Preclinical dosing optimization: Dosing schedule, tissue

成功的药物开发需要一种安全和最有效的治疗方案。在早期的临床试验中，吗啡介导的外显子跳跃已经在动物模型(小鼠和狗)和DMD男孩中被证明实现了Dstrophin的恢复。在优化AO治疗的剂量选择时，除了疗效之外，还必须考虑毒性。PMO-AO的经验表明，尽管与其他AO化学药物相比，这些化合物具有更好的治疗指数，但肾脏效应预计是主要的剂量限制毒性。建立可靠的临床前和临床再生性方法 监测吗莫利诺对肾脏的影响对于最佳剂量选择和“分类”批准的调控策略至关重要。初步的GLP毒理学研究和针对外显子51(12wk，960 mg/kg/wk IV)使用大剂量吗啡的非GLP研究表明，该药物在肾脏近端小管细胞中显著蓄积。然而，一旦停止药物治疗，肾脏积聚就会消失，这表明优化剂量可以将肾脏毒性降至最低，或者可能需要在治疗方案中放假。在这里，我们假设，在DMD患者中建立一个适当的治疗窗口将涉及到AO药物的肌肉滞留(疗效)和肾脏滞留(毒性)之间的平衡。因此，本项目的目标是通过以下两个目标系统地研究实现这一平衡的配药计划。目的1使用iPRECIO遥测泵(颈静脉插管)，以尿液和上皮细胞生化终点(核心B)、肾脏组织学和新的生物标志物(项目2)为终点，在大鼠身上进行6个月的吗啡药物慢性比较剂量试验。在目标2中，我们将在小鼠疾病模型(MDX)中执行类似的静脉给药方案，目的是测试药物疗效、药物在肌肉中的生物利用度(RT-PCR和dystrophin数量)以及肾脏蓄积(核心B和项目2)。这种对多个品种的多种药物的系统比较将提供对优化治疗指数的评估。使用吗啉氧来纠正遗传缺陷正在迅速发展到其他几种人类疾病，如FSHD，囊性纤维化，强直性肌营养不良，LGMD等。因此，这项建议产生的疗效和毒性数据比治疗Duchenne肌营养不良具有更广泛的意义。

Successful drug development requires a therapeutic regimen that is safe and maximally effective. Morpholino-mediated exon skipping has been demonstrated to achieve restoration of dystrophin in both animal models (mice and dogs) and in DMD boys in early clinical trials. In addition to efficacy, toxicity must be considered in optimizing dose selection for AO therapy. Experience with PMO-AO suggests that while these compounds have a better therapeutic index compared to other AO chemistries, renal effects are anticipated to be the primary dose limiting toxicity. Establishment of reliable and reproductive methods for pre-clinical and clinical monitoring of renal effects of morpholino AO is critical for optimal dose selection and a "class" approval regulatory strategy. Preliminary GLP toxicology studies, and non-GLP studies using high dose morpholino AO directed at exon 51 (12 wk, 960 mg/kg/wk IV murine) showed significant accumulation of the drug in kidney proximal tubule cells. However, kidney accumulation resolved once drug treatment was stopped, suggesting optimized dosing could minimize renal toxicity or that a drug "holiday" may be needed in the regimen. Here we hypothesize that establishing an appropriate therapeutic window in DMD patients will involve a balance of muscle retention of AO drug (efficacy) vs. kidney retention (toxicity). Therefore, the goal of this project is to systematically investigate the dosing schedules that achieve this balance through the following two aims. Aim 1 will carry out 6 month chronic comparative dosing of morpholino drug in rats using iPRECIO telemetry pumps (jugular vein cannulation), with endpoints of urine and epithelial cell biochemistry (Core B), kidney histology and novel biomarkers (Project 2). In Aim 2, we will carry out a similar IV dosing schedule in the mouse disease model (mdx), with the goal of testing drug efficacy, drug bioavailability in muscle (RT-PCR and dystrophin quantity), and kidney accumulation (Core B, and Project 2). This systematic comparison of multiple drugs in multiple species will provide an assessment of the optimized therapeutic Index. Use of morpholino AO to correct genetic defects is rapidly advancing to several other human diseases such as FSHD, Cystic fibrosis, myotonic dystrophy, LGMD etc. Therefore the efficacy and toxicity data generated from this proposal have much broader implications than treatment for Duchenne muscular dystrophy.