Department of Bioengineering, University of California San Diego, San Diego, CA 92093, USA.
Department of Anesthesiology, University of California San Diego, San Diego, CA 92093, USA.
Sci Transl Med. 2021 Mar 10;13(584). doi: 10.1126/scitranslmed.aay9056.
Current treatments for chronic pain rely largely on opioids despite their substantial side effects and risk of addiction. Genetic studies have identified in humans key targets pivotal to nociceptive processing. In particular, a hereditary loss-of-function mutation in Na1.7, a sodium channel protein associated with signaling in nociceptive sensory afferents, leads to insensitivity to pain without other neurodevelopmental alterations. However, the high sequence and structural similarity between Na subtypes has frustrated efforts to develop selective inhibitors. Here, we investigated targeted epigenetic repression of Na1.7 in primary afferents via epigenome engineering approaches based on clustered regularly interspaced short palindromic repeats (CRISPR)-dCas9 and zinc finger proteins at the spinal level as a potential treatment for chronic pain. Toward this end, we first optimized the efficiency of Na1.7 repression in vitro in Neuro2A cells and then, by the lumbar intrathecal route, delivered both epigenome engineering platforms via adeno-associated viruses (AAVs) to assess their effects in three mouse models of pain: carrageenan-induced inflammatory pain, paclitaxel-induced neuropathic pain, and BzATP-induced pain. Our results show effective repression of Na1.7 in lumbar dorsal root ganglia, reduced thermal hyperalgesia in the inflammatory state, decreased tactile allodynia in the neuropathic state, and no changes in normal motor function in mice. We anticipate that this long-lasting analgesia via targeted in vivo epigenetic repression of Na1.7 methodology we dub pain LATER, might have therapeutic potential in management of persistent pain states.
目前，慢性疼痛的治疗主要依赖于阿片类药物，尽管它们有很大的副作用和成瘾风险。遗传研究已经在人类中确定了对伤害性处理至关重要的关键靶点。特别是，与伤害性感觉传入有关的钠通道蛋白 Na1.7 的遗传性功能丧失突变导致对疼痛不敏感，而没有其他神经发育改变。然而，Na 亚型之间的高序列和结构相似性挫败了开发选择性抑制剂的努力。在这里，我们通过基于 clustered regularly interspaced short palindromic repeats (CRISPR)-dCas9 和锌指蛋白的脊髓水平表观基因组工程方法，研究了在初级传入神经中靶向 Na1.7 的表观遗传抑制作为慢性疼痛的潜在治疗方法。为此，我们首先优化了 Na1.7 在 Neuro2A 细胞中的体外抑制效率，然后通过腰椎鞘内途径，通过腺相关病毒 (AAV) 递送这两种表观基因组工程平台，以评估它们在三种疼痛模型中的效果：卡拉胶诱导的炎症性疼痛、紫杉醇诱导的神经病理性疼痛和 BzATP 诱导的疼痛。我们的结果显示，Na1.7 在腰椎背根神经节中的抑制作用有效，减轻了炎症状态下的热痛觉过敏，减轻了神经病理性状态下的触觉过敏，而对小鼠的正常运动功能没有影响。我们预计，通过靶向体内表观遗传抑制 Na1.7 的方法实现的这种持久镇痛，可能在慢性疼痛状态的管理中具有治疗潜力。
