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Abstract Background Medulloblastoma (MB) is the most common malignant pediatric brain tumor that originates in the cerebellum and brainstem. Frequent somatic mutations and deregulated expression of epigenetic regulators in MB highlight the substantial role of epigenetic alterations. 5-hydroxymethylcytosine (5hmC) is a highly abundant cytosine modification in the developing cerebellum and is regulated by ten-eleven translocation (TET) enzymes. Results We investigate the alterations of 5hmC and TET enzymes in MB and their significance to cerebellar cancer formation. We show total abundance of 5hmC is reduced in MB, but identify significant enrichment of MB-specific 5hmC marks at regulatory regions of genes implicated in stem-like properties and Nanog-binding motifs. While TET1 and TET2 levels are high in MBs, only knockout of Tet1 in the smoothened (SmoA1) mouse model attenuates uncontrolled proliferation, leading to a favorable prognosis. The pharmacological Tet1 inhibition reduces cell viability and platelet-derived growth factor signaling pathway-associated genes. Conclusions These results together suggest a potential key role of 5hmC and indicate an oncogenic nature for TET1 in MB tumorigenesis, suggesting it as a potential therapeutic target for MBs.

Cancer stem cells (CSCs) have been identified in a multiple of cancer types and resistant to traditional cancer therapies such as chemotherapeutic agents and radiotherapy, which may destroy bulk tumor cells but not all CSCs, contributing to reformation tumor masses and subsequent relapse. Moreover, it is very difficult to effectively identify and eliminate CSCs because they share some common phenotypic and functional characteristics of normal stem cells. Therefore, finding better therapeutic strategies to selectively target CSCs might be helpful to reduce subsequent malignancies. In the present study, we found that caffeic acid effectively suppresses self-renewal capacity, stem-like characteristics, and migratory capacity of CD44+ and CD133+ colorectal CSCs in vitro and in vivo. In addition, we also revealed that PI3K/Akt signaling may be linked to multiple colorectal CSC-associated characteristics, such as radio-resistance, stem-like property, and tumorigenic potential. To the best of our knowledge, this is the first study demonstrating that caffeic acid effectively targets colorectal CSC populations by inhibiting the growth and/or self-renewal capacity of colorectal CSCs through PI3K/Akt signaling in vitro and in vivo.

Background: Nasopharyngeal carcinoma (NPC) is a malignant tumor that occurs in the nasopharyngeal mucosa. Clinically, radiotherapy is the preferred treatment for NPC, and cervical lymph node metastasis is easy to emerge in the early stage. Therefore, this study aimed to investigate the role and potential molecular mechanisms of miR-96-5p in NPC cells to develop new therapeutic horizons. Methods: The expression of miR-96-5p and CDK1 was measured by RT-qPCR or Western blot. The target relationship between miR-96-5p and CDK1 was confirmed by luciferase reporter assay. CCK-8, sphere formation, flow cytometry and colony formation assay were employed to examine cell viability, stem-like property, apoptosis and cycle, respectively. Male BALB/c nude mice model (6– 8 weeks, weigh 18– 20 g) was used to evaluate the effect of miR-96-5p on tumor growth in vivo. Results: miR-96-5p was lowly expressed and CDK1 was highly expressed in NPC tissues and cell lines. CDK1 was identified as a direct target of miR-96-5p, and its expression was negatively regulated by miR-96-5p. By targeting CDK1, miR-96-5p overexpression significantly inhibited tumor sphere formation, promoted apoptosis and cell cycle arrest in CNE-2Z cells. Importantly, CCK-8 and colony formation assay demonstrated that elevated miR-96-5p enhanced the radiotherapy and chemotherapy sensitivity of CNE-2Z cells. Animal experiments showed that the overexpression of miR-96-5p reduced tumor weight and size in tumor-bearing mice and inhibited the expression of stem-like marker proteins and apoptosis-related proteins. Conclusion: These results, together, suggested that miR-96-5p induced cell cycle arrest and apoptosis, inhibited stem-like property, and enhanced the radiochemical sensitivity of NPC by targeting CDK1. In short, miR-96-5p may be a diagnostic and therapeutic target for NPC. [ABSTRACT FROM AUTHOR]

Abstract Background Ring finger protein 43 (RNF43) is a member of the transmembrane E3 ubiquitin ligase family that was originally found in stem cells and plays important roles in tumor formation and progression. Our previous study indicated that RNF43 might be a tumor suppressor protein in gastric cancer. Given its antagonistic relationship with leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5), one of the gastric cancer stem cell markers, investigation of the potential role of RNF43 in gastric stem cancer cells is necessary. Methods Immunohistochemistry staining, western blot analysis, and quantitative reverse transcription polymerase chain reaction were used to determine the mRNA and protein expression level of RNF43 and other Wnt pathway factors. Gastric cancer stem-like cells were obtained from gastric cancer tumor and cell lines by tumorsphere culture. The adeno-associated virus system was used to upregulate RNF43 expression in cancer cells. Functional experiments including tumorsphere formation, chemotherapy resistance, surface marker detection, and tumor xenograft assay were performed to measure stem-like properties in gastric cancer stem-like cells after RNF43 overexpression. Results RNF43 loss was significantly associated with TNM stage, distant metastasis, and Lauren classification, and predicted worse prognosis in gastric cancer patients. RNF43 expression was even lower in tumorspheres derived from tumor tissues or cell lines compared with adherent cancer cells and normal gastric cells. Overexpression of RNF43 in gastric cancer cells impaired their stem-like properties, including sphere formation ability, chemoresistance in vitro, and tumorigenicity in vivo. Moreover, Wnt pathway-related proteins were decreased in RNF43-overexpressing cells, while Wnt pathway activators could reverse the trend to some extent. Conclusions Our findings indicated that RNF43 might not only participate in gastric cancer progression, but also attenuate the stemness of gastric cancer stem-like cells through the Wnt/β-catenin pathway.

The refractory pulmonary adenocarcinoma is characterized by its metastasis and resistance to cytotoxic agents. While the underlying molecular mechanism is unclear, the property of chemoresistance may mainly lie in the presence of highly resistant cancer stem cells. We examined the function of Wnt/β-catenin signaling in maintaining cancer stem cells (CSCs) in lung adenocarcinoma. Lentivirus-mediated knockdown of β-catenin expression accelerated cell cycle. Subsequently, β-catenin knockdown PC9 cells improve the sensitivity to chemotherapy. Further focusing on Wnt signal by administrating PP and EGFR-TKIs as Wnt antagonists can decrease metastasis and induce apoptosis. Collectively, these results indicate that Wnt signaling pathway plays an essential role in maintaining highly resistant CSCs, regulation of cell cycle, metastasis and apoptosis in lung adenocarcinoma. [ABSTRACT FROM AUTHOR]

Traditionally, it was presumed that gastric cancer was derived from tumor cells with stem-like properties. In the present study, stem-like cells from the gastric cancer cell lines MKN45 and SGC7901 were enriched by growing them as spheres in a defined serum-free medium. Following enrichment for stem-like cells, cluster of differentiation (CD)24 and CD44 were applied as candidate stem cell markers to examine the expression profile. It was revealed that the sphere-derived cells contained a higher proportion of cells expressing the stem cell surface markers CD24 and CD44 when compared with the parental cells. It was also identified that the expression of cytokeratin 18 in sphere-derived cells was decreased and the expression of vimentin and aldehyde dehydrogenase 1 (ALDH1) was increased compared with the parental cells. This finding supports the existence of a population of tumor sphere-forming cells with stem cell properties in the MKN45 and SGC7901 cell lines. Furthermore, the stem cell population was enriched in cells expressing CD24, CD44, vimentin and ALDH1 cell surface markers. These results support the existence of gastric cancer stem cells and provide an alternative approach to the diagnosis and treatment of gastric cancer. [ABSTRACT FROM AUTHOR]

Cancer stem cells (CSCs) have been identified in a multiple of cancer types and resistant to traditional cancer therapies such as chemotherapeutic agents and radiotherapy, which may destroy bulk tumor cells but not all CSCs, contributing to reformation tumor masses and subsequent relapse. Moreover, it is very difficult to effectively identify and eliminate CSCs because they share some common phenotypic and functional characteristics of normal stem cells. Therefore, finding better therapeutic strategies to selectively target CSCs might be helpful to reduce subsequent malignancies. In the present study, we found that caffeic acid effectively suppresses self-renewal capacity, stem-like characteristics, and migratory capacity of CD44 + and CD133 + colorectal CSCs in vitro and in vivo . In addition, we also revealed that PI3K/Akt signaling may be linked to multiple colorectal CSC-associated characteristics, such as radio-resistance, stem-like property, and tumorigenic potential. To the best of our knowledge, this is the first study demonstrating that caffeic acid effectively targets colorectal CSC populations by inhibiting the growth and/or self-renewal capacity of colorectal CSCs through PI3K/Akt signaling in vitro and in vivo ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Park, Kim, Hong and Lee.)

Background: Taxifolin is a natural flavonoid with anti-oxidant and anti-proliferative properties. In this study, we investigated the stemness-related inhibitory effects of taxifolin in two lung cancer cell lines, A549 and H1975, as well as in A549 xenografts., Methods: A549 and H1975 cells, as well as A549 xenograft BALB/c mice were treated with taxifolin. Cell viability, stemness, mobility and protein expression were tested with Cell counting kit-8 (CCK-8), Colony formation assay, Flow cytometry, Transwell, Western blot and Immunohistochemistry, respectively., Results: CCK-8 exhibited an obvious toxicity of taxifolin to both cell lines at higher dose. Then taxifolin of 0, 25, 50, and 100 µM/L were subsequently used. Taxifolin exhibited inhibitory effects on stemness and sphere formation, reduced protein expression of SOX2 and OCT4, and reduced CD133-positive cells. Furthermore, taxifolin decreased invasive cells, reduced N-cadherin and vimentin while increased E-cadherin expression, indicating that epithelial-mesenchymal transition (EMT) was inhibited. All of the effects observed were exhibited in a dose-dependent manner, and A549 cells proved to be more sensitive to taxifolin than H1975 cells. Taxifolin inactivated PI3K and TCF4 protein phosphorylation; however, taxifolin was not observed to have an effect on NF-κB P65 or STAT3. Taxifolin also suppressed tumor growth in A549 xenograft BALB/c mice, with decreased SOX2 and OCT4 expression and inhibited PI3K and TCF4., Conclusions: In summary, taxifolin inhibited stemness and EMT in lung cancer cells possibly via the inactivation of PI3K and OCT4. Taxifolin could be a potential prodrug or serve as an adjuvant in lung cancer treatment., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/atm-20-3329). The authors have no conflicts of interest to declare., (2020 Annals of Translational Medicine. All rights reserved.)

Background: Gastric cancer (GC) is the second most significant contributor to cancer-related mortality in China. GC treatment is often hindered by metastasis and chemoresistance, which leads to poor prognosis. This study set out to investigate the role of miR-567 on the stem-like properties and chemo-resistance of GC cells, as well as its potential molecular mechanism., Methods: The expression of miR-567 in GES-1, AGS, SCG-7901, MGC-803, SUN-16, and MKN1 cell lines was detected by Real-time PCR. AGS cells were transfected with NC or miR-567 mimics and RPL15 3'UTR (wt) or RPL15 3'UTR (mut) using Lipofectamine 2000. CCK-8, 5-ethynyl-2'-deoxyuridine (EdU) assay were detected the effect of miR-567/RPL15/TGF-β/Smad on gastric cancer cell viability and proliferation, respectively. Western blot were used to analyze the effects of miR-567/RPL15/TGF-β/Smad on protein levels of SOX2, NANOG, ALDH1A1, TGF-β1, TGFβ-R1, SMAD1, P-SMAD1, SMAD2 and P-SMAD2., Results: The results showed that the expression of miR-567 was down-regulated in GC cell lines. TargetScan and luciferase report assay indicated that RPL15 was the target of miR-567. Functional analysis discovered that the overexpression of miR-567 inhibited the microsphere formation of AGS stem cells, while PRL15 overexpression promoted the formation of microspheres in AGS cells. Through Western blot analysis, miR-567 overexpression was further revealed to inhibit the expression of stem-like marker proteins (SOX2, NANOG, and ALDH1A1). Furthermore, PRL15 overexpression was found to significantly promote the growth of AGS/DDP cells, while miR-567 overexpression reversed the effect of PRL15 on cisplatin-resistant cell growth. The relationship between miR-567 and the TGF-β1/TGFβ-R1/Smad2/Smad3 pathway was also shown. The addition of TGF-β/Smad pathway inhibitor LY 3200882 significantly inhibited the expression of PRL-15, TGF-β1, TGF-R1, p-Smad1, and p-sSmad2, while reversing the effect of miR-567 inhibitor on stem-like properties and chemical resistance., Conclusions: Finally, this study elucidated the effect of miR-567 on the stem-like properties and chemical resistance of GC cells via the RPL15 /TGF-beta/Smad axis., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/tcr.2020.04.13). The authors have no conflicts of interest to declare., (2020 Translational Cancer Research. All rights reserved.)

Programmed cell death protein 1 (PD-1) and its ligand PD-L1 blockade have been identified to target immune checkpoints to treat human cancers with durable clinical benefit. Several studies reveal that the response to PD-1-PD-L1 blockade might correlate with PD-L1 expression levels in tumor cells. However, the mechanistic pathways that regulate PD-L1 protein expression are not understood. Here, we reported that PD-L1 protein is regulated by ATG7-autophagy with an ATG7-initiated positive feedback loop in bladder cancer (BC). Mechanistic studies revealed that ATG7 overexpression elevates PD-L1 protein level mainly through promoting autophagy-mediated degradation of FOXO3a, thereby inhibiting its initiated miR-145 transcription. The lower expression of miR-145 increases pd-l1 mRNA stability due to the reduction of its direct binding to 3′-UTR of pd-l1 mRNA, in turn leading to increasing in pd-l1 mRNA stability and expression, and finally enhancing stem-like property and invasion of BC cells. Notably, overexpression of PD-L1 in ATG7 knockdown cells can reverse the defect of autophagy activation, FOXO3A degradation, and miR-145 transcription attenuation. Collectively, our results revealed a positive feedback loop to promoting PD-L1 expression in human BC cells. Our study uncovers a novel molecular mechanism for regulating pd-l1 mRNA stability and expression via ATG7/autophagy/FOXO3A/miR-145 axis and reveals the potential for using combination treatment with autophagy inhibitors and PD-1/PD-L1 immune checkpoint blockade to enhance therapeutic efficacy for human BCs. [ABSTRACT FROM AUTHOR]

Background: Ring finger protein 43 (RNF43) is a member of the transmembrane E3 ubiquitin ligase family that was originally found in stem cells and plays important roles in tumor formation and progression. Our previous study indicated that RNF43 might be a tumor suppressor protein in gastric cancer. Given its antagonistic relationship with leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5), one of the gastric cancer stem cell markers, investigation of the potential role of RNF43 in gastric stem cancer cells is necessary., Methods: Immunohistochemistry staining, western blot analysis, and quantitative reverse transcription polymerase chain reaction were used to determine the mRNA and protein expression level of RNF43 and other Wnt pathway factors. Gastric cancer stem-like cells were obtained from gastric cancer tumor and cell lines by tumorsphere culture. The adeno-associated virus system was used to upregulate RNF43 expression in cancer cells. Functional experiments including tumorsphere formation, chemotherapy resistance, surface marker detection, and tumor xenograft assay were performed to measure stem-like properties in gastric cancer stem-like cells after RNF43 overexpression., Results: RNF43 loss was significantly associated with TNM stage, distant metastasis, and Lauren classification, and predicted worse prognosis in gastric cancer patients. RNF43 expression was even lower in tumorspheres derived from tumor tissues or cell lines compared with adherent cancer cells and normal gastric cells. Overexpression of RNF43 in gastric cancer cells impaired their stem-like properties, including sphere formation ability, chemoresistance in vitro, and tumorigenicity in vivo. Moreover, Wnt pathway-related proteins were decreased in RNF43-overexpressing cells, while Wnt pathway activators could reverse the trend to some extent., Conclusions: Our findings indicated that RNF43 might not only participate in gastric cancer progression, but also attenuate the stemness of gastric cancer stem-like cells through the Wnt/β-catenin pathway.