Melanoma Cancer Stem Cells Research Articles

261 Distinct Melanoma Cancer Stem Cell Profile Defines Response to Immune Checkpoint Inhibitors in Melanomas

This study examined the relationship between cancer stem cells (CSCs) and the susceptibility of melanomas to immunotherapy. Immunotherapy has emerged as a pillar of cancer therapy, demonstrating marked survival benefits in patients with melanoma. However, there remains a significant number of patients who fail to respond to treatment. To improve patient outcomes and optimise appropriate drug selection, it is a priority to unravel the mechanisms underlying the emergence of drug resistance. One of the most important key players to have emerged in the development of drug resistance are the CSCs. Transcriptomic profiles of 158 melanoma biopsies from patients treated with anti-programmed cell death 1 ligand-1 (PD-1) monotherapy (nivolumab or pembrolizumab; n = 63) or combined anti-PD-1 and anti-cytotoxic T lymphocyte antigen 4 (CTLA-4; ipilimumab; n = 57) were obtained. Patients were categorised as responders or non-responders using the Response Evaluation Criteria in Solid Tumors (RECIST) criteria, where responders had a complete response (CR), partial response (PR), or stable disease (SD) of greater than 6 months with no disease progression. Samples were analysed for differential gene expression after normalisation using DESeq2 and HTSeq. Gene levels of well-defined CSCs were compared between responders and non-responders. Changes to gene expression levels were also compared pre and post immunotherapy treatment. This analysis identified a unique melanoma CSC marker gene expression profile “signature” that corresponds to a positive response to immune checkpoint inhibitors. This gene expression profile was associated with longer progression-free survival in patients treated with immunotherapy agents. Therefore, we have identified a CSC gene expression profile that can be used to predict patient response to the current available anti-PD-1 or anti-CTLA4 based therapies. Furthermore, this helps with identifying patients in need of other therapeutic strategies.

Expression of cathepsins B and D by cancer stem cells in head and neck metastatic malignant melanoma.

We have previously demonstrated cancer stem cell (CSC) subpopulations in head and neck metastatic malignant melanoma (HNmMM), and the expression of components of the renin-angiotensin system (RAS) by these CSCs. Cathepsins B, D and G are involved in carcinogenesis and constitute bypass loops of the RAS. This study investigated the expression and localization of cathepsins B, D and G, in relation to these CSCs. Immunohistochemical staining demonstrated expression of cathepsins B, D and G in HNmMM sections from all 20 patients. Western blotting confirmed the presence of cathepsins B and D proteins in all six HNmMM tissue samples and four HNmMM-derived primary cell lines. RT-qPCR showed transcript expression of cathepsins B, D and G in all six HNmMM tissue samples, and cathepsins B and D but not cathepsin G in all four HNmMM-derived primary cell lines. Enzymatic activity assays demonstrated cathepsins B and D were active in all six HNmMM tissue samples. Immunofluorescence staining performed on two of the HNmMM tissue samples demonstrated expression of cathepsins B and D by the CSCs, and cathepsin G by cells within the peritumoral stroma. Our novel findings suggest the possibility of targeting these CSCs by modulation of paracrine RAS signaling.

Suitability of CD133 as a Marker for Cancer Stem Cells in Melanoma.

Objectives:CD133 is considered a cancer stem cell (CSC) marker in various malignancies; however, its role as a biomarker of malignant melanoma remains controversial. The present study was conducted to evaluate the suitability of CD133 surface antigen as a CSC marker in melanoma. Methods:Human melanoma cells were fractionally separated by magnetic cell separation depending on the CD133 phenotype and transplanted into immunodeficient mice to evaluate their tumorigenic capacity. Furthermore, the time until the development of a palpable tumor and the growth rate were measured, and the final tumor volume was assessed after 8 weeks. The immunohistochemical expression of CD133 in the induced neoplasia was then compared using histomorphometry. Results:Notably, neoplasms were induced in all the groups (n = 48), including in the CD133-negative group. Tumors induced by unsorted cells had the largest volume (p = 0.014) but were detected significantly later in this group (p ≤ 0.001). Interestingly, all explanted tumors expressed CD133, with no significant differences among groups. Conclusions:In contrast to the results obtained in prior studies, the suitability of CD133 as a CSC marker could not be demonstrated. The current encouraging progress in targeted therapy for malignant melanoma highlights the need to identify more effective targets.

CD133 Antigen as a Potential Marker of Melanoma Stem Cells: In Vitro and In Vivo Studies.

Melanoma is the most dangerous type of skin cancer. Cancer stem cells (CSCs) are suspected to be responsible for the cancer recurrence and in the consequence for cancer therapy failure. CD133 is a potential marker for detection of melanoma CSCs. Experiments were performed on the B16-F10 mouse melanoma cell line. CD133+ cells were isolated using an immunomagnetic cell sorting technique. After isolation proliferative and clonogenic potential of CD133+, CD133- and CD133+/- were evaluated. The potential of CD133+ and CD133- cells for tumor induction was conducted on C57BL/6J mouse model. Three different cell quantities (100, 1000, 10000) were tested. Tumor morphology, number of mitoses, and tumor necrosis area were analyzed. Average 0.12% CD133+ cells were isolated. Compared to CD133- and unsorted CD133+/- cells, CD133+ cells were characterized by the higher proliferative and clonogenic potential. These properties were not confirmed in vivo, as both CD133+ and CD133- cells induced tumor growth in mouse model. No statistical differences in mitosis number and tumor necrosis area were observed. Simultaneous detection of CD133 antigen with other markers is necessary for accurate identification of these melanoma cancer stem cells.

Metabolomic profile of cancer stem cell-derived exosomes from patients with malignant melanoma.

Malignant melanoma (MM) is the most aggressive and life-threatening form of skin cancer. It is characterized by an extraordinary metastasis capacity and chemotherapy resistance, mainly due to melanoma cancer stem cells (CSCs). To date, there are no suitable clinical diagnostic, prognostic or predictive biomarkers for this neoplasia. Therefore, there is an urgent need for new MM biomarkers that enable early diagnosis and effective disease monitoring. Exosomes represent a novel source of biomarkers since they can be easily isolated from different body fluids. In this work, a primary patient-derived MM cell line enriched in CSCs was characterized by assessing the expression of specific markers and their stem-like properties. Exosomes derived from CSCs and serums from patients with MM were characterized, and their metabolomic profile was analysed by high-resolution mass spectrometry (HRMS) following an untargeted approach and applying univariate and multivariate statistical analyses. The aim of this study was to search potential biomarkers for the diagnosis of this disease. Our results showed significant metabolomic differences in exosomes derived from MM CSCs compared with those from differentiated tumour cells and also in serum-derived exosomes from patients with MM compared to those from healthy controls. Interestingly, we identified similarities between structural lipids differentially expressed in CSC-derived exosomes and those derived from patients with MM such as the glycerophosphocholine PC 16:0/0:0. To our knowledge, this is the first metabolomic-based study aimed at characterizing exosomes derived from melanoma CSCs and patients' serum in order to identify potential biomarkers for MM diagnosis. We conclude that metabolomic characterization of CSC-derived exosomes sets an open door to the discovery of clinically useful biomarkers in this neoplasia.

Research Progress of Cancer Stem Cells in Uveal Melanoma.

Uveal melanoma is the most common malignant tumor in adult eyes, mostly in the choroid, but also in the iris and ciliary body. Distant metastasis is found in nearly half of the patients. Cancer stem cells are a kind of cells with the ability of self-renewal and multidirectional differentiation, which are related to tumor invasion and metastasis. Although the concept of cancer stem cells is relatively mature in other tumors, its existence and verification methods in uveal melanoma are still uncertain. A more in-depth understanding of cancer stem cells and their mechanism may reveal new strategies to treat uveal melanoma. This article reviews the concept of cancer stem cells and their research progress in uveal melanoma, including identification, probable markers, cancer stem cell targeted drug therapy and the controversies and prospects in this field.

The Evaluation of Effect of Aurora Kinase Inhibitor CCT137690 in Melanoma and Melanoma Cancer Stem Cell.

Dysregulation of the cell cycle is one of the main causes of melanomagenesis. Genomewide studies showed that the expression of Aurora -A and -B significantly has been upregulated in melanoma. However, there is no FDA approved drug targeting aurora kinases in the treatment of melanoma. In addition, the development of resistance to chemotherapeutic agents in the treatment of melanoma and, as a result, the relapse due to heterogeneous cell groups in patients is a second phenomenon that causes treatment failure. Therefore, there is an urgent need for therapeutic alternatives targeting both melanoma and Melanoma Cancer Stem Cells (MCSCs) in treatments. At this stage, cell cycle regulators become promising targets. In this study, we aimed to identify the effects of Aurora kinase inhibitor CCT137690 on the cytotoxicity, apoptosis, cell cycle, migration, and colony formation and expression changes of genes related to proliferation, cell death and cell cycle in melanoma and melanoma cancer stem cell. In addition, we investigated the apoptotic and cytostatic effects of CCT137690 in normal fibroblast cells. We evaluated the cytotoxic effect of CCT137690 in MCSCs, NM2C5 referring as melanoma model cells and WI-38 cells by using the WST-1 test. The effect of CCT137690 on apoptosis was detected via Annexin V and JC-1 method; on cell cycle progression by cell cycle test; on gene expression by using RT-PCR, on migration activity by wound healing assay and clonal growth by clonogenic assay in NM2C5 cells and MCSCs. The effects of CCT137690 in WI-38, referring as healthy fibroblast cell, were assessed through Annexin V and cell cycle method. CCT137690 was determined to have a cytotoxic and apoptotic effect in MCSCs and melanoma. It caused polyploidy and cell cycle arrest at the G2/M phase in MCSCs and melanoma cells. The significant decrease in the expression of MMP2, MMP7, MMP10, CCNB1, IRAK1, PLK2 genes, and the increase in the expression of PTEN, CASP7, p53 genes were detected. Aurora kinases inhibitor CCT137690 displays promising anticancer activity in melanoma and especially melanoma cancer stem cells. The effect of CCT137690 on melanoma and MCSC may provide a new approach to treatment protocols.

Cisplatin-Induced Giant Cells Formation Is Involved in Chemoresistance of Melanoma Cells.

Melanoma is notoriously resistant to current cancer therapy. However, the chemoresistance mechanism of melanoma remains unclear. The present study unveiled that chemotherapy drug cisplatin induced the formation of giant cells, which exhibited enlargement in cell diameter and nucleus in mice and human melanoma cells. Giant cells were positive with melanoma maker S100 and cancer stem cell markers including ABCB5 and CD133 in vitro and in vivo. Moreover, giant cells retained the mitotic ability with expression of proliferation marker Ki-67 and exhibited multiple drug resistance to doxorubicin and actinomycin D. The mitochondria genesis/activities and cellular ATP level were significantly elevated in giant cells, implicating the demand for energy supply. Application of metabolic blockers such as sodium azide or 2-deoxy glucose abolished the cisplatin-induced giant cells formation and expression of cancer stemness markers. The present study unveils a novel chemoresistance mechanism of melanoma cells via size alteration and the anti-neoplastic strategy by targeting giant cells.

Aggressiveness Potential of Spontaneous Canine Mucosal Melanoma Can Dictate Distinct Cancer Stem Cell Compartment Behaviors in Regard to Their Initial Size and Expansion Abilities.

Mucosal melanoma represents one of the most highly metastatic and aggressive subtypes of melanoma. The biology of mucosal melanoma is poorly documented, and the lack of experimental models makes it difficult to design and test new therapies. Dogs are frequently affected by melanomas of the oral cavity, making spontaneous canine melanoma a potentially predictable model for their human counterpart. We recently established and characterized two new canine mucosal melanoma cell lines named OCR_OCMM1 and OCR_OCMM2. Here, we identified quiescent cancer stem cell (CSC) subpopulations in both canine cell lines that displayed similarities to human quiescent CSCs: canine melanoma CSCs had the ability to self-renew, produced nonstem cell (SC) progeny, and formed melanospheres that recapitulated the phenotypic profile of the parental tumor. These CSCs also formed melanoma in immunodeficient mice, and the inhibition of PI3K/AKT signaling expanded the CSC pool. A subset of non-CSCs transitioned to become CSCs. OCR_OCMM1 and OCR_OCMM2 displayed different CSC compartment behaviors in regard to their initial size and expansion abilities. Collectively, this study showed that the OCR_OCMM1 and OCR_OCMM2 canine melanoma cell lines are powerful cellular tools to study melanoma SCs, not only for mucosal but also for the more common human cutaneous melanoma.

SOX2 upregulates side population cells and enhances their chemoresistant ability by transactivating ABCC1 expression contributing to intrinsic resistance to paclitaxel in melanoma.

Paclitaxel is the last choice for the treatment of advanced melanoma as a second-line chemotherapeutic agent, but there are still many cases of intrinsic resistance to paclitaxel in melanoma and the reasons that cause paclitaxel resistance remain unclear. Here, we identified that high expression of SRY-box transcription factor 2 (SOX2) and high ratio of side population (SP) cells reduced the sensitivity to paclitaxel in melanoma cells. The knockout and the induction of SOX2 completely depleted and significantly upregulated the ratios of melanoma SP cells, respectively. These data suggest that SOX2, a pluripotent transcription factor for inducing cancer stem cells in melanoma, is also sufficient and necessary for the induction of melanoma SP cells. ATP-binding cassette (ABC) subfamily C member 1 (ABCC1) is one of ABC transporters which causes SP cells to be resistance to chemotherapeutic agents by efficiently pumping drugs out of cells. The knockout and the induction of ABCC1 significantly increased and decreased the sensitivity of melanoma cells to paclitaxel. High expression of ABCC1 was identified in melanoma cell lines with high expression of SOX2 and in their SP cells. SOX2 was identified to induce ABCC1 transcription. Taken together, SOX2 upregulates SP cells and enhances their chemoresistant ability by increasing ABCC1 expression, which contributes to intrinsic resistance to paclitaxel in melanoma. Our findings will lead to new insights into melanoma biology and therapy resistance, and eventually to new therapeutic targets.

A novel oral micellar fenretinide formulation with enhanced bioavailability and antitumour activity against multiple tumours from cancer stem cells

BackgroundAn increasing number of anticancer agents has been proposed in recent years with the attempt to overcome treatment-resistant cancer cells and particularly cancer stem cells (CSC), the major culprits for tumour resistance and recurrence. However, a huge obstacle to treatment success is the ineffective delivery of drugs within the tumour environment due to limited solubility, short circulation time or inconsistent stability of compounds that, together with concomitant dose-limiting systemic toxicity, contribute to hamper the achievement of therapeutic drug concentrations. The synthetic retinoid Fenretinide (4-hydroxy (phenyl)retinamide; 4-HPR) formerly emerged as a promising anticancer agent based on pre-clinical and clinical studies. However, a major limitation of fenretinide is traditionally represented by its poor aqueous solubility/bioavailability due to its hydrophobic nature, that undermined the clinical success of previous clinical trials.MethodsHere, we developed a novel nano-micellar fenretinide formulation called bionanofenretinide (Bio-nFeR), based on drug encapsulation in an ion-pair stabilized lipid matrix, with the aim to raise fenretinide bioavailability and antitumour efficacy.ResultsBio-nFeR displayed marked antitumour activity against lung, colon and melanoma CSC both in vitro and in tumour xenografts, in absence of mice toxicity. Bio-nFeR is suitable for oral administration, reaching therapeutic concentrations within tumours and an unprecedented therapeutic activity in vivo as single agent.ConclusionAltogether, our results indicate Bio-nFeR as a novel anticancer agent with low toxicity and high activity against tumourigenic cells, potentially useful for the treatment of solid tumours of multiple origin.

Melanoma cancer stem-like cells: Optimization method for culture, enrichment and maintenance

Malignant Melanoma is known for being one of the most aggressive cancers with an incidence that increases every year. Cancer stem cells (CSCs) are involved in the resistance to therapeutic treatments, the metastatic process, and the patient’s relapses. Thus, it is of vital importance for researchers to find the methodology that allows us to obtain enriched subpopulations that maintain their stem-like properties without differentiating over time. In the present manuscript, our objective was to compare the ability of conditioned medium obtained from human mesenchymal stem cells (MSCs) isolated by enzymatic and non-enzymatic methods for the enrichment and maintenance of melanoma CSCs. Our results showed for the first time that MSCs isolated by less aggressive methodology displayed higher CSCs enrichment and maintenance capacity. Because they do not undergo enzymatic and proteolytic stress, MSCs produce a greater amount of signalling molecules, helping to improve the phenotype characteristics of CSCs and keep them over time.

MicroRNA-92 Expression in CD133+ Melanoma Stem Cells Regulates Immunosuppression in the Tumor Microenvironment via Integrin-Dependent Activation of TGFβ.

In addition to being refractory to treatment, melanoma cancer stem cells (CSC) are known to suppress host antitumor immunity, the underlying mechanisms of which need further elucidation. In this study, we established a novel role for miR-92 and its associated gene networks in immunosuppression. CSCs were isolated from the B16-F10 murine melanoma cell line based on expression of the putative CSC marker CD133 (Prominin-1). CD133+ cells were functionally distinct from CD133- cells and showed increased proliferation in vitro and enhanced tumorigenesis in vivo. CD133+ CSCs also exhibited a greater capacity to recruit immunosuppressive cell types during tumor formation, including FoxP3+ Tregs, myeloid-derived suppressor cells (MDSC), and M2 macrophages. Using microarray technology, we identified several miRs that were significantly downregulated in CD133+ cells compared with CD133- cells, including miR-92. Decreased expression of miR-92 in CSCs led to higher expression of target molecules integrin αV and α5 subunits, which, in turn, enhanced TGFβ activation, as evidenced by increased phosphorylation of SMAD2. CD133+ cells transfected with miR-92a mimic and injected in vivo showed significantly decreased tumor burden, which was associated with reduced immunosuppressive phenotype intratumorally. Using The Cancer Genome Atlas database of patients with melanoma, we also noted a positive correlation between integrin α5 and TGFβ1 expression levels and an inverse association between miR-92 expression and integrin alpha subunit expression. Collectively, this study suggests that a miR-92-driven signaling axis involving integrin activation of TGFβ in CSCs promotes enhanced tumorigenesis through induction of intratumoral immunosuppression. SIGNIFICANCE: CD133+ cells play an active role in suppressing melanoma antitumor immunity by modulating miR-92, which increases influx of immunosuppressive cells and TGFβ1 expression.

Shrimp miRNA suppresses the stemness of human cancer stem cellsviathe PIN1 pathway

A tumor can be regarded as a cell mass with a biologic hierarchy that is orchestrated by cancer stem cells. The specific features of cancer stem cells may result from a series of gene expression regulatory mechanisms. As important regulatory factors of gene expression, microRNAs (miRNAs) have critical roles in the self-renewal, pluripotency, differentiation, and tumorigenicity of cancer stem cells. However, the effects of animal miRNAs on cancer stem cells have not been investigated extensively. Here, we report that miRNA of a shrimp, Marsupenaeus japonicus (mja-miR-35-3p), which possesses antiviral activity in shrimp, could suppress the proliferation of melanoma and breast cancer stem cells, but not cancer nonstem cells, by arresting the cell cycle in the G1 phase and inducing apoptosis. Shrimp mja-miR-35-3p (named mja-miR-35) targets the human peptidylprolyl cis/trans isomerase, never in mitosis gene a-interacting 1 (PIN1) gene, which is up-regulated in cancer stem cells. The results indicated that PIN1 silencing caused cell cycle arrest in the G1 phase, induced apoptosis, and decreased the sphere-forming capacity of cancer stem cells, but not cancer nonstem cells, showing that PIN1 had beneficial effects against the stemness of cancer stem cells. The in vivo data revealed that shrimp mja-miR-35 suppressed the stemness of cancer stem cells in mice by inhibiting the PIN1 antiapoptotic-cell cycle pathway. These findings demonstrated that the miRNAs of aquatic animals might be an important source for discovering human antitumor drugs.-Zhang, S., Zhang, X. Shrimp miRNA suppresses the stemness of human cancer stem cells via the PIN1 pathway.

Cellular and molecular biology of cancer stem cells in melanoma: Possible therapeutic implications.

Malignant melanoma is a tumor characterized by a very high level of heterogeneity, responsible for its malignant behavior and ability to escape from standard therapies. In this review we highlight the molecular and biological features of the subpopulation of cancer stem cells (CSCs), well known to be characterized by self-renewal properties, deeply involved in triggering the processes of tumor generation, metastasis, progression and drug resistance. From the molecular point of view, melanoma CSCs are identified and characterized by the expression of stemness markers, such as surface markers, ATP-binding cassette (ABC) transporters, embryonic stem cells and intracellular markers. These cells are endowed with different functional features. In particular, they play pivotal roles in the processes of tumor dissemination, epithelial-to-mesenchymal transition (EMT) and angiogenesis, mediated by specific intracellular signaling pathways; moreover, they are characterized by a unique metabolic reprogramming. As reported for other types of tumors, the CSCs subpopulation in melanoma is also characterized by a low immunogenic profile as well as by the ability to escape the immune system, through the expression of a negative modulation of T cell functions and the secretion of immunosuppressive factors. These biological features allow melanoma CSCs to escape standard treatments, thus being deeply involved in tumor relapse. Targeting the CSCs subpopulation is now considered an attractive treatment strategy; in particular, combination treatments, based on both CSCs-targeting and standard drugs, will likely increase the therapeutic options for melanoma patients. The characterization of CSCs in liquid biopsies from single patients will pave the way towards precision medicine.

Retinoic acid increases the anticancer effect of paclitaxel by inducing differentiation of cancer stem cells in melanoma.

Malignant melanoma is a highly lethal disease, and advanced stages of melanoma have proven to be resistant to many chemotherapeutic drugs including temozolomide and paclitaxel. Cancer stem cells (CSCs) have been identified and isolated in different cancers including melanoma, and have been proven playing important role in the drug resistance. Retinoic acid (RA) is a promising anticancer agent, which can induce differentiation of CSCs. The main purpose of the present study was to evaluate the possible RA-induced differentiation of melanoma CSCs and sensitization of melanoma CSCs to paclitaxel. Our results show that CSCs of human melanoma A375 cells was more tolerant to paclitaxel than other non-CSCs melanoma cells. On the contrary, RA had stronger inhibitory effect on melanoma CSCs than on non-CSCs. At the same time, RA could arrest the cell cycle of CSCs and reduce the expression of Sox-2, Oct-4 in CSCs of melanoma, thereby induced the differentiation of CSCs and increased its sensitivity to paclitaxel. With this study we concluded that RA increases the anticancer effect of paclitaxel by inducing differentiation of cancer stem cells in melanoma, and the combined application of RA and paclitaxel may be more effective in the treatment of melanoma.

Inhibition of Stearoyl-CoA desaturase 1 reverts BRAF and MEK inhibition-induced selection of cancer stem cells in BRAF-mutated melanoma

BackgroundCombination therapy with BRAF and MEK inhibitors significantly improves survival in BRAF mutated melanoma patients but is unable to prevent disease recurrence due to the emergence of drug resistance. Cancer stem cells (CSCs) have been involved in these long-term treatment failures. We previously reported in lung cancer that CSCs maintenance is due to altered lipid metabolism and dependent upon Stearoyl-CoA-desaturase (SCD1)-mediated upregulation of YAP and TAZ. On this ground, we investigated the role of SCD1 in melanoma CSCs.MethodsSCD1 gene expression data of melanoma patients were downloaded from TCGA and correlated with disease progression by bioinformatics analysis and confirmed on patient’s tissues by qRT-PCR and IHC analyses. The effects of combination of BRAF/MEKi and the SCD1 inhibitor MF-438 were monitored by spheroid-forming and proliferation assays on a panel of BRAF-mutated melanoma cell lines grown in 3D and 2D conditions, respectively. SCD1, YAP/TAZ and stemness markers were evaluated in melanoma cells and tissues by qRT-PCR, WB and Immunofluorescence.ResultsWe first observed that SCD1 expression increases during melanoma progression. BRAF-mutated melanoma 3D cultures enriched for CSCs overexpressed SCD1 and were more resistant than 2D differentiated cultures to BRAF and MEK inhibitors. We next showed that exposure of BRAF-mutated melanoma cells to MAPK pathway inhibitors enhanced stemness features by upregulating the expression of YAP/TAZ and downstream genes but surprisingly not SCD1. However, SCD1 pharmacological inhibition was able to downregulate YAP/TAZ and to revert at the same time CSC enrichment and resistance to MAPK inhibitors.ConclusionsOur data underscore the role of SCD1 as prognostic marker in melanoma and promote the use of SCD1 inhibitors in combination with MAPK inhibitors for the control of drug resistance.

A novel Anti-Cancer Stem Cells compound optimized from the natural symplostatin 4 scaffold inhibits Wnt/β-catenin signaling pathway

Cancer stem cells (CSCs) are responsible for carcinogenesis, cancer progression, relapse, metastasis and drug resistance. Therefore, the development of drug molecules targeting CSCs plays a vital role in medicinal researching field. However, there are extremely rare molecules that selectively ablate CSCs. The research and development of drugs targeting CSCs is limited due to a lack of anti-CSCs lead compounds. In this study, an anti-CSCs lead compound 35b was discovered, which was derived from the natural chemical scaffold of Symplostatin 4. This compound exhibited a significantly suppressive effect on tumor growth both in vitro and in vivo. Additionally, 35b could significantly reduce the number of melanoma tumor spheres and decrease the percentage of ALDH+ melanoma cells. Further mechanism study illustrated that compound 35b could eliminate the melanoma CSCs by efficiently blocking Wnt/β-catenin signaling pathway. Collectively, our findings would provide a novel chemical scaffold and alternative idea of molecular design for development of anti-CSCs drugs.

Mesenchymal stem cell's secretome promotes selective enrichment of cancer stem-like cells with specific cytogenetic profile

Cancer stem cells (CSCs) are responsible for tumor initiation, metastasis and cancer recurrence, however the involvement of microenvironment is crucial. Here, we have analyzed how human mesenchymal stem cells (MSCs)-derived conditioned medium (CM) affect colon and melanoma CSCs enrichment and maintenance. Our results strongly suggest that the secretome of CM-MSCs selects and maintains subpopulations with high expression of CSCs markers and ALDH1 activity, low proliferation rates with G1 phase arrest, and notably retain in vivo these properties. Cytogenetic analyses indicated that CM-cultured cells contain alterations in chromosome 17 (17q25). Subsequent SKY-FISH analyses suggested that genes located in 17q25 might be involved in stem-cell maintenance. The characterization of secreted proteins present in CM-MSCs revealed that four cytokines and seven growth factors are directly linked to the CSCs enrichment reported in this study. Further analyses revealed that the combination of just IL6 and HGF is enough to provide cancer cells with better stemness properties. In conclusion, this study demonstrates how specific chromosomal alterations present in CSCs subpopulations might represent an advantage for their in vitro maintenance and in vivo stemness properties.

Targeting melanoma stem cells with the Vitamin E derivative \u03b4-tocotrienol

The prognosis of metastatic melanoma is very poor, due to the development of drug resistance. Cancer stem cells (CSCs) may play a crucial role in this mechanism, contributing to disease relapse. We first characterized CSCs in melanoma cell lines. We observed that A375 (but not BLM) cells are able to form melanospheres and show CSCs traits: expression of the pluripotency markers SOX2 and KLF4, higher invasiveness and tumor formation capability in vivo with respect to parental adherent cells. We also showed that a subpopulation of autofluorescent cells expressing the ABCG2 stem cell marker is present in the A375 spheroid culture. Based on these data, we investigated whether δ-TT might target melanoma CSCs. We demonstrated that melanoma cells escaping the antitumor activity of δ-TT are completely devoid of the ability to form melanospheres. In contrast, cells that escaped vemurafenib treatment show a higher ability to form melanospheres than control cells. δ-TT also induced disaggregation of A375 melanospheres and reduced the spheroidogenic ability of sphere-derived cells, reducing the expression of the ABCG2 marker. These data demonstrate that δ-TT exerts its antitumor activity by targeting the CSC subpopulation of A375 melanoma cells and might represent a novel chemopreventive/therapeutic strategy against melanoma.