Modulating bioreductive drug activity with antivascular agents

Modulation of tumor hypoxia to increase bioreductive drug antitumor activity was investigated. The antivascular agent 5,6-dimethylxanthenone acetic acid (DMXAA) was used in combination studies with the bioreductive drugs Tirapazamine (TPZ) and Mitomycin C (MMC). Blood perfusion studies with DMXAA showed a maximal reduction of 66% in tumor blood flow 4 hours post drug administration. This tumor specific decrease in perfusion was also found to be dose-dependent, with 25 and 30 mg/kg DMXAA yielding greater than 50% reduction in tumor blood flow. Increases in antitumor activity with combination therapy (bioreductive drugs $+$ DMXAA) were significant over individual therapies, suggesting an increased activity due to increased hypoxia induced by DMXAA. Combination studies yielded the following significant tumor growth delays over control: MMC (5mg/kg) $+$ DMXAA (25mg/kg) = 20 days, MMC (2.5mg/kg) $+$ DMXAA (25 mg/kg) = 8 days, TPZ (21.4mg/kg) $+$ DMXAA (17.5mg/kg) = 4 days. The mechanism of interaction of these drugs was investigated by measuring metabolite production and DNA damage. 'Real time' microdialysis studies indicated maximal metabolite production at 20-30 minutes post injection for individual and combination therapies. DNA double strand breaks induced by TPZ $\pm$ DMXAA (20 minutes post injection) were analyzed by pulsed field gel electrophoresis (PFGE). Southern blot analyses and quantification showed TPZ induced DNA double strand breaks, but this effect was not evident in combination studies with DMXAA. Based on these data, combination studies of TPZ $+$ DMXAA showed increased antitumor activity over individual drug therapies. The mechanism of this increased activity, however, does not appear to be due to an increase in TPZ bioreduction at this time point. ^

THE INTERACTION BETWEEN INTERNAL AND EXTERNAL INFORMATION ON RELATIONAL DATA SEARCH

People often use tools to search for information. In order to improve the quality of an information search, it is important to understand how internal information, which is stored in user’s mind, and external information, represented by the interface of tools interact with each other. How information is distributed between internal and external representations significantly affects information search performance. However, few studies have examined the relationship between types of interface and types of search task in the context of information search. For a distributed information search task, how data are distributed, represented, and formatted significantly affects the user search performance in terms of response time and accuracy. Guided by UFuRT (User, Function, Representation, Task), a human-centered process, I propose a search model, task taxonomy. The model defines its relationship with other existing information models. The taxonomy clarifies the legitimate operations for each type of search task of relation data. Based on the model and taxonomy, I have also developed prototypes of interface for the search tasks of relational data. These prototypes were used for experiments. The experiments described in this study are of a within-subject design with a sample of 24 participants recruited from the graduate schools located in the Texas Medical Center. Participants performed one-dimensional nominal search tasks over nominal, ordinal, and ratio displays, and searched one-dimensional nominal, ordinal, interval, and ratio tasks over table and graph displays. Participants also performed the same task and display combination for twodimensional searches. Distributed cognition theory has been adopted as a theoretical framework for analyzing and predicting the search performance of relational data. It has been shown that the representation dimensions and data scales, as well as the search task types, are main factors in determining search efficiency and effectiveness. In particular, the more external representations used, the better search task performance, and the results suggest the ideal search performance occurs when the question type and corresponding data scale representation match. The implications of the study lie in contributing to the effective design of search interface for relational data, especially laboratory results, which are often used in healthcare activities.

The roles of costimulatory molecules in the cooperative effects of combination epinephrine and dexamethasone on type-1/type-2 cytokine production in tetanus-toxoid specific stimulated human peripheral blood mononuclear cells

A growing number of studies show strong associations between stress and altered immune function. In vivo studies of chronic and acute stress have demonstrated that cognitive stressors are strongly correlated with high circulating levels of catecholamines (CT) and corticosteroids (CS) that are associated with changes in type-1/type-2 cytokine expression. Although individual pharmacologic doses of CS and CT can inhibit the expression of T-helper 1 (Th1, type-1 like) and promote the production of T-helper 2 (Th2, type-2 like) cytokines in antigen-specific and mitogen stimulated human leukocyte cultures in vitro, little attention has been focused on the effects of combination physiologic-stress doses of CT and CS that may be more physiologically relevant. In addition, both in-vivo and in-vitro studies suggest that the differential expression of the B7 family of costimulatory molecules CD80 and CD86 may promote the expression of type-1 or type-2 cytokines, respectively. Furthermore, corticosteroids can influence the expression of β2-adrenergic receptors in various human tissues. We therefore investigated the combined effects of physiologic-stress doses of in vitro CT and CS upon the type-1/type-2 cytokine balance and expression of B7 costimulatory molecules of human peripheral blood mononuclear cells (PBMC) as a model to study the immunomodulatory effects of physiologic stress. Results demonstrated a significant decrease in type-1 cytokine expression and a significant increase in type-2 cytokine production in our CS+CT incubated cultures when compared to either CT or CS agents alone. In addition, we demonstrated the differential expression of CD80/CD86 in favor of CD86 at the cellular and population level as determined by flow cytometry in lipopolysaccharide stimulated human Monocytes. Furthermore, we developed flow cytometry based assays to detect total β2AR in human CD4+ T-lymphocytes that demonstrated decreased expression of β2AR in mitogen stimulated CD4+ T-lymphocytes in the presence of physiologic stress levels of CS and CT as single in vitro agents, however, when both CS and CT were combined, significantly higher expression of β2AR was observed. In summary, our in vitro data suggest that both CS and CT work cooperatively to shift immunity towards type-2 responses. ^

Identifying and Targeting Molecular Deregulations in Uterine Leiomyosarcoma: A Role for mTOR Inhibition-Based Combination Therapies

Uterine leiomyosarcoma (ULMS) is an aggressive malignancy characterized by marked chemoresistance, frequent relapses, and poor outcome. Despite efforts to improve survival over the past several decades, only minimal advances have been made. Hence, there is an urgent and unmet need for better understanding of the molecular deregulations that underlay ULMS and development of more effective therapeutic strategies. This work identified several common deregulations in a large (n=208) tissue microarray of ULMS compared to GI smooth muscle, myometrium, and leiomyoma controls. Our results suggest that significant loss of smooth muscle and gynecological differentiation markers is common in ULMS, a finding that could help render improved ULMS diagnosis, especially for advanced disease. Similarly to reports in other malignancies, we found that several cancer-related proteins were differentially expressed; these could be useful together as biomarkers for ULMS. Notably, we identified significant upregulation and overexpression of the mTOR pathway in ULMS, examined the possible contribution of tyrosine kinase receptor deregulation promoting mTOR activation, and unraveled a role for pS6RP and p4EBP1 as molecular disease prognosticators. The significance of mTOR activation in ULMS and its potential as a therapeutic target were further investigated. Rapamycin abrogated ULMS cell growth and cell cycle progression in vitro but induced only sight growth delay in vivo. Given that effective mTOR therapies likely require combination mTOR blockade with inhibition of other targets, coupled with recent observations suggesting that Aurora A kinase (Aurk A) deregulations commonly occur in ULMS, the preclinical impact of dually targeting both pathways was evaluated. Combined therapy with rapamycin (an mTORC1 inhibitor) and MLN8237 (an investigational Aurk A inhibitor) profoundly and synergistically abrogated ULMS growth in vitro. Interestingly, the superior effects were noted only when MLN8237 was pre-administered. This novel therapeutic combination and scheduling regimen resulted in marked tumor growth inhibition in vivo. Together, these data support further exploration of dual mTOR and Aurk A blockade for the treatment of human ULMS.

Pathway analysis of genome-wide association study data in primary biliary cirrhosisa

Genome-wide association studies (GWAS) have successfully identified several genetic loci associated with inherited predisposition to primary biliary cirrhosis (PBC), the most common autoimmune disease of the liver. Pathway-based tests constitute a novel paradigm for GWAS analysis. By evaluating genetic variation across a biological pathway (gene set), these tests have the potential to determine the collective impact of variants with subtle effects that are individually too weak to be detected in traditional single variant GWAS analysis. To identify biological pathways associated with the risk of development of PBC, GWAS of PBC from Italy (449 cases and 940 controls) and Canada (530 cases and 398 controls) were independently analyzed. The linear combination test (LCT), a recently developed pathway-level statistical method was used for this analysis. For additional validation, pathways that were replicated at the P <0.05 level of significance in both GWAS on LCT analysis were also tested for association with PBC in each dataset using two complementary GWAS pathway approaches. The complementary approaches included a modification of the gene set enrichment analysis algorithm (i-GSEA4GWAS) and Fisher's exact test for pathway enrichment ratios. Twenty-five pathways were associated with PBC risk on LCT analysis in the Italian dataset at P<0.05, of which eight had an FDR<0.25. The top pathway in the Italian dataset was the TNF/stress related signaling pathway (p=7.38×10 -4, FDR=0.18). Twenty-six pathways were associated with PBC at the P<0.05 level using the LCT in the Canadian dataset with the regulation and function of ChREBP in liver pathway (p=5.68×10-4, FDR=0.285) emerging as the most significant pathway. Two pathways, phosphatidylinositol signaling system (Italian: p=0.016, FDR=0.436; Canadian: p=0.034, FDR=0.693) and hedgehog signaling (Italian: p=0.044, FDR=0.636; Canadian: p=0.041, FDR=0.693), were replicated at LCT P<0.05 in both datasets. Statistically significant association of both pathways with PBC genetic susceptibility was confirmed in the Italian dataset on i-GSEA4GWAS. Results for the phosphatidylinositol signaling system were also significant in both datasets on applying Fisher's exact test for pathway enrichment ratios. This study identified a combination of known and novel pathway-level associations with PBC risk. If functionally validated, the findings may yield fresh insights into the etiology of this complex autoimmune disease with possible preventive and therapeutic application.^

PIM KINASE INHIBITION: SINGLE AGENT AND COMBINATION APPROACHES IN B-CELL LYMPHOMA

Proviral integration site for Moloney murine leukemia virus (Pim) kinases are Ser/Thr/Tyr kinases. They modulate B-cell development but become oncoproteins and promote cancer development once overexpressed. Containing three isoforms, Pim-1, -2 and -3 are known to phosphorylate various substrates that regulate transcription, translation, cell cycle, and survival pathways in both hematological and solid tumors. Mantle cell lymphoma (MCL) is an aggressive B-cell lymphoma. Elevated Pim kinase levels are common in MCL, and it negatively correlates with patient outcome. SGI-1776 is a small molecule inhibitor selective for Pim-1/-3. We hypothesize that SGI-1776 treatment in MCL will inhibit Pim kinase function, and inhibition of downstream substrates phosphorylation will disrupt transcriptional, translational, and cell cycle processes while promoting apoptosis. SGI-1776 treatment induced moderate to high levels of apoptosis in four MCL cell lines (JeKo-1, Mino, SP-53 and Granta-519) and peripheral blood mononuclear cells (PBMCs) from MCL patients. Phosphorylation of transcription and translation regulators, c-Myc and 4E-BP1 declined in both model systems. Additionally, levels of short-lived Mcl-1 mRNA and protein also decreased and correlated with decline of global RNA synthesis. Collectively, our investigations highlight Pim kinases as viable drug targets in MCL and emphasize their roles in transcriptional and translational regulation. We further investigated a combination strategy using SGI-1776 with bendamustine, an FDA-approved DNA-damaging alkylating agent for treating non-Hodgkin’s lymphoma. We hypothesized this combination will enhance SGI-1776-induced transcription and translation inhibition, while promoting bendamustine-triggered DNA damage and inducing additive to synergistic cytotoxicity in B-cell lymphoma. Bendamustine alone resulted in moderate levels of apoptosis induction in MCL cell lines (JeKo-1 and Mino), and in MCL and splenic marginal zone lymphoma (a type of B-cell lymphoma) primary cells. An additive effect in cell killing was observed when combined with SGI-1776. Expectedly, SGI-1776 effectively decreased global RNA and protein synthesis levels, while bendamustine significantly inhibited DNA synthesis and generated DNA damage response. In combination, intensified inhibitory effects in DNA, RNA and protein syntheses were observed. Together, these data suggested feasibility of using Pim kinase inhibitor in combination with chemotherapeutic agents such as bendamustine in B-cell lymphoma, and provided foundation of their mechanism of actions in lymphoma cells.

The SanaViz: Human Centered Geovisualization to facilitate visual exploration of Public Health data

These three manuscripts are presented as a PhD dissertation for the study of using GeoVis application to evaluate telehealth programs. The primary reason of this research was to understand how the GeoVis applications can be designed and developed using combined approaches of HC approach and cognitive fit theory and in terms utilized to evaluate telehealth program in Brazil. First manuscript The first manuscript in this dissertation presented a background about the use of GeoVisualization to facilitate visual exploration of public health data. The manuscript covered the existing challenges that were associated with an adoption of existing GeoVis applications. The manuscript combines the principles of Human Centered approach and Cognitive Fit Theory and a framework using a combination of these approaches is developed that lays the foundation of this research. The framework is then utilized to propose the design, development and evaluation of “the SanaViz” to evaluate telehealth data in Brazil, as a proof of concept. Second manuscript The second manuscript is a methods paper that describes the approaches that can be employed to design and develop “the SanaViz” based on the proposed framework. By defining the various elements of the HC approach and CFT, a mixed methods approach is utilized for the card sorting and sketching techniques. A representative sample of 20 study participants currently involved in the telehealth program at the NUTES telehealth center at UFPE, Recife, Brazil was enrolled. The findings of this manuscript helped us understand the needs of the diverse group of telehealth users, the tasks that they perform and helped us determine the essential features that might be necessary to be included in the proposed GeoVis application “the SanaViz”. Third manuscript The third manuscript involved mix- methods approach to compare the effectiveness and usefulness of the HC GeoVis application “the SanaViz” against a conventional GeoVis application “Instant Atlas”. The same group of 20 study participants who had earlier participated during Aim 2 was enrolled and a combination of quantitative and qualitative assessments was done. Effectiveness was gauged by the time that the participants took to complete the tasks using both the GeoVis applications, the ease with which they completed the tasks and the number of attempts that were taken to complete each task. Usefulness was assessed by System Usability Scale (SUS), a validated questionnaire tested in prior studies. In-depth interviews were conducted to gather opinions about both the GeoVis applications. This manuscript helped us in the demonstration of the usefulness and effectiveness of HC GeoVis applications to facilitate visual exploration of telehealth data, as a proof of concept. Together, these three manuscripts represent challenges of combining principles of Human Centered approach, Cognitive Fit Theory to design and develop GeoVis applications as a method to evaluate Telehealth data. To our knowledge, this is the first study to explore the usefulness and effectiveness of GeoVis to facilitate visual exploration of telehealth data. The results of the research enabled us to develop a framework for the design and development of GeoVis applications related to the areas of public health and especially telehealth. The results of our study showed that the varied users were involved with the telehealth program and the tasks that they performed. Further it enabled us to identify the components that might be essential to be included in these GeoVis applications. The results of our research answered the following questions; (a) Telehealth users vary in their level of understanding about GeoVis (b) Interaction features such as zooming, sorting, and linking and multiple views and representation features such as bar chart and choropleth maps were considered the most essential features of the GeoVis applications. (c) Comparing and sorting were two important tasks that the telehealth users would perform for exploratory data analysis. (d) A HC GeoVis prototype application is more effective and useful for exploration of telehealth data than a conventional GeoVis application. Future studies should be done to incorporate the proposed HC GeoVis framework to enable comprehensive assessment of the users and the tasks they perform to identify the features that might be necessary to be a part of the GeoVis applications. The results of this study demonstrate a novel approach to comprehensively and systematically enhance the evaluation of telehealth programs using the proposed GeoVis Framework.