Doctor of Philosophy (PhD), Wright State University, 2022, Biomedical Sciences PhD

Diffuse midline glioma, K27M-mutant (DMG) are intractable brain tumors, primarily occurring in the pediatric and adolescent population. Patients have a median survival of less than one year after diagnosis. A lack of therapeutic targets has been a barrier to improvement in patient survival. Irradiation therapy improves symptoms while chemotherapy and surgical intervention, for the most part, have not yet demonstrated utility for treatment. DMG are characterized by a histone H3 mutation that results in the genome-wide loss of epigenetic-repressive marks on K27 and is associated with the misexpression of genes, like Cancer/Testis antigens. A member of this group, transketolase-like 1 (TKTL1), is an enzyme that catalyzes reactions bridging glycolysis and the pentose phosphate pathway. This work demonstrates that DMG express TKTL1, which reprograms cellular metabolism in support of tumor cell growth and regulates the expression of hypoxia-inducible factor 1 alpha (HIF-1α). HIF-1α in tumor cells is associated with increased invasion, metastasis, increased survival, and resistance to therapy. Inhibition of HIF-1α in DMG cells decreased hypoxia-induced gene expression, glycolytic capacity, mitochondrial respiration, and tumor cell proliferation. TKTL1 knockdown similarly reduced tumor cell proliferation and the population of mitotic cells and decreased glycolytic rate. Loss of TKTL1 did not increase sensitivity to the chemotherapeutic agents, panobinostat or carboplatin, but rather slowed DMG cell growth independently of their cytotoxic effects. These data indicate TKTL1 expression in DMG alters metabolism and proliferation and is a potential therapeutic target.

Committee: Robert M. Lober M.D., Ph.D. (Committee Chair); Thomas L. Brown Ph.D. (Committee Co-Chair); Weiwen Long Ph.D. (Committee Member); David R. Ladle Ph.D. (Committee Member); Lynn K. Hartzler Ph.D. (Committee Member) Subjects: Biomedical Research; Cellular Biology; Oncology

Doctor of Philosophy, Case Western Reserve University, 2019, Epidemiology and Biostatistics

Gliomas are the most common primary malignant brain tumor. Glioblastoma (GBM), World Health Organization (WHO) Grade IV, is the most common, aggressive, and deadly glioma and has extremely poor prognosis. Lower grade gliomas (LGG), comprised of WHO Grades II and III tumors (sometimes referred to as low and intermediate grade gliomas), are invasive and may progress to higher-grade lesions. In 2016, the WHO reclassified the definition of GBM, dividing these tumors into isocitrate dehydrogenase (IDH)-wildtype and IDH-mutant GBM, where the vast majority are IDH-wildtype. A nomogram accounts is an easily accessible tool for physicians to use on behalf of their patients for predicting survival, developing individualized cancer prognosis, and deciding the interval for follow-up and/or imaging. Sex disparities in cancer survival have been established for several cancers but have remained inconclusive for gliomas. Therefore, we aimed to (1) develop and independently validate a nomogram for patients with newly diagnosed LGG, (2) develop and independently validate a nomogram for patients with newly diagnosed IDH-wildtype GBM, and (3) determine whether sex differences exist in glioma survival. The LGG nomogram was trained using data from The Cancer Genome Atlas (TCGA) and validated using OBTS data. The GBM nomogram was trained using data from the Ohio Brain Tumor Study (OBTS) and validated using data from the University of California San Francisco (UCSF). For both nomograms, survival was assessed using Cox proportional hazards regression, random survival forests, and recursive partitioning analysis. Nomograms provide an individualized estimate of survival rather than a group estimate and can be useful tools to patients and healthcare providers for counseling patients and their families regarding treatment decisions, follow-up, and prognosis. Free software for implementing these nomograms have been developed. To assess sex differences in glioma survival, data were obt (open full item for complete abstract)

Committee: Chun Li Ph.D. (Committee Chair); Jill Barnholtz-Sloan Ph.D. (Advisor); Marta Couce M.D., Ph.D. (Committee Member); Curtis Tatsuoka Ph.D. (Committee Member) Subjects: Biostatistics; Epidemiology

Master of Sciences, Case Western Reserve University, 2017, Clinical Research

Gliomas are primary brain tumors and are challenging to treat. Traditionally, gliomas are characterized by their histologic appearance and grade. Recently, molecular data have complemented histopathologic data and provided new insights on glioma biology and behavior. Vasorin is a transmembrane protein that is commonly overexpressed in glioblastoma, the most aggressive type of glioma. The function of Vasorin is not well understood but involves regulation of apoptosis induced by inflammation or hypoxia and regulation of transforming growth factor-beta (TGF-beta) signaling. The purpose of this study was to use a bioinformatics approach to discern potential roles for Vasorin in glioma biology. Vasorin expression correlated strongly with tumor grade, glioblastoma, IDH wild type gliomas, TERT promoter mutation and unmethylated MGMT promoter. Proteomic analyses revealed potential interactions between Vasorin and developmental programs, including Notch, TGF-beta and homeobox proteins, and IDH1. Additional studies are needed to validate these findings.

Committee: Jeremy Rich (Committee Chair) Subjects: Biology; Medicine

Master of Science (MS), Bowling Green State University, 2023, Biological Sciences

Although there are many therapies against cancer, they involve devastating side effects. Hence, we need to understand the nature of cancer cells and the unique markers that exist within these cells that allow them to evade the immune system. These cell properties could be exploited to our advantage. CD47 is a cell membrane receptor protein widely expressed in most cells and is a versatile and crucial target in the tumor microenvironment for creating novel therapeutic approaches for cancer treatment. However, few studies have examined CD47 in glioma. CD47 is found on the surface of multiple cell types, and it usually protects the cells from being removed by phagocytes. It has been found that most cancer cells have high CD47 expression that prevents them from being engulfed by macrophages or activated microglial cells, essentially acting as a “don't eat me signal.” A second protein expressed on cancer cells, calreticulin (CALR), facilitates cell removal by phagocytes, serving as an “eat me signal.” In this project, we compared CD47 expression in glioma cancer stem cells (CSCs), which are negative for Hoechst 33342 nuclear staining (H-), and non-stem glioma cells (Hoechst-positive, H+) of the C6 cell line derived from a rat astrocytoma. We examined the colocalization of CALR with CD47 in both C6 cell types using immunocytochemistry and compared CALR and CD47 gene expression reported in the NCI-60 database of multiple human cancers. We found a significant difference in CD47 expression, with more CD47 in the H+ cells than the H- cells, which could imply that GSCs are more susceptible to CD47 immunotherapy. The highest expression of CD47 ( 50% above signal range) appeared to be in exosomes related to both cell types. We found a positive correlation between CD47 and CALR distribution in the H+ cells (p = 0.0204) and in both H+ and H- cells combined (p = 0.0121), suggesting that the cells might protect themselves from CALR-induced phagocytosis by increasing CD47. We al (open full item for complete abstract)

Committee: Michael Geusz PhD (Committee Chair); Julia Halo PhD (Committee Member); Paul Morris PhD (Committee Member) Subjects: Biology; Medicine

Master of Science (MS), Bowling Green State University, 2023, Biological Sciences

An important mechanism that cancer cells use to survive adverse conditions is upregulating their unfolded protein response (UPR) to secrete more chaperone proteins including calreticulin (CALR). CALR is a resident endoplasmic reticulum protein that has various functions spanning from the nucleus, cytoplasm, and up to the cell membrane. Evidence indicates that cancer cells and especially cancer stem cells (CSCs) have adapted an elevated UPR causing CALR to be produced abundantly such that some of the protein relocates to the cell membrane. At the cell membrane CALR functions as an extracellular signal to immune cells that triggers removal of the cell by microglia and macrophages through phagocytosis. This project tested whether CSCs of gliomas have higher surface CALR expression than non-stem-like cancer cells that form most of the tumor. CSCs previously identified in the C6 rat glioma cell line were further characterized by immunocytochemistry. Based on previous reports from our lab and others, we identified the CSCs in C6 rat cell cultures along with immunofluorescence imaging of cell-surface CALR. Significantly higher surface CALR expression was present in CSCs within the cell clusters forming early tumorspheres. We also extracted RNA-Seq results from the IVY Glioblastoma Atlas Project database describing gene expression for CALR and genes coding for 25 proteins closely related by function to CALR. We compared this activity across five glioblastoma tumor structures and found elevated gene expression in the Pseudopalisading Cells Around Necrosis and Microvascular Proliferation. The known prevalence of CSCs in these tumor structures and the transcriptome data agree with our C6 stem cell results indicating elevated CALR in glioma CSCs. When we induced epithelial-mesenchymal transition (EMT) in C6 cells, an event leading to invasive CSCs, surface CALR was significantly elevated in the resulting cells. We also examined by immunofluorescence the expression of ATF4, a tr (open full item for complete abstract)

Committee: Michael Geusz Ph.D. (Committee Chair); Carol Heckman Ph.D. (Committee Member); Vipa Phuntumart Ph.D. (Committee Member) Subjects: Biology; Biomedical Research; Health; Immunology; Medicine; Neurosciences

PhD, University of Cincinnati, 2022, Medicine: Pathobiology and Molecular Medicine

Malignant gliomas such as glioblastoma are highly heterogeneous with distinct cells of origin and varied genetic alterations. Analysis of TCGA databases reveals that co-mutations of PTEN and TP53 are most significantly enriched in human high-grade gliomas. Mice with PTEN/p53 ablation mediated by multilineage-expressing human GFAP (hGFAP)-promoter-driven Cre lines develop glioma but with incomplete penetrance and long latency. However, the specific cell lineages and states that are susceptible to PTEN/p53-loss-induced malignant transformation remain elusive. Unexpectedly, ablation of Pten and Trp53 in Nestin+ neural stem cells (NSCs) or Pdgfra+/NG2+ committed oligodendrocyte precursor cells (OPCs), two major cells of origin in glioma, did not induce glioma formation in mice. Strikingly, mice lacking Pten and Trp53 in Olig1+/Olig2+ intermediate-lineage precursors prior to the committed OPCs develop high-grade gliomas with 100% penetrance and short latency. The resulting tumors exhibit distinct tumor phenotypes and drug sensitivities from NSC- or OPC-derived glioma subtypes. Integrated transcriptomic and epigenomic analyses revealed that PTEN/p53-loss induced activation of oncogenic pathways including HIPPO-YAP and PI3K signaling to promote malignant transformation. Targeting the core regulatory circuitries YAP and PI3K signaling effectively inhibited tumor cell growth. Thus, our multi-cell state in vivo mutagenesis analyses suggests that transit-amplifying states of Olig1/2-intermediate-lineage precursors predispose to PTEN/p53-loss-induced transformation during gliomagenesis, pointing to subtype-specific treatment strategies for gliomas with distinct genetic alterations. The establishment of glioma mouse models is critical for defining cellular alterations and initiating processes during glioma development. I have developed a virus-induced glioma animal model with a retrovirus that overexpresses oncogenic Pdgfb or activating Ras mutant in conjunction with domin (open full item for complete abstract)

Committee: Qing Richard Lu Ph.D. (Committee Member); Vladimir Bogdanov Ph.D. (Committee Member); Timothy Phoenix (Committee Member); Jose Cancelas-Perez M.D. (Committee Member); Kenneth Campbell Ph.D. (Committee Member) Subjects: Oncology

MS, University of Cincinnati, 2022, Medicine: Cancer and Cell Biology

Microglia is the primary brain resident macrophage and is an integral component of brain development and functions including neurogenesis, angiogenesis, and synapse formation. The microglia exhibit spatial and temporal diversity in their morphology and functions. The microglial functions depend on the region and stage of development. Despite the advancement in research on microglia, the diversity profile of the microglia has yet been fully understood. Besides microglia, immune cells in the normal brain also include macrophages like the choroid plexus, meningeal, and perivascular macrophages. The role of microglia and different brain resident macrophages in normal brain physiology, neurological diseases, and brain tumors remains to be poorly defined. As the deadliest immune-privileged brain tumor, glioblastomas are largely treatment-resistant, which is in part due to the immunosuppressive tumor microenvironment. Specifically, the tumor-associated macrophages (TAM) act as a pro- and anti-tumorigenic component in GBM and are emerging as a potential target for developing anti-tumor drugs. Here, we discuss the origin, biomarkers, regulatory factors, and physiology of disease-associated microglia/macrophages. A better understanding of disease-associated microglia/macrophages and tumor microenvironment may facilitate the development of effective treatment strategies for neurodegenerative diseases and brain cancers.

Committee: Qing Lu Ph.D. (Committee Member); Ty Troutman Ph.D. (Committee Member); Ziyuan Guo Ph.D. (Committee Member); Theresa Alenghat V.M.D. Ph.D. (Committee Member) Subjects: Cellular Biology

Doctor of Philosophy, Case Western Reserve University, 2020, Biomedical Engineering

Glioblastoma multiforme (GBM) is resilient to the current standard of care treatment of surgical resection followed by concurrent radiotherapy and temozolomide (TMZ) chemotherapy. GBM patient responses are poor and variable, resulting in more than 90% tumor recurrence and grim survival. The high mortality of GBM is attributed to its invasive peripheral growth, partially intact blood-brain barrier (BBB), regions of hypoxia, and high cellular heterogeneity that includes brain tumor initiating cells (BTICs) and immunosuppressive cells. These features of GBM work together to restrict the delivery of drugs throughout the tumor, suppress immune recognition of tumor cells, and facilitate tumor progression. Nanoparticles are well-suited to address limitations associated with the treatment of GBM by enhancing drug delivery to the tumor and reducing side effects. The overall objective of the work in this dissertation is to develop systemically administered nanoparticles that overcome barriers to drug distribution and cellular heterogeneity in GBM to improve therapeutic responses. In murine GBM models, the effective delivery of Doxorubicin (DOX) chemotherapy, BTIC inhibitor, and immune-stimulating agonists were evaluated using two distinct mesoporous silica nanoparticles (MSNs): 1) the Fe@MSN particle and 2) the immuno-MSN particle. First, drug release from the Fe@MSN particle was triggered using an external radiofrequency (RF) field to enhance the distribution of DOX and/or BTIC inhibitor across the partially intact BBB and into the tumor interstitium. The effective delivery of drugs facilitated by Fe@MSN particles translated into suppressed GBM growth, depleted stem-like cell phenotypes in hypoxic regions, and prolonged or cancer-free survival. Second, towards further improving GBM treatment strategies, the immuno-MSN particle delivered immune-stimulating agonists to dysfunctional immune cells in GBM to reverse the effects of immunosuppression. Immuno-MSN particles facilitat (open full item for complete abstract)

Committee: Efstathios Karathanasis Ph.D. (Advisor); Agata Exner Ph.D. (Advisor); Dominique Durand Ph.D. (Committee Chair); Jennifer Yu M.D., Ph.D. (Committee Member) Subjects: Biomedical Engineering; Immunology

Doctor of Philosophy (Ph.D.), University of Dayton, 2020, Biology

The interaction between the tumor cells and the surrounding normal cells constitutes the Tumor microenvironment (TME). The Toll-like Receptor (TLR), Jun N-terminal Kinase (JNK), and Tumor Necrosis Factor (TNF) produce inflammatory components in the TME and are thought to play a critical role in tumor survival and progression. However, the exact nature and mechanism of interactions within the TME remain poorly understood. These core inflammatory pathways are conserved in Drosophila. As 90% of tumors are epithelial in origin, we used an epithelial tumor model in the wing imaginal discs of Drosophila melanogaster to study the interaction of these key inflammatory pathways in the TME. We established a new TME model by creating FLP-out clones of oncogenic forms of Yki or RasV12 in polarity deficient (scribble mutant) cells marked by GFP surrounded by normal cells. These mosaic clones allow us to test changes in intercellular and signaling interactions within the tumor, surrounding its microenvironment and in distant normal cells. We hypothesized that crosstalk between the TLR and TNF pathways in the TME promotes tumor survival and progression through JNK pathway. We observed Drosophila IκB Cactus (TLR component) is upregulated in the tumor cells and downregulating Cactus negatively affect tumor growth (JNK expression) and invasion (JNK target, MMP1 expression). The genetic epistasis experiments between JNK and TNF revealed that downregulation of the TNF receptors in the tumor does not affect the metastatic abilities of the tumor cells. Further, we report Hippo pathway effector, Yki as the ultimate regulator, that transcriptionally regulates Cactus expression which in turn mediates tumor promoting JNK signaling in the tumor cells. We also studied the tumorigenesis in the Drosophila glioma model generated by co-activation of the epidermal growth factor receptor (EGFR) and Phosphatidylinositol 3-kinase (PI3K) pathway. Glioma is an aggressive form of adult brain tumor (open full item for complete abstract)

Committee: Madhuri Kango-Singh Ph.D. (Advisor); Amit Singh Ph.D. (Committee Member); Mark Nielsen Ph.D. (Committee Member); Pothitos Pitychoutis Ph.D. (Committee Member); Ichiro Nakano M.D., Ph.D. (Committee Member) Subjects: Biology; Biomedical Research; Experiments; Genetics; Immunology; Molecular Biology; Oncology

Doctor of Philosophy, Case Western Reserve University, 2019, Pathology

Malignant gliomas are one of the gravest diagnoses to deliver in oncology. Recent integrative multiomics studies of multicenter cohorts have deepened our understanding of complex disease processes, but also highlighted the daunting task of crippling these aggressive neoplasms with extensive inter- and intratumoral heterogeneity. The cancer stem cell hypothesis enriched opportunities to test therapeutic vulnerabilities in contexts that best phenocopy the original diseases. Among biological outputs, metabolic profiles resolved by global metabolomics remain difficult to analyze in high throughput studies. Nevertheless, integration of metabolic fluxes and cellular states informed by epigenetic and transcriptional unearthed critical metabolic dependencies in the cancer stem cell state. Advances in preclinical studies and genomics have informed promising clinical trials. Developments in immunotherapy hold further promise in improving patient outcomes. Closer integration of clinical observations and preclinical studies will uncover new opportunities to find therapeutic targets and improve treatment protocols.

Committee: Jeremy Rich M.D., M.H.Sc., M.B.A (Advisor); Parameswaran Ramakrishnan Ph.D., M.S (Committee Chair); Nima Sharifi M.D. (Committee Member); Zhenghe Wang Ph.D. (Committee Member); Clive Hamlin Ph.D. (Committee Member) Subjects: Biomedical Research; Medicine; Neurology; Oncology

Doctor of Philosophy (Ph.D.), Bowling Green State University, 2019, Biological Sciences

Circadian rhythms control cancer cell behavior in tumors and in vitro. Epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) are two key events of metastasis that lead to cancer progression and aggressiveness. Studies suggest that genes controlling circadian rhythms also regulate EMT, which generates mesenchymal cells (M-cells) with cancer stem cell (CSC) properties through dedifferentiation. MET enables post-EMT cells to differentiate back to epithelial cells at metastatic sites and initiate secondary tumors. If a circadian clock can be identified in M-cells, then it could be manipulated pharmacologically to more effectively target the cells with novel anticancer agents specific to CSCs or cause their differentiation into more easily treated and less aggressive cells before metastasis occurs. We tested for any role of circadian clocks in EMT and MET events by using cancer cell lines from two different tissues. We compared C6 rat glioma cells that have a well-established circadian clock with MCF-7 human breast tumor cells that are considered lacking a functional clock. EMT was induced in cell cultures by exchanging standard serum-containing medium (SM) with stem cell medium (SCM), a non-serum medium containing specific growth factors promoting CSC survival. Single-cell behavior and morphological states were quantified microscopically through time-lapse imaging. Expression of EMT markers ZEB1 and TWIST, mesenchymal markers vimentin and PDGFRA, and stem cell markers OCT4, nestin, MSI1 and CD133 were validated by immunocytochemistry. Both C6 and MCF-7 cultures showed circadian oscillations in the population size of post-EMT M-cells. MET was then induced by returning the cultures to SM from SCM. MET events observed in glioma CSCs clustered significantly at a particular phase of the circadian cycle. The cellular microenvironment also influenced migration properties of C6 cells with SM promoting faster closure in a standard wound healing assa (open full item for complete abstract)

Committee: Michael Geusz PhD (Advisor); Lynn Darby PhD (Other); Paul Morris PhD (Committee Member); Vipaporn Phuntumart PhD (Committee Member); Amit Tiwari PhD (Committee Member) Subjects: Biology; Biomedical Research; Cellular Biology; Health Sciences; Medicine; Molecular Biology; Neurosciences; Oncology; Pharmacology

Doctor of Philosophy, Case Western Reserve University, 2018, Epidemiology and Biostatistics

Glioma is the most commonly occurring malignant brain tumor in the United States, with highest incidence in males and persons over 60. There are no well validated risk factors for these tumors that explain a large proportion of cases. To date, genome-wide association studies have identified 25 validated risk loci which explain 30% of overall heritable risk. The goal of this dissertation is to utilize demographic differences in incidence to increase power for detection of variants with varying effects by age and sex. Sex-stratified logistic regression models were used to generate betas, standard errors, and p-values. A significant association was detected at 7p11.2 (EGFR), a SNP previously associated with glioma risk, in males only. A previously identified intergenic SNP in 8q24.21 had an effect size in females that was approximately 2-fold that of males. Additionally, I identified a new large region on 3p21.31 with significant association in females only. Gene-specific p-values were generated by combining single SNP p-values for all SNPs within gene boundaries using three different scoring algorithms. Genes were prioritized that were identified by at least 2 of 3 algorithms. Pathway scores were generated by combined gene-specific p-values using Pascal. Significant associations were found in genes containing SNPs previously associated with glioma (EGFR, and TERT) which remained nominally significant after conditioning on known SNPs, suggesting these regions may contain additional sources of genetic risk. There were significant associations in the telomere maintenance pathway in both sexes. Age-stratified logistic regression models were used to generate betas, standard errors, and p-values. A previously identified SNP in 8q24.21 was found to have a significant effect in the youngest age group, with no detectable effect in older age groups. An examination of glioblastoma patients within The Cancer Genome Atlas found higher prevalence of `LGG'-like characteristics withi (open full item for complete abstract)

Committee: Jill Barnholtz-Sloan Ph.D. (Advisor); William Bush Ph.D. (Committee Chair); Frederick Schumacher Ph.D. (Committee Member); Farren Briggs Ph.D. (Committee Member) Subjects: Epidemiology

Master of Science (MS), Bowling Green State University, 2017, Biological Sciences

Circadian rhythms influence tumorigenesis and the rate of tumor growth. A fundamental question is whether the circadian timing system alters cancer cell properties through neurosecretion at a neural-tumor synapse (NTS). The master clock of the circadian system is located in the suprachiasmatic nucleus (SCN) of the hypothalamus and regulates multiple circadian rhythms in behavior and physiology including rhythms in neurotransmitter secretion in the brain. Like cells throughout the body, many cancer cell types also contain a circadian clock. The clock can modulate responsiveness of cancer cells to chemotherapeutic treatments. The question addressed in this study was whether glioma cancer cells show evidence of a circadian rhythm in their response to serotonin (5-HT), a neurotransmitter that could provide signaling of timing information through the NTS. This potential control of gliomas by the circadian clock might be used clinically to improve the timing and effectiveness of anticancer treatments. A dose-response curve was created to find an optimal dosage of 5-HT for further testing. This dosage (10 µM) significantly suppressed area and width of C6 rat glioma cells while also increasing cell circularity, resulting in a morphology resembling cells that are more invasive and motile following an epithelial-to-mesenchymal transition (EMT). A second neurotransmitter, glutamate, produced an opposite effect on area with increasing dosage. The optimized 5-HT treatment was then given to C6 cells at four-hour intervals for 48 hours to determine whether there are circadian effects of 5-HT on cell morphology. Before treatment, circadian clocks within the cells were synchronized using forskolin. The suppression of average cell area and width by 5-HT showed a circadian rhythm with a peak responsiveness just before the minimum of the previously described daily rhythm in 5-HT levels of rat brain. Treatments with 5-HT also caused a slowing of ongoing oscillations in intracellular Ca2 (open full item for complete abstract)

Committee: Michael Geusz Dr. (Advisor); Verner Bingman Dr. (Committee Member); Robert Huber Dr. (Committee Member) Subjects: Biology; Cellular Biology; Experiments; Health Sciences; Molecular Biology; Morphology; Neurobiology; Neurosciences; Oncology

PhD, University of Cincinnati, 2017, Medicine: Systems Biology and Physiology

Background: Malignant gliomas currently have a poor prognosis as recurrence remains nearly inevitable despite aggressive treatment. Gliomas are histologically graded as I-IV and the grade IV glioblastoma (GBM) is the most common and lethal malignant glioma with a median survival time of just over one year. Primary GBM arises spontaneously while secondary GBM develops from a recurrent, progressive, lower grade tumor. While primary and secondary GBM are histologically indistinguishable, their genetic profiles and molecular characteristics differ substantially. Primary GBM usually displays amplification or mutation of at least one receptor tyrosine kinase, most often the epidermal growth factor receptor (EGFR). Secondary GBM is typified by the presence of a mutant isocitrate dehydrogenase 1 (IDH1) allele and rarely exhibits EGFR amplification. Despite the striking differences in the molecular development of primary and secondary GBM, they are currently treated with the same general strategy which usually involves surgical resection, radiation therapy, and treatment with temozolomide, a DNA alkylating agent. While this is the most effective treatment plan currently available, it almost universally results in fatal disease recurrence. Objective: The objective of this dissertation was to further our understanding of treatment resistance in adult gliomas and identify targetable vulnerabilities conferred by the genetic characteristics of the tumor to improve treatment efficacy. Results: To evaluate GBM-specific mechanisms of resistance to the EGFR tyrosine kinase inhibitor, gefitinib, EGFR-overexpressing U87 cells were treated with increasing doses of gefitinib. RNA sequencing of gefitinib resistant cells revealed overexpression of the receptor tyrosine kinase ROS1, which was confirmed in multiple resistant clones by western blot. A cell viability assay showed gefitinib resistant cells to be sensitive to treatment with a combination of gefitinib and a smal (open full item for complete abstract)

Committee: El Mustapha Bahassi Ph.D. (Committee Chair); Zalfa Abdelmalek Ph.D. (Committee Member); Nelson Horseman Ph.D. (Committee Member); Carolyn Price Ph.D. (Committee Member); Peter Stambrook Ph.D. (Committee Member); Yana Zavros Ph.D. (Committee Member) Subjects: Oncology

Master of Sciences, Case Western Reserve University, 2017, Systems Biology and Bioinformatics

Motivation: The prognosis of low-grade-glioma (LGG) patients is very poor. Identifying subnetworks related to LGG can better describe the genetic make-up of the tumor. Methods: n-Node Subnetwork Enumeration Algorithm (nSEA) was developed to identify significantly dysregulated subnetworks. We utilized a filtered protein network to enumerate n-node subnetworks exhaustively and score each subnetwork to carry out feature selection. These subnetwork seeds were expanded to identify tumor-specific subnetworks. Clustering these subnetworks provided patient groups with different subnetwork states. Results: We identified 92 subnetwork features, 8 subnetwork groups and 5 patient groups. A new patient group was identified with favorable outcomes. By decision tree modeling, the new group were characterized as down-regulated MAPK/B-Raf pathway and up-regulated Notch pathway. It had fewer mutations of candidate genes, hypomethylation of NIPBL and hypermethylation of KALRN. Conclusions: These results could provide opportunities for improved treatment options and personalized interventions of LGG.

Committee: Gurkan Bebek (Committee Chair); Mark Cameron (Committee Member); Jean-Eudes Dazard (Committee Member) Subjects: Bioinformatics; Oncology

Master of Science, University of Toledo, 2016, Chemistry

Glioma is a type of tumor which originates in the glial cells that surround and support neurons in the brain. According to the Central Brain Tumor Registry of the United States (CBTRUS), 29% of all central nervous system tumors diagnosed are gliomas. Rat glioma models and cell lines reflect high-grade brain tumors. Proteomics is commonly used to discover novel glioma biomarkers and study drug-protein interactionsin rat glioma models and cell lines. Proteomic studies are important because they provide a wealth of information about biochemical properties of brain tumors, and assist in discovery and validation of glioma biomarkers and drug targets. Here, proteomic techniques were used to explore the differences in the proteomes of malignant rat glioma 2 (RG2) cells in the presence and the absence of a sulphur-containing drug, lanthionine ketimine ethyl ester (LKE). In 2010, Hensley et al. synthesized LK from 3-bromopyruvate and L-cysteine hydrochloride as described by Cavallini et al., and LKE was synthesized using L-cysteine ethyl ester. It was shown that LKE promotes neurite elongation in neurons at nanomolar concentrations, protects neurons against oxidative stress, and suppresses microglial activation. Recently, Hensley et al. have analyzed the expression of proteins in RG2 cells that were treated with LKE using mostly immunoblotting techniques. A few proteins that are involved in the process of autophagy (cellular recycling) in RG2 cells were influenced by LKE. The objective of this project was to apply proteomic techniques and mass spectrometry (MS) to further study the differences in the protein expression levels in LKE-treated and untreated RG2 cells. Electrophoretic methods (e.g., SDS-PAGE), chromatographic techniques (e.g., nano-HPLC), and MS (e.g., MALDI-MS/MS and ESI-MS/MS) were used to study proteins and phosphoproteins from RG2 cells. Using nano-HPLC-MALDI-MS/MS, 22 proteins were identified from LKE-treated RG2 cells, and 18 proteins were ident (open full item for complete abstract)

Committee: Dragan Isailovic (Committee Chair); Donald Ronning (Committee Member); Jon Kirchhoff (Committee Member) Subjects: Chemistry

Doctor of Philosophy (Ph.D.), Bowling Green State University, 2014, Biological Sciences

Increased cancer risk is linked to disruption of circadian rhythms. Cancer stem cells (CSCs) are a known cause of cancer aggressiveness, but their circadian properties have not been described. In this study we describe the circadian properties of C6 rat glioma tumorspheres and MCF-7 human breast CSCs. We discovered circadian rhythms in gene expression within C6 glioma tumorspheres enriched in CSCs and found that the circadian clock is particularly robust in medium lacking any growth factors. A method is introduced for identifying individual CSCs in culture for single-cell analysis. CSCs in monolayer, attached cell cultures failed to show a circadian rhythm in nuclear localization of mPER2 protein, suggesting that cell interactions or the tumor-like microenvironment within tumorspheres enable circadian timing. The MCF-7 cancer cell line, derived from an epithelial breast tumor, has been widely studied because of its aggressiveness and high percentage of CSCs. Although several cancer cell lines have distinct circadian rhythms in gene expression, the reason why many other lines apparently lack circadian clocks remains unclear. Similarly, circadian rhythms of cells within tumors are also often poorly organized or absent. Considering the cell heterogeneity of cancer cell lines, including CSCs within these lines, it seemed likely that some of the cells could retain a functional circadian clock. To test this idea, we probed the circadian properties of MCF-7 cultures with a reporter gene that expresses a functional mPER2 protein fused with firefly luciferase under the control of the mouse Per2 promoter. MCF-7 cells grew as small clusters in medium containing fetal bovine serum or one containing growth factors stimulating CSC proliferation. The percentage of clusters expressing the mPer2 gene was surprisingly high, and at least 60% of these expressed a circadian rhythm. Reporter gene expression and Per2 mRNA were elevated in response to growth factors that preven (open full item for complete abstract)

Committee: Michael Geusz Ph.D. (Advisor); Rogers Scott Ph.D. (Committee Member); Moris Paul Ph.D. (Committee Member); Maltese William Ph.D. (Committee Member); Anzenbacher Pavel Ph.D. (Other) Subjects: Biology; Cellular Biology; Molecular Biology

Doctor of Philosophy, Case Western Reserve University, 2014, Molecular Medicine

Glioblastoma (GBM) remains one of the most lethal human cancers, with conventional treatment offering only palliation. Like all cancers, GBMs display the Warburg effect, a preferential utilization of aerobic glycolysis for energy supplies. This metabolic shift reduces the cells oxygen dependence and provides a steady supply of anabolic material yet requires a steady supply of glucose. In this dissertation, I show that nutrient restriction contributes to tumor progression by enriching for glioblastoma stem cells (GSCs), the most treatment-resistant and regenerative of GBM cells. This enrichment is due to both preferential GSC survival and adaptation of non-GSCs through acquisition of stem-like features. GSCs outcompete for glucose uptake by co-opting the high-affinity neuronal glucose transporter isoform, type 3 (GLUT3). In the normal brain, glucose is an essential fuel for neuronal metabolism; yet vascular glucose delivery is physiologically stymied by the blood brain barrier. In response to this, neurons express GLUT3, allowing steady glucose uptake from a glucose-poor microenvironment. GSCs preferentially express GLUT3 and targeting GLUT3 inhibits GSC growth and tumorigenic potential, suggesting GLUT3 is a marker of CSCs. GLUT3, but not GLUT1, correlates with poor survival in brain tumors and other solid cancers in clinical patient datasets; thus, CSCs in many cancer types may extract nutrients with high affinity. Given the restricted expression profile of GLUT3 in non-malignant tissue and the critical role of GLUT3 in CSC biology, therapeutic targeting of GLUT3 may prove to be a viable treatment across multiple cancer types.

Committee: Jeremy Rich MD (Advisor); Xiongwei Zhu PhD (Committee Chair); Jan Jensen PhD (Committee Member); Robert Silverman PhD (Committee Member); Robert Weil MD (Committee Member); Anita Hjelmeland PhD (Committee Chair) Subjects: Biology; Biomedical Research; Cellular Biology; Molecular Biology; Neurobiology

MS, University of Cincinnati, 2007, Allied Health Sciences : Genetic Counseling

Neurofibromatosis type 1 (NF1) frequently involves the central nervous system (CNS), but there is extreme variability in the expression of CNS complications, even within families with the same causative mutation. It has been proposed, but not proven, that CNS complications tend to cluster within individuals with NF1. We conducted a retrospective medical record review to determine if CNS abnormalities occur more frequently in individuals with NF1 who have an optic pathway glioma (OPG). Seventy-two (72) subjects with OPG and 189 without OPG were screened for CNS complications. OPGs were found to be associated with a diagnosis of additional CNS tumors and T2 hyperintensities, supporting a difference in CNS complications manifested by patients who develop OPG versus those who do not. Additional understanding of the biology of brain lesions in NF1 is necessary to further elucidate why patients with OPG are more likely to develop additional CNS lesions.

Committee: Dr. Elizabeth Schorry (Advisor) Subjects:

MS, University of Cincinnati, 2006, Medicine : Epidemiology (Environmental Health)

Neurofibromatosis type-1 (NF-1) is characterized by widely variable clinical features. These patients are at an increased risk of developing wide variety of benign and malignant neoplasia. Optic pathway glioma (OPG) occurs in almost 20% of patients. There is poor genotype-phenotype correlation and limited data to quantify the risk of tumor development when preceded by specific clinical features. We hypothesize that presence of specific clinical features in NF-1 patients can predict their risk for subsequent OPG development. Our data suggests that patients with ages 4 years or less at diagnosis of NF-1, presence of T-2 hyperintense lesions on imaging (UBOs), developmental delay and seizures appear to be at an increased risk of developing subsequent OPG compared to the patients who do not have these clinical features. Race appears to have a disease modifying effect on progression. Patients with these risk factors may be stratified as “high risk” and may benefit from frequent scanning and follow up for early detection of OPG.

Committee: John Perentesis MD (Committee Chair); Bandana Chakraborty Dr.PH (Other); Ranajit Chakraborty PhD (Other); Elizabeth Schorry MD (Other) Subjects: Health Sciences, Oncology