Daniela A. Bota, Ph.D. - US grants

DESCRIPTION (provided by applicant): This application is focused on the study of the mechanisms by which chemotherapeutic drugs impact the cognitive function of cancer patients. Millions of people are diagnosed with cancer every year, and more than 60% of these now survive for 20 years, with severely diminished quality of life due to treatment-induced cognitive impairments. I am a fully trained neuro-oncologist, with a solid background in the lab, where I studied both mitochondrial and brain tumor biology. Over the past years, I recognized the importance of chemotherapy- induced cognitive defects and became passionate about finding the biological explanations for this major pathology. A. The specific aims of this proposal will focus on two DNA-targeting compounds that are widely used in oncology - cisplatin and temozolomide;we aim to explore the mechanisms by which these drugs provoke learning and memory defects. The ultimate goal of these studies would be to prevent or counteract these adverse effects. Aim 1) To determine the relative vulnerability of neural progenitor cells and mature neurons to clinically-relevant doses of cisplatin and temozolomide, using in vitro systems. Aim 2) To determine the mechanism by which cisplatin and temozolomide injure neuronal cell populations, testing if these mechanisms involve mitochondrial dysfunction. Aim 3) To examine the effects of acute and chronic graded cisplatin and temozolomide doses on vulnerable neuronal populations in vivo, and to study the role of this cellular injury in learning and memory defects. Aim 4) To examine if chemotherapy-induced cognitive deficits can be ameliorated by an intervention that augments neurogenesis and dendritic spine growth / stability, i.e., BDNF enhancement using ampakines. B. My career plan is to conduct the research proposed in the nurturing environment offered by the University of California, Irvine. This includes my mentor, an internationally known neuroscientist/clinician (Prof. Tallie Z. Baram), my co-mentor, an international leader in oncology (Prof. Frank Meyskens), my supportive chair and dean, protected research time, and excellent collaboration from my clinical colleagues. My immediate career goal is to immerse myself in cutting-edge neuroscience that will facilitate my understanding of the mechanisms by which cancer treatments impact the brain. This will be accomplished via basic neuroscience courses, hands-on methods, lab meetings, national meetings and intensive self-study. My long-term goals are to assume a senior role in my lab, acquire the skills necessary for productive publications, enlarge my research group, apply successfully for R01 funding and receive tenure. Finally, I want to enhance my involvement in the neuroscience community and to generate an independent, creative, translational research program. In summary, my goal is to develop cutting-edge bench-to bedside research focused on the biological mechanisms underlying the prominent cognitive deficits caused by chemotherapy, and to reverse this process. This grant will provide me with the necessary funding and mentorship to become a successful, independent researcher. PUBLIC HEALTH RELEVANCE: Learning and memory defects induced by chemotherapeutic drugs are rapidly emerging as major clinical problem, as one and a half million people are diagnosed with cancer every year in the US, and more than 60% of these now survive for 20 years or more. Chemotherapeutic drugs may affect cognitive function via several potential mechanisms, such as killing sensitive neural progenitor cells (stem cells) or injuring existing neurons, especially the vulnerable parts of brain cells that are involved in learning and memory formation (dendrites and dendritic spines). The specific aims of this proposal focus on two medications used widely in cancer care and explore the potential mechanisms by which they provoke learning and memory defects, with the ultimate goal to prevent or counteract these adverse effects of cancer therapy with interventions that can be used in the clinic.

PROJECT SUMMARY Glioblastoma (GBM) is the most aggressive primary brain tumor with a two years survival rate of less than 50% following surgical resection, radiation, and chemotherapy. Recurrence is nearly universal after the first-line treatment, and there is currently no therapy proven to prolong survival after tumor recurrence. Thus, there is an urgent need for more effective GBM therapies. The overarching goal of this project is to further develop and validate new chemotherapeutic agents for the treatment of GBM. GBM's resistance to radiation and chemotherapy heavily correlates with extensive hypoxia-induced, mitochondria-dependent phenotypic changes such as glycolytic respiration, decreased the ability to undergo apoptosis and extensive invasiveness. Mitochondrial LonP1 is an ATP-stimulated protease, directly up-regulated by HIF-1?. LonP1 is overexpressed in human malignant gliomas and its elevated expression levels are associated with high glioma tumor grade and poor patient survival. Therefore, regulation of mitochondrial function by inhibiting LonP1 protease could represent a novel approach for GBM and potentially other fast-growing malignancies which heavily depend on hypoxic adaptation. The proposed project is based on our published and preliminary results obtained from in vitro (cell- based) studies with LonP1 inhibition using siRNA and the inhibitor compounds CC4 and BT317 and in vivo LonP1-overexpression xenograft models studies. BT317 is a small molecule compound, able to cross the blood- brain barrier and to achieve promising concentrations in the brain. BT317 is highly effective in inducing cell death in multiple glioma lines and patient-derived glioblastoma stem cell cultures, with an IC50 value of 60-100 µM (temozolomide ? the main FDA approved therapy and has minimal toxicity in normal lines. identifying BT317 as a potentially new therapy for this universally fatal disease. In this project, we propose to: (1) examine the effect of mitochondrial LonP1 knockout in distinct patient-derived primary glioma stem-like cells (GSC), glioblastoma cell lines and xenograft models, (2) identify microenvironment cues and LonP1-induced mitochondrial changes that drive GSC invasiveness, and (3) examine the drug-target inhibition and molecular mechanisms for anti- cancer efficacy of the LonP1 inhibitor, BT317. The studies outlined here are the first to explore a very promising avenue ? mitochondrial Lon protease inhibition ? as a treatment for GBM.

1 Chemotherapy-related cognitive impairment (CRCI, chemobrain), chemotherapy-induced peripheral neuropathy 2 (CIPN) and gait changes are debilitating side-effects of cancer treatment with platinum agents (e.g., cisplatin), 3 taxanes, and vinca alkaloids. Cisplatin is widely used as a chemotherapeutic agent to treat ovarian malignancies. 4 Over 70% of women report experiencing CRCI, CIPN and/or falls during treatment or after completion, impairing 5 their quality of life. These neurotoxic impairments can also compromise treatment with cisplatin, influencing 6 disease progression. Currently, there are no FDA-approved clinical interventions for the treatment of CRCI and 7 CIPN. Mechanistically, cisplatin-induced neuronal toxicity derives from nuclear and mitochondrial DNA damage, 8 and oxidative stress, which induce the activation of the mitogen-activated protein kinases (MAPK), p38MAPK 9 and c-Jun N-terminal kinase (JNK), leading to neuronal apoptosis. Our preliminary data show that in vitro 10 pharmacological inhibition with small molecule inhibitors, i.e., neflamapimod for p38MAPK and SP600125 for 11 JNK, prevents cisplatin-induced reduction in dendritic spine branching and density. Based on these data, we 12 hypothesize that inhibition of the p38MAPK/JNK pathways will prevent cisplatin-induced neuronal 13 apoptosis and damage, leading to attenuation of cognitive impairments, gait changes, and neuropathic 14 pain associated with CRCI and CIPN. In this project, we propose to determine if: (1) cisplatin-induced p38 15 MAPK/JNK signaling underlies structural and functional neuronal damage, using in vitro pharmacological 16 inhibition and siRNA silencing; (2) neflamapimod and SP600125 prevent cisplatin-induced neuropathy and gait 17 alterations in the ID8 syngeneic epithelial ovarian cancer in C57BL/6 mice and the transgenic breast cancer 18 model C3TAg in FVBN mice; and (3) cisplatin-induced neurotoxicity is attenuated by p38MAPK/JNK inhibition 19 without compromising its anti-cancer activity. Our Approach includes in vitro analysis of 2 separate neuronal cell 20 lines, behavioral analysis using sensory testing for CIPN, testing of cognitive impairment, and novel 21 MouseWalker for gait changes in female mice using the two mouse cancer models. The proposed studies will 22 demonstrate the role of the p38MAPK and JNK in cisplatin induced CRCI/CIPN, and translational potential for 23 novel strategies to treat CRCI and CIPN. Due to health disparities, women suffer more disproportionately from 24 cancer and pain-related treatment than men. Therefore, testing our hypothesis in female mice is expected to 25 significantly advance the understanding and treatment of cisplatin-induced neurotoxic side effects and improve 26 the quality of life for women with cancer. Nevertheless, we expect that these findings may also apply to cisplatin- 27 induced neurotoxicity in males and to other cancers than ovarian and breast cancers. 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58