Melanotan-II (MT-II) is a cyclic heptapeptide analogue of α-MSH (α-melanocyte stimulating hormone) which tans the skin rapidly and is currently being evaluated for the prevention of sunlight-induced skin cancers. In these preformulation studies, the degradation of MT-II followed apparent first-order kinetics. The degradation rate increased as the temperature or phosphate buffer concentration were increased. The pH-rate profile of MT-II degradation showed that MT-II was most stable at approximately pH 5. The degradation of MT-II was not affected by ionic strength. The pKa1 and pKa2 were estimated to be 6.53 and 11.74, respectively. The partition coefficient was studied at various pH values. The Δ log PC at pH 7.35 was 1.05 which indicated that MT-II could pass the intestinal membrane relatively easily. The preformulation data presented here can be used to help develop an appropriate dosage form for MT-II.

<p> Chronic pain is a huge economic burden and a devastating complication requiring therapeutic interventions. This dissertation evaluated the analgesic potency, efficacy, tolerance and cross-tolerance profile of morphine and its ester derivative, morphine-6-<i>O</i>-sulfate (M6S) in a rat model of diabetic neuropathy, and in normal rats utilizing a multimodal pain domain approach (e.g., burning pain, pricking pain, and deep muscle pain). Diabetes was induced in male Sprague-Dawley rats with streptozotocin, 65mg/kg (intraperitoneally). Hot water tail-flick latency, paw pressure, pinprick sensitivity and hot plate withdrawal thresholds (HTL, PPT, PST and HPT) were measured at various time points and doses after i.p. injection (N=6) on days 1, 3, 7, 9, 12, 19, 25 and 29 of treatment in both normal and diabetic rats. Affinity binding studies, GTP&gamma; S assays and intracellular cAMP assays were performed in Chinese hamster ovary cells transfected with human mu (&mu;) - or delta (&delta;)-opioid receptors (N=5). HPLC-DAD stability studies were performed <i>in vitro </i> in various pH buffers and biological fluids (N=4). Pharmacokinetic parameters for M6S were studied using intravenous (1 mg/kg), intraperitoneal (i.p. 5.6, 10 mg/kg) and oral routes (10, 30 mg/kg) of drug administration (N=4 to 7 for each dose). Compared to morphine, in diabetic animals, M6S was 12-fold more potent in the HPT test (ED₅₀ M6S = 1.1 &plusmn; 0.02 &mu;mol/kg Vs ED₅₀ MOR=12.1 &plusmn; 1.8 &mu;mol/kg; p&lt; 0.05), and 20-times more potent in the HTL test (in diabetic rats, ED₅₀ M6S = 1.1 &plusmn; 0.02 &mu;mol/kg Vs ED₅₀ MOR = 12.1 &plusmn; 1.8 &mu;mol/kg; p&lt; 0.001). In the PST test, M6S was 3-fold more potent than morphine in diabetic rats (ED₅₀ M6S = 6.9 &plusmn; 1.46 &mu;mol/kg Vs ED₅₀ MOR = 16.2 &plusmn; 1.24 &mu;mol/kg). Similar potency differences were also observed between morphine and M6S in normal rats. Diabetes caused a decrease in potency (about 3-5 fold) of morphine in the HPT and HTF tests and a loss of efficacy in the PST and PPT tests (4^th week of diabetes) without significantly effecting M6S potency and efficacy in all these tests. Furthermore, 9 <i>vs.</i> 28 days of chronic treatment were required for rats to become tolerant to morphine and M6S, respectively, in the HTF and HPT tests. M6S also sustained its potent analgesic effects in morphine-tolerant rats (i.e. no cross-tolerance) and demonstrated clinical potential as an opioid rotation drug in morphine-tolerant subjects. With no significant differences in binding, activation of GTP&gamma; and inhibition of cAMP at the &mu;-opioid receptor, M6S activated G-proteins via &delta;-ORs more potently than morphine <i> (e.g.,</i> M6S-ED₅₀ = 101 &plusmn; 41.6 nM; MOR-ED₅₀ = 785 &plusmn; 140 nM), and modulated adenylyl cyclase activity via &delta;-ORs in intact CHO-hDOR cells more potently than morphine <i>(e.g.,</i> M6S-ED₅₀ = 55.1 &plusmn; 17.5 nM; MOR-ED₅₀ = 372 &plusmn; 11.9 nM). Naltrindole (1mg/kg; i.p.) blocked M6S analgesia by 50% in HTF, HPT tests and 90% in the PST test but was without effect on morphine analgesia. In vitro stability studies and pharmacokinetic studies showed no susceptibility of M6S to hydrolyze to morphine. The bioavailability of M6S after the i.p. route of administration was above 93%, while oral bioavailability was ~ 5%. Our <i>in vitro</i> stability assays and pharmacokinetic studies have demonstrated that M6S is not a prodrug of morphine and that the molecule has difficulty in passing the epithelial barrier of gastrointestinal tract, due to its polar and zwitterionic nature. Our computational and receptor docking studies suggested that the sulfate moiety of M6S and 214 lysine residue of delta opioid receptor is a critical binding interaction. Collectively, our studies have provided a set of direct and indirect evidence supporting the ability of the stable M6S molecule to interact with and activate both &mu;, and most importantly, &delta;-opioid receptors to modulate their respective pain control pathways. In addition, evidence has been provided that clearly demonstrates the superiority of M6S over morphine in the treatment of multidomain neuropathic pain.</p>

Objectives: To identify and characterize response patterns in patients with wet age-related macular degeneration (AMD) over 24 month after ranibizumab therapy; to determine demographic and clinical predictors of response patterns at the initial time of treatment (baseline); and to quantify and compare resource utilization across response patterns at the end of treatment (month 24).Methods: We performed a secondary data analysis using existing data from a prospective, observational, multicenter, open-label trial of 0.5 mg of ranibizumab administered by intravitreal injection. Patients with wet AMD were followed over 24 +/- 3 months with intermediate data points at 6 +/- 2 months and 12 +/- 2 months, and a few data points at 2.5 +/- 1 month that coincided with the end of the loading phase. The primary outcome of interest was change in visual acuity (as measured by Early Treatment Diabetic Retinopathy Study [ETDRS] letters). Latent class growth analysis (LCGA) was used to examine treatment response variability in the data and to identify latent classes (unobserved groups) of response patterns. A multinomial logistic regression was specified to identify predictors of the response patterns. Variables related to resource utilization at the end of treatment were also examined across response patterns. Results: LCGA demonstrated a large variability in visual acuity change. We identified three clusters of patients for each response pattern. Patients in cluster 1 (partial responders, 56.68% of the total sample) had stable improvement in visual acuity that plateaued at month 3 and then gradually diminished over time. Patients in cluster 2 (optimal responders, 23.50%) showed progressive improvement in visual acuity that plateaued at month 6 and then remained stable over time. Patients in cluster 3 (non-responders, 19.82%) had the worst performing course in visual acuity and showed drastic decline for the first 12 months that tapered off. Multinomial logistic regression revealed significant differences across clusters in terms of age, baseline visual acuity, and certain lesion types. Resource utilization at the end of treatment also varied significantly across clusters, with non-responders on average receiving the lowest total number of ranibizumab per patient. Conclusions: LCGA identified three response patterns to ranibizumab among patients with wet AMD. The patterns were significantly associated with age, baseline visual acuity, and certain lesion types. Non-responders on average received the lowest total number of ranibizumab per patient. Identifying differential responders has obvious advantages for understanding treatment effects and may help provide a basis of classification for intervention.

Pharmacological induction of proteotoxic stress is rapidly emerging as a promising strategy for cancer cell-directed chemotherapeutic intervention. Due to the essential role of the cellular heat shock response in cytoprotection through the maintenance of proteostasis and suppression of apoptosis, small molecule heat shock protein (Hsp) inhibitors can be harnessed for targeted induction of cytotoxic effects in cancer cells. Recent research strongly suggests that melanoma is a malignant tumor amenable to therapeutic modulation of proteotoxic stress, particularly in situations where traditional chemotherapeutics and novel targeted therapies have failed. Based on this rationale, my graduate research has focused on the identification of redox-directed electrophilic pharmacophores capable of modulating the heat shock response for the therapeutic induction of proteotoxic stress targeting malignant human melanoma cells. The following specific aims were pursued: (1) to identify redox-directed heat shock response modulators active in malignant melanoma cells; (2) to explore melanoma cell directed activity of our lead heat shock response inducer, aurin (4-[bis(p-hydroxyphenyl)methylene]-2,5-cyclohexadien-1-one; CAS #143-74-8); and (3) to explore melanoma cell directed activity of our lead heat shock response antagonist, methylene blue (3,7-bis(dimethylamino)-phenothiazin-5-ium chloride; CAS#: 61-73-4). First, we demonstrate that the quinone methide, aurin, is a targeted Hsp90 inhibitor that induces apoptosis in human malignant melanoma cells but not in non-malignant human skin cells. Second, we have identified methylene blue as a functional antagonist of the global heat shock response which is active at the mRNA and protein levels, and have discovered that it sensitizes melanoma cells to the apoptogenic activity of the Hsp90 antagonist, geldanamycin. Taken together, these data suggest the feasibility of aurin and methylene blue as functional therapeutic heat shock response modulators targeting melanoma cells through pharmacological induction of proteotoxic stress.

We now know from genomics that many disease states lead to changes in expressed proteins (adaptation/plasticity). Therefore, drug design and discovery based on normal states and single targets often is inadequate or even counter-indicated. Therefore, the "system changes" that have occurred must be considered in any treatment for the disease. Such "systems changes" are clearly evident in neuropathic pain where opioids can actually heighten pain. In these pain states there are increased levels of neurotransmitters such as cholecystokinin (CCK) in which both the peptides and their receptors are increased in pain states. To effectively treat diseases involving "systems changes" a new paradigm for the design of compounds was proposed. In this new approach single peptide or peptidomimetic molecules are designed to interact with multiple receptor targets. For the treatment of pain, a series of linear and cyclic peptides and peptidomimetics were designed based on the overlapping pharmacophores of opioid and CCK ligands. The CCK/opioid analogues were synthesized and evaluated for their biological activities. Several of the CCK/opioid analogues were found to simultaneously interact with opioid receptors as agonists and CCK receptors as antagonists. In addition, the lead compounds have been tested in several pain models and were found to be promising in the treatment of neuropathic pain. Further, the structure-activity relationships of these novel peptides have provided new insights into the requirements for binding and bioactivity at opioid and CCK receptors, as well as the overlapping pharmacophores of CCK and enkephalin.

Pentaazapentacosane pentahydrochloride (PAPC-HCl) is a synthetically produced aliphatic pentaamine that is being investigated for use as an anticancer agent. As part of this research project a rapid high-performance liquid chromatographic method for determination of the dansyl derivative of PAPC was developed. The chromatographic system uses a reverse phase C-8 column, a mobile phase of acetic acid buffer and acetonitrile and UV detection. The dansylation conditions were optimized with a pH of 11.0 and a 20 fold dansyl chloride excess. The yield of dansyl PAPC increased 10 fold as the reaction pH was changed from 9.5 to 10.5. An investigation of the products formed in the dansylation reaction revealed that, even under conditions of pH and dansyl chloride concentration most likely to produce partially dansylated products, only perdansyl PAPC is present. This unexpected finding is explained by a mechanism whereby (1) only completely unionized amine molecules will dansylate and (2) the ratio of unionized molecules to ionized molecules increases as dansylation proceeds. The proposed mechanism is verified by comparing the dansylation vs. pH profile of PAPC to that of a reference monoamine (piperidine ·HCl). After 4 hours at room temperature and pH 9.5, 100% of piperidine is dansylated while under the same conditions only 10% of PAPC is derivatized. A pH greater than 10.5 is required to completely dansylate PAPC. This difference is significantly greater than would be predicted from the pKₐ values but it is consistent with the proposed mechanism.

Carbendazim (methyl-2-benzimidazolecarbamate), a well known anti-fungal agent that may have significant anti-cancer activity, is a poorly water-soluble ampholyte. Unfortunately its projected oral dose up to hundred mg per day is far greater than its water solubility of 6 μg/ml. The overall aim of this research was to conduct preformulation studies and develop therapeutically viable oral and parenteral formulations. The solubility of carbendazim was altered by using both solute and solvent modification approaches. The solvent modification was carried out by investigating the solubilization of carbendazim by pH in combination with cosolvents, surfactants or complexants. At pH 7 the total drug solubility is 6.11 ± 0.45 μg/ml which increases by 1 to 7 fold with cosolvent, surfactant or complexant. However, at pH 2 the solubility increases by 400 times and at pH 1.3 over 3000 times. Both cosolvents and non-ionic surfactants have a negligible effect on the total drug solubility at pH 2, whereas the total drug solubility increases by combining pH 2 with anionic surfactants or complexants. The solute modification was accomplished by preparing different salts of carbendazim. In all six different salts, viz., hydrochloride dihydrate, phosphate, hemisulfate hydrate, mesylate, besylate, and tosylate were prepared. Their structures were determined using single crystal x-ray diffraction. The developability attributes (i.e. hygroscopicity, thermal behavior, aqueous solubility, solid state stability, and dissolution rate) of these compounds were evaluated. The dissolution studies showed that all the salts had better dissolution rate than the free base, the mesylate salt been the best. Based on the preformulation studies, two formulations were chosen for oral dosing in mice. The proposed oral formulation of 9.4 mg/ml carbendazim at pH 1 in phosphate buffer and 20% SBEβCD phosphate buffer, respectively, did not precipitate on in vitro static serial dilution with water and seven up. However, the formulation containing only phosphate buffer showed activity in mice by significantly reducing the tumor growth in B-16 melanoma injected mice. Also, it gave active blood levels, which were comparable to IV dosing.

Purposes: The purpose of this study was to estimate the incidence rate of epilepsy in 2009 among various patient populations of Arizona seniors enrolled in Medicaid. Additionally, this study aimed to assess the incremental and relative economic and healthcare resource burden of new-onset epilepsy compared with those without newly diagnosed epilepsy. Methods: Arizona Medicaid claims data years 2008 through 2010 were used for this analysis. Patients who were at least 65 years of age as of January 1, 2009 and were continuously enrolled in any coverage group pertaining to Arizona Medicaid for at least 12 months during 2008 and 2009 were considered eligible for the 2009 cohort; patients who were continuously enrolled during 2008 but not at all during 2009 were excluded. In addition to meeting the criteria for the 2009 cohort, incident cases of 2009 must also have epilepsy-related healthcare claims, a one-year clean period during which they cannot have any epilepsy-related healthcare claim, and that their first epilepsy-related healthcare claim in 2009 cannot have with a diagnosis code of 345.x1. The outcome variables assessed included incidence rate of being diagnosed with epilepsy, total monthly healthcare costs, total monthly inpatient costs, total monthly outpatient costs, total monthly prescription costs, incidence rate of inpatient stay, and incidence rate of physician visits. Results: The incidence rate of epilepsy in 2009 for this population was 7.9 per 1,000. Patients with epilepsy-related disease-based risk factors and of younger age had significantly greater incidence rate compared with their counterparts. In general, total monthly healthcare costs and incidence rate of inpatient stay were approximately three times greater in patients with newly diagnosed epilepsy compared with their counterpart (p < 0.001); incremental total monthly healthcare costs in patients with newly diagnosed epilepsy reached 2,077 US dollars. Conclusion: The Arizona Medicaid population appeared to have higher incidence rate compared with the US Medicare and the general US population. Additionally, Arizona Medicaid beneficiaries with newly diagnosed epilepsy had significantly greater healthcare costs and resource utilization compared with those without epilepsy.

Purpose: The purpose of this research was to examine and compare readmissions between the rural and urban areas for dual eligible patients in Arizona. The study also examined if living in an area of high socioeconomic status as opposed to living in an area of low socioeconomic status affects patient's risk of all-cause-30-day readmission. Methods: The study used data on dual eligible members who were continuously enrolled in AHCCCS and the University of Arizona Health Network between January 1st, 2011 and November 30th, 2012. The outcomes of interest in this study were risk of all-cause-30-day readmission, length of stay at index admission, and cost of readmission (i.e. amount paid by the University of Arizona Health Network). Log-binomial regression, Poison regression, and gamma regressions were used to model risk of readmission, length of stay at index admission, and cost of readmissions respectively. Results: Readmission of dual eligible patients in this sample was not related to residential location of patients. All-cause-30-day readmission did not differ for rural dual eligible patients and urban dual eligible patients. Readmission of dual eligible patients in this sample was not related to socioeconomic conditions. Dual eligible patients discharged in the year 2011 had a significantly higher risk of all-cause-30-day readmission than those discharged in the year 2012 (RR=1.05; p=0.03). Dual eligible patients discharged to skilled nursing facilities had a higher all-cause-30-day readmission risk compared to those discharged home. Length of stay at index admission was not associated with residential location. Patients admitted in the year 2011 were more likely to stay longer than those admitted in 2012 (IRR=1.13; p<.0001).The longer a dual eligible patient stayed during their readmission, the more their cost of readmission. (RR=1.33; p=0.0024). Conclusions: Readmission was neither associated with residential location nor socioeconomic condition. Cost associated with all-cause-30-day readmissions was not associated with residential location of the patients. The study also concludes that length of stay at index admission did not differ with socioeconomic conditions of the areas patients lived. It also concludes that year of discharge was associated with their risk of all-cause-30-day readmission.

Targeted cancer therapeutics represent the advent of a new therapeutic age, brought forth by the small molecule tyrosine kinase inhibitor (TKI) imatinib (Gleevec®). Imatinib is able to cause complete and sustained remissions in patients with chronic myelogenous leukemia (CML) driven by the Abelson (ABL) kinase, which caused a massive paradigm shift in how cancer is treated. The following research has been completed to extend the principles of imatinib therapy to the rearranged during transfection (RET) kinase. The RET kinase is involved in driving the pathology of medullary thyroid cancer (MTC), papillary thyroid carcinoma (PTC), certain non-small cell lung cancers (NSCLC), chronic myelomonocytic leukemia (CMML), tamoxifen resistant breast cancer, and Spitz melanoma. A heavily diverse population of solid and liquid carcinomas are driven by the RET oncogene, and patients presenting with these cancers could significantly benefit from a RET inhibitor. Previous drug discovery campaigns identified RET activity after therapeutic development for an unrelated kinase, as the case with vandetanib (Calpresa®) and cabozantinib (Cometriq®). Both agents fail to achieve dominant activity on RET and are more active on the vascular endothelial growth factor receptor 2 (VEGFR2), yet still achieve efficacy in RET driven tumors. This likely results from interrupting the oncogene cooperation between RET and VEGFR2; VEGFR2 provides the nutrients through angiogenesis that RET requires to promote proliferation and survival. We hypothesized that an equipotent RET/VEGFR2 dual inhibitor could maximize inhibiting the cooperation between RET and VEGFR2 in RET driven cancers. The inhibitor should be developed to maintain activity on all known RET mutations for treatment durability. In that case, the RET oncogene, despite mutating, will always be inhibited. Through research efforts, Pz-1 was identified as a sub-nanomolar, equipotent inhibitor of both RET (IC₅₀<0.001 µM) and VEGFR2 (IC₅₀<0.001 µM). Pz-1 was found active on every known, clinically relevant RET mutant tested at an IC₅₀≤0.001 µM. Through RET-driven xenograft models, Pz-1 was found active at an oral dose as low as 0.3 mg/kg/day.