The PI3-K/AKT-pathway and radiation resistance mechanisms in non-small cell lung cancer.
Schuurbiers Olga C J,Kaanders Johannes H A M,van der Heijden Henricus F M,Dekhuijzen Richard P N,Oyen Wim J G,Bussink Johan
Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer
The phosphatidylinositol-3-kinase (PI3-K)/protein kinase B (AKT) pathway is associated with all three major radiation resistance mechanisms: intrinsic radiosensitivity, tumor cell proliferation, and hypoxia. In cell signaling cascades, the PI3-K/AKT signaling pathway is a key regulator of normal and cancerous growth and cell fate decisions by processes such as proliferation, invasion, apoptosis, and induction of hypoxia-related proteins. Activation of this pathway can be the result of stimulation of receptor tyrosine kinases such as epidermal growth factor receptor or vascular endothelial growth factor receptor or from mutations or amplification of PI3-K or AKT itself which are frequently found in non-small cell lung cancer (NSCLC). Furthermore, several treatment modalities such as radiotherapy can stimulate this survival pathway. Monitoring and manipulation of this signal transduction pathway may have important implications for the management of NSCLC. Strong and independent associations were found between expression of activated AKT (pAKT) and treatment outcome in clinical trials. Direct targeting and inhibition of this pathway may increase radiosensitivity by antagonizing the radiation induced cellular defense mechanisms especially in tumors that have activated the PI3-K/AKT cascade. To successfully implement these treatments in daily practice, there is a need for molecular predictors of sensitivity to inhibitors of PI3-K/AKT activation. In conclusion, the PI3-K/AKT pathway plays a crucial role in cellular defense mechanisms. Therefore, quantification of the activation status is a potential parameter for predicting treatment outcome. More importantly, specific targeting of this pathway in combination with radiotherapy or chemotherapy may enhance tumor control in NSCLC by antagonizing cellular defense in response to treatment.
Non-canonical NF-κB Antagonizes STING Sensor-Mediated DNA Sensing in Radiotherapy.
Hou Yuzhu,Liang Hua,Rao Enyu,Zheng Wenxin,Huang Xiaona,Deng Liufu,Zhang Yuan,Yu Xinshuang,Xu Meng,Mauceri Helena,Arina Ainhoa,Weichselbaum Ralph R,Fu Yang-Xin
The NF-κB pathway plays a crucial role in supporting tumor initiation, progression, and radioresistance of tumor cells. However, the role of the NF-κB pathway in radiation-induced anti-tumor host immunity remains unclear. Here we demonstrated that inhibiting the canonical NF-κB pathway dampened the therapeutic effect of ionizing radiation (IR), whereas non-canonical NF-κB deficiency promoted IR-induced anti-tumor immunity. Mechanistic studies revealed that non-canonical NF-κB signaling in dendritic cells (DCs) was activated by the STING sensor-dependent DNA-sensing pathway. By suppressing recruitment of the transcription factor RelA onto the Ifnb promoter, activation of the non-canonical NF-κB pathway resulted in decreased type I IFN expression. Administration of a specific inhibitor of the non-canonical NF-κB pathway enhanced the anti-tumor effect of IR in murine models. These findings reveal the potentially interactive roles for canonical and non-canonical NF-κB pathways in IR-induced STING-IFN production and provide an alternative strategy to improve cancer radiotherapy.
ATM-mediated stabilization of ZEB1 promotes DNA damage response and radioresistance through CHK1.
Zhang Peijing,Wei Yongkun,Wang Li,Debeb Bisrat G,Yuan Yuan,Zhang Jinsong,Yuan Jingsong,Wang Min,Chen Dahu,Sun Yutong,Woodward Wendy A,Liu Yongqing,Dean Douglas C,Liang Han,Hu Ye,Ang K Kian,Hung Mien-Chie,Chen Junjie,Ma Li
Nature cell biology
Epithelial-mesenchymal transition (EMT) is associated with characteristics of breast cancer stem cells, including chemoresistance and radioresistance. However, it is unclear whether EMT itself or specific EMT regulators play causal roles in these properties. Here we identify an EMT-inducing transcription factor, zinc finger E-box binding homeobox 1 (ZEB1), as a regulator of radiosensitivity and DNA damage response. Radioresistant subpopulations of breast cancer cells derived from ionizing radiation exhibit hyperactivation of the kinase ATM and upregulation of ZEB1, and the latter promotes tumour cell radioresistance in vitro and in vivo. Mechanistically, ATM phosphorylates and stabilizes ZEB1 in response to DNA damage, ZEB1 in turn directly interacts with USP7 and enhances its ability to deubiquitylate and stabilize CHK1, thereby promoting homologous recombination-dependent DNA repair and resistance to radiation. These findings identify ZEB1 as an ATM substrate linking ATM to CHK1 and the mechanism underlying the association between EMT and radioresistance.
miR-205 acts as a tumour radiosensitizer by targeting ZEB1 and Ubc13.
Zhang Peijing,Wang Li,Rodriguez-Aguayo Cristian,Yuan Yuan,Debeb Bisrat G,Chen Dahu,Sun Yutong,You M James,Liu Yongqing,Dean Douglas C,Woodward Wendy A,Liang Han,Yang Xianbin,Lopez-Berestein Gabriel,Sood Anil K,Hu Ye,Ang K Kian,Chen Junjie,Ma Li
Tumour cells associated with therapy resistance (radioresistance and drug resistance) are likely to give rise to local recurrence and distant metastatic relapse. Recent studies revealed microRNA (miRNA)-mediated regulation of metastasis and epithelial-mesenchymal transition; however, whether specific miRNAs regulate tumour radioresistance and can be exploited as radiosensitizing agents remains unclear. Here we find that miR-205 promotes radiosensitivity and is downregulated in radioresistant subpopulations of breast cancer cells, and that loss of miR-205 is highly associated with poor distant relapse-free survival in breast cancer patients. Notably, therapeutic delivery of miR-205 mimics via nanoliposomes can sensitize the tumour to radiation in a xenograft model. Mechanistically, radiation suppresses miR-205 expression through ataxia telangiectasia mutated (ATM) and zinc finger E-box binding homeobox 1 (ZEB1). Moreover, miR-205 inhibits DNA damage repair by targeting ZEB1 and the ubiquitin-conjugating enzyme Ubc13. These findings identify miR-205 as a radiosensitizing miRNA and reveal a new therapeutic strategy for radioresistant tumours.
The brain-penetrant clinical ATM inhibitor AZD1390 radiosensitizes and improves survival of preclinical brain tumor models.
Durant Stephen T,Zheng Li,Wang Yingchun,Chen Kan,Zhang Lingli,Zhang Tianwei,Yang Zhenfan,Riches Lucy,Trinidad Antonio G,Fok Jacqueline H L,Hunt Tom,Pike Kurt G,Wilson Joanne,Smith Aaron,Colclough Nicola,Reddy Venkatesh Pilla,Sykes Andrew,Janefeldt Annika,Johnström Peter,Varnäs Katarina,Takano Akihiro,Ling Stephanie,Orme Jonathan,Stott Jonathan,Roberts Caroline,Barrett Ian,Jones Gemma,Roudier Martine,Pierce Andrew,Allen Jasmine,Kahn Jenna,Sule Amrita,Karlin Jeremy,Cronin Anna,Chapman Melissa,Valerie Kristoffer,Illingworth Ruth,Pass Martin
Poor survival rates of patients with tumors arising from or disseminating into the brain are attributed to an inability to excise all tumor tissue (if operable), a lack of blood-brain barrier (BBB) penetration of chemotherapies/targeted agents, and an intrinsic tumor radio-/chemo-resistance. Ataxia-telangiectasia mutated (ATM) protein orchestrates the cellular DNA damage response (DDR) to cytotoxic DNA double-strand breaks induced by ionizing radiation (IR). ATM genetic ablation or pharmacological inhibition results in tumor cell hypersensitivity to IR. We report the primary pharmacology of the clinical-grade, exquisitely potent (cell IC, 0.78 nM), highly selective [>10,000-fold over kinases within the same phosphatidylinositol 3-kinase-related kinase (PIKK) family], orally bioavailable ATM inhibitor AZD1390 specifically optimized for BBB penetration confirmed in cynomolgus monkey brain positron emission tomography (PET) imaging of microdosed C-labeled AZD1390 (, 0.33). AZD1390 blocks ATM-dependent DDR pathway activity and combines with radiation to induce G cell cycle phase accumulation, micronuclei, and apoptosis. AZD1390 radiosensitizes glioma and lung cancer cell lines, with p53 mutant glioma cells generally being more radiosensitized than wild type. In in vivo syngeneic and patient-derived glioma as well as orthotopic lung-brain metastatic models, AZD1390 dosed in combination with daily fractions of IR (whole-brain or stereotactic radiotherapy) significantly induced tumor regressions and increased animal survival compared to IR treatment alone. We established a pharmacokinetic-pharmacodynamic-efficacy relationship by correlating free brain concentrations, tumor phospho-ATM/phospho-Rad50 inhibition, apoptotic biomarker (cleaved caspase-3) induction, tumor regression, and survival. On the basis of the data presented here, AZD1390 is now in early clinical development for use as a radiosensitizer in central nervous system malignancies.
Lovastatin sensitizes lung cancer cells to ionizing radiation: modulation of molecular pathways of radioresistance and tumor suppression.
Sanli Toran,Liu Caiqiong,Rashid Ayesha,Hopmans Sarah N,Tsiani Evangelia,Schultz Carrie,Farrell Thomas,Singh Gurmit,Wright James,Tsakiridis Theodoros
Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer
INTRODUCTION:In this study, we investigated the effect of the 3-hydroxy-3-methylgutaryl-CoA reductase inhibitor lovastatin, as a sensitizer of lung cancer cells to ionizing radiation (IR). METHODS:A549 lung adenocarcinoma cells were treated with 0 to 50 μM lovastatin alone or in combination with 0 to 8 Gy IR and subjected to clonogenic survival and proliferation assays. To assess the mechanism of drug action, we examined the effects of lovastatin and IR on the epidermal growth factor (EGF) receptor and AMP-activated kinase (AMPK) pathways and on apoptotic markers and the cell cycle. RESULTS:Lovastatin inhibited basal clonogenic survival and proliferation of A549 cells and sensitized them to IR. This was reversed by mevalonate, the product of 3-hydroxy-3-methylgutaryl-CoA reductase. Lovastatin attenuated selectively EGF-induced phosphorylation of EGF receptor and Akt, and IR-induced Akt phosphorylation, in a mevalonate-sensitive fashion, without inhibition on extracellular signal-regulated kinase 1/2 phosphorylation by either stimulus. IR phosphorylated and activated the metabolic sensor and tumor suppressor AMPK, but lovastatin enhanced basal and IR-induced AMPK phosphorylation. The drug inhibited IR-induced expression of p53 and the cyclin-dependent kinase inhibitors p21(cip1) and p27(kip1), but caused a redistribution of cells from G1-S phase (control and radiated cells) and G2-M phase (radiated cells) of cell cycle into apoptosis. The latter was also evident by induction of nuclear fragmentation and cleavage of caspase 3 by lovastatin in both control and radiated cells. CONCLUSIONS:We suggest that lovastatin inhibits survival and induces radiosensitization of lung cancer cells through induction of apoptosis, which may be mediated by a simultaneous inhibition of the Akt and activation of the AMPK signaling pathways.
Targeting PFKFB3 radiosensitizes cancer cells and suppresses homologous recombination.
Gustafsson Nina M S,Färnegårdh Katarina,Bonagas Nadilly,Ninou Anna Huguet,Groth Petra,Wiita Elisee,Jönsson Mattias,Hallberg Kenth,Lehto Jemina,Pennisi Rosa,Martinsson Jessica,Norström Carina,Hollers Jessica,Schultz Johan,Andersson Martin,Markova Natalia,Marttila Petra,Kim Baek,Norin Martin,Olin Thomas,Helleday Thomas
The glycolytic PFKFB3 enzyme is widely overexpressed in cancer cells and an emerging anti-cancer target. Here, we identify PFKFB3 as a critical factor in homologous recombination (HR) repair of DNA double-strand breaks. PFKFB3 rapidly relocates into ionizing radiation (IR)-induced nuclear foci in an MRN-ATM-γH2AX-MDC1-dependent manner and co-localizes with DNA damage and HR repair proteins. PFKFB3 relocalization is critical for recruitment of HR proteins, HR activity, and cell survival upon IR. We develop KAN0438757, a small molecule inhibitor that potently targets PFKFB3. Pharmacological PFKFB3 inhibition impairs recruitment of ribonucleotide reductase M2 and deoxynucleotide incorporation upon DNA repair, and reduces dNTP levels. Importantly, KAN0438757 induces radiosensitization in transformed cells while leaving non-transformed cells unaffected. In summary, we identify a key role for PFKFB3 enzymatic activity in HR repair and present KAN0438757, a selective PFKFB3 inhibitor that could potentially be used as a strategy for the treatment of cancer.
Tumor-reprogrammed resident T cells resist radiation to control tumors.
Arina Ainhoa,Beckett Michael,Fernandez Christian,Zheng Wenxin,Pitroda Sean,Chmura Steven J,Luke Jason J,Forde Martin,Hou Yuzhu,Burnette Byron,Mauceri Helena,Lowy Israel,Sims Tasha,Khodarev Nikolai,Fu Yang-Xin,Weichselbaum Ralph R
Successful combinations of radiotherapy and immunotherapy depend on the presence of live T cells within the tumor; however, radiotherapy is believed to damage T cells. Here, based on longitudinal in vivo imaging and functional analysis, we report that a large proportion of T cells survive clinically relevant doses of radiation and show increased motility, and higher production of interferon gamma, compared with T cells from unirradiated tumors. Irradiated intratumoral T cells can mediate tumor control without newly-infiltrating T cells. Transcriptomic analysis suggests T cell reprogramming in the tumor microenvironment and similarities with tissue-resident memory T cells, which are more radio-resistant than circulating/lymphoid tissue T cells. TGFβ is a key upstream regulator of T cell reprogramming and contributes to intratumoral Tcell radio-resistance. These findings have implications for the design of radio-immunotherapy trials in that local irradiation is not inherently immunosuppressive, and irradiation of multiple tumors might optimize systemic effects of radiotherapy.
MicroRNA 23b regulates autophagy associated with radioresistance of pancreatic cancer cells.
Wang Peng,Zhang Juan,Zhang Li,Zhu Zhengfei,Fan Jie,Chen Lianyu,Zhuang Liping,Luo Jianmin,Chen Hao,Liu Luming,Chen Zhen,Meng Zhiqiang
BACKGROUND & AIMS:Tumor resistance to radiation is a challenge in the treatment of patients with pancreatic cancer. Improving our understanding of the mechanisms of radioresistance could lead to strategies to increase patients' response to therapy. We investigated the roles of microRNAs (miRNAs) involved in radioresistance of pancreatic cancer cells. METHODS:We established radioresistant pancreatic cancer cell lines and used array analysis to compare levels of different miRNAs between radioresistant cell lines and the parental cell lines from which they were derived. We transfected pancreatic cancer cells with miRNA mimics or inhibitors and evaluated their effects on cell radiosensitivity using a clonogenic survival assay. The effects of miRNA on autophagy were determined by transmission electron microscopy and immunoblot analysis. We used a luciferase reporter assay to identify messenger RNA targets of specific miRNAs. RESULTS:Radioresistant pancreatic cancer cells had reduced levels of the miRNA miR-23b and increased autophagy compared with cells that were not radioresistant. Overexpression of miR-23b inhibited radiation-induced autophagy, whereas an inhibitor of miR-23b promoted autophagy in pancreatic cancer cells. Overexpression of miR-23b sensitized pancreatic cancer cells to radiation. The target of miR-23b, ATG12, was overexpressed in radioresistant cells; levels of ATG12 protein correlated with the occurrence of autophagy. Expression of miR-23b blocked radiation-induced autophagy and sensitized pancreatic cancer cells to radiation. We observed an inverse correlation between the level of miR-23b and autophagy in human pancreatic cancer tissue samples. CONCLUSIONS:In pancreatic cancer cells, reduced levels of the miRNA miR-23b increase levels of ATG12 and autophagy to promote radioresistance. miR-23b might be used to increase the sensitivity of pancreatic cancer cells to radiation therapy.
Mesenchymal differentiation mediated by NF-κB promotes radiation resistance in glioblastoma.
Bhat Krishna P L,Balasubramaniyan Veerakumar,Vaillant Brian,Ezhilarasan Ravesanker,Hummelink Karlijn,Hollingsworth Faith,Wani Khalida,Heathcock Lindsey,James Johanna D,Goodman Lindsey D,Conroy Siobhan,Long Lihong,Lelic Nina,Wang Suzhen,Gumin Joy,Raj Divya,Kodama Yoshinori,Raghunathan Aditya,Olar Adriana,Joshi Kaushal,Pelloski Christopher E,Heimberger Amy,Kim Se Hoon,Cahill Daniel P,Rao Ganesh,Den Dunnen Wilfred F A,Boddeke Hendrikus W G M,Phillips Heidi S,Nakano Ichiro,Lang Frederick F,Colman Howard,Sulman Erik P,Aldape Kenneth
Despite extensive study, few therapeutic targets have been identified for glioblastoma (GBM). Here we show that patient-derived glioma sphere cultures (GSCs) that resemble either the proneural (PN) or mesenchymal (MES) transcriptomal subtypes differ significantly in their biological characteristics. Moreover, we found that a subset of the PN GSCs undergoes differentiation to a MES state in a TNF-α/NF-κB-dependent manner with an associated enrichment of CD44 subpopulations and radioresistant phenotypes. We present data to suggest that the tumor microenvironment cell types such as macrophages/microglia may play an integral role in this process. We further show that the MES signature, CD44 expression, and NF-κB activation correlate with poor radiation response and shorter survival in patients with GBM.
Stromal PTEN determines mammary epithelial response to radiotherapy.
Sizemore Gina M,Balakrishnan Subhasree,Thies Katie A,Hammer Anisha M,Sizemore Steven T,Trimboli Anthony J,Cuitiño Maria C,Steck Sarah A,Tozbikian Gary,Kladney Raleigh D,Shinde Neelam,Das Manjusri,Park Dongju,Majumder Sarmila,Krishnan Shiva,Yu Lianbo,Fernandez Soledad A,Chakravarti Arnab,Shields Peter G,White Julia R,Yee Lisa D,Rosol Thomas J,Ludwig Thomas,Park Morag,Leone Gustavo,Ostrowski Michael C
The importance of the tumor-associated stroma in cancer progression is clear. However, it remains uncertain whether early events in the stroma are capable of initiating breast tumorigenesis. Here, we show that in the mammary glands of non-tumor bearing mice, stromal-specific phosphatase and tensin homolog (Pten) deletion invokes radiation-induced genomic instability in neighboring epithelium. In these animals, a single dose of whole-body radiation causes focal mammary lobuloalveolar hyperplasia through paracrine epidermal growth factor receptor (EGFR) activation, and EGFR inhibition abrogates these cellular changes. By analyzing human tissue, we discover that stromal PTEN is lost in a subset of normal breast samples obtained from reduction mammoplasty, and is predictive of recurrence in breast cancer patients. Combined, these data indicate that diagnostic or therapeutic chest radiation may predispose patients with decreased stromal PTEN expression to secondary breast cancer, and that prophylactic EGFR inhibition may reduce this risk.
Radiation-induced acid ceramidase confers prostate cancer resistance and tumor relapse.
Cheng Joseph C,Bai Aiping,Beckham Thomas H,Marrison S Tucker,Yount Caroline L,Young Katherine,Lu Ping,Bartlett Anne M,Wu Bill X,Keane Barry J,Armeson Kent E,Marshall David T,Keane Thomas E,Smith Michael T,Jones E Ellen,Drake Richard R,Bielawska Alicja,Norris James S,Liu Xiang
The Journal of clinical investigation
Escape of prostate cancer (PCa) cells from ionizing radiation-induced (IR-induced) killing leads to disease progression and cancer relapse. The influence of sphingolipids, such as ceramide and its metabolite sphingosine 1-phosphate, on signal transduction pathways under cell stress is important to survival adaptation responses. In this study, we demonstrate that ceramide-deacylating enzyme acid ceramidase (AC) was preferentially upregulated in irradiated PCa cells. Radiation-induced AC gene transactivation by activator protein 1 (AP-1) binding on the proximal promoter was sensitive to inhibition of de novo ceramide biosynthesis, as demonstrated by promoter reporter and ChIP-qPCR analyses. Our data indicate that a protective feedback mechanism mitigates the apoptotic effect of IR-induced ceramide generation. We found that deregulation of c-Jun induced marked radiosensitization in vivo and in vitro, which was rescued by ectopic AC overexpression. AC overexpression in PCa clonogens that survived a fractionated 80-Gy IR course was associated with increased radioresistance and proliferation, suggesting a role for AC in radiotherapy failure and relapse. Immunohistochemical analysis of human PCa tissues revealed higher levels of AC after radiotherapy failure than those in therapy-naive PCa, prostatic intraepithelial neoplasia, or benign tissues. Addition of an AC inhibitor to an animal model of xenograft irradiation produced radiosensitization and prevention of relapse. These data indicate that AC is a potentially tractable target for adjuvant radiotherapy.
Atm deletion with dual recombinase technology preferentially radiosensitizes tumor endothelium.
Moding Everett J,Lee Chang-Lung,Castle Katherine D,Oh Patrick,Mao Lan,Zha Shan,Min Hooney D,Ma Yan,Das Shiva,Kirsch David G
The Journal of clinical investigation
Cells isolated from patients with ataxia telangiectasia are exquisitely sensitive to ionizing radiation. Kinase inhibitors of ATM, the gene mutated in ataxia telangiectasia, can sensitize tumor cells to radiation therapy, but concern that inhibiting ATM in normal tissues will also increase normal tissue toxicity from radiation has limited their clinical application. Endothelial cell damage can contribute to the development of long-term side effects after radiation therapy, but the role of endothelial cell death in tumor response to radiation therapy remains controversial. Here, we developed dual recombinase technology using both FlpO and Cre recombinases to generate primary sarcomas in mice with endothelial cell-specific deletion of Atm to determine whether loss of Atm in endothelial cells sensitizes tumors and normal tissues to radiation. Although deletion of Atm in proliferating tumor endothelial cells enhanced the response of sarcomas to radiation, Atm deletion in quiescent endothelial cells of the heart did not sensitize mice to radiation-induced myocardial necrosis. Blocking cell cycle progression reversed the effect of Atm loss on tumor endothelial cell radiosensitivity. These results indicate that endothelial cells must progress through the cell cycle in order to be radiosensitized by Atm deletion.
Host STING-dependent MDSC mobilization drives extrinsic radiation resistance.
Liang Hua,Deng Liufu,Hou Yuzhu,Meng Xiangjiao,Huang Xiaona,Rao Enyu,Zheng Wenxin,Mauceri Helena,Mack Matthias,Xu Meng,Fu Yang-Xin,Weichselbaum Ralph R
Radiotherapy induces and promotes innate and adaptive immunity in which host STING plays an important role. However, radioresistance in irradiated tumors can also develop, resulting in relapse. Here we report a mechanism by which extrinsic resistance develops after local ablative radiation that relies on the immunosuppressive action of STING. The STING/type I interferon pathway enhances suppressive inflammation in tumors by recruiting myeloid cells in part via the CCR2 pathway. Germ-line knockouts of CCR2 or treatment with an anti-CCR2 antibody results in blockade of radiation-induced MDSC infiltration. Treatment with anti-CCR2 antibody alleviates immunosuppression following activation of the STING pathway, enhancing the anti-tumor effects of STING agonists and radiotherapy. We propose that radiation-induced STING activation is immunosuppressive due to (monocytic) M-MDSC infiltration, which results in tumor radioresistance. Furthermore, the immunosuppressive effects of radiotherapy and STING agonists can be abrogated in humans by a translational strategy involving anti-CCR2 antibody treatment to improve radiotherapy.
Radiation and checkpoint blockade immunotherapy: radiosensitisation and potential mechanisms of synergy.
Sharabi Andrew B,Lim Michael,DeWeese Theodore L,Drake Charles G
The Lancet. Oncology
Checkpoint blockade immunotherapy has received mainstream attention as a result of striking and durable clinical responses in some patients with metastatic disease and a reasonable response rate in many tumour types. The activity of checkpoint blockade immunotherapy is not restricted to melanoma or lung cancer, and additional indications are expected in the future, with responses already reported in renal cancer, bladder cancer, and Hodgkin's lymphoma among many others. Additionally, the interactions between radiation and the immune system have been investigated, with several studies describing the synergistic effects on local and distant tumour control when radiation therapy is combined with immunotherapy. Clinical enthusiasm for this approach is strengthened by the many ongoing trials combining immunotherapy with definitive and palliative radiation. Herein, we discuss the biological and mechanistic rationale behind combining radiation with checkpoint blockade immunotherapy, with a focus on the preclinical data supporting this potentially synergistic combination. We explore potential hypotheses and important considerations for clinical trial designs. Finally, we reintroduce the notion of radiosensitising immunotherapy, akin to radiosensitising chemotherapy, as a potential definitive therapeutic modality.
C-Met inhibitor MK-8003 radiosensitizes c-Met-expressing non-small-cell lung cancer cells with radiation-induced c-Met-expression.
Bhardwaj Vikas,Zhan Yanai,Cortez Maria Angelica,Ang Kie Kian,Molkentine David,Munshi Anupama,Raju Uma,Komaki Ritsuko,Heymach John V,Welsh James
Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer
INTRODUCTION:The radiation doses used to treat unresectable lung cancer are often limited by the proximity of normal tissues. Overexpression of c-Met, a receptor tyrosine kinase, occurs in about half of non-small-cell lung cancers (NSCLCs) and has been associated with resistance to radiation therapy and poor patient survival. We hypothesized that inhibiting c-Met would increase the sensitivity of NSCLC cells to radiation, enhancing the therapeutic ratio, which may potentially translate into improved local control. METHODS:We tested the radiosensitivity of two high-c-Met-expressing NSCLC lines, EBC-1 and H1993, and two low-c-Met-expressing lines, A549 and H460, with and without the small-molecule c-Met inhibitor MK-8033. Proliferation and protein expression were measured with clonogenic survival assays and Western blotting, respectively. γ-H2AX levels were evaluated by immunofluorescence staining. RESULTS:MK-8033 radiosensitized the high-c-Met-expressing EBC-1 and H1993 cells but not the low-c-Met-expressing cell lines A549 and H460. However, irradiation of A549 and H460 cells increased the expression of c-Met protein at 30 minutes after the irradiation. Subsequent targeting of this up-regulated c-Met by using MK-8033 followed by a second radiation dose reduced the clonogenic survival of both A549 and H460 cells. MK-8033 reduced the levels of radiation-induced phosphorylated (activated) c-Met in A549 cells. CONCLUSIONS:These results suggest that inhibition of c-Met could be an effective strategy to radiosensitize NSCLC tumors with high basal c-Met expression or tumors that acquired resistance to radiation because of up-regulation of c-Met.
Reprogramming the immunological microenvironment through radiation and targeting Axl.
Aguilera Todd A,Rafat Marjan,Castellini Laura,Shehade Hussein,Kariolis Mihalis S,Hui Angela Bik-Yu,Stehr Henning,von Eyben Rie,Jiang Dadi,Ellies Lesley G,Koong Albert C,Diehn Maximilian,Rankin Erinn B,Graves Edward E,Giaccia Amato J
Increasing evidence suggests that ionizing radiation therapy (RT) in combination with checkpoint immunotherapy is highly effective in treating a subset of cancers. To better understand the limited responses to this combination we analysed the genetic, microenvironmental, and immune factors in tumours derived from a transgenic breast cancer model. We identified two tumours with similar growth characteristics but different RT responses primarily due to an antitumour immune response. The combination of RT and checkpoint immunotherapy resulted in cures in the responsive but not the unresponsive tumours. Profiling the tumours revealed that the Axl receptor tyrosine kinase is overexpressed in the unresponsive tumours, and Axl knockout resulted in slower growth and increased radiosensitivity. These changes were associated with a CD8 T-cell response, which was improved in combination with checkpoint immunotherapy. These results suggest a novel role for Axl in suppressing antigen presentation through MHCI, and enhancing cytokine release, which promotes a suppressive myeloid microenvironment.
Inhibition of Nuclear PTEN Tyrosine Phosphorylation Enhances Glioma Radiation Sensitivity through Attenuated DNA Repair.
Ma Jianhui,Benitez Jorge A,Li Jie,Miki Shunichiro,Ponte de Albuquerque Claudio,Galatro Thais,Orellana Laura,Zanca Ciro,Reed Rachel,Boyer Antonia,Koga Tomoyuki,Varki Nissi M,Fenton Tim R,Nagahashi Marie Suely Kazue,Lindahl Erik,Gahman Timothy C,Shiau Andrew K,Zhou Huilin,DeGroot John,Sulman Erik P,Cavenee Webster K,Kolodner Richard D,Chen Clark C,Furnari Frank B
Ionizing radiation (IR) and chemotherapy are standard-of-care treatments for glioblastoma (GBM) patients and both result in DNA damage, however, the clinical efficacy is limited due to therapeutic resistance. We identified a mechanism of such resistance mediated by phosphorylation of PTEN on tyrosine 240 (pY240-PTEN) by FGFR2. pY240-PTEN is rapidly elevated and bound to chromatin through interaction with Ki-67 in response to IR treatment and facilitates the recruitment of RAD51 to promote DNA repair. Blocking Y240 phosphorylation confers radiation sensitivity to tumors and extends survival in GBM preclinical models. Y240F-Pten knockin mice showed radiation sensitivity. These results suggest that FGFR-mediated pY240-PTEN is a key mechanism of radiation resistance and is an actionable target for improving radiotherapy efficacy.
Role of KEAP1/NRF2 and TP53 Mutations in Lung Squamous Cell Carcinoma Development and Radiation Resistance.
Jeong Youngtae,Hoang Ngoc T,Lovejoy Alexander,Stehr Henning,Newman Aaron M,Gentles Andrew J,Kong William,Truong Diana,Martin Shanique,Chaudhuri Aadel,Heiser Diane,Zhou Li,Say Carmen,Carter Justin N,Hiniker Susan M,Loo Billy W,West Robert B,Beachy Philip,Alizadeh Ash A,Diehn Maximilian
Lung squamous cell carcinoma (LSCC) pathogenesis remains incompletely understood, and biomarkers predicting treatment response remain lacking. Here, we describe novel murine LSCC models driven by loss of Trp53 and Keap1, both of which are frequently mutated in human LSCCs. Homozygous inactivation of Keap1 or Trp53 promoted airway basal stem cell (ABSC) self-renewal, suggesting that mutations in these genes lead to expansion of mutant stem cell clones. Deletion of Trp53 and Keap1 in ABSCs, but not more differentiated tracheal cells, produced tumors recapitulating histologic and molecular features of human LSCCs, indicating that they represent the likely cell of origin in this model. Deletion of Keap1 promoted tumor aggressiveness, metastasis, and resistance to oxidative stress and radiotherapy (RT). KEAP1/NRF2 mutation status predicted risk of local recurrence after RT in patients with non-small lung cancer (NSCLC) and could be noninvasively identified in circulating tumor DNA. Thus, KEAP1/NRF2 mutations could serve as predictive biomarkers for personalization of therapeutic strategies for NSCLCs. SIGNIFICANCE:We developed an LSCC mouse model involving Trp53 and Keap1, which are frequently mutated in human LSCCs. In this model, ABSCs are the cell of origin of these tumors. KEAP1/NRF2 mutations increase radioresistance and predict local tumor recurrence in radiotherapy patients. Our findings are of potential clinical relevance and could lead to personalized treatment strategies for tumors with KEAP1/NRF2 mutations. Cancer Discov; 7(1); 86-101. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 1.
Antagonism of EGFR and Notch limits resistance to EGFR inhibitors and radiation by decreasing tumor-initiating cell frequency.
Hu Shi,Fu Wenyan,Li Tian,Yuan Qingning,Wang Feifei,Lv Gaojian,Lv Yuanyuan,Fan Xiaoyan,Shen Yafeng,Lin Fangxing,Tang Ying,Ye Xuting,Yang Yongji,Lei Changhai
Science translational medicine
Epidermal growth factor receptor (EGFR) blockade and radiation are efficacious in the treatment of cancer, but resistance is commonly reported. Studies have suggested that dysregulation of Notch signaling and enrichment of the cancer stem cell population underlie these treatment challenges. Our data show that dual targeting of EGFR and Notch2/3 receptors with antibody CT16 not only inhibited signaling mediated by these receptors but also showed a strong anti-stem cell effect both in vitro and in vivo. Treatment with CT16 prevented acquired resistance to EGFR inhibitors and radiation in non-small cell lung cancer (NSCLC) cell line models and patient-derived xenograft tumors. CT16 also had a superior radiosensitizing impact compared with EGFR inhibitors. CT16 in combination with radiation had a larger antitumor effect than the combination of radiation with EGFR inhibitors or tarextumab. Mechanistically, CT16 treatment inhibits the stem cell-like subpopulation, which has a high mesenchymal gene expression and DNA repair activity, and reduces tumor-initiating cell frequency. This finding highlights the capacity of a combined blockade of EGFR and Notch signaling to augment the response to radiation and suggests that CT16 may achieve clinical efficacy when combined with radiation in NSCLC treatment.
Polyoxometalate-Based Radiosensitization Platform for Treating Hypoxic Tumors by Attenuating Radioresistance and Enhancing Radiation Response.
Yong Yuan,Zhang Chunfang,Gu Zhanjun,Du Jiangfeng,Guo Zhao,Dong Xinghua,Xie Jiani,Zhang Guangjin,Liu Xiangfeng,Zhao Yuliang
Radioresistance is one of the undesirable impediments in hypoxic tumors, which sharply diminishes the therapeutic effectiveness of radiotherapy and eventually results in the failure of their treatments. An attractive strategy for attenuating radioresistance is developing an ideal radiosensitization system with appreciable radiosensitization capacity to attenuate tumor hypoxia and reinforce radiotherapy response in hypoxic tumors. Therefore, we describe the development of Gd-containing polyoxometalates-conjugated chitosan (GdW@CS nanosphere) as a radiosensitization system for simultaneous extrinsic and intrinsic radiosensitization, by generating an overabundance of cytotoxic reactive oxygen species (ROS) using high-energy X-ray stimulation and mediating the hypoxia-inducible factor-1a (HIF-1a) siRNA to down-regulate HIF-1α expression and suppress broken double-stranded DNA self-healing. Most importantly, the GdW@CS nanospheres have the capacity to promote the exhaustion of intracellular glutathione (reduced GSH) by synergy W-triggered GSH oxidation for sufficient ROS generation, thereby facilitating the therapeutic efficiency of radiotherapy. As a result, the as-synthesized GdW@CS nanosphere can overcome radioresistance of hypoxic tumors through a simultaneous extrinsic and intrinsic strategy to improve radiosensitivity. We have demonstrated GdW@CS nanospheres with special radiosensitization behavior, which provides a versatile approach to solve the critical radioresistance issue of hypoxic tumors.
Understanding the functions of tumor stroma in resistance to ionizing radiation: emerging targets for pharmacological modulation.
Chargari Cyrus,Clemenson Céline,Martins Isabelle,Perfettini Jean-Luc,Deutsch Eric
Drug resistance updates : reviews and commentaries in antimicrobial and anticancer chemotherapy
Maintenance of both normal epithelial tissues and their malignant counterparts is supported by the host tissue stroma. The tumor stroma mainly consists of the basement membrane, fibroblasts, extracellular matrix, immune cells, and vasculature. Although most host cells in the stroma possess certain tumor-suppressing abilities, the stroma will change during malignancy and eventually promote growth, invasion, and metastasis. There is growing evidence that the stroma influences importantly the response to radiation therapy (RT). On the one hand, irradiation releases numerous inflammatory cytokines within the extracellular matrix and activates tumor specific antigens presentation, triggering an immune reaction that contributes to the antitumor effect seen after RT. On the other hand, the stroma significantly contributes to radioresistance but also increases the metastatic risk. Indeed, fibroblasts, which are major actors of the impact of stroma on tumor response, are involved in activation of autocrine and paracrine molecular signaling pathways regulating tumor cell proliferation, cell death, response to hypoxia, DNA repair systems and mesenchymal-epithelial transition. They are also actors of the peritumoral desmoplastic reaction, which decreases tumor radiosensitivity. The irradiated stroma can also contribute to tumor relapse after RT through recruitment of bone marrow-derived progenitors that contribute to local tumor relapse through neovascularization. It is therefore time to question preclinical models that would not take into account this impact of stroma. The increasing knowledge of the relationship between stroma and response to IR could help developing innovative strategies for potentially improve antitumor effect of RT.
An IRAK1-PIN1 signalling axis drives intrinsic tumour resistance to radiation therapy.
Liu Peter H,Shah Richa B,Li Yuanyuan,Arora Arshi,Ung Peter Man-Un,Raman Renuka,Gorbatenko Andrej,Kozono Shingo,Zhou Xiao Zhen,Brechin Vincent,Barbaro John M,Thompson Ruth,White Richard M,Aguirre-Ghiso Julio A,Heymach John V,Lu Kun Ping,Silva Jose M,Panageas Katherine S,Schlessinger Avner,Maki Robert G,Skinner Heath D,de Stanchina Elisa,Sidi Samuel
Nature cell biology
Drug-based strategies to overcome tumour resistance to radiotherapy (R-RT) remain limited by the single-agent toxicity of traditional radiosensitizers (for example, platinums) and a lack of targeted alternatives. In a screen for compounds that restore radiosensitivity in p53 mutant zebrafish while tolerated in non-irradiated wild-type animals, we identified the benzimidazole anthelmintic oxfendazole. Surprisingly, oxfendazole acts via the inhibition of IRAK1, a kinase thus far implicated in interleukin-1 receptor (IL-1R) and Toll-like receptor (TLR) immune responses. IRAK1 drives R-RT in a pathway involving IRAK4 and TRAF6 but not the IL-1R/TLR-IRAK adaptor MyD88. Rather than stimulating nuclear factor-κB, radiation-activated IRAK1 prevented apoptosis mediated by the PIDDosome complex (comprising PIDD, RAIDD and caspase-2). Countering this pathway with IRAK1 inhibitors suppressed R-RT in tumour models derived from cancers in which TP53 mutations predict R-RT. Moreover, IRAK1 inhibitors synergized with inhibitors of PIN1, a prolyl isomerase essential for IRAK1 activation in response to pathogens and, as shown here, in response to ionizing radiation. These data identify an IRAK1 radiation-response pathway as a rational chemoradiation therapy target.
Hypoxia-induced p53 modulates both apoptosis and radiosensitivity via AKT.
Leszczynska Katarzyna B,Foskolou Iosifina P,Abraham Aswin G,Anbalagan Selvakumar,Tellier Céline,Haider Syed,Span Paul N,O'Neill Eric E,Buffa Francesca M,Hammond Ester M
The Journal of clinical investigation
Restoration of hypoxia-induced apoptosis in tumors harboring p53 mutations has been proposed as a potential therapeutic strategy; however, the transcriptional targets that mediate hypoxia-induced p53-dependent apoptosis remain elusive. Here, we demonstrated that hypoxia-induced p53-dependent apoptosis is reliant on the DNA-binding and transactivation domains of p53 but not on the acetylation sites K120 and K164, which, in contrast, are essential for DNA damage-induced, p53-dependent apoptosis. Evaluation of hypoxia-induced transcripts in multiple cell lines identified a group of genes that are hypoxia-inducible proapoptotic targets of p53, including inositol polyphosphate-5-phosphatase (INPP5D), pleckstrin domain-containing A3 (PHLDA3), sulfatase 2 (SULF2), B cell translocation gene 2 (BTG2), cytoplasmic FMR1-interacting protein 2 (CYFIP2), and KN motif and ankyrin repeat domains 3 (KANK3). These targets were also regulated by p53 in human cancers, including breast, brain, colorectal, kidney, bladder, and melanoma cancers. Downregulation of these hypoxia-inducible targets associated with poor prognosis, suggesting that hypoxia-induced apoptosis contributes to p53-mediated tumor suppression and treatment response. Induction of p53 targets, PHLDA3, and a specific INPP5D transcript mediated apoptosis in response to hypoxia through AKT inhibition. Moreover, pharmacological inhibition of AKT led to apoptosis in the hypoxic regions of p53-deficient tumors and consequently increased radiosensitivity. Together, these results identify mediators of hypoxia-induced p53-dependent apoptosis and suggest AKT inhibition may improve radiotherapy response in p53-deficient tumors.
The anti-malarial atovaquone increases radiosensitivity by alleviating tumour hypoxia.
Ashton Thomas M,Fokas Emmanouil,Kunz-Schughart Leoni A,Folkes Lisa K,Anbalagan Selvakumar,Huether Melanie,Kelly Catherine J,Pirovano Giacomo,Buffa Francesca M,Hammond Ester M,Stratford Michael,Muschel Ruth J,Higgins Geoff S,McKenna William Gillies
Tumour hypoxia renders cancer cells resistant to cancer therapy, resulting in markedly worse clinical outcomes. To find clinical candidate compounds that reduce hypoxia in tumours, we conduct a high-throughput screen for oxygen consumption rate (OCR) reduction and identify a number of drugs with this property. For this study we focus on the anti-malarial, atovaquone. Atovaquone rapidly decreases the OCR by more than 80% in a wide range of cancer cell lines at pharmacological concentrations. In addition, atovaquone eradicates hypoxia in FaDu, HCT116 and H1299 spheroids. Similarly, it reduces hypoxia in FaDu and HCT116 xenografts in nude mice, and causes a significant tumour growth delay when combined with radiation. Atovaquone is a ubiquinone analogue, and decreases the OCR by inhibiting mitochondrial complex III. We are now undertaking clinical studies to assess whether atovaquone reduces tumour hypoxia in patients, thereby increasing the efficacy of radiotherapy.
A signature motif in LIM proteins mediates binding to checkpoint proteins and increases tumour radiosensitivity.
Xu Xiaojie,Fan Zhongyi,Liang Chaoyang,Li Ling,Wang Lili,Liang Yingchun,Wu Jun,Chang Shaohong,Yan Zhifeng,Lv Zhaohui,Fu Jing,Liu Yang,Jin Shuai,Wang Tao,Hong Tian,Dong Yishan,Ding Lihua,Cheng Long,Liu Rui,Fu Shenbo,Jiao Shunchang,Ye Qinong
Tumour radiotherapy resistance involves the cell cycle pathway. CDC25 phosphatases are key cell cycle regulators. However, how CDC25 activity is precisely controlled remains largely unknown. Here, we show that LIM domain-containing proteins, such as FHL1, increase inhibitory CDC25 phosphorylation by forming a complex with CHK2 and CDC25, and sequester CDC25 in the cytoplasm by forming another complex with 14-3-3 and CDC25, resulting in increased radioresistance in cancer cells. FHL1 expression, induced by ionizing irradiation in a SP1- and MLL1-dependent manner, positively correlates with radioresistance in cancer patients. We identify a cell-penetrating 11 amino-acid motif within LIM domains (eLIM) that is sufficient for binding CHK2 and CDC25, reducing the CHK2-CDC25 and CDC25-14-3-3 interaction and enhancing CDC25 activity and cancer radiosensitivity accompanied by mitotic catastrophe and apoptosis. Our results provide novel insight into molecular mechanisms underlying CDC25 activity regulation. LIM protein inhibition or use of eLIM may be new strategies for improving tumour radiosensitivity.