XIAP Limits Autophagic Degradation of Sox2 and Is A Therapeutic Target in Nasopharyngeal Carcinoma Stem Cells

Abstract

Rationale: Nasopharyngeal carcinoma (NPC) is the most frequent head and neck tumor in South China. The presence of cancer stem cells (CSCs) in NPC contributes to tumor maintenance and therapeutic resistance, while the ability of CSCs to escape from the apoptosis pathway may render them the resistant property to the therapies. Inhibitor of apoptosis proteins family proteins (IAPs), which are overexpressed in nasopharyngeal carcinoma stem cells, may play an important role in maintaining nasopharyngeal cancer stem cell properties. Here, we develop a novel CSC-targeting strategy to treat NPC through inhibiting IAPs.Methods: Human NPC S-18 and S-26 cell lines were used as the model system in vitro and in vivo. Fluorescence activated cell sorting (FACS) assay was used to detect nasopharyngeal SP cells and CD44+ cells. The characteristics of CSCs were defined by sphere suspension culture, colony formation assay and cell migration. The role of XIAP on the regulation of Sox2 protein stability and ERK1-mediated phosphorylation of Sox2 signaling pathway were analyzed using immunoblotting, immunoprecipitation, immunofluorescence, phosphorylation mass spectrometry, siRNA silencing and plasmid overexpression. The correlation between XIAP and Sox2 in NPC biopsies and their role in prognosis was performed by immunohistochemistry. APG-1387 or chemotherapies-induced cell death and apoptosis in S-18 and S-26 were determined by WST, immunoblotting and flow cytometry assay.Results: IAPs, especially X chromosome-linked IAP (XIAP), were elevated in CSCs of NPC, and these proteins were critically involved in the maintenance of CSCs properties by enhancing the stability of Sox2. Mechanistically, ERK1 kinase promoted autophagic degradation of Sox2 via phosphorylation of Sox2 at Ser251 and further SUMOylation of Sox2 at Lys245 in non-CSCs. However, XIAP blocked autophagic degradation of Sox2 by inhibiting ERK1 activation in CSCs. Additionally, XIAP was positively correlated with Sox2 expression in NPC tissues, which were associated with NPC progression. Finally, we discovered that a novel antagonist of IAPs, APG-1387, exerted antitumor effect on CSCs. Also, the combination of APG-1387 with CDDP /5-FU has a synergistic effect on NPC.Conclusion: Our study highlights the importance of IAPs in the maintenance of CSCs in NPC. Thus, XIAP is a promising therapeutic target in CSCs and suggests that NPC patients may benefit from a combination treatment of APG-1387 with conventional chemotherapy.

The SMAC Mimetic APG-1387 Sensitizes Immune-Mediated Cell Apoptosis in Hepatocellular Carcinoma

“…Also, the expression of the CSC marker Sox2 in HCCLM3 cells was reduced by APG-1387 as monotherapy or combination treatment (Figure 3I ). These findings are consistent with previous findings in nasopharyngeal carcinoma cells in vitro ( Wu et al, 2013 ; Ji et al, 2018 ). To examine whether the combination of APG-1387 with TNF-α or TRAIL specifically targets SP cells, we sorted SP and MP cells from the HCCLM3 cell line.…”

Functional characterization of SOX2 as an anticancer target

“…91 A later study confirms that SOX2 phosphorylation on S251 site by ERK1/2 could promote SOX2 SUMOylation in nasopharyngeal carcinoma (NPC) cells. 92 Pias2 was identified as the SUMO E3 ligase to promote SOX2 SUMOylation, which inhibits SOX2 transcriptional activity in ESCs. 90 The SUMOylated SOX2 has impaired property in its DNA binding and is readily subjected to autophagic degradation, thus SOX2 SUMOylation results in reduced stability and activity of SOX2, thereby reducing the stemness of NPC.…”

“…Currently, the approaches undertaken to target SOX2 includes: (1) to alter the endogenous SOX2 gene expression by direct gene targeting using the zinc-finger (ZF)-based artificial transcription factor (ATF); (2) to generate the peptide aptamer for SOX2 targeting; (3) to target the SOX2-DNA binding, thus inhibiting SOX2 transcriptional activity; (4) to target SOX2 via inducing immune responses; (4) to use small molecules inhibitors of signaling pathways that affect SOX2, thus indirectly inhibiting SOX2; (6) to target protein ubiquitylation and degradation to specifically shut down SOX2 expression; (7) to develop the PROteolysis Targeting Chimera (PROTAC) method to directly degrade SOX2 autophagic degradation in NPC. 92 An XIAP inhibitor (APG-1387) has an antitumor effect on CSCs expressing high level of SOX2 with a synergistic effect in combination with CDDP/5-FU. 92 In LSCCs, the inhibitor of histone demethylase LSD1 (CBB1007) suppresses growth of SOX2-expressing cancer cells, since LSD1 likely serves as an epigenetic target in SOX2-expressing cancers.…”

Genetic approaches in mice demonstrate that neuro-mesodermal progenitors express T/Brachyury but not Sox2

“…On the other hand, T is expressed not only in the pre-somitic mesoderm and CNH, but also in the forming neural tube at both E8.5 (Figure 5a1 with lower intensity, in correspondence with previous reports 9,11,20 . Presumably, high Sox2 protein stability, which is estimated at > 48 h [28][29][30][31][32] , is responsible for this residual Sox2 immunostaining, that likely stems from gene expression at the epiblast stage.…”