HIF2alpha inhibition promotes p53 pathway activity, tumor cell death, and radiation responses.
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ABSTRACT:
SUBMITTER: Bertout JA
PROVIDER: S-EPMC2726037 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Glioblastoma is the most frequent and malignant brain tumor. Treatment includes chemotherapy with temozolomide concomitant with surgical resection and/or irradiation. However, a number of cases are resistant to temozolomide, as well as the human glioblastoma cell line U138-MG. We investigated doxazosin's (an antihypertensive drug) activity against glioblastoma cells (C6 and U138-MG) and its neurotoxicity on primary astrocytes and organoptypic hippocampal cultures. For this study, the following methods were used: citotoxicity assays, flow cytometry, western-blotting and confocal microscopy. We showed that doxazosin induces cell death on C6 and U138-MG cells. We observed that doxazosin's effects on the PI3K/Akt pathway were similar as LY294002 (PI3K specific inhibitor). In glioblastoma cells treated with doxasozin, Akt levels were greatly reduced. Upon examination of activities of proteins downstream of Akt we observed upregulation of GSK-3? and p53. This led to cell proliferation inhibition, cell death induction via caspase-3 activation and cell cycle arrest at G0/G1 phase in glioblastoma cells. We used in this study Lapatinib, a tyrosine kinase inhibitor, as a comparison with doxazosin because they present similar chemical structure. We also tested the neurocitotoxicity of doxazosin in primary astrocytes and organotypic cultures and observed that doxazosin induced cell death on a small percentage of non-tumor cells. Aggressiveness of glioblastoma tumors and dismal prognosis require development of new treatment agents. This includes less toxic drugs, more selective towards tumor cells, causing less damage to the patient. Therefore, our results confirm the potential of doxazosin as an attractive therapeutic antiglioma agent.
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Project description:Inactivation of p53 found in more than half of human cancers is often associated with increased tumor resistance to anti-cancer therapy. We have previously shown that overexpression of the phosphatase Wip1 in p53-negative tumors sensitizes them to chemotherapeutic agents, while protecting normal tissues from the side effects of anti-cancer treatment. In this study, we decided to search for kinases that prevent Wip1-mediated sensitization of cancer cells, thereby interfering with efficacy of genotoxic anti-cancer drugs. To this end, we performed a flow cytometry-based screening in order to identify kinases that regulated the levels of γH2AX, which were used as readout. Another criterion of the screen was increased sensitivity of p53-negative tumor cells to cisplatin (CDDP) in a Wip1-dependent manner. We have found that a treatment with a low dose (75 nM) of MK-1775, a recently described specific chemical inhibitor of Wee1, decreases CDDP-induced H2AX phosphorylation in p53-negative cells and enhances the Wip1-sensitization of p53-negative tumors. We were able to reduce CDDP effective concentration by 40% with a combination of Wip1 overexpression and Wee1 kinase inhibition. We have observed that Wee1 inhibition potentiates Wip1-dependent tumor sensitization effect by reducing levels of Hipk2 kinase, a negative regulator of Wip1 pathway. In addition, during CDDP treatment, the combination of Wee1 inhibition and Wip1 overexpression has a mild but significant protective effect in normal cells and tissues. Our results indicate that inhibition of the negative regulators of Wip1 pathway, Wee1 and Hipk2, in p53-negative tumors could potentiate efficiency of chemotherapeutic agents without concomitant increase of cytotoxicity in normal tissues. The development and clinical use of Wee1 and Hipk1 kinase chemical inhibitors might be a promising strategy to improve anti-cancer therapy.
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Project description:Tumor regression in sites distant to the irradiated field are thought to be associated with emission of damage-associated molecular patterns (DAMPs) molecules and generation of immunogenic cell death (ICD). Glioma stem cells (GSCs) are resistant to high doses of radiation, and ultimately select the outgrowth of a more aggressive tumor. This study showed high-dose IR triggered fewer DAMPs molecules exposure and release in GSCs comparing to matched non-GSCs. Downregulation of binding immunoglobulin protein (Bip) promoted IR-mediated endoplasmic reticulum stress to generate DAMPs molecules by PERK and IRE1-α phosphorylation, and increased dendritic cells mature and effector T lymphocytes activation. GSCs treated with Bip knockdown and IR efficiently prevented tumor generation, and reduced post-radiotherapy tumor recurrence. These data suggest that Bip plays a critical role in inhibition of IR-induced ICD in GSCs, and Bip inhibition may be a promising strategy on adjuvant therapy by ameliorating tumor immune microenvironment.
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Project description:Tetraploidy constitutes an adaptation to stress and an intermediate step between euploidy and aneuploidy in oncogenesis. Tetraploid cells are particularly resistant against genotoxic stress including radiotherapy and chemotherapy. Here, we designed a strategy to preferentially kill tetraploid tumor cells. Depletion of checkpoint kinase-1 (Chk1) by siRNAs, transfection with dominant-negative Chk1 mutants or pharmacological Chk1 inhibition killed tetraploid colon cancer cells yet had minor effects on their diploid counterparts. Chk1 inhibition abolished the spindle assembly checkpoint and caused premature and abnormal mitoses that led to p53 activation and cell death at a higher frequency in tetraploid than in diploid cells. Similarly, abolition of the spindle checkpoint by knockdown of Bub1, BubR1 or Mad2 induced p53-dependent apoptosis of tetraploid cells. Chk1 inhibition reversed the cisplatin resistance of tetraploid cells in vitro and in vivo, in xenografted human cancers. Chk1 inhibition activated p53-regulated transcripts including Puma/BBC3 in tetraploid but not in diploid tumor cells. Altogether, our results demonstrate that, in tetraploid tumor cells, the inhibition of Chk1 sequentially triggers aberrant mitosis, p53 activation and Puma/BBC3-dependent mitochondrial apoptosis.
| S-EPMC2131784 | biostudies-literature
Project description:BackgroundMalignant peripheral nerve sheath tumors (MPNSTs) are aggressive sarcomas that arise most commonly in the setting of the Neurofibromatosis Type 1 (NF1) cancer predisposition syndrome. Despite aggressive multimodality therapy, outcomes are dismal and most patients die within 5 years of diagnosis. Prior genomic studies in our laboratory identified tyrosine kinase 2 (TYK2) as a frequently mutated gene in MPNST. Herein, we explored the function of TYK2 in MPNST pathogenesis.MethodsImmunohistochemistry was utilized to examine expression of TYK2 in MPNSTs and other sarcomas. To establish a role for TYK2 in MPNST pathogenesis, murine and human TYK2 knockdown and knockout cells were established using shRNA and CRISPR/Cas9 systems, respectively.ResultsWe have demonstrated that TYK2 was highly expressed in the majority of human MPNSTs examined. Additionally, we demonstrated that knockdown of Tyk2/TYK2 in murine and human MPNST cells significantly increased cell death in vitro. These effects were accompanied by a decrease in the levels of activated Stats and Bcl-2 as well as an increase in the levels of Cleaved Caspase-3. In addition, Tyk2-KD cells demonstrated impaired growth in subcutaneous and metastasis models in vivo.ConclusionTaken together, these data illustrate the importance of TYK2 in MPNST pathogenesis and suggest that the TYK2 pathway may be a potential therapeutic target for these deadly cancers.
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Project description:Autophagy has emerged as a critical pathway in tumor development. S100A4 plays important roles in tumor metastasis, but its role in regulating autophagy has not been well characterized. In this study, we found that S100A4 was significantly upregulated in lung adenocarcinoma tissues. Clinical investigation demonstrated that high expression level of S100A4 was associated with tumor size and advanced tumor grades of lung adenocarcinoma patients. Moreover, our results revealed that extracellular S100A4 or overexpression of S100A4 inhibited starvation-induced autophagy and promoted cell proliferation in lung cancer cells in vitro; whereas small interfering RNA (siRNA)-mediated suppression of S100A4 increased autophagy and reduced cell viability in both A549 and LLC cells. Additionally, S100A4 inhibited starvation-induced autophagy to promote tumor cell viability via the Wnt pathway. Increased expression of ?-catenin consistently led to a decreased LC3-II protein abundance. Further, the inhibitory effect of S100A4 on autophagy and its promotion role in cell proliferation was abolished in A549 and LLC cells using the receptor for advanced glycation end products (RAGE)-specific inhibitor (FPS-ZM1). S100A4-deficient mice showed retarded tumor development. This effect was well correlated with increased expression of autophagy markers. Our findings demonstrate that S100A4 promotes lung tumor development through inhibiting autophagy in a ?-catenin signaling and S100A4 receptor RAGE-dependent manner, which provides a novel mechanism of S100A4-associated promotion of tumor development.
| S-EPMC5833421 | biostudies-literature
Project description:PurposeThe radiosensitivity of the normal intestinal epithelium is the major limiting factor for definitive radiotherapy against abdominal malignancies. Radiosensitizers, which can be used without augmenting radiation toxicity to normal tissue, are still an unmet need. Inhibition of proteosomal degradation is being developed as a major therapeutic strategy for anticancer therapy as cancer cells are more susceptible to proteasomal inhibition-induced cytotoxicity compared with normal cells. Auranofin, a gold-containing antirheumatoid drug, blocks proteosomal degradation by inhibiting deubiquitinase inhibitors. In this study, we have examined whether auranofin selectively radiosensitizes colon tumors without promoting radiation toxicity in normal intestine.Experimental designThe effect of auranofin (10 mg/kg i.p.) on the radiation response of subcutaneous CT26 colon tumors and the normal gastrointestinal epithelium was determined using a mouse model of abdominal radiation. The effect of auranofin was also examined in a paired human colonic organoid system using malignant and nonmalignant tissues from the same patient.ResultsBoth in the mouse model of intestinal injury and in the human nonmalignant colon organoid culture, auranofin pretreatment prevented radiation toxicity and improved survival with the activation of p53/p21-mediated reversible cell-cycle arrest. However, in a mouse model of abdominal tumor and in human malignant colonic organoids, auranofin inhibited malignant tissue growth with inhibition of proteosomal degradation, induction of endoplasmic reticulum stress/unfolded protein response, and apoptosis.ConclusionsOur data suggest that auranofin is a potential candidate to be considered as a combination therapy with radiation to improve therapeutic efficacy against abdominal malignancies.
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Project description:ObjectivesBoth heat shock (HS) and ionizing radiation have an impact on the cell cycle and may induce cell cycle arrest or apoptosis. Mutations of the p53 gene are observed at a high frequency in human tumours and are recognized in about half of all human cancers. Sensitivity to radiation, heat and anticancer agents has been observed in p53(+/+) cells, but not in mutated or p53-deficient cells. Moreover, enhancement of radiosensitivity by HS has been observed in wild-type p53 cells but not in p53-deficient cells. The molecular mechanism of the differential cell response to HS or ionizing radiation is not yet understood.Materials and methodsDifferences in cellular response to radiation (200 kV X-ray, 1, 2, 5 Gy) and HS (39 degrees C, 41 degrees C and 43 degrees C for 30 min) on cell cycle progression of cultures of human p53 mutant cells were investigated by flow cytometry. In addition, the effects of stressors used on the expression of several heat shock genes (HSP27, HSP60, HSP70, HSC70, HSP75, HSP78, HSP90) were studied by reverse transcriptase-polymerase chain reaction.Results and conclusionsYet, with respect to HSP gene expression, different stressors produced similar effects. Combination of HS and radiation treatment significantly induced the transcription of the HSP70 gene above the level induced by each stressor alone. Cell cycle analysis, however, revealed striking differences in prolonged dynamics of cell division in response to each stressor. Thus, p53 status could be a useful indicator in predictive assays for hyperthermia cancer treatment in combination with radiation and/or chemotherapy.
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Project description:Maintenance of genomic integrity is essential for adult tissue homeostasis and defects in the DNA-damage response (DDR) machinery are linked to numerous pathologies including cancer. Here, we present evidence that the DDR exerts tumor suppressor activity in gliomas. We show that genes encoding components of the DDR pathway are frequently altered in human gliomas and that loss of elements of the ATM/Chk2/p53 cascade accelerates tumor formation in a glioma mouse model. We demonstrate that Chk2 is required for glioma response to ionizing radiation in vivo and is necessary for DNA-damage checkpoints in the neuronal stem cell compartment. Finally, we observed that the DDR is constitutively activated in a subset of human GBMs, and such activation correlates with regions of hypoxia.
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Project description:Pheochromocytomas and paragangliomas (PCCs/PGLs) are rare neuroendocrine tumors arising from chromaffin tissue located in the adrenal or ganglia of the sympathetic or parasympathetic nervous system. The treatment of non-resectable or metastatic PCCs/PGLs is still limited to palliative measures, including somatostatin type 2 receptor radionuclide therapy with [177Lu]Lu-DOTA-TATE as one of the most effective approaches to date. Nevertheless, the metabolic and molecular determinants of radiation response in PCCs/PGLs have not yet been characterized. This study investigates the effects of hypoxia-inducible factor 2 alpha (HIF2α) on the susceptibility of PCCs/PGLs to radiation treatments using spheroids grown from genetically engineered mouse pheochromocytoma (MPC) cells. The expression of Hif2α was associated with the significantly increased resistance of MPC spheroids to external X-ray irradiation and exposure to beta particle-emitting [177Lu]LuCl3 compared to Hif2α-deficient controls. Exposure to [177Lu]LuCl3 provided an increased long-term control of MPC spheroids compared to single-dose external X-ray irradiation. This study provides the first experimental evidence that HIF2α-associated pseudohypoxia contributes to a radioresistant phenotype of PCCs/PGLs. Furthermore, the external irradiation and [177Lu]LuCl3 exposure of MPC spheroids provide surrogate models for radiation treatments to further investigate the metabolic and molecular determinants of radiation responses in PCCs/PGLs and evaluate the effects of neo-adjuvant-in particular, radiosensitizing-treatments in combination with targeted radionuclide therapies.
| S-EPMC7865577 | biostudies-literature