IRE1a-XBP1 Signaling as a Driver of Chemotherapy-Induced Peripheral Neuropathy

摘要 在接受细胞毒治疗的患者中，高达75%的患者会发生化疗引起的周围神经病变(CIPN 紫杉醇(PTX)等药物，是停止化疗的主要原因。这些患者经历了 疼痛、对冷的敏感和不平衡。PTX激活Toll样受体4(TLR4)诱导CIPN相关疼痛 单核细胞上，诱导促炎细胞因子的表达。将TLR4作为目标可能有风险，因为 TLR4在抗癌免疫防御中的作用有趣的是，TLR4的激活触发了肌醇需求酶 1α-X盒结合蛋白1(IRE1α-XBP1)在免疫细胞中的信号转导。IRE1基因α-XBP1的抑制作用 通过减少促炎因子来增强PTX的抗肿瘤作用，提示它可能是一种 有吸引力的靶点是预防CIPN和提高PTX对癌症的疗效。我们已经确认了PTX引起 IRE1α-XBP1激活并诱导原代人类白细胞的致炎表型。值得注意的是， 免疫细胞(Ern1/Xbp1f/f-Vav1cre)缺乏IRE1α-XBP1的小鼠的白细胞不显示这一点 PTX暴露后的促炎表型。这些条件性基因敲除(CKO)小鼠表现出更少的感冒 神经病理性疼痛(部分坐骨神经结扎-PSNL)模型中的痛觉过敏和后爪失衡。 此外，通过转录分析，我们发现在白细胞中ir1α-xbp1信号是至关重要的。 诱导与CIPN相关的前列腺素和细胞因子所需的。因此，我们 假设PTX通过激活白细胞中的IRE1α-XBP1信号促进CIPN，并且 靶向这一途径可以用来预防CIPN。我们将实现以下具体目标：1) 明确免疫内源性IRE1α-XBP1在PTX诱导的CIPN中的作用。我们将给小鼠注射PTX IRE1α-XBP1(CKO)及其野生型(WT)的白细胞特异性缺失。我们预料到 CKO小鼠将免受PTX诱导的CIPN相关行为的影响。然后，我们将对WT小鼠进行选择性地 IRE1CIPN抑制剂(MKC8866或KIRA8)，以从药理上阻止α诱导的行为。 这些研究将确定靶向IRE1α治疗CIPN的可行性和治疗潜力。2)确定如何 PTX影响免疫细胞内IRE1α-XBP1信号，从而驱动CIPN。我们将利用内质网应激激活 指标(ERAI)转基因小鼠，当Ire 1α时，其细胞表达黄色荧光蛋白变体(Venus) 已被激活。免疫荧光将用于识别血液、脾、坐骨神经中的内质网应激白细胞 在CIPN过程中，神经和背根神经节。白细胞将根据报告阳性率和 通过RNA-Seq、免疫表型和功能分析进行分析。这些实验将揭示PTX是如何- 诱导的IRE1XBP1XBP1激活影响白细胞在整个α进展过程中的功能。我们的神经科团队- 肿瘤学家、内科临床医生和在疼痛生物学、神经免疫学、 免疫学、癌症生物学、IRE1α-XBP1、基因组学和生物信息学是测试这一点的独特位置 创新假说，并有助于开发治疗慢性疼痛的新型非麻醉性药物。

Summary Chemotherapy-induced peripheral neuropathy (CIPN) occurs in up to 75% of patients who receive cytotoxic agents such as paclitaxel (PTX), and is a major reason to discontinue chemotherapy. These patients experience pain, sensitivity to cold, and imbalance. PTX induces CIPN-related pain by activating Toll-like receptor 4 (TLR4) on monocytes, which induces expression of pro-inflammatory cytokines. Targeting TLR4 may be risky due to the role of TLR4 in immune defense against cancer. Interestingly, TLR4 activation triggers inositol-requiring enzyme 1 alpha-X-Box Binding Protein 1 (IRE1α-XBP1) signaling in immune cells. The inhibition of IRE1α-XBP1 enhances PTX antineoplastic effects by reducing pro-inflammatory factors, suggesting that it could be an attractive target to prevent CIPN and improve the efficacy of PTX for cancer. We have confirmed that PTX causes IRE1α-XBP1 activation and induces a pro-inflammatory phenotype in primary human leukocytes. Notably, leukocytes from mice lacking IRE1α-XBP1 specifically in immune cells (Ern1/Xbp1f/f-Vav1cre) do not display this pro-inflammatory phenotype upon PTX exposure. These conditional knockout (cKO) mice exhibit reduced cold allodynia and hind paw unbalance in a model of neuropathic pain (partial sciatic nerve ligation - PSNL). Additionally, through transcriptomic analyses we found that IRE1α-XBP1 signaling in leukocytes is critically required for the induction of prostanoids and cytokines that have been associated with CIPN. Therefore, we hypothesize that PTX promotes CIPN by activating IRE1α-XBP1 signaling in leukocytes, and that targeting this pathway could be used to prevent CIPN. We will accomplish the following specific aims: 1) Define the role of immune-intrinsic IRE1α-XBP1 in PTX-induced CIPN. We will administer PTX to mice with leukocyte-specific deletion of IRE1α-XBP1 (cKO) as well as their wild type (WT) counterparts. We anticipate that cKO mice will be protected from PTX-induced CIPN related behaviors. Then, we will treat WT mice with selective IRE1α inhibitors (MKC8866 or KIRA8) in order to pharmacologically prevent PTX-induced CIPN behaviors. These studies will define the feasibility and therapeutic potential of targeting IRE1α for CIPN. 2) Establish how PTX influences IRE1α-XBP1 signaling in immune cells to drive CIPN. We will exploit the ER stress-activated indicator (ERAI) transgenic mouse, whose cells express a yellow fluorescent protein variant (Venus) when IRE1α is activated. Immunofluorescence will be used to identify ER-stressed leukocytes in the blood, spleen, sciatic nerves, and dorsal root ganglia during CIPN. Leukocytes will be sorted according to reporter positivity and analyzed via RNA-Seq, immunophenotyping, and functional assays. These experiments will unearth how PTX- induced IRE1α-XBP1 activation influences leukocyte function throughout CIPN progression. Our team of neuro- oncologists, internist clinicians, and basic scientists with expertise in pain biology, neuroimmunology, immunology, cancer biology, IRE1α-XBP1, genomics and bioinformatics is uniquely positioned to test this innovative hypothesis and contribute to the development of novel non-narcotic treatments for chronic pain.