Liver cancer remains difficult to treat, owing to a paucity of drugs that target critical dependencies
; broad-spectrum kinase inhibitors such as sorafenib provide only a modest benefit to patients ...with hepatocellular carcinoma
. The induction of senescence may represent a strategy for the treatment of cancer, especially when combined with a second drug that selectively eliminates senescent cancer cells (senolysis)
. Here, using a kinome-focused genetic screen, we show that pharmacological inhibition of the DNA-replication kinase CDC7 induces senescence selectively in liver cancer cells with mutations in TP53. A follow-up chemical screen identified the antidepressant sertraline as an agent that kills hepatocellular carcinoma cells that have been rendered senescent by inhibition of CDC7. Sertraline suppressed mTOR signalling, and selective drugs that target this pathway were highly effective in causing the apoptotic cell death of hepatocellular carcinoma cells treated with a CDC7 inhibitor. The feedback reactivation of mTOR signalling after its inhibition
is blocked in cells that have been treated with a CDC7 inhibitor, which leads to the sustained inhibition of mTOR and cell death. Using multiple in vivo mouse models of liver cancer, we show that treatment with combined inhibition of of CDC7 and mTOR results in a marked reduction of tumour growth. Our data indicate that exploiting an induced vulnerability could be an effective treatment for liver cancer.
Hepatocellular carcinoma (HCC)-the most common form of liver cancer-is an aggressive malignancy with few effective treatment options
. Lenvatinib is a small-molecule inhibitor of multiple receptor ...tyrosine kinases that is used for the treatment of patients with advanced HCC, but this drug has only limited clinical benefit
. Here, using a kinome-centred CRISPR-Cas9 genetic screen, we show that inhibition of epidermal growth factor receptor (EGFR) is synthetic lethal with lenvatinib in liver cancer. The combination of the EGFR inhibitor gefitinib and lenvatinib displays potent anti-proliferative effects in vitro in liver cancer cell lines that express EGFR and in vivo in xenografted liver cancer cell lines, immunocompetent mouse models and patient-derived HCC tumours in mice. Mechanistically, inhibition of fibroblast growth factor receptor (FGFR) by lenvatinib treatment leads to feedback activation of the EGFR-PAK2-ERK5 signalling axis, which is blocked by EGFR inhibition. Treatment of 12 patients with advanced HCC who were unresponsive to lenvatinib treatment with the combination of lenvatinib plus gefitinib (trial identifier NCT04642547) resulted in meaningful clinical responses. The combination therapy identified here may represent a promising strategy for the approximately 50% of patients with advanced HCC who have high levels of EGFR.
The dependency of cancer cells on glutamine may be exploited therapeutically as a new strategy for treating cancers that lack druggable driver genes. Here we found that human liver cancer was ...dependent on extracellular glutamine. However, targeting glutamine addiction using the glutaminase inhibitor CB-839 as monotherapy had a very limited anticancer effect, even against the most glutamine addicted human liver cancer cells. Using a chemical library, we identified V-9302, a novel inhibitor of glutamine transporter ASCT2, as sensitizing glutamine dependent (GD) cells to CB-839 treatment. Mechanically, a combination of CB-839 and V-9302 depleted glutathione and induced reactive oxygen species (ROS), resulting in apoptosis of GD cells. Moreover, this combination also showed tumor inhibition in HCC xenograft mouse models in vivo. Our findings indicate that dual inhibition of glutamine metabolism by targeting both glutaminase and glutamine transporter ASCT2 represents a potential novel treatment strategy for glutamine addicted liver cancers.
Cellular senescence is characterized as a stable proliferation arrest that can be triggered by multiple stresses. Most knowledge about senescent cells is obtained from studies in primary cells. ...However, senescence features may be different in cancer cells, since the pathways that are involved in senescence induction are often deregulated in cancer. We report here a comprehensive analysis of the transcriptome and senolytic responses in a panel of 13 cancer cell lines rendered senescent by two distinct compounds. We show that in cancer cells, the response to senolytic agents and the composition of the senescence-associated secretory phenotype are more influenced by the cell of origin than by the senescence trigger. Using machine learning, we establish the SENCAN gene expression classifier for the detection of senescence in cancer cell samples. The expression profiles and senescence classifier are available as an interactive online Cancer SENESCopedia.
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•Senescent cancer cells respond differently to senolytic ABT-263•SASP expression in cancer is heterogeneous and influenced by cell origin•The SENCAN classifier detects cancer cell senescence in vitro•The Cancer SENESCopedia contains transcriptome data from 37 senescence models
Jochems et al. define common vulnerabilities of senescent cancer cells and shared features for the unequivocal detection of cancer cell senescence. Comprehensive analysis in a cancer cell panel reveals the context dependency of cancer cell senescence and allows the establishment of a SENCAN classifier to detect cancer cell senescence.
Senescence is a proliferation arrest that can result from a variety of stresses. Cancer cells can also undergo senescence, but the stresses that provoke cancer cells to undergo senescence are ...unclear. Here, we use both functional genetic and compound screens in cancer cells harboring a reporter that is activated during senescence to find targets that induce senescence. We show that suppression of the SWI/SNF component SMARCB1 induces senescence in melanoma through strong activation of the MAP kinase pathway. From the compound screen, we identified multiple aurora kinase inhibitors as potent inducers of senescence in RAS mutant lung cancer. Senescent melanoma and lung cancer cells acquire sensitivity to the BCL2 family inhibitor ABT263. We propose a one-two punch approach for the treatment of cancer in which a drug is first used to induce senescence in cancer cells and a second drug is then used to kill senescent cancer cells.
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•CRISPR and chemical screens identify senescence inducers in cancer cells•SMARCB1 knockout induces senescence in melanoma•Aurora kinase inhibition induces senescence in multiple cancer types•Senescent cancer cells become vulnerable to killing by ABT263
Wang et al. find that CRISPR-mediated genetic screens and chemical screens serve as two types of high-throughput methods to identify senescence inducers in cancer cells. They also show that senescent cancer cells can be killed selectively by the BCL2-family inhibitor ABT263, providing a potential sequential drug treatment strategy for cancer.
Inducing senescence in cancer cells is emerging as a new therapeutic strategy. In order to find ways to enhance senescence induction by palbociclib, a CDK4/6 inhibitor approved for treatment of ...metastatic breast cancer, we performed functional genetic screens in palbociclib-resistant cells. Using this approach, we found that loss of CDK2 results in strong senescence induction in palbociclib-treated cells. Treatment with the CDK2 inhibitor indisulam, which phenocopies genetic CDK2 inactivation, led to sustained senescence induction when combined with palbociclib in various cell lines and lung cancer xenografts. Treating cells with indisulam led to downregulation of cyclin H, which prevented CDK2 activation. Combined treatment with palbociclib and indisulam induced a senescence program and sensitized cells to senolytic therapy. Our data indicate that inhibition of CDK2 through indisulam treatment can enhance senescence induction by CDK4/6 inhibition.
Abstract
Glioblastoma (GBM) is a fatal brain tumor with a great unmet need for better therapies. Research on the genomic landscape of GBM has revealed that three major proliferation pathways — the ...PI3K-mTOR-AKT, the MAPK, and the CDK4/6-RB pathways — are overactive in the majority of GBM patients. Based on that knowledge we have developed a multi-target combination therapy (MTCT) by employing inhibitors of these three pathways. When these inhibitors are used in combination, lower, clinically more relevant concentrations of each drug are sufficient for target inhibition and proliferation arrest than with single agents. Excitingly, we find that MTCT not only shuts down proliferation but also causes permanent growth inhibition by the induction of senescence and the secretion of a senescence-associated secretory phenotype (SASP). MTCT-senescent cells have increased mitochondrial reactive oxygen species (mtROS). However, they do not acquire nuclear DNA damage but only local mitochondrial DNA damage. Addition of antioxidants that deplete mtROS resulted in senescence rescue both in terms of morphology and proliferation arrest. MTCT-senescent cells display an enhancement in total mitochondrial mass, mitochondrial DNA copy numbers, and the transcription of mitochondrial proteins. Despite this enhancement, the basal respiration rate of these cells is comparable to untreated cells, indicating an impaired maximal respiration capacity in MTCT-senescent cells. Concordantly, these cells display striking disturbances of their mitochondrial network. Intriguingly, depletion of functional mitochondria prevents the induction of mtROS and senescence by MTCT but not by classical senescence inducers such as ionizing radiation or high concentrations of CDK4/6 inhibitors. To conclude, MTCT induces senescence in GBM cells through a unique nuclear DNA damage-independent mechanism for which the mitochondria are essential.
Pro-senescence therapies are increasingly being considered for the treatment of cancer. Identifying additional targets to induce senescence in cancer cells could further enable such therapies. ...However, screening for targets whose suppression induces senescence on a genome-wide scale is challenging, as senescent cells become growth arrested, and senescence-associated features can take 1 to 2 weeks to develop. For a screen with a whole-genome CRISPR library, this would result in billions of undesirable proliferating cells by the time the senescent features emerge in the growth arrested cells. Here, we present a suicide switch system that allows genome-wide CRISPR screening in growth-arrested subpopulations by eliminating the proliferating cells during the screen through activation of a suicide switch in proliferating cells. Using this system, we identify in a genome-scale CRISPR screen several autophagy-related proteins as targets for senescence induction. We show that inhibiting macroautophagy with a small molecule ULK1 inhibitor can induce senescence in cancer cell lines of different origin. Finally, we show that combining ULK1 inhibition with the senolytic drug ABT-263 leads to apoptosis in a panel of cancer cell lines. IMPLICATIONS: Our suicide switch approach allows for genome-scale identification of pro-senescence targets, and can be adapted to simplify other screens depending on the nature of the promoter used to drive the switch.
Senolytics, drugs that kill senescent cells, have been proposed to improve the response to pro-senescence cancer therapies; however, this remains challenging due to a lack of broadly acting senolytic ...drugs. Using CRISPR/Cas9-based genetic screens in different senescent cancer cell models, we identify loss of the death receptor inhibitor cFLIP as a common vulnerability of senescent cancer cells. Senescent cells are primed for apoptotic death by NF-κB-mediated upregulation of death receptor 5 (DR5) and its ligand TRAIL, but are protected from death by increased cFLIP expression. Activation of DR5 signaling by agonistic antibody, which can be enhanced further by suppression of cFLIP by BRD2 inhibition, leads to efficient killing of a variety of senescent cancer cells. Moreover, senescent cells sensitize adjacent non-senescent cells to killing by DR5 agonist through a bystander effect mediated by secretion of cytokines. We validate this 'one-two punch' cancer therapy by combining pro-senescence therapy with DR5 activation in different animal models.
Drug-tolerant persisters (DTPs) are a rare subpopulation of cells within a tumor that can survive therapy through nongenetic adaptive mechanisms to develop relapse and repopulate the tumor following ...drug withdrawal. Using a cancer cell line with an engineered suicide switch to kill proliferating cells, we perform both genetic screens and compound screens to identify the inhibition of bromodomain and extraterminal domain (BET) proteins as a selective vulnerability of DTPs. BET inhibitors are especially detrimental to DTPs that have reentered the cell cycle (DTEPs) in a broad spectrum of cancer types. Mechanistically, BET inhibition induces lethal levels of ROS through the suppression of redox-regulating genes highly expressed in DTPs, including GPX2, ALDH3A1, and MGST1. In vivo BET inhibitor treatment delays tumor relapse in both melanoma and lung cancer. Our study suggests that combining standard of care therapy with BET inhibitors to eliminate residual persister cells is a promising therapeutic strategy.
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•Unbiased genetic screen and compound screen identify BRD2 as a vulnerability of DTPs•BET inhibitors suppress outgrowth of DTPs in a broad spectrum of cancer types•BET inhibitors suppress DTEPs through transcriptional repression of GPX2/ALDH3A1/MGST1•BET inhibitors delay tumor relapse
Chen et al. uses high-throughput screening approaches to identify BRD2 inhibition as a vulnerability of drug-tolerant persisters (DTPs). They further demonstrate that BET inhibitors suppress the emergence of DTPs in multiple cancer cell lines in vitro and can forestall drug resistance through the eradication of DTPs in vivo.
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