Chemo-enzymatic Approach to Late-stage Aromatic Fluorination of Drug Scaffolds

药物支架后期芳香氟化的化学酶法

• 批准号: 8323260
• 负责人: Ryan Mark Lauchli
• 金额: $ 5.57万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8323260
• 关键词: Address; Adverse effects; Biological; Biological Assay; Catalysis; Chemicals; Chemistry; Complex; Cytochrome P450; Diclofenac; Eligibility Determination; Engineering; Enzymes; Fluorides; Fluorine; Flurbiprofen; Generations; Hydroxylation; Metabolic; Methods; Palladium; Patients; Pharmaceutical Preparations; Pharmacologic Substance; Phenols; Property; Protocols documentation; Reaction; Research Personnel; Staging; Testing; Time; Warfarin; abstracting; appendage; directed evolution; drug candidate; drug market; drug modification; enzyme activity; high throughput screening; improved; novel; novel strategies; scaffold; scale up; small molecule

Project Summary/Abstract The incorporation of fluorine into small molecule pharmaceutical candidates is a powerful method to improve drug properties.[1] Currently, the late-stage introduction of fluorine atoms onto active drug-candidate scaffolds is a significant challenge for synthetic chemists.[2,3] This proposal describes a novel strategy to address the significant challenge of accessing phenols by selective enzymatic aromatic hydroxylation. This enzymatic step will be followed by chemical transformation of the resulting phenols to aryl fluorides using, and improving upon, recently developed chemistry.[4] This strategy will allow chemo-enzymatic aromatic C-H fluorination to be used for the late-stage modification of drug leads. Successful implementation of this strategy will fulfill an existing gap in biocatalytic aromatic hydroxylation, as well as facilitate rapid access to valuable complex fluorinated drugs and drug candidates. The specific aims of this project are 1) to engineer cytochrome P450 enzymes to conduct aromatic hydroxylation, 2) to scale up enzymatic aromatic hydroxylation and complete the chemical introduction of the aryl fluoride, and 3) to expand the strategy developed in aims 1 and 2 to the drugs warfarin and diclofenac. Using directed evolution, cytochrome P450 BM3 will be developed to carry out aromatic hydroxylation not only in high yield, but also with unprecedented regioselectivity. As ortho, meta, and para hydroxylation are all possible, enzymes with each type of regioselectivity will be engineered. To aid in this engineering effort, a novel high-throughput screening (HTS) protocol will be developed for enzyme activity, with a unique combination of assays that reveal the regioselectivites of the reactions. The enzymes resulting from directed evolution efforts will be used to carry out aromatic hydroxylations on a drug, flurbiprofen, on a gram scale. The complex phenols isolated from these hydroxylations will be transformed to aryl triflates, and finally to the sought-after aryl fluorides using chemistry recently demonstrated on simple aromatic molecules. The aryl fluorides will be assayed to demonstrate improved drug properties. The enzymes evolved for flurbiprofen aryl hydroxylation will be used for the selective hydroxylation of the important drugs warfarin and diclofenac, and the phenolic products will be converted to aryl fluorides and tested for enhanced properties.

项目总结/文摘

项目成果

Improved selectivity of an engineered multi-product terpene synthase.

• DOI: 10.1039/c4ob00479e
• 发表时间: 2014-05
• 期刊: Organic & biomolecular chemistry
• 影响因子: 3.2
• 作者: Ryan Lauchli;Julia Pitzer;Rebekah Z. Kitto;K. Z. Kalbarczyk;Kersten S. Rabe

High-throughput screening for terpene-synthase-cyclization activity and directed evolution of a terpene synthase.

• DOI: 10.1002/anie.201301362
• 发表时间: 2013-05-17
• 期刊: ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
• 影响因子: 16.6
• 作者: Lauchli, Ryan;Rabe, Kersten S.;Kalbarczyk, Karolina Z.;Tata, Amulya;Heel, Thomas;Kitto, Rebekah Z.;Arnold, Frances H.
• 通讯作者: Arnold, Frances H.