This article detailed the research progress of PROTAC technology in the past two years, summarized the new targets and technologies of PROTACs targeting cancer, viral infections, immune diseases, neurodegenerative diseases.

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Overview of New Targets And Technologies of
PROTAC
Most of the drugs used in clinical practice are based on small molecules. Different from
traditional small molecule inhibitors and antagonists, protein degradation technology has
developed rapidly in recent years due to its ability to induce the degradation of therapeutic
target proteins, which provides a new idea for the development of new drugs.
The concept of PROTACs (PROteolysis-TArgeting Chimeras) was first proposed by
Crews et al. in 2001. PROTACs can use the natural protein cleaning system in the body to
reduce protein level rather than inhibit protein function, so as to cure the disease.
PROTAC is a heterobifunctional molecule that looks like a dumbbell, with one end of the
molecule connected to a ligand that binds a target protein, one end to an E3 ubiquitin
ligase, and a suitable linker in the middle. PROTAC degradation of target proteins is
achieved through the ubiquitin proteasome system (UPS) : PROTAC molecules bind
target protein (POI) and E3 ligase to form a teradical complex, which tags the target
protein with ubiquitination, and the ubiquitinated protein is recognized and degraded by
intracellular proteasome 26S.
Figure 1. Mechanism of PROTAC-mediated protein degradation
(Signal Transduct Target Ther, 7(1): 181.)
On June 9, 2022, the team of Professor Rao Yu of Tsinghua University published a review
article entitled "PROTACs: great opportunities for academia and industry (an update from

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2020 to 2021)" in the journal Signal Transduction and Targeted Therapy under Nature,
which detailed the research progress of PROTAC technology in the past two years,
summarized the representative new targets of PROTACs targeting cancer, viral infections,
immune diseases, neurodegenerative diseases.
Research progress of new targets of PROTAC
In the past 20 years, the field of PROTAC has entered a period of rapid development,
especially since the successful degradation of BET protein by dBET1 PROTAC with
Pomalidomide as E3 ligase ligand in 2015. In the past two years, research papers related
to PROTAC have experienced explosive growth. In 2019, Rao Yu's team had summarized
more than 40 protein targets that were reported to be degraded by PROTAC. In the past
two years, about 90 protein targets that can be degraded by PROTAC have been added,
covering cancer, immune disorders, viral infections, neurodegenerative diseases and
other disease fields, among which cancer is the main application field.
Figure 2. The rapid development of PROTAC-related research in the past two years
(Signal Transduct Target Ther, 7(1): 181.)
According to incomplete statistics, different degradation agents based on PROTAC
technology can degrade about 54 kinases, accounting for 45% of the total target. Kinases
have been preferred as targets for protein degradation, primarily because most kinases

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have known and potent inhibitors or ligands that can be easily modified to link the linkers
and maintain sufficient binding affinity. In addition, kinases have deep binding pockets
that facilitate the binding of PROTACs, which induce the interaction of the kinases with E3
ligases, which in turn ubiquitinate and ultimately degrade the kinases. Furthermore,
despite the high homology of kinase proteins, PROTACs can selectively degrade different
isoforms of kinases.
So far, 518 kinds of kinases have been discovered, which are involved in various
physiological regulatory processes such as cell survival, proliferation, differentiation,
apoptosis, and metabolism. As shown in the figure below, these kinases can be divided
into nine categories according to their structure and function, namely receptor tyrosine
kinases (RTKs), TKL kinases (TKLs), STE kinases (STEs), CAMK kinases (CAMKs),
AGC kinases (AGCs) , CMGC kinases (CMGCs), atypical protein kinases, CK1 kinases
(CK1) and others. Among them, those marked in red are human degradable kinases with
existing PROTAC degraders. Receptor tyrosine kinases (RTKs) and CMGC kinases
(CMGCs) have the most developed PROTAC degraders, with 19 and 14 respectively,
accounting for more than half of the total. In contrast, CK1 kinase (CK1) and CAMK
kinases (CAMKs) have not developed a PROTAC degrader.

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Figure 3. Degradable human kinases based on PROTAC technology and their classification
(Signal Transduct Target Ther, 7(1): 181.)
New technology of PROTACs
Since PROTACs were developed on the basis of POI inhibitors, they still have a certain
degree of off-target effects. Due to the large molecular weight of PROTAC, its poor cell
membrane permeability and poor pharmacokinetic (PK) properties greatly reduce its
biological and therapeutic effects. In addition, although some PROTACs can effectively
induce the degradation of target proteins, their biological effects are weak and have no
effective effect on disease. Finally, most proteins do not have corresponding
small-molecule conjugates to design PROTACs, such as transcription factors that play an
important role in disease development. Since there are few inhibitors of transcription
factors, there is no binder available when designing PROTACs targeting transcription
factors. This greatly limits the application of PROTAC technology. In order to solve
the above problems, different types of PROTAC technologies have emerged in
recent years, such as Antibody-PROTAC, Aptamer-PROTAC conjugates,
Dual-target PROTACs, Folate-caged PROTACs and TF-PROTACs.
Antibody-PROTAC
Antibody-PROTAC is a new strategy to explore the assembly of new antibody-PROTAC
conjugates in combination with antibodies. This technology enables specific degradation
of proteins in different cells and tissues, thereby optimizing and maximizing the
therapeutic window, reducing the side effects of broad-spectrum PROTACs, and
increasing their potential as drugs or chemical tools.
Aptamer-PROTAC conjugates

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Aptamers are single-stranded nucleic acids with complex three-dimensional structures,
mainly including stems, loops, hairpins and G4 polymers. They bind to target proteins
through special effects such as hydrogen bonds, van der Waals forces, base stacking
forces, electrostatic effects, etc., with high specificity and affinity, and can improve the
water solubility, membrane permeability, and tumor targeting of traditional PROTACs.
Dual-target PROTACS
In the occurrence and development of cancer, there are usually multiple factors that work
together, including different kinds of kinases and growth factors, which can act
independently or interfere with each other through signaling networks. This method is
mainly to design a single molecule that binds two or more pharmacophore, and
simultaneously targets two or more anti-tumor targets.
Folate-caged PROTACs
Folate receptor alpha (FOLR1) is low in normal tissues but highly expressed in many
human cancers. Folate caged PROTACs are another technology to improve the targeting
specificity of PROTACs. The basic principle is to introduce folate groups into PROTAC
molecules to achieve release in target cells and tissues. In this technique, folic acid
releases active PROTAC through the action of endogenous hydrolase in the cell, and then
the degradant induces the degradation of the target protein.
TF-PROTACs
Transcription factors (TFs) are a class of proteins involved in gene expression and
regulation, which are also potential targets for cancer therapy. Unlike conventional
kinases, transcription factors do not possess the active pockets or allosteric regulatory
sites commonly found in kinases or other enzymes, making them difficult to target
by small molecule inhibitors. Since TFs can bind to specific DNA sequences and regulate
the transcription process, it is theoretically possible to target TFs with different DNA

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sequences instead of small molecule inhibitors. Therefore, TF-PROTAC replaces the
small-molecule ligand of the targeting protein with the corresponding DNA sequence, so
that it forms TF-PROTAC, targets specific TF, and induces its degradation, thereby
regulating the level and biological function of specific TF.
Clinical Trial Study of PROTAC
In addition to being a research tool, PROTACs also have great potential for
application in disease treatment. It has emerged as a new mode of drug discovery that
has the potential to transform traditional drug discovery into a new blockbuster therapy. As
of March 2022, as shown in the table below, more than ten PROTAC drugs have
entered the clinical development stage worldwide. Among them, ARV-110 and
ARV-471 of Arvinas have entered clinical phase II, which are the fastest clinical progress
among PROTAC drugs.

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Table 1. The summary of protein-degradation drug candidates based on PROTAC technology in
the global clinical and IND stages
(Signal Transduct Target Ther, 7(1): 181.)
Summary and Prospect
Compared with other drugs and therapies, PROTACs have many potential advantages,
such as wide tissue distribution and oral administration. Compared with other therapies
(such as cell therapy, antibody drugs, etc.), the production process of PROTAC is
simpler. Compared with small molecule drugs, PROTAC can target more targets that
cannot be targeted by small molecule drugs, resulting in better effects. Therefore,
PROTAC technology has received high attention from the industry, and has begun to be
used in drug research and development for cancer, immune disorders, viral infections,
neurodegenerative diseases, etc., among which the application in the field of cancer is the
main one. Arivinas, founded by Craig Crews, was the first to carry out the
drug development of PROTAC technology. In the past two years, ARV-110, a PROTAC
inhibitor targeting the androgen receptor (AR) and ARV-471, a PROTAC inhibitor
targeting the estrogen receptor (ER), developed by Arivinas, have been clinically
validated in prostate and breast cancer, respectively, which are milestones in the
application of PROTAC technology.
Although PROTAC has grown rapidly over the past 20 years, there are still many
challenges to be addressed. These challenges mainly come from two aspects,
namely, the optimization of PROTAC molecular design and druggability, and the
comprehensive evaluation of biological activity. The first is about the molecular design
and druggability of PROTAC, involving target protein ligands, new E3 ligase ligands and
new linkers. The second is bioactivity evaluation, involving the screening, druggability
evaluation and pharmacological evaluation of PROTAC molecules. There are no ready
answers to these questions at present, but it is believed that with the development of more
biological, pharmacological and clinical research, new evaluation methods and systems

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will be gradually established to solve these problems. It is believed that more and more
PROTACs will enter preclinical and clinical studies in the future, which will further test the
therapeutic effect of PROTACs. It is expected that PROTAC technology will bring benefits
to human disease treatment and life health in the future.
The commonly used linkers in the development of PROTACs are PEGs. Biopharma
PEG is a professional PEG derivatives supplier that provides multi functionalized PEG
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Reference:
PROTACs: great opportunities for academia and industry (an update from 2020 to 2021)
Related articles:
[1] Summary of PROTAC Degraders in Clinical Trials
[2] Four Major Trends In The Development of PROTAC
[3] PROTAC And Other Protein Degradation Technology
[4] PROTACs VS. Traditional Small Molecule Inhibitors