Most drugs block or modulate the function of the target protein by binding to a specific site of the target protein. However, the activity of many proteins cannot be changed in this way. Protein degradation agents represented by Proteolis-Targeting Chimaeras (PROTACs) can change the function of proteins in an unconventional way. This new type of drug molecule can promote the ubiquitination of the target protein, and then the labeled target protein will be degraded into small peptide molecules by the cell’s proteasome mechanism. However, because the ubiquitin-mediated degradation pathway occurs in the cell, protein degradation agents currently mainly attack targets in the cell.
On July 29, in a study published in the journal Nature, a team of scientists from Stanford University reported on a different protein degradation technology-lysosome-targeting chimaeras (LYTACs), It opens up new possibilities for targeted degradation of extracellular and membrane-bound proteins.
Researchers report that lysosomal targeting chimeras (LYTACs) are bifunctional molecules with two binding domains (below), and one end carries a cell surface transmembrane receptor CI-M6PR (cation-independent mannose- The oligoglycopeptide group bound by 6-phosphate receptor, and the other end carries an antibody or small molecule that binds to the target protein. The two binding domains are connected by a chemical linker.
The mechanism of action of lysosomal targeted chimeras (LYTACs) | Banik et al. reported in a recent Nature paper that a LYTAC molecule composed of an oligoglycopeptide group, an antibody, and a linker. The oligosaccharide peptide group can bind to the cell surface receptor named CI-M6PR, and the antibody can bind to a specific transmembrane or extracellular protein. The antibody can also be replaced by a small molecule (not shown in the figure). When LYTAC binds to CI-M6PR and target protein at the same time, the resulting CI-M6PR-LYTAC-target protein complex will be “engulfed” by the cell membrane to form a transport vesicle. The vesicles transport the complex to the lysosome (an organelle containing protein degrading enzymes), after which the target protein is degraded and the receptor enters the cycle for reuse. Whether LYTAC is also degraded remains to be seen. Researchers believe that LYTACs have potential application value in treatment. (Source: Nature)
The trimeric CI-M6PR–LYTAC–target protein complex formed on the plasma membrane will be “engulfed” by the cell membrane to form a transport vesicle. The vesicles transport the complex to the lysosome (an organelle containing protein degrading enzymes), after which the target protein is degraded.
Specifically, in this new study, scientists such as Steven M. Banik first designed LYTACs of different sizes and with different linkers, and used a small molecule called biotin as the target protein binding domain. Researchers have observed that these LYTACs can quickly transport extracellular fluorescent avidin to intracellular lysosomes in a way that requires binding to CI-M6PR. When they replaced biotin with an antibody that recognizes apolipoprotein E4, a protein associated with neurodegenerative diseases, the protein was also internalized and degraded by lysosomes.
Next, Banik et al. investigated whether LYTACs can induce the degradation of membrane proteins as drug targets. The results showed that in multiple cancer cell lines, LYTACs did induce the internalization and lysosomal degradation of epidermal growth factor receptor (EGFR, a membrane protein that drives cell proliferation by activating signaling pathways). Compared with the use of antibodies to block EGFR, the use of LYTACs to reduce EGFR levels in these cancer cell lines reduces the signal activation downstream of EGFR. This result confirms the previously reported advantages of using “target degradation” in therapeutic applications.
Similar results have been observed using LYTACs to target other single-pass transmembrane proteins, including targeted degradation of PD-L1 that helps cancer cells escape from the immune system.
According to the researchers, in the next step, they want to know whether LYTACs can also induce multiple transmembrane proteins (including the ubiquitous G protein-coupled receptors) and proteins that transport substances across the membrane (such as ion channels and solution carrier proteins). ) Degradation.
A review article issued by Nature in response to this new research progress pointed out that if LYTACs can degrade these proteins, then it will be interesting to compare the performance of this new technology with PROTACs. Because LYTACs can bind to the extracellular domains of these proteins, and PROTACs can bind to the intracellular domains of these proteins.
In addition, the article also mentioned that, like any new drug model, LYTACs have many areas for improvement. For example, Banik and colleagues’ first LYTACs targeting PD-L1 caused only partial degradation of the protein, and they attributed this result to the low expression of CI-M6PR in the cell line used. Considering this situation, the design of LYTACs can also use other lysosome-shuttling receptors (Lysosome-shuttling receptors) to replace CI-M6PR.
Targeted protein degradation is a promising treatment strategy, and the first PROTACs are currently being evaluated in clinical trials. In terms of application, LYTACs and PROTACs complement each other. PROTACs is a bifunctional protein degradation agent that mainly targets intracellular proteins. It connects the “target protein ligand” and “the recruitment ligand of E3 ubiquitin ligase” through a linker, that is, one end of PROTACs is connected to the target protein. Binding, the other end is combined with E3 ubiquitin ligase. The E3 ubiquitin ligase can mark the target protein as defective or damaged by “sticking” it to the target protein. After that, the cell’s protein shredder (ie 26S proteasome) will recognize and degrade the labeled target protein.
Affected by molecular size (larger than traditional drugs), PROTACs usually do not penetrate well into biological membranes, but for LYTACs, molecular size should not be a big issue because they do not need to penetrate the cell membrane.
In general, LYTACs, as an effective tool that can expand the range of degradable proteins, have very promising prospects. Of course, if a new therapy for humans is to be developed, it is necessary to further reveal the pharmacokinetics and toxicity of LYTACs, as well as how they are metabolized, distributed and excreted.
With the continuous development and maturity of technologies such as PROTACs and LYTACs, and the emergence of new protein degradation technologies, maybe one day, no protein cannot be targeted for degradation? What follows may be a milestone in the treatment of multiple diseases.
1# Steven M. Banik et al. Lysosome-targeting chimaeras for degradation of extracellular proteins. Nature (2020).
2# New class of molecule targets proteins outside cells for degradation (Source: Nature)