Dysregulation of the Cyclin D/E2F activity as a core mechanism driving cancer stem cell plasticity and cell cycle dynamics

The hypothesis that tumorigenic stem cells arises from the mutated stem cells suggests that cancer can be viewed as an aberrant organ initiated by tumorigenic cancer cells with stem-cell qualities. The tumor initiator cell or tumor stem cell/ cancer stem cell (CSC) is possible origins ofnormal stem cell or progenitor cells. The existence of such a CSC population may represent only a small fraction of a tumor possessing the ability to regenerate tumors through aberrant proliferation and self-renewal, but most cancer cells lack this ability. A central question in stem cell and CSC biology is, which cells can be transformed to generate tumors? With the growing evidence that CSC exist in malignancies of haematopoietic origin and in most solid tumors, it is becoming increasingly important to understand the molecular mechanisms that regulate self-renewal and differentiation because mutation of genes involved in these pathways likely participates in tumor growth. Studies of normal stem cell and CSC from the same tissue have suggested that the ontogeny of tumors. That signaling pathways such as Wnt, BMP and Notch have similar effects on normal stem cell and CSC self-renewal implys that common molecular pathways regulate both populations. Stem cell biology has provided a platform to address many questions in developmental biology. Further understanding the biology of stem cell, cancer cell, and CSC will contribute to the identification of molecular targets important for future therapies. To know the biology of CSC, we first should identify the unique characteristics of stem cells. A normal adult stem cell is defined as a somatic cell undergoing extensive cell division and has the potential to give rise to both stem cells and cells that differentiate into specialized cells. A normal stem cell must possess two qualities to play its natural function: self-renewal and differentiation. Self-renewal is an important ability of stem cells, indicating a special cell division that enables a stem cell to develop another stem cell with essentially the same development and replication potential. The capability to self-renewal causes the stem cell expansion in response to local or systemic signals, which start massive proliferation and maintenance of a tissue specific undifferentiated pool of cells in the organ or tissue. Differentiation is the second function of a stem cell and involves the development of daughter cells that become tissue specific specialized cells. Studies have demonstrated that the CSCs are likely to share many of the properties of normal stem cells, and that the CSCs are source of tumor recurrences and/or metastasis after apparently complete remissions. Although there is an argument about CSC model that may overlook the complexity of the interactions which occur within a tumor and between a tumor and its environment, evidence that many cancers are driven by CSC has important clinical implications. For example, conventional therapies may shrink tumors by killing mainly cells with limited proliferative potential, however, CSCs are less sensitive to these therapies, then they will remain viable after therapy and re-establish the tumor. Therefore, CSC model has favored the conceptual development of targeted therapies towards CSC’s “stemness”. However, how to achieve CSC eradication is emergent to overcome the major barrier. Here we describe latest therapeutic approaches based on the “stemness” of CSC that may allow us to discover the effective methods for eradicating seeds of malignant tumor cells. This includes targeted therapy directed toward CSC by surface specific markers, survival niche, blocking CSC critical signal pathways, inhibiting CSC efficient DNA repair, manipulation of ncRNAs, screening drug-resistance of CSC in three dimensional culture, CSC vaccines as well as CSC nanotheranostics.

Simple SummaryCancer stem cells are thought to cause a poor response to chemotherapies. The aim of our study was to explore the still unknown biology of hepatoblastoma cancer stem cells that are essential for tumorigenesis. With our investigations, we aimed to gain more insight into cancer stem cell characteristics and to provide directions for new therapeutic approaches to treat refractory hepatoblastoma. We showed that hepatoblastoma cancer stem cells express PD-L1, a factor which helps tumors escape immune attacks. Furthermore, we detected cancer stem cell progeny evolving from non-cancer stem cells. Finally, we revealed that another subset of cancer stem cells is induced during chemotherapy. Our findings give a possible explanation why chemotherapies fail in certain hepatoblastoma cases and why new therapeutic approaches should consider the plasticity of hepatoblastoma cancer stem cells.The biology of cancer stem cells (CSCs) of pediatric cancers, such as hepatoblastoma, is sparsely explored. This is mainly due to the very immature nature of these tumors, which complicates the distinction of CSCs from the other tumor cells. Previously, we identified a CSC population in hepatoblastoma cell lines expressing the CSC markers CD34 and CD90, cell surface Vimentin (csVimentin) and binding of OV-6. In this study, we detected the co-expression of the immune escape factor PD-L1 in the CSC population, whereas the other tumor cells remained negative. FACS data revealed that non-CSCs give rise to CSCs, reflecting plasticity of CSCs and non-CSCs in hepatoblastoma as seen in other tumors. When we treated cells with cisplatin and decitabine, a new CD34+/lowOV-6lowCD90+ population emerged that lacked csVimentin and PD-L1 expression. Expression analyses showed that this new CSC subset shared similar pluripotency and EMT features with the already-known CSCs. FACS results further revealed that this subset is also generated from non-CSCs. In conclusion, we showed that hepatoblastoma CSCs express PD-L1 and that the biology of hepatoblastoma CSCs is of a plastic nature. Chemotherapeutic treatment leads to another CSC subset, which is highly chemoresistant and could be responsible for a poor prognosis after postoperative chemotherapy.

Cancer stem cells (CSCs) are a small subset within the tumor mass significantly contributing to cancer progression through dysregulation of various oncogenic pathways, driving tumor growth, chemoresistance and metastasis formation. The aggressive behavior of CSCs is guided by several intracellular signaling pathways such as WNT, NF-kappa-B, NOTCH, Hedgehog, JAK-STAT, PI3K/AKT1/MTOR, TGF/SMAD, PPAR and MAPK kinases, as well as extracellular vesicles such as exosomes, and extracellular signaling molecules such as cytokines, chemokines, pro-angiogenetic and growth factors, which finely regulate CSC phenotype. In this scenario, tumor microenvironment (TME) is a key player in the establishment of a permissive tumor niche, where CSCs engage in intricate communications with diverse immune cells. The "oncogenic" immune cells are mainly represented by B and T lymphocytes, NK cells, and dendritic cells. Among immune cells, macrophages exhibit a more plastic and adaptable phenotype due to their different subpopulations, which are characterized by both immunosuppressive and inflammatory phenotypes. Specifically, tumor-associated macrophages (TAMs) create an immunosuppressive milieu through the production of a plethora of paracrine factors (IL-6, IL-12, TNF-alpha, TGF-beta, CCL1, CCL18) promoting the acquisition by CSCs of a stem-like, invasive and metastatic phenotype. TAMs have demonstrated the ability to communicate with CSCs via direct ligand/receptor (such as CD90/CD11b, LSECtin/BTN3A3, EPHA4/Ephrin) interaction. On the other hand, CSCs exhibited their capacity to influence immune cells, creating a favorable microenvironment for cancer progression. Interestingly, the bidirectional influence of CSCs and TME leads to an epigenetic reprogramming which sustains malignant transformation. Nowadays, the integration of biological and computational data obtained by cutting-edge technologies (single-cell RNA sequencing, spatial transcriptomics, trajectory analysis) has significantly improved the comprehension of the biunivocal multicellular dialogue, providing a comprehensive view of the heterogeneity and dynamics of CSCs, and uncovering alternative mechanisms of immune evasion and therapeutic resistance. Moreover, the combination of biology and computational data will lead to the development of innovative target therapies dampening CSC-TME interaction. Here, we aim to elucidate the most recent insights on CSCs biology and their complex interactions with TME immune cells, specifically TAMs, tracing an exhaustive scenario from the primary tumor to metastasis formation.

Phenotypic and functional heterogeneity is one of the most relevant features of cancer cells within different tumor types and is responsible for treatment failure. Cancer stem cells (CSCs) are a population of cells with stem cell-like properties that are considered to be the root cause of tumor heterogeneity, because of their ability to generate the full repertoire of cancer cell types. Moreover, CSCs have been invoked as the main drivers of metastatic dissemination and therapeutic resistance. As such, targeting CSCs may be a useful strategy to improve the effectiveness of classical anticancer therapies. Recently, metabolism has been considered as a relevant player in CSC biology, and indeed, oncogenic alterations trigger the metabolite-driven dissemination of CSCs. More interestingly, the action of metabolic pathways in CSC maintenance might not be merely a consequence of genomic alterations. Indeed, certain metabotypic phenotypes may play a causative role in maintaining the stem traits, acting as an orchestrator of stemness. Here, we review the current studies on the metabolic features of CSCs, focusing on the biochemical energy pathways involved in CSC maintenance and propagation. We provide a detailed overview of the plastic metabolic behavior of CSCs in response to microenvironment changes, genetic aberrations, and pharmacological stressors. In addition, we describe the potential of comprehensive metabolic approaches to identify and selectively eradicate CSCs, together with the possibility to ‘force’ CSCs within certain metabolic dependences, in order to effectively target such metabolic biochemical inflexibilities. Finally, we focus on targeting mitochondria to halt CSC dissemination and effectively eradicate cancer.

CD44, a multifunctional cell surface protein, has emerged as a pivotal regulator in cancer stem cell (CSC) biology, orchestrating processes such as stemness, metabolic reprogramming, and therapeutic resistance. Recent studies have identified a critical role of CD44 in ferroptosis resistance by stabilizing SLC7A11 (xCT), a key component of the antioxidant defense system, enabling CSCs to evade oxidative stress and sustain tumorigenic potential. Additionally, CD44 regulates intracellular iron metabolism and redox balance, further supporting CSC survival and adaptation to stressful microenvironments. Therapeutic strategies targeting CD44, including ferroptosis inducers and combination therapies, have shown significant potential in preclinical and early clinical settings. Innovations such as CD44-mediated nanocarriers and metabolic inhibitors present novel opportunities to disrupt CSC-associated resistance mechanisms. Furthermore, the dynamic plasticity of CD44 isoforms governed by transcriptional, post-transcriptional, and epigenetic regulation underscores the importance of context-specific therapeutic approaches. This review highlights the multifaceted roles of CD44 in CSC biology, focusing on its contribution to ferroptosis resistance, iron metabolism, and redox regulation. Targeting CD44 offers a promising avenue for overcoming therapeutic resistance and improving the outcomes of refractory cancers. Future studies are needed to refine these strategies and enable their clinical translation.

Click to increase image sizeClick to decrease image sizeKeywords:: cancer stem cellbreast cancerEMTZEB1bivalent chromatincell plasticitynon-cancer stem cell

An alginate-based platform for cancer stem cell research

Cancer stem cells (CSCs) can drive tumor growth, and their maintenance may rely on post-transcriptional regulation of gene expression, including that mediated by microRNAs (miRNAs). The let-7 miRNA family has been shown to induce differentiation by silencing stem cell programs. Let-7-mediated target gene suppression is prevented by LIN28A/B, which reduce let-7 biogenesis in normal embryonic and some cancer stem cells and ensure maintenance ofstemness. Here, we find that glioblastoma stem cells (GSCs) lack LIN28 and express both let-7 and their target genes, suggesting LIN28-independent protection from let-7 silencing. Using photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP), we show that insulin-like growth factor 2 mRNA-binding protein 2 (IMP2) binds to let-7 miRNA recognition elements (MREs) and prevents let-7 target gene silencing. Our observations define the RNA-binding repertoire of IMP2 and identify a mechanism whereby it supports GSC and neural stem cell specification.

Background/Objectives: The present article summarizes and analyzes the current knowledge about the role of markers and dysregulated signaling pathways in pancreatic cancer stem cells (CSCs) and their value for possible therapeutic approaches. Method: An electronic search of PubMed/MEDLINE was used to identify relevant original articles and reviews. Results: Despite significant effort and research funds, pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest diseases. Because of the lack of symptoms, the majority of patients presents with advanced stage. Patients with advanced disease will receive systemic therapy. However, such therapy only eradicates tumor bulk, but does not eliminate so-called cancer stem cells. These CSCs are thought to be the cause of resistance, metastasis, and recurrence. This review will focus on recent insights into the biology of pancreatic CSCs. It further highlights the importance of PDAC stem cell markers as prognostic indicators and targets for therapies specifically eliminating PDAC stem cells. Conclusions: Pancreatic CSCs appear to be crucial for the processes of cancer cell invasion and metastasis. Therefore, the understanding of the molecular mechanisms implicated in the biology of CSCs as well as the identification of specific markers may generate novel therapeutic strategies and contribute in the reduction of metastasis.

The hallmarks of ovarian cancer stem cells and niches: Exploring their harmonious interplay in therapy resistance.

Background: Cancer stem cells (CSCs) contribute to tumor metastasis, chemotherapy resistance, and are highly tumorigenic when injected into immunodeficient mice. High levels of aldehyde dehydrogenase (ALDH1) are characteristic of stem cells and allow the isolation of subpopulations of CSCs, including those derived from breast cancer. HER2+ tumors, which are more aggressive than tumors with no amplification of this oncogene, are enriched in CSCs, a finding that suggests the participation of this oncogene in the generation and maintenance of CSCs. miR-4728-3p is a poorly investigated miRNA, which is located inside an intron of HER2. Its expression is increased in HER2+ tumors and cell lines, but its role in CSC formation and tumor behavior has not been investigated. Aim: to contribute to the better understanding of the role of miRNA-4728-3p in the biology of CSCs in HER2+ breast cancer cell lines. Methods: miR-4728-3p levels were evaluated in breast cancer cell lines MCF-7, BT-474 and SKBR3 grown in adherent conditions and in conditions that favor the formation of mammospheres for 7, 14, 21 and 28 days. Overexpression of miR-4728-3p was performed using lentiviral transduction with plasmids for the overexpression of this miRNA. Flow cytometry was performed to analyze ALDH1 expression in adherent cells and cells derived from mammospheres and also in cells overexpressing miRNA-4728-3p. Results: Mammospheres in culture for 7 days showed high levels of ALDH1 for all cells lines evaluated. The overexpression of miR-4728-3p induced a significant increment (>10-fold) of ALDH1+ subpopulations in MCF-7 and SKBR3, reinforcing its possible role in the biology of CSCs. Conclusion: HER2-amplification is an event that intrinsically leads to the amplification of miR-4728-3p. The effects of the amplification of this oncogene and this miRNA have to be evaluated together as both may have important consequences for tumor behavior and CSC formation. This increased expression of ALDH1 after miR-4728-3p overexpression suggest the importance of this miRNA in the biology of CSC formation. Note: This abstract was not presented at the meeting. Citation Format: Juliana Laino do Val Carneiro, Diana Noronha Nunes, Emmanuel Dias-Neto. Evaluation of the role of miRNA-4728-3p, a miRNA located in an intron of HER2 gene, in breast cancer stem cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 532. doi:10.1158/1538-7445.AM2014-532

Ovarian cancer is most lethal among gynecological cancers with often fatal consequences due to lack of effective biomarkers and relapse, which propels ovarian cancer research into unique directions to establish solid targeted therapeutics. "Ovarian stem cells" expressing germline pluripotent markers serve as novel paradigm with potential to address infertility, menopause, and probably influence tumor initiation. Cancer stem cells (CSCs) pose vital role in tumor recurrence and hence it is extremely important to study them with respect to ovarian stem cells across various cancer stages and normal ovaries. Pluripotent (OCT4, NANOG, SOX2, SSEA1, and SSEA4), germline (IFITM3, VASA/DDX4), and cancer stem (CD44, LGR5) cell specific markers were characterized for protein and mRNA expression in tumor tissues to understand their distribution in the surface epithelium and ovarian cortex in benign, borderline, and high-grade malignant stages. To elucidate whether pluripotent ovarian germline stem cells and CSCs are common subset of stem cells in tumor tissues, VASA was colocalized with known pluripotent stem (OCT4, SSEA1, SSEA4) and CSC (CD44, LGR5) specific markers by confocal microscopy. Single, smaller spherical (≤5 μm), and larger elliptical fibroblast like (≥10 μm) cells (also in clusters or multiples) were detected implying probable functional behavioral significance of cells in tumor initiation and metastasis across various cancer stages. Cells revealed characteristic staining pattern in ovarian surface epithelium (OSE) and cortex regions exclusive for each marker. Co-expression studies revealed specific subpopulations existing simultaneously in OSE and cortex and that a dynamic hierarchy of (cancer) stem cells with germline properties prevails in normal ovaries and cancer stages. Novel insights into CSC biology with respect to ovarian and germline stem cell perspective were obtained. Understanding molecular signatures and distribution within ovarian tissue may enable identification of precise tumor-initiating CSC populations and signaling pathways thus improving their efficient targeting and strategies to prevent their dissemination causing fatal relapse.

Purpose: Tumours are composed of a heterogeneous cell population. Cancer stem cells, which make up a minor fraction of a tumour, may be the cells that initiate and sustain tumour growth. Cancer stem cells are believed to share many properties with normal stem cells that render them relatively insensitive to classical radio- and chemotherapy.Conclusions: We discuss what those (cancer) stem cell properties are and how the interactions with the microenvironment – ‘the niche’ – control those aspects of (cancer) stem cell biology. We also describe possible strategies to target cancer stem cells in order to prevent cancers from escaping therapy.

p97/VCP, an evolutionarily concerned ATPase, partakes in multiple cellular proteostatic processes, including the endoplasmic reticulum (ER)-associated protein degradation (ERAD). Elevated expression of p97 is common in many cancers and is often associated with poor survival. Here we report that the levels of p97 positively correlated with the histological grade, tumor size, and lymph node metastasis in breast cancers. We further examined p97 expression in the stem-like cancer cells or cancer stem cells (CSCs), a cell population that purportedly underscores cancer initiation, therapeutic resistance, and recurrence. We found that p97 was consistently at a higher level in the CD44+/CD24−, ALDH+, or PKH26+ CSC populations than the respective non-CSC populations in human breast cancer tissues and cancer cell lines and p97 expression also positively correlated with that of SOX2, another CSC marker. To assess the role of p97 in breast cancers, cancer proliferation, mammosphere, and orthotopic growth were analyzed. Similarly as p97 depletion, two pharmacological inhibitors, which targets the ER-associated p97 or globally inhibits p97’s ATPase activity, markedly reduced cancer growth and the CSC population. Importantly, depletion or inhibition of p97 greatly suppressed the proliferation of the ALDH+ CSCs and the CSC-enriched mammospheres, while exhibiting much less or insignificant inhibitory effects on the non-CSC cancer cells. Comparable phenotypes produced by blocking ERAD suggest that ER proteostasis is essential for the CSC integrity. Loss of p97 gravely activated the unfolded protein response (UPR) and modulated the expression of multiple stemness and pluripotency regulators, including C/EBPδ, c-MYC, SOX2, and SKP2, which collectively contributed to the demise of CSCs. In summary, p97 controls the breast CSC integrity through multiple targets, many of which directly affect cancer stemness and are induced by UPR activation. Our findings highlight the importance of p97 and ER proteostasis in CSC biology and anticancer therapy.

This article provides a brief overview of cancer-preventive phytochemicals specifically targeting pancreatic cancer (PC) stem cells for prevention and treatment. Cancer stem cells (CSCs) represent a small proportion of the total cells of a given tumor, and contribute to tumor growth, recurrence, metastasis, and treatment resistance. Many intertwined pathways, including hedgehog, Wnt Signaling, and NOTCH, have been shown to play a role in the formation of CSCs. Recently, numerous chemopreventive agents, such as genistein, (-)-epigallocatechin-3-gallate (EGCG), sulforaphane, curcumin, resveratrol, and quercetin, have been shown to target CSCs mediated through the inhibition of multiple signalling pathways, to avoid toxicity and the side effects of chemical compounds. A growing body of research suggests that CSCs are the drivers in treatment resistance, cancer recurrence, and metastasis, in addition to tumor initiation and heterogeneity. Patient survival depends on these CSCs, which are one cause of tumor recurrence after surgery and chemotherapy. Therefore, target selection; an improved understanding of CSC biology, the genetic and molecular profiles of CSCs, and their key signaling pathways, and; appropriate clinical trials endpoints that are designed to target CSCs will help in the development of drugs that will specifically target this small population of stem cells.