Abstract 2886: Osteoclast and neuronal cross-talk as a mechanism for chest-wall pain caused by thoracic radiosurgery for lung cancer
Authors: Park Sun , Peters Megan , Moore Joseph , Reno Kaitlyn , Farris Michael , Peters Christopher , Willey Jeffery 
Published: March 2024

Abstract
Radiation therapy (RT)-induced chest wall pain (CWP) is a severe complication after thoracic stereotactic body radiation therapy (SBRT) for lung tumors; with increased survivorship, ~30% of patients develop severe CWP. The causes of CWP are unknown, but could involve: 1] damage to intercostal peripheral nerves; 2] bone loss from affected osteoclast activity, or 3] both (secreted factors from activated osteoclasts can affect nerves). This study tested the potential for radiation to affect osteoclasts and/or sensory neurons to induce pain. Mature osteoclasts differentiated from RAW264.7 cells plated on Osteoassay surfaces were exposed to an acute 10 Gy Cs137 γ-rays and bone resorption was assessed. We collected conditioned media (CM) from these cells at 48h and applied to cultured primary mouse dorsal root ganglia neurons (irradiated and non-irradiated neuron), assessing neurite growth and pain-associated marker (CGRP, SP) expression through immunohistochemistry and RT-qPCR. Risedronate (50µM), an anti-resorptive agent, was administered pre-radiation to inhibit osteoclast activity. Unpaired t-tests were utilized for statistical analyses (α=0.05). Osteoclast resorption activity was increased by 48h after irradiation. These irradiated, activated osteoclasts exhibited increased mRNA expression of differentiation (RANK) and activity (CTSK, MMP9) markers. Neurons treated with CM from irradiated osteoclasts exhibited elevated levels of CGRP (+38.5%) and SP (+29.5%) compared to cells treated with CM from non-irradiated cells. Consistently, in co-cultures of osteoclasts and neurons exposed to radiation, irradiated neurons exhibited significantly increased neuropeptide levels. Risedronate not only reduced osteoclast activity but also inhibited neuropeptide expression in irradiated neurons within these co-cultures. Moreover, neuropeptide levels were similar between directly irradiated neurons and non-irradiated neurons. However, irradiated neurons exposed to CM from either irradiated or non-irradiated osteoclasts showed a marked 4-fold increase in neuropeptide levels vs neurons that were non-irradiated and then received osteoclast CM. Our findings indicate that radiation activates osteoclasts and leads to direct bone loss in vitro and signaling between irradiated osteoclasts and non-irradiated neurons increases pain responses. Importantly, this response is enhanced when neurons were also previously irradiated. During SBRT, intercostal nerves may be influenced by adjacent irradiated osteoclasts, increasing expression of pain-related neuropeptides as the osteoclasts break down bone, thus enhancing the risk of CWP as treatment progresses across time. Understanding these interactions could reveal potential targets for preventing CWP, allowing higher tumor radiation doses with less toxicity. Support: The Milton Raben Foundation.
Citation Format: Sun Park, Megan Peters, Joseph Moore, Kaitlyn Reno, Michael Farris, Christopher Peters, Jeffery Willey. Osteoclast and neuronal cross-talk as a mechanism for chest-wall pain caused by thoracic radiosurgery for lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2886.