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Tumor-initiating cells (TICs) play a key role in cancer progression and immune escape. However, how TICs evade immune elimination remains poorly characterized. Combining single-cell RNA sequencing (scRNA-seq), dual-recombinase-based lineage tracing, and other approaches, we identified a WNT-activated subpopulation of malignant cells that act as TICs in vivo. We found intensive reciprocal interactions between TICs and immune regulatory tumor-associated macrophages (Reg-TAMs) via GAS6-AXL/MERTK signaling pathways, which facilitated the immune escape of TICs. Our study employed chemical inhibitors and Axl/Mertk conditional double knockout mice to demonstrate that inhibiting the interaction between TIC-derived GAS6 and AXL/MERTK in Reg-TAMs reactivated anti-tumor immune responses. We identified CCL8 as a critical mediator of the GAS6/AXL/MERTK pathway, primarily by inhibiting regulatory T cell (Treg) infiltration into the tumor. Furthermore, the AXL/MERTK signaling blockade sensitized tumor cells to anti-PD-1 treatment. Thus, we elucidated a detailed mechanism by which TICs evade tumor immunity, providing insights into strategies to eradicate TICs that escape conventional immunotherapy.

Abstract Tumor initiation represents the first step in tumorigenesis during which normal progenitor cells undergo cell fate transition to cancer. Capturing this process as it occurs in vivo, however, remains elusive. Here we employ spatiotemporally controlled oncogene activation and tumor suppressor inhibition together with multiomics to unveil the processes underlying oral epithelial progenitor cell reprogramming into tumor initiating cells at single cell resolution. Tumor initiating cells displayed a distinct stem-like state, defined by aberrant proliferative, hypoxic, squamous differentiation, and partial epithelial to mesenchymal invasive gene programs. YAP-mediated tumor initiating cell programs included activation of oncogenic transcriptional networks and mTOR signaling, and recruitment of myeloid cells to the invasive front contributing to tumor infiltration. Tumor initiating cell transcriptional programs are conserved in human head and neck cancer and associated with poor patient survival. These findings illuminate processes underlying cancer initiation at single cell resolution, and identify candidate targets for early cancer detection and prevention.

Immunotherapy has become a key new pillar of cancer treatment, and this has sparked interest in understanding mechanisms of cancer immune evasion. It has long been appreciated that cancers are constituted by heterogeneous populations of tumour cells. This feature is often fuelled by specialized cells that have molecular programs resembling tissue stem cells. Although these cancer stem cells (CSCs) have capacity for unlimited self-renewal and differentiation, it is increasingly evident that some CSCs are capable of achieving remarkable immune resistance. Given that most immunotherapy regiments have overlooked CSC-specific immune-evasive mechanisms, many current treatment strategies often lead to cancer relapse. This Review focuses on advancements in understanding how CSCs in solid tumours achieve their unique immune-evasive properties, enabling them to drive tumour regrowth. Moreover, as cancers often arise from tissue stem cells that acquired oncogenic mutations, we discuss how tissue stem cells undergoing malignant transformation activate intrinsic immune-evasive mechanisms and establish close interactions with suppressive immune cells to escape immune surveillance. In addition, we summarize how in advanced disease stages, CSCs often hijack features of normal stem cells to resist antitumour immunity. Finally, we provide insights in how to design a new generation of cancer immunotherapies to ensure elimination of CSCs., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. Springer Nature Limited.)

[Display omitted] • Theoretical in-silico modeling was used to predict in vitro radiation cytotoxicity. • Advanced imaging strategies were used to assess radiosensitivity longitudinally. • Radiation dose order differentially impacts radioresistant and radiosensitive PDX. • Ramping down dose induces cytotoxicity in all PDX with differential radiosensitivity. Current radiotherapy (RT) in glioblastoma (GBM) is delivered as constant dose fractions (CDF), which do not account for intratumoral-heterogeneity and radio-selection in GBM. These factors contribute to differential treatment response complicating the therapeutic efficacy of this principle. Our study aims to investigate an alternative dosing strategy to overcome radio-resistance using a novel longitudinal radiation cytotoxicity assay. Theoretical In-silico mathematical assumptions were combined with an in-vitro experimental strategy to investigate alternative radiation regimens. Patient-derived xenograft (PDX) brain tumor-initiating cells (BTICs) with differential radiation-sensitivities were tested individually with sham control and three regimens of the same nominal and average dose of 16 Gy (over four fractions), but with altered doses per fraction. Fractions were delivered conventionally (CDF: 4, 4, 4, 4 Gy), or as dynamic dose fractions (DDF) "ramped down" (RD: 7, 5, 3, 1 Gy), or DDF "ramped up" (RU: 1, 3, 5, 7 Gy), every 4 days. Interfraction-longitudinal data were collected by imaging cells every 5 days, and endpoint viability was taken on day 20. The proposed method of radiosensitivity assessment allows for longitudinal-interfraction investigation in addition to endpoint analysis. Delivering four-fraction doses in an RD manner proves to be most effective at overcoming acquired radiation resistance in BTICs (Relative cell viability: CDF vs. RD: P < 0.0001; Surviving fraction: CDF: vs. RD: P < 0.0001). Using in-silico cytotoxicity prediction modeling and an altered radiosensitivity assessment, we show DDF-RD is effective at inducing cytotoxicity in three BTIC lines with differential radiosensitivity. [ABSTRACT FROM AUTHOR]

Pancreatic intraepithelial neoplasia (PanIN) is the most common precursor lesion of pancreatic ductal adenocarcinoma (PDAC), which has poor prognosis with a short median overall survival of 6-12 months and a low 5-year survival rate of approximately 3%. It is crucial to remove PanIN lesions to prevent the development of invasive PDAC, as PDAC spreads rapidly outside the pancreas. This review aims to provide the latest knowledge on PanIN risk, pathology, cellular origin, genetic susceptibility, and diagnosis, while identifying research gaps that require further investigation in this understudied area of precancerous lesions. PanINs are classified into PanIN 1, PanIN 2, and PanIN 3, with PanIN 3 having the highest likelihood of developing into invasive PDAC. Differentiating between PanIN 2 and PanIN 3 is clinically significant. Genetic alterations found in PDAC are also present in PanIN and increase with the grade of PanIN. Imaging methods alone are insufficient for distinguishing PanIN, necessitating the use of genetic and molecular tests for identification. In addition, metabolomics technologies and miRNAs are playing an increasingly important role in the field of cancer diagnosis, offering more possibilities for efficient identification of PanIN. Although detecting and stratifying the risk of PanIN poses challenges, the combined utilization of imaging, genetics, and metabolomics holds promise for improving patient survival in this field. [ABSTRACT FROM AUTHOR]

Purpose of review: The role of the microbiome in prostate cancer is an emerging subject of research interest. Certain lifestyle factors, such as obesity and diet, can also impact the microbiome, which has been implicated in many diseases, such as heart disease and diabetes. However, this link has yet to be explored in detail in the context of prostate cancer. The purpose of this review is to explore the cross-talk between obesity, dietary interventions, and microbiome alterations in the development and progression of prostate cancer. Recent findings: Many possible mechanisms exist linking obesity and dietary interventions to microbiome alterations and prostate cancer. The gut microbiome produces metabolites that could play a role in prostate cancer oncogenesis, including short-chain fatty acids, cholesterol derivatives, and folic acid. The microbiome also plays a pivotal role in the prostate tumor microenvironment (TME), contributing to inflammation, local tissue hypoxia, and epithelial-mesenchymal transition. A bidirectional relationship exists between obesity and the microbiome, and certain diets can enact changes to the microbiome, its associated metabolites, and prostate cancer outcomes. Summary: Cross-talk exists between obesity, dietary interventions, and the role of the microbiome in the development and progression of prostate cancer. To further our understanding, future human studies in prostate cancer should investigate microbiome changes and incorporate an assessment of microbiome-derived metabolites and cellular/immune changes in the TME. [ABSTRACT FROM AUTHOR]

Background: Growing evidence shows that the reprogramming of fatty acid (FA) metabolism plays a key role in HER2-positive (HER2 +) breast cancer (BC) aggressiveness, therapy resistance and cancer stemness. In particular, HER2 + BC has been defined as a "lipogenic disease" due to the functional and bi-directional crosstalk occurring between HER2-mediated oncogenic signaling and FA biosynthesis via FA synthase activity. In this context, the functional role exerted by the reprogramming of CD36-mediated FA uptake in HER2 + BC poor prognosis and therapy resistance remains unclear. In this study, we aimed to elucidate whether enhanced CD36 in mesenchymal HER2 + cancer stem cells (CSCs) is directly involved in anti-HER2 treatment refractoriness in HER2 + BC and to design future metabolism-based approaches targeting both FA reprogramming and the “root” of cancer. Methods: Molecular, biological and functional characterization of CD36-mediated FA uptake was investigated in HER2 + BC patients, cell lines, epithelial and mesenchymal CSCs. Cell proliferation was analyzed by SRB assay upon treatment with lapatinib, CD36 inhibitor, or Wnt antagonist/agonist. Engineered cell models were generated via lentivirus infection and transient silencing. CSC-like properties and tumorigenesis of HER2 + BC cells with or without CD36 depletion were examined by mammosphere forming efficiency assay, flow cytometry, cell sorting, ALDH activity assay and xenograft mouse model. FA uptake was examined by flow cytometry with FA BODIPY FL C16. Intratumor expression of CSC subsets was evaluated via multiplex immunostaining and immunolocalization analysis. Results: Molecular data demonstrated that CD36 is significantly upmodulated on treatment in therapy resistant HER2 + BC patients and its expression levels in BC cells is correlated with FA uptake. We provided evidence of a consistent enrichment of CD36 in HER2 + epithelial-mesenchymal transition (EMT)-like CSCs from all tested resistant cell models that mechanistically occurs via Wnt signaling pathway activation. Consistently, both in vitro and in vivo dual blockade of CD36 and HER2 increased the anti-CSC efficacy of anti-HER2 drugs favoring the transition of the therapy resistant mesenchymal CSCs into therapy-sensitive mesenchymal-epithelial transition (MET)-like epithelial state. In addition, expression of CD36 in intratumor HER2 + mesenchymal CSCs is significantly associated with resistance to trastuzumab in HER2 + BC patients. Conclusions: These results support the metabolo-oncogenic nature of CD36-mediated FA uptake in HER2 + therapy-refractory BC. Our study provides evidence that targeting CD36 might be an effective metabolic therapeutic strategy in the treatment of this malignancy. [ABSTRACT FROM AUTHOR]

This study explores the influence of miR-21 and its interaction with the target gene Neurotrophin-3 (NTF3) in cervical cancer (CC). We employed bioinformatics tools, including DIANA, Targetscan, miRDB, and miRDIP, to predict the target genes of miR-21. Immunohistochemistry, RT-qPCR, and Western blotting were performed to quantify the expression levels of miR-21-5p and NTF3 in cervical cancer cells. Additionally, a dual luciferase reporter assay was conducted to examine the specific relationship between miR-21-5P and NTF3. We assessed cell behavior through various tests, including cell viability, scratch wound assays, colony formation, cell invasion experiments, and flow cytometry assays. The dual luciferase reporter assay confirmed that NTF3 is a direct target of miR-21. Overexpression of NTF3 inhibited cell proliferation and migration, while promoting apoptosis, as demonstrated by flow cytometry. Transcriptome sequencing and enrichment analyses (KEGG and GO) revealed NTF3's involvement in key oncogenic pathways, including PI3K-AKT, MAPK, and calcium signaling. This study underscores the critical role of miR-21 in regulating the proliferation, migration, and apoptosis of cervical cancer cells by targeting NTF3. [ABSTRACT FROM AUTHOR]

Glioblastoma is the deadliest primary brain tumor, largely due to inevitable recurrence of the disease after treatment. While most recurrences are local, patients rarely present with a new discontiguous focus of glioblastoma. Little is currently known about the genetic profile of discontiguous recurrences. In our institutional database, we identified 22 patients with targeted exome sequencing of pairs of initial and recurrent IDH-wildtype glioblastoma. Recurrences were classified as contiguous or discontiguous based on the presence or absence of T2 FLAIR signal connection to the initial site of disease on MRI. Exome analysis revealed shared driver and passenger mutations between discontiguous recurrences and initial tumors, supporting a common origin. Discontiguous recurrences were more likely to be hypermutated compared to contiguous recurrences (p = 0.038). Analysis of 2 glioblastoma cases with discontiguous recurrence at a collaborating institution also exhibited hypermutation. In conclusion, discontiguous glioblastoma recurrences share a common origin with the initial tumor and are more likely to be hypermutated than contiguous recurrences. [ABSTRACT FROM AUTHOR]

Objective: Cervical cancer is a common malignancy among women, and radiotherapy remains a primary treatment modality across all disease stages. However, resistance to radiotherapy frequently results in treatment failure, highlighting the need to identify novel therapeutic targets to improve clinical outcomes. Methods: The expression of molecule interacting with CasL-2 (MICAL2) was confirmed in cervical cancer tissues and cell lines through western blotting (WB) and immunohistochemistry (IHC). Siha and Hela cells were used to examine the regulatory and biological functions of MICAL2 via knockdown and overexpression experiments. Assays including MTT, colony formation, wound healing, transwell migration, and sphere formation were employed, along with WB analysis. DNA damage in irradiated cells with MICAL2 knockdown or overexpression was evaluated using the comet assay, while γ-H2AX and Rad51 protein levels were detected by WB. In vivo experiments validated the tumorigenic and radioresistance functions of MICAL2. Additionally, the relationship between MICAL2 expression and radiotherapy response was analyzed in 62 patients with cervical cancer by assessing tumor regression and MICAL2 levels six months post-treatment. Results: MICAL2 expression was significantly elevated in cervical cancer tissues and cells. Functional analyses demonstrated that MICAL2 promotes cell proliferation, migration, and invasion by activating the MAPK and PI3K/AKT pathways, as confirmed through both in vitro and in vivo experiments. Silencing MICAL2 increased DNA damage, impeded DNA repair, and enhanced radiosensitivity. Among the 62 patients with cervical cancer, elevated MICAL2 expression was associated with a lower complete response rate to radiotherapy (25.6% vs. 60.9% in those with low expression), reduced progression-free survival, and advanced cancer stage (*p < 0.05). Conclusion: MICAL2 plays a critical role in tumor progression and radiotherapy resistance in cervical cancer. These findings provide a foundation for developing targeted therapies to improve treatment outcomes in this population. [ABSTRACT FROM AUTHOR]

High-grade soft tissue sarcomas (STS) are a heterogeneous and aggressive set of cancers. Failure to respond anthracycline chemotherapy, standard first-line treatment, is associated with poor outcomes. We investigated the contribution of STS cancer stem cells (STS-CSCs) to doxorubicin resistance. We identified a positive correlation between CSC abundance and doxorubicin IC50. Utilizing patient-derived samples from five sarcoma subtypes we investigated if a common genetic signature across STS-CSCs could be targeted. We identified Enhancer of Zeste homolog 2 (EZH2), a member of the polycomb repressive complex 2 (PRC2) responsible for H3K27 methylation as being enriched in CSCs. EZH2 activity and a shared epigenetic profile was observed across subtypes and targeting of EZH2 ablated the STS-CSC population. Treatment of doxorubicin-resistant cell lines with tazemetostat resulted in a decrease in the STS-CSC population. These data confirm the presence of shared genetic programs across distinct subtypes of CSC-STS that can be therapeutically targeted. [ABSTRACT FROM AUTHOR]

INhibitor of Growth (ING1-5) proteins are epigenetic readers that target histone acetyltransferase (HAT) or histone deacetylase (HDAC) complexes to the H3K4Me3 mark of active transcription. ING5 targets Moz/Morf and HBO1 HAT complexes that alter acetylation of H3 and H4 core histones, affecting gene expression. Previous experiments in vitro indicated that ING5 functions to maintain stem cell character in normal and in cancer stem cells. Here we find that CRISPR/Cas9 ING5 knockout (KO) mice are sub-fertile but show no decrease in lifespan or ability to heal wounds despite indications of depleted stem cell pools in several tissues. ING5 KO mouse embryo fibroblasts accumulate in G2 of the cell cycle, have high levels of abnormal nuclei and show high basal levels of the γH2AX indicator of DNA damage. KO animals also develop severe dermatitis at a 5-fold higher rate that wild-type littermates. Consistent with ING5 serving a tumor suppressive role, ING5 KO mice developed germinal centre diffuse large B-cell lymphomas at a rate 6-fold higher than control mice at 18 months of age. These data suggest that ING5 functions in vivo to maintain stem cell character in multiple organs, that reduction of stem cell populations is not limiting for murine lifespan and that like a subset of other ING family members, ING5 functions as a tumor suppressor in hematopoietic cells in vivo. [ABSTRACT FROM AUTHOR]

Purpose: Tumor-associated macrophages (TAMs) are pivotal immune cells within the tumor microenvironment (TME), exhibiting dual roles across various cancer types. Depending on the context, TAMs can either suppress tumor progression and weaken drug sensitivity or facilitate tumor growth and drive therapeutic resistance. This study explores whether targeting TAMs can suppress the progression of head and neck squamous cell carcinoma (HNSCC) and improve the efficacy of chemotherapy. Methods: Bioinformatics analyses were performed to evaluate TAMs infiltration levels in HNSCC tumor tissues and examine their associations with patients' clinicopathological characteristics and prognosis. Flow cytometry was utilized to measure the expression of key macrophage markers and assess apoptosis following treatment with colony stimulating factor 1 receptor (CSF1R) inhibitors (BLZ945, PLX3397). Additionally, immunohistochemistry was employed to detect CD68 and CD8 expression. In vivo, the antitumor efficacy of CSF1R inhibitors was tested in mouse HNSCC tumor model, both as monotherapy and in combination with cisplatin, to evaluate potential synergistic effects. Results: Bioinformatic analysis identified TAMs as the predominant infiltrating immune cells in the TME of HNSCC, with significantly higher infiltration levels in tumor tissues compared to adjacent non-tumor tissues. High TAMs infiltration was associated with poorer overall survival (OS), disease-free survival (DFS), human papillomavirus (HPV) infection status, and advanced disease stages. The TAMs-related genes prediction model demonstrated high prognostic accuracy. CSF1R is primarily expressed in TAMs, where high CSF1R expression may suppress antigen binding and activation. In vitro experiments showed that CSF1R inhibitors induce TAMs apoptosis, enhance their phagocytic activity, and reduce CD206 expression and IL-10 secretion, thereby diminishing their immunosuppressive function. In vivo experiments revealed that while CSF1R inhibitors alone had limited efficacy in suppressing tumor growth, their combination with cisplatin significantly enhanced therapeutic efficacy, as evidenced by increased CD8+ T cells infiltration within the TME. Conclusion: Targeting TAMs via CSF1R inhibition enhances the therapeutic efficacy of cisplatin in HNSCC. These findings suggest that CSF1R inhibitors hold promise as a component of combination therapy for HNSCC. [ABSTRACT FROM AUTHOR]

Background: The refinement of risk stratification in lung adenocarcinoma (LUAD) plays a pivotal role in advancing precision medicine; however, the current staging classification falls short of comprehensiveness, particularly in the case of stage IA patients. We aimed to molecularly stratify LUAD patients especially for stage IA. Methods: We analysed tumour heterogeneity and identified highly proliferating cancer cells (HPCs) in LUAD by performing single-cell RNA sequencing (scRNA-seq) analysis, immunohistochemical (IHC) staining using a tissue microarray, flow cytometry and biological experiments. Then, we quantified the content of HPCs in nine LUAD datasets by single-sample gene set enrichment analysis and evaluated the relationship between the percentage of HPCs and overall survival (OS). Next, we analysed the OS predictive effect of HPCs at different LUAD stages, especially for stage I risk stratification. Furthermore, we established a prognostic prediction model based on HPC-associated genes for clinical application. The above findings were validated in another five LUAD datasets. Finally, we explored the relationship between HPCs and the progressive pathological evolution of early-stage LUAD and the driving mutations by scRNA-seq, bulk RNA-seq and IHC staining. Results: LUAD tissues carry a small proportion of HPCs, which show potential for malignant proliferation and intense interactions with the microenvironment. A high HPC content is an independent risk factor for OS in LUAD patients, even in stage IA patients. HPCs can be used to establish a cut-off point for the prognosis of stage IA disease, with patients with a higher risk showing a prognosis similar to that of patients with stage IB disease. We built an R package (HSurADs) based on HPC-associated genes, which exhibited good efficacy for the prognostic prediction of LUAD. HPCs gradually increase with the pathological evolution of early-stage LUAD, which may be affected by TP53 mutations. Conclusion: The HPC content can be used as a novel prognostic factor for LUAD, especially for stage IA risk stratification. [ABSTRACT FROM AUTHOR]

Background: Prostaglandin D2 (PGD2) inhibits the development of different malignant tumors; however, the underlying mechanism of inhibiting tumor development is not yet clear. This study aimed to elucidate how PGD2 inhibits the stemness of gastric cancer stem cells (GCSCs) via autophagy and its underlying molecular mechanism to provide a theoretical basis for the treatment of gastric cancer. Methods: In this study, GCSCs were enriched in vitro by serum-free incubation. Furthermore, the effects of PGD2 and PGD2 receptor (PTGDR2) on autophagy were detected by Western blotting, immunofluorescence analysis, and transmission electron microscopy. Moreover, the ATG4B ubiquitination levels were assessed via immunoprecipitation and other methods. Results: The results indicated that PGD2 induced LC3I/LC3II conversion in GCSCs to activate autophagy, while PGD2 promoted the expression of PTGDR2, thereby further activating autophagy. Furthermore, PTGDR2 competes with ATG4B for binding with E3 ligase RNF5 (also known as RMA1) to promote autophagy protein ATG4B expression. Moreover, PTGDR2 knockdown blocked the activation of autophagy by PGD2 and the level of ATG4B ubiquitination in GCSCs. Conclusions: In summary, it was elucidated that the PGD2/PTGDR2 signaling cascade affects GCSCs stemness by regulating autophagy, suggesting that the PGD2/PTGDR2 signaling pathway could serve as a novel target for cancer therapy. Schematic diagram of the study. [ABSTRACT FROM AUTHOR]

Thyroid cancer has become the most common endocrine malignancy. Although the majority of differentiated thyroid cancers have a favorable prognosis, advanced thyroid cancers, iodine-refractory thyroid cancers, and highly malignant undifferentiated carcinomas still face a serious challenge of poor prognosis and even death. Cancer stem cells are recognized as one of the central drivers of tumor evolution, recurrence and treatment resistance. A fresh viewpoint on the oncological aspects of thyroid cancer, including proliferation, invasion, recurrence, metastasis, and therapeutic resistance, has been made possible by the recent thorough understanding of the defining and developing features as well as the plasticity of thyroid cancer stem cells (TCSCs). The above characteristics of TCSCs are complicated and regulated by cell-intrinsic mechanisms (including activation of key stem signaling pathways, somatic cell dedifferentiation, etc.) and cell-extrinsic mechanisms. The complex communication between TCSCs and the infiltrating immune cell populations in the tumor microenvironment (TME) is a paradigm for cell-extrinsic regulators. This review introduces the current advances in the studies of TCSCs, including the origin of TCSCs, the intrinsic signaling pathways regulating the stemness of TCSCs, and emerging biomarkers; We further highlight the underlying principles of bidirectional crosstalk between TCSCs and immune cell populations driving thyroid cancer progression, recurrence, or metastasis, including the specific mechanisms by which immune cells maintain the stemness and other properties of TCSCs and how TCSCs reshape the immune microenvironmental landscape to create an immune evasive and pro-tumorigenic ecological niche. Finally, we outline promising strategies and challenges for targeting key programs in the TCSCs-immune cell crosstalk process to treat thyroid cancer. [ABSTRACT FROM AUTHOR]

Cancer remains a leading cause of mortality globally and a major health burden, with chemotherapy often serving as the primary therapeutic option for patients with advanced-stage disease, partially compensating for the limitations of non-curative treatments. However, the emergence of chemotherapy resistance significantly limits its efficacy, posing a major clinical challenge. Moreover, heterogeneity of resistance mechanisms across cancer types complicates the development of universally effective diagnostic and therapeutic approaches. Understanding the molecular mechanisms of chemoresistance and identifying strategies to overcome it are current research focal points. This review provides a comprehensive analysis of the key molecular mechanisms underlying chemotherapy resistance, including drug efflux, enhanced DNA damage repair (DDR), apoptosis evasion, epigenetic modifications, altered intracellular drug metabolism, and the role of cancer stem cells (CSCs). We also examine specific causes of resistance in major cancer types and highlight various molecular targets involved in resistance. Finally, we discuss current strategies aiming at overcoming chemotherapy resistance, such as combination therapies, targeted treatments, and novel drug delivery systems, while proposing future directions for research in this evolving field. By addressing these molecular barriers, this review lays a foundation for the development of more effective cancer therapies aimed at mitigating chemotherapy resistance. [ABSTRACT FROM AUTHOR]

Background: Glioblastoma multiforme (GBM) exhibits a cellular hierarchy with a subpopulation of stem-like cells known as glioblastoma stem cells (GSCs) that drive tumor growth and contribute to treatment resistance. NAD(H) emerges as a crucial factor influencing GSC maintenance through its involvement in diverse biological processes, including mitochondrial fitness and DNA damage repair. However, how GSCs leverage metabolic adaptation to obtain survival advantage remains elusive. Methods: A multi-step process of machine learning algorithms was implemented to construct the glioma stemness-related score (GScore). Further in silico and patient tissue analyses validated the predictive ability of the GScore and identified a potential target, CYP3A5. Loss-of-function or gain-of-function genetic experiments were performed to assess the impact of CYP3A5 on the self-renewal and chemoresistance of GSCs both in vitro and in vivo. Mechanistic studies were conducted using nontargeted metabolomics, RNA-seq, seahorse, transmission electron microscopy, immunofluorescence, flow cytometry, ChIP‒qPCR, RT‒qPCR, western blotting, etc. The efficacy of pharmacological inhibitors of CYP3A5 was assessed in vivo. Results: Based on the proposed GScore, we identify a GSC target CYP3A5, which is highly expressed in GSCs and temozolomide (TMZ)-resistant GBM patients. This elevated expression of CYP3A5 is attributed to transcription factor STAT3 activated by EGFR signaling or TMZ treatment. Depletion of CYP3A5 impairs self-renewal and TMZ resistance of GSCs. Mechanistically, CYP3A5 maintains mitochondrial fitness to promote GSC metabolic adaption through the NAD⁺/NADH-SIRT1-PGC1α axis. Additionally, CYP3A5 enhances the activity of NAD-dependent enzyme PARP to augment DNA damage repair. Treatment with CYP3A5 inhibitor alone or together with TMZ effectively suppresses tumor growth in vivo. Conclusion: Together, this study suggests that GSCs activate STAT3 to upregulate CYP3A5 to fine-tune NAD⁺/NADH for the enhancement of mitochondrial functions and DNA damage repair, thereby fueling tumor stemness and conferring TMZ resistance, respectively. Thus, CYP3A5 represents a promising target for GBM treatment. [ABSTRACT FROM AUTHOR]

Background/Aim: Ifosfamide is used clinically with doxorubicin as first-line chemotherapy for soft-tissue sarcoma. However, ifosfamide efficacy for soft-tissue sarcoma is limited due to frequent occurence of ifosfamide resistance and thus more effective therapy is needed. The present study aimed to determine the synergy of recombinant methioninase (rMETase) plus ifosfamide against HT1080 human fibrosarcoma cells in vitro. Additionally, the present study also investigated the efficacy of a methionine-restricted diet combined with ifosfamide in nudemouse models of ifosfamide-resistant HT1080 (IR-HT1080). Materials and Methods: Cell viability for HT1080 human fibrosarcoma cells was determined in four groups in vitro: No treatment control; ifosfamide alone; rMETase alone; and a combination of ifosfamide plus rMETase. HT1080 tumors were established in nude mice subcutaneously. The HT1080 tumor models were treated by administering ifosfamide by intraperitoneal injection twice a week, for a total of 11 doses. Surviving tumors were considered ifosfamide resistant (IRHT1080). Four groups of IR-HT1080 nude-mouse models were subsequently established: Group 1 was a no-treatment control, Group 2 received ifosfamide, Group 3 was given a methioninerestricted diet (MR), and Group 4 received ifosfamide plus MR. Additionally, two groups of nude mice with parental HT1080 subcutaneous tumors were included: Group 5 was a notreatment control, and Group 6 received ifosfamide for comparison. Results: The 50% inhibitory concentration (IC50) for ifosfamide against HT1080 cells was 0.38 mM. The IC50 for rMETase was 0.75 U/ml for HT1080 cells (data from [4]). The combination of rMETase (0.75 U/ml) plus ifosfamide (0.38 mM) was synergistic against HT1080 fibrosarcoma cells in vitro. The combination of ifosfamide plus MR eradicated the IR-HT1080 tumors in nude-mouse models, while each treatment alone achieved limited tumor inhibition. Conclusion: The present results suggest the combination of MR and ifosfamide has promising potential for overcoming ifosfamide resistance in future clinical applications. [ABSTRACT FROM AUTHOR]

Objective: This study aims to explore the potential role of mesenchymal stem cells (MSCs) in the treatment of osteoarthritis (OA), particularly the function of the NOTCH1 signaling pathway in maintaining the stemness of MSCs and in chondrocyte differentiation. Methods: Utilizing diverse analytical techniques on an osteoarthritis dataset, we unveil distinct gene expression patterns and regulatory relationships, shedding light on potential mechanisms underlying the disease. Techniques used include the culture of MSCs, induction of differentiation into chondrocytes, establishment of stable cell lines, Western Blot, and immunofluorescence. Through the construction of lentiviruses overexpressing and knocking out NOTCH1, the effects of NOTCH1 on the stemness of MSCs and chondrocyte differentiation were investigated. Additionally, the effects of NOTCH1 on chondrocyte homeostasis and apoptosis were evaluated by adding the EZH2 inhibitor GSK126 and the endoplasmic reticulum stress inducer tunicamycin. Results: Experimental results demonstrated that NOTCH1 expression can influence the maintenance of MSC stemness and chondrocyte differentiation by regulating EZH2. Knockout of NOTCH1 decreased the expression of chondrocyte markers, while overexpression increased their expression. Under conditions of endoplasmic reticulum stress, NOTCH1 expression helped reduce the expression of stress-related proteins, maintain chondrocyte homeostasis, and inhibit apoptosis. Conclusion: The NOTCH1 signaling pathway plays a crucial role in maintaining the stemness of MSCs, differentiating into chondrocytes, and in the treatment of osteoarthritis. NOTCH1 influences the differentiation fate of MSCs and the homeostasis of chondrocytes by regulating EZH2 and other related genes, offering new targets and strategies for the treatment of diseases like osteoarthritis. [ABSTRACT FROM AUTHOR]

Antibody-drug conjugate (ADC) therapy has transformed treatment for several solid tumors, including small cell lung cancer (SCLC). However, significant challenges remain, including systemic toxicity, acquired resistance, and the lack of reliable biomarkers for patient selection. To enhance the effectiveness of ADC therapies in SCLC, we focused on target selection in this study by investigating the expression of ADC targets - SEZ6, DLL3, CD276, and TACSTD2 - in cell lines and patient samples. SEZ6 expression was significantly elevated in various SCLC transcriptional subtypes, particularly ASCL1, and exhibited gender-specific differences, being lower in women. DLL3 was primarily observed in the ASCL1 subtype, while CD276 showed high expression in non-neuroendocrine subtypes. TACSTD2 levels were generally low and attenuated in lymph nodes and brain metastases compared to primary tumors. Our findings underscore the importance of understanding target expression patterns to optimize ADC therapy and advance precision medicine in SCLC treatment. [ABSTRACT FROM AUTHOR]

Background: Pancreatic cancer is a highly aggressive neoplasm characterized by poor diagnosis. Amino acids play a prominent role in the occurrence and progression of pancreatic cancer as essential building blocks for protein synthesis and key regulators of cellular metabolism. Understanding the interplay between pancreatic cancer and amino acid metabolism offers potential avenues for improving patient clinical outcomes. Methods: A comprehensive analysis integrating 10 machine learning algorithms was executed to pinpoint amino acid metabolic signature. The signature was validated across both internal and external cohorts. Subsequent GSEA was employed to unveil the enriched gene sets and signaling pathways within high- and low-risk subgroups. TMB and drug sensitivity analyses were carried out via Maftools and oncoPredict R packages. CIBERSORT and ssGSEA were harnessed to delve into the immune landscape disparities. Single-cell transcriptomics, qPCR, and Immunohistochemistry were performed to corroborate the expression levels and prognostic significance of this signature. Results: A four gene based amino acid metabolic signature with superior prognostic capabilities was identified by the combination of 10 machine learning methods. It showed that the novel prognostic model could effectively distinguish patients into high- and low-risk groups in both internal and external cohorts. Notably, the risk score from this novel signature showed significant correlations with TMB, drug resistance, as well as a heightened likelihood of immune evasion and suboptimal responses to immunotherapeutic interventions. Conclusion: Our findings suggested that amino acid metabolism-related signature was closely related to the development, prognosis and immune microenvironment of pancreatic cancer. [ABSTRACT FROM AUTHOR]

Background: Epidemiological studies associate an increase in breast cancer risk, particularly triple-negative breast cancer (TNBC), with lack of breastfeeding. This is more prevalent in African American women, with significantly lower rate of breastfeeding compared to Caucasian women. Prolonged breastfeeding leads to gradual involution (GI), whereas short-term or lack of breastfeeding leads to abrupt involution (AI) of the breast. Our previous study utilizing a murine model demonstrated precancerous changes, specifically hyperplasia, a non-obligate precursor of breast cancer in the mammary glands of AI mice. Here we investigated mechanisms during early events of AI that prompts precancerous changes in mouse mammary glands. Methods: Uniparous FVB/N mice were randomized to AI and GI on postpartum day 7 when all pups were removed from AI dams. GI dams were allowed to nurse the pups till day 31. Cell death kinetics and gene expression were assessed by TUNEL assay and qPCR respectively. Immune cell changes were investigated by flow cytometry, cytokine array and multiplex immunofluorescence. 3D-organoid cultures were used for in vitro assay of luminal progenitor cells. Results: AI results in rapid cell death, DNA repair response, and immunosuppressive myeloid cells infiltration, leading to a chronically inflamed microenvironment. GI elicits a more controlled immune response and extended cell death. At the peak of cell death, AI glands harbored more immunosuppressive myeloid-derived suppressor cells (MDSCs) and CD206 + M2-like macrophages, known to promote oncogenic events, compared to GI glands. AI glands exhibit an enrichment of CCL9-producing MDSCs and CD206 + M2-like macrophages that promote expansion of ELF5 + /ERα- luminal cells, both in vitro and in vivo. Multiplex imaging of AI glands demonstrated an increase in ELF5 + /WNT5a + luminal cells alongside a reduction in the ELF5 + /ERα + population when involution appeared histologically complete. A significantly higher number of CD206 + cells in post involution AI gland attests to a chronically inflamed state induced by AI. Conclusions: Our findings reveal significant disparities between AI and GI gland dynamics at the early phase of involution. CCL9, secreted by immune cells at the peak of cell death promotes expansion of Elf5 + /ERα- luminal progenitor cells, the putative precursors of TNBC connecting early events of AI with increased breast cancer risk. [ABSTRACT FROM AUTHOR]

Head and neck squamous cell carcinoma (HNSCC) originates from the mucosal epithelium of the oral cavity, pharynx, and larynx, and is marked by high rates of recurrence and metastasis. Calcium signaling is associated with the progression of HNSCC and the development of drug resistance. Changes in calcium ion flow can trigger severe pathophysiological processes, including malignant transformation, tumor proliferation, epithelial-mesenchymal transition, and apoptosis evasion. Calcium channels regulate and facilitate these processes. Remodeling of calcium signaling has become one of the most prevalent adaptive mechanisms in cancer cells. Preclinical and clinical evidence indicates that alterations in calcium signaling are crucial for the progression of HNSCC. This review examines the role of calcium channels in HNSCC development and evaluates current clinical trials targeting these channels to assess the feasibility of calcium signaling-based therapies for HNSCC. [ABSTRACT FROM AUTHOR]

Yes-associated protein (YAP) activation confers resistance to chemotherapy and targeted therapy. Methionine participates in cellular processes by converting to methyl donor for the methylation of DNA, RNA and protein. However, it remains unclear whether methionine affects drug resistance by influencing YAP activity. In this study, we report that methionine deprivation remarkably suppresses the transcriptional activity of YAP–TEAD in cancer cells. Methionine promotes PRMT1-catalyzed asymmetric dimethylation at R124 of YAP (YAP R124me2a). Mimicking of YAP methylation abolishes the reduction effect of methionine-restricted diet on YAP-induced drug resistance. YAP activates the transcription of SLC43A2, the methionine transporter, to increase methionine uptake in cancer cells. Knockdown of SLC43A2 decreases the level of YAP R124me2a. BCH, the inhibitor of SLC43A2, sensitizes tumors to anticancer drugs. Thus, our results unravel the positive feedback between YAP R124 methylation and SLC43A2 that contributes to anticancer drug resistance. Disrupting this positive feedback could be a potential strategy for cancer therapy. While methionine deficiency can sensitize cancer cells to therapy, YAP activation contributes to drug resistance. Here, authors discover that the positive feedback loop present between YAP R124 methylation and the methionine transporter SLC43A2 is involved in drug resistance to multiple therapies. [ABSTRACT FROM AUTHOR]

Emerging evidence suggests that fusion of cancer cells with leucocytes, such as macrophages, plays a significant role in cancer metastasis and results in tumor hybrid cells that acquire resistance to chemo- and radiation therapy. However, the precise mechanisms behind the leukocyte-cancer cell fusion remain unclear. The present in vitro study explores the presence of fusion between the monocyte cell line (THP-1) and the breast cancer cell line (MCF-7) in relation to the expression of CD36 and phosphatidylserine with and without treatment of these cells with ionizing radiation. The study reveals that spontaneous THP-1/MCF-7 cell fusion increases significantly from 2.8% to 6% after irradiation. The interaction between CD36 and phosphatidylserine plays a pivotal role in THP-1/MCF-7 cell fusion, as inhibiting this interaction using anti-CD36 antibodies significantly reduces cell fusion. While irradiation leads to a dose-dependent escalation in phosphatidylserine expression in MCF-7 cells, it does not impact the expression of CD36 in either THP-1 or MCF-7 cells. To the best of our knowledge, this is the first study to demonstrate the involvement of the CD36-phosphatidylserine interaction in the fusion between monocytes and cancer cells, shedding light on a novel explanatory mechanism for the roles of CD36 and phosphatidylserine in tumor progression. [ABSTRACT FROM AUTHOR]

Background: Cluster of differentiation 44 (CD44) is the most commonly reported immune marker of the cancer stem cells in renal cell carcinoma. Objectives: To assess the frequency of CD44 expression in renal tumors, to assess the expression of CD44 in different histological types, and to detect its association with variable clinicopathological parameters. Materials and methods: In this retrospective case series study, 76 cases of primary renal tumors were obtained by nephrectomy, and a study of CD44 was done by using the immunohistochemical technique. Results: The age of the patients ranged from 29–82 years with a mean of 54.8 ± 11.96 with a male to female ratio of 1.05:1. Tumor size ranged from 2–17 cm with a mean of 6.2 ± 3.23. Clear cell renal cell carcinoma was forming 61.83% of the cases. Among all cases, a positive CD44 expression was observed in 35 (46.05%) cases; among the malignant cases, CD44 was positive in 32 (45.71%). CD44 immunohistochemical stain showed significantly higher expression in cases with higher nuclear grade and tumor stage (P-value of 0.0376 and 0.0075, respectively). CD44 cannot differentiate between benign and malignant renal tumors. Conclusion: CD44 immune marker can be used as a prognostic factor to predict the aggressive behavior of renal tumors. It can also be used with standard prognostic markers to add prognostic information for subgrouping cases within the same grade and stage. [ABSTRACT FROM AUTHOR]

Colon cancer is one of the most common cancer-related deaths. Drug resistance is one of the biggest challenges in cancer treatment. Numerous pharmacological and biochemical investigations have documented the benzimidazole ring's anticancer, anti-inflammatory, and antioxidant properties. Within the scope of our project, the effect of newly synthesized benzimidazolium salt (BS) on cell proliferation was tested with MTT assay, and its effect on apoptosis and cell cycle was tested with annexin V and PI in the two different colon cancer cell lines (HT-29 and DLD-1). Our study examined the expressions of some genes related to apoptosis and, additionally, caspase activities with the multicaspase kit. BS showed an antiproliferative effect at lower doses in HT-29 colon cancer cells. When HT-29 cells were exposed to a 20 µM dosage, they showed increased caspase activity and apoptosis compared to DLD-1 cells. HT-29 accumulated in the G2/M phase of the cell cycle, whereas DLD-1 cells accumulated more in the S phase. In HT-29 cells, colony formation was inhibited; however, in DLD-1 cells, this effect was insufficient. Based on the apoptosis-death pathway, BS is expected to have anti-cancer effects. As a result of this work, this chemical was thoroughly examined in two different colon cancer cell lines, and additional, more comprehensive initiatives are being planned in light of the information obtained from this study., (© 2024 Wiley Periodicals LLC.)

Evidence for folate's protective effects on neural tube defects led the USA and Chile to start mandatory folic acid (FA) fortification programs, decreasing up to 50%. However, ∼30% of the population consuming fortified foods reach supraphysiologic serum levels. Although controversial, several epidemiological and clinical observations suggest that folate increases cancer risk, giving concern about the risks of FA supplementation. The Cancer stem cells (CSCs) model has been used to explain survival to anticancer therapies. The Notch-1 pathway plays a role in several cancers and is associated with the stemness process. Different studies show that modulation of metabolic pathways regulates stemness capacity in cancer. Supraphysiologic concentrations of FA increase the proliferation of HT-29 cells by Notch-1 activation. However, whether folate can induce a stemness-like phenotype in cancer is not known. We hypothesized that FA protects from glucose deprivation-induced cell death through Notch-1 activation. HT-29 cells were challenged with glucose deprivation at basal (20 nM) and supraphysiological (400 nM) FA and 5-MTHF concentrations. We analyzed changes in stemness-like gene expression, cell death and different energetic metabolic functions. Supraphysiological concentrations of FA increased stemness-like genes, and improved survival and oxygen consumption, inducing AMPK phosphorylation and HSP-70 protein expression. We evaluated the Notch-1 pathway using the DAPT and siRNA as inhibitors, decreasing the stemness-like gene expression and preventing the FA protection against glucose deprivation-induced cell death. Moreover, they decreased oxygen consumption and AMPK phosphorylation. These results suggest that FA protects against glucose deprivation. These effects were associated with AMPK activation, a critical metabolic mediator in nutrient consumption and availability that activates the Notch-1 pathway., Competing Interests: Declarations. Conflict of interest: The authors declare no conflicts of interest., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Sine oculis homeobox homolog 1 (SIX1) is a new identified cancer driver in the development of prostate cancer (PC). However, the upstream regulatory mechanisms for SIX1 reactivation in cancer remains elusive. Here, we found that Ku70 robustly interacts with SIX1 in the nucleus of PC cells. The HD domain of SIX1 and the DBD domain of Ku70 are required for formation of Ku70-SIX1 complex. 20 groups of hydrogen bonds were identified in this complex by molecular dynamics simulation. Depletion of Ku70/SIX1 notably abrogates the proliferation and migration of PC. Further studies revealed that SIX1 is recruited to the promoter region on glutamate-pyruvate transaminase 2 (GPT2). Ku70 enhances the SIX1-mediated transcriptional activation on GPT2, thereby facilitating the generation of alpha-ketoglutarate (α-KG). In addition, formation of the Ku70-SIX1 complex promotes GPT2-dependent cell proliferation and migration in PC. Moreover, the expression of GPT2 is upregulated and strongly correlated with the expression of Ku70/SIX1 in PC tissues. In summary, our findings not only provide insight into the mechanistic interactions between Ku70 and SIX1, but also highlight the significance of the Ku70-SIX1-GPT2 axis for α-KG metabolism and PC carcinogenesis., Competing Interests: Competing interests: The authors declare no competing interests. Ethics approval and consent to participate: The authors confirm that all methods were performed in accordance with the relevant guidelines and regulations. Informed consent was obtained from all the subjects. All animal experiments were approved by the institutional animal care and use committees of Guangzhou Medical University(GY2022-137). All human subjects were performed with the approval of the Medical Ethics Committee of the First People’s Hospital of Foshan (ethics approval number: L [2024] No. 3)., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Glycine decarboxylase (GLDC) has been identified to be dysregulated and plays pivotal roles in various cancers. Besides, studies have suggested that GLDC expression is elevated in oral squamous cell carcinoma (OSCC) and associated with a worse prognosis, but the precise role and molecular mechanism of GLDC in OSCC remain unexplored. The current study first confirmed the high expression of GLDC in OSCC and its correlation with worse survival in patients with OSCC. By knocking down GLDC, it was discovered that the growth and colony formation of OSCC cells, as well as the development of xenograft tumors, were effectively suppressed. In addition, GLDC deficiency inhibited the migration and invasion of OSCC cells in vitro through regulating EMT markers and attenuated lung metastasis in vivo. Mechanistically, GLDC was found to affect the activity of the p53 signaling pathway. GLDC depletion retarded the progression of OSCC by activating the p53 signaling pathway. Moreover, p300 co-functioned with TFAP2A to induce acetylation of GLDC, which resulted in the upregulation of GLDC in OSCC. To conclude, acetylation-induced GLDC upregulation facilitated the tumorigenesis and metastasis of OSCC by its inhibition of the activity of the p53 signaling pathway., (© 2024 Wiley Periodicals LLC.)

As an essential branched amino acid, valine is pivotal for protein synthesis, neurological behaviour, haematopoiesis and leukaemia progression 1-3 . However, the mechanism by which cellular valine abundancy is sensed for subsequent cellular functions remains undefined. Here we identify that human histone deacetylase 6 (HDAC6) serves as a valine sensor by directly binding valine through a primate-specific SE14 repeat domain. The nucleus and cytoplasm shuttling of human, but not mouse, HDAC6 is tightly controlled by the intracellular levels of valine. Valine deprivation leads to HDAC6 retention in the nucleus and induces DNA damage. Mechanistically, nuclear-localized HDAC6 binds and deacetylates ten-eleven translocation 2 (TET2) to initiate active DNA demethylation, which promotes DNA damage through thymine DNA glycosylase-driven excision. Dietary valine restriction inhibits tumour growth in xenograft and patient-derived xenograft models, and enhances the therapeutic efficacy of PARP inhibitors. Collectively, our study identifies human HDAC6 as a valine sensor that mediates active DNA demethylation and DNA damage in response to valine deprivation, and highlights the potential of dietary valine restriction for cancer treatment., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Luciferase-based reporter assay is an important tool that employs bioluminescence to quickly and precisely investigate the gene of interest's promoter activity by reporter gene expression at the transcriptional level. The promoter of the gene of interest is fused with the reporter gene (a gene that produces luciferase enzymes) and then transfected into the cells. Luciferase is an enzyme that catalyzes a chemical reaction to produce light. The bioluminescence activity of the luciferase gene in the transfected cells is directly proportional to the expression of the gene of interest, which is measured by using a luminometer. In this chapter, we outline the use of a dual-reporter luciferase assay to measure Aurora A kinase activity during the mitotic cycle., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Tumorigenesis embodies the formation of a heterotypic tumour microenvironment (TME) that, among its many functions, enables the evasion of T cell-mediated immune responses. Remarkably, most TME cell types, including cancer cells, fibroblasts, myeloid cells, vascular endothelial cells and pericytes, can be stimulated to deploy immunoregulatory programmes. These programmes involve regulatory inducers (signals-in) and functional effectors (signals-out) that impair CD8 + and CD4 + T cell activity through cytokines, growth factors, immune checkpoints and metabolites. Some signals target specific cell types, whereas others, such as transforming growth factor-β (TGFβ) and prostaglandin E 2 (PGE 2 ), exert broad, pleiotropic effects; as signals-in, they trigger immunosuppressive programmes in most TME cell types, and as signals-out, they directly inhibit T cells and also modulate other cells to reinforce immunosuppression. This functional diversity and redundancy pose a challenge for therapeutic targeting of the immune-evasive TME. Fundamentally, the commonality of regulatory programmes aimed at abrogating T cell activity, along with paracrine signalling between cells of the TME, suggests that many normal cell types are hard-wired with latent functions that can be triggered to prevent inappropriate immune attack. This intrinsic capability is evidently co-opted throughout the TME, enabling tumours to evade immune destruction., Competing Interests: Competing interests: None of the research results reviewed and discussed here reflect bona fide competing interests. In the interest of full disclosure, D.H. currently serves on the scientific advisory boards of Pfizer Oncology, Opna Bio, 4D Molecular Therapeutics and Cellestia, and is a founder of Opna Bio, which has licensed an EPFL patent describing the RNA-binding protein FMRP as a new cancer target. O.M. has served as an adviser for Roche, Novartis, GSK, BMS, MSD, Regeneron, Pierre-Fabre, Merck and Amgen, and is a scientific board member of Cellula Therapeutics. M.J.P. has served as an adviser for AstraZeneca, ImmuneOncia, LegoChem Bio, MaxiVax, Merck, Molecular Partners, Third Rock Ventures, Tidal and Unikum., (© 2024. Springer Nature Limited.)

Objective: To investigate the expression features of common anti-glioma CAR-T targets (B7H3, CSPG4, EGFRv III, HER2 and IL-13Ra2) in gliomas with different grades and molecular subtypes, and explore the association of target expression with glioma malignant or immune phenotypes including immune evasion, stemness, antigen presentation, and tumor angiogenesis., Methods: Opal™ Multiplex immunofluorescence staining was performed on glioma tissues to detect the expression of targets, and biomarkers related to the phenotypes., Results: High variety of CAR-T target expression among glioma subtypes was observed. GBMs exhibited the highest expression level of all the examined targets among glioma subtypes. In all glioma cases, CSPG4 was the most prevalent target covering over 84% glioma cases, followed by B7H3 at over 64%. B7H3 exhibited the highest coverage (94%) in GBMs while CSPG4 was the most popular target in both oligodendrogliomas and astrocytomas, covering 94% and 80% cases, respectively. Bi or tri-target combination strategies markedly expanded the tumor coverage across glioma cases while increased tumor-cell coverage within tumor. PD-L1 expression was significantly enriched in all the target-positive cells (except the EGFRvIII + cells); CD133 expression was higher in the CSPG4 + or IL-13Ra2 + cells, and CD31 elevated in the B7H3 + cells, as compared with their negative cell populations., Conclusion: Anti-glioma CAR-T targets have heterogenous expression and distinct tumor coverage among glioma subtypes, and closely correlate with glioma malignant or immune phenotypes., Competing Interests: Declarations. Competing interests: No potential conflict of interest was disclosed., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Extensive research into gene fusions in cancer and other diseases has led to the discovery of novel biomarkers and therapeutic targets. Concurrently, various bioinformatics tools have been developed for fusion detection in RNA sequencing data, which, in the age of increasing affordability of sequencing, have delivered a large-scale identification of transcriptomic abnormalities. Historically, the focus of fusion transcript research was predominantly on coding RNAs and their resultant proteins, often overlooking non-coding RNAs (ncRNAs). This chapter discusses how ncRNAs are integral players in the landscape of gene fusions, detailing their contributions to the formation of gene fusions and their presence in chimeric transcripts. We delve into both linear and the more recently identified circular fusion RNAs, providing a comprehensive overview of the computational methodologies used to detect ncRNA-involved gene fusions. Additionally, we examine the inherent biases and limitations of these bioinformatics approaches, offering insights into the challenges and future directions in this dynamic field., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)