In this review, we explore key innovations in photoactivated therapeutic programming of carbon-based nanomaterials (CBNs), focusing on their diverse nanostructural configurations and their exceptional photothermal, photochemical, and photoacoustic properties. These attributes position CBNs as remarkable phototherapeutic agents, capable of addressing critical challenges in targeted cancer therapy through their precision, multifunctionality, and adaptability to specific therapeutic modalities. We will explore their diverse derivatives, and the role of chemical augmentation and site-specific surface functionalisation, which are pivotal in optimising the targeting and efficacy of phototherapeutic interventions. The biological and physical relevance of this ever-growing library of nanomaterials in targeted phototherapy will be thoroughly explored. Dynamic photo-triggering of the underlying molecular mechanisms of action e.g., energy conversion modalities lie at the heart of these therapeutic innovations. We will further discuss the tunability and programming of these carriers and structure-function alterations at specific therapeutic wavelengths. The application space of phototherapies is thoroughly mapped exploring the three primary approaches of photothermal therapy, photodynamic therapy and photochemical internalisation as well as emerging techniques and promising multimodal approaches that combine two or more of these processes. The specificity of the target tissue site and the approach under study forms another critical focus area of this review, with an emphasis on three types of cancer-breast cancer, lung cancer, and gliomas-that have demonstrated some of the most promising outcomes from photomedicine. We also provide a perspective on in vitro and in vivo validation and preclinical testing of CBNs for phototherapeutic applications. Finally, we reflect on the potential of CBNs to revolutionise targeted cancer therapy through data-driven materials design and integration with computational tools for biophysical performance optimisation. The exciting integration of machine learning into nanoparticle research and phototherapy has potential to fundamentally transform the landscape of nanomedicine. These techniques ranging from supervised learning algorithms such as random forests and support vector machines to more advanced neural networks and deep learning, can enable unprecedented precision in predicting, optimising, and tailoring the properties of nanoparticles for targeted applications. The transformative impact of photoactivated CBNs in advancing cancer treatment, paves the way for their clinical application and widespread adoption in personalised photomedicine. We conclude with a section on the current challenges facing the reproducibility, manufacturing throughput, and biocompatibility of these nanostructured materials including their long-term effects in trials and degradation profiles in biological systems as evaluated in vitro and in vivo.

The Wnt signaling pathway is critically involved in orchestrating cellular functions such as proliferation, migration, survival, and cell fate determination during development. Given its pivotal role in cellular communication, aberrant Wnt signaling has been extensively linked to the pathogenesis of various diseases. This review offers an in-depth analysis of the Wnt pathway, detailing its signal transduction mechanisms and principal components. Furthermore, the complex network of interactions between Wnt cascades and other key signaling pathways, such as Notch, Hedgehog, TGF-β, FGF, and NF-κB, is explored. Genetic mutations affecting the Wnt pathway play a pivotal role in disease progression, with particular emphasis on Wnt signaling's involvement in cancer stem cell biology and the tumor microenvironment. Additionally, this review underscores the diverse mechanisms through which Wnt signaling contributes to diseases such as cardiovascular conditions, neurodegenerative disorders, metabolic syndromes, autoimmune diseases, and cancer. Finally, a comprehensive overview of the therapeutic progress targeting Wnt signaling was given, and the latest progress in disease treatment targeting key components of the Wnt signaling pathway was summarized in detail, including Wnt ligands/receptors, β-catenin destruction complexes, and β-catenin/TCF transcription complexes. The development of small molecule inhibitors, monoclonal antibodies, and combination therapy strategies was emphasized, while the current potential therapeutic challenges were summarized. This aims to enhance the current understanding of this key pathway.

Cancer is a heterogeneous disease, which contributes to its rapid progression and therapeutic failure. Besides interpatient tumor heterogeneity, tumors within a single patient can present with a heterogeneous mix of genetically and phenotypically distinct subclones. These unique subclones can significantly impact the traits of cancer. With the plasticity that intratumoral heterogeneity provides, cancers can easily adapt to changes in their microenvironment and therapeutic exposure. Indeed, tumor cells dynamically shift between a more differentiated, rapidly proliferating state with limited tumorigenic potential and a cancer stem cell (CSC)-like state that resembles undifferentiated cellular precursors and is associated with high tumorigenicity. In this context, CSCs are functionally located at the apex of the tumor hierarchy, contributing to the initiation, maintenance, and progression of tumors, as they also represent the subpopulation of tumor cells most resistant to conventional anti-cancer therapies. Although the CSC model is well established, it is constantly evolving and being reshaped by advancing knowledge on the roles of CSCs in different cancer types. Here, we review the current evidence of how CSCs play a pivotal role in providing the many traits of aggressive tumors while simultaneously evading immunosurveillance and anti-cancer therapy in several cancer types. We discuss the key traits and characteristics of CSCs to provide updated insights into CSC biology and highlight its implications for therapeutic development and improved treatment of aggressive cancers.

Cancer stem cells (CSCs) represent a small, yet pivotal subpopulation of tumor cells that play significant roles in tumor initiation, progression and therapeutic resistance. Circular RNAs (circRNAs) are a distinct class of RNAs characterized by their closed‑loop structures, lacking 5' to 3'ends. There is growing evidence that circRNAs are integral to the development and regulation of CSCs. Aberrant expression of circRNAs in CSCs can contribute to oncogenic properties and drug resistance. Specifically, oncogenic circRNAs modulate CSC behavior via key signaling pathways, thereby promoting CSC self‑renewal and maintenance, as well as tumor progression. This review summarizes the latest research on the functional roles and regulatory mechanisms of circRNAs in CSC behavior and discusses potential applications and challenges of targeting circRNAs in CSCs. Understanding the intricate interactions between circRNAs and CSCs may lead to novel therapeutic strategies that effectively combat treatment resistance and improve patient outcomes.

Extracellular membrane vesicles (EVs) offer promising values in various medical fields, e.g., as biomarkers in liquid biopsies or as native (or bioengineered) biological nanocarriers in tissue engineering, regenerative medicine and cancer therapy. Based on their cellular origin EVs can vary considerably in composition and diameter. Cell biological studies on mammalian prominin-1, a cholesterol-binding membrane glycoprotein, have helped to reveal new donor membranes as sources of EVs. For instance, small EVs can originate from microvilli and primary cilia, while large EVs might be produced by transient structures such as retracting cellular extremities of cancer cells during the mitotic rounding process, and the midbody at the end of cytokinesis. Here, we will highlight the various subcellular origins of prominin-1+ EVs, also called prominosomes, and the potential mechanism(s) regulating their formation. We will further discuss the molecular and cellular characteristics of prominin-1, notably those that have a direct effect on the release of prominin-1+ EVs, a process that might be directly implicated in donor cell reprogramming of stem and cancer stem cells. Prominin-1+ EVs also mediate intercellular communication during embryonic development and adult homeostasis in healthy individuals, while disseminating biological information during diseases.

Glioblastoma stem cells (GSCs) are potent tumor initiators resistant to radiochemotherapy, and this subpopulation is hypothesized to re-populate the tumor milieu due to selection following conventional therapies. Here, we show that 5-aminolevulinic acid (ALA) treatment-a pro-fluorophore used for fluorescence-guided cancer surgery-leads to elevated levels of fluorophore conversion in patient-derived GSC cultures, and subsequent red light-activation induces apoptosis in both intrinsically temozolomide chemotherapy-sensitive and -resistant GSC phenotypes. Red light irradiation of ALA-treated cultures also exhibits the ability to target mesenchymal GSCs (Mes-GSCs) with induced temozolomide resistance. Furthermore, sub-lethal light doses restore Mes-GSC sensitivity to temozolomide, abrogating GSC-acquired chemoresistance. These results suggest that ALA is not only useful for fluorescence-guided glioblastoma tumor resection, but that it also facilitates a GSC drug-resistance agnostic, red light-activated modality to mop up the surgical margins and prime subsequent chemotherapy.

Glioblastoma (GB) is a cancerous brain tumor that originates from glial cells and leads to thousands of deaths each year and a five-year survival of only 6.8 %. Treatments for GB include surgery, chemotherapy, radiation, and immunotherapy. GB is an incurable fatal disease, necessitating the development of innovative strategies to find a developing effective therapy. Genetic therapies may be crucial in treating GB by identifying the mutations and amplifications of multiple genes, which drive its proliferation and spread. Use of small interfering RNAs (siRNAs) provides a novel technology used to suppress the genes associated with disease, which forms a basis for targeted therapy in GB and its stem cell population, which are recognized for their ability to develop resistance to chemotherapy and tumorigenic capabilities. This review examines the use of siRNAs in GB, emphasizing their effectiveness in suppressing key oncogenes and signaling pathways associated with tumor development, invasion, stemness, and resistance to standard treatments. siRNA-based gene silencing is a promising approach for developing targeted therapeutics against GB and associated stem cell populations, potentially enhancing patient outcomes and survival rates in this devastating disease.

Neuroblastoma is the most common extra-cranial solid tumor diagnosed mostly in children below the age of five years and comprises of about 15 % of all paediatric cancer deaths. Tumor initiating cancer stem cells (CSCs) can be targeted for better treatment approaches. BASP1-AS1 is a long non coding (Lnc) RNA that is a divergent LncRNA for its coding gene brain abundant membrane attached signal protein 1 (BASP1). We had earlier demonstrated it to be expressed in foetus derived human neural progenitor cells (hNPCs), where it was a positive regulator of BASP1 and was critical for neural differentiation. In this study, we have investigated the role of BASP1-AS1 in CSCs derived from the human neuroblastoma cell line SH-SY5Y. We cultured SH-SY5Y cells on Poly-d-Lysine coated flasks in serum free media supplemented with growth factors, which led to the enrichment of CSCs as determined by marker expression. When grown on ultra-low attachment flasks, these cells formed CSCs enriched neurospheres. We examined the effects of BASP1-AS1 siRNA mediated knockdown on CSCs enriched SH-SY5Y cells and SH-SY5Y derived neurospheres. BASP1-AS1 knockdown decreased the levels of the corresponding gene BASP1 and the rate of cell proliferation of CSCs enriched cells along with low expression of Ki67. It also reduced the mRNA levels of stem cell and pluripotency gene markers (CD133, CD44, c-KIT, SOX2, OCT4 and NANOG), as also Wnt 2 and the Wnt pathway effector β catenin. It also abrogated the formation of neurospheres in ultra-low attachment flasks. A similar effect on proliferation and stemness related properties was seen on BASP1 knockdown. BASP1-AS1 and its related pathways may provide a point of intervention for the CSCs population in neuroblastoma.

Glioblastoma multiforme (GBM) is among the most aggressive brain cancers, and it contains glioma stem cells (GSCs) that drive tumor initiation, progression, and recurrence. These cells resist conventional therapies, contributing to high recurrence rates in GBM patients. Developing in vitro models that mimic the tumor microenvironment (TME), particularly the GSC niche, is crucial for understanding GBM growth and therapeutic resistance. Three-dimensional (3D) spheroid models provide a more physiologically relevant approach than traditional two-dimensional (2D) cultures, recapitulating key tumor features like hypoxia, cell heterogeneity, and drug resistance. This review examines scaffold-free and scaffold-based methods for generating 3D GBM spheroids, focusing on their applications in studying the cancer stem cell niche. The discussion encompasses methods such as the hanging drop, low-adhesion plates, and magnetic levitation, alongside advancements in embedding spheroids within extracellular matrix-based hydrogels and employing 3D bioprinting to fabricate more intricate tumor models. These 3D culture systems offer substantial potential for enhancing our understanding of GBM biology and devising more effective targeted therapies.

Cancer stem cells (CSCs) are a major hindrance to clinical cancer treatment. Owing to their high tumorigenic and metastatic potential, CSCs are vital in malignant tumor initiation, growth, metastasis, and therapeutic resistance, leading to tumorigenesis and recurrence. Compared with normal tumor cells, CSCs express high levels of surface markers (CD44, CD90, CD133, etc.) and activate specific signaling pathways (Wnt/β-catenin, Notch, and Hedgehog). Although Current drug delivery systems (DDS) precisely target CSCs, the heterogeneity and multidrug resistance of CSCs impede CSC isolation and screening. Conversely, hydrogel DDSs exhibit good biocompatibility and high drug delivery efficiency. Hydrogels are three-dimensional (3D) spatial structures for drug encapsulation that facilitate the controlled release of bioactive molecules. Hence, hydrogels can be loaded with drugs to precisely target CSCs. Their 3D structure can also culture non-CSCs and facilitate their transformation into CSCs. for identification and isolation. Given that their elastic modulus and stiffness characteristics reflect those of the cellular microenvironment, hydrogels can simulate extracellular matrix pathways and markers to regulate CSCs, disrupting the equilibrium between CSC and non-CSC transformation. This article reviews the CSC microenvironment, metabolism, signaling pathway, and surface markers. Additionally, we summarize the existing CSC targeting strategies and explore the application of hydrogels for CSC screening and treatment. Finally, we discuss potential advances in CSC research that may lead to curative measures for tumors through targeted and precise attacks on CSCs.

ALDH1A3 is a marker for mesenchymal glioblastomas characterized by a greater degree of aggressiveness compared to other major subtypes. ADH1A3 has been implicated in the regulation of stemness and radioresistance mediated by glioblastoma stem cells. Mechanisms by which ALDH1A3 promotes malignant progression of glioblastoma remain elusive posing a challenge for rationalization of ALDH1A3 targeting in glioblastoma, and it is also unclear how ALDH1A3 regulates glioblastoma cells stemness. Usage of different models with diverse genetic backgrounds and often unknown degree of stemness is one possible reason for discrepant views on the role of ALDH1A3 in glioblastoma stem cells. This study clarifies ALDH1A3 impacts on glioblastoma stem cells by modelling ALDH1A3 expression in an otherwise invariable genetic background with consideration of the impacts of inherent plasticity and proliferative changes associated with transitions between cell states. Our main finding is that ALDH1A3 exerts cell-state dependent impact on proliferation of glioblastoma stem cells. We provide evidence that ALDH1A3 augments radiation-induced inhibition of self-renewal and promotes the proliferation of differentiated GSC progenies. Congruent effects ALDH1A3 and radiation on self-renewal and proliferation provides a framework for promoting glioblastoma growth under radiation treatment.

Gliomas are one of the most aggressive types of brain tumors and are associated with high morbidity and mortality rates. Currently, conventional treatments for gliomas such as surgical resection, radiotherapy, and chemotherapy have limited effectiveness, and new approaches are needed to improve patient outcomes. mRNA-based vaccines represent a promising therapeutic strategy for cancer treatment, including gliomas. Recent advances in immunotherapy using mRNA-based dendritic cell vaccines have shown great potential in preclinical and clinical trials. Dendritic cells are professional antigen-presenting cells that play a crucial role in initiating and regulating immune responses. In this review, we summarize the current progress of mRNA-based vaccines for gliomas, with a focus on recent advances in dendritic cell-based mRNA vaccines. We also discuss the feasibility and safety of mRNA-based clinical applications for gliomas.

Glioblastoma multiforme (GBM) is a highly malignant brain tumor, and glioblastoma stem cells (GSCs) are the primary cause of GBM heterogeneity, invasiveness, and resistance to therapy. Sirtuin 3 (SIRT3) is mainly localized in the mitochondrial matrix and plays an important role in maintaining GSC stemness through cooperative interaction with the chaperone protein tumor necrosis factor receptor-associated protein 1 (TRAP1) to modulate mitochondrial respiration and oxidative stress. The present study aimed to further elucidate the specific mechanisms by which SIRT3 influences GSC stemness, including whether SIRT3 serves as an autophagy substrate and the mechanism of SIRT3 degradation. We first found that SIRT3 is enriched in CD133+ GSCs. Further experiments revealed that in addition to promoting mitochondrial respiration and reducing oxidative stress, SIRT3 maintains GSC stemness by epigenetically regulating CD133 expression via succinate. More importantly, we found that SIRT3 is degraded through the autophagy-lysosome pathway during GSC differentiation into GBM bulk tumor cells. GSCs are highly dependent on glutamine for survival, and in these cells, we found that glutamine deprivation triggers autophagic SIRT3 degradation to restrict CD133 expression, thereby disrupting the stemness of GSCs. Together our results reveal a novel mechanism by which SIRT3 regulates GSC stemness. We propose that glutamine restriction to trigger autophagic SIRT3 degradation offers a strategy to eliminate GSCs, which combined with other treatment methods may overcome GBM resistance to therapy as well as relapse.

Cancer stem cells (CSCs), a small subset of cells in tumors that are characterized by self-renewal and continuous proliferation, lead to tumorigenesis, metastasis, and maintain tumor heterogeneity. Cancer continues to be a significant global disease burden. In the past, surgery, radiotherapy, and chemotherapy were the main cancer treatments. The technology of cancer treatments continues to develop and advance, and the emergence of targeted therapy, and immunotherapy provides more options for patients to a certain extent. However, the limitations of efficacy and treatment resistance are still inevitable. Our review begins with a brief introduction of the historical discoveries, original hypotheses, and pathways that regulate CSCs, such as WNT/β-Catenin, hedgehog, Notch, NF-κB, JAK/STAT, TGF-β, PI3K/AKT, PPAR pathway, and their crosstalk. We focus on the role of CSCs in various therapeutic outcomes and resistance, including how the treatments affect the content of CSCs and the alteration of related molecules, CSCs-mediated therapeutic resistance, and the clinical value of targeting CSCs in patients with refractory, progressed or advanced tumors. In summary, CSCs affect therapeutic efficacy, and the treatment method of targeting CSCs is still difficult to determine. Clarifying regulatory mechanisms and targeting biomarkers of CSCs is currently the mainstream idea.

Glioblastoma (GBM), the most aggressive and fatal brain malignancy, is largely driven by a subset of tumor cells known as cancer stem cells (CSCs). CSCs possess stem cell-like properties, including self-renewal, proliferation, and differentiation, making them pivotal for tumor initiation, invasion, metastasis, and overall tumor progression. The regulation of CSCs is primarily controlled by transcription factors (TFs) which regulate the expressions of genes involved in maintaining stemness and directing differentiation. This review aims to provide a comprehensive overview of the role of TFs in regulating CSCs in GBM. The discussion encompasses the definitions of CSCs and TFs, the significance of glioma stem cells (GSCs) in GBM, and how TFs regulate GSC self-renewal, proliferation, differentiation, and transformation. The potential for developing TF-targeted GSC therapies is also explored, along with future research directions. By understanding the regulation of GSCs by TFs, we may uncover novel diagnostic and therapeutic strategies against this devastating disease of GBM.

CD133, a cancer stem cell (CSC) marker in tumors, including melanoma, is associated with tumor recurrence, chemoresistance, and metastasis. Patient-derived melanoma cell lines were transduced with a Tet-on vector expressing CD133, generating doxycycline (Dox)-inducible cell lines. Cells were exposed to Dox for 24 h to induce CD133 expression, followed by RNA-seq and bioinformatic analyses, revealing genes and pathways that are significantly up- or downregulated by CD133. The most significantly upregulated gene after CD133 was amphiregulin (AREG), validated by qRT-PCR and immunoblot analyses. Induced CD133 expression significantly increased cell growth, percentage of cells in S-phase, BrdU incorporation into nascent DNA, and PCNA levels, indicating that CD133 stimulates cell proliferation. CD133 induction also activated EGFR and the MAPK pathway. Potential mechanisms highlighting the role(s) of CD133 and AREG in melanoma CSC were further delineated using AREG/EGFR inhibitors or siRNA knockdown of AREG mRNA. Treatment with the EGFR inhibitor gefitinib blocked CD133-induced cell growth increase and MAPK pathway activation. Importantly, siRNA knockdown of AREG reversed the stimulatory effects of CD133 on cell growth, indicating that AREG mediates the effects of CD133 on cell proliferation, thus serving as an attractive target for novel combinatorial therapeutics in melanoma and cancers with overexpression of both CD133 and AREG.

Cancer treatments are advancing to harness the body's immune system against tumours, aiming for lasting effects. This progress involves combining potent chemotherapy drugs with immunogens to kill cancer cells and trigger lasting immunity. Developing new prodrugs that integrate both chemotherapy and immune-boosting elements could significantly improve anticancer outcomes by activating multiple mechanisms to kill cancer cells. While bacterial polysaccharides are typically not used in therapy due to their immune-stimulating properties, we propose a safe application of an extremophilic bacterial polysaccharide, Mauran (MR), modified with the anticancer drug 5-fluorouracil (5FU) to create a novel prodrug. This obtained prodrug, chloracetyl-MR-5FU, is specifically targeted using gold nanocages to CD133+ glioma cells. Test results have shown a high encapsulation efficiency of the drug during the polysaccharide modification process; its anticancer activity was demonstrated in vitro and the release of the prodrug was demonstrated in ex vivo studies.

Prominin-1 (CD133) is a cholesterol-binding membrane glycoprotein selectively associated with highly curved and prominent membrane structures. It is widely recognized as an antigenic marker of stem cells and cancer stem cells and is frequently used to isolate them from biological and clinical samples. Recent progress in understanding various aspects of CD133 biology in different cell types has revealed the involvement of CD133 in the architecture and dynamics of plasma membrane protrusions, such as microvilli and cilia, including the release of extracellular vesicles, as well as in various signaling pathways, which may be regulated in part by posttranslational modifications of CD133 and its interactions with a variety of proteins and lipids. Hence, CD133 appears to be a master regulator of cell signaling as its engagement in PI3K/Akt, Src-FAK, Wnt/β-catenin, TGF-β/Smad and MAPK/ERK pathways may explain its broad action in many cellular processes, including cell proliferation, differentiation, and migration or intercellular communication. Here, we summarize early studies on CD133, as they are essential to grasp its novel features, and describe recent evidence demonstrating that this unique molecule is involved in membrane dynamics and molecular signaling that affects various facets of tissue homeostasis and cancer development. We hope this review will provide an informative resource for future efforts to elucidate the details of CD133's molecular function in health and disease.

Glioblastoma multiform (GBM) is among the commonly diagnosed brain malignancies with poor prognosis. CD133 has been introduced as an oncogene in various cancers, like GBM. This study aimed to investigate the significance of CD133 in GBM development using in silico and in vitro techniques.

The TCGA-GBM database was analyzed for the correlational and comparative studies. After selecting the U87MG cell line, CD133-siRNA was transfected into U87MG cells and treated with temozolomide. The cell viability, cell cycle, migration, clonogenicity, and apoptosis of groups were investigated using MTT, flow cytometry, wound-healing, colony formation, and annexin V/PI assays. Using qRT-PCR method, the mRNA expression levels of MMP16, SOX2, RAF1, MAP2K1, MAPK3, PIK3CA, AKT3, mTOR, CDK4, and BCL2 were studied.

CD133 silencing improves apoptosis rate, arrests the cell cycle at the sub-G1 phase, suppresses the clonogenicity of U87MG cells, and inhibits the PI3K/Akt and MAPK pathways via downregulating the RAF1, MAP2K1, MAPK3, PIK3CA, AKT3, and mTOR expression. Besides, combining CD133 silencing with temozolomide treatment considerably inhibits the migration of U87MG cells compared to temozolomide monotherapy.

CD133 can regulate the PI3K/Akt and MAPK pathways and modulate the clonogenicity, apoptosis, and cell cycle of GBM. Combining CD133 silencing with temozolomide treatment considerably increases apoptosis, arrests the cell cycle at the sub-G1, and suppresses migration of U87MG cells compared to temozolomide monotherapy.

Glioblastoma multiforme (GBM) is the most common and deadly type of brain tumor originating from glial cells. Despite decades of clinical trials and research, there has been limited success in improving survival rates. However, molecular pathology studies have provided a detailed understanding of the genetic alterations associated with the formation and progression of glioblastoma-such as Kirsten rat sarcoma viral oncogene homolog (KRAS) signaling activation (5%), P53 mutations (25%), and adenomatous polyposis coli (APC) alterations (2%)-laying the groundwork for further investigation into the biological and biochemical basis of this malignancy. These analyses have been crucial in revealing the sequential appearance of specific genetic lesions at distinct histopathological stages during the development of GBM. To further explore the pathogenesis and progression of glioblastoma, here, we developed the glial-fibrillary-acidic-protein (GFAP)-Cre-driven mouse model and demonstrated that activated KRAS and p53 deficiencies play distinct and cooperative roles in initiating glioma tumorigenesis. Additionally, the combination of APC haploinsufficiency with mutant Kras activation and p53 deletion resulted in the rapid progression of GBM, characterized by perivascular inflammation, large necrotic areas, and multinucleated giant cells. Consequently, our GBM models have proven to be invaluable resources for identifying early disease biomarkers in glioblastoma, as they closely mimic the human disease. The insights gained from these models may pave the way for potential advancements in the diagnosis and treatment of this challenging brain tumor.

Glioblastoma (GBM) is a devastating cancer of the brain with an extremely poor prognosis. While X-ray radiotherapy and chemotherapy remain the current standard, proton beam therapy is an appealing alternative as protons can damage cancer cells while sparing the surrounding healthy tissue. However, the effects of protons on in vitro GBM models at the cellular level, especially when co-cultured with endothelial cells, the building blocks of brain micro-vessels, are still unexplored. In this work, novel 3D-engineered scaffolds inspired by the geometry of brain microvasculature are designed, where GBM cells cluster and proliferate. The architectures are fabricated by two-photon polymerization (2PP), pre-cultured with endothelial cells (HUVECs), and then cultured with a human GBM cell line (U251). The micro-vessel structures enable GBM in vivo-like morphologies, and the results show a higher DNA double-strand breakage in GBM monoculture samples when compared to the U251/HUVECs co-culture, with cells in 2D featuring a larger number of DNA damage foci when compared to cells in 3D. The discrepancy in terms of proton radiation response indicates a difference in the radioresistance of the GBM cells mediated by the presence of HUVECs and the possible induction of stemness features that contribute to radioresistance and improved DNA repair.

Glioblastoma multiforme (GBM) is regarded as a highly aggressive brain cancer with a poor prognosis. There is an increase in the expression of P-glycoprotein (P-gp), responsible for multidrug resistance (MDR), making it a potential target for improving drug responses. Additionally, glioblastoma stem cells (GSCs) increase resistance to chemo- and radiotherapy and play a major role in cancer relapse. In this study, we targeted P-gp using a small molecule inhibitor, reversan (RV), to inhibit MDR that prolonged the retention of drugs in the cytosolic milieu. To eliminate GBM and GSCs, we have used two well-established anti-cancer drugs, regorafenib (RF) and curcumin (CMN). To improve the pharmacokinetics and decrease systemic delivery of drugs, we developed nanostructure hybrid lipid capsules (nHLCs), where hydrophobic drugs can be loaded in the core, and their physicochemical properties were determined by dynamic light scattering (DLS) and cryo-scanning electron microscopy (SEM). Inhibition of MDR by RV has also shown enhanced retention of nHLC in GBM cells. Co-delivery of drug-loaded nHLCs, pre-treated with RV, exhibited superior cytotoxicity in both GBM and GSCs than their individual doses and effectively reduced the size and stemness of tumor spheres and accelerated the rate of apoptosis, suggesting a promising treatment for glioblastoma.

Despite maximally safe resection of the magnetic resonance imaging (MRI)-defined contrast-enhanced (CE) central tumor area and chemoradiotherapy, most patients with glioblastoma (GBM) relapse within a year in peritumoral FLAIR regions. Magnetic resonance spectroscopy imaging (MRSI) can discriminate metabolic tumor areas with higher recurrence potential as CNI+ regions (choline/N-acetyl-aspartate index >2) can predict relapse sites. As relapses are mainly imputed to glioblastoma stem-like cells (GSCs), CNI+ areas might be GSC enriched. In this prospective trial, 16 patients with GBM underwent MRSI/MRI before surgery/chemoradiotherapy to investigate GSC content in CNI-/+ biopsies from CE/FLAIR. Biopsy and derived-GSC characterization revealed a FLAIR/CNI+ sample enrichment in GSC and in gene signatures related to stemness, DNA repair, adhesion/migration, and mitochondrial bioenergetics. FLAIR/CNI+ samples generate GSC-enriched neurospheres faster than FLAIR/CNI-. Parameters assessing biopsy GSC content and time-to-neurosphere formation in FLAIR/CNI+ were associated with worse patient outcome. Preoperative MRI/MRSI would certainly allow better resection and targeting of FLAIR/CNI+ areas, as their GSC enrichment can predict worse outcomes.

Malignant brain tumors are highly heterogeneous tumors with a poor prognosis and a high morbidity and mortality rate in both children and adults. The cancer stem cell (CSC, also named tumor-initiating cell) model states that tumor growth is driven by a subset of CSCs. This model explains some of the clinical observations of brain tumors, including the almost unavoidable tumor recurrence after initial successful chemotherapy and/or radiotherapy and treatment resistance. Over the past two decades, strategies for the identification and characterization of brain CSCs have improved significantly, supporting the design of new diagnostic and therapeutic strategies for brain tumors. Relevant studies have unveiled novel characteristics of CSCs in the brain, including their heterogeneity and distinctive immunobiology, which have provided opportunities for new research directions and potential therapeutic approaches. In this review, we summarize the current knowledge of CSCs markers and stemness regulators in brain tumors. We also comprehensively describe the influence of the CSCs niche and tumor microenvironment on brain tumor stemness, including interactions between CSCs and the immune system, and discuss the potential application of CSCs in brain-based therapies for the treatment of brain tumors.

Since their discovery at the beginning of this millennium, glioma stem cells (GSCs) have sparked extensive research and an energetic scientific debate about their contribution to glioblastoma (GBM) initiation, progression, relapse, and resistance. Different molecular subtypes of GBM coexist within the same tumor, and they display differential sensitivity to chemotherapy. GSCs contribute to tumor heterogeneity and recapitulate pathway alterations described for the three GBM subtypes found in patients. GSCs show a high degree of plasticity, allowing for interconversion between different molecular GBM subtypes, with distinct proliferative potential, and different degrees of self-renewal and differentiation. This high degree of plasticity permits adaptation to the environmental changes introduced by chemo- and radiation therapy. Evidence from mouse models indicates that GSCs repopulate brain tumors after therapeutic intervention, and due to GSC plasticity, they reconstitute heterogeneity in recurrent tumors. GSCs are also inherently resilient to standard-of-care therapy, and mechanisms of resistance include enhanced DNA damage repair, MGMT promoter demethylation, autophagy, impaired induction of apoptosis, metabolic adaptation, chemoresistance, and immune evasion. The remarkable oncogenic properties of GSCs have inspired considerable interest in better understanding GSC biology and functions, as they might represent attractive targets to advance the currently limited therapeutic options for GBM patients. This has raised expectations for the development of novel targeted therapeutic approaches, including targeting GSC plasticity, chimeric antigen receptor T (CAR T) cells, and oncolytic viruses. In this review, we focus on the role of GSCs as drivers of GBM and therapy resistance, and we discuss how insights into GSC biology and plasticity might advance GSC-directed curative approaches.

Glioma is the most common primary brain tumor. Glioma stem cells (GSCs) are the origin of gliomagenesis and may develop from normal neural progenitor cells (NPCs). However, how neoplastic transformation occurs in normal NPCs and the role of the Ras/Raf/MAPK pathway in NPC transformation is unclear. The present study generated NPCs from human embryonic stem cells (ESCs) carrying gene alterations in the Ras/Raf/MAPK pathway. The CCK‑8 proliferation, single‑cell clonal expansion, cell migration, RT‑qPCR, immunofluorescence staining, western blotting, transcriptome and Seahorse analyses, and intracranial implantation assay were performed to identify the characterization of transformed NPCs in vitro and in vivo. Brain organoids were used to verify the phenotypes transforming in NPCs. KRAS‑activated NPCs exhibited increased proliferation and migration in vitro. KRAS‑activated NPCs showed atypical morphology and formed aggressive tumors in immunodeficient mice. At the molecular level, KRAS‑activated NPCs displayed neoplasm‑associated metabolic and gene expression profiles. Moreover, activation of KRAS led to substantial cell proliferation and abnormal structure in ESC‑derived brain organoids. The present study showed that activated KRAS transformed normal NPCs to GSC‑like cells and established a simple cellular model to investigate gliomagenesis.

Boron neutron capture therapy (BNCT) is a high-LET particle radiotherapy clinically tested for treating malignant gliomas. Boronophenylalanine (BPA), a boron-containing phenylalanine derivative, is selectively transported into tumor cells by amino acid transporters, making it an ideal agent for BNCT. In this study, we investigated whether the amino acid 5-aminolevulinic acid (ALA) could sensitize glioma stem cells (GSCs) to BNCT by enhancing the uptake of BPA. Using human and mouse GSC lines, pre-incubation with ALA increased the intracellular accumulation of BPA dose-dependent. We also conducted in vivo experiments by intracerebrally implanting HGG13 cells in mice and administering ALA orally 24 h before BPA administration (ALA + BPA-BNCT). The ALA preloading group increased the tumor boron concentration and improved the tumor/blood boron concentration ratio, resulting in improved survival compared to the BPA-BNCT group. Furthermore, we found that the expression of amino acid transporters was upregulated following ALA treatment both in vitro and in vivo, particularly for ATB0,+. This suggests that ALA may sensitize GSCs to BNCT by upregulating the expression of amino acid transporters, thereby enhancing the uptake of BPA and improving the effectiveness of BNCT. These findings have important implications for strategies to improve the sensitivity of malignant gliomas to BPA-BNCT.

Owing to the strong absorption of water in the near-infrared (NIR) region near 1.0 μm, this wavelength is considered unsuitable as an imaging and analytical signal in biological environments. However, 1.0 μm NIR can be converted into heat and used as a local water-molecular heating strategy for the photothermal therapy of biological tissues. Herein, we describe a Nd-Yb co-doped nanomaterial (water-heating nanoparticles (NPs)) as strong 1.0 μm emissive NPs to target the absorption band of water. Furthermore, introducing Tm ions into the water-heating NPs improve the NIR lifetime, enabling the development of a NIR imaging-guided water-heating probe (water-heating NIR NPs). In the glioblastoma multiforme male mouse model, tumor-targeted water-heating NIR NPs reduce the tumor volume by 78.9% in the presence of high-resolution intracranial NIR long-lifetime imaging. Hence, water-heating NIR NPs can be used as a promising nanomaterial for imaging and photothermal ablation in deep-tissue-bearing tumor therapy.

Targeting glioblastoma (GBM) stem-like cells (GSCs) is a common interest in both the laboratory investigation and clinical treatment of GBM. Most of the currently applied GBM stem-like markers lack validation and comparison with common standards regarding their efficiency and feasibility in various targeting methods. Using single-cell RNA sequencing datasets from 37 GBM patients, we obtained a large pool of 2173 GBM stem-like marker candidates. To evaluate and select these candidates quantitatively, we characterized the efficiency of the candidate markers in targeting the GBM stem-like cells by their frequencies and significance of being the stem-like cluster markers. This was followed by further selection based on either their differential expression in GBM stem-like cells compared with normal brain cells or their relative expression level compared with other expressed genes. The cellular location of the translated protein was also considered. Different combinations of selection criteria highlight different markers for different application scenarios. By comparing the commonly used GSCs marker CD133 (PROM1) with markers selected by our method regarding their universality, significance, and abundance, we revealed the limitations of CD133 as a GBM stem-like marker. Overall, we propose BCAN, PTPRZ1, SOX4, etc. for laboratory-based assays with samples free of normal cells. For in vivo targeting applications that require high efficiency in targeting the stem-like subtype, the ability to distinguish GSCs from normal brain cells, and a high expression level, we recommend the intracellular marker TUBB3 and the surface markers PTPRS and GPR56.

Glioblastoma stem cells (GSCs) are subpopulations of tumor-initiating cells responsible for glioblastoma (GBM) tumorigenesis and recurrence. Dual inhibition of vascular endothelium and GSCs is still a challenge due to their different pathological features. Here we present a combined all-in-control strategy to realize a local photothermal therapy (PTT). We designed T-cell-mimic nanoparticles with aggregation-induced emission (AIE) characteristics by coating the genetically engineered T cell membrane (CM) onto AIE nanoparticles (CM@AIE NPs). The CM shell was designed against CD133 and epidermal growth factor receptor (EGFR) which provides the possibility to target both GBM cells and GSCs for cancer therapy. CM@AIE NPs can serve as the tight junction (TJ) modulators to trigger an intracellular signaling cascade, causing TJ disruption and actin cytoskeleton reorganization to allow CM@AIE NPs to cross the blood-brain barrier (BBB) silently. The 980 nm excitation-triggered PTT can completely inhibit tumorigenesis and recurrence. The combination of CM-coating nanotechnology and genetic editing technique can inspire further development of synergetic techniques for preventing GBM recurrence.

Glioblastoma (GBM), classified as a grade IV glioma, is a rapidly growing, aggressive, and most commonly occurring tumor of the central nervous system. Despite the therapeutic advances, it carries an ominous prognosis, with a median survival of 14.6 months after diagnosis. Accumulating evidence suggests that cancer stem cells in GBM, termed glioma stem cells (GSCs), play a crucial role in tumor propagation, treatment resistance, and tumor recurrence. GSCs, possessing the capacity for self-renewal and multilineage differentiation, are responsible for tumor growth and heterogeneity, leading to primary obstacles to current cancer therapy. In this respect, increasing efforts have been devoted to the development of anti-GSC strategies based on targeting GSC surface markers, blockage of essential signaling pathways of GSCs, and manipulating the tumor microenvironment (GSC niches). In this review, we will discuss the research knowledge regarding GSC-based therapy and the underlying mechanisms for the treatment of GBM. Given the rapid progression in nanotechnology, innovative nanomedicines developed for GSC targeting will also be highlighted from the perspective of rationale, advantages, and limitations. The goal of this review is to provide broader understanding and key considerations toward the future direction of GSC-based nanotheranostics to fight against GBM.

Exosomes participate in intercellular communication by transporting functionally active molecules. Such cargo from the original cells comprising proteins, micro-RNA, mRNA, single-stranded (ssDNA) and double-stranded DNA (dsDNA) molecules pleiotropically transforms the target cells. Although cancer cells secrete exosomes carrying a significant level of DNA capable of modulating oncogene expression in a recipient cell, the regulatory mechanism is unknown. We have previously reported that cancer cells produce exosomes containing NANOGP8 DNA. NANOGP8 is an oncogenic paralog of embryonic stem cell transcription factor NANOG and does not express in cells since it is a pseudogene. However, in this study, we evaluated NANOGP8 expression in glioblastoma multiforme (GBM) tissue from a surgically removed brain tumor of a patient. Significantly higher NANOGP8 transcription was observed in GBM cancer stem cells (CSCs) than in GBM cancer cells or neural stem cells (NSCs), despite identical sequences of NANOGP8-upstream genomic region in all the cell lines. This finding suggests that upstream genomic sequences of NANOGP8 may have environment-dependent promoter activity. We also found that the regulatory sequences upstream of exosomal NANOGP8 GBM DNA contain multiple core promoter elements, transcription factor binding sites, and segments of human viruses known for their oncogenic role. The exosomal sequence of NANOGP8-upstream GBM DNA is different from corresponding genomic sequences in CSCs, cancer cells, and NSCs as well as from the sequences reported by NCBI. These sequence dissimilarities suggest that exosomal NANOGP8 GBM DNA may not be a part of the genomic DNA. Exosomes possibly acquire this DNA from other sources where it is synthesized by an unknown mechanism. The significance of exosome-bestowed regulatory elements in the transcription of promoter-less retrogene such as NANOGP8 remains to be determined.

Spermatogenesis is a complex process orchestrated by several essential genes. Prominin-1 (Prom1/PROM1) is a gene that is expressed in the testis but with a poorly understood role in spermatogenesis.

We used Prom1 knockout (Prom1 KO) mice to assess the role of Prom1 in spermatogenesis. To this end, we performed immunohistochemistry, immunofluorescence, western blotting, β-galactosidase staining, and apoptosis assay. Additionally, we analyzed the morphology of sperm and assessed litter sizes.

We observed that PROM1 is localized to the dividing spermatocytes in seminiferous epithelial cells, sperm, and columnar epithelium in the epididymis. In the Prom1 KO testis, an aberrant increase in apoptotic cells and a decrease in proliferating seminiferous epithelial cells were observed. Cellular FLICE-like inhibitory protein (c-FLIP) and extracellular signal-regulated kinase 1/2 (ERK1/2) expression were also significantly decreased in Prom1 KO testis. In addition, a significantly increased number of epididymal spermatozoa with abnormal morphology and less motility was found in Prom1 KO mice.

PROM1 maintains spermatogenic cell proliferation and survival via c-FLIP expression in the testis. It is also involved in sperm motility and fertilization potential. The mechanism underlying the effect of Prom1 on sperm morphology and motility remains to be identified.

Cancer is not a hard-wired phenomenon but an evolutionary disease. From the onset of carcinogenesis, cancer cells continuously adapt and evolve to satiate their ever-growing proliferation demands. This results in the formation of multiple subtypes of cancer cells with different phenotypes, cellular compositions, and consequently displaying varying degrees of tumorigenic identity and function. This phenomenon is referred to as cancer plasticity, during which the cancer cells exist in a plethora of cellular states having distinct phenotypes. With the advent of modern technologies equipped with enhanced resolution and depth, for example, single-cell RNA-sequencing and advanced computational tools, unbiased cancer profiling at a single-cell resolution are leading the way in understanding cancer cell rewiring both spatially and temporally. In this review, the processes and mechanisms that give rise to cancer plasticity include both intrinsic genetic factors such as epigenetic changes, differential expression due to changes in DNA, RNA, or protein content within the cancer cell, as well as extrinsic environmental factors such as tissue perfusion, extracellular milieu are detailed and their influence on key cancer plasticity hallmarks such as epithelial-mesenchymal transition (EMT) and cancer cell stemness (CSCs) are discussed. Due to therapy evasion and drug resistance, tumor heterogeneity caused by cancer plasticity has major therapeutic ramifications. Hence, it is crucial to comprehend all the cellular and molecular mechanisms that control cellular plasticity. How this process evades therapy, and the therapeutic avenue of targeting cancer plasticity must be diligently investigated.

Glioblastoma multiforme (GBM) is the most aggressive and most common malignant brain tumor with poor patient survival despite therapeutic intervention. On the cellular level, GBM comprises a rare population of glioblastoma stem cells (GSCs), driving therapeutic resistance, invasion, and recurrence. GSCs have thus come into the focus of therapeutic strategies, although their targeting remains challenging. In the present study, we took advantage of three GSCs-populations recently established in our lab to investigate key signaling pathways and subsequent therapeutic strategies targeting GSCs. We observed that NF-κB, a crucial transcription factor in GBM progression, was expressed in all CD44+/CD133+/Nestin+-GSC-populations. Exposure to TNFα led to activation of NF-κB-RELA and/or NF-κB-c-REL, depending on the GBM type. GSCs further expressed the proto-oncogene MYC family, with MYChigh GSCs being predominantly located in the tumor spheres ("GROW"-state) while NF-κB-RELAhigh GSCs were migrating out of the sphere ("GO"-state). We efficiently targeted GSCs by the pharmacologic inhibition of NF-κB using PTDC/Bortezomib or inhibition of MYC by KJ-Pyr-9, which significantly reduced GSC-viability, even in comparison to the standard chemotherapeutic drug temozolomide. As an additional cell-therapeutic strategy, we showed that NK cells could kill GSCs. Our findings offer new perspectives for developing efficient patient-specific chemo- and immunotherapy against GBM.

Glioma stem cells (GSC) exhibit plasticity in response to environmental and therapeutic stress leading to tumor recurrence, but the underlying mechanisms remain largely unknown. Here, we employ single-cell and whole transcriptomic analyses to uncover that radiation induces a dynamic shift in functional states of glioma cells allowing for acquisition of vascular endothelial-like and pericyte-like cell phenotypes. These vascular-like cells provide trophic support to promote proliferation of tumor cells, and their selective depletion results in reduced tumor growth post-treatment in vivo. Mechanistically, the acquisition of vascular-like phenotype is driven by increased chromatin accessibility and H3K27 acetylation in specific vascular genes allowing for their increased expression post-treatment. Blocking P300 histone acetyltransferase activity reverses the epigenetic changes induced by radiation and inhibits the adaptive conversion of GSC into vascular-like cells and tumor growth. Our findings highlight a role for P300 in radiation-induced stress response, suggesting a therapeutic approach to prevent glioma recurrence.

The quiescent/slow-cycling state preserves the self-renewal capacity of cancer stem cells (CSCs) and leads to the therapy resistance of CSCs. The mechanisms maintaining CSCs quiescence remain largely unknown. Here, it is demonstrated that lower expression of MAN1A1 in glioma stem cell (GSC) resulted in the formation of high-mannose type N-glycan on CD133. Furthermore, the high-mannose type N-glycan of CD133 is necessary for its interaction with DNMT1. Activation of p21 and p27 by the CD133-DNMT1 interaction maintains the slow-cycling state of GSC, and promotes chemotherapy resistance and tumorigenesis of GSCs. Elimination of the CD133-DNMT1 interaction by a cell-penetrating peptide or MAN1A1 overexpression inhibits the tumorigenesis of GSCs and increases the sensitivity of GSCs to temozolomide. Analysis of glioma samples reveals that the levels of high-mannose type N-glycan are correlated with glioma recurrence. Collectively, the high mannose CD133-DNMT1 interaction maintains the slow-cycling state and tumorigenic potential of GSC, providing a potential strategy to eliminate quiescent GSCs.

Glioblastoma is one of the most aggressive types of cancer with success of therapy being hampered by the existence of treatment resistant populations of stem-like Tumour Initiating Cells (TICs) and poor blood-brain barrier drug penetration. Therapies capable of effectively targeting the TIC population are in high demand. Here, we synthesize spherical diketopyrrolopyrrole-based Conjugated Polymer Nanoparticles (CPNs) with an average diameter of 109 nm. CPNs were designed to include fluorescein-conjugated Hyaluronic Acid (HA), a ligand for the CD44 receptor present on one population of TICs. We demonstrate blood-brain barrier permeability of this system and concentration and cell cycle phase-dependent selective uptake of HA-CPNs in CD44 positive GBM-patient derived cultures. Interestingly, we found that uptake alone regulated the levels and signaling activity of the CD44 receptor, decreasing stemness, invasive properties and proliferation of the CD44-TIC populations in vitro and in a patient-derived xenograft zebrafish model. This work proposes a novel, CPN- based, and surface moiety-driven selective way of targeting of TIC populations in brain cancer.