Evaluation of TRPM8-expressing neurons as novel regulators of acute radiotherapy-associated pain in patients with head and neck cancer

该提案的目的是确定ARTN/GFRTRPM3/信号通路的每个组成部分的作用 放射相关疼痛(RAP)。放射治疗头颈癌(HNC)的常见后果是 急性口面部说唱。这种疼痛通常很严重，很难用目前的止痛药控制。对RAP的管制 在这些病例中，80%依赖于阿片类药物。这些药物对大多数患者并不完全有效，6个月后 放疗结束后，三分之一的HNC患者仍将依赖阿片类药物。迫切需要找出 更安全、更有效、更少上瘾的疼痛缓解策略，适用于接受治疗性放射治疗的HNC患者。 解决这一需求被我们知识中的一个关键鸿沟所阻碍：分子和细胞机制 导致口腔面部急性说唱的原因尚不为人所知。在RAP的小鼠模型中，我们的初步数据表明 急性口面部RAP的一个重要组成部分是信号通路，通常被认为是“冷痛”。 路径。在这项提议中要检验的假设是，局部照射的组织通过 一种涉及“冷痛”通路的神经回路。这一信号通路主要由神经元介导。 它们表达一种称为TRPM8的特定离子通道。TRPM8激活的典型机制包括释放 损伤组织中的青蒿素；Artn与其神经元受体结合，称为GfRα3，该受体进而激活 TRPM8.这项拨款的目的是：(1)界定TRPM8在调节急性口面部RAP中的作用；以及 (2)确定RAP信令是否需要ARTN/GFRα3信令。我们将检验这一假设并 通过三个特定目标(SA)实现我们的目标：(SA1)确定TRPM8表达神经元 在急性口面部RAP的发生中起关键作用；(Sa_2)以确定GfRα_3受体是否激活 RAP中的TRPM8信号；以及(SA3)确定ARTN是否是TRPM8(和急性 RAP)在口腔照射后。这一贡献意义重大，因为它将促进我们对 驱动急性口面部说唱的分子和细胞机制。结果将为以下工作奠定基础 确定既能改善HNC的舒适度又能减少阿片依赖的新靶向治疗 患有RAP的患者。重要的是，如果这种RAP途径在不同的解剖部位之间是保守的，我们的结果 可能远远超出HNC的范围，并积极影响患者的止痛选择 放射治疗适用于多种癌症。这项拟议的研究是创新的：还没有关于 RAP的分子和细胞机制，而不是试图通过减轻疼痛 炎症，我们将确定直接抑制RAP的策略。我们的方法使用了复杂的方法 用于追踪神经回路，并通过使用在体小鼠的体外研究最大化翻译相关性 人体细胞培养实验，以及患者来源样本的检测。不管每个人 我们所建议的机制的组件完全符合所建议的信令级联，我们的工作将 产生新的、相关的信息，推动该领域朝着急性口面部说唱的新疗法迈进。

The goal of this proposal is to define the role of each component of the ARTN/GFR3/TRPM8 signaling pathway in radiation-associated pain (RAP). A common consequence of radiotherapy for head and neck cancer (HNC) is acute orofacial RAP. This pain is often severe, and difficult to control with current analgesics. Control of RAP relies on opioids in >80% of these cases. These drugs are not fully effective in most patients, and 6 months after finishing radiotherapy, one-third of HNC patients will still be opioid-dependent. There is an urgent need to identify safer, more effective, and less addictive pain relief strategies for HNC patients undergoing therapeutic irradiation. Addressing this need is prevented by a critical gap in our knowledge: the molecular and cellular mechanisms that drive acute orofacial RAP are unknown. In a mouse model of RAP, our preliminary data have indicated that a critical component of acute orofacial RAP is a signaling pathway which is usually identified as a “cold pain” pathway. The hypothesis to be tested in this proposal is that locally irradiated tissues generate extreme pain via a neural circuit that involves the “cold pain” pathway. This signaling pathway is principally mediated by neurons which express a specific ion channel called TRPM8. A typical mechanism for TRPM8 activation involves release of artemin (ARTN) from injured tissues; ARTN binds its neuronal receptor, called GFRα3, which in turn activates TRPM8. The objectives of this grant are: (1) to define the role of TRPM8 in mediating acute orofacial RAP; and (2) to determine whether ARTN/GFRα3 signaling is required for RAP signaling. We will test the hypothesis and achieve our objectives with three specific aims (SA): (SA1) to determine whether TRPM8-expressing neurons are critical to development of acute orofacial RAP; (SA2) to determine whether the GFRα3 receptor activates TRPM8 signaling in RAP; and (SA3) to determine whether ARTN is a critical activator of TRPM8 (and acute RAP) after oral irradiation. This contribution is significant because it will advance our understanding of the molecular and cellular mechanisms that drive acute orofacial RAP. The results will lay the groundwork for identification of new targeted treatments that will both improve comfort and reduce opioid-dependency for HNC patients with RAP. Importantly, if this RAP pathway is conserved between various anatomic sites, our results may extend well beyond the realm of HNC, and positively impact the analgesic options for patients who undergo radiotherapy for a wide range of cancers. The proposed research is innovative: there has been no research on the molecular and cellular mechanisms of RAP, and rather than attempting to alleviate pain by mitigating inflammation, we will identify strategies for direct inhibition of RAP. Our approach uses sophisticated methods for tracing neural circuits, and translational relevance is maximized through use of in vivo mouse studies, in vitro human cell culture experiments, and examination of patient-derived samples. Regardless of whether each component of our proposed mechanism fits together into a signaling cascade exactly as proposed, our work will generate novel, relevant information that advances the field toward novel therapies for acute orofacial RAP.