Pre-clinical and clinical studies of NTBC and other compounds as potential treatments for albinism

NTBC 和其他化合物作为白化病潜在治疗方法的临床前和临床研究

• 批准号: 10266890
• 负责人: Brian Brooks
• 金额: $ 102.37万
• 依托单位: NATIONAL EYE INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10266890
• 关键词: Adult; Affect; Albinism; Alleles; Amblyopia; Axon; Blindness; Carrier Proteins; Cells; Characteristics; Child; Clinical Protocols; Clinical Research; Clinical Trials; Collaborations; Collection; Defect; Development; Disease; Dose; Early treatment; Electron Microscopy; Eye; FDA approved; Fibroblasts; Future; Glare; Goals; Hair; Human; Hypopigmentation; In Vitro; Iris; Length; Libraries; Manuscripts; Measurement; Melanins; Melanosomes; Modeling; Monophenol Monooxygenase; Morphology; Mus; Natural Products; Oculocutaneous Albinism; Operative Surgical Procedures; Optic Chiasm; Oral; Pathologic Nystagmus; Patients; Pharmaceutical Preparations; Pharmacologic Substance; Photophobia; Physiology; Pigmentation physiologic function; Pigments; Pilot Projects; Plasma; Preparation; Production; Proteins; Protocols documentation; Publishing; Refractive Errors; Research; Role; Skin; Skin Pigmentation; Structure of retinal pigment epithelium; Testing; Therapeutic; Tissues; Transillumination; Tyrosine; Validation; Vision; Visual impairment; Work; Zebrafish; experience; ganglion cell; high-throughput drug screening; human subject; improved; in vivo; in vivo evaluation; induced pluripotent stem cell; inhibitor/antagonist; insight; macula; melanocyte; mouse model; open label; orbit muscle; postnatal; preclinical study; side effect; stem cell differentiation; stem cell technology; vision aid; visual cycle

1. Clinical protocol for studying the effect of nitisinone (NTBC) treatment in human subjects with OCA1B Our previously-published work had established that nitisinone (NTBC) can increase melanin pigmentation in a mouse model of OCA1B. We performed an open-label pilot study with five adult patients with OCA-1B (Adams D.R., et al., JCI Insight. 2019). After establishing baseline measurements of iris, skin, and hair pigmentation, the patients were treated over 12 months with 2 mg/d oral NTBC. Changes in pigmentation and visual function were evaluated at 3-month intervals. Iris melanin remained unchanged after NTBC treatment, however, hair and skin pigmentation appeared to have increased in some patients with OCA-1B. The iris transillumination grading scale used in this study proved robust, with potential for use in future clinical trials. 2. Effect of NTBC in mouse models of OCA 2, OCA3 and OCA4 Treatment of a mouse model of OCA3 with oral NTBC does not have a favorable effect on melanin production and minimally affects the number of pigmented melanosomes in the iris stroma (Onojafe I.F., et al., Invest Ophthalmol Vis Sci. 2019). We conclude that treatment of OCA3 patients with NTBC is unlikely to be therapeutic. We further hypothesized that NTBC might improve melanization in mouse models of OCA2 (melanocyte-specific transporter protein) and of OCA4 (the so-called "underwhite" allele of SLC45A2). Similar to OCA3 mice, plasma tyrosine concentration was increased in NTBC-treated OCA2 and OCA4 mice after a month of nitisinone treatment with no overt toxic side-effects. Unlike the OCA3 mice, fur pigmentation was augmented in both mouse lines. Iris pigmentation was augmented predominantly in OCA4 mice. Electron microscopy also confirmed a small increase in choroidal pigmentation only in treated OCA4 mice, whereas, no effect was observed in the RPE in both lines. In a manuscript in preparation, we suggest that NTBC treatment might be beneficial in partially restoring pigmentation in OCA4 but not OCA2. 3. High-throughput drug screening to identify compounds that regulate Tyr activity We used purified, truncated Tyr protein (previously tested and validated to have equivalent enzymatic activity to full length protein) in a high-throughput drug screening. In collaboration with NCATS, we screened 34,000 compounds from the Genesis Drug Collection, the Natural Products Library, and the NCATS Pharmaceutical Collection. We identified >100 new inhibitors and a few activators of tyrosinase. After validation in a secondary enzymatic screen in vitro and in zebrafish in vivo, we are testing two doses of the top candidate compound on the OCA1B mouse model. At the lowest dose tested, the drug was well tolerated when administered i.p. for 30 days. 4. In vitro disease-in-a-dish modeling of OCA We developed a disease-in-a-dish model of oculocutaneous albinism type 1A (OCA1A) and type 2 (OCA2) using induced pluripotent stem cell (iPSC) technology. OCA patient fibroblasts were reprogrammed to iPSCs and differentiated to retinal pigment epithelium (OCA-RPE) using a developmentally-guided differentiation protocol. Tissue morphology and physiology as well as expression and functionality of the visual cycle apparatus were comparable to control pigmented iPSC-derived RPE. Furthermore, pigmentation defects comprising immature melanosomes in the OCA-RPE cells in vitro faithfully replicated tissue characteristics in vivo.

1。研究硝酸（NTBC）治疗对人类受试者的临床方案 我们先前发表的工作已经确定，尼炎（NTBC）可以在OCA1B的小鼠模型中增加黑色素色素沉着。我们对五名OCA-1B患者进行了一项开放标签试验研究（Adams D.R.等，JCIInsight。2019）。在建立了虹膜，皮肤和毛发色素沉着的基线测量后，用2 mg/d口服NTBC对患者进行了12个月的治疗。以3个月的间隔评估了色素沉着和视觉功能的变化。 NTBC治疗后，虹膜黑色素保持不变，一些OCA-1B患者的头发和皮肤色素沉着似乎增加了。这项研究中使用的虹膜透射分级量表证明了强大的稳定性，并有可能在以后的临床试验中使用。 2。NTBC在OCA 2，OCA3和OCA4的鼠标模型中的影响 用口服NTBC对OCA3的小鼠模型的处理对黑色素的产生没有良好的影响，并且最小影响了虹膜基质中有色黑色素体的数量（Onojafe I.F.等人，投资Ophthalmol VissSci。2019）。我们得出的结论是，OCA3 NTBC患者的治疗不太可能是治疗性的。 我们进一步假设NTBC可能会改善OCA2（黑色素细胞特异性转运蛋白）和OCA4（SLC45A2的所谓“下白色”等位基因）的小鼠模型中的黑色素化。与OCA3小鼠相似，在经过一个月的尼炎治疗后，NTBC处理的OCA2和OCA4小鼠的血浆酪氨酸浓度升高，没有明显的毒性副作用。与OCA3小鼠不同，两种小鼠系列的皮肤色素沉着均增强。在OCA4小鼠中，虹膜色素沉着主要增加。电子显微镜还证实，仅在处理过的OCA4小鼠中，脉络膜色素沉着的增加很小，而在两条线中，RPE中均未观察到效果。在准备手稿中，我们建议NTBC治疗可能有益于在OCA4中部分恢复色素沉着，而不是OCA2。 3。高通量药物筛查以识别调节Tyr活性的化合物 我们在高通量药物筛选中使用了纯化的，截断的Tyr蛋白（先前测试并经过验证具有等效酶活性与全长蛋白）。与NCAT合作，我们从Genesis药物收集，天然产品库和NCATS Pharmaceutical Collection中筛选了34,000种化合物。我们确定了> 100个新抑制剂和一些酪氨酸酶激活剂。在体外和体内斑马鱼中进行二级酶筛选验证后，我们正在测试OCA1B小鼠模型上的两剂顶级候选化合物。在测试的最低剂量下，当服用I.P.时，该药物耐受性良好30天。 4。OCA的体外疾病建模 我们使用诱导的多能干细胞（IPSC）技术开发了一种疾病的眼皮白化病（OCA1A）和2型（OCA2）的模型。 OCA患者成纤维细胞被重编程为IPSC，并使用开发引导的分化方案区分了视网膜色素上皮（OCA-RPE）。组织形态和生理学以及视觉周期设备的表达和功能与对照色素化的IPSC衍生的RPE相当。此外，色素沉着缺陷在体外忠实地复制了OCA-RPE细胞中未成熟的黑素体的缺陷。