UC Irvine

ABSTRACT OF THE DISSERTATION

Radiation- and Chemotherapy-Induced Cognitive Deficits: Neural-Derived Extracellular Vesicles as Translational Therapiesby Sarah Mae Smith Doctor of Philosophy in Environmental Health Sciences University of California, Irvine, 2022 Professor Charles L. Limoli, Chair

Approximately 24,000 patients are diagnosed with brain tumors each year, which represents an estimated 166,000 people living with brain cancer as of 2015. Radiotherapy constitutes one of the principle therapies for primary and metastatic brain tumors along with chemotherapy and surgery, with roughly 200,000 patients receiving brain radiation treatment each year in the United States. One of the most common and most damaging iatrogenic effects of cancer treatment is cognitive dysfunction, including impairments in working memory, learning ability, executive function, and attention. The deleterious effects of cranial ionizing radiation exposure are exacerbated by concomitant use of chemotherapeutic agents that elevate neurotoxicity. While the mechanisms underlying radiation-induced cognitive dysfunction have not yet been elucidated, much of the underlying pathology believed to be contributory is related to decreased hippocampal neurogenesis, increased neuroinflammation, microvascular injury and alterations in neuronal structure that disrupts dendritic morphology, spine density and synaptic proteins. Stem cell-based transplantation strategies have also shown considerable promise in ameliorating the negative effects of cranial irradiation. My initial work in the Limoli lab sought to examine the impact of bilateral and unilateral hemisphere transplantations of human neural stem cell (hNSC) and hNSC-derived extracellular vesicles on the structural integrity of hippocampal neurons in the irradiated rodent brain, in order to ascertain the extent and range of the beneficial effects of cell-based therapies as a function of distance from the transplant site (which has important therapeutic implications for dosing and administration). We found that stem cell transplantation decreases the number of activated microglia at one month post-irradiation, signifying a reduction in neuroinflammation. In addition, grafted stem cells preserved host neuronal structure one month and four months after irradiation. Stem cell transplantation also allayed the increased expression of the major synaptic scaffolding protein PSD-95 caused by irradiation. Further, as clinical utility of stem cell transplantation strategies to offset normal tissue damage caused by irradiation may be limited by the downstream potential for teratoma formation and immune rejection, we also attempted to circumvent such caveats by evaluating the promise of using transplanted human neural stem cell-derived extracellular vesicles in place of stem cells. We showed that unilateral transplantation of both stem cells and extracellular vesicles 48 hours post-irradiation signals for the renormalization of synaptic proteins such as PSD-95 in the contralateral hippocampus, and found that unilateral transplantation has the same protective effects on neuroinflammation, dendritic morphology, and spine density as bilateral transplantation. My later studies evaluate the impact of GABAergic neuron-derived extracellular vesicles on irradiation- and chemotherapy-associated neurocognitive deficits associated with clinically-relevant treatments, including fractionated radiation and the chemotherapeutic agent temozolomide (TMZ). We first examined whether retro-orbital injection of extracellular vesicles (EV) derived from GABAergic neurons can alleviate radiation-associated deficits in cognition at one month following treatment. GABAergic neuron-derived EVs are proposed to improve cognition after cancer therapy through providing an inhibitory impetus to compensate for irradiation- and chemotherapy-induced hippocampal hyperexcitation, as is apparent in preliminary electrophysiology studies in irradiated animals. Our behavioral data suggests that fractionated irradiation has deleterious neurological effects as determined by performance on hippocampus-, prefrontal cortex- and amygdala-dependent behavioral tasks, and that retro-orbital injection of GABAergic EVs attenuates some of these effects and rescues cognition. Intriguingly, these benefits were not replicated with the application of glutamatergic neuron-derived EVs, which did not reverse the observed decrements in cognitive function after irradiation. In exploring the neurobiochemical basis for this rescue in cognition on neuronal morphology, synaptic integrity, and neurotrophic factors, we found that dendritic spine density in the dentate gyrus of the hippocampus is significantly reduced following irradiation, but statistically similar to controls following the application of retro-orbital GABAergic EVs. We also determined that irradiation reduced levels of neurotrophins in the hippocampus, but retro-orbital injections of GABAergic EVs ameliorated this decrease. We built upon these studies by next exploring whether retro-orbital injection of GABAergic neuron-derived EVs can ameliorate the adverse effects associated with a systemic insult to the brain, intraperitoneal injections of the chemotherapeutic agent temozolomide (TMZ), in the context of a clinically-relevant treatment protocol also including fractionated radiation. Temozolomide is an alkylating agent that crosses the blood-brain barrier and is used primarily to treat glioblastoma. Again, several of the behavioral tasks show neurocognitive decrements following the TMZ+IRR protocol, which were improved with the GABAergic EV injections. Likewise, tissue from the hippocampi of these animals was analyzed for neurotrophic factor levels, which revealed that glial cell-derived neurotrophic factor was reduced after irradiation but improved in the GABAergic EV cohort. Finally, we have been evaluating the optimal timecourse for delivery of GABAergic neuron-derived EVs being employed to rescue cognitive decrements associated with irradiation by examining whether retro-orbitally injecting GABAergic neuron-derived EVs the week before the first fractionated radiation dose could result in a preemptive protective effect. We have identified that the protective paradigm of GABAergic EV application mitigates radiation-induced cognitive injury and enhances the quantity of neurotrophins in the hippocampus relative to irradiated animals.