In situ imaging and proteome profiling indicate andrographolide is a highly promiscuous compound.

Lin Li,Sanjay Samanta,Hadhi Wijaya,Shao Q Yao,Yulin Lam

doi:10.1038/srep11522

Abstract

Natural products represent an enormous source of pharmacologically useful compounds, and are often used as the starting point in modern drug discovery. Many biologically interesting natural products are however not being pursued as potential drug candidates, partly due to a lack of well-defined mechanism-of-action. Traditional in vitro methods for target identification of natural products based on affinity protein enrichment from crude cellular lysates cannot faithfully recapitulate protein-drug interactions in living cells. Reported herein are dual-purpose probes inspired by the natural product andrographolide, capable of both reaction-based, real-time bioimaging and in situ proteome profiling/target identification in live mammalian cells. Our results confirm that andrographolide is a highly promiscuous compound and engaged in covalent interactions with numerous previously unknown cellular targets in cell type-specific manner. We caution its potential therapeutic effects should be further investigated in detail.

Highlights

Fluorescently quenched activity-based probes based on peptide acyloxymethylketones which were capable of real-time imaging of protease activities in mammalian cells[17]
Synthetic access to these probes was significantly simplified by adopting semi-synthetic strategies based on previously reported procedures (Fig. 2 & Supplementary Information)[21,24]
AP1NP were obtained by using similar methods from AP1 in 65% and 36% yields, respectively

Summary

Introduction

Fluorescently quenched activity-based probes based on peptide acyloxymethylketones which were capable of real-time imaging of protease activities in mammalian cells[17]. Detailed in situ bioimaging and proteome profiling confirmed that andrographolide is a highly promiscuous compound engaged in covalent interactions with numerous previously unidentified cellular targets in cell type-specific manner. Andrographolide ( referred to as WT) is a bicyclic diterpenoid isolated from leaves of Andrographis paniculata[18], which is used extensively in traditional Chinese medicine (TCM) It possesses a wide spectrum of biological activities including antibacterial, anti-inflammatory, antimalarial, anticancer and others[19], and is currently used in clinic in China for the treatment of paediatric pneumonia and respiratory tract infection[20]. Michael addition at the C-12 position of these probes by a nucleophilic residue (i.e. cysteine) from the intended target would result in the formation of a covalent protein-probe adduct, which undergoes rapid and spontaneous β -elimination to release the highly fluorescent AMC and AAN (Fig. 2 & Supplementary Information). 9 different probes of WT were designed for the current study (Table 1)

Methods

Results

Conclusion