Abstract
BackgroundTumor cell repopulation after radiotherapy is a major cause for the tumor radioresistance and recurrence. This study aims to investigate the underlying mechanism of tumor repopulation after radiotherapy, with focus on whether and how necroptosis takes part in this process.MethodsNecroptosis after irradiation were examined in vitro and in vivo. And the growth-promoting effect of necroptotic cells was investigated by chemical inhibitors or shRNA against necroptosis associated proteins and genes in in vitro and in vivo tumor repopulation models. Downstream relevance factors of necroptosis were identified by western blot and chemiluminescent immunoassays. Finally, the immunohistochemistry staining of identified necroptosis association growth stimulation factor was conducted in human colorectal tumor specimens to verify the relationship with clinical outcome.ResultsRadiation-induced necroptosis depended on activation of RIP1/RIP3/MLKL pathway, and the evidence in vitro and in vivo demonstrated that the inhibition of necroptosis attenuated growth-stimulating effects of irradiated tumor cells on living tumor reporter cells. The JNK/IL-8 were identified as downstream molecules of pMLKL during necroptosis, and inhibition of JNK, IL-8 or IL-8 receptor significantly reduced tumor repopulation after radiotherapy. Moreover, the high expression of IL-8 was associated with poor clinical prognosis in colorectal cancer patients.ConclusionsNecroptosis associated tumor repopulation after radiotherapy depended on activation of RIP1/RIP3/MLKL/JNK/IL-8 pathway. This novel pathway provided new insight into understanding the mechanism of tumor radioresistance and repopulation, and MLKL/JNK/IL-8 could be developed as promising targets for blocking tumor repopulation to enhance the efficacy of colorectal cancer radiotherapy.
Highlights
Tumor cell repopulation after radiotherapy is a major cause for the tumor radioresistance and recurrence
We provided evidence that radiation-induced necroptosis is dependent on sequential activation of receptor interacting protein 1 (RIP1)/receptor interacting protein 3 (RIP3)/Mixed lineage kinase domain-like protein (MLKL), and necroptosis contributes to tumor repopulation through the MLKL/c-jun N-terminal kinase (JNK)/IL-8 axis
To examine whether necroptosis involved in the process, we first evaluated the levels of RIP1, RIP3 and MLKL endogenous expression in HT29 and HCT116 cells by use of western blot analysis
Summary
Tumor cell repopulation after radiotherapy is a major cause for the tumor radioresistance and recurrence. This study aims to investigate the underlying mechanism of tumor repopulation after radiotherapy, with focus on whether and how necroptosis takes part in this process. It is noteworthy that more than 50% of cancer patients received RT in the course of their disease treatment, and in which 40% can be cured by RT [1,2,3]. Numerous studies are dedicated to explore the molecular mechanisms of this process, and a few growth-related signal pathways, cancer stem cells and tumor microenvironment such as hypoxia, tumor educated macrophages or fibroblasts are reported to be implicated [5,6,7]. Apoptosis and necrosis have been recognized as positive processes for cancer treatment.
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