Molecular Cancer

ObjectiveRNA adenosine-to-inosine (A-to-I) editing, catalyzed by adenosine deaminases acting on RNA (ADARs), is a pervasive post-transcriptional mechanism that expands transcriptomic and proteomic diversity. However, the global landscape of RNA A-to-I editing, as well as its functional and clinical significance in esophageal squamous cell carcinoma (ESCC), remains largely unexplored. This study aimed to systematically characterize the RNA editome of ESCC and elucidate its biological and clinical significance. MethodsWhole-transcriptome sequencing was performed on 121 primary ESCC tumors, with or without lymph node metastasis, together with matched normal tissues, to construct a high-resolution RNA editome. ADAR1-regulated RNA editing events were identified, and their functional consequences were investigated using integrated transcriptomic, phosphoproteomic, and RNA immunoprecipitation sequencing (RIP-seq) analyses. Associations between CDK13 editing, cGAS-STING-interferon-stimulated gene (ISG) signaling, and patient survival were further evaluated. ResultsA total of 222,020 high-confidence RNA editing sites were identified, of which approximately 98% were A-to-I events, including 124,486 ESCC-specific edits predominantly enriched in introns, 3' untranslated regions, and non-coding RNAs, highlighting a pervasive post-transcriptional regulatory layer. RNA A-to-I editing was extensively remodeled and globally up-regulated in non-metastatic ESCC, whereas only minimal changes were observed during lymph node metastasis, indicating that RNA editing alterations predominantly occur during early tumorigenesis. CDK13 emerged as a recurrent ADAR1 target, with RNA editing inversely correlated with CDK13 expression. ADAR1-mediated CDK13 editing established a positive feedback loop associated with enhanced interferon-stimulated gene (ISG) signaling and poorer survival in non-metastatic ESCC. Mechanistically, partial attenuation of CDK13 induced chronic, tumor cell-intrinsic activation of the cGAS-STING-ISG pathway. Integrated multi-omics analyses further revealed that CDK13 regulates phosphorylation networks governing cytoskeleton organization, intracellular trafficking, RNA homeostasis, and immune signaling. ConclusionRNA A-to-I editing represents a dynamic regulatory mechanism driving early ESCC progression and remodeling tumor cell-intrinsic immune signaling. ADAR1-mediated editing of CDK13 provides a mechanistic link between RNA editing and cGAS-STING-ISG pathway activation, revealing potential therapeutic vulnerabilities and supporting its utility as an early prognostic biomarker in ESCC.

Erlotinib resistance remains a critical barrier in treating EGFR-mutant non-small cell lung cancer (NSCLC). While distinct resistance mechanisms have been identified, the temporal evolution of transcriptional states and the role of non-coding RNAs in this process remain poorly understood. To address this, we performed a secondary single-cell RNA sequencing (scRNA-seq) analysis of PC9 cells treated with Erlotinib (GEO Accession: GSE149383). We employed pseudotime trajectory inference (Monocle3) and rigorous in silico modeling to map resistance evolution and predict miRNA-lncRNA interactions. Our trajectory analysis revealed a biphasic evolution of resistance: an early phase characterized by ribosomal stress responses (RPS5, RPL21) followed by a late proliferative phase driven by cell cycle regulators (CENPF, HMGB2). Notably, the long non-coding RNA NEAT1 showed dynamic upregulation during this transition. Computational modeling identified miR-124-3p as a high-confidence regulator of NEAT1, with structural analysis confirming a thermodynamically stable interaction ({Delta}G = - 14.8 kcal/mol). These findings suggest that Erlotinib resistance is not a static state but a dynamic process involving sequential transcriptional reprogramming. We propose the miR-124-3p/NEAT1 axis as a potential therapeutic target to disrupt the stress-adaptation phase of drug resistance.

Enhancer is a critical epigenetic feature in head and neck squamous cell carcinoma (HNSCC), yet the functional roles of individual enhancers remain poorly understood. Here, we conducted an integrative multi-omics analysis based on publicly available ATAC-seq, H3K27ac ChIP-seq, transcriptomic profiling, and genetic association datasets to systematically map HNSCC-associated enhancers and their target genes. Integrating ATAC-seq and H3K27ac ChIP-seq, we identified 20,362 enhancers and 18,040 enhancer-associated genes, highlighting widespread regulatory complexity. Functional characterization of the TERT-associated enhancer GH05J001312 (GRCh38/hg38: chr5:1312099-1317743) revealed strong transcriptional activity in HNSCC. CRISPR-mediated deletion of a core sequence significantly reduced TERT expression, impaired cellular proliferation in vitro, and suppressed tumor growth in vivo, confirming its role as a key cis-regulatory element. RNA-seq analysis of enhancer-edited cells uncovered 742 differentially expressed genes enriched in cancer-related pathways, including MAPK and IL-17 signaling, indicating a broad transcriptional impact. Collectively, our findings establish GH05J001312 as a functional enhancer driving oncogenic programs in HNSCC and suggest enhancer-targeted strategies as a potential therapeutic avenue.

Aggressive cancers such as triple-negative breast cancer (TNBC) and pancreatic cancer remain difficult to treat because their malignant behavior is driven by complex gene networks rather than single oncogenic targets. Our study identifies a NFAT3 transcription factor-dependent miRNAs signature that suppresses tumor aggressiveness through extracellular vesicles (EVs). Functional analyses demonstrated that a specific combination of fifteen miRNAs (miR-Comb 15) is required to inhibit cancer cell proliferation and invasion across TNBC and pancreatic cancer models, whereas individual miRNA are insufficient to reproduce the full anti-tumoral effect. To enable therapeutic translation, we have engineered EVs derived from HEK 293T, a non-tumoral and scalable vesicle source. Using an optimized exogenous pH-gradient loading strategy, miR-Comb 15 was efficiently incorporated into EVs without altering their integrity or intrinsic bioactivity. Among multiple delivery platforms tested, miR-Comb 15 loaded HEK 293T EVs consistently exhibited superior anti-tumoral efficacy both in vitro and in vivo. Together, these findings establish a strong mechanistic link between transcriptional regulation and EVs-mediated RNA delivery and demonstrates that rationally designed miRNA combinations delivered by EVs represent a promising therapeutic strategy for aggressive cancers.

Small-cell neuroendocrine carcinoma (SCNC) is a rare but highly malignant tumor subtype that primarily arises in the lung, also rarely in other organs, and as a consequence of treatment induced lineage transdifferentiation of prostate adenocarcinomas. The molecular convergence of SCNC across diverse tissues enables its identification through conserved SCNC-specific molecular markers, facilitating tumor subtype classification. As a critical post-transcriptional regulatory mechanism, alternative polyadenylation (APA) modulates 3'UTR length and significantly impacts tumor progression. However, its role in SCNC remains largely unclear. Here, we report a global 3'UTR lengthening pattern driven by APA in SCNC. We identified a set of conserved 3'UTR lengthening events across SCNCs of different tissue origins, which are strongly associated with neural development and related signaling pathways. Furthermore, we developed a neural network-based prediction model to classify SCNC by leveraging these specific APA signatures. Our study provides new insights into the post-transcriptional landscape of SCNCs and highlights APA signatures as promising biomarkers for SCNC identification.

A key limitation in cancer transcriptomics is the lack of accompanying genomic profiling such as whole-genome sequencing (WGS) or copy number alteration (CNA) data. Here we address this by showing that RNA-seq alone can be used to infer chromosomal aberrations and identify biologically meaningful patterns in lung cancer. Through a large-scale meta-analysis of publicly available RNA-seq datasets from non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), and matched controls, we reconstructed large scale CNA profiles and identified deletions in 3p, 9p, and 17p as the most frequent genomic events. Validation against paired WGS data confirmed a high degree of accuracy for RNA-seq-based inference. Our analyses revealed that while deletion-associated transcriptional heterogeneity exists, approximately 25% of differentially expressed genes were shared across all three deletion classes, indicating a conserved oncogenic program in lung cancer. Enrichment analysis linked these shared genes to pathways governing cell division, DNA replication, and extracellular matrix organization, while deletion-specific effects reflected disruption of tumor suppressor pathways, notably p53 signaling in 17p-deleted tumors, leading to deregulation of NOS2 and PLOD2. Integrating gene-level expression data, we identified both shared and deletion-specific biomarkers: TPX2 was consistently overexpressed across all deletion groups, while PTPRZ1 and CLDN9 were uniquely associated with del3p and del9p, respectively. Experimental validation in lung cancer cell lines confirmed these predictions, particularly the upregulation of CLDN9 in del9p carriers. By demonstrating that RNA-seq data can capture large-scale chromosomal events and reveal their transcriptional consequences, this study establishes an efficient framework for genomic inference and biomarker discovery, introducing CLDN9 as a novel, deletion-specific marker with potential prognostic and therapeutic value for 9p-deleted lung cancers.

The TGF-{beta} signaling pathway has both tumor-suppressing and metastasis-promoting effects in cancer. However, the molecular determinants governing this switch remain unclear. Here, we explored the miR-16-1-3p/MDM2/p53 axis as a critical conductor of the TGF-{beta}-Smad pathway in osteosarcoma. Although miR-16-1-3p overexpression by itself markedly reduces proliferative and clonogenic potential of U2OS cells, when paired with TGF-{beta} treatment, it significantly increases arrest cells in G1 phase and nearly extinguishing the growth capability of these cells. MiR-16-1-3p overexpression inhibited TGF-induced actin remodeling and EMT featuring, significantly decreasing vimentin levels. TGF-{beta} enhances both 2D and 3D migration, but miR-16-1-3p overexpression, alone or with TGF-{beta}, strongly counteracts its pro-migratory effects. MiR-16-1-3p restored p53 stability by targeting MDM2, redirecting TGF-{beta}-Smad signaling toward p21 activation and proliferation inhibition while attenuating its EMT-promoting capacity. Administration of TGF-{beta} together with miR-16-1-3p dramatically increases the sensitivity of wild-type U2OS cells to cisplatin, exceeding that of TGF-{beta} therapy alone by more than an order of magnitude. Administering TGF-{beta} and miR-16-1-3p together significantly reduces the tumor nodule volume and Ki67 expression, while effectively eradicates metastases in the chicken chorioallantoic membrane (CAM) in vivo model. For the first time, our research demonstrates that miR-16-1-3p shifts TGF-{beta}1 signaling from a facilitator of metastasis to a promoter of anti-growth effects through MDM2 inhibition and p53 stabilization, effectively reducing the self-renewal and invasiveness of cancer stem cells in human osteosarcoma model. This process preserves TGF-{beta}s tumor-suppressive role while limiting its associated cancer risks.

Circular RNAs (circRNAs) constitute a novel class of noncoding RNAs showcasing distinct tissue- and cell-specific expression patterns. Despite the extensive profiling of circRNAs, their individual functions remain poorly understood. To fill this gap, we designed a genome-wide library of 65,300 shRNAs, targeting 9,663 clinically relevant circRNAs and 3,981 of their linear parental genes, and conducted functional screening in seven types of human cancer. We identified a total of 1,342 essential circRNAs (13.9% screened) that impact cell proliferation in at least one cell line, and in 96.5% of the cases, the linear counterparts are not essential. While a shared common subset emerges as functional regulators across all examined cell lines, the majority of circRNAs are functional in a cell type-specific manner. For a comprehensive presentation of the functional circRNA landscape in cancer, we introduce FunCirc, an online database encompassing functional circRNAs across cancer cell lines, coupled with circRNA expression profiles from diverse cancer and tissue types. Our work enhances the understanding of circRNA functions in cancer and provides the scientific community with a resource to further investigate their intricate roles.

Reliable non-invasive biomarkers for tumor grading and disease monitoring in pediatric brain tumors are an unmet clinical need. Circulating extracellular vesicles (EVs) carrying molecular cargo reflective of tumor biology, offer promise as liquid biopsy tools. We have previously discovered EV-associated Syndecan-1 (SDC1) to be overexpressed in malignant brain tumors, but its value as a biomarker in pediatric disease remains unclear. In this study, plasma EVs were isolated from pediatric brain tumor patients (n=60) by size-exclusion chromatography and characterized using cryo-electron microscopy, nanoflow cytometry, immunoblotting, and single-vesicle total internal reflection fluorescence imaging. EV-associated SDC1 (EV-SDC1) was quantified and analyzed in relation to tumor grade, subtype, surgical resection status, and tumor volume. EV-SDC1 levels were significantly elevated in high-grade (ependymoma, diffuse midline glioma, and atypical teratoid/rhabdoid tumor (AT/RT)) compared with low-grade pilocytic astrocytoma tumors and robustly discriminated grade 3 tumors from pilocytic astrocytoma (AUROC 1.00). Independent validation using transcriptomic data from the Open Pediatric Brain Tumor Atlas showed SDC1 mRNA levels to effectively distinguish high grade (ependymoma, medulloblastoma, diffuse midline glioma, and AT/RT) from pilocytic astrocytoma patients. Furthermore, EV-SDC1 levels decreased following complete tumor resection but remained elevated in patients with residual disease or recurrence. Collectively, circulating SDC1-positive EVs represents a clinically informative biomarker reflecting tumor aggressiveness and treatment response in pediatric brain tumors, supporting their potential for non-invasive disease stratification and monitoring.

Neuroblastoma (NB) is the most common extracranial solid tumor in children. Relapsed or refractory (R/R) high-risk (HR) NB tumors continue to exhibit poor outcomes despite intensive and protractive multimodal therapy. Activating mutations in the RAS- mitogen-activated protein kinase (MAPK) pathway are frequently observed in R/R HRNB. The early promise of ALK inhibitors to treat ALK-mutant NB underscores the ability of appropriate targeted therapies to improve outcomes for HRNB patients. While MAPK pathway activation is prominent in HRNB, FDA-approved MEK inhibitors and KRAS G12C inhibitors have failed to demonstrate significant preclinical single-agent activity. Daraxonrasib (RMC-6236), a potent and selective RAS(ON) inhibitor, has demonstrated activity in both preclinical models and early phase clinical trials of RAS-mutant adult cancers. A subset of R/R HRNB tumors is noteworthy for containing diverse RAS-mutations, providing rationale for RMC-6236 investigation. In this study, we evaluated the therapeutic efficacy and oncogenic signaling modulation of RMC-6236 across NB models harboring RAS pathway activation. RMC-6236 as a single-agent treatment led to a significant decrease in cell viability, suppression of downstream MAPK signaling, upregulation of the MAPK pathway effector protein BIM, and increased cell death in RAS-mutant NB models as well as in NF1-mutant NB models. In vivo studies evidenced that RMC-6236 had on-target activity that significantly reduced tumor growth and extended survival in RAS-mutant HRNB mouse models. Furthermore, RMC-6236-induced both BCL-2 and BIM upregulation and enhancement of BIM:BCL-2 complexes in RAS-mutant NB. As such, the BCL-2 inhibitor venetoclax further enhanced RMC-6236-mediated killing by disrupting RMC-6236 enhanced BIM:BCL-2 complexes. These findings demonstrate that RMC-6236 is a rationale targeted therapy for RAS-mutant NB, a subset of NB that is progressively understood as conferring particularly poor outcomes. RMC-6236 is a clinically relevant drug that can successfully target the MAPK pathway in these cancers. This study supports expanded clinical testing of this novel therapy to this important subset of neuroblastoma.

Glioblastoma (GBM) is an aggressive, high-grade glioma with a near-universally fatal prognosis. Therapeutic failure is often attributed to the highly selective blood brain barrier (BBB), the diffuse infiltrative nature of the tumor, and the marked intratumoral heterogeneity of GBM. Although antibody drug conjugates ADCs have shown promise for high grade gliomas such as GBM, efficacy is limited by ADC size. Aptamers--short, synthetic, single-stranded DNA or RNA molecules--can be [~]6-fold lower in molecular weight than IgG antibodies and have the potential to cross the intact BBB. Unlike other nucleic acid-based therapies, aptamer function arises from three-dimensional shape rather than genetic coding. Here we aim to replace the targeting component of the ADC paradigm with a DNA aptamer, thus creating an aptamer-drug conjugate (ApDC). We employed in vivo SELEX using an orthotopic patient-derived xenograft (PDX) GBM mouse model and a vast ([~]100 trillion 80-mer sequences) ApDC library. We report the results from this first in vivo ApDC selection of its kind. We characterize target tissue binding ex vivo, cell association, biodistribution, and pharmacokinetics from this selection. This study exemplifies an unbiased approach to a problem that rational design has yet to overcome, offering a new direction for GBM therapeutic development.

Differentiation-based therapies represent a promising strategy for the treatment of neuroblastoma; however, single-agent approaches frequently yield incomplete and transient responses due to the robustness of underlying gene regulatory networks. MicroRNAs (miRNAs) are endogenous regulators of gene expression that modulate entire gene programs rather than individual molecular targets, making them attractive candidates for network-level therapeutic intervention. While individual miRNAs have been investigated as therapeutic agents, the potential for synergistic interactions between miRNAs remains largely unexplored. Here, we developed a scalable high-content phenotypic screening platform to identify synergistic miRNA combinations that promote neuronal differentiation and growth arrest in neuroblastoma cells. Using SK-N-BE(2)-C cells and automated quantification of neurite outgrowth and confluence, we screened pairwise combinations of differentiation-associated miRNAs at submaximal doses. Candidate synergistic interactions were identified using the Highest Single Agent framework and subsequently validated by dose-response interaction modeling. We identified a robust synergistic interaction between miR-124-3p and miR-363-3p that exceeded zero-interaction potency expectations by approximately 20.9% and increased maximal differentiation-associated phenotypic response by 73% relative to single-miRNA treatments. Target gene and pathway enrichment analyses revealed that miR-124-3p and miR-363-3p regulate largely distinct but functionally complementary target gene sets. These complementary targets converged on neuronal differentiation and cell cycle control pathways, providing a mechanistic basis for their cooperative activity. Together, these findings establish miRNA combinations as programmable network regulators capable of inducing complex cellular phenotypes with greater efficacy than single agents. This work provides a conceptual and experimental framework for the rational discovery of synergistic miRNA therapeutics and suggests new avenues for differentiation-based treatment strategies in neuroblastoma and other diseases driven by dysregulated regulatory networks.

tRNA-derived fragments (tRFs) are relatively recently discovered class of small RNAs implicated in gene-regulatory processes in diverse biological contexts but there have been very few reports of a clear phenotypic role of these small RNAs in cancer progression. By analyzing small RNA-seq data from The Cancer Genome Atlas (TCGA), we found that high expression of three 3' tRFs (tRF-3a), tRF-3009a, tRF-3021a or tRF-3030a, is significantly associated with poor overall survival in low-grade glioma (LGG). In glioblastoma cells, tRF-3009a, tRF-3021a and tRF-3030a enhance cell invasion and migration but tRF-3021a was uniquely required for cell proliferation and suppression of apoptosis. Interestingly, tRF-3021a knockdown decreases global protein synthesis prior to and independent of apoptosis. These data indicate that tRF-3021a supports glioma cell survival and particularly protein synthesis while promoting cellular invasion and migration. Given its association with poor outcome in LGG patients, tRF-3021a represents a promising biomarker and potential therapeutic target in gliomas and these results provide a foundation for future studies to define its molecular interactors and downstream pathways controlling protein synthesis and apoptosis in cancer cells. ImplicationtRF-3021a promotes malignant glioma phenotypes, sustains global protein synthesis and prevents spontaneous apoptosis, motivating efforts to evaluate it as a biomarker and therapeutic target.

Diffuse hemispheric gliomas (DHGs) are highly aggressive and infiltrative CNS tumors that are refringent to treatment, and with a 5-year overall survival of around 20%. A fraction of DHGs is driven by mutations in the histones H3.1 and H3.3. In this study, we demonstrate that the expression of histone H3.3 glycine 34 to arginine mutations (H3.3-G34R) result in the epigenetic and transcriptional activation of the NF-{kappa}B signaling pathway in DHG. To target this vulnerability, we designed high density lipoprotein (HDL) nanoparticles loaded with unmethylated CpG dinucleotides, which mimic the immune stimulatory activity of bacterial DNA. CpG are recognized by Toll-like receptor 9 (TLR9), activating the NF-{kappa}B signaling. The CpG-mediated NF-{kappa}B activation results in the release of immuno-stimulating cytokines that promote an antitumoral response. As we previously established that G34-mutant DHGs are characterized by DNA repair impairment, we combined CpG dinucleotides with a PARP (poly (ADP-ribose) polymerase) inhibitor, olaparib, in the HDL nanoparticles.

Thoracic malignancies, including lung adenocarcinoma (ADC) and malignant pleural mesothelioma (MPM), remain associated with poor prognosis and limited durable therapeutic responses in advanced stages. Although targeted therapies and immunotherapy have improved outcomes in selected patients, systemic chemotherapy continues to play a central role in routine clinical practice. However, treatment response is highly heterogeneous, and reliable predictive biomarkers of chemotherapy sensitivity are lacking. Both ADC and MPM frequently involve the pleural cavity and are commonly associated with malignant pleural effusion (MPE), which contributes to symptoms such as dyspnea and chest pain and requires therapeutic drainage. Importantly, MPE represents a clinically accessible source of viable tumor cells obtained through minimally invasive procedures. In this study, we established patient-derived organoids (PDOs) from malignant pleural effusion samples obtained from five patients with advanced lung adenocarcinoma and, as an exploratory extension, from one patient with malignant pleural mesothelioma. Organoids were characterized by immunohistochemistry and subjected to systematic chemotherapy drug screening. Inter-model variability in treatment response was assessed, and selected drug sensitivities were further validated through dose-response assays. Pleural effusion-derived organoids successfully recapitulated tumor-specific phenotypic features and revealed marked heterogeneity in chemotherapy sensitivity across models. Secondary validation confirmed the reproducibility of selected responses. Our findings support the feasibility of generating functional organoid models from malignant pleural effusions and highlight their potential as translational platforms for individualized chemotherapy profiling in advanced thoracic malignancies.

Oncolytic virotherapy exploits viruses to selectively infect and destroy cancer cells while sparing normal tissues and represents a promising strategy in oncology. Human adenovirus type 5 (HAd5), although widely used, shows limited clinical efficacy due to high levels of preexisting immunity and suboptimal tumor selectivity. In this study, we evaluated novel gorilla-derived adenoviruses (GRAd) as alternative oncolytic vectors. Two distinct GRAd groups, GRAdBs and GRAdCs, were characterized for replication and cytopathic activity. GRAd25 (GRAdB group) exhibited robust replication in both tumor and normal cells, whereas GRAd32 (GRAdC group) demonstrated selective replication in tumor cells. To broaden tumor tropism while preserving selectivity, we generated a chimeric GRAd32 vector, GRAd32Fk25, by replacing its native fiber knob with that of GRAd25, potentially shifting receptor usage from CAR to CD46, which is more abundantly expressed in tumor cells. The vector was further armed with a therapeutic antibody by inserting the coding sequence for the single-chain Fc form (scFv-Fc) of EV20, a humanized anti-HER3 antibody, under endogenous viral regulatory control. In vitro analyses showed that GRAd32Fk25 maintained tumor-restricted replication and produced functional EV20 capable of binding HER3 and inhibiting downstream PI3K/Akt signaling. These results indicate that engineered GRAd vectors, exemplified by GRAd32Fk25 armed with EV20, provide a selective and versatile platform for oncolytic virotherapy with potential advantages over HAd5-based approaches.

Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive sarcomas with poor prognosis and a strong tendency for metastasis and relapse. Surgical removal remains the mainstay of treatment but is frequently ineffective or impractical. Currently, no effective targeted therapy exists for this type of malignancy. PRMT5 has recently emerged as a promising therapeutic target in various human cancers with MTAP loss, which results in cancer cell dependency on PRMT5 activity. The frequent loss of MTAP in MPNSTs suggests that PRMT5 inhibition is a promising therapeutic option and enables the stratification of cancer patients with few treatment options. We first examined human nerve sheath tumor samples and found that increased PRMT5 expression and activity correlated with MTAP loss in 86.8% (33/38) of MPNSTs and in atypical neurofibromatous neoplasm with uncertain biologic potential (ANNUBP) (5/5). When PRMT5 activity was inhibited genetically and chemically, the cell growth of MTAP-deficient MPNST cell lines was suppressed, but not that of MTAP-proficient MPNST cell lines. Moreover, in the PRMT5-inhibited MTAP-deficient MPNST cell lines, spontaneous DNA damage accumulation was observed following G2/M cell cycle arrest. The DNA replication stress marker RPA32 decreased, and CHK1 was activated early after PRMT5 knockdown, likely contributing to the accumulation of DNA damage. In addition, we combined PRMT5 inhibition with the DNA-damaging agents doxorubicin and gemcitabine, resulting in synergistic effects and increased cancer cell death in MTAP-deficient MPNST cell lines. Together, these findings identify PRMT5 as a compelling therapeutic target in MTAP-deficient MPNSTs. This PRMT5 inhibition strategy has strong translational potential for MPNSTs. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=181 SRC="FIGDIR/small/710638v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@15abc35org.highwire.dtl.DTLVardef@1fa4ebborg.highwire.dtl.DTLVardef@470c51org.highwire.dtl.DTLVardef@79cc3f_HPS_FORMAT_FIGEXP M_FIG C_FIG

Glioblastoma (GBM) is a highly aggressive brain tumor characterized by extensive heterogeneity, diffuse invasion, and recurrence despite multimodal therapy. Aberrant transcriptional programs driven by oncogenic signaling pathways sustain GBM growth, stemness, and therapy resistance, yet targeting individual molecular nodes has yielded limited clinical benefit. Here, we introduce a transcriptional rewiring strategy based on an engineered epigenetic silencer factor (ESF) targeting the YAP/TAZ-TEAD axis. We developed a TEAD1 Epigenetic Silencer (TES) by fusing the DNA-binding domain of TEAD1 to repressive epigenetic modules. TES selectively binds TEAD genomic targets and imposes stable transcriptional repression of YAP/TAZ-dependent gene programs through chromatin remodeling and DNA methylation. Genome-wide analyses revealed that TES preserves TEAD1 DNA-binding specificity while converting an oncogenic transcriptional platform into a repressive state. Functionally, TES impaired proliferation, induced cell death, and reduced migratory and invasive properties in glioma cell lines and patient-derived cancer stem-like cells. In vivo, TES significantly reduced tumor growth in orthotopic GBM xenograft models and enhanced the therapeutic efficacy of temozolomide. Importantly, TES was well tolerated by normal neural cells in vitro and in the adult mouse brain in vivo. These findings establish TES as a proof-of-concept epigenetic therapy to durably suppress oncogenic transcriptional networks in GBM.

Cancer is the leading cause of disease-related deaths among children in high-income countries. Tumor heterogeneity and lack of mechanism-of-action-based therapeutic options are key challenges to overcome in order to improve pediatric cancer patients survival. Here, we report the EU-IMI-2 funded public-private partnership "ITCC-Pediatric Preclinical Proof-of-Concept Platform" (ITCC-P4), which has built a large repertoire of patient-derived xenograft (PDX) models, representing all major solid pediatric cancer types, for in vivo drug testing. Three-hundred-fifty-three PDX models from diagnostic and relapsed pediatric cancers have been established and molecularly characterized, together with matched germline/tumor samples. As proof-of-concept, we present in vivo drug screening data in neuroblastoma and rhabdomyosarcoma models. PDX data, accessible at http://r2platform.com/itcc-p4, allow the selection of models based on oncogenic drivers and/or potential biomarkers for preclinical testing. Operated by a non-profit entity (www.itccp4.com), this sustainable platform aids academic and industrial researchers in developing and prioritizing innovative therapies for pediatric cancer. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/703023v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@195ba30org.highwire.dtl.DTLVardef@f2c2d9org.highwire.dtl.DTLVardef@1d63f4dorg.highwire.dtl.DTLVardef@d60027_HPS_FORMAT_FIGEXP M_FIG C_FIG

Head and neck squamous cell carcinoma (HNSCC) remains associated with substantial morbidity and a 5-year overall survival rate of approximately 60%, reflecting persistent radio- and chemo-resistance and the lack of effective precision medicine strategies. Patient-Derived Tumor Organoids (PDTO) constitute promising functional models that may predict individual treatment response. In this study, we generated PDTO from surgically resected HNSCC of the oral cavity, oropharynx, larynx, and hypopharynx. A total of 20 long-term PDTO lines were established, maintaining growth over seven passages and successfully cryopreserved, capturing the molecular and clinical diversity of the patient cohort. These PDTO faithfully recapitulated histological features, major tumor marker expression, and the genomic and transcriptomic landscapes of their tumors of origin, with stability over time. Functional assays revealed heterogeneous responses to cisplatin and X-rays. Importantly, in vitro sensitivity of PDTO was associated with clinical outcome of patients at 24 months. Cisplatin response of PDTO predicted prognosis with 66.7% sensitivity and 100% specificity, while X-ray response showed 91.7% sensitivity and 75% specificity. Notably, all patients whose PDTO were classified as resistant to both cisplatin and X-rays experienced relapse and/or death within 24 months. Collectively, the successful long-term expansion and cryopreservation of HNSCC PDTO establish a stable and scalable preclinical resource that captures the molecular and clinical heterogeneity of the disease. This biobank provides a valuable platform for mechanistic studies and for the evaluation of innovative therapeutic strategies. This cohort represents one of the largest clinically annotated HNSCC PDTO collections to date, demonstrating a robust association between PDTO response to cisplatin and X-rays and patient prognosis. These findings support the predictive potential of PDTO-based functional assays and argue for their integration into standardized, rapid, and miniaturized precision oncology workflows for HNSCC.