Development of Anti-OLAM Aptamers as Novel Analgesics (Phase 2)

描述(由申请人提供)：作为新型止痛药的抗Olam适体的开发(第二阶段)由于对疼痛机制的不完全理解，疼痛的管理仍然是一个主要的卫生保健问题。TRPV1是配体门控离子通道的瞬时受体电位(Trp)家族中的重要成员，可以检测周围组织中的有害化学和物理刺激。药理学和基因缺失研究都表明，TRPV1在炎症、热、痛觉过敏和其他疼痛条件下发挥着关键作用。氧化亚油酸代谢物(OLAM)最近被证明包含一个新的内源性TRPV1激动剂家族，有助于急性和炎症性疼痛条件。因此，阻断OLAM系统的化合物很可能构成一个新的止痛药家族。为了直接支持这一预测，本文提供的第一阶段数据表明，注射针对两种主要OLAM：9-HODE和13-HODE的高亲和力DNA适配子，在膜片钳和热痛大鼠行为模型中产生了显著的镇痛作用。尽管这些数据为概念验证提供了证据，但适配子必须发展成更重的结合物，以避免肾脏和其他主要器官的快速清除。因此，运营技术公司(Optech)建议继续开发其成功的高亲和力抗Olam适配子，从第一阶段发展为体内长寿命的适配子结合物，专门与9-HODE和13-HODE结合，并在较长时间内中和它们的疼痛产生活性。这将允许用更特异、更便宜和更高亲和力的DNA适体来取代抗HODE多克隆抗体。在第二阶段，Optech希望通过在3‘端添加布洛芬来实现几个旨在增强药代动力学(PK)的特定目标。80)，从而能够与血清白蛋白结合以增加体重并延缓肾脏和肝脏的清除。还将研究减缓体内清除和保护体内适配子的替代但已被证实的方法，包括3‘-聚乙二醇(PEG)和共价3’大鼠白蛋白附着。此外，西南研究院(SwRI)将建立适体-HODE相互作用的三维分子模型，以实现潜在的分子工程，如有可能，使用修饰或非天然脱氧核苷酸来改善适体亲和力(已经表现出低NMKd值)和特异性。除了在存在和不存在不同剂量适体结合物的情况下进行体外膜片钳研究外，还将研究大鼠在注射和暴露于43oC有害热和各种适体-3‘-结合物前后的行为变化。如果观察到相对长期的PK(大约几个小时)，并且可以再次证明热/灼伤止痛，Optech将寻求向FDA提交IND或Pre-IND申请。

DESCRIPTION (provided by applicant): Development of Anti-OLAM Aptamers as Novel Analgesics (Phase 2) The management of pain remains a major health care problem due to an incomplete understanding of pain mechanisms. TRPV1, a prominent member of the transient receptor potential (TRP) family of ligand-gated ion channels, detects noxious chemical and physical stimuli in peripheral tissues. Both pharmacological and gene deletion studies have demonstrated a pivotal role for TRPV1 in inflammatory heat hyperalgesia and other pain conditions. Oxidized linoleic acid metabolites (OLAMs) have been recently demonstrated to comprise a novel family of endogenous TRPV1 agonists that contribute to acute and inflammatory pain conditions. Therefore, compounds that block the OLAM system are likely to constitute a novel family of analgesics. In direct support of this prediction, Phase 1 data provided herein demonstrate that injection of high affinity DNA aptamers developed against two of the major OLAMs: 9-HODE and 13-HODE, produced significant analgesia in patch- clamp and rat behavioral models of heat pain. Although these data provide evidence for proof-of-concept, the aptamers must be developed into heavier conjugates in order to avoid rapid clearance by the kidneys and other major organs. Accordingly, Operational Technologies Corporation (OpTech) proposes to continue developing its successful high affinity anti-OLAM aptamers from Phase 1 into long-lived aptamer conjugates in vivo which specifically bind to 9-HODE and 13-HODE and neutralize their pain-producing activities for extended periods of time. This would permit replacing anti-HODE polyclonal antibodies with more specific, less expensive and higher affinity DNA aptamers. In Phase 2, OpTech expects to complete several specific aims targeted toward enhancing pharmacokinetics (PK) by addition of ibuprofen to the 3' end (ref. 80), thereby enabling association with serum albumin to add weight and retard renal and hepatic clearance. Alternative, but proven, methods to slow in vivo clearance and protect aptamers in vivo including 3'-polyethylene glycol (PEG) and covalent 3' rat albumin attachment will also be investigated. Moreover, 3-dimensional molecular models of aptamer-HODE interactions will be generated by the Southwest Research Institute (SwRI) to enable potential molecular engineering of improvements to aptamer affinity (already exhibiting low nM KD values) and specificity, if possible, using modified or unnatural deoxynucleotides. In addition to in vitro patch clamp studies in the presence and absence of various doses of the aptamer conjugates, rats will be studied for behavioral changes before and after injection and exposure to 43oC noxious heat with the various aptamer-3'-conjugates. If relatively long- term (on the order of hours) PK is observed and heat/burn analgesia can again be demonstrated, OpTech will seek to file an IND or pre-IND application with the FDA.