Senescence as a therapeutically relevant response to CDK4/6 inhibitors

Abstract

Cyclin-dependent kinases 4 and 6 (CDK4/6) phosphorylate and inhibit retinoblastoma (RB) family proteins. Hyperphosphorylated RB releases E2F transcription factors, activating a transcriptional program that initiates S phase. Due to the critical role that this pathway has in regulating cell cycle progression, inhibiting CDK4/6 is an attractive therapeutic strategy. Indeed, CDK4/6 inhibitors in combination with antiestrogens produce a significant benefit in patients with ER⁺/HER2^- breast cancer. Clinical trials are currently investigating if the use of CDK4/6 inhibitors alone or in combination can be extended to other cancer types. Inhibition of CDK4/6 can result in different cell fates such as quiescence, senescence, or apoptosis. Senescence is a stress response that can be induced by stimuli that include oncogenic activation, chemotherapy, irradiation, and targeted therapies such as CDK4/6 inhibitors. Senescent cells undergo a stable cell cycle arrest and produce a bioactive secretome that remodels their microenvironment and engages the immune system. In this review, we analyze the therapeutic relevance of senescence induction by CDK4/6 inhibitors. We also discuss how different therapies, including checkpoint inhibitors and drugs targeting MEK or PI3K, can be used in combination with CDK4/6 inhibitors to reinforce or exploit senescence. Recently, a lot of effort has been put into identifying compounds that selectively kill senescent cells (termed senolytics). Thus, sequential treatment with senolytics might be an additional strategy to potentiate the antitumor effects of CDK4/6 inhibitors.

Figure 1. Inhibition of CDK4/6 induces cell cycle arrest in G1 phase.

After M phase, RB is hypophosphorylated and can bind and inhibit transcription factors of the E2F family. When activated by Cyclin D, CDK4/6 phosphorylate RB. RB can subsequently be phosphorylated by CDK2/Cyclin E. Hyperphosphorylated RB releases E2F transcription factors that activate a transcriptional programme promoting transition to S phase. Cyclin dependent kinase inhibitors of the CIP/KIP family (such as p21^CIP) can inhibit both CDK2 and CDK4/6 activity. INK4 proteins, like p16^INK4a, specifically inhibit CDK4/6 activity leading to cell cycle arrest in G1 phase. CDK4/6 inhibitors (such as Palbociclib, Abemaciclib or Ribociclib) exert similar effects, also inducing an arrest in G1.

Figure 2. Different cell fates caused by CDK4/6 inhibitors influence treatment outcome.

In cancer cells with a functional RB pathway, treatment with CDK4/6 inhibitors usually induces quiescence or senescence. In some tumor types, senescence can be enforced by combined treatment with MEK inhibitors. The combination of a CDK4/6 inhibitor with a PI3K inhibitor can change the cell fate towards apoptosis, which is rarely seen in a monotherapy setting. Senescent cells have a characteristic phenotype that includes an enlarged and flattened morphology, increased senescence-associated β-Galactosidase activity (“blue cells”) and the senescence-associated secretory phenotype (SASP). The SASP promotes changes in the tumor microenvironment, like vascular remodeling (allowing for increased uptake of chemotherapy in pancreatic cancer) or recruitment of cytotoxic T cells. CDK4/6 inhibitors also induce enhanced expression of MHC-I complexes on cancer cells, facilitating activation of cytotoxic T cells. Combination treatment with checkpoint inhibitors (e.g. an anti-PD-1 antibody) can further improve the immune-mediated clearance of senescent cancer cells. In some tumors, treatment with CDK4/6 inhibitors can also reduce levels of immunosuppressive regulatory T cells or attract NK cells and promote NK-cell mediated cancer cell death.

Figure 3. Sequential treatment with CDK4/6 inhibitors and senolytics.

Treatment with CDK4/6 inhibitors induces senescence in many cancer cells. Senescent cells share specific vulnerabilities that can be targeted by senolytic drugs. Examples of senolytic drugs include BCL2-family inhibitors such as navitoclax (ABT-263) or cardiac glycosides (e.g. digoxin). Senolytic drugs induce apoptosis in senescent cells. The stepwise combination of drugs able to induce senescence (pro-senescence therapy) such as CDK4/6 inhibitors and a senolytic drug constitutes an emerging therapeutic strategy.