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A Colorimetric Assay that Specifically Measures Granzyme B Proteolytic Activity: Hydrolysis of Boc-Ala-Ala-Asp-S-Bzl
Published on: November 28, 2014
NK cell-derived GZMB (granzyme B) suppresses glioblastoma radioresistance by blocking SDC1-mediated autophagosome
Jingze Yan1, Ruishen Feng1, Qin Qin1
1Department of Radiation Oncology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, China.
Abstract:
Radiotherapy is a fundamental step in the combined treatment of glioblastoma (GBM), while radioresistance of GBM causes limitation of therapeutic efficacy. Natural killer (NK) cells, a potential target of immunotherapy, have attracted considerable attention due to the robust cancer cell-targeted cytotoxicity in combined treatment with radiotherapy, suggesting NK cell regulation might be a radiosensitization strategy. Here we show that a cytotoxic subset of NK cells could be stimulated by ionizing radiation (IR) and accumulate in the GBM tumor microenvironment (TME). Co-culturing with NK cells significantly enhances the GBM cell response to IR, and pharmaceutically depleting NK cells in mice elevates IR-induced tumor growth delay. Specifically, GZMB should be the radiosensitization effector secreted by NK cells. Suppressing GZMB activity remarkably impairs NK-mediated GBM radiosensitization. Meanwhile, administrating exogenous GZMB improves irradiation dose-survival response in vitro or in a xenograft model. Mechanically, GZMB blocks autophagosome-lysosome fusion in GBM cells by directly recognizing and cleaving SDC1, a key regulator of autophagosome maturation, at the valine 225 and aspartate 228 sites. Uncleavable mutation of SDC1 reverses GZMB-mediated radiosensitization in GBM. Further studies demonstrate that cleavage of SDC1 obstructs the localization of TGM2, a key MAP1LC3/LC3 recognizer, on the lysosome surface. Clinical data reveal GBM patients with an SDC1 valine 225 or aspartate 228 mutation display lower response to radiotherapy. In this study, we disclose the critical role of NK cells in tumor radiotherapy through secreting GZMB and impeding autophagosome maturation, as well as propose a potential strategy combining radiotherapy and NK-based immunotherapy against radioresistant GBM.Abbreviations: DEGs: differentially expressed genes; GBM: glioblastoma; GZMB: granzyme B; IL: interleukin; IR: ionizing radiation; IRS: immunoreactive score; LAMP: lysosomal associated membrane protein; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; mSDC1: mutant SDC1; NK: natural killer; PRF1: perforin 1; SDC1: syndecan 1; SNAP29: synaptosome associated protein 29; SQSTM1: sequestosome 1; STX17: syntaxin 17; TGM2: transglutaminase 2; TME: tumor microenvironment; TGD: tumor growth delay; VAMP8: vesicle associated membrane protein 8; WT: wild type.
Insights
Natural killer (NK) cells enhance glioblastoma radiotherapy by secreting granzyme B (GZMB), which blocks autophagosome-lysosome fusion. This NK cell-mediated radiosensitization offers a promising immunotherapy strategy for radioresistant glioblastomas.
Area of Science:
- Oncology
- Immunology
- Cell Biology
Background:
- Glioblastoma (GBM) radiotherapy efficacy is limited by radioresistance.
- Natural killer (NK) cells show potential in cancer immunotherapy, especially combined with radiotherapy.
- NK cell regulation is a potential strategy for GBM radiosensitization.
Purpose of the Study:
- To investigate the role of NK cells in GBM radiosensitization.
- To identify the effector molecule responsible for NK-mediated radiosensitization.
- To elucidate the molecular mechanism by which NK cells enhance GBM response to radiotherapy.
Main Methods:
- Co-culturing GBM cells with NK cells and exposing to ionizing radiation (IR).
- Pharmacological depletion of NK cells in a mouse model.
- Assessing the role of granzyme B (GZMB) in NK-mediated radiosensitization.
- Investigating the interaction between GZMB and syndecan 1 (SDC1) in GBM cells.
- Analyzing clinical data from GBM patients.
Main Results:
- Ionizing radiation (IR) stimulates and accumulates cytotoxic NK cells in the GBM tumor microenvironment (TME).
- NK cell co-culture enhances GBM cell response to IR; NK cell depletion reduces IR-induced tumor growth delay.
- Granzyme B (GZMB) is the effector molecule mediating NK cell radiosensitization.
- GZMB cleaves SDC1 at specific sites, blocking autophagosome-lysosome fusion and impairing GBM cell autophagy.
- Clinical data show SDC1 mutations correlate with lower radiotherapy response in GBM patients.
Conclusions:
- NK cells play a critical role in enhancing glioblastoma radiotherapy efficacy.
- GZMB secreted by NK cells radiosensitizes GBM by inhibiting autophagosome maturation via SDC1 cleavage.
- Combining radiotherapy with NK cell-based immunotherapy is a potential strategy against radioresistant GBM.
