β-Tubulin Isotype, TUBB4B, Regulates The Maintenance of Cancer Stem Cells

Recent advancements in cancer research have shown that cancer stem cell (CSC) niche is a crucial factor modulating tumor progression and treatment outcomes. It sustains CSCs by orchestrated regulation of several cytokines, growth factors, and signaling pathways. Although the features defining adult stem cell niches are well-explored, the CSC niche is poorly characterized. Since membrane trafficking proteins have been shown to be essential for the localization of critical proteins supporting CSCs, we investigated the role of TUBB4B, a probable membrane trafficking protein that was found to be overexpressed in the membranes of stem cell enriched cultures, in sustaining CSCs in oral cancer. Here, we show that the knockdown of TUBB4B downregulates the expression of pluripotency markers, depletes ALDH1A1+ population, decreases in vitro sphere formation, and diminishes the tumor initiation potential in vivo. As TUBB4B is not known to have any role in transcriptional regulation nor cell signaling, we suspected that its membrane trafficking function plays a role in constituting a CSC niche. The pattern of its expression in tissue sections, forming a gradient in and around the CSCs, reinforced the notion. Later, we explored its possible cooperation with a signaling protein, Ephrin-B1, the abrogation of which reduces the self-renewal of oral cancer stem cells. Expression and survival analyses based on the TCGA dataset of head and neck squamous cell carcinoma (HNSCC) samples indicated that the functional cooperation of TUBB4 and EFNB1 results in a poor prognosis. We also show that TUBB4B and Ephrin-B1 cohabit in the CSC niche. Moreover, depletion of TUBB4B downregulates the membrane expression of Ephrin-B1 and reduces the CSC population. Our results imply that the dynamics of TUBB4B is decisive for the surface localization of proteins, like Ephrin-B1, that sustain CSCs by their concerted signaling.

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LncRNA ASAP1-IT1 enhances cancer cell stemness via regulating miR-509-3p/YAP1 axis in NSCLC

Cancer Cell International ◽

10.1186/s12935-021-02270-7 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Yantao Liu ◽

Yuping Yang ◽

Lingli Zhang ◽

Jiaqiang Lin ◽

Bin Li ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Growth ◽

Cancer Stem Cells ◽

Tumor Growth ◽

Cancer Cell ◽

Qrt Pcr

Abstract Background Non-small cell lung cancer (NSCLC) is a major cause of cancer-related death worldwide, and cancer stem cell is responsible for the poor clinical outcome of NSCLC. Previous reports indicated that long noncoding RNAs (lncRNAs) play important roles in maintaining cancer stemness, however, the underlying mechanisms remain unclear. This study investigates the role of ASAP1 Intronic Transcript 1 (ASAP1-IT1) in cancer cell stemness of NSCLC. Methods The expression of ASAP1-IT1, microRNA-509-3p (miR-509-3p) and apoptosis-/stemness-related genes was analyzed by qRT-PCR in NSCLC tissues, cancer cells and spheres of cancer stem cells. Knockdown of ASAP1-IT1 or overexpression of miR-509-3p in NSCLC cells by infection or transfection of respective plasmids. Sphere formation and colony formation were used to detect NSCLC stem cell-like properties and tumor growth in vitro. Luciferase reporter assays, RNA immunoprecitation (RIP) and qRT-PCR assays were used to analyze the interaction between lncRNA and miRNA. The expression of expression of regulated genes of ASAP1-IT1/miR-509-3p axis was evaluated by qRT-PCR and Western blot. The NSCLC xenograft mouse model was used to validate the role of ASAP1-IT1 in NSCLC stemness and tumor growth in vivo. Results ASAP1-IT1 was up-regulated in NSCLC tissues, cancer cells, and in spheres of A549-derived cancer stem cells. Downregulation of ASAP1-IT1 or overexpression of miR-509-3p significantly decreased cell colony formation and stem cell-like properties of A549-dereived stem cells with decreased expression of stem cell biomarkers SOX2, CD34, and CD133, and suppressing the expression of cell growth-related genes, Cyclin A1, Cyclin B1, and PCNA. Furthermore, knockdown of ASAP1-IT1 or overexpression of miR-509-3p repressed tumor growth in nude mice via reducing expression of tumorigenic genes. ASAP1-IT1 was found to interact with miR-509-3p. Moreover, overexpression of ASAP1-IT1 blocked the inhibition by miR-509-3p on stem cell-like properties and cell growth of A549-dereived stem cells both in vitro and in vivo. Finally, the level of YAP1 was regulated by ASAP1-IT1 and miR-509-3p. Conclusions YAP1-involved ASAP1-IT1/miR-509-3p axis promoted NSCLC progression by regulating cancer cell stemness, and targeting this signaling pathway could be is a promising therapeutic strategy to overcome NSCLC stemness.

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pERK-mediated IL8 secretion can enhance the migration, invasion, and cisplatin resistance of CD10-positive oral cancer cells

BMC Cancer ◽

10.1186/s12885-021-09025-7 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Yinfei Pu ◽

Qingxiang Li ◽

Yifei Wang ◽

Le Xu ◽

Qiao Qiao ◽

...

Keyword(s):

Stem Cells ◽

Oral Cancer ◽

Cancer Stem Cells ◽

Signaling Pathway ◽

Cisplatin Resistance ◽

Erk Signaling ◽

Spheroid Formation ◽

Oral Cancer Cells

Abstract Background Cancer stem cells (CSCs) drive tumor initiation and progression and participate in tumor chemoresistance. We recently discovered that oral squamous cell carcinoma (OSCC) cells that highly express CD10 (CD10H cells) present cancer stem cells (CSC)-associated characteristics, which, in turn, affect the tumor growth, epithelial-mesenchymal transition (EMT), and resistance to cisplatin. In this study, we further investigated this mechanism in vitro and in vivo. We hypothesized that IL8 might regulate migration, invasion, and cisplatin resistance of CD10-positive oral cancer cells through the ERK pathway. Methods CD10 MicroBead Kit was used to select HN6 cells with high and low expression of CD10. The target protein IL8 was screened via protein chip assay. Lentiviral transduction and specific inhibitor were applied to investigate the signaling pathway. Real-time PCR, Western blot, and immunohistochemistry were used to analyze the mRNA and protein expression; transwell assay, spheroid formation assay, and cell viability assay were used to study the cell biological behavior in vitro; xenograft animal model was used to evaluate the tumor formation rate in vivo. Results Overexpression of CD10 promoted CSC-related genes expression and enhanced migration, invasion, spheroid formation, and chemoresistance in HN6 cells. Moreover, the overexpression of IL8 was detected in OSCC tumor tissue and cell lines (HN6 and CAL27) overexpressing CD10. IL8 secreted by CD10H HN6 promoted migration and invasion and restored tumor chemosensitivity via the p-ERK signaling pathway, while the inhibition of IL8 increased the chemosensitivity to cisplatin. Conclusions IL8 secretion by CD10 positive cells promotes migration, invasion, and cisplatin resistance of OSCC via the p-ERK signaling pathway.

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High Frequency of Tumor Propagating Cells in Fusion-Positive Rhabdomyosarcoma

Genes ◽

10.3390/genes12091373 ◽

2021 ◽

Vol 12 (9) ◽

pp. 1373

Author(s):

Melanie Generali ◽

Sampoorna Satheesha ◽

Peter K. Bode ◽

Debora Wanner ◽

Beat W. Schäfer ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cancer Stem Cells ◽

High Frequency ◽

Cell Clone ◽

Hierarchical Organization ◽

Stem Cell Markers ◽

Worse Prognosis

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children. Fusion-positive RMS (FPRMS), expressing the PAX3/7-FOXO1, has a worse prognosis compared to the more common fusion-negative RMS (FNRMS). Although several studies reported hierarchical organization for FNRMS with the identification of cancer stem cells, the cellular organization of FPRMS is not yet clear. In this study we investigated the expression of key stem cell markers, developed a sphere assay, and investigated the seven most common FPRMS cell lines for subpopulations of tumor propagating cancer stem-like cells, also called cancer stem cells (CSCs). Moreover, loss- and gain-of-functions of the stem cell genes SOX2, OCT4, and NANOG were investigated in the same cells. Single-cell clonal analysis was performed in vitro as well as in vivo. We found that no stable CSC subpopulation could be enriched in FPRMS. Unlike depletion of PAX3-FOXO1, neither overexpression nor siRNA-mediated downregulation of SOX2, OCT4, and NANOG affected physiology of RMS cells. Every single subclone-derived cell clone initiated tumor growth in mice, despite displaying considerable heterogeneity in gene expression. FPRMS appears to contain a high frequency of tumor propagating stem-like cells, which could explain their higher propensity for metastasis and relapse. Their dependency on PAX3-FOXO1 activity reinforces the importance of the fusion protein as the key therapeutic target.

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LncRNA ASAP1-IT1 Enhances Cancer Cell Stemness via Regulating miR-509-3p/YAP1 Axis in NSCLC

10.21203/rs.3.rs-445483/v2 ◽

2021 ◽

Author(s):

Yantao Liu ◽

Yuping Yang ◽

Lingli Zhang ◽

Jiaqiang Lin ◽

Bin Li ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Growth ◽

Cancer Stem Cells ◽

Tumor Growth ◽

Cancer Cells ◽

Cancer Cell ◽

Qrt Pcr

Abstract Background: Non-small cell lung cancer (NSCLC) is a major cause of cancer-related death worldwide, and cancer stem cell is mainly responsible for the poor clinical outcome of NSCLC. Although previous reports indicated long noncoding RNAs (lncRNAs) play important roles in cancer stemness maintain, the underlying causes mechanisms remain is still mysterious. Our study aims to investigate the role of ASAP1 Intronic Transcript 1 (ASAP1-IT1) in cancer cell stemness of NSCLC. Methods: qRT-PCR analysis the expression of ASAP1-IT1 and microRNA-509-3p (miR-509-3p) in NSCLC tissues, cancer cells and spheres of cancer cells-derived cancer stem cells. Knockdown of ASAP1-IT1 or overexpression of miR-509-3p in NSCLC cells by infection or transfection. Sphere formation and colony formation detects NSCLC stem cell-like properties and tumor growth in vitro. Luciferase reporter assays, RNA immunoprecitation (RIP) and qRT-PCR assays analyzed the interaction between lncRNA and miRNA, qRT-PCR and Western blot detect ASAP1-IT1/miR-509-3p axis's regulation on the expression of regulated genes. NSCLC xenograft mice model validates ASAP1-IT1 role in NSCLC stemness and tumor growth in vivo. Results: ASAP1-IT1 was up-regulated in NSCLC tissues, cancer cells, and in spheres of A549-derived cancer stem cells. Downregulated ASAP1-IT1 or miR-509-3p significantly decreased cell colony formation and stem cell-like properties of A549-dereived stem cells with decreased expression of stem cell biomarkers SOX2, CD34, and CD133, and suppressing the expression of cell growth-related genes, Cyclin A1, Cyclin B1, and PCNA. Furthermore, knockdown of ASAP1-IT1 or overexpression of miR-509-3p repressed tumor growth in nude mice via reducing expression of tumorigenic genes. ASAP1-IT1 was found to interact with miR-509-3p. Moreover, overexpression of ASAP1-IT1 blocked miR-509-3p mediated regulation of stem cell-like properties, cell growth, and cell apoptosis of A549-dereived stem cells both in vitro and in vivo. Finally, the level of YAP1 was regulated by ASAP1-IT1 and miR-509-3p.Conclusions: YAP1-involved ASAP1-IT1/miR-509-3p axis promoted NSCLC progression by regulating cancer cell stemness，and targeting related signaling is a promising therapeutic strategy to overcome NSCLC stemness.

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LncRNA ASAP1-IT1 Enhances Cancer Cell Stemness via Regulating miR-509-3p/YAP1 Axis in NSCLC

10.21203/rs.3.rs-445483/v1 ◽

2021 ◽

Author(s):

Yantao Liu ◽

Yuping Yang ◽

Lingli Zhang ◽

Jiaqiang Lin ◽

Bin Li ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Growth ◽

Cancer Stem Cells ◽

Tumor Growth ◽

Cancer Cells ◽

Cancer Cell ◽

Qrt Pcr

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STEM-26. SMALL MOLECULE DRUG CBL0137 INHIBITS THE FACT COMPLEX AND SENSITIZES GLIOBLASTOMA CANCER STEM CELLS TO RADIOTHERAPY

Neuro-Oncology ◽

10.1093/neuonc/noaa215.843 ◽

2020 ◽

Vol 22 (Supplement_2) ◽

pp. ii201-ii202

Author(s):

Miranda Tallman ◽

Abigail Zalenski ◽

Amanda Deighen ◽

Morgan Schrock ◽

Sherry Mortach ◽

...

Keyword(s):

Stem Cells ◽

Cancer Stem Cells ◽

High Rate ◽

Orthotopic Model ◽

In Vivo Models ◽

Specific Cytotoxicity ◽

Treatment Paradigm ◽

Cancer Agent

Abstract Glioblastoma (GBM) is a malignant brain tumor with nearly universal recurrence. GBM cancer stem cells (CSCs), a subpopulation of radio- and chemo-resistant cancer cells capable of self-renewal, contribute to the high rate of recurrence. The anti-cancer agent, CBL0137, inhibits the FACT (facilitates chromatin transcription) complex leading to cancer cell specific cytotoxicity. Here, we show that CBL0137 sensitized GBM CSCs to radiotherapy using both in vitro and in vivo models. Treatment of CBL0137 combined with radiotherapy led to increased DNA damage in GBM patient specimens and failure to resolve the damage led to decreased cell viability. Using clonogenic assays, we confirmed that CBL0137 radiosensitized the CSCs. To validate that combination therapy impacted CSCs, we used an in vivo subcutaneous model and showed a decrease in the frequency of cancer stem cells present in tumors as well as decreased tumor volume. Using an orthotopic model of GBM, we confirmed that treatment with CBL0137 followed by radiotherapy led to significantly increased survival compared to either treatment alone. Radiotherapy remains a critical component of patient care for GBM, even though there exists a resistant subpopulation. Radio-sensitizing agents, including CBL0137, pose an exciting treatment paradigm to increase the efficacy of irradiation, especially by inclusively targeting CSCs.

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Regulation of sarcomagenesis by the empty spiracles homeobox genes EMX1 and EMX2

Cell Death and Disease ◽

10.1038/s41419-021-03801-w ◽

2021 ◽

Vol 12 (6) ◽

Author(s):

Manuel Pedro Jimenez-García ◽

Antonio Lucena-Cacace ◽

Daniel Otero-Albiol ◽

Amancio Carnero

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Transcription Factors ◽

Neural Crest ◽

Tumor Suppressors ◽

Homeobox Genes ◽

Neuronal Cell ◽

Cell Gene Expression

AbstractThe EMX (Empty Spiracles Homeobox) genes EMX1 and EMX2 are two homeodomain gene members of the EMX family of transcription factors involved in the regulation of various biological processes, such as cell proliferation, migration, and differentiation, during brain development and neural crest migration. They play a role in the specification of positional identity, the proliferation of neural stem cells, and the differentiation of certain neuronal cell phenotypes. In general, they act as transcription factors in early embryogenesis and neuroembryogenesis from metazoans to higher vertebrates. The EMX1 and EMX2’s potential as tumor suppressor genes has been suggested in some cancers. Our work showed that EMX1/EMX2 act as tumor suppressors in sarcomas by repressing the activity of stem cell regulatory genes (OCT4, SOX2, KLF4, MYC, NANOG, NES, and PROM1). EMX protein downregulation, therefore, induced the malignance and stemness of cells both in vitro and in vivo. In murine knockout (KO) models lacking Emx genes, 3MC-induced sarcomas were more aggressive and infiltrative, had a greater capacity for tumor self-renewal, and had higher stem cell gene expression and nestin expression than those in wild-type models. These results showing that EMX genes acted as stemness regulators were reproduced in different subtypes of sarcoma. Therefore, it is possible that the EMX genes could have a generalized behavior regulating proliferation of neural crest-derived progenitors. Together, these results indicate that the EMX1 and EMX2 genes negatively regulate these tumor-altering populations or cancer stem cells, acting as tumor suppressors in sarcoma.

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“Empowering” Cardiac Cells via Stem Cell Derived Mitochondrial Transplantation- Does Age Matter?

International Journal of Molecular Sciences ◽

10.3390/ijms22041824 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1824

Author(s):

Matthias Mietsch ◽

Rabea Hinkel

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Treatment Strategies ◽

Cardiac Cells ◽

Aging Effects ◽

Beneficial Effects ◽

Micro Rnas ◽

New Treatment

With cardiovascular diseases affecting millions of patients, new treatment strategies are urgently needed. The use of stem cell based approaches has been investigated during the last decades and promising effects have been achieved. However, the beneficial effect of stem cells has been found to being partly due to paracrine functions by alterations of their microenvironment and so an interesting field of research, the “stem- less” approaches has emerged over the last years using or altering the microenvironment, for example, via deletion of senescent cells, application of micro RNAs or by modifying the cellular energy metabolism via targeting mitochondria. Using autologous muscle-derived mitochondria for transplantations into the affected tissues has resulted in promising reports of improvements of cardiac functions in vitro and in vivo. However, since the targeted treatment group represents mainly elderly or otherwise sick patients, it is unclear whether and to what extent autologous mitochondria would exert their beneficial effects in these cases. Stem cells might represent better sources for mitochondria and could enhance the effect of mitochondrial transplantations. Therefore in this review we aim to provide an overview on aging effects of stem cells and mitochondria which might be important for mitochondrial transplantation and to give an overview on the current state in this field together with considerations worthwhile for further investigations.

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STEM-20. RADIO-SENSITIZING GLIOBLASTOMA CANCER STEM CELLS BY INHIBITION OF FACT

Neuro-Oncology ◽

10.1093/neuonc/noz175.993 ◽

2019 ◽

Vol 21 (Supplement_6) ◽

pp. vi237-vi238

Author(s):

Miranda Montgomery ◽

Abigail Zalenski ◽

Amanda Deighen ◽

Sherry Mortach ◽

Treg Grubb ◽

...

Keyword(s):

Stem Cells ◽

Dna Damage ◽

Cancer Stem Cells ◽

Combination Therapy ◽

High Rate ◽

Primary Role ◽

Cancer Cell Growth ◽

Treatment Paradigm

Abstract Glioblastoma (GBM) has a particularly high rate of recurrence with a 5-year overall survival rate of approximately 5%. This is in part due to a sub-population of cancer stem cells (CSC), which are both radioresistant and chemotherapeutically resistant to conventional treatments. Here we investigated CBL0137, a small molecule form of curaxin, in combination with radiotherapy as a means to radiosensitize CSCs. CBL0137 sequesters FACT (facilitates chromatin transcription) complex to chromatin, which leads to activation of p53 and inhibition of NF-κB. This sequestering of FACT results in cytotoxicity especially within tumor cells and prevents FACT from performing its primary role as a histone chaperone, as well as inhibits its part in the DNA damage response pathway. We show that when combined with radiotherapy, CBL0137 administration limited the ability of CSCs to identify and repair damaged DNA. CSCs treated in vitro with CBL0137 and irradiation showed an increased inhibition of cancer cell growth and decreased viability compared to irradiation or drug alone. Combination therapy also showed more DNA damage in the CSCs than with either agent alone. Based on our in vitro evidence for the efficacy of combination therapy to target CSCs, we moved forward to test the treatment in vivo. Using a subcutaneous model, we show that the amount of CD133+ cells (a marker for GMB CSCs) was reduced in irradiation plus CBL0137 compared to either treatment alone. Survival studies demonstrated that irradiation plus CBL0137 compared to irradiation alone or CBL0137 alone increase lifespan. Here we show the ability of CBL0137, in combination with irradiation, to target patient GBM CSCs both in vitro and in vivo. This work establishes a new treatment paradigm for GBM that inclusively targets CSCs and may ultimately reduce tumor recurrence.

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