Targeted Cholera Toxin for Treatment of Hyperalgesia

描述（由申请人提供）：美国疼痛协会进行的一项研究表明，9%的美国人患有中度至重度慢性疼痛。目前治疗慢性疼痛的方法在许多情况下无效或产生广泛的副作用，限制了它们的效用。在这个项目中，我们将利用最近的发现，在慢性疼痛的啮齿动物模型中，鞘内注射霍乱毒素可以阻断痛觉过敏和异常痛觉，并且毒素可以通过神经激肽1 （NK1）受体直接靶向脊髓中的伤害性神经元。我们将把霍乱毒素的催化部分偶联到P物质上，以便将霍乱毒素引导到脊髓中表达NK1受体的细胞。该偶联物将在表达NK1的细胞系和慢性疼痛的啮齿动物模型中进行活性测试，以验证其刺激cAMP生成、抑制Gs蛋白表达和抑制痛觉过敏和异位性疼痛的能力。这种偶联物的独特之处在于，霍乱毒素不会像皂苷、白喉毒素和假单胞菌外毒素那样杀死脊髓中表达NK1的细胞，这些细胞以前被用于NK1受体靶向策略。相反，霍乱毒素会使阿片受体与g偶联，从而增强其抑制作用，并会降低g偶联受体系统的活性，从而进一步抑制伤害性传递。因此，据我们所知，该项目将是第一个通过直接操纵体内已确定的神经元群体中的g蛋白功能来产生治疗效果的项目。如果成功，该项目将产生一种控制疼痛的新型药物，并将通过使特定细胞中的g蛋白成为药理靶点，为治疗开辟一个全新的维度。

DESCRIPTION (provided by applicant): In a study conducted by the American Pain Society it was demonstrated that 9 percent of the United States population suffers from moderate to severe chronic pain. Current methods to treat chronic pain are in many instances not effective or produce a wide range of side effects that limit their utility. In this project we are going to take advantage of the recent discoveries that intrathecally administered cholera toxin blocks hyperalgesia and allodynia in rodent models of chronic pain, and that toxins can be directly targeted to nociceptive neurons in the spinal cord via the neurokinin 1 (NK1) receptor. We will conjugate the catalytic portion of cholera toxin to substance P in order to direct the cholera toxin to NK1 receptor expressing cells in the spinal cord. This conjugate will be tested for activity in NK1 expressing cell lines and in rodent models of chronic pain for its ability to stimulate cAMP production, suppress the expression of Gs g-proteins and to inhibit hyperalgesia and allodynia. The unique aspect of this conjugate is that cholera toxin will not kill the NK1 expressing cells in the spinal cord like saporin, diphtheria toxin and pseudomonas exotoxin, which have previously been used in NK1 receptor targeting strategies. Instead, cholera toxin will uncouple opioid receptors from Gs, thus enhancing their inhibitory actions, and will reduce the activity of Gs coupled receptor systems, which would further suppress nociceptive transmission. Thus, this project will be the first, to the best our knowledge, to produce a therapeutic effect by directly manipulating g-protein function in an identified population of neurons in vivo. If successful, this project will produce a novel agent for the control of pain and will open up an entire new dimension in therapeutics by making g-proteins in specific cells the pharmacological target.