Non-stem Cancer Cells Research Articles

Abstract 372: Macrophage contact-dependent stemness induction and progressive CSC enrichment during metastatic dissemination in breast cancer

Abstract Background: Cancer stem cells (CSCs) play an important role during metastatic progression of breast cancer. However, little is known, at the single cell level, about the process of stemness induction in non-stem cells or the dynamic behavior of CSCs during hematogenous dissemination. Methods: Here, we employed high-resolution intravital multiphoton microscopy with a SOX2/OCT4 responsive fluorescent biosensor for stemness to directly observe the induction of stemness in single non-stem cells and their evolution through the metastatic cascade in living animals using orthotopic breast cancer xenograft model. We confirmed our findings in vitro using tumor cell-macrophage co-culture assays. Results: We report that, both in vitro and in vivo, direct physical contact with macrophages induces stemness in non-stem cancer cells via juxtacrine Notch-Jagged1 signaling. In vivo, macrophage depletion with clodronate treatment showed a significant decrease in stem cells. In vitro, using either the fate mapping of non-stem cells with or without macrophage contact, or the origin-mapping of stem cells to find whether they originated from non-stem cells or pre-existing stem cells, we found that there was four-fold increase in new CSC induction after direct macrophage contact. In contrast, we did not see any role of macrophages in the expansion of pre-existing CSCs, both in vivo and in vitro, indicating that macrophage contact-dependent stem induction is the primary mechanism of CSC generation. Using immunohistochemical staining in fixed tissue and live imaging of primary tumors and lungs in mice using optical windows, we found that during the course of dissemination of tumor cells from the primary site, CSCs become progressively enriched in the tumor cell population as they approach dissemination doorways (known as TMEM, Tumor MicroEnvironment of Metastasis), intravasate, circulate and arrive at the lung. Association with and passage through TMEM doorways is the step that generates the greatest enrichment in CSCs (~ 60-fold). On arrival in the lung, CSCs represent more than 75% of the disseminated tumor cell population, greatly enriched compared with their representation in the bulk primary tumor of ~ 1%. Conclusion: Overall, these data indicate, for the first time, that macrophages associated with TMEM induce CSCs and promote TMEM-mediated CSC intravasation and early metastatic seeding. Our results are consistent with the dramatic enrichment of cancer stem cell markers in association with TMEM in breast cancer patients (Kim et al 2020 AACR abstract) and support a strategy for anti-metastatic therapy. Citation Format: Ved P. Sharma, Yarong Wang, Binwu Tang, George S. Karagiannis, Emily A. Xue, David Entenberg, Lucia Borriello, Anouchka Coste, Joan G. Jones, Chinmay R. Surve, Dominic Esposito, Maja H. Oktay, Lalage M. Wakefield, John S. Condeelis. Macrophage contact-dependent stemness induction and progressive CSC enrichment during metastatic dissemination in breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 372.

Broccoli extract increases drug-mediated cytotoxicity towards cancer stem cells of head and neck squamous cell carcinoma

BackgroundHead and neck squamous cell carcinomas (HNSCC) are malignant neoplasms with poor prognosis. Treatment-resistant cancer stem cell (CSC) is one reason for treatment failure. Considerable attention has been focused on sulforaphane (SF), a phytochemical from broccoli possessing anticancer properties. We investigated whether SF could enhance the chemotherapeutic effects of cisplatin (CIS) and 5-fluorouracil (5-FU) against HNSCC–CSCs, and its mechanisms of action.MethodsCD44+/CD271+ FACS-isolated CSCs from SCC12 and SCC38 human cell lines were treated with SF alone or combined with CIS or 5-FU. Cell viability, colony- and sphere-forming ability, apoptosis, CSC-related gene and protein expression and in vivo tumour growth were assessed. Safety of SF was tested on non-cancerous stem cells and in vivo.ResultsSF reduced HNSCC–CSC viability in a time- and dose-dependent manner. Combining SF increased the cytotoxicity of CIS twofold and 5-FU tenfold, with no effects on non-cancerous stem cell viability and functions. SF-combined treatments inhibited CSC colony and sphere formation, and tumour progression in vivo. Potential mechanisms of action included the stimulation of caspase-dependent apoptotic pathway, inhibition of SHH pathway and decreased expression of SOX2 and OCT4.ConclusionsCombining SF allowed lower doses of CIS or 5-FU while enhancing these drug cytotoxicities against HNSCC–CSCs, with minimal effects on healthy cells.

Glycosphingolipid expression at breast cancer stem cells after novel thieno[2,3-b]pyridine anticancer compound treatment

Glycosphingolipid expression differs between human breast cancer stem cells (CSC) and cancer non-stem cells (non-CSC). We performed studies of viability, type of cell death, cancer stem cell percent and glycosphingolipid expression on CSC and non-CSC after treatment of MDA-MB-231 and MDA-MB-453 triple-negative breast cancer cells with a newly developed thienopyridine anticancer compound (3-amino-N-(3-chloro-2-methylphenyl)-5-oxo-5,6,7,8-tetrahydrothieno[2,3-b]quinoline-2-carboxamide, 1). Compound 1 was cytotoxic for both breast cancer cell lines and the majority of cells died by treatment-induced apoptosis. The percent of cancer stem cells and number of formed mammospheres was significantly lower. Glycosphingolipids IV6Neu5Ac-nLc4Cer and GalNAc-GM1b (IV3Neu5Ac-Gg5Cer) not reported previously, were identified in both CSCs and non-CSCs. IV6Neu5Ac-nLc4Cer had increased expression in both CSCs and non-CSCs of both cell lines after the treatment with 1, while GM3 (II3Neu5Ac-LacCer) had increased expression only on both cell subpopulations in MDA-MB-231 cell line. GalNAc-GM1b, Gb4Cer (GalNAcβ1-3Galα1-4Galβ1-4Glcβ1-1Cer) and GM2 (II3Neu5Ac-GalNAcβ1-4Galβ1-4Glcβ1-1Cer) were increased only in CSCs of both cell lines while GD3 was decreased in CSC of MDA-MB-231 cell line. Due to its effect in reducing the percentage of cancer stem cells and number of mammospheres, and its influence upon several glycosphingolipid expressions, it can be concluded that compound 1 deserves attention as a potential new drug for triple-negative breast cancer therapy.

Chaetocin Abrogates the Self-Renewal of Bladder Cancer Stem Cells via the Suppression of the KMT1A-GATA3-STAT3 Circuit.

Bladder cancer stem cells (BCSCs) have the abilities of self-renewal, differentiation, and metastasis; confer drug resistance; and exhibit high tumorigenicity. We previously identified that the KMT1A–GATA3–STAT3 axis drives the self-renewal of BCSCs. However, the therapeutic effect of targeting KMT1A in BCSCs remains unknown. In this study, we confirmed that the expression of KMT1A was remarkably higher in BCSCs (3–5-fold) than those in bladder cancer non-stem cells or normal bladder epithelial cells. Among the six KMT1A inhibitors, chaetocin significantly suppressed the cell propagation (inhibition ratio: 65%–88%, IC50 = 24.4–32.5 nM), induced apoptosis (2–5-fold), and caused G1 phase cell cycle arrest (68.9 vs 55.5%) of bladder cancer (BC) cells, without influencing normal bladder epithelial cells. More importantly, chaetocin abrogated the self-renewal of BCSCs (inhibition ratio: 80.1%) via the suppression of the KMT1A–GATA3–STAT3 circuit and other stemness-related pathways. Finally, intravesical instillation of chaetocin remarkably inhibited the growth of xenograft tumors (inhibition ratio: 71–82%) and prolonged the survival of tumor-bearing mice (70 vs 53 days). In sum, chaetocin abrogated the stemness maintenance and tumor growth of BCSCs via the suppression of the KMT1A–GATA3–STAT3 circuit. Chaetocin is an effective inhibitor targeting KMT1A in BCSCs and could be a promising therapeutic strategy for BC.

Abstract B04: Using PSA dynamics to forecast individual responses to intermittent androgen deprivation

Abstract Intermittent androgen-deprivation therapy (IADT) is an attractive treatment approach for biochemically recurrent prostate cancer (PCa), whereby cycling treatment on and off has been shown to limit toxicities and reduce cumulative dose. While IADT has been shown to delay the development of treatment resistance, underlying mechanisms and actionable biomarkers are required to accurately predict when resistance will emerge and how to maximally delay time to progression. We developed a quantitative framework to simulate the enrichment of prostate cancer stem cell dynamics during treatment as a plausible mechanism of resistance evolution. Using simulated dynamics of cancer stem cells and nonstem cancer cells, we derive longitudinal blood serum prostate-specific antigen (PSA) concentrations that were calibrated to and validated with data of 70 patients undergoing multiple cycles of IADT. Model analysis suggests that cancer stem cell proliferation patterns correlate with PSA dynamics and patient outcomes. Learning these dynamics adaptively from the earlier treatment cycles in individual patients can predict the evolution of resistance in subsequent IADT cycles with a predictive power of 90%, which warrants prospective evaluation in a clinical setting. The model is used to simulate and predict alternative treatment options. In particular, simulations show that response dynamics from the first IADT cycle can be used to identify patients who would benefit from concurrent docetaxel. Citation Format: Renee Brady, Heiko Enderling. Using PSA dynamics to forecast individual responses to intermittent androgen deprivation [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B04.

Targeting Tumorigenicity of Breast Cancer Stem Cells Using SAHA/Wnt-b Catenin Antagonist Loaded Onto Protein Corona of Gold Nanoparticles.

BackgroundAmong various theories for the origin of cancer, the “stemness phenotype model” suggests a dynamic feature for tumor cells in which non-cancer stem cells (non-CSCs) can inter-convert to CSCs. Differentiation with histone-deacetylase inhibitor, vorinostat (SAHA), can induce stem cells to differentiate as well as enforces non-CSCs to reprogram to CSCs. To avoid this undesirable effect, one can block the Wnt-βcatenin pathway. Thus, a dual delivery system of SAHA and a Wnt-βcatenin blocker will be beneficial in the induction of differentiation of CSCs. Protein corona (PC) formation in nanoparticle has a biologic milieu, and despite all problematic properties, it can be employed as a medium for dual loading of the drugs.Materials and MethodsWe prepared sphere gold nanoparticles (GNPs) with human plasma protein corona loaded with SAHA as differentiating agent and PKF118-310 (PKF) as a Wnt-βcatenin antagonist. The MCF7 breast cancer stem cells were treated with NPs and the viability and differentiation were evaluated by Western blotting and sphere formation assay.ResultsWe found that both drugs loaded onto corona-capped GNPs had significant cytotoxicity in comparison to bare GNP-corona. Data demonstrated an increase in stem cell population and upregulation of mesenchymal marker, Snail by SAHA-loaded GNPs treatment; however, the combination of PKF loaded GNPs along with SAHA-loaded GNPs resulted in a reduction of stem cell populations and Snail marker. We have shown that in MCF7 and its CSCs simultaneous treatment with SAHA and PKF118-310 induced differentiation and inhibition of Snail induction.ConclusionOur study reveals the PC-coated GNPs as a biocompatible career for both hydrophilic (PKF) and hydrophobic (SAHA) agents which can decrease breast cancer stem cell populations along with reduced stemness state regression.

Blood and Cancer: Cancer Stem Cells as Origin of Hematopoietic Cells in Solid Tumor Microenvironments.

The concepts of hematopoiesis and the generation of blood and immune cells from hematopoietic stem cells are some steady concepts in the field of hematology. However, the knowledge of hematopoietic cells arising from solid tumor cancer stem cells is novel. In the solid tumor microenvironment, hematopoietic cells play pivotal roles in tumor growth and progression. Recent studies have reported that solid tumor cancer cells or cancer stem cells could differentiate into hematopoietic cells. Here, we discuss efforts and research that focused on the presence of hematopoietic cells in tumor microenvironments. We also discuss hematopoiesis from solid tumor cancer stem cells and clarify the notion of differentiation of solid tumor cancer stem cells into non-cancer hematopoietic stem cells.

C-KIT regulates stability of cancer stemness in CD44-positive colorectal cancer cells

Cancer stem cells (CSCs) are subpopulations of cancer cells with high self-renewal potential that are involved in tumor progression and recurrence. It has been postulated that CSCs and non-stem cancer cells are inter-convertible. However, precise mechanisms for the plasticity and stability of cancer stemness remain elusive. Here, we demonstrate that CD44-positive colorectal CSC fractions contain two types of cancer cells: “CD44-stable” cells, in which CD44 expression is stably sustained, and “CD44-trasnsient” cells, which are rapidly converted to CD44-negative cells. CD44-stable cells expressed higher levels of c-KIT tyrosine kinase than CD44-transient cells. c-KIT knockdown by siRNAs converted the CD44-positive cells to CD44-negative cells, which expressed lower levels of stem cell markers such as ASCL2 and EPCAM. In the CD44-positive cells, c-KIT phosphorylation level was very low whereas stem cell factor, a c-KIT ligand, elevated c-KIT phosphorylation without affecting stem cell marker expression. CRISPR-Cas9-mediated knockout of the c-KIT gene in CD44 stable cells attenuated the CSC properties including expression of CD44 and other stem cell markers, clonogenicity and in vivo tumorigenic potential in a mouse xenograft model. These observations suggest that the colorectal CSC fractions contain cancer cells with differential plasticity, which is determined by c-KIT.

Molecular Chaperones in Cancer Stem Cells: Determinants of Stemness and Potential Targets for Antitumor Therapy.

Cancer stem cells (CSCs) are a great challenge in the fight against cancer because these self-renewing tumorigenic cell fractions are thought to be responsible for metastasis dissemination and cases of tumor recurrence. In comparison with non-stem cancer cells, CSCs are known to be more resistant to chemotherapy, radiotherapy, and immunotherapy. Elucidation of mechanisms and factors that promote the emergence and existence of CSCs and their high resistance to cytotoxic treatments would help to develop effective CSC-targeting therapeutics. The present review is dedicated to the implication of molecular chaperones (protein regulators of polypeptide chain folding) in both the formation/maintenance of the CSC phenotype and cytoprotective machinery allowing CSCs to survive after drug or radiation exposure and evade immune attack. The major cellular chaperones, namely heat shock proteins (HSP90, HSP70, HSP40, HSP27), glucose-regulated proteins (GRP94, GRP78, GRP75), tumor necrosis factor receptor-associated protein 1 (TRAP1), peptidyl-prolyl isomerases, protein disulfide isomerases, calreticulin, and also a transcription heat shock factor 1 (HSF1) initiating HSP gene expression are here considered as determinants of the cancer cell stemness and potential targets for a therapeutic attack on CSCs. Various approaches and agents are discussed that may be used for inhibiting the chaperone-dependent development/manifestations of cancer cell stemness.

Sphingosine Kinase 1 Regulates the Survival of Breast Cancer Stem Cells and Non-stem Breast Cancer Cells by Suppression of STAT1.

Cancer stem cells (CSCs) represent rare tumor cell populations capable of self-renewal, differentiation, and tumor initiation and are highly resistant to chemotherapy and radiotherapy. Thus, therapeutic approaches that can effectively target CSCs and tumor cells could be the key to efficient tumor treatment. In this study, we explored the function of SPHK1 in breast CSCs and non-CSCs. We showed that RNAi-mediated knockdown of SPHK1 inhibited cell proliferation and induced apoptosis in both breast CSCs and non-CSCs, while ectopic expression of SPHK1 enhanced breast CSC survival and mammosphere forming efficiency. We identified STAT1 and IFN signaling as key regulatory targets of SPHK1 and demonstrated that an important mechanism by which SPHK1 promotes cancer cell survival is through the suppression of STAT1. We further demonstrated that SPHK1 inhibitors, FTY720 and PF543, synergized with doxorubicin in targeting both breast CSCs and non-CSCs. In conclusion, we provide important evidence that SPHK1 is a key regulator of cell survival and proliferation in breast CSCs and non-CSCs and is an attractive target for the design of future therapies.

Glioblastoma Multiforme Stem Cell Cycle Arrest by Alkylaminophenol Through the Modulation of EGFR and CSC Signaling Pathways.

Cancer stem cells (CSCs), a small subpopulation of cells existing in the tumor microenvironment promoting cell proliferation and growth. Targeting the stemness of the CSC population would offer a vital therapeutic opportunity. 3,4-Dihydroquinolin-1(2H)-yl)(p-tolyl)methyl)phenol (THTMP), a small synthetic phenol compound, is proposed to play a significant role in controlling the CSC proliferation and survival. We assessed the potential therapeutic effects of THTMP on glioblastoma multiforme (GBM) and its underlying mechanism in various signaling pathways. To fully comprehend the effect of THTMP on the CSCs, CD133+ GBM stem cell (GSC) and CD133- GBM Non-stem cancer cells (NSCC) population from LN229 and SNB19 cell lines was used. Cell cycle arrest, apoptosis assay and transcriptome analysis were performed for individual cell population. THTMP strongly inhibited NSCC and in a subtle way for GSC in a time-dependent manner and inhibit the resistance variants better than that of temozolomide (TMZ). THTMP arrest the CSC cell population at both G1/S and G2/M phase and induce ROS-mediated apoptosis. Gene expression profiling characterize THTMP as an inhibitor of the p53 signaling pathway causing DNA damage and cell cycle arrest in CSC population. We show that the THTMP majorly affects the EGFR and CSC signaling pathways. Specifically, modulation of key genes involved in Wnt, Notch and Hedgehog, revealed the significant role of THTMP in disrupting the CSCs’ stemness and functions. Moreover, THTMP inhibited cell growth, proliferation and metastasis of multiple mesenchymal patient-tissue derived GBM-cell lines. THTMP arrests GBM stem cell cycle through the modulation of EGFR and CSC signaling pathways.

Cancer stem cell-specific expression profiles reveal emerging bladder cancer biomarkers and identify circRNA_103809 as an important regulator in bladder cancer.

Bladder cancer stem cells (BCSCs), exhibiting self-renewal and differentiation capacities, may contribute to the tumor initiation, metastasis, recurrence and drug resistance of bladder cancer. However, the underlying functional mechanisms of BCSCs remain to be clarified. In this study, we describe the differentially-expressed mRNAs, lncRNAs, and circRNAs in BCSCs compared with that in bladder cancer non-stem cells (BCNSCs) through the transcriptome microarray data analysis using bladder cancer patients’ specimens. CircRNA_103809, the top one among the highly expressed circRNA identified in BCSCs, promotes the self-renewal, migration and invasion capabilities of bladder cancer by acting as a miR-511 sponge. Additionally, GO and KEGG pathway analysis suggest the differentially expressed genes identified may be involved in the cellular metabolism, differentiation and metastasis regulation of the cancer cells. Co-expression networks of lncRNAs/mRNAs and circRNAs/mRNAs constructed by WGCNA give a picture of the non-coding/coding RNAs regulating patterns in BCSCs. Notably, as core genes in the networks, AHCY, C6orf136 and LRIG1 show high potential to be prognosticators for bladder cancer. Therefore, further studies of non-coding RNA functional mechanisms in BCSCs is valuable for detecting the pathogenic mechanisms and discovering novel biomarkers in bladder cancer.

Transcompp: understanding phenotypic plasticity by estimating Markov transition rates for cell state transitions.

Gradual population-level changes in tissues can be driven by stochastic plasticity, meaning rare stochastic transitions of single-cell phenotype. Quantifying the rates of these stochastic transitions requires time-intensive experiments, and analysis is generally confounded by simultaneous bidirectional transitions and asymmetric proliferation kinetics. To quantify cellular plasticity, we developed Transcompp (Transition Rate ANalysis of Single Cells to Observe and Measure Phenotypic Plasticity), a Markov modeling algorithm that uses optimization and resampling to compute best-fit rates and statistical intervals for stochastic cell-state transitions. We applied Transcompp to time-series datasets in which purified subpopulations of stem-like or non-stem cancer cells were exposed to various cell culture environments, and allowed to re-equilibrate spontaneously over time. Results revealed that commonly used cell culture reagents hydrocortisone and cholera toxin shifted the cell population equilibrium toward stem-like or non-stem states, respectively, in the basal-like breast cancer cell line MCF10CA1a. In addition, applying Transcompp to patient-derived cells showed that transition rates computed from short-term experiments could predict long-term trajectories and equilibrium convergence of the cultured cell population. Freely available for download at http://github.com/nsuhasj/Transcompp. Supplementary data are available at Bioinformatics online.

The effects of hyperthermia on human hepatocellular carcinoma stem and mature cancer cells

Introduction and objectivesHepatocellular carcinoma (HCC) can recur following radiofrequency ablation and other hyperthermic treatment modalities. Cancer stem cells (CSCs) are a subpopulation of HCC cells that are difficult to eradicate and largely responsible for tumor recurrences. Thus, the principal objective of this study was to determine whether human HCC CSCs are relatively thermal-resistant compared to non-stem or mature cancer cells (MCCs). Materials and methodsEpithelial cell adhesion molecule (EpCAM) positive enriched CSCs and EpCAM− MCCs were derived from a human HCC cell line using fluorescence activated cell sorting. Each cell population was exposed to 65°C heat for 0–16min and survival documented at various time points. ResultsCell survival curves were similar in CSC and MCCs throughout the 16min heat exposure period. Maximum killing was obtained after 12–14min of heat exposure. Cytoprotective, heat shock proteins-70 (HSP70) and -90 (HSP90) mRNA expression were not disproportionately increased in CSCs. ConclusionsThese results suggest that human HCC CSCs are not more thermal resistant than MCCs and therefore, do not support the hypothesis that HCC recurrences following hyperthermic treatment reflect CSC thermal-resistance.

Immune vulnerability of ovarian cancer stem-like cells due to low CD47 expression is protected by surrounding bulk tumor cells

ABSTRACT Recurrence of advanced epithelial ovarian cancer is common despite optimal surgical debulking and initial favorable responses to chemotherapy. Evidences suggest that cancer stem cells (CSCs) have inherent resistance to conventional therapies such as chemotherapy and play a decisive role in cancer recurrence. Cancer stem cells are also believed to be able to evade immunological attack. However, this study showed a different scenario in which cancer stem-like cells are more vulnerable to immunosurveillance. Our study demonstrated that isolated murine cancer stem-like cells, stem cell antigen (SCA)-1+ ID8 and CD133+ HM-1 cells, were susceptible to phagocytosis by macrophages and consequent CD8+ T cell immunity. The increased phagocytosis of these stem cell-like cells is attributed to low CD47 protein expression. SCA-1+ ID8 cells were able to grow in syngeneic mice but were soon rejected. Restoring CD47 expression delayed this immune-mediated rejection. SCA-1+ ID8 cells showed rapid growth by mixing with bulk ID8 cells. These results suggest that stem-like cells could be protected by surrounding non-stem cancer cells from immune attack. Similarly, both isolated human CD24−/low SKOV3 stem-like cells and spheroid OVCAR3 cells expressed lower CD47 levels. Our study provided novel insights into the immune characteristics of CSCs within a tumor microenvironment. The results might lead to the design of more effective treatment strategies for ovarian cancer.

Chaetocin Abrogates the Self-Renewal of Bladder Cancer Stem Cells <i>via</i> the Suppression of KMT1A-GATA3-STAT3 Circuit

Background: Bladder cancer stem cells (BCSCs) have the abilities of self-renewal, differentiation, metastasis, conferring drug resistance and exhibiting high tumorigenicity. KMT1A-GATA3-STAT3 axis was proved to maintain the stemness of BCSCs, in which KMT1A was highly expressed in BCSCs rather than bladder cancer non-stem cells (BCNSCs) and normal bladder epithelial cells. However, the therapeutic effect of targeting KMT1A in bladder cancer (BC) or BCSCs remains unknown. Methods: In this study, we confirmed that the expression of KMT1A was remarkably higher in BCSCs than those in BCNSCs. Six histone methyltransferase inhibitors AMI-1, BIX-01294, chaetocin, DZNeP, GSK343 and UNC0631 were investigated targeting KMT1A through cell proliferation assay. Findings: Among those, chaetocin suppressed the cell propagation, induced apoptosis and caused G1 phase cell cycle arrest of BC cells, without influencing normal bladder epithelial cells. More importantly, chaetocin abrogated the self-renewal of BCSCs via the suppression of KMT1A-GATA3-STAT3 circuit and other stemness-related pathways. Finally, intravesical instillation of chaetocin inhibited the formation of BCSCs tumor spheres and xenograft tumors. Chaetocin is an effective histone methyltransferase inhibitor targeting KMT1A in BCSCs. Furthermore, chaetocin abrogated the stemness maintenance and tumor growth of BCSCs via the suppression of KMT1A-GATA3-STAT3 circuit. In future, chaetocin can be a potential therapeutic strategy for BC. Funding Statement: This work was supported by the Basic Scientific Research Operating Expenses for Central Universities (No. buctrc201910), National Natural Science Foundation of China (81602644), the project of Basic Research Cooperation of Beijing-Tianjin-Hebei (H2019104018) and the grant from the Ministry of Science and Technology of China (2010ZX09401-403). Declaration of Interests: The authors have declared that no competing interest exists. Ethics Approval Statement: Human tissues were obtained from The Second Affiliated Hospital of Kunming Medical University College (Kunming, China) with informed consent and approved by the Research Ethics Board at The Second Affiliated Hospital of Kunming Medical University. The mice were maintained under standard conditions according to the institutional guidelines for animal care. All animal experiments were approved by the Animal Ethics Committee of The Second Affiliated Hospital of Kunming Medical University College (Kunming, China) and were carried out in compliance with the Animal Management Rules of the Ministry of Health of the People's Republic of China.

Amino Acid-Mediated Metabolism: A New Power to Influence Properties of Stem Cells.

The self-renewal and differentiation potentials of stem cells are dependent on amino acid (AA) metabolism. We review the literature on the metabolic preference of both cancer and noncancer stem cells. The balance in AA metabolism is responsible for maintaining the functionality of noncancer stem cells, and altering the levels of AAs can influence the malignant biological behavior of cancer stem cells. AAs are considered nutrients participating in metabolism and playing a critical role in maintaining the activity of normal stem cells and the effect of therapy of cancer stem cells. Targeting AA metabolism helps inhibit the stemness of cancer stem cells and remodels the function of normal stem cells. This review summarizes the metabolic characteristics and regulation pathways of AA in different stem cells, not only from the nutritional perspective but also from the genomic perspective that have been reported in the recent five years. In addition, we briefly survey new therapeutic modalities that may help eradicate cancer stem cells by exploiting nutrient deprivation. Understanding AA uptake characteristics helps researchers define the preference for AA in different stem cells and enables clinicians make timely interventions to specifically target the cell behavior.

Histone deacetylase (HDAC) inhibitors and doxorubicin combinations target both breast cancer stem cells and non-stem breast cancer cells simultaneously.

Breast cancer stem cells (CSCs) are a small subpopulation of cancer cells that have high capability for self-renewal, differentiation, and tumor initiation. CSCs are resistant to chemotherapy and radiotherapy, and are responsible for cancer recurrence and metastasis. By utilizing a panel of breast cancer cells and mammospheres culture as cell-based screening platforms, we performed high-throughput chemical library screens to identify agents that are effective against breast CSCs and non-CSCs. The hit molecules were paired with conventional chemotherapy to evaluate the combinatorial treatment effects on breast CSCs and non-CSCs. We identified a total of 193 inhibitors that effectively targeting both breast CSCs and non-CSCs. We observed that histone deacetylase inhibitors (HDACi) synergized conventional chemotherapeutic agents (i.e., doxorubicin and cisplatin) in targeting breast CSCs and non-CSCs simultaneously. Further analyses revealed that quisinostat, a potent inhibitor for class I and II HDACs, potentiated doxorubicin-induced cytotoxicity in both breast CSCs and non-CSCs derived from the basal-like (MDA-MB-468 and HCC38), mesenchymal-like (MDA-MB-231), and luminal-like breast cancer (MCF-7). It was also observed that the basal-like breast CSCs and non-CSCs were more sensitive to the co-treatment of quisinostat with doxorubicin compared to that of the luminal-like breast cancer subtype. In conclusion, this study demonstrates the potential of HDACi as therapeutic options, either as monotherapy or in combination with chemotherapeutics against refractory breast cancer.

Abstract NT-107: CHEMORESISTANT OVARIAN CANCER STEM CELLS REVEAL NOVEL THERAPEUTIC TARGETS

Abstract More than 60% of patients with ovarian cancer respond to first-line therapy, often achieving complete remission. However, these encouraging clinical outcomes are quickly overshadowed by high rates of treatment failure. More than half of advanced-stage patients relapse within five years, and these patients face poor prognoses. We sought to better understand the cause of ovarian cancer treatment failure with the hope of improving patient outcomes. We hypothesized that ovarian cancer stem cells may resist chemotherapy allowing them to regrow tumors causing relapse, metastasis and ultimately death. We further postulated that characterization of these cells would reveal therapeutic targets that may be exploited to achieve more durable remissions. To begin to test this hypothesis, we proposed that high aldehyde dehydrogenase I activity (ALDH1high) would identify stem-like ovarian cancer cells that would resist chemotherapy treatment and be capable of regenerating tumors. We compared ALDH1high cells to ALDH1low cells in three ovarian cancer cell lines: the human SKOV3 cell line, the rat FNAR-C1 cell line, and the murine ID8 cell line. Consistent with our predictions, we found that ALDH1high cells were resistant to several drugs commonly used to treat ovarian cancer. The ALDH1high population was also the only fraction of cells capable of forming tumors in vivo. In keeping with their proposed cancer stem cell phenotype, ALDH1high cells were smaller in size, relatively quiescent, and able to regenerate the phenotypic diversity of the cell line. Furthermore, they lacked contact inhibition and were capable of nonadherent growth. Significant controversy exists concerning how best to identify ovarian cancer stem cells. Many markers have been proposed with data to both support and refute their use in the literature. Resolving these uncertainties is hampered by the lack of systematic evaluation of each marker in proposed cancer stem cell pools. We used microarray analysis and quantitative real-time RT-PCR to measure the expression of ABCG2, CD24, CD44, CD133 and KIT in our ALDH1high cells compared to their ALDH1low counterparts and found no consistent differential regulation in the ALDH1high cells across all three cell lines. Further work remains needed to more clearly define the relationships between each of these proposed ovarian cancer stem cell markers. Gene expression profiles identified several possible vulnerabilities that could be exploited therapeutically. ALDH1high cells have upregulation of the mTOR pathway, FGF18 and CD47. They also express Her-2/neu at similarly high levels as ALDH1low cells. Drugs are already clinically available to target mTOR and Her-2/neu, which would speed their implementation. FGF18 signals through the FGFR3 receptor. Although FGF18 is upregulated in the cancer stem cell pool, both cancer stem and non-stem cells express its receptor, FGFR3, at similar levels. Several approaches to target FGFR3 are in clinical trials, as are antibodies targeting CD47. mTOR and CD47 inhibition would potentially be selective for ovarian cancer stem cells, but Her-2/neu and FGFR3 inhibition could also target the more differentiated, non-stem cancer cells. Further work is necessary to validate these treatment approaches. Citation Format: Allison C. Sharrow, Richard J Jones, Lily Wu. CHEMORESISTANT OVARIAN CANCER STEM CELLS REVEAL NOVEL THERAPEUTIC TARGETS [abstract]. In: Proceedings of the 12th Biennial Ovarian Cancer Research Symposium; Sep 13-15, 2018; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2019;25(22 Suppl):Abstract nr NT-107.

Mitochondrial Damage Mediated by miR-1 Overexpression in Cancer Stem Cells

It is well known that cells rely on mitochondrial respiration for survival. However, the effect of microRNAs (miRNAs) on mitochondria of cells has not been extensively explored. Our results indicated that the overexpression of a miRNA (miR-1) could destroy mitochondria of cancer stem cells. miR-1 was downregulated in melanoma stem cells (MSCs) and breast cancer stem cells (BCSCs) compared with cancer non-stem cells. However, the upregulation of miR-1 in cancer non-stem cells did not induce mitochondrial damage. miR-1 overexpression caused mitochondrial damage of cancer stem cells by directly targeting the 3′ UTRs of MINOS1 (mitochondrial inner membrane organizing system 1) and GPD2 (glycerol-3-phosphate dehydrogenase 2) genes and interacting with LRPPRC (leucine-rich pentatricopeptide-repeat containing) protein, a protein localized in mitochondria. MINOS1, GPD2, and LRPPRC in mitochondria were required for mitochondrial inner membrane. The results of in vitro and in vivo assays demonstrated that miR-1 overexpression induced mitophagy of cancer stem cells. Therefore, our study contributed novel insights into the mechanism of miRNA-mediated regulation of mitochondria morphology of cancer stem cells.