Multiple effects of CDK4/6 inhibition in cancer: From cell cycle arrest to immunomodulation. Bonelli Mara,La Monica Silvia,Fumarola Claudia,Alfieri Roberta Biochemical pharmacology Dysregulation of the cell cycle is a hallmark of cancer that leads to aberrant cellular proliferation. CDK4/6 are cyclin-dependent kinases activated in response to proliferative signaling, which induce RB hyper-phosphorylation and hence activation of E2F transcription factors, thus promoting cell cycle progression through the S phase. Pharmacologic inhibition of CDK4/6 by palbociclib, ribociclib, or abemaciclib has been showing promising activity in multiple cancers with the best results achieved in combination with other agents. Indeed, CDK4/6 inhibitors are currently approved in combination with endocrine therapy for the treatment of estrogen receptor-positive, human epidermal growth factor receptor 2-negative advanced or metastatic breast cancer. Moreover, a number of clinical trials are currently underway to test the efficacy of combining CDK4/6 inhibitors with different drugs not only in breast but also in other types of cancer. Beyond the inhibition of cell proliferation, CDK4/6 inhibitors have recently revealed new effects on cancer cells and on tumor microenvironment. In particular, it has been reported that these agents induce a senescent-like phenotype, impact on cell metabolism and exert both immunomodulatory and immunogenic effects. Here we describe recent data on the anti-tumor effects of CDK4/6 inhibitors as single agents or in combined therapies, focusing in particular on their metabolic and immunomodulatory activities. 10.1016/j.bcp.2019.113676
    Cell cycle regulators in cancer cell metabolism. Leal-Esteban Lucia C,Fajas Lluis Biochimica et biophysica acta. Molecular basis of disease Cancer proliferation and progression involves altered metabolic pathways as a result of continuous demand for energy and nutrients. In the last years, cell cycle regulators have been involved in the control of metabolic processes, such as glucose and insulin pathways and lipid synthesis, in addition to their canonical function controlling cell cycle progression. Here we describe recent data demonstrating the role of cell cycle regulators in the metabolic control especially in studies performed in cancer models. Moreover, we discuss the importance of these findings in the context of current cancer therapies to provide an overview of the relevance of targeting metabolism using inhibitors of the cell cycle regulation. 10.1016/j.bbadis.2020.165715
    Two-way communication between the metabolic and cell cycle machineries: the molecular basis. Kaplon Joanna,van Dam Loes,Peeper Daniel Cell cycle (Georgetown, Tex.) The relationship between cellular metabolism and the cell cycle machinery is by no means unidirectional. The ability of a cell to enter the cell cycle critically depends on the availability of metabolites. Conversely, the cell cycle machinery commits to regulating metabolic networks in order to support cell survival and proliferation. In this review, we will give an account of how the cell cycle machinery and metabolism are interconnected. Acquiring information on how communication takes place among metabolic signaling networks and the cell cycle controllers is crucial to increase our understanding of the deregulation thereof in disease, including cancer. 10.1080/15384101.2015.1044172
    Metabolic control of the cell cycle. Kalucka Joanna,Missiaen Rindert,Georgiadou Maria,Schoors Sandra,Lange Christian,De Bock Katrien,Dewerchin Mieke,Carmeliet Peter Cell cycle (Georgetown, Tex.) Cell division is a metabolically demanding process, requiring the production of large amounts of energy and biomass. Not surprisingly therefore, a cell's decision to initiate division is co-determined by its metabolic status and the availability of nutrients. Emerging evidence reveals that metabolism is not only undergoing substantial changes during the cell cycle, but it is becoming equally clear that metabolism regulates cell cycle progression. Here, we overview the emerging role of those metabolic pathways that have been best characterized to change during or influence cell cycle progression. We then studied how Notch signaling, a key angiogenic pathway that inhibits endothelial cell (EC) proliferation, controls EC metabolism (glycolysis) during the cell cycle. 10.1080/15384101.2015.1090068
    PFKFB3 regulates oxidative stress homeostasis via its S-glutathionylation in cancer. Seo Minsuh,Lee Yong-Hwan Journal of molecular biology Whereas moderately increased cellular oxidative stress is supportive for cancerous growth of cells, excessive levels of reactive oxygen species (ROS) are detrimental to their growth and survival. We demonstrated that high ROS levels, via increased oxidized glutathione (GSSG), induce isoform-specific S-glutathionylation of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) at residue Cys206, which is located near the entrance to the 6-phosphofructo-2-kinase catalytic pocket. Upon this ROS-dependent, reversible, covalent modification, a marked decrease in its catalytic ability to synthesize fructose-2,6-bisphosphate (Fru-2,6-P₂), the key glycolysis allosteric activator, was observed. This event was coupled to a decrease in glycolytic flux and an increase in glucose metabolic flux into the pentose phosphate pathway. This shift, in turn, caused an increase in reduced glutathione (GSH) and, ultimately, resulted in ROS detoxification inside HeLa cells. The ability of PFKFB3 to control the Fru-2,6-P₂ levels in an ROS-dependent manner allows the PFKFB3-expressing cancer cells to continue energy metabolism with a reduced risk of excessive oxidative stress and, thereby, to support their cell survival and proliferation. This study provides a new insight into the roles of PFKFB3 as switch that senses and controls redox homeostasis in cancer in addition to its role in cancer glycolysis. 10.1016/j.jmb.2013.11.021
    Endothelial pyruvate kinase M2 maintains vascular integrity. Kim Boa,Jang Cholsoon,Dharaneeswaran Harita,Li Jian,Bhide Mohit,Yang Steven,Li Kristina,Arany Zolt The Journal of clinical investigation The M2 isoform of pyruvate kinase (PKM2) is highly expressed in most cancer cells, and has been studied extensively as a driver of oncogenic metabolism. In contrast, the role of PKM2 in nontransformed cells is little studied, and nearly nothing is known of its role, if any, in quiescent cells. We show here that endothelial cells express PKM2 almost exclusively over PKM1. In proliferating endothelial cells, PKM2 is required to suppress p53 and maintain cell cycle progression. In sharp contrast, PKM2 has a strikingly different role in quiescent endothelial cells, where inhibition of PKM2 leads to degeneration of tight junctions and barrier function. Mechanistically, PKM2 regulates barrier function independently of its canonical activity as a pyruvate kinase. Instead, PKM2 suppresses NF-kB and its downstream target, the vascular permeability factor angiopoietin 2. As a consequence, loss of endothelial cell PKM2 in vivo predisposes mice to VEGF-induced vascular leak, and to severe bacteremia and death in response to sepsis. Together, these data demonstrate new roles of PKM2 in quiescent cells, and highlight the need for caution in developing cancer therapies that target PKM2. 10.1172/JCI120912
    Stimulation of glycolysis promotes cardiomyocyte proliferation after injury in adult zebrafish. Fukuda Ryuichi,Marín-Juez Rubén,El-Sammak Hadil,Beisaw Arica,Ramadass Radhan,Kuenne Carsten,Guenther Stefan,Konzer Anne,Bhagwat Aditya M,Graumann Johannes,Stainier Didier Yr EMBO reports Cardiac metabolism plays a crucial role in producing sufficient energy to sustain cardiac function. However, the role of metabolism in different aspects of cardiomyocyte regeneration remains unclear. Working with the adult zebrafish heart regeneration model, we first find an increase in the levels of mRNAs encoding enzymes regulating glucose and pyruvate metabolism, including pyruvate kinase M1/2 (Pkm) and pyruvate dehydrogenase kinases (Pdks), especially in tissues bordering the damaged area. We further find that impaired glycolysis decreases the number of proliferating cardiomyocytes following injury. These observations are supported by analyses using loss-of-function models for the metabolic regulators Pkma2 and peroxisome proliferator-activated receptor gamma coactivator 1 alpha. Cardiomyocyte-specific loss- and gain-of-function manipulations of pyruvate metabolism using Pdk3 as well as a catalytic subunit of the pyruvate dehydrogenase complex (PDC) reveal its importance in cardiomyocyte dedifferentiation and proliferation after injury. Furthermore, we find that PDK activity can modulate cell cycle progression and protrusive activity in mammalian cardiomyocytes in culture. Our findings reveal new roles for cardiac metabolism and the PDK-PDC axis in cardiomyocyte behavior following cardiac injury. 10.15252/embr.201949752
    PFK15, a Small Molecule Inhibitor of PFKFB3, Induces Cell Cycle Arrest, Apoptosis and Inhibits Invasion in Gastric Cancer. Zhu Wei,Ye Liang,Zhang Jianzhao,Yu Pengfei,Wang Hongbo,Ye Zuguang,Tian Jingwei PloS one PFKFB3 (6-phosphofructo-2-kinase) synthesizes fructose 2,6-bisphosphate (F2,6P2), which is an allosteric activator of 6-phosphofructo-1-kinase (PFK-1), the rate-limiting enzyme of glycolysis. Overexpression of the PFKFB3 enzyme leads to high glycolytic metabolism, which is required for cancer cells to survive in the harsh tumor microenvironment. The objective of this study was to investigate the antitumor activity of PFK15 (1-(4-pyridinyl)-3-(2-quinolinyl)-2-propen-1-one), a small molecule inhibitor of PFKFB3, against gastric cancer and to explore its potential mechanisms. The effects of PFK15 on proliferation, apoptosis and cell cycle progression in gastric cancer cells were evaluated by cytotoxicity and apoptosis assays, flow cytometry, and western blotting. In addition, the invasion inhibition effects of PFK15 were measured by transwell invasion assay and western blot analysis, and a xenograft tumor model was used to verify the therapeutic effect of PFK15 in vivo. Results showed that PFK15 inhibited the proliferation, caused cell cycle arrest in G0/G1 phase by blocking the Cyclin-CDKs/Rb/E2F signaling pathway, and induced apoptosis through mitochondria in gastric cancer cells. Tumor volume and weight were also significantly reduced upon intraperitoneal injection with PFK15 at 25 mg/kg. In addition, PFK15 inhibited the invasion of gastric cancer cells by downregulating focal adhesion kinase (FAK) expression and upregulating E-cadherin expression. Taken together, our findings indicate that PFK15 is a promising anticancer drug for treating gastric cancer. 10.1371/journal.pone.0163768
    E2F-dependent mitogenic stimulation of the splicing of transcripts from an S phase-regulated gene. Darville M I,Rousseau G G Nucleic acids research There is one class of genes whose expression increases at the G1/S transition of the cell cycle. One of these genes codes for 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFK-2), an enzyme that controls glycolysis. The cell-cycle regulation of the PFK-2 gene depends on a binding site for the transcription factor E2F located at the 5'end of the first exon and involves not only a transcriptional, but also a post-transcriptional, mechanism. We have investigated this mechanism by studying in Rat-1 fibroblasts mature and immature mRNAs from the endogenous PFK-2 gene and from stably expressed transgenes containing PFK-2 gene regions. An increase in precursor mRNA half-life and processing took place at the G1/S transition. Transgenes with a mutated E2F binding site or with mutated splice sites lost the regulation by serum, indicating that both an intact E2F binding site and an efficient splicing reaction are necessary for proper mitogenic stimulation. In quiescent cells, the transgene lacking the E2F binding site was more efficiently spliced than the wild-type construct. These results indicate that, in the wild-type gene, precursor mRNA splicing is blocked in G0and that this block requires the E2F binding site. The data provide evidence for a coupling between stimulation of promoter activity and increased mRNA splicing in the mitogenic regulation of S phase-regulated genes. 10.1093/nar/25.14.2759
    Hypoxia-Sensitive COMMD1 Integrates Signaling and Cellular Metabolism in Human Macrophages and Suppresses Osteoclastogenesis. Murata Koichi,Fang Celestia,Terao Chikashi,Giannopoulou Eugenia G,Lee Ye Ji,Lee Min Joon,Mun Se-Hwan,Bae Seyeon,Qiao Yu,Yuan Ruoxi,Furu Moritoshi,Ito Hiromu,Ohmura Koichiro,Matsuda Shuichi,Mimori Tsuneyo,Matsuda Fumihiko,Park-Min Kyung-Hyun,Ivashkiv Lionel B Immunity Hypoxia augments inflammatory responses and osteoclastogenesis by incompletely understood mechanisms. We identified COMMD1 as a cell-intrinsic negative regulator of osteoclastogenesis that is suppressed by hypoxia. In human macrophages, COMMD1 restrained induction of NF-κB signaling and a transcription factor E2F1-dependent metabolic pathway by the cytokine RANKL. Downregulation of COMMD1 protein expression by hypoxia augmented RANKL-induced expression of inflammatory and E2F1 target genes and downstream osteoclastogenesis. E2F1 targets included glycolysis and metabolic genes including CKB that enabled cells to meet metabolic demands in challenging environments, as well as inflammatory cytokine-driven target genes. Expression quantitative trait locus analysis linked increased COMMD1 expression with decreased bone erosion in rheumatoid arthritis. Myeloid deletion of Commd1 resulted in increased osteoclastogenesis in arthritis and inflammatory osteolysis models. These results identify COMMD1 and an E2F-metabolic pathway as key regulators of osteoclastogenic responses under pathological inflammatory conditions and provide a mechanism by which hypoxia augments inflammation and bone destruction. 10.1016/j.immuni.2017.06.018
    An E2F-dependent late-serum-response promoter in a gene that controls glycolysis. Darville M I,Antoine I V,Mertens-Strijthagen J R,Dupriez V J,Rousseau G G Oncogene The F-type mRNA of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase is found in proliferating, but not in quiescent, cells. This bifunctional enzyme catalyses the synthesis and degradation of fructose-2,6-bisphosphate, a potent stimulator of glycolysis. F-type mRNA concentration decreased upon differentiation of rat rhabdomyosarcoma cells; it increased in Rat-1 fibroblasts stimulated to proliferate by serum, by epidermal growth factor, or by the v-src oncogene product. This increase resulted, at least in part, from a stimulation of F promoter activity. The stimulation occurred at the G1/S transition of the cell cycle. It depended on a binding site for the oncogenic transcription factor E2F located in the first exon of F-type mRNA. This effect was inhibited by agents that increase cAMP concentration. The data provide the first evidence that a gene involved in the control of glycolysis can be regulated by a late-serum-response promoter in an E2F-dependent way.
    E2F1 mediates sustained lipogenesis and contributes to hepatic steatosis. Denechaud Pierre-Damien,Lopez-Mejia Isabel C,Giralt Albert,Lai Qiuwen,Blanchet Emilie,Delacuisine Brigitte,Nicolay Brandon N,Dyson Nicholas J,Bonner Caroline,Pattou François,Annicotte Jean-Sébastien,Fajas Lluis The Journal of clinical investigation E2F transcription factors are known regulators of the cell cycle, proliferation, apoptosis, and differentiation. Here, we reveal that E2F1 plays an essential role in liver physiopathology through the regulation of glycolysis and lipogenesis. We demonstrate that E2F1 deficiency leads to a decrease in glycolysis and de novo synthesis of fatty acids in hepatocytes. We further demonstrate that E2F1 directly binds to the promoters of key lipogenic genes, including Fasn, but does not bind directly to genes encoding glycolysis pathway components, suggesting an indirect effect. In murine models, E2F1 expression and activity increased in response to feeding and upon insulin stimulation through canonical activation of the CDK4/pRB pathway. Moreover, E2F1 expression was increased in liver biopsies from obese, glucose-intolerant humans compared with biopsies from lean subjects. Finally, E2f1 deletion completely abrogated hepatic steatosis in different murine models of nonalcoholic fatty liver disease (NAFLD). In conclusion, our data demonstrate that E2F1 regulates lipid synthesis and glycolysis and thus contributes to the development of liver pathology. 10.1172/JCI81542
    Cancer immune control needs senescence induction by interferon-dependent cell cycle regulator pathways in tumours. Brenner Ellen,Schörg Barbara F,Ahmetlić Fatima,Wieder Thomas,Hilke Franz Joachim,Simon Nadine,Schroeder Christopher,Demidov German,Riedel Tanja,Fehrenbacher Birgit,Schaller Martin,Forschner Andrea,Eigentler Thomas,Niessner Heike,Sinnberg Tobias,Böhm Katharina S,Hömberg Nadine,Braumüller Heidi,Dauch Daniel,Zwirner Stefan,Zender Lars,Sonanini Dominik,Geishauser Albert,Bauer Jürgen,Eichner Martin,Jarick Katja J,Beilhack Andreas,Biskup Saskia,Döcker Dennis,Schadendorf Dirk,Quintanilla-Martinez Leticia,Pichler Bernd J,Kneilling Manfred,Mocikat Ralph,Röcken Martin Nature communications Immune checkpoint blockade (ICB)-based or natural cancer immune responses largely eliminate tumours. Yet, they require additional mechanisms to arrest those cancer cells that are not rejected. Cytokine-induced senescence (CIS) can stably arrest cancer cells, suggesting that interferon-dependent induction of senescence-inducing cell cycle regulators is needed to control those cancer cells that escape from killing. Here we report in two different cancers sensitive to T cell-mediated rejection, that deletion of the senescence-inducing cell cycle regulators p16/p19 (Cdkn2a) or p21 (Cdkn1a) in the tumour cells abrogates both the natural and the ICB-induced cancer immune control. Also in humans, melanoma metastases that progressed rapidly during ICB have losses of senescence-inducing genes and amplifications of senescence inhibitors. Metastatic cells also resist CIS. Such genetic and functional alterations are infrequent in metastatic melanomas regressing during ICB. Thus, activation of tumour-intrinsic, senescence-inducing cell cycle regulators is required to stably arrest cancer cells that escape from eradication. 10.1038/s41467-020-14987-6
    Cyclin D-Cdk4,6 Drives Cell-Cycle Progression via the Retinoblastoma Protein's C-Terminal Helix. Topacio Benjamin R,Zatulovskiy Evgeny,Cristea Sandra,Xie Shicong,Tambo Carrie S,Rubin Seth M,Sage Julien,Kõivomägi Mardo,Skotheim Jan M Molecular cell The cyclin-dependent kinases Cdk4 and Cdk6 form complexes with D-type cyclins to drive cell proliferation. A well-known target of cyclin D-Cdk4,6 is the retinoblastoma protein Rb, which inhibits cell-cycle progression until its inactivation by phosphorylation. However, the role of Rb phosphorylation by cyclin D-Cdk4,6 in cell-cycle progression is unclear because Rb can be phosphorylated by other cyclin-Cdks, and cyclin D-Cdk4,6 has other targets involved in cell division. Here, we show that cyclin D-Cdk4,6 docks one side of an alpha-helix in the Rb C terminus, which is not recognized by cyclins E, A, and B. This helix-based docking mechanism is shared by the p107 and p130 Rb-family members across metazoans. Mutation of the Rb C-terminal helix prevents its phosphorylation, promotes G1 arrest, and enhances Rb's tumor suppressive function. Our work conclusively demonstrates that the cyclin D-Rb interaction drives cell division and expands the diversity of known cyclin-based protein docking mechanisms. 10.1016/j.molcel.2019.03.020
    Interconnection between Metabolism and Cell Cycle in Cancer. Icard Philippe,Fournel Ludovic,Wu Zherui,Alifano Marco,Lincet Hubert Trends in biochemical sciences Cell cycle progression and division is regulated by checkpoint controls and sequential activation of cyclin-dependent kinases (CDKs). Understanding of how these events occur in synchrony with metabolic changes could have important therapeutic implications. For biosynthesis, cancer cells enhance glucose and glutamine consumption. Inactivation of pyruvate kinase M2 (PKM2) promotes transcription in G1 phase. Glutamine metabolism supports DNA replication in S phase and lipid synthesis in G2 phase. A boost in glycolysis and oxidative metabolism can temporarily furnish more ATP when necessary (G1/S transition, segregation of chromosomes). Recent studies have shown that a few metabolic enzymes [PKM2, 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase (PFKFB3), GAPDH] also periodically translocate to the nucleus and oversee cell cycle regulators or oncogene expression (c-Myc). Targeting these metabolic enzymes could increase the response to CDK inhibitors (CKIs). 10.1016/j.tibs.2018.12.007
    Cyclin F Controls Cell-Cycle Transcriptional Outputs by Directing the Degradation of the Three Activator E2Fs. Clijsters Linda,Hoencamp Claire,Calis Jorg J A,Marzio Antonio,Handgraaf Shanna M,Cuitino Maria C,Rosenberg Brad R,Leone Gustavo,Pagano Michele Molecular cell E2F1, E2F2, and E2F3A, the three activators of the E2F family of transcription factors, are key regulators of the G1/S transition, promoting transcription of hundreds of genes critical for cell-cycle progression. We found that during late S and in G2, the degradation of all three activator E2Fs is controlled by cyclin F, the substrate receptor of 1 of 69 human SCF ubiquitin ligase complexes. E2F1, E2F2, and E2F3A interact with the cyclin box of cyclin F via their conserved N-terminal cyclin binding motifs. In the short term, E2F mutants unable to bind cyclin F remain stable throughout the cell cycle, induce unscheduled transcription in G2 and mitosis, and promote faster entry into the next S phase. However, in the long term, they impair cell fitness. We propose that by restricting E2F activity to the S phase, cyclin F controls one of the main and most critical transcriptional engines of the cell cycle. 10.1016/j.molcel.2019.04.010
    Pyruvate-Carboxylase-Mediated Anaplerosis Promotes Antioxidant Capacity by Sustaining TCA Cycle and Redox Metabolism in Liver. Cappel David A,Deja Stanisław,Duarte João A G,Kucejova Blanka,Iñigo Melissa,Fletcher Justin A,Fu Xiaorong,Berglund Eric D,Liu Tiemin,Elmquist Joel K,Hammer Suntrea,Mishra Prashant,Browning Jeffrey D,Burgess Shawn C Cell metabolism The hepatic TCA cycle supports oxidative and biosynthetic metabolism. This dual responsibility requires anaplerotic pathways, such as pyruvate carboxylase (PC), to generate TCA cycle intermediates necessary for biosynthesis without disrupting oxidative metabolism. Liver-specific PC knockout (LPCKO) mice were created to test the role of anaplerotic flux in liver metabolism. LPCKO mice have impaired hepatic anaplerosis, diminution of TCA cycle intermediates, suppressed gluconeogenesis, reduced TCA cycle flux, and a compensatory increase in ketogenesis and renal gluconeogenesis. Loss of PC depleted aspartate and compromised urea cycle function, causing elevated urea cycle intermediates and hyperammonemia. Loss of PC prevented diet-induced hyperglycemia and insulin resistance but depleted NADPH and glutathione, which exacerbated oxidative stress and correlated with elevated liver inflammation. Thus, despite catalyzing the synthesis of intermediates also produced by other anaplerotic pathways, PC is specifically necessary for maintaining oxidation, biosynthesis, and pathways distal to the TCA cycle, such as antioxidant defenses. 10.1016/j.cmet.2019.03.014