Differentiation Of Cancer Stem Cells Research Articles

Epigenomic Profiling of Epithelial Ovarian Cancer Stem-Cell Differentiation Reveals GPD1 Associated Immune Suppressive Microenvironment and Poor Prognosis.

Intraperitoneal metastasis is a challenging clinical scenario in epithelial ovarian cancer (EOC). As they are distinct from hematogenous metastasizing tumors, epithelial ovarian cancer cells primarily disseminate within the peritoneal cavity to form superficially invasive carcinomas. Unfavorable pharmacokinetics for peritoneal tumors and gut toxicity collectively lead to a narrow therapeutic window and therefore limit the opportunities for a favorable clinical outcome. New insights into tumor metastasis in the peritoneal microenvironment are keenly awaited to develop new therapeutic strategies. Epithelial ovarian cancer stem cell (OCSC) seeding is considered to be a critical component of the peritoneal spread. Using a unique and stepwise process of the OCSC differentiation model may provide insight into the intraperitoneal metastasis. The transcriptome and epigenome of OCSC differentiation were characterized by expression array and MethylCap-Seq. The TCGA, AOCS, and KM-Plotter databases were used to evaluate the association between survival outcomes and the methylation/expression levels of candidate genes in the EOC datasets. The STRING database was used to investigate the protein–protein interaction (PPI) for candidates and their associated genes. The infiltration level of immune cells in EOC patients and the association between clinical outcome and OCSCs differentiation genes were estimated using the TIDE and TIME2.0 algorithms. We established an EOC differentiation model using OCSCs. After an integrated transcriptomics and methylomics analysis of OCSCs differentiation, we revealed that the genes associated with earlier OCSC differentiation were better able to reflect the patient’s outcome. The OCSC differentiation genes were involved in regulating metabolism shift and the suppressive immune microenvironment. High GPD1 expression with high pro-tumorigenic immune cells (M2 macrophage, and cancer associated fibroblast) had worst survival. Moreover, we developed a methylation signature, constituted by GNPDA1, GPD1, GRASP, HOXC11, and MSLN, that may be useful for prognostic prediction in EOC. Our results revealed a novel role of epigenetic plasticity OCSC differentiation and suggested metabolic and immune intervention as a new therapeutic strategy.

Histone Deacetylase Inhibitors Impair Glioblastoma Cell Motility and Proliferation.

Simple SummaryGlioblastoma (GBM) is considered the deadliest brain tumor; with patients displaying a high incidence of relapse and a 3-year survival of only 3–5%. For these reasons, investigation of the molecular basis of the disease could provide novel targets for therapy and improve patient prognoses. Based on our previous data, demonstrating that high levels of the transcription factor TCF4 (TCF7L2) sustain the aggressiveness and the stem cell features of these tumors, in this study we tested the ability of the histone deacetylase inhibitors (HDI) Trichostatin-A and Vorinostat to suppress TCF4 levels. We demonstrated that HDI treatment impairs proliferation and viability of GBM cells. Moreover, molecular analysis of HDI effects disclosed their ability to counteract tumor cell motility by affecting the RhoA-GTPase and the interferon pathways, supporting their further characterization as potential anti-GBM agents.Despite being subjected to high-dose chemo and radiotherapy, glioblastoma (GBM) patients still encounter almost inevitable relapse, due to the capability of tumor cells to disseminate and invade normal brain tissues. Moreover, the presence of a cancer stem cell (CSC) subpopulation, already demonstrated to better resist and evade treatments, further frustrates potential therapeutic approaches. In this context, we previously demonstrated that GBM is characterized by a tightly-regulated balance between the β-catenin cofactors TCF1 and TCF4, with high levels of TCF4 responsible for sustaining CSC in these tumors; thus, supporting their aggressive features. Since histone deacetylase inhibitors (HDI) have been reported to strongly reduce TCF4 levels in colon cancer cells, we hypothesized that they could also exert a similar therapeutic action in GBM. Here, we treated primary GBM cultures with Trichostatin-A and Vorinostat, demonstrating their ability to strongly suppress the Wnt-dependent pathways; thus, promoting CSC differentiation and concomitantly impairing GBM cell viability and proliferation. More interestingly, analysis of their molecular effects suggested a prominent HDI action against GBM cell motility/migration, which we demonstrated to rely on the inhibition of the RhoA-GTPase and interferon intracellular cascades. Our results suggest HDI as potential therapeutic agents in GBM, through their action on multiple cancer hallmarks.

Bone morphogenetic protein receptor inhibitors suppress the growth of glioblastoma cells.

Glioblastomas (GBMs) are aggressive brain tumors that are resistant to chemotherapy and radiation. Bone morphogenetic protein (BMP) ligand BMP4 is being examined as a potential therapeutic for GBMs because it induces differentiation of cancer stem cells (CSCs) to an astrocyte phenotype. ID1 is reported to promote self-renewal and inhibit CSC differentiation. In most cancers, ID1 is transcriptionally upregulated by BMP4 promoting invasion and stemness. This conflicting data bring into question whether BMP signaling is growth suppressive or growth promoting in GBMs. We utilized BMP inhibitors DMH1, JL5, and Ym155 to examine the role of BMP signaling on the growth of GBMs. DMH1 targets BMP type 1 receptors whereas JL5 inhibits both the type 1 and type 2 BMP receptors. Ym155 does not bind the BMP receptors but rather inhibits BMP signaling by inducing the degradation of BMPR2. We show that JL5, DMH1, and Ym155 decreased the expression of ID1 in SD2 and U87 cells. JL5 and Ym155 also decreased the expression of BMPR2 and its downstream target inhibitor of apoptosis protein XIAP. JL5 treatment resulted in significant cell death and suppressed self-renewal to a greater extent than that induced by BMP4 ligand. The lysosome inhibitor chloroquine increases the localization of BMPR2 to the plasma membrane enhancing JL5-induced downregulation of ID1 and cell death in SD2 cells. We show that BMP signaling is growth promoting in GBMs. These studies suggest the need for development of BMP inhibitors and evaluation as potential therapeutic for GBMs.

Protein Degradation by E3 Ubiquitin Ligases in Cancer Stem Cells.

Simple SummaryThe aim of this review was to discuss the fundamental role of E3 ubiquitin ligases in controlling cancer stem cells. It will be surmised that protein degradation controlled by the E3 ubiquitin ligases plays a fundamental role in the self-renewal, maintenance and differentiation of cancer stem cells, highlighting its potential as an effective therapeutic target for anticancer drug development.Cancer stem cells are a small subpopulation within the tumor with high capacity for self-renewal, differentiation and reconstitution of tumor heterogeneity. Cancer stem cells are major contributors of tumor initiation, metastasis and therapy resistance in cancer. Emerging evidence indicates that ubiquitination-mediated post-translational modification plays a fundamental role in the maintenance of cancer stem cell characteristics. In this review, we will discuss how protein degradation controlled by the E3 ubiquitin ligases plays a fundamental role in the self-renewal, maintenance and differentiation of cancer stem cells, highlighting the possibility to develop novel therapeutic strategies against E3 ubiquitin ligases targeting CSCs to fight cancer.

The IVF-generated human embryonic microenvironment reverses progestin resistance in endometrial cancer cells by inducing cancer stem cell differentiation.

Progestin resistance is a critical factor that prevents patients with endometrial cancer (EC) from receiving conservative therapy. However, the etiology remains elusive. Cancer stem cells (CSCs) may be a contributing factor to progestin resistance in EC. These cells share similar stemness properties with embryonic stem cells that have a multipotent but differential naïve phenotype. Embryonic stem cells are programed to self-renew, to differentiate and to show plasticity toward a normal cellular phenotype in their defined microenvironment. However, whether this microenvironment may promote CSC differentiation toward a better responsive phenotype and reverse progestin resistance has not yet been clarified. In the current study, we found that progestin resistance of endometrial CSCs can be improved or reversed by using in vitro fertilization (IVF)-generated embryonic sac-derived fluid containing the embryonic microenvironment. Furthermore, suppression or reversal of progestin resistance was mediated by placental alkaline phosphatase (ALPP), a factor secreted into the embryonic microenvironment by IVF-generated blastocysts. ALPP significantly reversed progestin resistance by facilitating endometrial CSC differentiation through downregulating the stemness genes NANOG, OCT4 and SOX2. We further showed that the downregulation of NANOG, OCT4 and SOX2 by ALPP was carried out by TET1/2-mediated epigenetic modulation of the promoter regions of these genes. Such changes at the molecular level initiated endometrial CSC differentiation and promoted a better responsive endometrial cancer phenotype. In fact, their response to progestin treatment was similar to that of well-differentiated endometrioid carcinoma cells without CSCs. ALPP could be a novel target in the process to overcome progestin resistance, and such findings may provide a new approach for the conservative treatment of endometrial cancer.

An active RUNX1-ID1/ID3 axis governs differentiation and chemoresistance of cancer stem cell population in epithelial ovarian cancer cells

Progression, relapse, and therapy resistance are the most challenging features of cancer therapy that have been postulated to be driven by Cancer Stem Cell (CSC) population. This enigmatic subpopulation of cancer cells has therefore emerged as promising therapeutic candidate. We earlier reported enrichment of CSC-like side population (SP) with increasing resistance towards Cisplatin and Paclitaxel either alone or in combination in epithelial ovarian cancer (EOC) cells. This SP population is a small proportion of the total population of cancer cells characterised with high expression of drug transporters, a unique feature of stem cells and thereby can be isolated through their efflux properties of DNA binding dyes. While the bulk non-SP (NSP) population of the cancer cells lack overexpression of the drug transporters and thus can be identified as the dye containing population. In this study, we show that increased expression of Runt related transcription factor 1 (RUNX1) maintains undifferentiated state of CSC-like SP cells through upregulation of inhibitors of DNA binding/differentiation genes (ID1 and ID3) in late cisplatin-paclitaxel resistant cells. Higher RUNX1 expression was found to correlate with decreased median overall survival and disease-free survival in The Cancer Genome Atlas (TCGA) data set of high grade serous ovarian cancer (HGSOC) patients. The protein-protein interaction network analysis of 397 upregulated genes in RUNX1-high samples of TCGA data show significant enrichment of pathways known to negatively regulate CSC differentiation. Intriguingly RUNX1 inhibition not only induces CSC differentiation but also downregulates anti-apoptotic protein BCL2 in both SP and NSP cells and potentiates cytotoxic effects of Cisplatin-Paclitaxel in chemoresistant EOC cells. Inhibition of BCL2 through Venetoclax treatment, a small molecule BH3 mimic, sensitized these cells to platinum taxol treatment. Altogether, our data reveal new regulatory roles by RUNX1 to modulate CSC differentiation via ID1 and ID3 and to promote chemoresistance through BCL2 upregulation.

ERBB Signaling Pathway in Cancer Stem Cells.

The epidermal growth factor receptor (EGFR) was first tyrosine kinase receptor linked to human cancers. EGFR or ERBB1 is a member of ERBB subfamily, which consists of four type I transmembrane receptor tyrosine kinases, ERBB1, 2, 3 and 4. ERBBs form homo/heterodimers after ligand binding except ERBB2 and consequently becomes activated. Different signal pathways, such as phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT), RAS/RAF/MEK/ERK, phospholipase Cγ and JAK-STAT, are triggered by ERBB activation. Since ERBBs, through these pathways, regulate stemness and differentiation of cancer stem cells (CSCs), their roles in CSC tumorigenicity have extensively been investigated. The hyperactivation of ERBBs and its downstream pathways stimulated by either genetic and/or epigenetic factors are frequently described in many types of human cancers. Their dysregulations make cells acquiring CSC characters such as survival, tumorigenicity and stemness. Because of the roles in tumor growth and progress, ERBBs are considered to be one of the drug targets as cancer treatment strategy. In this chapter, we will summarize the structure, function and roles of ERBB subfamily along with their relative pathways regulating the stemness and tumorigenicity of CSCs. Finally, we will discuss the targeting therapy strategies of cancer along with ERBBs in addition to some challenges and future perspectives.

Identifying the E2F3-MEX3A-KLF4 signaling axis that sustains cancer cells in undifferentiated and proliferative state.

Rationale: Dysregulation of signaling that governs self-renewal and differentiation of intestinal stem cells (ISCs) is a major cause of colorectal cancer (CRC) initiation and progression.Methods: qRT-PCR, western blotting, in situ hybridization, immunohistochemistry and immunofluorescence assays were used to detect the expression levels of MEX3A, KLF4 and E2F3 in CRC tissues. The biological functions of MEX3A were studied using Mex3a knockout (KO) and intestinal epithelium specific conditional knockout (cKO) mice, AOM-DSS mouse colorectal tumor model, Apc floxed mouse tumor model and intestinal and tumor organoids. Transcriptomic RNA sequencing (RNA-seq), RNA crosslinking immunoprecipitation (CLIP) and luciferase reporter assays were performed to explore the molecular mechanisms of MEX3A.Results: RNA-binding protein MEX3A, a specific ISC marker gene, becomes ectopically upregulated upon CRC and its levels negatively correlate with patient survival prognosis. MEX3A functions as an oncoprotein that retains cancer cells in undifferentiated and proliferative status and it enhances their radioresistance to DNA damage. Mechanistically, a rate limiting factor of cellular proliferation E2F3 induces MEX3A, which in turn activates WNT pathway by directly suppressing expression of its pro-differentiation transcription factor KLF4. Knockdown of MEX3A with siRNA or addition of KLF4 agonist significantly suppressed tumor growth both by increasing differentiation status of cancer cells and by suppressing their proliferation.Conclusions: It identifies E2F3-MEX3A-KLF4 axis as an essential coordinator of cancer stem cell self-renewal and differentiation, representing a potent new druggable target for cancer differentiation therapy.

Differentiation of cancer stem cells into erythroblasts in the presence of CoCl2

Cancer stem cells (CSCs) are subpopulations in the malignant tumors that show self-renewal and multilineage differentiation into tumor microenvironment components that drive tumor growth and heterogeneity. In previous studies, our group succeeded in producing a CSC model by treating mouse induced pluripotent stem cells. In the current study, we investigated the potential of CSC differentiation into blood cells under chemical hypoxic conditions using CoCl2. CSCs and miPS-LLCcm cells were cultured for 1 to 7 days in the presence of CoCl2, and the expression of VEGFR1/2, Runx1, c-kit, CD31, CD34, and TER-119 was assessed by RT-qPCR, Western blotting and flow cytometry together with Wright-Giemsa staining and immunocytochemistry. CoCl2 induced significant accumulation of HIF-1α changing the morphology of miPS-LLCcm cells while the morphological change was apparently not related to differentiation. The expression of VEGFR2 and CD31 was suppressed while Runx1 expression was upregulated. The population with hematopoietic markers CD34+ and c-kit+ was immunologically detected in the presence of CoCl2. Additionally, high expression of CD34 and, a marker for erythroblasts, TER-119, was observed. Therefore, CSCs were suggested to differentiate into erythroblasts and erythrocytes under hypoxia. This differentiation potential of CSCs could provide new insight into the tumor microenvironment elucidating tumor heterogenicity.

Chronic exposure to FGF2 converts iPSCs into cancer stem cells with an enhanced integrin/focal adhesion/PI3K/AKT axis.

We previously demonstrated the conversion of normal stem cells, including induced pluripotent stem cells (iPSCs), into cancer stem cells (CSCs) without genetic manipulation. Herein, we designed a meta-analysis to assess gene expression profiles in different breast cancer cell lines focusing on the secretory factors responsible for conversion. As a result, fibroblast growth factor 2 (FGF2) was found to be the best candidate in T47D and BT549cells, of which conditioned medium was previously successful in inducing CSCs. When treated with 3.1μg/ml FGF2, mouse iPSCs not only maintained survival without LIF for three weeks but also acquired growth ability independent of FGF2. The resultant cells exhibited expression of stemness and cancer stem cell markers, sphere-forming ability, differentiation, and tumorigenicity with malignancy. The primary cultures of the tumor confirmed the signatures of CSCs with two different phenotypes with or without GFP expression under control of the Nanog promoter. Bioinformatic analysis of gene expression profiles suggested constitutive autocrine activation of the FGF receptor, integrins, focal adhesions, and PI3K/AKT pathways. FGF2 could potently initiate cancer as a component of the inflammatory microenvironment.

Inhibition of anti-cancer stem cells drug salinomycin on Wnt/β-catenin signaling pathway

<p indent=0mm>Cancer stem cells (CSCs) are a small subset of cells able to self-renew, differentiate, and generate heterogenetic cells, which play a vital role in the tumorigenesis, development, recurrence, metastasis and resistance of tumors. Therefore, developing agents targeting CSCs holds promise to eradicate cancer cells in the future, which provides a novel direction for anti-tumor drug research. Recently, a substantial number of experiments have confirmed the killing effect of salinomycin on CSCs in many types of cancers, including liver cancer, breast cancer, gastric cancer, colorectal cancer, prostate cancer, and osteosarcoma. Salinomycin not only inhibits the proliferation, differentiation and metastasis of CSCs, but also induces their apoptosis. The occurrence and development of cancers and CSCs are crucially influenced by the aberrant activation of the Wnt/β-catenin signaling pathway, a highly conservative signaling system in the process of biological evolution closely associated with biological processes such as tissue development and organogenesis. Accordingly, targeting the abnormally activated Wnt signaling pathway could inhibit the proliferation of cancer cells and CSCs, and induce their apoptosis. Wnt/β-catenin signaling pathway plays an essential part in the effect of salinomycin on CSCs. This paper reviews the research progress that salinomycin inhibits CSCs via regulating the Wnt/β-catenin signaling pathway.

Caveolin1: its roles in normal and cancer stem cells.

Stem cells are characterized by the capability of self-renewal and multi-differentiation. Normal stem cells, which are important for tissue repair and tissue regeneration, can be divided into embryonic stem cells (ESCs) and somatic stem cells (SSCs) depending on their origin. As a subpopulation of cells within cancer, cancer stem cells (CSCs) are at the root of therapeutic resistance. Tumor-initiating cells (TICs) are necessary for tumor initiation. Caveolin1 (Cav1), a membrane protein located at the caveolae, participates in cell lipid transport, cell migration, cell proliferation, and cell signal transduction. The purpose of this review was to explore the relationship between Cav1 and stem cells. In ESCs, Cav1 is beneficial for self-renewal, proliferation, and migration. In SSCs, Cav1 exhibits positive or/and negative effects on stem cell self-renewal, differentiation, proliferation, migration, and angiogenic capacity. Cav1 deficiency impairs normal stem cell-based tissue repair. In CSCs, Cav1 inhibits or/and promotes CSC self-renewal, differentiation, invasion, migration, tumorigenicity ability, and CSC formation. And suppressing Cav1 promotes chemo-sensitivity in CSCs and TICs. Cav1 shows dual roles in stem cell biology. Targeting the Cav1-stem cell axis would be a new way for tissue repair and cancer drug resistance.

The high expression of miR-31 in lung adenocarcinoma inhibits the malignancy of lung adenocarcinoma tumor stem cells

Therapies for lung adenocarcinoma (LUAD) are mainly limited by drug resistance, metastasis or recurrence related to cancer stem cells (CSCs) with high proliferation and self-renewing. This research validated that miR-31 was over-expressed in LUAD by the analysis of generous clinical samples data. And the results of clinical data analysis showed that high expression of miR-31 was more common in patients with worse prognosis. The genes differentially expressed in LUAD tissues compared with normal tissues and A549CD133+ cells (LUAD CSCs) compared with A549 cells were separately screened from Gene Expression Profiling Interactive Analysis and GEO datasets. The target genes that may play a role in the regulation of lung adenocarcinoma was screened by comparison between the differential genes and the target genes of miR-31. The functional enrichment analysis of GO Biological Processes showed that the expression of target genes related to cell proliferation was increased, while the expression of target genes related to cell invasion and metastasis was decreased in LUAD tissues and A549CD133+ cells. The results suggested that miR-31 may have a significant inhibitory effect on the differentiation, invasion, metastasis and adhesion of LUAD CSCs, which was verified in vivo and in vitro experiments. Knock down of miR-31 accelerated xenograft tumor growth and liver metastasis in vivo. Likewise, the carcinogenicity, invasion and metastasis of A549CD133+ CSCs were promoted after miR-31 knockdown. The study validated that miR-31 was up regulated in LUAD and its expression may affect the survival time of patients with lung adenocarcinoma, which indicated that miR-31 may have potential value for diagnosis and prognosis of LUAD. However, the inhibitory effect of miR-31 on tumorigenesis, invasion and metastasis of lung adenocarcinoma CSCs suggested its complexity in the regulation of lung adenocarcinoma, which may be related to its extensive regulation of various target genes.

Retinoic Acid-Loaded Dendritic Polyglycerol-Conjugated Gold Nanostars for Targeted Photothermal Therapy in Breast Cancer Stem Cells.

The existence of cancer stem cells (CSCs) poses a major obstacle for the success of current cancer therapies, especially the fact that non-CSCs can spontaneously turn into CSCs, which lead to the failure of the treatment and tumor relapse. Therefore, it is very important to develop effective strategies for the eradication of the CSCs. In this work, we have developed a CSCs-specific targeted, retinoic acid (RA)-loaded gold nanostars-dendritic polyglycerol (GNSs-dPG) nanoplatform for the efficient eradication of CSCs. The nanocomposites possess good biocompatibility and exhibit effective CSCs-specific multivalent targeted capability due to hyaluronic acid (HA) decorated on the multiple attachment sites of the bioinert dendritic polyglycerol (dPG). With the help of CSCs differentiation induced by RA, the self-renewal of breast CSCs and tumor growth were suppressed by the high therapeutic efficacy of photothermal therapy (PTT) in a synergistic inhibitory manner. Moreover, the stemness gene expression and CSC-driven tumorsphere formation were significantly diminished. In addition, the in vivo tumor growth and CSCs were also effectively eliminated, which indicated superior anticancer activity, effective CSCs suppression, and prevention of relapse. Taken together, we developed a CSCs-specific targeted, RA-loaded GNSs-dPG nanoplatform for the targeted eradication of CSCs and for preventing the relapse.

Nanoliposomal Delivery of MicroRNA-203 Suppresses Migration of Triple-Negative Breast Cancer through Distinct Target Suppression.

Triple-negative breast cancers affect thousands of women in the United States and disproportionately drive mortality from breast cancer. MicroRNAs are small, non-coding RNAs that negatively regulate gene expression post-transcriptionally by inhibiting target mRNA translation or by promoting mRNA degradation. We have identified that miRNA-203, silenced by epithelial–mesenchymal transition (EMT), is a tumor suppressor and can promote differentiation of breast cancer stem cells. In this study, we tested the ability of liposomal delivery of miR-203 to reverse aspects of breast cancer pathogenesis using breast cancer and EMT cell lines. We show that translationally relevant methods for increasing miR-203 abundance within a target tissue affects cellular properties associated with cancer progression. While stable miR-203 expression suppresses LASP1 and survivin, nanoliposomal delivery suppresses BMI1, indicating that suppression of distinct mRNA target profiles can lead to loss of cancer cell migration.

Human prostate epithelial cells and prostate-derived stem cells malignantly transformed in vitro with sodium arsenite show impaired Toll like receptor -3 (TLR3)-associated anti-tumor pathway

A therapeutic strategy for prostate cancer (PCa) involves the use of 9-cis-retinoic acid (9cRA) to induce cancer stem cells (CSCs) differentiation and apoptosis. Polyinosinic:polycytidylic acid (PIC) is a Toll-like receptor 3 (TLR3) agonist that induces tumor cells apoptosis after activation. PIC+9cRA combination activates retinoic acid receptor β (RARβ) re-expression, leading to CSC differentiation and growth arrest. Since inorganic arsenic (iAs) targets prostatic stem cells (SCs), we hypothesized that arsenic-transformed SCs (As-CSCs) show an impaired TLR3-associated anti-tumor pathway and, therefore, are unresponsive to PIC activation. We evaluated TLR3-mediated activation of anti-tumor pathway based in RARβ expression, on As-CSC and iAs-transformed epithelial cells (CAsE-PE). As-CSCs and CAsE-PE showed lower TLR3 and RARβ basal expression compared to their respective isogenic controls WPE-Stem and RWPE-1. Also, iAs transformants showed reduced expression of mediators in TLR3 pathway. Importantly, As-CSCs were irresponsive to PIC+9cRA in terms of increased RARβ and decreased SC-markers expression, while CAsE-PE, a heterogeneous cell line having a small SC population, were partially responsive. These observations indicate that iAs can impair TLR3 expression and anti-tumor pathway activated by PIC+9cRA in SCs and prostatic epithelial cells. These findings suggest that TLR3-activation based therapy may be an ineffective therapeutic alternative for iAs-associated PCa.

Abstract 3090: Targeting WNT and retinoid signaling pathways to promote differentiation of cancer stem cells along the neuroendocrine lineage in CRC

Abstract Since APC mutations lead to overpopulation of ALDH+ stem cells (SCs) in colorectal cancers (CRCs) and that ALDH is a key component of the retinoic acid (RA) signaling pathway, the hypothesis was developed that human CRC evolves due to an imbalance between WNT and RA signaling. Indeed, we previously discovered that progressive overpopulation of ALDH+ SCs occurs in normal, mutant, adenomatous, and malignant colon crypt tissues, which coincided with the change in the APC mutation zygosity state in familial adenomatous polyposis (FAP) patients. Therefore, the SC overpopulation that drives development of CRC appears to result from dysregulated WNT signaling, of which APC is a negative regulator. Normal colonic SC differentiation is feedback regulated by neuroendocrine cells (NECs) that reside adjacent to SCs in the bottom of the colon crypt. Our goal was to determine if APC mutation and subsequent dysregulated WNT signaling contributes to the inability of SCs to differentiate into NECs, leading to overpopulation of cancer SCs (CSCs) during tumor development. Although RA signaling is altered in CRC, we still found that all- trans-retinoic acid (ATRA) could reduce the number of ALDH+ CSCs in several CRC cell lines. We also found that administering ATRA promotes differentiation of CSCs along the neuroendocrine lineage. Moreover, induction of wild-type (wt)-APC expression in HT29 CRC cells led to increased expression of several NEC markers including CGA, GLP2R, NSE, SSTR1; and decreased WNT/β-catenin signaling when tested via TCF reporter assay. Inducing wt-APC also led to reduced β-catenin and change in protein expression of several proteins known to be targeted by WNT signaling including c-MYC, c-JUN, MET, and CD44. Our preliminary results also show that inducing wt-APC expression increases the number of GLPR2+ NECs. Thus, a link between WNT and RA signaling provides a possible mechanism that might explain how mutant APC leads to decreased maturation of ALDH+ SCs along the NEC lineage and an increased the number of ALDH+ SCs. Therefore, discovering ways to increase differentiation of SCs along the NEC lineage in vivo might lead to new more effective treatment strategies for CRC patients. Citation Format: Caroline O. Facey, Victoria O. Hunsu, Brian T. Osmond, Lynn M. Opdenaker, Bruce M. Boman. Targeting WNT and retinoid signaling pathways to promote differentiation of cancer stem cells along the neuroendocrine lineage in CRC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3090.

Abstract 1071: Studying the ability of retinoids to inhibit growth of CRC cells based on their retinoid pathway genotype

Abstract Colorectal cancer (CRC) tumorigenesis arises through acquisition of mutations in a colonic stem cell (SC) that transforms it into a cancer stem cell (CSC). There are no curative treatments for advanced CRC because CSCs appear to be resistant to chemotherapy and/or radiation. Because all-trans retinoic acid (ATRA) is a highly effective and often curative treatment for acute promyelocytic leukemia patients, we conjecture that retinoids may be an efficacious treatment for CRC. Retinoic acid (RA) is a metabolite of vitamin A which is known to induce differentiation of SCs and inhibit cell proliferation. Our goal is to discover how to therapeutically target human CRC CSCs using the differentiation-inducing effects of retinoid agents. Our previous study on the RA metabolizing enzyme aldehyde dehydrogenase (ALDH) shows that ALDH marks normal and malignant colonic SCs and that RA signaling mainly occurs through colonic ALDH+ SCs. Hypothesis: The ability of retinoid agents, ATRA, 9-cis retinoic acid, 13-cis retinoic acid and Liarozole, to induce differentiation of CSCs depends on the RA pathway genotype according to mutations in RA signaling components in CRC cells. Indeed, our bioinformatics analysis shows that many RA pathway genes are overexpressed and often mutated in CRC. We have also evaluated the ability of the different RA ligands to inhibit cell growth and induce differentiation using a panel of CRC cell lines that have specific RA signaling mutations. Further investigation of RA signaling mechanisms that regulate colon SCs and how dysregulation contributes to the SC overpopulation that drives CRC growth should provide insight into strategies for designing new SC-targeted therapies for CRC. Citation Format: Victoria O. Hunsu, Caroline O. Facey, Lynn M. Opdenaker, Bruce M. Boman. Studying the ability of retinoids to inhibit growth of CRC cells based on their retinoid pathway genotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1071.

Exosomal lncRNA UCA1 modulates cervical cancer stem cell self-renewal and differentiation through microRNA-122-5p/SOX2 axis

BackgroundThere is growing evidence discussing the role of long non-coding RNAs (lncRNAs) in cervical cancer (CC). We performed this study to explore the impact of exosomal lncRNA urothelial cancer-associated 1 (UCA1) in CC stem cells by sponging microRNA-122-5p (miR-122-5p) and regulating SOX2 expression.MethodsCC stem cells (CD133+CaSki) and exosomes were extracted and identified. The synthesized UCA1- and miR-122-5p-related sequences were transfected into CaSki cells, CaSki cells-derived exosomes were extracted and then co-cultured with CD133+CaSki cells. The functional roles of UCA1 and miR-122-5p in self-renewal and differentiation ability of CC stem cells were determined using ectopic expression, knockdown/depletion and reporter assay experiments. An in vivo experiment was performed to verify the in vitro results.ResultsUp-regulated UCA1 and SOX2 and down-regulated miR-122-5p were found in CaSki-Exo. Exosomes promoted invasion, migration, proliferation and restrained apoptosis of CD133+CaSki cells. Silencing UCA1 or up-regulating miR-122-5p degraded SOX2 expression, and reduced invasion, migration and proliferation of CD133+CaSki cells while advanced apoptosis and suppressed the tumor volume and weight in nude mice.ConclusionOur study provides evidence that CaSki-Exo can promote the self-renewal and differentiation ability of CC stem cells while silencing UCA1 or up-regulating miR-122-5p restrains self-renewal and differentiation of CC stem cells.

Natural products targeting cancer stem cells: Implications for cancer chemoprevention and therapeutics.

Cancer, being the leading cause of death in the globe, has been one of the major thrust areas of research worldwide. In a new paradigm about neoplastic transformations, the initiation and recurrence of disease is attributed to few mutated cells in bulk of tumor called cancer stem cells (CSCs). CSCs have capacity of self-renewal and differentiation, which are known for resistance to radio and chemotherapy leading to recurrence of the disease even after treatment. Most of traditional drugs implicated in cancer therapy targeting primary tumors have substantial toxicity to the physiological system and have not been efficient in targeting these CSCs leading to poor prognosis. Targeting these CSCs in bulk of tumor might be novel strategy for cancer chemoprevention and therapeutics. Diet-derived interventions and diverse natural products are known to target these CSCs and related signaling pathways, namely, Wnt, Notch, and Hedgehog pathways, which are implicated for CSC self-renewal. PRACTICAL APPLICATIONS: Cancer remains a global challenge even in this century. Poor prognosis, survival rate, and recurrence of the disease have been the major concerns in traditional cancer therapy regimes. Targeting cancer stem cells might be novel strategy for elimination and cure of the chronic disease as they are known to modulate all stages of carcinogenesis and responsible for recurrence and resistance to chemotherapy and radiotherapy. The evidence support that natural products might inhibit, delay, or reverse the process of tumorigenesis and modulate the different signaling pathways implicated for cancer stem cells self-renewal and differentiation. Natural products have minimal toxicity compared to traditional cancer therapy drugs since they have long been utilized in our food habits without any major side effects reported. Thus, targeting cancer stem cells with natural product might be a novel strategy for drug development in cancer chemoprevention and therapeutics.