Short peptide domains of the Wnt inhibitor sFRP4 target ovarian cancer stem cells by neutralizing the Wnt β-catenin pathway, disrupting the interaction between β-catenin and CD24 and suppressing autophagy

Aims: One of the hallmarks of cancer stem cells (CSC) is hyperactive Wnt beta-catenin signaling due to the decreased presence of Wnt antagonists such as secreted frizzled related protein 4 (SFRP4). Cysteine-rich domain (CRD) and netrin-like domain (NLD) are the two functional domains of SFRP4 having anti-tumor properties. In this study, we have explored the effectiveness of short micropeptides SC-301 (from CRD) and SC-401 (from NLD) on CSC properties, EMT, apoptosis and autophagy in ovarian CSCs enriched from PA-1 and SKOV-3 cell lines. Main methods: Gene expression analysis, Western blot and immunocytochemistry were performed on ovarian CSCs to evaluate the inhibitory potential of micropeptides to various CSC associated oncogenic properties. Co-immunoprecipitation was performed to detect the binding of CD24 to beta-catenin protein complex. CYTO-ID Autophagy Detection Kit 2.0 was used to monitor autophagic flux in peptide treated CSCs. Key findings: It is clearly seen that the micropeptides derived from both the domains inhibit Wnt pathway, initiate apoptosis, inhibit migration and chemosensitize CSCs. Specifically, CD24, a defining marker of ovarian CSC was suppressed by peptide treatment. Notably, interaction between CD24 and beta-catenin was disrupted upon peptide treatment. SFRP4 peptide treatment also suppressed the autophagic process which is crucial for CSC survival. Significance: The study demonstrated that although both peptides have inhibitory effects, SC-401 was emphati-cally more effective in targeting CSC properties and down regulating the Wnt beta-catenin machinery.