Gliomas are the most common primary brain tumors, yet the molecular circuits that drive their malignancy remain incompletely defined. Here, using an integrative, multi-dimensional approach, we aimed to pinpoint key molecular drivers having both functional and clinical relevance to disease progression and tumor aggressiveness in gliomas. Genome-wide CRISPR-Cas9 dependency screen across 70 glioma cell lines was paired with tumor aggressiveness-targeted transcriptomic differential expression and survival analyses to pinpoint critical drivers of disease progression in gliomas. Functional and gene set enrichments as well as protein-protein interaction network analyses were used to identify dominant pathways and key hub genes, followed by independent validation across external transcriptomic and proteomic datasets. Upstream regulator analyses and alternative splicing profiling were performed to nominate regulatory drivers and derive a small nuclear ribonucleoprotein D2 polypeptide (SNRPD2)-associated splicing signature. Initial screening uncovered 222 essential genes (Chronos<-1) in gliomas, 87 of which were overexpressed in tumors displaying proliferative, epithelial-mesenchymal transition, glycolytic, hypoxic, and inflammatory signatures, and were associated with poor overall survival, consistent with aggressive disease biology. These genes converged on alternative splicing regulation, proteasome function, and cell cycle, with spliceosome core component, SNRPD2 emerging as the top hub gene. High SNRPD2 expression was associated with disease aggressiveness, tumor progression, and adverse clinical outcomes. MYC was identified as a putative transcriptional driver of SNRPD2. High SNRPD2 expression was also linked to differential (oncogenic) alternative splicing of multiple cancer-associated genes, correlating with disease aggressiveness and poor clinical outcomes. These data establish SNRPD2 and its associated alternatively spliced repertoire as a central adaptive node linked to disease aggressiveness in gliomas, highlighting it as a potential therapeutic target in glioma patients.

Reprogramming somatic cells to an embryonic state opens a transformative pathway to convert cancer cells into benign ones. By delving into the changes that occur during this process, we can enhance our understanding of tumor development and unlock groundbreaking therapeutic strategies. In this study, we successfully reprogrammed the bladder cancer cell line using Yamanaka factors and conducted a stage-specific, comprehensive proteomic analysis of the resulting molecular changes. The bladder cancer cell line HTB-4 was reprogrammed and cultured on vitronectin-coated surfaces following Sendai virus reprogramming, enabling a thorough evaluation of pluripotent marker expression. Both parental and reprogrammed cells were tested for proliferation, migration, invasion, and colony formation. nLC-MS/MS analysis was performed to identify molecular differences between parental bladder cancer cells and reprogrammed cells across initial passages. Reprogrammed HTB-4 cells retain their ability to adhere and exhibit significant expression of pluripotency-associated proteins, forming colony-like structures. Stage-specific proteomic analyses reveal notable differences between reprogrammed cells and progenitor cells, particularly in pathways related to epithelial-mesenchymal transition, stem cell maintenance, and differentiation. We developed an in vitro model of bladder cancer reprogramming that identifies biomarkers associated with the induction of stem-like states and cellular plasticity. Our findings reveal significant stage-specific proteomic changes offering insights into the hierarchical organization of bladder cancer and the molecular mechanisms underlying the cancer stem cell phenotype. These results facilitate the development of more precise, patient-specific in vitro models for studying tumor recurrence and treatment resistance. However, further mechanistic studies are needed to translate effectively potential biomarkers into clinical practice.

Cancer stem cells (CSCs) play key roles in hepatocellular carcinoma (HCC) initiation, progression, therapeutic resistance, and recurrence, yet their cellular and spatial heterogeneity remains poorly understood. This study aimed to systematically characterize HCC-associated CSCs and identify prognostic biomarkers and potential therapeutic strategies using single-cell omics. Single-cell RNA sequencing and spatial transcriptomics data were obtained from HCCDB v2.0. Malignant cells were re-clustered using Harmony-based batch correction, followed by uniform manifold approximation and projection (UMAP) and Louvain clustering. Copy number variation analysis validated malignant identities. CSC-associated molecular features were characterized using differential expression, gene regulatory network analysis (SCENIC), pathway enrichment (GSVA), pseudotime trajectory inference (Monocle, CytoTRACE2), and cell-cell communication analysis (CellChat). CSC-specific genes were integrated with GEO survival datasets (GSE76427, GSE14520) to construct a prognostic model, and potential CSC-targeting compounds were predicted using Connectivity Map. Six malignant subpopulations were identified, including a progenitor-like CSC subset expressing EPCAM, SOX9, and SOX4. Spatial transcriptomics revealed CSC enrichment at the tumor-stroma interface. CSCs exhibited strong stemness, metabolic plasticity, and invasive potential, with activation of WNT/β-catenin, TGF-β, Notch, EMT, MYC, and mTORC1 pathways. Key transcription factors (TEAD2, SOX4, HNF1B, KLF7) were identified. A 12-gene CSC-derived signature stratified patients into distinct risk groups with significantly different overall survival. Several candidate compounds, including fluspirilene, genistein, and daunorubicin, showed potential CSC-suppressive activity. This study provides a comprehensive single-cell-based atlas of CSCs in HCC, highlighting their spatial niches, regulatory programs, and clinical relevance. The identified prognostic signature and candidate drugs offer promising avenues for CSC-targeted therapies.

Melanoma is a highly aggressive cancer in which metastatic dissemination remains the primary cause of mortality. This study aimed to define the role of the splicing factor 3b subunit 4 (SF3B4) in melanoma progression and its downstream regulatory mechanisms. SF3B4 expression was analyzed in public datasets. Its functional role was assessed by knockdown or inhibition in melanoma cells using proliferation, wound healing, and transwell assays. Talin1 expression and splicing were evaluated by RT-qPCR and immunoblotting, and FAK phosphorylation was measured as a downstream readout. SF3B4 is significantly upregulated in melanoma, particularly in metastatic lesions, and its expression correlates with poor patient survival. SF3B4 depletion suppresses melanoma cell growth and migration. Talin1 was identified as a downstream target of SF3B4, as SF3B4 knockdown reduced Talin1 mRNA and protein levels and impaired its splicing, leading to increased intron retention. Consistently, SF3B4 loss reduced phosphorylation of focal adhesion kinase (FAK), indicating attenuation of Talin1-mediated signaling. Talin1 knockdown recapitulated the migration defects observed upon SF3B4 depletion, and combined knockdown showed no additive effect, supporting a shared regulatory pathway. SF3B4 promotes melanoma cell migration through splicing-dependent regulation of Talin1. The SF3B4-Talin1 axis represents a potential therapeutic target in metastatic melanoma.

Lung cancer is the most lethal malignancy worldwide, and there remains an urgent need for reliable biomarkers to improve diagnosis and treatment. Nucleophosmin 1 (NPM1), a nucleolar phosphoprotein, has been implicated in hematological cancers, but its significance in lung cancer is less clear. This study investigated the oncogenic role of NPM1 in lung cancer and its involvement in ERK1/2 pathway activation in lung cancer cells. Transcriptomic data from TCGA were analyzed to assess NPM1 expression in lung cancer and normal tissues. In vitro assays using A549 and H1299 cells were conducted following siRNA-mediated silencing of NPM1. Cell proliferation, soft agar colony formation, and western blot analyses were performed. In vivo tumorigenicity was tested using a nude mouse xenograft model. NPM1 expression was significantly elevated in lung cancer tissues compared with normal samples. Silencing NPM1 reduced proliferation, colony formation, and tumor growth. Mechanistic studies revealed that NPM1 knockdown decreased phosphorylation of ERK1/2, indicating its role in activating this pathway. NPM1 contributes to lung cancer progression via ERK1/2 signaling. These results highlight NPM1 as a novel oncogene and suggest its potential as a diagnostic and prognostic biomarker in lung cancer.

Fibronectin 1 (FN1), located on chromosome 2q35, encodes fibronectin, a high molecular weight glycoprotein of the extracellular matrix. Several histologically overlapping chondroid matrix-producing tumors are known to harbor FN1 rearrangements, including soft tissue chondroma, synovial chondromatosis, calcifying aponeurotic fibroma, calcified chondroid mesenchymal neoplasm and phosphaturic mesenchymal tumor. Over the past 10 years, fusions involving the FN1 gene have also been identified in other mesenchymal neoplasms such as lipofibromatosis and inflammatory myofibroblastic tumor. The current World Health Organization Classification of Soft Tissue and Bone Tumors suggests that FN1-rearranged lesions are typically benign or intermediate. This review provides an updated overview of the clinical, histological and molecular genetic features of FN1-rearranged mesenchymal neoplasms and discusses their relationships with one another.

γ-Glutamylcyclotransferase (GGCT) depletion suppresses breast cancer cell proliferation by inducing cellular senescence. However, the underlying molecular mechanisms have not been fully elucidated. Therefore, the objective of this study was to elucidate the mechanisms by which GGCT depletion suppresses cancer cell proliferation. Human breast cancer MCF-7 cells were transfected with GGCT-specific or control siRNAs. Transcriptomic profiling by RNA sequencing identified differentially expressed genes (q<0.01, |log2 fold change|>1), and Gene Ontology and KEGG analyses characterized affected pathways. Key genes and functional effects on the TGF-β2/SMAD3 axis, cell-cycle progression, and senescence were validated by qRT-PCR, western blotting, and SA-β-Gal assays. Comprehensive gene expression analysis revealed that depletion of GGCT increases the expression levels of the cell cycle arrest factors CDKN1A (p21Cip1) and CDKN2B (p15INK4b), accompanied by elevated transforming growth factor-β2 (TGFB2) expression. Blocking this pathway through the simultaneous knockdown of TGFB2 was found to significantly restore the growth-inhibitory effect mediated by cellular senescence induced by GGCT depletion. This finding demonstrated that these phenotypes depend on the TGF-β2 pathway. Furthermore, we identified SMAD3 as a TGF-β2 downstream factor essential for the increase in p21Cip1 and p15INK4b and the growth-inhibitory effect induced by GGCT depletion. Activation of the TGF-β2/SMAD3 pathway is a mechanism by which cellular senescence is induced through GGCT depletion, suggesting that GGCT inhibition represents a promising therapeutic strategy for the treatment of breast cancer.

The association between the expression of protein kinase C zeta (PKCζ) and chemotherapeutic responses among different subtypes of breast cancer has yet to be fully elucidated. The present study aimed to investigate the association between PKCζ expression and disease-specific survival (DSS) rates, with a particular focus on the influence of chemotherapy and cancer stem cell (CSC)-associated ALDH1A3 expression. Clinical and gene expression data from the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) dataset (n=2,509) were analyzed using Kaplan-Meier and Cox proportional hazards models. The findings were validated using The Cancer Genome Atlas (TCGA) Pan-Cancer Atlas dataset (n=1,084). In the METABRIC dataset, high PKCζ expression (PKCζ high) was associated with poor DSS rates in patients with the normal-like, claudin-low and basal-like subtypes treated with chemotherapy. Consistent results were obtained in TCGA dataset, where PKCζ high expression in basal-like breast cancer predicted poor prognosis, especially among patients treated with cyclophosphamide, an alkylating agent. Combined analysis further revealed that patients with high expression of both PKCζ and ALDH1A3 (PKCζ high/ALDH1A3 high) basal-like tumors treated with cyclophosphamide, doxorubicin, or fluorouracil had the worst DSS rates compared with other groups. In addition, PKCζ high/ALDH1A3 high basal-like tumors treated with anthracycline- or taxane-based regimens also exhibited poorer prognoses compared with other groups. PKCζ contributes to chemotherapeutic resistance, especially in patients with ALDH1A3-positive basal-like breast cancer, possibly through the regulation of CSC survival and proliferation. Moreover, PKCζ, either alone or in combination with ALDH1A3 expression, may serve as a prognostic biomarker for predicting the therapeutic efficacy of chemotherapy in basal-like breast cancer.

Upper tract urothelial carcinoma (UTUC) has a notably high incidence and aggressiveness in East Asian populations; however, its molecular mechanisms remain poorly defined. Our previous study identified 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC), a bifunctional enzyme involved in de novo purine biosynthesis, as a tumor-promoting factor regulated by microRNA-145-5p in UTUC. Therefore, this study aimed to systematically investigate the functional interplay between ATIC and its downstream effectors in UTUC. To elucidate ATIC-regulated signaling pathways, we performed tandem mass tag (TMT)-based quantitative proteomics in BFTC909 cells with ATIC knockdown, followed by functional, biochemical, and drug-sensitivity assays. Proteomic profiling identified B7-H3 (CD276), prion protein (PRNP), RAC2, and NT5E (CD73) as downstream molecules downregulated by ATIC silencing. Functional assays revealed that suppressing the expression of these proteins inhibited cell proliferation, migration, and invasion, and enhanced cisplatin sensitivity. RNA interference analysis indicated that B7-H3 may lie upstream of prion protein and RAC2. Mechanistically, the ATIC/B7-H3 axis were shown to modulate mTOR, AKT, ERK, and p38 phosphorylation, linking metabolic activity to oncogenic and chemoresistant signaling. These findings revealed an ATIC-associated metabolic-immunoregulatory network in UTUC, through which ATIC supports mTOR-related signaling and promotes tumor progression and cisplatin resistance. Targeting the ATIC-driven network may offer new therapeutic opportunities for UTUC management.