Chemoproteomic Analysis of Covalent Drug Target

Covalent drugs generally refer to drug molecules that form covalent bonds with amino acid residues on target proteins, such as cysteine, lysine, serine, etc. Aspirin is the earliest discovered covalent drug molecule, and there are also representative examples found in natural products, such as oridonin, which exhibits anti-inflammatory bioactivity. In recent years, covalent drugs have attracted the attention of pharmaceutical research and development companies. Currently, there are at least six covalent drug targeting kinase that have been approved by the FDA and are on the market, such as ibrutinib, which targets the BTK kinase.

 

Technical Principles

When conducting target discovery for covalent drugs in cells or tissues, the active small molecule can be chemically modified by attaching reporter groups (such as biotin, bio-orthogonal group, etc.). Under the premise of retaining the original activity of small molecules, protein targets that interact with them can be directly captured within live cells or tissues. Through enrichment, enzymolysis, mass spectrometry identification and data analysis, the target of the omics information and information of small molecule interaction amino acid can be obtained.

 

However, many active molecules are difficult to modify, or the products that react with amino acid residues are unstable, so that they are not suitable for mass spectrometry detection. For this kind of active molecules, competitive site identification is an excellent solution. The core technology of this method is based on a universal chemical probe. When the active small molecule reacts with an amino acid residue and occupies the site, a signal difference is generated compared to a blank control sample. By analyzing this signal difference, information about the target protein and amino acid site of the active molecule can be obtained, including both on-target and off-target information.

 

At ChemPro Innovations, our biological core technology platform DIA-ABPP can be used to comprehensively screen the targets of covalent drugs in cells, down to the amino acid residues. This method is applicable to various sample systems, such as live cells, tissues, cell lysates, etc. Active small molecules encompass a broad range of compounds, including but not limited to endogenous metabolites, active ingredients of natural products, and covalent drugs, etc.

 

Workflow

 

1.Labeling proteins using active molecular probes

 

2.Protein enrichment

 

3.Protein digestion

 

4.Multiplex quantitative labeling

 

5.Mass spectrometry detection

 

6.Analysis of small molecule targets and amino acid sites

 

Service at ChemPro Innovations

1.Design and modification of covalent drug probes

 

2.Omics analysis of covalent drug proteins (based on gel imaging and biomacromolecule mass spectrometry)

 

3.Analysis of covalent drug action site (based on mass spectrometry)

 

4.Target verification of covalent drugs (targeted quantitative protein analysis/ western blotting)

 

5.Screening of lead compounds of covalent drugs