Figure 1. Wee1 signaling pathway
The Weel family includes Wee1A, Wee1B, and Myt1. Wee1A is present in somatic cells; Wee1B is present in blast cells; Myt1 is expressed in both somatic and blast cells. So far, the Wee1 protein is the most well-known Weel protein, and Wee1 protein kinases are members of the serine/threonine family and are highly conserved in evolution and are abundantly present in many eukaryotes and are very active in the S and G2 phases of the cell cycle (Figure 2). Wee1 was originally isolated from the fission yeast cells by Nurse. Because it inhibits cell division control protein 2 (Cdc), it is called Cdc28 in budding yeast. It is called CDK1 activity in humans, thereby inhibiting cell mitosis and eukaryotic cells. The biological cell is reduced in size, so it is named the "Wee" family. Wee1 protein kinase is a key regulatory center in DNA replication, chromosomal concentration, and histone transcription. These biological behavioral abnormalities can lead to genomic instability and cause malignant tumors, but inhibition or down-regulation of Wee1 protein kinase expression in malignant tumors can trigger mitotic catastrophe and apoptosis leads to tumor cell death.
Figure 2. Schematic outline of Wee1 functions
Wee1 family includes Wee1A, Wee1B, and Myt1. Wee1A is present in somatic cells; Wee1B is present in blast cells; Myt1 is expressed in both somatic and embryonic cells, but there are few studies on Myt1, and the only studies are now limited to lytic yeast, which may be related to its low expression. We are mainly to introduce the Wee1 protein. The human wee1 gene is located at 11p15.3~p15.1 and can transcribe a 3 kb mRNA with a coding content of 647 amino acids and a relative molecular mass of 94 KD. The protein kinase comprises three domains: the N-terminal regulatory domain, the central kinase domain, and the C-terminal regulatory domain. There are two kinds of untranslated regions of different lengths at the 3' and 5' ends of the mRNA. There are two phosphorylation sites on the 3' end sequence, which can be recognized and degraded by the ubiquitination system, and the 5' end will affect its half-life. During the interphase, the maintenance of Wee1 protein kinase activity depends on its binding to 14-3-3β protein and its autophosphorylation; in the G2/M phase, Wee1 protein kinase passes CDK1-mediated phosphorylation negative feedback mechanism, and 14-3-3β protein binding and other phosphorylation kinases down-regulate its activity; in cell M phase, β-TrCP binds to Wee1 protein kinase phosphorylation sites Ser53 and PS121, resulting in degradation of Wee1 protein kinase.
Wee1 signaling pathway
The conduction of the wee1 signaling pathway can be described in two ways: 1). The role of Wee1 protein kinase in the cell cycle. The cell cycle refers to the whole process that eukaryotic cells undergo from the completion of one division to the end of the next division. The core component of the cell cycle regulatory system is the cyclin-dependent kinase. To ensure the phase and synergy of cell cycle events, the phase activation of CDKs is particularly important in the cell cycle, specific or temporal expression, accumulation, and breakdown of cyclins. The CDKs/Cyclins complex formed by the combination of CDKs and cyclins is called mitosis promoting factor (MPF), which regulates the initiation and progression of various stages of the cell cycle, thereby controlling cell proliferation and apoptosis. CDKs are catalytic subunits of MPF. Cyclin is a regulatory subunit of MPF. In the G2/M phase of the cell, Wee1 protein kinase inactivates the CDK1/Cyclin B complex by inhibiting CDK1 activity by phosphorylating the Tyr15 site of CDK1, thereby inhibiting the cell cycle into the mitosis phase; in the S phase, Wee1 protein kinase that acts as the chromatin synthesis sensor has two consecutive phosphorylation events. One is to phosphorylate the Tyr15 site of CDK1 throughout the S phase of the cell, preventing cells from entering the next cell cycle phase until DNA replication or repair is complete; The second is to phosphorylate the Tyr37 site of histone H2B, stop histone synthesis, and maintain the correct histone-to-DNA ratio before entering the mitosis phase. Since the coupling of DNA synthesis to histone transcription is critical for chromosome formation, these two processes make Wee1 protein kinase a major regulator of chromosome integrity. 2). The role of Wee1 protein kinase in DNA damage response. DNA stores the genetic information on which organisms depend for survival and reproduction. The external environment and factors within the organism can cause damage or alteration of DNA molecules if DNA damage or genetic information changes. Failure to correct will affect the stability of the genome. ATR can be stimulated by a wide range of genotoxicities, which in turn leads to the activation of DNA single-strand breaks. In addition, ATR can be activated by the activation of ATM, which occurs during the process of double-stranded DNA breaks. ATR is the major kinase responsible for the phosphorylation and activation of Chk1. Compared with Chk2, Chk1 can be activated simultaneously by ATM and ATR, which can occur normally during the cell cycle or in response to cell replication stress (such as during the replication fork stagnation). Chk1 can be phosphorylated and activated simultaneously. Wee1 protein kinase and Cdc25c phosphatase, Wee1 protein kinase phosphorylates the Tyr15 site of CDK1/CyclinB complex CDK1, which inactivates the cell cycle to prevent DNA repair in the G2 phase. Cdc25c phosphatase dephosphorylates the Tyr15 site of CDK1, allowing it to regain its activity, resulting in the cell cycle entering the M phase. It can be seen that whether the cell cycle can enter the M phase depends on the equilibrium state of Wee1 protein kinase and Cdc25c phosphatase.
Nowadays, the regulation of Wee1 signaling pathway has been a clinical hotspot. Of course, the most important concern is the inhibitor of Wee1, which can be used for the treatment of various cancers. Wee1 inhibitors are AZD-1775, PD0166285 and PD0407824. PD0166285 inhibits both Wee1 and Myt1; PD0407824 is a co-inhibitor of Wee1 and Chk1; AZD1775 is a specific small molecule inhibitor of Wee1. In addition, miR-194 also inhibits Wee1. The most studied is AZD-1775, which has entered Phase II clinical trials and is used in tumors with high expression of Wee1, which are well tolerated and have fewer adverse reactions. The HepG2/DDP cell line is a hepatoma cell that has been knocked out of the Wee1 gene, and the Wee1 protein kinase is silenced, which leads to enhanced sensitivity of the HepG2/DDP cell line to cisplatin and also increases the rate of apoptosis; GST, MRP1, and LRP will increase the expression levels of BCL-2. Pgp and survivin were significantly decreased in HepG2/DDP cell line, and the phosphorylation level of MEK/ERK was also significantly decreased, indicating that Wee1 can regulate the expression of drug-related genes and the activity of MEK/ERK pathway, and control the drug resistance and proliferation of human liver cancer cells. Studies have shown that Wee1 acts as a promoter of branch vessels in liver metastases of colorectal cancer and plays an important role in tumor growth and development. AZD-1775 can reduce the range of tumor angiogenesis without relying on tumor cells; when comparing the vascular endothelial cells adjacent to the normal liver tissue in the metastatic lesions, it was found that the endothelial cells were similar in morphology but different in molecular expression. Wee1 protein kinase has a high expression level and low survival rate in patients with stage IV gastric cancer and male gastric cancer with lymph node metastasis; for gastric cancer cell lines (AGS, YCC-2, MKN28, KATO III, SNU), studies with SNU-5, SNU-16, SNU-216, SNU-601, SNU-638, SNU-668, and SNU-719 have shown that AZD-1775 significantly inhibits gastric cancer cell proliferation, and induces apoptosis, and cell cycle arrest is more effective in gastric cancer cell lines with high expression of Wee1, and Wee1 inhibitor combined with other anticancer drugs (5-FU, paclitaxel, etc.) is more effective than a single application. In the pancreatic ductal adenocarcinoma cell line, the anti-tumor effect of AZD-1775 is affected by DNA repair status and is superior to DNA-containing repair genes in DDR-P (DNA repair proficient) cell lines (MIA PaCa2 and PANC-1), and FANCC, FANCG, and BRCA2 mutated cell lines (PL11, Hs 766T, and Capan-1). The inhibition of Wee1 alone may not provide clinical help, especially for FANCC, FANCG, and BRCA2 gene mutations. Triple-negative breast cancer studies have shown that cisplatin-induced DNA damage can activate DNA replication detection sites ATR, Chk1, and Wee1 regulation, cell cycle arrest in S phase for DNA repair, preventing cisplatin-induced cell death and resistance. The use of Wee1 inhibitor or siRNA can cause DNA replication to reignite, further increase replication stress and lead to more DNA damage, eventually killing cancer cells, while using cisplatin and AZD-1775 inhibitors to treat triple-negative breast cancer, is expected to overcome cisplatin drug resistance and better treatment results. miR-194 expression was significantly down-regulated in laryngeal squamous cell carcinoma tissues and cells. Overexpression of miR-194 in laryngeal squamous cell carcinoma cells inhibited tumor proliferation, metastasis, invasion and drug resistance, while Wee1 was shown to be miR-194. As the new direct-acting target, its overexpression overcomes the effect of miR-194 on tumor suppression to some extent. The above studies indicate that Wee1 inhibitors can reduce tumor cell resistance and inhibit tumor growth and metastasis. Wee1 inhibitors use Wee1 inhibitors in combination with other anti-tumor drugs for better anti-tumor effect, but the effect depends on DNA repair status.
It has now been found that the expression of Wee1 is up-regulated in many cancers, and it has been clinically found that treatment with an inhibitor of Wee1 can effectively improve the occurrence and development of cancer.
As in the case of cancer, it is possible to reduce the expansion of malignant cells by inhibiting the expression of wee1, which can be used as a target for clinical treatment of leukemia.