ALDH-positive Cells Research Articles

Fractionated irradiation promotes radioresistance and decreases oxidative stress by increasing Nrf2 of ALDH-positive nasopharyngeal cancer stem cells

Radiotherapy is widely regarded as the primary therapeutic modality for nasopharyngeal cancer (NPC). Studies have shown that cancer cells with high resistance to radiation, known as radioresistant cancer cells, may cause residual illness, which in turn might contribute to the occurrence of cancer recurrence and metastasis. It has been shown that cancer stem-like cells (CSCs) exhibit resistance to radiation therapy. In the present study, fractionated doses of radiation-induced epithelial-mesenchymal transition (EMT) and ALDH+ CSCs phenotype of NPC tumor spheroids. Furthermore, it has been shown that cells with elevated ALDH activity have increased resistance to the effects of fractionated irradiation. Nuclear factor erythroid-2-related factor 2 (Nrf2) plays a pivotal role in regulating cellular antioxidant systems. A large body of evidence suggests that Nrf2 plays a significant role in the development of radioresistance in cancer. The authors’ research revealed that the application of fractionated irradiation resulted in a decline in Nrf2-dependent reactive oxygen species (ROS) levels, thereby mitigating DNA damage in ALDH+ stem-like NPC cells. In addition, immunofluorescence analysis revealed that subsequent to the process of fractionated irradiation of ALDH+ cells, activated Nrf2 was predominantly localized inside the nucleus. Immunofluorescent analysis also revealed that the presence of the nuclear Nrf2+/NQO1+/ALDH1+ axis might potentially serve as an indicator of poor prognosis and resistance to radiotherapy in patients with NPC. Thus, the authors’ findings strongly suggest that the radioresistance of ALDH-positive NPC CSCs to fractionated irradiation is regulated by nuclear Nrf2 accumulation. Nrf2 exerts its effects through the downstream effector NQO1/ALDH1, which depends on ROS attenuation.

Abstract A001: Silencing of EYA4 regulates stemness in H3K27M-mutant diffuse midline glioma

Abstract A001: Silencing of EYA4 regulates stemness in H3K27M-mutant diffuse midline glioma

Altered Phenotypes of Breast Epithelial × Breast Cancer Hybrids after ZEB1 Knock-Out.

ZEB1 plays a pivotal role in epithelial-to-mesenchymal transition (EMT), (cancer) cell stemness and cancer therapy resistance. The M13HS tumor hybrids, which were derived from spontaneous fusion events between the M13SV1-EGFP-Neo breast epithelial cells and HS578T-Hyg breast cancer cells, express ZEB1 and exhibit prospective cancer stem cell properties. To explore a possible correlation between the ZEB1 and stemness/ EMT-related properties in M13HS tumor hybrids, ZEB1 was knocked-out by CRISPR/Cas9. Colony formation, mammosphere formation, cell migration, invasion assays, flow cytometry and Western blot analyses were performed for the characterization of ZEB1 knock-out cells. The ZEB1 knock-out in M13HS tumor cells was not correlated with the down-regulation of the EMT-related markers N-CADHERIN (CDH2) and VIMENTIN and up-regulation of miR-200c-3p. Nonetheless, both the colony formation and mammosphere formation capacities of the M13HS ZEB1 knock-out cells were markedly reduced. Interestingly, the M13HS-2 ZEB1-KO cells harbored a markedly higher fraction of ALDH1-positive cells. The Transwell/ Boyden chamber migration assay data indicated a reduced migratory activity of the M13HS ZEB1-knock-out tumor hybrids, whereas in scratch/ wound-healing assays only the M13SH-8 ZEB1-knock-out cells possessed a reduced locomotory activity. Similarly, only the M13HS-8 ZEB1-knock-out tumor hybrids showed a reduced invasion capacity. Although the ZEB1 knock-out resulted in only moderate phenotypic changes, our data support the role of ZEB1 in EMT and stemness.

Histone H3 K27M-mediated regulation of cancer cell stemness and differentiation in diffuse midline glioma

Therapeutic resistance remains a major obstacle to preventing progression of H3K27M-altered Diffuse Midline Glioma (DMG). Resistance is driven in part by ALDH-positive cancer stem cells (CSC), with high ALDH1A3 expression observed in H3K27M-mutant DMG biopsies. We hypothesized that ALDH-mediated stemness and resistance may in part be driven by the oncohistone itself. Upon deletion of H3K27M, ALDH1A3 expression decreased dramatically and was accompanied by a gain in astrocytic marker expression and a loss of neurosphere forming potential, indicative of differentiation. Here we show that the oncohistone regulates histone acetylation through ALDH1A3 in a Wnt-dependent manner and that loss of H3K27M expression results in sensitization of DMGs to radiotherapy. The observed elevated Wnt signaling in H3K27M-altered DMG likely stems from a dramatic suppression of mRNA and protein expression of the Wnt inhibitor EYA4 driven by the oncohistone. Thus, our findings identify EYA4 as a bona fide tumor suppressor in DMG that upon suppression, results in aberrant Wnt signaling to orchestrate stemness and differentiation. Future studies will explore whether overexpression of EYA4 in DMG can impede growth and invasion. In summary, we have gained mechanistic insight into H3K27M-mediated regulation of cancer stemness and differentiation, which provides rationale for exploring new therapeutic targets for DMG.

ΔNp63/p73 drive metastatic colonization by controlling a regenerative epithelial stem cell program in quasi-mesenchymal cancer stem cells.

Cancer stem cells (CSCs) may serve as the cellular seeds of tumor recurrence and metastasis, and they can be generated via epithelial-mesenchymal transitions (EMTs). Isolating pure populations of CSCs is difficult because EMT programs generate multiple alternative cell states, and phenotypic plasticity permits frequent interconversions between these states. Here, we used cell-surface expression of integrin β4 (ITGB4) to isolate highly enriched populations of human breast CSCs, and we identified the gene regulatory network operating in ITGB4+ CSCs. Specifically, we identified ΔNp63 and p73, the latter of which transactivates ΔNp63, as centrally important transcriptional regulators of quasi-mesenchymal CSCs that reside in an intermediate EMT state. We found that the transcriptional program controlled by ΔNp63 in CSCs is largely distinct from the one that it orchestrates in normal basal mammary stem cells and, instead, it more closely resembles a regenerative epithelial stem cell response to wounding. Moreover, quasi-mesenchymal CSCs repurpose this program to drive metastatic colonization via autocrine EGFR signaling.

High Expression of CD58 and ALDH1A3 Predicts a Poor Prognosis in Basal-like Breast Cancer.

CD58 is an immune adhesion molecule on the cellular surface. It was previously found that a high expression of CD58 predicted a poor prognosis of patients with lower-grade gliomas. Therefore, the aim of this paper was to investigate the association between CD58 and breast cancer. CD58 gene expression data downloaded from cBioPortal was compared between the different subtypes of breast cancer. Clinical prognosis was examined using Kaplan-Meier analysis and multivariable Cox regression analysis. The association between CD58 expression and immune cell infiltration was estimated using the TIMER 2.0 web platform. Finally, the tumour sphere formation of aldehyde dehydrogenase 1 (ALDH1)high basal-like breast cancer stem cells in which CD58 was knocked down using siRNA was measured. CD58 mRNA was mainly enriched in claudin-low and basal-like subtypes. The high expression of CD58 predicted a good prognosis in patients with luminal A and luminal B breast cancer. This prediction may be due to the association of immune cell infiltration with CD58. Notably, patients with luminal A breast cancer with a high expression of CD58 in association with ALDH1A3 exhibited a good prognosis; however, this did not apply to patients with basal-like breast cancer. The in vitro experiments revealed that knockdown of CD58 inhibited the tumour sphere formation ability of ALDH1high basal-like cancer cells. CD58 may function as a potential prognostic biomarker and therapeutic target in ALDH-positive basal-like cancer stem cells.

TNFAIP3 promotes ALDH-positive breast cancer stem cells through FGFR1/MEK/ERK pathway.

Breast cancer stem cells (BCSCs) are a tiny population of self-renewing cells that may contribute to cancer initiation, progression, and resistance to therapy in patients. In our prior study, we found that tumor necrosis factor alpha-induced protein 3 (TNFAIP3) is necessary for fibroblast growth factors receptor 1 (FGFR1) signaling-promoted tumor growth and progression in breast cancer (BC). This study aims to investigate the involvement of TNFAIP3 in regulating BCSCs. In this work, we showed that ALDH-positive BCSCs were increased by activating the FGFR1-MEK-ERK pathway, meanwhile utilizing FGFR1 inhibitor, MEK inhibitor, or ERK inhibitor reversed the phenomenon in BC cells. Moreover, ALDH-positive BCSCs were decreased in TNFAIP3-knockout or TNFAIP3-depressing cells. In vivo analysis displayed that TNFAIP3-silenced MDA-MB-231 xenografts developed smaller tumors and ALDH immunostaining levels were significantly lower in TNFAIP3-depressing or TNFAIP3-knockout tumor tissues. Besides, our results also revealed that TNFAIP3 influences the transcription stemness factors gene expression. Taken together, TNFAIP3 could be a potential regulator in FGFR1-MEK-ERK-promoted ALDH-positive BCSCs.

An anti-alcoholism drug, disulfiram and copper complex improves radio-resistance of tumor-initiating cells in esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is a malignant cancer with a poor prognosis. Chemoradiotherapy is one of the most important strategies for patients with locally advanced unresectable ESCC; however, its therapeutic effect is unsatisfactory. Tumor-initiating cells (TICs) have been reported to be resistant to conventional chemotherapy and radiotherapy so far. Therefore, we aimed to develop a treatment strategy targeting TICs in ESCC to improve radiosensitivity. First, we validated aldehyde dehydrogenase 1 (ALDH1) as a TIC marker and investigated its ability to mediate resistance in human ESCC cell lines using flow cytometry, Western blotting, and functional analyses. Then, we focused on disulfiram (DSF), an aldehyde dehydrogenase inhibitor, used to treat alcohol use disorder. We investigated the effect of DSF and copper (II) D-gluconate (Cu) on the radiosensitivity of ESCC in xenograft mouse models. ALDH1-positive cells showed an upregulation of SOX2 and Nanog, exhibiting much stronger tumor-initiating properties than ALDH1-negative cells. Furthermore, inhibition of ALDH1 attenuated the tumor-initiating properties of ESCC cell lines. Our results also showed that ALDH1-positive cells were resistant to chemotherapy and radiotherapy, and the inhibition of ALDH1 led to the mitigation of therapeutic resistance. Our in vitro and in vivo studies revealed that the DSF/Cu complex could radiosensitize ALDH1-positive ESCC cells and downregulate the phosphoinositide 3-kinase/Akt pathway. ALDH1 inhibition by the DSF/Cu complex enhances the radiosensitivity of TICs in ESCC. The drug repositioning approach using disulfiram is a potential treatment option to overcome radioresistance in patients with locally advanced ESCC.

Isothiocyanates (ITCs) 1-(Isothiocyanatomethyl)-4-phenylbenzene and 1-Isothiocyanato-3,5-bis(trifluoromethyl)benzene-Aldehyde Dehydrogenase (ALDH) Inhibitors, Decreases Cisplatin Tolerance and Migratory Ability of NSCLC.

One of the main treatment modalities for non-small-cell lung cancer (NSCLC) is cisplatin-based chemotherapy. However, the acquisition of cisplatin resistance remains a major problem. Existing chemotherapy regimens are often ineffective against cancer cells expressing aldehyde dehydrogenase (ALDH). As such, there is an urgent need for therapies targeting ALDH-positive cancer cells. The present study compares the anticancer properties of 36 structurally diverse isothiocyanates (ITCs) against NSCLC cells with the ALDH inhibitor disulfiram (DSF). Their potential affinity to ALDH isoforms and ABC proteins was assessed using AutoDockTools, allowing for selection of three compounds presenting the strongest affinity to all tested proteins. The selected ITCs had no impact on NSCLC cell viability (at tested concentrations), but significantly decreased the cisplatin tolerance of cisplatin-resistant variant of A549 (A549CisR) and advanced (stage 4) NSCLC cell line H1581. Furthermore, long-term supplementation with ITC 1-(isothiocyanatomethyl)-4-phenylbenzene reverses the EMT phenotype and migratory potential of A549CisR to the level presented by parental A549 cells, increasing E-Cadherin expression, followed by decreased expression of ABCC1 and ALDH3A1. Our data indicates that the ALDH inhibitors DSF and ITCs are potential adjuvants of cisplatin chemotherapy.

On the mechanisms of photodynamic action of photosensitizers based on polycationic derivatives of synthetic bacteriochlorin against human lung cancer cells A549 (in vitro study)

BackgroundOne of the tasks of anticancer photodynamic therapy is increasing the efficacy of treatment of cancer nodes with large (clinically relevant) sizes using near-infrared photosensitizers (PS). We study the photodynamic action against A549 human lung cancer cells using PS based on polycationic derivatives of synthetic bacteriochlorin. MethodsThe efficacy and mechanisms of the photodynamic action of PS based on polycationic derivatives of synthetic bacteriochlorin against A549 lung cancer cells were studied in vitro using immunocytochemical and morphological methods. ResultsIt was found that PS based on tetracationic and octacationic derivatives of synthetic bacteriochlorin induce necrosis, apoptosis, decreasing of proliferative and mitotic activity, as well as reducing the number of ALDH1-positive cancer cells with signs of stem cells in A549 human lung cancer cell culture. The IC50 values (concentration of a PS that reduces cells survival by 50%) were about 0.69 μM for tetracationic PS and 0.57 μM for octacationic PS under irradiation at 30 J/cm2 while in the “dark” control they were higher than 100 μM for both PSs. ConclusionsPhotosensitizers based on polycationic derivatives of synthetic bacteriochlorin have high phototoxicity against A549 cancer cells caused by the induction of necrosis and apoptosis of cancer cells, including cells with signs of stemness, and a sharp decrease of mitotic and proliferative activity.

BCL6 and the Notch pathway: a signaling axis leading to a novel druggable biotarget in triple negative breast cancer.

The transcriptional repressor B-cell lymphoma 6 (BCL6) is dysregulated in several neoplasms, but its role in triple negative breast cancer (TNBC), a highly aggressive subtype which lacks effective treatment, is unclear. The presence of intratumoral cancer stem cells (CSCs) is a main cause of tumor relapse. The Notch signaling pathway is crucial for regulating CSC self-renewal and promoting breast cancer (BC) development and resistance to anticancer therapies. Here, we investigated signaling cascades of BCL6 in the CSC compartment of TNBCs, and the mechanisms that govern its activity, mainly through Notch signaling. Gene expression, somatic copy number alterations and clinical data from the Cancer Genome Atlas and METABRIC were accessed through theXena and cbioportal browsers. Public transcriptome profiles from TNBC datasets were retrieved from theGene Expression Omnibus. Mammosphere formation efficiency was calculated after BCL6 knockdown via transient siRNAtransfection, stable silencing or pharmacological inhibition. The effects exhibited via BCL6 inhibition in putative TNBC stem-like cells were evaluated by immunofluorescence and qRT-PCR analyses. Chromatin immunoprecipitation experiments were performed to validate a putative BCL6 responsive element located in the first intron of the Numb gene and to define the circuit of corepressors engaged by BCL6 following its inhibition. Immunoprecipitation assays were carried out to investigate a novel interaction at the basis of BCL6 control of CSC activity in TNBC. In silico analyses of benchmarked public datasets revealed a significant enrichment of BCL6 in cancer stemness related pathways, particularly of Notch signaling in TNBC. In vitro stable inhibition of BCL6 significantly reduced tumor cell growth and, accordingly, we found thatthe mammosphere formation efficiency of BCL6 silencedcells was significantly impaired by pharmacological inhibition of Notch signaling. BCL6 was found to be expressed at significantly higher levels in TNBC mammospheres than in their adherent counterparts, and loss of BCL6 function significantly decreased mammosphere formation with preferential targeting of CD44-positive versus ALDH-positive stem-like cells. Functional interplay between BCL6 and the chromatin remodeling factor EZH2 triggered the BCL6/Notch stemness signaling axis via inhibition of Numb transcription. Our results may be instrumental for the prospective design of combination treatment strategies that selectively target novel TNBC-associated biomarker(s) whose activity is implicated in the regulation of cancer stemness (such as BCL6) and molecules in developmentally conserved signaling pathways (such as Notch) to achieve long-lasting tumor control and improve patient outcomes.

Niclosamide induces miR-148a to inhibit PXR and sensitize colon cancer stem cells to chemotherapy.

SummaryTumor recurrence is often attributed to cancer stem cells (CSCs). We previously demonstrated that down-regulation of Pregnane X Receptor (PXR) decreases the chemoresistance of CSCs and prevents colorectal cancer recurrence. Currently, no PXR inhibitor is usable in clinic. Here, we identify miR-148a as a targetable element upstream of PXR signaling in CSCs, which when over-expressed decreases PXR expression and impairs tumor relapse after chemotherapy in mouse tumor xenografts. We then develop a fluorescent reporter screen for miR-148a activators and identify the anti-helminthic drug niclosamide as an inducer of miR-148a expression. Consequently, niclosamide decreased PXR expression and CSC numbers in colorectal cancer patient-derived cell lines and synergized with chemotherapeutic agents to prevent CSC chemoresistance and tumor recurrence in vivo. Our study suggests that endogenous miRNA inducers is a viable strategy to down-regulate PXR and illuminates niclosamide as a neoadjuvant repurposing strategy to prevent tumor relapse in colon cancer.

Efficient DNA Repair Mitigates Replication Stress Resulting in Less Immunogenic Cytosolic DNA in Radioresistant Breast Cancer Stem Cells

Cancer stem cells (CSCs) are a major cause of tumor therapy failure. This is mainly attributed to increased DNA repair capacity and immune escape. Recent studies have shown that functional DNA repair via homologous recombination (HR) prevents radiation-induced accumulation of DNA in the cytoplasm, thereby inhibiting the intracellular immune response. However, it is unclear whether CSCs can suppress radiation-induced cytoplasmic dsDNA formation. Here, we show that the increased radioresistance of ALDH1-positive breast cancer stem cells (BCSCs) in S phase is mediated by both enhanced DNA double-strand break repair and improved replication fork protection due to HR. Both HR-mediated processes lead to suppression of radiation-induced replication stress and consequently reduction of cytoplasmic dsDNA. The amount of cytoplasmic dsDNA correlated significantly with BCSC content (p=0.0002). This clearly indicates that HR-dependent avoidance of radiation-induced replication stress mediates radioresistance and contributes to its immune evasion. Consistent with this, enhancement of replication stress by inhibition of ataxia telangiectasia and RAD3 related (ATR) resulted in significant radiosensitization (SER37 increase 1.7-2.8 Gy, p<0.0001). Therefore, disruption of HR-mediated processes, particularly in replication, opens a CSC-specific radiosensitization option by enhancing their intracellular immune response.

Lysophosphatidic Acid Promotes the Expansion of Cancer Stem Cells via TRPC3 Channels in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is a highly aggressive cancer for which targeted therapeutic agents are limited. Growing evidence suggests that TNBC originates from breast cancer stem cells (BCSCs), and elucidation of the molecular mechanisms controlling BCSC proliferation will be crucial for new drug development. We have previously reported that the lysosphingolipid sphingosine-1-phosphate mediates the CSC phenotype, which can be identified as the ALDH-positive cell population in several types of human cancer cell lines. In this study, we have investigated additional lipid receptors upregulated in BCSCs. We found that lysophosphatidic acid (LPA) receptor 3 was highly expressed in ALDH-positive TNBC cells. The LPAR3 antagonist inhibited the increase in ALDH-positive cells after LPA treatment. Mechanistically, the LPA-induced increase in ALDH-positive cells was dependent on intracellular calcium ion (Ca2+), and the increase in Ca2+ was suppressed by a selective inhibitor of transient receptor potential cation channel subfamily C member 3 (TRPC3). Moreover, IL-8 production was involved in the LPA response via the activation of the Ca2+-dependent transcriptional factor nuclear factor of activated T cells. Taken together, our findings provide new insights into the lipid-mediated regulation of BCSCs via the LPA-TRPC3 signaling axis and suggest several potential therapeutic targets for TNBC.

Zinc-finger protein 750 mitigates malignant biological behavior of oral CSC-like cells enriched from parental CAL-27 cells.

Oral squamous cell carcinoma (OSCC) is the most commonly occurring oral malignancy. Cancer stem cells (CSCs) are known to be responsible for cancer recurrence and metastasis. Zinc-finger protein 750 (ZNF750) has been reported to inhibit OSCC cell proliferation and invasion. The present study aimed to elucidate the role of ZNF750 in the inhibition of the renewal ability of CSCs derived from the OSCC cell line, CAL-27. The effects of ZNF750 on CSC-like properties were examined using aldehyde dehydrogenase (ALDH), tumor sphere formation and colony formation assays. Reverse transcription-quantitative PCR and western blotting were performed to detect the expression levels of octamer-binding transcription factor 4, sex-determining region Y-box 2, the enhancer of zeste homolog 2 (Ezh2), embryonic ectoderm development (EED) and SUZ12 polycomb repressive complex 2 subunit (SUZ12), and for the identification of genes associated with metastasis. ZNF750 effectively attenuated CSC-like cell self-renewal abilities; ZNF750 decreased the ALDH-positive cell population, tumor sphere and colony formation abilities, cell viability and stemness factors. Furthermore, the expression levels of Ezh2, EED and SUZ12 were decreased by ZNF750. ZNF750 inhibited MMP1, 3, 9 and 13 expression levels, and decreased the cell invasion and migratory abilities. Moreover, the expression of tissue inhibitors of matrix metalloproteinases-1 was increased by ZNF750. However, opposite effects were observed following the knockdown of the ZNF750 gene. Overall, the present study demonstrated that ZNF750 has the potential to inhibit the renewal of CSC-like cells enriched from parental CAL-27 cells.

SOD2 Enhancement by Long-Term Inhibition of the PI3K Pathway Confers Multi-Drug Resistance and Enhanced Tumor-Initiating Features in Head and Neck Cancer.

The phosphoinositide-3-kinase (PI3K) pathway has widely been considered as a potential therapeutic target for head and neck cancer (HNC); however, the application of PI3K inhibitors is often overshadowed by the induction of drug resistance with unknown mechanisms. In this study, PII3K inhibitor resistant cancer cells were developed by prolonged culturing of cell lines with BEZ235, a dual PI3K and mammalian target of rapamycin (mTOR) inhibitor. The drug resistant HNC cells showed higher IC50 of the proliferation to inhibitors specifically targeting PI3K and/or mTOR, as compared to their parental cells. These cells also showed profound resistance to drugs of other classes. Molecular analysis revealed persistent activation of phosphorylated AKT at threonine 308 in the drug resistant cells and increased expression of markers for tumor-initiating cells. Interestingly, increased intra-cellular ROS levels were observed in the drug resistant cells. Among anti-oxidant molecules, the expression of SOD2 was increased and was associated with the ALDH-positive tumor-initiating cell features. Co-incubation of SOD inhibitors and BEZ235 decreased the stemness feature of the cells in vitro, as shown by results of the spheroid formation assay. In conclusion, dysregulation of SOD2 might contribute to the profound resistance to PI3K inhibitors and the other drugs in HNC cells.

ALDH1 and SALL4 Expression in Cell Block Samples from Patients with Lung Adenocarcinoma and Malignant Pleural Effusion.

Malignant pleural effusion (MPE) can accompany advanced lung adenocarcinoma. Recent studies suggest that MPE could contain a heterogeneous subpopulation of cells with stem-like properties, such as tumorigenicity and self-renewal, indicating that they could be the source of metastasis. Although previous studies analyzed the correlation between cancer stem cell (CSC) marker expression and clinical outcomes using lung cancer tissues, investigations regarding the association of MPE with CSC marker expression are limited. We performed immunohistochemistry to examine the expression of aldehyde dehydrogenase 1 (ALDH1) and Sal-like 4 (SALL4) in 46 cell block samples of MPE from patients with lung adenocarcinoma. ALDH1-positive and SALL4-positive cancer cells in MPE were detected in 30 (65.2%) and 21 samples (45.7%), respectively. Cluster formation was detected in 26 samples (56.5%). The number of clusters was significantly higher in ALDH1-positive/SALL4-negative samples. SALL4 expression was inversely correlated with the cluster ratio (r = −0.356) and positively associated with the Ki-67 index (r = 0.326), suggesting that MPE cells with high SALL4 expression comprised the proliferative subpopulation. In conclusion, we demonstrated that MPE contains an ALDH1-positive/SALL4-negative subpopulation exhibiting cluster formation and a SALL4-positive proliferative subpopulation.

FTY720 Inhibits Expansion of Breast Cancer Stem Cells via PP2A Activation.

Growing evidence suggests that breast cancer originates from a minor population of cancer cells termed cancer stem cells (CSCs), which can be identified by aldehyde dehydrogenase (ALDH) activity-based flow cytometry analysis. However, novel therapeutic drugs for the eradication of CSCs have not been discovered yet. Recently, drug repositioning, which finds new medical uses from existing drugs, has been expected to facilitate drug discovery. We have previously reported that sphingosine kinase 1 (SphK1) induced proliferation of breast CSCs. In the present study, we focused on the immunosuppressive agent FTY720 (also known as fingolimod or Gilenya), since FTY720 is known to be an inhibitor of SphK1. We found that FTY720 blocked both proliferation of ALDH-positive cells and formation of mammospheres. In addition, we showed that FTY720 reduced the expression of stem cell markers such as Oct3/4, Sox2 and Nanog via upregulation of protein phosphatase 2A (PP2A). These results suggest that FTY720 is an effective drug for breast CSCs in vitro.

Magnolol inhibits cancer stemness and IL-6/Stat3 signaling in oral carcinomas

Cancer stem cells (CSCs) have been known to be implicated in tumorigenesis, metastasis, and drug resistance in oral squamous cell carcinomas (OSCC). In this study, we aimed to investigate whether magnolol, a polyphenolic component derived from Magnolia officinalis, exhibited the anti-CSCs properties. The cytotoxicity of magnolol was tested using normal gingival epithelioid SG cells and sphere-forming OSCC-CSCs isolated from SAS, OECM1, and GNM cells. Secondary sphere-forming ability, the proportion of ALDH1 positive cells, Transwell migration, and invasion capacities were examined as well. The chemosensitive effects of magnolol were investigated using MTT, secondary sphere-forming, and invasion assays. Magnolol exerted a higher cytotoxicity of OSCC-CSCs and cancer stemness features, including self-renewal ability, the expression CSC marker, migration, and invasion capacities were all downregulated in magnolol-treated OSCC-CSCs. Moreover, administration of magnolol potentiated the effect of cisplatin, including a decrease in cell viability, self-renewal, and invasion activities. In addition, we observed that the secretion of IL-6 and phosphorylation of Stat3 were decreased in OSCC-CSCs treated with magnolol. Our data suggest that magnolol is able to target CSCs and suppress the cancer stemness properties, at least in part, via IL-6/Stat3 signaling. Besides, a dietary supplement of magnolol may function as an adjunct to cisplatin treatment.

Identification and Quantification of “Cancer Stem Cells” in Potentially Malignant and Malignant Disorders of Head and Neck Region: A Flow-Cytometric Study

Background: Cancer stem cells (CSCs) are subpopulation of cells existent in a cancerous mass of cells. These CSCs hijack the properties of stem cells like self-renewal as well as being resistant to any conventional cancer therapies. The objective of this study was to identify and quantify the presence of these CSCs by using surface markers CD44, CD133, and ALDH1 among three groups of subjects who were age and gender matched (Normal controls, Oral cancers, Potentially malignant disorders). Methodology: This study was conducted in a sample of 108 subjects who were divided into three groups: Group I- Controls (C), Group II- Oral cancer (OC), Group III- Potentially Malignant Disorders (PMDs). Among them, 40 subjects each were present in Group I &amp; II and 28 subjects were included in group III, and they were respectively diagnosed histopathologically as OC and PMDs. The identification of the sub-population of CSCs by means of above mentioned surface markers was done using Flow cytometry. Results interpretation: Non-parametric tests were applied. Median age limit was 59 years in group II, which was higher than Group I or Group III subjects. p-value was 0.002* which was significant. Group I included 19 females (47.5%) and 21 males (52.5%). In Group II, 18 subjects were females (45%) and 22 were males (55%), and in Group III, 17 subjects were females (60.7%) and 11 were males (39.3%). p-value was 0.409, which was not very significant. In group II, 23 subjects (57.5%) were in clinical stage 2, 11 (27,5%) were in clinical stage 1 and six (15%) were in clinical stage 3. Histopathologically in group II, 15 (37.5%) were in grade I, 20 (50%) were in grade II and five (12.5%) were in grade III. Results of the three groups were compared and correlated regarding the presence of cancer stem cells based on the surface markers CD44, CD133 and ALDH1. Unlike other similar studies, our study showed no statistically significant presence of CD44, ALDH1 positive cells, but only CD133 was slightly significant. Inference: The results of our study showed no statistically significant evidence in identification of the presence of cancer stem cells in the oral cancers as well as potentially malignant disorders based on the presence of surface markers.