Development of a Single-Neurosphere Culture to Assess Radiation Toxicity and Pre-Clinical Cancer Combination Therapy Safety

SIMPLE SUMMARY: Exposure to radiation during nuclear catastrophes, natural sources such as space travel, or during cancer treatment can lead to central nervous system toxicity. The high sensitivity of neural progenitor cells (NPCs) to radiation poses a significant obstacle to dose escalation for the treatment of brain cancer such as glioblastoma, yet studies suggest that higher (dose-escalated) radiation therapy (RT) could enhance local control. Unfortunately, this is presently infeasible due to the toxicity to neighboring normal tissue; thus, there is a critical need to understand the cellular mechanisms that determine the extent of radiation-induced toxicity in normal NPCs, to develop CNS toxicity radiation counter measures or develop new safe brain cancer chemo–radiation therapy strategies. ABSTRACT: Radiation therapy (RT) is a crucial treatment modality for central nervous system (CNS) tumors but toxicity to healthy CNS tissues remains a challenge. Additionally, environmental exposure to radiation during nuclear catastrophes or space travel presents a risk of CNS toxicity. However, the underlying mechanisms of radiation-induced CNS toxicity are not fully understood. Neural progenitor cells (NPCs) are highly radiosensitive, resulting in decreased neurogenesis in the hippocampus. This study aimed to characterize a novel platform utilizing rat NPCs cultured as 3D neurospheres (NSps) to screen the safety and efficacy of experimental drugs with and without radiation exposure. The effect of radiation on NSp growth and differentiation was assessed by measuring sphere volume and the expression of neuronal differentiation markers Nestin and GFAP and proliferation marker Ki67. Radiation exposure inhibited NSp growth, decreased proliferation, and increased GFAP expression, indicating astrocytic differentiation. RNA sequencing analysis supported these findings, showing upregulation of Notch, BMP2/4, S100b, and GFAP gene expression during astrogenesis. By recapitulating radiation-induced toxicity and astrocytic differentiation, this single-NSp culture system provides a high-throughput preclinical model for assessing the effects of various radiation modalities and evaluates the safety and efficacy of potential therapeutic interventions in combination with radiation.