Acute induction of oncogenic Ras provokes cellular senescence involving the retinoblastoma (Rb) pathway, but the tumour suppressive potential of senescence in vivo remains elusive. Recently, Rb-mediated silencing of growth-promoting genes by heterochromatin formation associated with methylation of histone H3 lysine 9 (H3K9me) was identified as a critical feature of cellular senescence, which may depend on the histone methyltransferase Suv39h1. Here we show that Emicro-N-Ras transgenic mice harbouring targeted heterozygous lesions at the Suv39h1, or the p53 locus for comparison, succumb to invasive T-cell lymphomas that lack expression of Suv39h1 or p53, respectively. By contrast, most N-Ras-transgenic wild-type ('control') animals develop a non-lymphoid neoplasia significantly later. Proliferation of primary lymphocytes is directly stalled by a Suv39h1-dependent, H3K9me-related senescent growth arrest in response to oncogenic Ras, thereby cancelling lymphomagenesis at an initial step. Suv39h1-deficient lymphoma cells grow rapidly but, unlike p53-deficient cells, remain highly susceptible to adriamycin-induced apoptosis. In contrast, only control, but not Suv39h1-deficient or p53-deficient, lymphomas senesce after drug therapy when apoptosis is blocked. These results identify H3K9me-mediated senescence as a novel Suv39h1-dependent tumour suppressor mechanism whose inactivation permits the formation of aggressive but apoptosis-competent lymphomas in response to oncogenic Ras.

p53 and INK4a/ARF mutations promote tumorigenesis and drug resistance, in part, by disabling apoptosis. We show that primary murine lymphomas also respond to chemotherapy by engaging a senescence program controlled by p53 and p16(INK4a). Hence, tumors with p53 or INK4a/ARF mutations-but not those lacking ARF alone-respond poorly to cyclophosphamide therapy in vivo. Moreover, tumors harboring a Bcl2-mediated apoptotic block undergo a drug-induced cytostasis involving the accumulation of p53, p16(INK4a), and senescence markers, and typically acquire p53 or INK4a mutations upon progression to a terminal stage. Finally, mice bearing tumors capable of drug-induced senescence have a much better prognosis following chemotherapy than those harboring tumors with senescence defects. Therefore, cellular senescence contributes to treatment outcome in vivo.

Here, we developed a new synthetic lethal strategy for further optimizing the eradication of cancer stem cells (CSCs). Briefly, we show that chronic treatment with the FDA-approved antibiotic Doxycycline effectively reduces cellular respiration, by targeting mitochondrial protein translation. The expression of four mitochondrial DNA encoded proteins (MT-ND3, MT-CO2, MT-ATP6 and MT-ATP8) is suppressed, by up to 35-fold. This high selection pressure metabolically synchronizes the surviving cancer cell sub-population towards a predominantly glycolytic phenotype, resulting in metabolic inflexibility. We directly validated this Doxycycline-induced glycolytic phenotype, by using metabolic flux analysis and label-free unbiased proteomics. Next, we identified two natural products (Vitamin C and Berberine) and six clinically-approved drugs, for metabolically targeting the Doxycycline-resistant CSC population (Atovaquone, Irinotecan, Sorafenib, Niclosamide, Chloroquine, and Stiripentol). This new combination strategy allows for the more efficacious eradication of CSCs with Doxycycline, and provides a simple pragmatic solution to the possible development of Doxycycline-resistance in cancer cells. In summary, we propose the combined use of i) Doxycycline (Hit-1: targeting mitochondria) and ii) Vitamin C (Hit-2: targeting glycolysis), which represents a new synthetic-lethal metabolic strategy for eradicating CSCs. This type of metabolic Achilles' heel will allow us and others to more effectively "starve" the CSC population.

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Derailed cytokine and immune cell networks account for organ damage and clinical severity of COVID-19 [1][2][3][4] . Here we show that SARS-CoV-2, like other viruses, evokes cellular senescence as a primary stress response in infected cells. Virus-induced senescence (VIS) is indistinguishable from other forms of cellular senescence and accompanied by a senescence-associated secretory phenotype (SASP), composed of pro-inflammatory cytokines, extracellular matrix-active factors and pro-coagulatory mediators [5][6][7] . COVID-19 patients displayed markers of senescence in their airway mucosa in situ and elevated serum levels of SASP factors. Mirroring COVID-19 hallmark features such as macrophage and neutrophil infiltration, endothelial damage and widespread thrombosis in affected lung tissue 1,8,9 , in vitro assays demonstrated macrophage activation with SASP-reminiscent secretion, complement lysis and SASP-amplifying secondary senescence of endothelial cells, neutrophil extracellular trap (NET) formation as well as activation of platelets and the clotting cascade in response to supernatant of VIS cells, including SARS-CoV-2-induced senescence. Senolytics such as Navitoclax and Dasatinib/Quercetin selectively eliminated VIS cells, mitigated COVID-19-reminiscent lung disease and reduced inflammation in SARS-CoV-2-driven hamster and mouse models. Our findings mark VIS as pathogenic trigger of COVID-19-related cytokine escalation and organ damage, and suggest senolytic targeting of virus-infected cells as a novel treatment option against SARS-CoV-2 and perhaps other viral infections.The pandemic human pathogenic SARS-CoV-2 coronavirus causes upper respiratory infections and subsequently COVID-19 lung disease that may get further complicated by septic multi-organ failure and comes with significant mortality 10,11 . Escalating immune activation with massive cytokine release seems to drive severe COVID-19 1-3 , possibly more than the virus infection itself. Mechanisms of viral

Activated RAS/BRAF oncogenes induce cellular senescence as a tumor-suppressive barrier in early cancer development, at least in part, via an oncogene-evoked DNA damage response (DDR). In contrast, Myc activation-although producing a DDR as well-is known to primarily elicit an apoptotic countermeasure. Using the Emu-myc transgenic mouse lymphoma model, we show here in vivo that apoptotic lymphoma cells activate macrophages to secrete transforming growth factor beta (TGF-beta) as a critical non-cell-autonomous inducer of cellular senescence. Accordingly, neutralization of TGF-beta action, like genetic inactivation of the senescence-related histone methyltransferase Suv39h1, significantly accelerates Myc-driven tumor development via cancellation of cellular senescence. These findings, recapitulated in human aggressive B cell lymphomas, demonstrate that tumor-prompted stroma-derived signals may limit tumorigenesis by feedback senescence induction.

SUMMARY Oncogene-induced senescence, e.g., in melanocytic nevi, terminates the expansion of pre-malignant cells via transcriptional silencing of proliferation-related genes due to decoration of their promoters with repressive tri-methylated histone H3 lysine 9 (H3K9) marks. We show here that structurally distinct H3K9-active demethylases—the lysine-specific demethylase-1 (LSD1) and several Jumonji C domain-containing moieties (such as JMJD2C)—disable senescence and permit Ras/Braf-evoked transformation. In mouse and zebrafish models, enforced LSD1 or JMJD2C expression promoted Braf-V600E-driven melanomagenesis. A large subset of established melanoma cell lines and primary human melanoma samples presented with a collective upregulation of related and unrelated H3K9 demethylase activities, whose targeted inhibition restored senescence, even in Braf inhibitor-resistant melanomas, evoked secondary immune effects and controlled tumor growth in vivo.

In malignancies, enhanced nuclear factor-kB (NF-kB) activity is largely viewed as an oncogenic property that also confers resistance to chemotherapy. Recently, NF-kB has been postulated to participate in a senescence-associated and possibly senescence-reinforcing cytokine response, thereby suggesting a tumor-restraining role for NF-kB. Using a mouse lymphoma model and analyzing transcriptome and clinical data from lymphoma patients, we show here that therapy-induced senescence presents with and depends on active NF-kB signaling, whereas NF-kB simultaneously promotes resistance to apoptosis. Further characterization and genetic engineering of primary mouse lymphomas according to distinct NF-kB-related oncogenic networks reminiscent of diffuse large B-cell lymphoma (DLBCL) subtypes guided us to identify Bcl2-overexpressing germinal center B-cell-like (GCB) DLBCL as a clinically relevant subgroup with significantly superior outcome when NF-kB is hyperactive. Our data illustrate the power of cross-species investigations to functionally test genetic mechanisms in transgenic mouse tumors that recapitulate distinct features of the corresponding human entity, and to ultimately use the mouse model-derived genetic information to redefine novel, clinically relevant patient subcohorts.