, tasteless and orally active treatment for cystinosis and, if possible, improve on the current dosing regimen of every 6h. By using pro-drugs, cysteamine will be chemically camouflaged and hence, the side effects associated with its administration will be minimised or even entirely abolished.

Nephropathic cystinosis is a rare autosomal recessive disease characterised by raised lysosomal levels of cystine in the cells of almost all organs. It is treated by regular oral and topical administration of the aminothiol, cysteamine(Cystagon™), which possesses an offensive taste and smell. The oral form frequently causes emesis,and should be administered every six hours to be maximally effective. The topical eye drop treatment requires hourly application to be most effective.In an attempt to reduce this frequency and improve the treatment, the preparation and evaluation of three alternative cysteamine containing formulations (suppositories, long-acting ophthalmic gels and an inhaler) was undertaken. The physiochemical properties, stability and release profiles of the active (cysteamine or phe conjugate) from the formulations were evaluated. The suppositories released cysteamine over a 20-40 minute period with a T75= 10-13minutes. They were most stable at 4°C. The analysis of the ophthalmic gels demonstrated that a weak gel network was formed at low shear stress, the bioadhesion of the gel was increased with inclusion of a cysteamine derivative (e.g.mean force of 0.067N compared to 0.107N with compound included) and eight-hour, first order release from the gel was observed. There was significant adhesion observed between the ophthalmic gels and bovine corneal tissue. The pulmonary microspheres were spherical and within the optimum size range for deep lung delivery (1-5μm). However, Andersen Cascade Impactor analysis revealed poor deep lung penetration. In conclusion, these results demonstrated that more development work was required to produce a useful pulmonary formulation of cysteamine, however, formulation of an ocular applicable gel or suppository was readily achievable. The suppository preparations may be particularly beneficial for the treatment of infants, whilst the ophthalmic gel preparations could be developed for daily or overnight use. With respect to pulmonary delivery, microspheres in the optimum size range were produced. However, deep lung targeting was prevented by static agglomeration, which requires further investigation.