PROTAC technology as a Novel tool to identify the target of lathyrane diterpenoids

Abstract

Active natural products are an important source of drug discovery due to their unique biocompatibility, novel-structural framework and extensive pharmacological activities. The target identification of natural products is the key to thoroughly understanding their mechanism of action and guiding their subsequent structure optimization. Lathyrane diterpenoids isolated from the seeds of Euphorbia lathyris in our laboratory have been proved to possess good anti-inflammatory activities in lipopolysaccharide (LPS)-induced macrophages, but their precise target and mechanism of action remain unclear. Herein, we employed PROTAC technology combined with quantitative proteomic analysis to identify the potential targets of lathyrane diterpenoids in macrophages. ZCY-PROTAC, synthesized based on Lathyrol, the core scaffold structure of the most active natural compound (2S,3S,4S,5R,9S,11R,15R)-15-acetoxy-3- cinnamoyloxy-5-hydroxy-14-oxolathyra-6(17),12E-diene, ZCY-001), intensively degraded a target protein MAFF in mouse and human cells. MST, CETSA and DARTS assays confirmed the direct binding of MAFF with Lathyrol or ZCY020, a natural product sharing the same core scaffold structure. Further mechanism studies showed that ZCY020 was capable of inhibiting the formation of a MAFF homodimer, promoting MAFF-Nrf2 heterodimerization, and thus regulating the transcription and expression of downstream protein HO-1, thereby exerting antioxidant, anti-inflammatory activity and promoting protective mitophagy in LPS-induced inflammation in macrophages and acute lung injury in mice. Together, our research indicates that MAFF is a potential target of ZCY020 and other lathyrane diterpenoids, and that PROTAC technology can be an innovative approach and a useful supplement for target identification of natural products.

Discovery of XL01126: A Potent, Fast, Cooperative, Selective, Orally Bioavailable, and Blood–Brain Barrier Penetrant PROTAC Degrader of Leucine-Rich Repeat Kinase 2

“…PROTAC is not only an emerging drug discovery modality but also offers new chemical tools for target identification and validation and for deciphering target biology. 44,45 For example, PROTACmediated degradation can reveal the noncatalytic activity of protein kinases. 46,47 Herein, we report the discovery and characterization of XL01126, a von Hippel−Lindau (VHL)based, fast, potent, cooperative, and selective LRRK2 PROTAC degrader that is also orally bioavailable and blood−brain barrier (BBB)-permeable.…”

Natural Product-Inspired Targeted Protein Degraders: Advances and Perspectives

“…140 Finally, in addition to their therapeutic properties, natural degraders also have the potential to be developed as potent chemical biology tools not only for protein chemical genetic knockdown, e.g., two-headed estradiol PROTACs used to deplete the ER, but also for target identification of bioactive compounds, e.g., lathyrane-based PROTACs used to target the MAFF protein. 113 In addition, AUTAC1 ( 102) is a paradigm for the application of NPs in new emerging types of TPD technologies. 168 Thus, the other recently developed new TPD strategies, such as LYTAC, AUTAC, ATTEC, PhosTAC, TF-PROTAC, RIBOTAC, and photo-PROTAC, could also be expanded to the NP-inspired TPD.…”

“…Through comparative quantitative proteomics, the downregulated POI degraded by the PROTAC was found to be the MAF BZIP transcription factor F (MAFF) protein in both RAW264.7 and HEK293T cells (1−40 μM, 24 h). 113 The key advantage of this method is different from that of the traditional affinity-based target identification method, as it has a strict requirement for affinity between the targets and small molecules, which is favorable for the moderate activity of NPs. Therefore, because of its excellent feasibility and reliability, this PROTAC-based method is a promising research tool for NP target identification.…”

Targeted protein degradation reveals BET bromodomains as the cellular target of Hedgehog Pathway Inhibitor-1

“…38,39 A recent preprint study furthermore describes the use of a PROTAC to identify the target of a natural product. 57 To convert a molecule into a PROTAC, it is necessary to first determine the structure-activity relationship for the parent molecule, which is often in itself part of a medicinal chemistry campaign and also a required step in the development of for example photo-affinity probes. Through our SAR analysis we arrived at HPP-9, a PROTAC version of HPI-1 that retained the inhibitory potency of the parent molecule, but was a partial rather than a full inhibitor.…”