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

该提案的目标是确定ARTN/GFR β 3/TRPM 8信号通路中每个组分的作用 辐射相关疼痛（RAP）头颈癌（HNC）放疗的一个常见后果是 急性口面RAP。这种疼痛通常很严重，并且难以用当前的止痛药控制。RAP的控制 依赖阿片类药物的病例超过80%这些药物对大多数患者并不完全有效， 即使完成放疗，三分之一的HNC患者仍将依赖阿片类药物。迫切需要查明 更安全，更有效，更少成瘾性疼痛缓解策略HNC患者接受治疗辐射。 解决这一需求是由我们的知识的一个关键差距：分子和细胞机制阻止 导致急性口面RAP的原因尚不清楚。在RAP的小鼠模型中，我们的初步数据表明， 急性口面RAP的一个关键成分是一种通常被认为是“冷痛”的信号通路， 通路本提案中待检验的假设是，局部照射的组织通过以下方式产生极度疼痛： 一个涉及“冷痛”通路的神经回路。这种信号通路主要由神经元介导 它们表达一种叫做TRPM 8的特殊离子通道。TRPM 8激活的典型机制涉及释放 arten（ARTN）从受伤的组织; ARTN结合其神经元受体，称为GFRα3，这反过来激活 TRPM 8.这项资助的目的是：（1）确定TRPM 8在介导急性口面RAP中的作用; (2)以确定RAP信号传导是否需要ARTN/GFRα3信号传导。我们将检验这个假设， 通过三个具体目标实现我们的目标（SA）：（SA 1）确定TRPM 8表达神经元是否 对急性口面RAP的发生至关重要;（SA 2）确定GFRα3受体是否激活 （SA 3）确定ARTN是否是TRPM 8的关键活化剂（和急性的TRPM 8活化剂）。 RAP）。这一贡献意义重大，因为它将促进我们对 分子和细胞机制驱动急性口面RAP。研究结果将为 确定新的靶向治疗，既改善舒适度，又减少HNC的阿片类药物依赖性 RAP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，并对接受HNC治疗的患者的镇痛选择产生积极影响。 放射治疗广泛的癌症。拟议的研究是创新的：没有研究 RAP的分子和细胞机制，而不是试图通过减轻疼痛来减轻疼痛， 炎症，我们将确定直接抑制RAP的策略。我们的方法使用复杂的方法 用于追踪神经回路，并通过使用体内小鼠研究、体外 人细胞培养实验和来自患者的样品的检查。不管每个人是否 我们提出的机制的组成部分完全按照提议的那样组合成一个信号级联，我们的工作将 产生新的相关信息，推动该领域向急性口面RAP的新疗法发展。

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.