The ‘ABC’ of split-nanoluciferase HIF heterodimerization bioassays: Applications, Benefits & Considerations

IF 5.3 2区医学 Q1 PHARMACOLOGY & PHARMACY

Biochemical pharmacology Pub Date : 2024-08-10 DOI:10.1016/j.bcp.2024.116478

Liesl K. Janssens, Christophe P. Stove

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Abstract

Hypoxia-inducible factors (HIF) are interesting targets for multiple therapeutic indications. While HIF activation is desired for the treatment of anemia-related and ischemic diseases, HIF inhibition is of tremendous interest to anti-cancer drug development. Different signaling events within the HIF pathway are being targeted by drug discovery programs, with a special interest in HIF-selective (possibly also HIF1/2 isoform-selective) compounds. In this study, we applied recently developed cell-based split-nanoluciferase HIF heterodimerization assays to study the effects of compounds, targeting HIF activity by various mechanisms of action. This study shows that the application of similar or diverse assay protocols allows to detect various influences on HIF heterodimerization as a key signaling event in the oxygen sensing pathway: increased HIF heterodimerization (roxadustat, MG-132), decreased HIF heterodimerization (PX-478, ibuprofen) and direct (HIF isoform-selective) heterodimerization inhibiting effects (PT-2385). Changes in treatment time and in the assay protocol allowed to assess direct and indirect effects on HIFα-HIFβ heterodimerization. In addition to the evaluation of applications of these new bioassays regarding pharmacological characterizations, benefits and considerations are discussed related to the use of cellular, luminescent-based bioassays. Briefly, benefits include the bidirectional nature of the biological readout, the upstream mechanism of detection, the differentiation between HIF1 and HIF2 effects and the simulation of various conditions. Specific and general considerations include cell-based, technical and disease/drug-related aspects (e.g., non-specific effects, color interference). In summary, the versatility of these bioassays offers benefits in widespread applications regarding drug discovery and pharmacological characterization of various therapeutics, applying either the same or optimized experimental protocols.

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分体荧光素酶 HIF 异源二聚体生物测定的 "ABC"：应用、益处和注意事项。

低氧诱导因子（HIF）是多种治疗适应症的有趣靶点。激活 HIF 是治疗贫血相关疾病和缺血性疾病的理想方法，而抑制 HIF 则是抗癌药物开发的巨大兴趣所在。HIF 通路中的不同信号事件正成为药物研发计划的目标，其中对 HIF 选择性（也可能是 HIF1/2 同工酶选择性）化合物特别感兴趣。在这项研究中，我们应用了最近开发的基于细胞的裂殖核素酶 HIF 异源二聚体测定法来研究化合物的作用，通过各种作用机制来靶向 HIF 活性。这项研究表明，应用类似或不同的检测方案可以检测出对作为氧传感途径中关键信号事件的 HIF 异源二聚化的各种影响：HIF 异源二聚化增加（roxadustat、MG-132）、HIF 异源二聚化减少（PX-478、布洛芬）以及直接（HIF 同工酶选择性）异源二聚化抑制作用（PT-2385）。通过改变处理时间和检测方案，可以评估对 HIFα-HIFβ 异源二聚化的直接和间接影响。除了评估这些新生物测定在药理学特征方面的应用外，还讨论了使用基于细胞发光的生物测定的好处和注意事项。简而言之，好处包括生物读数的双向性、检测的上游机制、HIF1 和 HIF2 效应的区分以及各种条件的模拟。具体和一般考虑因素包括基于细胞、技术和疾病/药物相关方面（如非特异性效应、颜色干扰）。总之，这些生物测定的多功能性为药物发现和各种治疗药物的药理学特征描述提供了广泛的应用优势，可应用相同或优化的实验方案。

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来源期刊

Biochemical pharmacology 医学-药学

CiteScore

10.30

自引率

1.70%

发文量

420

审稿时长

17 days

期刊介绍： Biochemical Pharmacology publishes original research findings, Commentaries and review articles related to the elucidation of cellular and tissue function(s) at the biochemical and molecular levels, the modification of cellular phenotype(s) by genetic, transcriptional/translational or drug/compound-induced modifications, as well as the pharmacodynamics and pharmacokinetics of xenobiotics and drugs, the latter including both small molecules and biologics. The journal''s target audience includes scientists engaged in the identification and study of the mechanisms of action of xenobiotics, biologics and drugs and in the drug discovery and development process. All areas of cellular biology and cellular, tissue/organ and whole animal pharmacology fall within the scope of the journal. Drug classes covered include anti-infectives, anti-inflammatory agents, chemotherapeutics, cardiovascular, endocrinological, immunological, metabolic, neurological and psychiatric drugs, as well as research on drug metabolism and kinetics. While medicinal chemistry is a topic of complimentary interest, manuscripts in this area must contain sufficient biological data to characterize pharmacologically the compounds reported. Submissions describing work focused predominately on chemical synthesis and molecular modeling will not be considered for review. While particular emphasis is placed on reporting the results of molecular and biochemical studies, research involving the use of tissue and animal models of human pathophysiology and toxicology is of interest to the extent that it helps define drug mechanisms of action, safety and efficacy.