In 2004, the Nobel Prize in Chemistry was awarded to Israeli scientists Aaron Sichanova, Afram Hershko and American scientist Irwin Ross for their discovery of ubiquitin-regulated protein degradation.
Protein degradation is rated as the next blockbuster therapy by Nature.
PQC & Intracellular Protein Degradation Pathway
A responsible protein quality control (PQC) system evolved in cells, including a variety of degradation pathways and molecular chaperones, which jointly complete the catalysis of the removal and refolding of abnormal proteins.
In protein homeostasis, the cell is in a dynamic equilibrium of protein synthesis, folding, and degradation.
Proteins are continuously synthesized and degraded in living cells. There are two ways to control the degradation of most proteins in mammals:
- Ubiquitin-proteasome system
In addition to degrading misfolded and damaged proteins, UPS is also responsible for controlling many key cellular processes (including cell cycle progression, cell proliferation and differentiation, cell signal transduction and transcription) and short-lived regulatory proteins.
- Autophagy-lysosome
The autophagy-lysosome pathway is mainly a stress response mechanism, which candegrade larger cell structures such as organelles, under starvation and other forms of stress.
Ubiquitin-proteasome Pathway
Ubiquitin is modified to the protein substrate through a series of enzyme-linked reactions:
- E1 activating enzyme primes ubiquitin via an ATP-dependent mechanism forming an E1~ubiquitin conjugate. (~; thioester bond)
- E2~ubiquitin conjugate via a transthiolation reaction with an E2 conjugating enzyme.
- One of the ~600 putative E3 ligases mediates the transfer of ubiquitin to a substrate protein.
The 26S proteasome is an ATP-dependent proteolytic complex composed of 20S core particles, 19S regulatory particles, and 11S regulatory factors. The hollow structure of the 20S cylinder has trypsin, chymotrypsin, and glutamyl-like peptide hydrolysis activities, which can cleave most peptide bonds.
The advantages and disadvantages of TPD compared to other protein silencing technologies
Compared with technologies such as CRISPER/Cas9 and RNA interference, targeted protein degradation (TPD) has the following advantages and disadvantages:
Pro
- TPD degrades the target protein in a shorter onset time, generally on the order of minutes/hour, and is less affected by molecular compensation and cell compensation.
- Possibility of reversibility and fine control.
- More precise and can selectively degrade the target protein.
Con
- Long development duration.
- The impact of off-target is not yet clear.
- The small molecules and E3 ligases that have been proven to connect to POI are relatively limited.
Main potentials of PROTAC
- Targeting the “Undruggable”
Protein degradation agents represented by PROTACs only need to weakly bind to the target protein to specifically label it, so that the target protein is degraded (traditional protein inhibitors need to have a strong binding to the active site of the target protein to be effective. However, it is estimated that 80% of proteins in human cells lack such sites).
- Higher Selectivity
PROTAC has higher selectivity than its single inhibitor, mainly due to:
- E3 Ligase tissue distribution has a certain specificity, which is equivalent to an extra layer of tissue selectivity.
- The spatial recognition of the target protein by E3 ubiquitin ligase can achieve selective profiling that cannot be achieved with traditional inhibitors.
- Controllability
The catalytic properties of PROTAC may lead to uncontrolled protein degradation, causing side effects (such as deterioration of the skin condition at the injection site), thereby limiting the clinical application of PROTAC. Photo-PROTAC provides a good solution for this, which is called the third-generation controllable PROTAC. The degradation of the target protein caused by it can be triggered by UVA or visible light.
- Catalytic mechanism
Small molecule inhibitors and macromolecular antibodies both need to continuously occupy the active site of the target protein to block the function, which is occupancy driven.
This requires the drug to meet the following conditions, which are a large enough dose to saturate the target, a long enough half-life to be continuously inhibited, and a high enough affinity to “get over” the original (ligand/receptor). As a result, many problems arise, such as large doses, excessive toxic and side effects, target mutation/overexpression, and drug resistance.
Advantages of the non-occupying event-driven mechanism are small dosage, overcoming drug target accumulation, overcoming drug resistance caused by target protein mutation/overexpression, independent of affinity, high selectivity, and elimination of protein accumulation.