Targeting CDK4/6 for Anticancer Therapy

Abstract

Cyclin-dependent kinase 4/6 (CDK4/6) are key regulators of the cell cycle and are deemed as critical therapeutic targets of multiple cancers. Various approaches have been applied to silence CDK4/6 at different levels, i.e., CRISPR to knock out at the DNA level, siRNA to inhibit translation, and drugs that target the protein of interest. Here we summarize the current status in this field, highlighting the mechanisms of small molecular inhibitors treatment and drug resistance. We describe approaches to combat drug resistance, including combination therapy and PROTACs drugs that degrade the kinases. Finally, critical issues and perspectives in the field are outlined.

Keywords: CDK4/6; PROTAC; abemaciclib; cancer; drug resistance; palbociclib; ribociclib; small molecular inhibitor; trilaciclib.

Figure 1

CDKs and cell cycle. Cell cycle consists of G1, S, G2 phase and mitosis. Independent of a cell cycle also exists a dormant G0 phase. Whether a cell steps into a cell cycle relies on balance between proliferative and antiproliferative signals. During a cell cycle, multiple CDKs are sequentially activated. In G1, activated CDK4/6 by cyclin D phosphorylate Rb, partially relieving inhibition of E2F by Rb. Meanwhile, CDK4/6 hijacks Cip/Kip proteins, which stimulate CDK2-cyclin E, facilitating complete phosphorylation of Rb. Thus, E2F activity is totally released, and transcription of S phase genes is initiated. In late G1, CDK2-cyclin E complex is formed, driving transition toward the S phase. Next, CDK2 and CDK1 are successively activated by cyclin A and contribute to S/G2 and G2/M conversion, respectively. Finally, CDK1-cyclin B complex functions during mitosis. Besides CDKs, checkpoints also participate in the cell cycle via regulation of CDKs activity, inducing cell cycle arrest when abnormal cell division is detected.

Figure 2

CDK4/6 serves as a hub in tumorigenesis. In cancer, multiple oncogenes may be activated, including those on JAK/STAT, PI3K/Akt/mTOR, RAS/RAF/MEK/ERK, BTK/NF-κB, Wnt/β-catenin pathways, etc., all of which meet at CDK4/6-cyclin D complex. Moreover, mutations on tumor suppressor genes such as p53 can enhance CDK4/6 activity via releasing P21^CIP1 inhibition. CDK4/6, therefore, serves as a hub in tumorigenesis.

Figure 3

Chemical structures of CDK4/6-specific small molecular inhibitors approved by the FDA.

Figure 4

Different cell fates of CDK4/6 small molecular inhibitor (SMIs) treatment. Cancer cells arrested in the G1 phase with SMIs lead to quiescence (at low concentration), senescence (at high concentration), or, in some cancer types, apoptosis. Quiescence is reversible, and quiescent cells can re-enter the cell cycle in response to mitogenic signaling. Senescence is irreversible, and senescent cells can be killed by senolytic drugs and step into apoptosis. Thus, cancer cells can be cleared in two sequential steps. In some cancer types, SMIs alone or the combined treatment with MEK inhibitor or PI3K/mTOR inhibitor can change cell fate toward senescence or apoptosis. Drug resistance is another outcome of SMIs treatment and can be surmounted by combined therapy and PROTACs.

Figure 5

Mode of protein degradation using PROTACs. PROTACs bring the protein of interest (POI) in the vicinity of E3 ligase (such as CRNB, VHL, MDM2, cIAP2, etc.), forming a stable ternary complex of POI-PROTAC-E3 ligase. This structure facilitates ubiquitin transfer from E2 conjugating enzyme to a lysine residue in POI, resulting in ubiquitination of POI. Ubiquitination can occur on ubiquitin itself, leading to polyubiquitination. Polyubiquitinated POI is then subjected to proteasome for degradation via ubiquitin proteasome system (UPS). PROTAC degrader can be recycled for the next round of degradation.

Figure 6

Mechanism of action of PROTACs in the degradation of CDK4/6 and chemical structures of selective CDK4/6 degraders.

Figure 7

Surface drawing of the human CDK4 (blue) and CDK6 (green) structures in two different orientations. Surface-exposed lysine residues are colored red and labeled.