Chemotherapy and CDK4/6 Inhibitors: Unexpected Bedfellows
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Author Manuscript Published OnlineFirst on June 16, 2020; DOI: 10.1158/1535-7163.MCT-18-1161 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. 1 Chemotherapy and CDK4/6 inhibitors: Unexpected bedfellows Patrick J. Roberts1, Vishnu Kumarasamy2, Agnieszka K. Witkiewicz2,3, Erik S. Knudsen*2,4 1G1 Therapeutics, Research Triangle Park, NC 2Center for Personalized Medicine, Roswell Park Cancer Institute, Buffalo NY 3Department of Pathology, Roswell Park Cancer Institute, Buffalo NY 4Department of Molecular and Cellular Biology, Roswell Park Cancer Institute, Buffalo NY CONFLICT OF INTEREST STATEMENT: At the time of the initiation of this manuscript PJR was an employee of G1 Therapeutics which is involved in the clinical development of CDK4/6 inhibitors. The other authors have no potential conflicts of interest to report. Corresponding Author *Erik S. Knudsen Department of Molecular and Cellular Biology Roswell Park Cancer Center [email protected] Downloaded from mct.aacrjournals.org on September 30, 2021. © 2020 American Association for Cancer Research. Author Manuscript Published OnlineFirst on June 16, 2020; DOI: 10.1158/1535-7163.MCT-18-1161 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. 2 Abstract: Cyclin-dependent kinases 4 and 6 (CDK4/6) have emerged as important therapeutic targets. Pharmacological inhibitors of these kinases function to inhibit cell cycle progression and exert other important effects on the tumor and host environment. Due to their impact on the cell cycle, CDK4/6 inhibitors (CDK4/6i) have been hypothesized to antagonize the anti-tumor effects of cytotoxic chemotherapy in tumors that are CDK4/6 dependent. However, there are multiple preclinical studies that illustrate potent cooperation between CDK4/6i and chemotherapy. Furthermore, the combination of CDK4/6i and chemotherapy is being tested in clinical trials to both enhance anti-tumor efficacy and limit toxicity. Exploitation of the non-canonical effects of CDK4/6i could also provide an impetus for future studies in combination with chemotherapy. Thus, while seemingly mutually exclusive mechanisms are at play, the combination of CDK4/6 inhibition and chemotherapy could exemplify rational medicine. Downloaded from mct.aacrjournals.org on September 30, 2021. © 2020 American Association for Cancer Research. Author Manuscript Published OnlineFirst on June 16, 2020; DOI: 10.1158/1535-7163.MCT-18-1161 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. 3 CDK4/6 in cell-cycle progression: Cyclin dependent kinases (CDKs) are serine/threonine kinases that regulate the sequential progression of the cell cycle in eukaryotic organisms. The molecular functions of these kinases in different phases of the cell cycle have been well characterized (1, 2). The cell cycle machinery in higher eukaryotes is tightly regulated by the presence of more than 10 proteins in the CDK family that can have overlapping and distinct functions (2). Cell-cycle initiation occurs in G1 phase, which is conventionally governed by the activation of CDK4 and CDK6 kinases that are downstream of mitogenic signals (3-5). The catalytic activity of CDK4 and CDK6 is positively regulated by the binding of D-type cyclins (D1, D2 and D3). Expression of D-type cyclins is induced in response to mitogenic stimuli and remains high as the cells progress to the G1/S phase boundary (6). Therefore, unlike other cyclins and CDKs that are regulated by other components of the cell-cycle machinery, the expression of D-type cyclins --and by extension CDK4/6 associated kinase activity--largely depend on mitogenic signaling pathways (7, 8). Transcription of D-type cyclins is intimately linked to multiple pathways that coalesce to lead to the accumulation of transcripts (7, 9, 10). Mitogenic signaling pathways also regulate the stability and localization of these proteins (11, 12). Importantly, a host of growth inhibitory mechanisms also impact CDK4/6 activity, including the induction of endogenous CDK4/6-specific inhibitors with specific stresses (e.g. CDKN2A which encodes p16INK4A), and active mechanisms of cyclin D1 degradation (13, 14). Thus, CDK4/6 activity acts as a sensor linking multiple-signaling pathways to the initiation of the cell cycle (15-17). Downloaded from mct.aacrjournals.org on September 30, 2021. © 2020 American Association for Cancer Research. Author Manuscript Published OnlineFirst on June 16, 2020; DOI: 10.1158/1535-7163.MCT-18-1161 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. 4 CDK4/6 regulates the cell cycle through phosphorylation of key substrates. Unlike the prototypical CDK1 and CDK2, which can phosphorylate many substrates, CDK4/6 has a very limited repertoire of targets (18). CDK4 and CDK6 selectively phosphorylate the RB tumor suppressor protein and additional members of the RB family (18-21). RB-family proteins function as transcriptional co-repressors and limit the expression of E2F target genes that include multiple genes required for cell cycle progression, DNA replication, and mitotic progression (22, 23). The phosphorylation of RB, which is initiated by CDK4 or CDK6 serves to limit transcriptional repression and enable progression through latter phases of the cell cycle defining the canonical CDK4/6-RB pathway (Fig 1A). The requirement for CDK4/6 in cell division has been interrogated utilizing multiple approaches and has illustrated important features of the cell cycle. The inhibition of CDK4/6 by the expression of endogenous inhibitors (e.g. p16INK4A) potently arrests cells that contain a functional RB protein and subsequently limits gene expression controlled by RB/E2F (Fig 1A). Multiple experimental methods (e.g. antibody injection, RNAi, etc.) have further suggested that D-type cyclins and/or CDK4/6 activity are generally important for progression from G1/S in normal cells as well as multiple cancer models (24). These findings contrast with studies in mouse models that clearly demonstrate that the cell cycle can proceed with genetic deletion of CDK4 and 6 or deletion of all D-type cyclins (25, 26). In this context, adaptation occurs in many tissues by enabling CDK2 or CDK1 activity to drive cell cycle entry. However, genetic suppression of CDK4/6 activity can limit or block tumor development in select models (27-30). Downloaded from mct.aacrjournals.org on September 30, 2021. © 2020 American Association for Cancer Research. Author Manuscript Published OnlineFirst on June 16, 2020; DOI: 10.1158/1535-7163.MCT-18-1161 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. 5 This was clearly shown in the context of HER2-driven breast cancer where CDK4/6 activity is required both for tumor etiology and maintenance (31). Pharmacological inhibitors of CDK4/6—mechanisms of action and resistance: Due to the function of CDK4/6 in coordinating cell division, pharmacological inhibitors have been developed as anti-cancer drugs. There are five selective CDK4/6 inhibitors (CDK4/6i); palbociclib (PD0332991), ribociclib (LEE011), abemaciclib (LY2835219), trilaciclib, (G1T28) and lerociclib (G1T38) (32-38). Currently, three of these drugs are FDA-approved for the treatment of ER+ metastatic breast cancer based on multiple randomized clinical trials (palbociclib, ribociclib, abemaciclib). While all of these compounds are selective for CDK4/6, palbociclib, ribociclib, abemaciclib, and lerociclib are formulated for oral long-term dosing. Trilaciclib was formulated specifically for intravenous delivery and short half-life with the intended goal of preventing chemotherapy-induced host toxicities. Consistent with their mechanism of action, all CDK4/6i have cytostatic activity that is associated with RB-dependent suppression of the G1/S transition (32, 36). Pharmacological CDK4/6i mimic the effect of RB activation (Fig 1B) and suppress the expression of genes that are conventionally regulated by the E2F-family of transcription factors (39, 40). Since many of these genes are involved in core functions of DNA replication and mitotic progression, and are considered essential for proliferation, the magnitude of transcriptional repression downstream from CDK4/6 inhibition is critical for cytostatic activity. Downloaded from mct.aacrjournals.org on September 30, 2021. © 2020 American Association for Cancer Research. Author Manuscript Published OnlineFirst on June 16, 2020; DOI: 10.1158/1535-7163.MCT-18-1161 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. 6 Multiple determinants of response to CDK4/6 inhibition are being elucidated through both preclinical investigation and the analysis of clinical specimens (Fig 1C). This work has illustrated that there are multiple cell cycle related alterations present in models or tumors (e.g. RB loss or overexpression of cyclin E) that are associated with resistance to CDK4/6 inhibitors (41-44) (Fig 1C). Conversely, a number of oncogenic signaling pathways (e.g. RAS/MAPK, PTEN/PI3K, or HIPPO) have emerged as contributing to resistance (45-47). These derangements enable escape from CDK4/6 inhibition by facilitating the inactivation/phosphorylation of RB even in the presence of the pharmacological CDK4/6i. This is believed to occur due to “plasticity”, which is associated with either incomplete inhibition of CDK4/6 or the ability of CDK2 to initiate