Preclinical evidence of PROTAC
The breakthrough in PROTAC’s design paves the way for its early clinical trials. According to statistics, as of January 8, 2023, 18 kinds of PROTAC were conducting phase I–III clinical trials.
The key preclinical study data are as follows:
- AR targeted protein degradant
Five kinds of PROTAC targeting androgen receptor (AR) (Bavdegalutamide (also known as ARV-110), CC-94676, AC176, HP518, and ARV-766) are conducting clinical trials in metastatic prostate cancer. ARV-110 preclinical data have been reported at academic conferences, while data on other AR degradants have not yet been disclosed and will not be discussed in this article.
Bavdegalutamide is a PROTAC proteolytic agent targeting AR. By recruiting E3 ubiquitin ligase to the vicinity of AR, AR is labeled with ubiquitin to guide it to be degraded by the proteasome.
The degradation activity of bavdegalutamide was tested in various animal models of prostate cancer. In castrated and non-ovariectomized resistant mice, the inhibitory effect of oral bavdegalutamide on tumor growth was significantly greater than that of enzalut. In the human tumor xenotransplantation model (PDX) expressing AR, bavdegalutamide inhibited tumor growth by 100%, and the level of PSA decreased by more than 90%.
- ER targeted protein degradant
Estrogen receptor (ER) positive breast cancer is the most common subtype of breast cancer. Endocrine therapy is a first-line treatment for ER positive advanced breast cancer and is often combined with CDK4/6 inhibitors. Fulvestrant is a new type of ER antagonist that can promote the degradation and down-regulation of ER, while the ER of up to 50% of patients treated with flurvist is still at the baseline level. In addition, during endocrine therapy, patients may have ESR1 gene mutations, resulting in reduced sensitivity to ER degradants such as flurvist. In order to solve the deficiency of flurvist group as an ER degrader, two kinds of PROTAC targeting ER (ARV-471 and AC682) are being developed clinically for ER positive breast cancer.
In the preclinical analysis of breast cancer cell lines, ARV-471 induced wild-type ER degradation, and the cell lines carrying ER-resistant mutations (Y537S and D538G) of DC50 1-2 ‘nM; also degraded after exposure to ARV-471. In addition, ARV-471 also showed encouraging anti-tumor activity in an ER positive breast cancer model. In the estrogen-dependent orthotopic MCF7 xenotransplantation model, the tumor growth inhibition rate was more than 90% at a daily dose of 10 mg/kg or 30 mg/kg (there was evidence that the tumor regressed, that is, at a dose of 30 mg/kg, the tumor inhibition rate was more than 100%), and the tumor ER level decreased by more than 90%.
- BTK targeted protein degradant
BTK is a key kinase in the signal transduction pathway of B cell receptor (BCR), which participates in the processes of B cell proliferation, differentiation, and apoptosis. A variety of BTK small molecule inhibitors, such as ibutini, have been approved or are in clinical trials for the treatment of B-cell malignant tumors, and drug-resistant mutations are the pain point of all covalent BTK inhibitors. Aiming at this pain point, four kinds of PTOTAC targeting BTK (NX-2127, NX-5948, BGB-16673, and HSK29116) are being developed for patients with B-cell malignant tumors. Preclinical data for NX-2127 and NX-5948 have been disclosed, but the results of BGB-16673 and HSK29116 studies are not known.
NX-2127 can induce BTK degradation in BTK wild-type cell lines of diffuse large B-cell lymphoma (DLBCL) and mantle cell lymphoma (MCL). In addition, BTK-C481S mutant cell lines are resistant to BTK inhibitors, while NX-2127 can effectively inhibit the proliferation of drug-resistant cells. NX-2127 can efficiently degrade BTK protein and inhibit cell proliferation more strongly than BTK inhibitors. RNA sequencing showed that NX-2127 down-regulated the expression of genes involved in DNA replication and repair, cell cycle, and survival signaling pathways. In addition, NX-2127 upregulates the expression of CD1c, and CD1c participates in the recognition of cancer cells by T cells.
- BRD9 targeted protein degradant
BRD9 is a subunit of the ncBAF complex, which also includes the carcinogenic fusion protein SS18-SSX, which is associated with synovial sarcoma. The efficacy of two BRD9-targeted PROTACs CFT8634 and FHD-609, in patients with advanced synovial sarcoma is being evaluated.
CFT8634 is a BRD9 protein degrader with oral activity, and its inhibitory activity is up to nanomoles (DC50 = 3 nM). The drug has been granted by the FDA as an orphan drug for the treatment of synovial sarcoma. CFT8634 in the dose range of 1–50 μ mg/kg once a day induced strong tumor growth inhibition in two different PDX models of synovial sarcoma. In one of the PDX models, the tumor subsided continuously after CFT8634 treatment.
- PROTAC targeting other proteins
DT2216 uses VHL as the ligand of the E3 ubiquitin ligase to target the anti-apoptotic protein BCL-xL, which is currently in phase I clinical trials.
KT-413 (formerly known as KTX-120) uses CRBN as an E3 ubiquitin ligase ligand targeting IRAK4, which participates in signal transduction downstream of Toll-like receptors. IRAK4 is a key molecule involved in the innate immune response.
KT-333 is a proteolytic agent targeting the transcriptional regulatory factor STAT3. STAT3 is an important transcriptional factor that participates in a variety of biological functions, including cell proliferation, survival, apoptosis, and inflammation. KT-333 induces effective degradation of STAT3 in solid tumor cell lines, lymphoma cell lines, and primary immune cells.
ASP3082 is a proteolytic agent targeting KRAS G12D. KRAS G12D is regulated by the mitogen-activated protein kinase pathway.