MicroRNA Therapeutics for Traumatic Brain Injury

越来越多的证据表明，在癌症中被广泛研究的癌基因/激酶可以 用于治疗创伤性脑损伤(TBI)。证据包括：1)癌基因/激酶(例如，Src，ROCK， ERK、CDK等)在脑损伤后被激活；2)癌基因/激酶的激活不仅导致神经元死亡 通过细胞周期重新进入成熟神经元，但也介导了白细胞的渗透和由此导致的炎症 3)癌基因/激酶抑制剂可以改善脑损伤的转归，如Src 抑制剂(PP2)、岩石抑制剂(Y-27632)、ERK抑制剂(PD98059)、CDK抑制剂(ROSCOVITE)等。 在这项拨款申请中，研究人员假设提升单个肿瘤抑制因子 MicroRNA(MiR)减少多个癌基因/激酶将改善脑损伤的预后。调查人员 靶向肿瘤抑制基因microNRA-125b(miR-125b)作为脑损伤治疗的候选药物，因为：1) Pilot miR表达研究表明miR-125b是显著改变的两个miRNAs之一 2)miR-125b可减少多癌基因(如Munck2，Alpk3，Neu1， BAP1、E2F、JNK、ERK等)，与miR-靶向算法(TargetScan)预测的一样 研究人员此前已经证明，SRC在改善脑外伤预后方面发挥着关键作用。这个 初步治疗研究表明，miR-125b模拟(2.4 mg/kg，静脉注射)和/或 (0.24 mg/kg，侧脑室注射)可改善急性期(24小时)的病理结果 在颅脑损伤后的较晚时间(11-15天)促进认知功能。 在这项提议中，研究人员将重点放在外围效应上，如静脉注射。治疗 对人类来说更容易翻译。使用全基因组测序，研究人员确定了前四个miR- 经miR-125b模拟处理后血液中降低的125b靶基因(Munck2、Alpk3、Neu1、BAP1) 在TBI之后。请注意，所有四个最重要的反应基因都是癌基因/激酶。初步的机械论 研究数据表明：1)miR-125b与Mknk2、Neu1和BAP1的3‘非翻译区(3’非编码区)结合； 吗啉寡核苷酸(Mos)-miR125b-mKnk2阻断miR-125b与mKnk2的3‘非编码区的结合。此外，他们还 将证明MO-miR125b-Mknk2/Alpk3/Neu1/BAP1在体内可阻止miR-125b模拟诱导的 这些靶基因存在于血细胞(白细胞、血小板)、内皮细胞和脑细胞(神经元、星形胶质细胞、 小胶质细胞)，从而阻断了miR-125b模拟脑损伤后的治疗效果。 总之，这项提议将表明miR-125b模拟物具有外围和中枢效应，以 通过降低miR-125b靶向癌基因/激酶(Munck2、Alpk3、Neu1、BAP1)来改善脑损伤的预后。这 这项研究将有助于脑损伤领域中癌基因/激酶病理生理学的文献研究。联合使用 静脉注射。MIR-125b模拟和脂质体治疗大鼠脑损伤是一种新颖的方法，可以转化为治疗人脑损伤。

Accumulating evidence shows oncogenes/kinases that have been widely studied in cancers can be leveraged to treat traumatic brain injury (TBI). The evidence includes: 1) oncogenes/kinases (e.g., Src, ROCK, ERK, CDK, others) are activated after TBI; 2) activation of oncogenes/kinases not only cause neuronal death via cell cycle re-entry in mature neurons, but also mediate leukocyte infiltration and inflammation which results in BBB disruption after TBI; and 3) oncogenes/kinases inhibitors can improve TBI outcome, such as Src inhibitor (PP2), ROCK inhibitor (Y-27632), ERK inhibitor (PD98059), CDK inhibitor (Roscovitine), and others. In this grant application, the investigators hypothesized that elevating a single tumor suppressor microRNA (miR) to decrease multiple oncogenes/kinases will improve TBI outcomes. The investigators targeted tumor suppressor microNRA-125b (miR-125b) as a candidate for TBI therapeutics, because: 1) the pilot miR expression study showed that miR-125b is one of the top two miRNAs that significantly altered in blood after both TBI and ICH; and 2) miR-125b decreases multiple oncogenes (e.g., Mknk2, Alpk3, Neu1, Bap1, E2F, JNK, ERK, others) as predicted by miR-target algorithm (TargetScan), in addition to the oncogene Src which the investigators have previously shown plays an critical role in improving TBI outcome. The preliminary therapeutic studies demonstrate that miR-125b mimic (2.4mg/kg, intravenously, i.v.) and/or (0.24mg/kg, intracerebroventricularly, i.c.v.) can improve pathological outcome at acute stage (24 hr) and promote cognitive function at later times (11-15 days) after TBI. The investigators focus the mechanistic study on peripheral effects in this proposal, as i.v. treatment is more translatable to humans. Using whole genome sequencing, the investigators identify the top four miR- 125b target genes (Mknk2, Alpk3, Neu1, Bap1) that are decreased in blood after miR-125b mimic treatment after TBI. Note that all of the four top responsive genes are oncogenes/kinases. The preliminary mechanistic study data show: 1) miR-125b binds to the 3’ untranslated regions (3’UTR) of Mknk2, Neu1 and Bap1; and 2) Morpholino Oligos (MOs)–miR125b–Mknk2 blocks the binding of miR-125b to 3’UTR of Mknk2. Moreover, they will prove that MO–miR125b–Mknk2/Alpk3/Neu1/Bap1 in vivo prevents miR-125b mimic-induced decrease of these target genes in blood cells (leucocytes, platelets), endothelium, and brain cells (neurons, astrocytes, microglia) after TBI, and thus blocks the therapeutic effects produced by miR-125b mimic after TBI. In summary, this proposal will show that miR-125b mimic has both peripheral and central effects to improve TBI outcome via decreasing miR-125b target oncogenes/kinases (Mknk2, Alpk3, Neu1, Bap1). This study will contribute to the literature of oncogenes/kinases pathophysiology in the TBI field. The combined use of i.v. miR-125b mimic and liposomes to treat TBI in rats is novel, and can be translated to treat human TBI.