Abstract
BackgroundGlioblastoma (GB) is the most common and highly malignant brain tumor characterized by aggressive growth and resistance to alkylating chemotherapy. Autophagy induction is one of the hallmark effects of anti-GB therapies with temozolomide (TMZ). However, the non-classical form of autophagy, autophagy-based unconventional secretion, also called secretory autophagy and its role in regulating the sensitivity of GB to TMZ remains unclear. There is an urgent need to illuminate the mechanism and to develop novel therapeutic targets for GB.MethodsCancer genome databases and paired-GB patient samples with or without TMZ treatment were used to assess the relationship between HMGB1 mRNA levels and overall patient survival. The relationship between HMGB1 protein level and TMZ sensitivity was measured by immunohistochemistry, ELISA, Western blot and qRT-PCR. GB cells were engineered to express a chimeric autophagic flux reporter protein consisting of mCherry, GFP and LC3B. The role of secretory autophagy in tumor microenvironment (TME) was analyzed by intracranial implantation of GL261 cells. Coimmunoprecipitation (Co-IP) and Western blotting were performed to test the RAGE-NFκB-NLRP3 inflammasome pathway.ResultsThe exocytosis of HMGB1 induced by TMZ in GB is dependent on the secretory autophagy. HMGB1 contributed to M1-like polarization of tumor associated macrophages (TAMs) and enhanced the sensitivity of GB cells to TMZ. Mechanistically, RAGE acted as a receptor for HMGB1 in TAMs and through RAGE-NFκB-NLRP3 inflammasome pathway, HMGB1 enhanced M1-like polarization of TAMs. Clinically, the elevated level of HMGB1 in sera may serve as a beneficial therapeutic-predictor for GB patients under TMZ treatment.ConclusionsWe demonstrated that enhanced secretory autophagy in GB facilitates M1-like polarization of TAMs to enhance TMZ sensitivity of GB cells. HMGB1 acts as a key regulator in the crosstalk between GB cells and tumor-suppressive M1-like TAMs in GB microenvironment and may be considered as an adjuvant for the chemotherapeutic agent TMZ.
Highlights
Glioblastoma (GB) is the most common and highly malignant brain tumor characterized by aggres‐ sive growth and resistance to alkylating chemotherapy
Exocytosis of High mobility group box 1 (HMGB1) induced by TMZ is dependent on the formation of autophagic vacuoles To determine whether TMZ promoted the exocytosis of leaderless proteins, the secretory autophagy proteins HMGB1, IL-33, Galectin-3, IL-1α, MIF, FGF1, AnnxinA1, S100A8, IL-1β, IL-37, IL-18 and FGF2, in supernatants of primary GB cells (GB1 and GB2 cells) and cell lines (U251 and LN229) under TMZ were analyzed
The mRNA of autophagy-related genes Autophagy-related 5 (ATG5), Autophagyrelated 7 (ATG7), Microtubuleassociated protein light chain (LC3B) and Beclin 1 (BECN1) were elevated in GB cells under TMZ treatment (Fig. S2A)
Summary
Glioblastoma (GB) is the most common and highly malignant brain tumor characterized by aggres‐ sive growth and resistance to alkylating chemotherapy. The standard treatment of GB consists of maximal resection followed by radiotherapy and concomitant chemotherapy with the alkylating agent temozolomide (TMZ) [3]. Autophagy, which recycles breakdown products to sustain cell metabolism and biosynthesis under stress conditions, has been proposed as a mechanism of chemoresistance to alkylating drugs [6]. The effect of autophagy inhibition in combination with TMZ did not improve the overall survival of patients with GB [5, 7]. The disputed effect of combination of autophagy inhibitors and TMZ in GB indicated that the role of autophagy in dependent on the context. The diverse roles of autophagy in cancer treatment have attracted considerable interest
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