PROTAC technology has a unique mechanism
- The PROTAC molecule specifically recognizes and binds to the target protein through the target protein ligand (POI Ligand) at one end and the E3 Ligase through the E3 Ubiquitin Ligase Ligand (E3 Ligase Ligand) at the other end.
- Forming the POI-PROTAC-E3 Ligase Ternary Complex
- In this ternary complex, the target protein POI is ubiquitinated by E3 ligase, and the ubiquitinated POI is subsequently recognized and degraded by the proteasome, thereby inhibiting the function of the target protein.
Advantages of PROTAC Technology in Drug Development
Change the Non-Drugability of the Target
The molecular mechanism of PROTAC is to degrade the target protein through the ubiquitin-proteasome system, and not to inhibit the function of the protein by blocking the functional area of the target protein through competitive binding. Therefore, PROTAC does not necessarily have to be the active region for the target protein to recognize the binding region, and the binding power does not necessarily have to be high-affinity. This makes some “non-druggable” target proteins that lack high-affinity binding of small molecule become “druggable”.
Traditional small molecule inhibitors inhibit the function of the target protein by competing with the active functional domain of the target protein and the amount of small molecules required is always large; while PROTAC degrades the target protein through the ubiquitin-proteasome system to relieve the function of the target protein, therefore PROTAC has the characteristics of recyclability, low dosage and high efficiency.
Compared with the antibody drugs, PROTAC does not trigger the production of anti-drug antibodies.
In conclusion, PROTAC has become an emerging weapon in the field of drug research and development. PROTAC is highly sought-after by scientific research institutions and pharmaceutical companies worldwide.
Introduction of Medicilon PROTAC Services
Design and Synthesis of Target Protein PROTAC-POI
Medicilon gathers the popular POI ligands and multiple tissue types of E3 ligase ligands, in addition of established a linker library containing hundreds of linking molecules. Moreover, Medicilon’s CADD technology platform has greatly improved the quality of PROTAC-POI design and synthesis.
In vitro Screening of PROTAC-POI
- HEK 293T Stable Transgenic Cell Line (POI-EGFP) High-Throughput Screening
The “endogenous protein-EGFP” 293T stable transgenic cell line constructed by gene knock-in technology was used for high-throughput screening of PROTAC molecules and DC50 was calculated.
The WB experiment was used to detect the degradation ability of the PROTAC molecules initially screened on the target protein and the DC50 value was analyzed.
- Cytotoxicity Test (CCK-8 or CTG method)
CTG detected the ability of PROTC-POI to inhibit the proliferation of tumor cells and analyzed the IC50 value.
- Mechanism Detection
a. Detect whether PROTAC degrades the target protein through E3 CRBN
b. Specificity Detection: Through the use of LC-MS/MS for proteomics analysis, to determine the specificity of PROTAC degradation protein; use KINOMEscan to analyze the specificity of PROTAC to kinase binding.
c. Tumor Cell Proliferation Inhibitory Effect
Use CCK-8 or CTG method to detect the inhibitory effect of PROTAC on tumor cell proliferation PROTAC
Screening of the Efficacy of PROTAC-POI in Animals
- CDX Mouse Tumor Model
Transplant the corresponding cancer cell lines into the nude mice or NSG mice to establish subcutaneous tumors or orthotopic tumor models, and test the inhibition effect of tumor growth by PROTAC-POI screened in vitro by oral, intragastric or intravenous administration
- PDX Mouse Tumor Model
The tumor tissues were transplanted into NSG mice in the form of tissue masses. These tumor tissues maintained the biological characteristics of the original tumors in the NSG mice, similar to clinical features. The PDX mouse model was used to test the inhibitory effect of lPROTAC-POI on in vivo tumor growth.
PK/PD Research, Pharmaceutical Analysis, Pharmacokinetic Research and Safety Evaluation
Perform PK/PD research, pharmaceutical analysis, pharmacokinetics research and safety evaluation on the obtained PROTAC-POI screened for in vivo drug efficacy testing in animals, and summarize the experimental results and documentations for IND filing to assist clients to accelerate the development process of PROTAC-POI drugs.