RESUMO
The assembly of organized colonies is the earliest manifestation in the derivation or induction of pluripotency in vitro. However, the necessity and origin of this assemblance is unknown. Here, we identify human pluripotent founder cells (hPFCs) that initiate, as well as preserve and establish, pluripotent stem cell (PSC) cultures. PFCs are marked by N-cadherin expression (NCAD+) and reside exclusively at the colony boundary of primate PSCs. As demonstrated by functional analysis, hPFCs harbor the clonogenic capacity of PSC cultures and emerge prior to commitment events or phenotypes associated with pluripotent reprogramming. Comparative single-cell analysis with pre- and post-implantation primate embryos revealed hPFCs share hallmark properties with primitive endoderm (PrE) and can be regulated by non-canonical Wnt signaling. Uniquely informed by primate embryo organization in vivo, our study defines a subset of founder cells critical to the establishment pluripotent state.
Assuntos
Antígenos CD/metabolismo , Caderinas/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Diferenciação Celular , Linhagem da Célula , Desenvolvimento Embrionário , Células-Tronco Embrionárias/metabolismo , Endoderma/metabolismo , Expressão Gênica/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Humanos , Análise de Célula Única , Via de Sinalização WntRESUMO
Sam68 (Src associated in mitosis of 68 kDa) is an RNA-binding and multifunctional protein extensively characterized in numerous cellular functions, such as RNA processing, cell cycle regulation, kinase- and growth factor signaling. Recent investigations highlighted Sam68 as a primary target of a class of reverse-turn peptidomimetic drugs, initially developed as inhibitors of Wnt/ß-catenin mediated transcription. Further investigations on such compounds revealed their capacity to selectively eliminate cancer stem cell (CSC) activity upon engaging Sam68. This work highlighted previously unappreciated roles for Sam68 in the maintenance of neoplastic self-renewal and tumor-initiating functions. Here, we discuss the implication of Sam68 in tumorigenesis, where central findings support its contribution to chromatin regulation processes essential to CSCs. We also review advances in CSC-targeting drug discovery aiming to modulate Sam68 cellular distribution and protein-protein interactions. Ultimately, Sam68 constitutes a vulnerability point of CSCs and an attractive therapeutic target to impede neoplastic stemness in human tumors.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Neoplasias , Humanos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Proteínas de Ligação a RNA/metabolismoRESUMO
We show that vitamin A (all-trans-retinol) (VA) is required both for the maintenance of pancreatic ß-cell and α-cell mass and for glucose-stimulated insulin secretion in adult mice. Dietary VA deprivation (VAD) causes greatly decreased pancreatic VA levels, hyperglycemia, and reduced insulin secretion. Adult mice fed VAD diets display remodeling of the endocrine pancreas, marked ß-cell apoptosis, shifts to smaller islet size distributions, decreased ß-cell mass, increased α-cell mass, and hyperglucagonemia. Importantly, although we induced VAD in the entire animal, the pancreatic ß-cells are exquisitely sensitive to VAD-associated apoptosis compared with other cell types in other organs. VAD causes major reductions in levels of the VA intracellular binding protein Crbp1 and the retinoic acid-metabolizing enzyme Cyp26a1 specifically in larger islets, suggesting the use of these proteins as biomarkers for early endocrine mass abnormalities. In the VAD mice, the reductions in pancreatic islet sizes and the associated aberrant endocrine functions, which show similarities to the phenotype in advanced type 2 diabetes, result from reductions in pancreatic VA signaling. Reintroduction of dietary VA to VAD mice restores pancreatic VA levels, glycemic control, normal islet size distributions, ß-cell to α-cell ratios, endocrine hormone profiles, and RARß2 and RARγ2 transcript levels. Restoration of ß-cell mass by reintroducing VA to VAD mice does not involve increased ß-cell proliferation or neogenesis. Pharmacologic modulation of pancreatic VA signaling should be explored for the preservation and/or restoration of pancreatic ß-cell mass and function in individuals with diabetes mellitus.
Assuntos
Apoptose , Hiperglicemia/patologia , Células Secretoras de Insulina/patologia , Deficiência de Vitamina A/patologia , Animais , Proliferação de Células , Cromatografia Líquida de Alta Pressão , Sistema Enzimático do Citocromo P-450/metabolismo , Diabetes Mellitus Tipo 2/patologia , Modelos Animais de Doenças , Glucagon/metabolismo , Insulina/metabolismo , Ilhotas Pancreáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Ácido Retinoico 4 Hidroxilase , Retinoides/metabolismo , Proteínas Celulares de Ligação ao Retinol/metabolismo , Transdução de SinaisRESUMO
A controlled balance between cell proliferation and differentiation is essential to maintain normal intestinal tissue renewal and physiology. Such regulation is powered by several intracellular pathways that are translated into the establishment of specific transcription programs, which influence intestinal cell fate along the crypt-villus axis. One important check-point in this process occurs in the transit amplifying zone of the intestinal crypts where different signaling pathways and transcription factors cooperate to manage cellular proliferation and differentiation, before secretory or absorptive cell lineage terminal differentiation. However, the importance of epigenetic modifications such as histone methylation and acetylation in the regulation of these processes is still incompletely understood. There have been recent advances in identifying the impact of histone modifications and chromatin remodelers on the proliferation and differentiation of normal intestinal crypt cells. In this review we discuss recent discoveries on the role of the cellular epigenome in intestinal cell fate, development, and tissue renewal. J. Cell. Physiol. 231: 2361-2367, 2016. © 2016 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.
Assuntos
Epigênese Genética , Células Epiteliais/citologia , Intestinos/citologia , Animais , Proliferação de Células , Histonas/metabolismo , Humanos , Processamento de Proteína Pós-TraducionalRESUMO
The small intestine consists of two histological compartments composed of the crypts and the villi. The function of the adult small intestinal epithelium is mediated by four different types of mature cells: enterocytes, goblet, enteroendocrine and Paneth. Undifferentiated cells reside in the crypts and produce these four types of mature cells. The niche-related Wnt and Bmp signaling pathways have been suggested to be involved in the regulation and maintenance of the stem cell microenvironment. In our laboratory, we isolated the first normal human intestinal epithelial crypt (HIEC) cell model from the human fetal intestine and in this study we investigated the expression of a panel of intestinal stem cell markers in HIEC cells under normal culture parameters as well as under conditions that mimic the stem cell microenvironment. The results showed that short term stimulation of HIEC cells with R-spondin 1 and Wnt-3a±SB-216763, a glycogen synthase kinase 3ß (GSK3ß) inhibitor, induced ß-catenin/TCF activity and expression of the WNT target genes, cyclin D2 and LGR5. Treatment of HIEC cells with noggin, an antagonist of BMP signaling, abolished SMAD2/5/8 phosphorylation. Inducing a switch from inactive WNT/active BMP toward active WNT/inactive BMP pathways was sufficient to trigger a robust intestinal primordial stem-like cell signature with predominant LGR5, PHLDA1, PROM1, SMOC2 and OLFM4 expression. These findings demonstrate that even fully established cultures of intestinal cells can be prompted toward a CBC stem cell-like phenotype. This model should be useful for studying the regulation of human intestinal stem cell self-renewal and differentiation.
Assuntos
Diferenciação Celular , Neoplasias Colorretais/patologia , Células Epiteliais/citologia , Intestinos/citologia , Células-Tronco/citologia , Proteínas Wnt/metabolismo , Via de Sinalização Wnt/fisiologia , Adulto , Western Blotting , Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Proliferação de Células , Células Cultivadas , Neoplasias Colorretais/metabolismo , Células Epiteliais/metabolismo , Feto/citologia , Feto/metabolismo , Técnica Indireta de Fluorescência para Anticorpo , Regulação da Expressão Gênica , Humanos , Mucosa Intestinal/metabolismo , Luciferases/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Regeneração/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células-Tronco/metabolismo , Proteínas Wnt/genéticaRESUMO
Polycomb proteins play key roles in mediating epigenetic modifications that occur during cell differentiation. The Polycomb repressive complex 2 (PRC2) mediates the tri-methylation of histone H3 lysine 27 (H3K27me3). In this study, we identify a distinguishing feature of two classes of PRC2 target genes, represented by the Nr2F1 (Coup-TF1) and the Hoxa5 gene, respectively. Both genes are transcriptionally activated by all-trans retinoic acid (RA) and display increased levels of the permissive H3K9/K14ac and tri-methylated histone H3 lysine 4 epigenetic marks in response to RA. However, while in response to RA the PRC2 and H3K27me3 marks are greatly decreased at the Hoxa5 promoter, these marks are initially increased at the Nr2F1 promoter. Functional depletion of the essential PRC2 protein Suz12 by short hairpin RNA (shRNA) technology enhanced the RA-associated transcription of Nr2F1, Nr2F2, Meis1, Sox9 and BMP2, but had no effect on the Hoxa5, Hoxa1, Cyp26a1, Cyp26b1 and RARß2 transcript levels in wild-type embryonic stem cells. We propose that PRC2 recruitment attenuates the RA-associated transcriptional activation of a subset of genes. Such a mechanism would permit the fine-tuning of transcriptional networks during differentiation.
Assuntos
Fator I de Transcrição COUP/genética , Complexo Repressor Polycomb 2/metabolismo , Ativação Transcricional , Tretinoína/farmacologia , Região 5'-Flanqueadora , Animais , Linhagem Celular , Linhagem Celular Tumoral , Metilação de DNA , Histonas/metabolismo , Proteínas de Homeodomínio/genética , Camundongos , Fosfoproteínas/genética , Complexo Repressor Polycomb 2/antagonistas & inibidores , Complexo Repressor Polycomb 2/genética , RNA Polimerase III/metabolismo , Receptores do Ácido Retinoico/metabolismo , Deleção de Sequência , Fatores de TranscriçãoRESUMO
The crypt-villus axis constitutes the functional unit of the small intestine, where mature absorptive cells are confined to the villi, and stem cells and transit amplifying and differentiating cells are restricted to the crypts. The polycomb group (PcG) proteins repress differentiation and promote self-renewal in embryonic stem cells. PcGs prevent transcriptional activity by catalysing epigenetic modifications, such as the covalent addition of methyl groups on histone tails, through the action of the polycomb repressive complex 2 (PRC2). Although a role for PcGs in the preservation of stemness characteristics is now well established, recent evidence suggests that they may also be involved in the regulation of differentiation. Using intestinal epithelial cell models that recapitulate the enterocytic differentiation programme, we generated a RNAi-mediated stable knockdown of SUZ12, which constitutes a cornerstone for PRC2 assembly and functionality, in order to analyse intestinal cell proliferation and differentiation. Expression of SUZ12 was also investigated in human intestinal tissues, revealing the presence of SUZ12 in most proliferative epithelial cells of the crypt and an increase in its expression in colorectal cancers. Moreover, PRC2 disruption led to a significant precocious expression of a number of terminal differentiation markers in intestinal cell models. Taken together, our data identified a mechanism whereby PcG proteins participate in the repression of the enterocytic differentiation program, and suggest that a similar mechanism exists in situ to slow down terminal differentiation in the transit amplifying cell population.
Assuntos
Trato Gastrointestinal/citologia , Complexo Repressor Polycomb 2/fisiologia , Células CACO-2 , Diferenciação Celular/fisiologia , Processos de Crescimento Celular/fisiologia , Células Cultivadas , Trato Gastrointestinal/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Histonas/genética , Histonas/metabolismo , Humanos , Complexo Repressor Polycomb 2/genética , Complexo Repressor Polycomb 2/metabolismo , Regiões Promotoras Genéticas , Transcrição GênicaRESUMO
All-trans retinoic acid (RA) signals via binding to retinoic acid receptors (RARs α, ß, and γ). RA directly influences expression of Pdx1, a transcription factor essential for pancreatic development and beta-cell (ß-cell) maturation. In this study we follow the differentiation of cultured wild-type (WT) vs. RARß knockout (KO) embryonic stem (ES) cells into pancreatic islet cells. We found that RARß KO ES cells show greatly reduced expression of some important endocrine markers of differentiated islet cells, such as glucagon, islet amyloid polypeptide (Iapp), and insulin 1 (Ins1) relative to WT. We conclude that RARß activity is essential for proper differentiation of ES cells to pancreatic endocrine cells.
Assuntos
Diferenciação Celular/genética , Deleção de Genes , Ilhotas Pancreáticas/citologia , Receptores do Ácido Retinoico/genética , Animais , Linhagem Celular , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Glucagon/genética , Glucagon/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Insulina/genética , Insulina/metabolismo , Polipeptídeo Amiloide das Ilhotas Pancreáticas/genética , Polipeptídeo Amiloide das Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Receptores do Ácido Retinoico/metabolismo , Transativadores/genética , Transativadores/metabolismo , Transcrição Gênica , Tretinoína/metabolismoRESUMO
Colorectal cancer is among the leading causes of cancer death in the USA. The polycomb repressive complex 2 (PRC2), including core components SUZ12 and EZH2, represents a key epigenetic regulator of digestive epithelial cell physiology and was previously shown to promote deleterious effects in a number of human cancers, including colon. Using colon cancer stem cells (CCSC) isolated from human primary colorectal tumors, we demonstrate that SUZ12 knockdown and treatment with DZNep, one of the most potent EZH2 inhibitors, increase apoptosis levels, marked by decreased Akt phosphorylation, in CCSCs, while embryonic stem (ES) cell survival is not affected. Moreover, DZNep treatments lead to increased PTEN expression in these highly tumorigenic cells. Taken together, our findings suggest that pharmacological inhibition of PRC2 histone methyltransferase activity may constitute a new, epigenetic therapeutic strategy to target highly tumorigenic and metastatic colon cancer stem cells.
Assuntos
Apoptose , PTEN Fosfo-Hidrolase/metabolismo , Complexo Repressor Polycomb 2/antagonistas & inibidores , Animais , Sobrevivência Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Epigênese Genética , Técnica Indireta de Fluorescência para Anticorpo , Células HT29 , Histonas/metabolismo , Humanos , Marcação In Situ das Extremidades Cortadas , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Proteínas de Neoplasias , PTEN Fosfo-Hidrolase/genética , Fosforilação , Complexo Repressor Polycomb 2/genética , Complexo Repressor Polycomb 2/metabolismo , Regiões Promotoras Genéticas , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Fatores de Transcrição , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Cancer stem cells (CSCs), functionally characterized by self-renewal and tumor-initiating activity, contribute to decreased tumor immunogenicity, while fostering tumor growth and metastasis. Targeting G9a histone methyltransferase (HMTase) effectively blocks CSC functions in colorectal tumors by altering pluripotent-like molecular networks; however, existing molecules directly targeting G9a HMTase activity failed to reach clinical stages due to safety concerns. Using a stem cell-based phenotypic drug-screening pipeline, we identified the dopamine transporter (DAT) antagonist vanoxerine, a compound with previously demonstrated clinical safety, as a cancer-specific downregulator of G9a expression. Here we show that gene silencing and chemical antagonism of DAT impede colorectal CSC functions by repressing G9a expression. Antagonizing DAT also enhanced tumor lymphocytic infiltration by activating endogenous transposable elements and type-I interferon response. Our study unveils the direct implication of the DAT-G9a axis in the maintenance of CSC populations and an approach to improve antitumor immune response in colon tumors.
Assuntos
Neoplasias do Colo , Histona-Lisina N-Metiltransferase , Piperazinas , Humanos , Histona-Lisina N-Metiltransferase/genética , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Dopamina/farmacologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologiaRESUMO
Colorectal cancer is ranked among the top leading causes of cancer death in industrialized populations. Polycomb group proteins, including Suz12 and Ezh2, are epigenetic regulatory proteins that act as transcriptional repressors of many differentiation-associated genes and are overexpressed in a large subset of colorectal cancers. Retinoic acid (RA) acts as a negative regulator of PcG actions in stem cells, but has shown limited therapeutic potential in some solid tumors, including colorectal cancer, in part because of retinoic acid receptor ß silencing. Through treatment with RA, Suz12 shRNA knockdown, or Ezh2 pharmacological inhibition with 3-deazaneplanocin A (DZNep), we increased TRAIL-mediated apoptosis in human colorectal cancer cell lines. This increased apoptosis in human colon cancer cells after RA or DZNep treatment was associated with a ~2.5-fold increase in TNFRSF10B (DR5) transcript levels and a 42% reduction in the H3K27me3 epigenetic mark at the TNFRSF10B promoter after DZNep addition. Taken together, our findings indicate that pharmacological inhibition of Polycomb repressive complex 2 histone methyltransferase activity may constitute a new epigenetic therapeutic strategy to overcome RA non-responsiveness in a subset of colorectal tumors by increasing TRAIL-mediated apoptosis sensitivity.
Assuntos
Apoptose/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Complexo Repressor Polycomb 2/antagonistas & inibidores , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Adenosina/análogos & derivados , Adenosina/farmacologia , Apoptose/genética , Linhagem Celular Tumoral , Neoplasias do Colo/genética , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste , Epigenômica , Células HT29 , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Células MCF-7 , Proteínas de Neoplasias , Complexo Repressor Polycomb 2/genética , Complexo Repressor Polycomb 2/metabolismo , Regiões Promotoras Genéticas/efeitos dos fármacos , RNA Interferente Pequeno/antagonistas & inibidores , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/genética , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/genética , Fatores de Transcrição , Tretinoína/farmacologiaRESUMO
Colorectal tumors are heterogenous cellular systems harboring small populations of self-renewing and highly tumorigenic cancer stem cells (CSCs). Understanding the mechanisms fundamental to the emergence of CSCs and colorectal tumor initiation is crucial for developing effective therapeutic strategies. Two recent studies have highlighted the importance of developmental gene expression programs as potential therapeutic targets to suppress pro-oncogenic stem cell populations in the colonic epithelium. Specifically, a subset of aberrant stem cells was identified in preneoplastic intestinal lesions sharing significant transcriptional similarities with fetal gut development. In such aberrant stem cells, Sox9 was shown as a cornerstone for altered cell plasticity, the maintenance of premalignant stemness, and subsequent colorectal tumor initiation. Independently, chemical genomics was used to identify FDA-approved drugs capable of suppressing neoplastic self-renewal based on the ontogenetic root of a target tumor and transcriptional programs embedded in pluripotency. Here, we discuss the joint conclusions from these two approaches, underscoring the importance of developmental networks in CSCs as a novel paradigm for identifying therapeutics targeting colorectal cancer stemness.
RESUMO
The frequency of somatic retrotranspositions of Long Interspersed Nuclear Elements 1 (LINE1) over a lifetime in healthy colonic epithelium and colorectal tumors has recently been reported. Indicative of a cell type-specific effect, LINE1 sequences in colonic epithelium showed lower levels of DNA methylation compared to other cell types examined in the study. Consistent with a role for DNA methylation in transposon silencing, the decreases in DNA methylation observed at LINE1 elements in colonic epithelium were accompanied by increases in LINE1 mRNA levels. In human primary colorectal tumors, LINE1 retrotransposition frequency was tenfold higher than in normal colonic tissues, with insertions potentially altering genomic stability and cellular functions. Here, we discuss the discoveries made by Nam and colleagues, emphasizing the intestinal-specific methylation signature regulating the LINE1 lifecycle and how this new information could shape future drug discovery endeavors against colorectal cancer.
RESUMO
Overlapping principles of embryonic and tumor biology have been described, with recent multi-omics campaigns uncovering shared molecular profiles between human pluripotent stem cells (hPSCs) and adult tumors. Here, using a chemical genomic approach, we provide biological evidence that early germ layer fate decisions of hPSCs reveal targets of human cancers. Single-cell deconstruction of hPSCs-defined subsets that share transcriptional patterns with transformed adult tissues. Chemical screening using a unique germ layer specification assay for hPSCs identified drugs that enriched for compounds that selectively suppressed the growth of patient-derived tumors corresponding exclusively to their germ layer origin. Transcriptional response of hPSCs to germ layer inducing drugs could be used to identify targets capable of regulating hPSC specification as well as inhibiting adult tumors. Our study demonstrates properties of adult tumors converge with hPSCs drug induced differentiation in a germ layer specific manner, thereby expanding our understanding of cancer stemness and pluripotency.
Assuntos
Neoplasias , Células-Tronco Pluripotentes , Humanos , Diferenciação Celular/fisiologia , Neoplasias/tratamento farmacológico , Neoplasias/genética , GenômicaRESUMO
Organoids can enable the study of solid tumors initiated from a single cancer stem cell (CSC) ex vivo. We describe a serial tumor organoid plating protocol using primary colorectal cancer (CRC) tissues as a rapid and cost-efficient approach to evaluate the impact of therapeutic interventions on CSC functions. We detail the isolation of primary colorectal CSCs, organoid embedding, serial passaging, and CSC-related analytical techniques. For complete details on the use and execution of this protocol, please refer to Masibag et al. (2021) and Bergin et al. (2021).
Assuntos
Neoplasias Colorretais , Organoides , Neoplasias Colorretais/patologia , Humanos , Células-Tronco Neoplásicas/patologiaRESUMO
The histone methyltransferase G9a is well-documented for its implication in neoplastic growth. However, recent investigations have demonstrated a key involvement of this chromatin writer in maintaining the self-renewal and tumor-initiating capacities of cancer stem cells (CSCs). Direct inhibition of G9a's catalytic activity was reported as a promising therapeutic target in multiple preclinical studies. Yet, none of the available pharmacological inhibitors of G9a activity have shown success at the early stages of clinical testing. Here, we discuss central findings of oncogenic expression and activation of G9a in CSCs from different origins, as well as the impact of the suppression of G9a histone methyltransferase activity in such contexts. We will explore the challenges posed by direct and systemic inhibition of G9a activity in the perspective of clinical translation of documented small molecules. Finally, we will discuss recent advances in drug discovery as viable strategies to develop context-specific drugs, selectively targeting G9a in CSC populations.
RESUMO
Colorectal tumors are hierarchically organized and governed by populations of self-renewing cancer stem cells, representing one of the deadliest types of cancers worldwide. Emergence of cancer stemness phenotype depends on epigenetic reprogramming, associated with profound transcriptional changes. As described for pluripotent reprogramming, epigenetic modifiers play a key role in cancer stem cells by establishing embryonic stem-like transcriptional programs, thus impacting the balance between self-renewal and differentiation. We identified overexpression of histone methyltransferase G9a as a risk factor for colorectal cancer, associated with shorter relapse-free survival. Moreover, using human transformed pluripotent cells as a surrogate model for cancer stem cells, we observed that G9a activity is essential for the maintenance of embryonic-like transcriptional signature promoting self-renewal, tumorigenicity, and undifferentiated state. Such a role was also applicable to colorectal cancer, where inhibitors of G9a histone methyltransferase function induced intestinal differentiation while restricting tumor-initiating activity in patient-derived colorectal tumor samples. Finally, by integrating transcriptome profiling with G9a/H3K9me2 loci co-occupancy, we identified the canonical Wnt pathway, epithelial-to-mesenchyme transition, and extracellular matrix organization as potential targets of such a chromatin regulation mechanism in colorectal cancer stem cells. Overall, our findings provide novel insights on the role of G9a as a driver of cancer stem cell phenotype, promoting self-renewal, tumorigenicity, and undifferentiated state.
Assuntos
Carcinógenos/metabolismo , Neoplasias Colorretais/genética , Antígenos de Histocompatibilidade/genética , Histona-Lisina N-Metiltransferase/genética , Células-Tronco Neoplásicas/metabolismo , Autorrenovação Celular/genética , Reprogramação Celular/genética , Neoplasias Colorretais/patologia , Epigênese Genética/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Células HCT116 , Humanos , Masculino , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Células-Tronco Neoplásicas/patologia , Intervalo Livre de Progressão , Transcriptoma/genéticaRESUMO
Cancer stem cells (CSCs) are documented to play a key role in tumorigenesis and therapy resistance. Despite significant progress in clinical oncology, CSC reservoirs remain elusive and difficult to eliminate. Reverse-turn peptidomimetics were characterized as disruptors of CBP/beta-Catenin interactions and represent a promising avenue to curb hyperactive canonical Wnt/beta-Catenin signaling in CSCs. Recent studies suggested Sam68 as a critical mediator of reverse-turn peptidomimetics response in CSC populations. Using computational and biochemical approaches we confirmed Sam68 as a primary target of reverse-turn peptidomimetics. Furthermore, we executed an in silico drug discovery pipeline to identify yet uncharacterized reverse-turn peptidomimetic structures displaying superior anti-CSC activity in transformed pluripotent and colorectal cancer cell models. Thus, we identified YB-0158 as a reverse-turn peptidomimetic small molecule with enhanced translational potential, altering key hallmarks of human colorectal CSCs in patient-derived ex vivo organoids and in vivo serial tumor transplantation.
RESUMO
The aberrant expression of dopamine receptors (DRDs) in acute myeloid leukemia (AML) cells has encouraged the repurposing of DRD antagonists such as thioridazine (TDZ) as anti-leukemic agents. Here, we access patient cells from a Phase I dose escalation trial to resolve the cellular and molecular bases of response to TDZ, and we extend these findings to an additional independent cohort of AML patient samples tested preclinically. We reveal that in DRD2+ AML patients, DRD signaling in leukemic progenitors provides leukemia-exclusive networks of sensitivity that spare healthy hematopoiesis. AML progenitor cell suppression can be increased by the isolation of the positive enantiomer from the racemic TDZ mixture (TDZ+), and this is accompanied by reduced cardiac liability. Our study indicates that the development of DRD-directed therapies provides a targeting strategy for a subset of AML patients and potentially other cancers that acquire DRD expression upon transformation from healthy tissue.
Assuntos
Hematopoese/fisiologia , Leucemia Mieloide Aguda/tratamento farmacológico , Células-Tronco Neoplásicas/metabolismo , Receptores Dopaminérgicos/metabolismo , Tioridazina/metabolismo , Humanos , Leucemia Mieloide Aguda/metabolismo , Transdução de Sinais/fisiologiaRESUMO
Natural products (NPs) encompass a rich source of bioactive chemical entities. Here, we used human cancer stem cells (CSCs) in a chemical genomics campaign with NP chemical space to interrogate extracts from diverse strains of actinomycete for anti-cancer properties. We identified a compound (McM25044) capable of selectively inhibiting human CSC function versus normal stem cell counterparts. Biochemical and molecular studies revealed that McM025044 exerts inhibition on human CSCs through the small ubiquitin-like modifier (SUMO) cascade, found to be hyperactive in a variety of human cancers. McM025044 impedes the SUMOylation pathway via direct targeting of the SAE1/2 complex. Treatment of patient-derived CSCs resulted in reduced levels of SUMOylated proteins and suppression of progenitor and stem cell capacity measured in vitro and in vivo. Our study overcomes a barrier in chemically inhibiting oncogenic SUMOylation activity and uncovers a unique role for SAE2 in the biology of human cancers.