Research Area

Cell Cycle Inhibitor


Introduction of cell cycle inhibitor

The process of cell cycle regulation is the activation or inactivation of various regulatory factors under the surveillance of checkpoints, thereby initiating the process of cell DNA replication and division into two daughter cells. Among many cell cycle regulators, cyclin-dependent kinase (CDK) is at the core, and it is a network system that regulates the cell cycle with cyclin and cyclin-dependent kinase inhibitors (CKIs). CDKs are a class of serine/threonine kinases, and currently, 13 species are found, including CDK1~13, which play a role in cell cycle regulation of CDKs and transcriptional regulation of CDKs. The regulation of the cell cycle is, in fact, the regulation of checkpoints, with G1/S regulatory points being the most important. When the cell cycle is stimulated by external signals such as growth factors, the catalytic subunit CDK4/CDK6 binds to the regulatory subunit CyclinD, and the CDKs residues are activated by phosphorylation/dephosphorylation. After activation of CDKs, the Rb protein is phosphorylated. The rb gene, also known as the retinoblastoma gene, is the first cloned tumor suppressor gene, and its ability to form a complex with transcription factors (such as E2F) after phosphorylation of the protein is lost. E2F plays an important role in cell cycle regulation and induces the expression of CyclinE and CDK2 and forms a CyclinE/CDK2 complex, which further phosphorylates Rb protein and fully releases E2F. Subsequently, E2F enters the nucleus to activate a series of cell cycles into S phase. In the late phase of DNA replication during S phase, CDK2 is activated by cyclinE, which inactivates transcription factor E2F in time, preventing apoptosis caused by persistently activated E2F. Research statistics show that more than 90% of human cancers have mutations in related genes in the CDK, Cyclin, CKI, and Rb pathways, with CDK and its corresponding regulatory subunit Cyclin being the most frequently dysfunctional. In addition, fluctuations in the cell cycle promote chemotherapy resistance and reduce the effects of chemotherapy. Therefore, the regulation of CDK/Cyclin activity, which restores normal cell cycle, is one of the strategies for treating tumors.

Cell cycle inhibitors are now used clinically

Drug researchers have focused on finding different types of CDK and Cyclin inhibitors as cutting-edge anti-cancer drugs. Currently, CDK inhibitors are mainly divided into the endogenous and the exogenous. The largest class of endogenous small molecule inhibitors is low molecular weight proteins, which are classified into two broad categories according to differences in structural functions, and one class is called the dual specific family INK4, including p15, p16, p18, p19, which inhibits the protein family. The inhibitory-dependent protein of CyclinD-associated kinase binds to the corresponding free CDK4, thereby blocking the binding of CDK4 to the corresponding cyclinD to form a catalytic dimer complex. The other class is called the Kip family, including P21, P27,P57. This protein family can form a trimer with a dimeric complex composed of cyclin E/CDK2 and cyclinA/CDK1, by blocking the catalytically active center of the dimer. Inhibition of these endogenous inhibitors, when combined with the kinase complex, specifically regulates its activity, thereby precisely regulating the cell's transformation from G1 to S phase. Studies have shown that the occurrence and development of multiple tumors are associated with decreased expression of CDKs/cyclins or decreased expression of endogenous inhibitors, such as deletion of P16, which has relationship with the development of melanoma, lung cancer, breast cancer, and colorectal cancer. Deletion of P27 protein is common in breast cancer, prostate cancer, colon cancer, and gastrointestinal cancer. Therefore, deletion of endogenous CDKs inhibitor or gene mutation is an important reference for tumor diagnosis. Endogenous small molecule inhibitors are also a class of important non-coding RNAs discovered in recent years. The target site regions bind to each other to rapidly and efficiently degrade mRNA or inhibit translation of the protein, controlling the protein at a lower or optimal level and requiring for life activities. More than 10 microRNAs have been discovered involved in cell cycle regulation. Among them, miR1-2 and miR3-4 target CDK4, respectively, and the cell cycle is arrested in the G1 phase, which inhibits tumor cell proliferation; miR-22 targets CDK6 cells. The cycle is stagnant in the G1 phase, which induces senescence in breast cancer cells. In different biological processes, these miRNAs regulate cell cycle progression by targeting E2F, CDK, Cyclin, P21, P27, DNA polymerase alpha, etc. to promote or block key regulators of the cell cycle. Exogenous inhibitors include antisense nucleic acids, antibodies, small interfering RNA interference (siRNA) and small molecule compounds. Small molecule compounds are the most important class of exogenous CDK inhibitors. In recent years, as the understanding of crystal structure allows people to conduct molecular simulation studies, breakthroughs have been made in the design and development of highly efficient and selective studies on chemical inhibitors of CDKs. It can be said that such compounds have new members every day. At present, small molecule CDK inhibitors can be divided into the following 13 categories, Roscovitine and Olomouc, Pyrimidines (PD-033299), Flavonoids (Flavopiridols), Thiazoles (SNS03), anthracene and its derivatives (SU951), piperidone (Paullones), imidazopyridine, pyrazolopyridine (AZ703), pyrazines ( AT751), butyrolactone-1 (butyrolactone-1), scorpionine (UCN-01) and other two species. Thirteen small molecule inhibitors have entered clinical trials. They are all small-molecule chemicals of planar heterocycles that compete with ATP for binding to the ATP binding site of CDK kinase. In vivo experiments showed that CYC202 has good drug resistance and good oral physiological activity and has obvious inhibitory effects on solid tumors in nude mice inoculated by human colon cancer and uterine cancer cells. In Phase Ib studies, 10 patients with ovarian cancers were taking CYC for more than 20 months, with no increase in tumors or severe treatment-related side effects, among which one patient’s tumor has shrunk by more than 30%, and some patients who have been treated for more than one year have a stable condition. Phase II clinical studies have found that CYC202 alone has a slightly poorer effect and is effective in combination with other chemotherapeutic drugs. Phase IIb clinical trials of CYC202 in combination with capecitabine for the treatment of breast cancer, combined with 2,2-difluorodeoxycytidine or cisplatin for the treatment of lung cancer and nasopharyngeal carcinoma are also underway. The development and application of small-molecule RNA interference technology have made it possible to study the gene expression of specific intervention target molecules, and many scientists have begun to intervene in the synthesis of CDK/Cyclin at the genetic level. Limaet al. transfected CyclinE-targeting siRNA into Hep3B, HepG2, SNU449 (CyclinE overexpression) and HuH7 (CyclinE overexpressed) and found that CyclinE expression was reduced by 90% in cells. DNA synthesis is significantly reduced, and cells undergo apoptosis. Galimberti et al. transfected siRNA targeting CyclinE, CDK2, and CDK1 into mouse lung cancer cells HOP-62, H-522 and H-23, respectively, and found that CyclinE/CDK2 can induce apoptosis and inhibit the proliferation of lung cancer cells. Decreased CDK1 expression caused by CDK1 siRNA interference only causes cell phase arrest and slows cell proliferation; while CDK1 and CDK2 siRNA co-interference lead to a simultaneous decrease of CDK1 and CDK2 expression, causing resistance in cell cycle S and G2/M phases. The stagnation also induced apoptosis of the cells. Cao Yinfang and other successful transfection of CDK2/CyclinE siRNA recombinant expression vector into HepG2 cells showed that CDK2 and CyclinE mRNA expression decreased significantly, cell cycle was arrested in S phase, G1 phase cells increased significantly, caspase-3 activity enhanced, HepG2 cells underwent apoptosis, and cell cycle changes were consistent with decreased proliferation of HepG2 cellsin vitro after transfection.

Function of cell cycle inhibitor

With the deepening of understanding of the important role of cell cycle regulation in tumor formation and apoptosis, cell cycle regulation has been further studied in tumor chemotherapy resistance. Cyclin-dependent kinases (Cdks), which play a role in driving the cell engine during the cell cycle, are ideal targets for tumor therapy. Most of the cancer cells have activation, overexpression of the cell division cycle (cdk) gene and defects in CDKIs function. The CDK inhibitor exerts a break action that inhibits the cell cycle. In recent years, CDKIs have become a major highlight of cancer therapy, which inhibits the activity of CDKs in the cell cycle. A series of clinical studies have also shown that a single application can have moderate effects. However, in combination with traditional cytotoxic chemotherapeutic drugs, CDKIs can significantly enhance the anti-tumor effect of traditional chemotherapeutic drugs. Therefore, research on the anticancer effects of CDKIs drugs and other chemotherapeutic drugs has become a hot spot in the current treatment of tumor resistance.

Reference

  1. Bendris N, Lemmers B, Blanchard J M. Cell cycle, cytoskeleton dynamics and beyond: the many functions of cyclins and CDK inhibitors. Cell Cycle. 2015, 14(12):1786-1798.
  2. Pitts T M, Davis S L, Eckhardt S G, et al. Targeting nuclear kinases in cancer: development of cell cycle kinase inhibitors. Pharmacology & Therapeutics. 2014, 142(2):258-269.
  3. Stone A, Sutherland R L, Musgrove E A. Inhibitors of cell cycle kinases: recent advances and future prospects as cancer therapeutics. Crit Rev Oncog. 2012, 17(2):175-198.
  4. Xu W, Mcarthur G. Cell Cycle Regulation and Melanoma. Current Oncology Reports. 2016, 18(6):34.
  5. Malumbres M, Barbacid M. Cell cycle, CDKs and cancer: a changing paradigm. Nature Reviews Cancer. 2009, 9(3):153-166.

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