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PubMed2026

Stem Cell Differentiation Disperses Transcriptional Clusters via a Conserved Surface-Condensate Trajectory.

Stem cells exhibit exceptionally prominent transcriptional clusters, which dissolve with progressing differentiation. Although these clusters are assigned central roles in embryonic gene regulation, their formation and loss during differentiation remain poorly understood. This study reveals that these prominent clusters disperse along a conserved trajectory in mouse embryonic stem cells, fruit fly testes, and zebrafish embryos. Imaging and lattice simulations show that these clusters form via surface condensation on H3K27ac-marked super-enhancer regions, which act as genomic scaffolds. Upon differentiation, partial loss of these active epigenetic marks and transcription-driven unfolding lead to dispersal of the prominent clusters. The block copolymer-based lattice simulations explain this process as a conserved trajectory through a three-dimensional state space, governed by surface condensation principles that extend beyond canonical liquid-liquid phase separation. This work marks surface condensation as a biophysical mechanism for the dynamic organization of stem cell-specific transcriptional hubs and demonstrates evolutionary conservation in several organisms. By uncovering a conserved biophysical mechanism for transcriptional organization in development, our work illustrates how polymer properties can contribute to the control of cell identity and fate.

Animals↗

PubMed2026

Selective control of HLA-DRB1 and HLA-DQA1 transcription through DR/DQ super enhancer microelements.

MHC-II gene expression requires promoter-proximal and distal organizing elements. A super enhancer located between the HLA-DR and -DQ genes was dissected into five microdomains (regions A-E) to define its mechanism of action. Regions A and B were required for maximal expression of HLA-DQA1. Region A facilitated HLA-DQA1 expression and substituted for region B in its absence, as well as mediating longer-range interactions with distal CTCF sites. Region D modulated HLA-DRB1 through specific 3D chromatin interactions, whereas a local insulator element (XL9) broadly interacted with the SE. Through deletion of HLA-DRB1 proximal-promoter elements, HLA-DQ expression increased by co-opting interactions with region D, indicating that competition between microelements regulates the absolute levels of MHC-II genes. Together, these data define an additional layer of control for MHC-II genes encoded in one of the most polymorphic and disease-associated regions of the human genome.

CP: immunology↗

PubMed2026

Genome-wide CRISPR screen identifies a cytokine-enhancer circuit driving HIF-2α activation in renal cancer.

Resistance to HIF-2α inhibitors such as belzutifan underscores the need to better understand how HIF-2α is transcriptionally regulated in clear cell renal cell carcinoma (ccRCC). Here, we uncover a cytokine-driven enhancer mechanism that sustains HIF-2α expression through the JAK1/STAT3 signaling pathway. Using a genome-wide CRISPR screen in von Hippel-Lindau-deficient (VHL-deficient) ccRCC cells, we identified SOCS3 as a key negative regulator of HIF-2α. Mechanistically, loss of SOCS3 activates JAK1/STAT3 signaling, leading to the recruitment of STAT3 to distal enhancers upstream of endothelial PAS domain-containing protein (EPAS1) that physically loop to its promoter to drive HIF-2α transcription. This cytokine-enhancer circuit was recapitulated in samples from patients with ccRCC and functionally validated using CRISPR interference (CRISPRi), which disrupted enhancer-promoter looping and reduced tumor growth in HIF-2α-dependent models. SOCS3 overexpression or pharmacologic inhibition of JAK1/STAT3 markedly suppressed HIF-2α expression and tumor progression both in vitro and in vivo. Unlike prior studies focusing on VHL/HIF occupancy-driven enhancer activation, this work defines a trans-acting cytokine-JAK1/STAT3 pathway that transcriptionally controls EPAS1. Together, these findings reveal a targetable enhancer mechanism that sustains HIF-2α expression and suggest that combined inhibition of JAK1/STAT3 and HIF-2α may overcome therapeutic resistance in kidney cancer.

Basic Helix-Loop-Helix Proteins↗

PubMed2026

Noncoding transcription controls the developmental dynamics of long-range gene regulation.

The genomic regions regulating gene expression are often themselves transcribed into a variety of noncoding RNAs (ncRNAs). However, the regulatory roles of this noncoding transcription remain largely unknown. By using live imaging, we reveal that the sequential transcription of ncRNAs emanating from distinct regulatory elements underlies gene activation in Drosophila embryos. Single-allele co-visualization uncovers that optimal gene activation is achieved by only moderate levels of enhancer activity. Disrupting enhancer-associated ncRNAs causes precocious gene activation, providing evidence that ncRNAs control the timing of gene expression in development. We further show that enhancer transcription can regulate long-range interactions within complex regulatory landscapes. We propose that ncRNAs locally modulate regulatory element activity in cis to shape genome organization and orchestrate the temporal control of gene expression in development.

Journal Article↗

PubMed2026

Deregulation of FOXF1/FENDRR from t(14;16)(q32;q24) defines a subtype of high-risk lineage ambiguous leukemia.

Despite great progress in understanding the genomic basis of immature T-cell acute lymphoblastic leukemia/lymphoblastic lymphoma (T-ALL) and acute leukemias of ambiguous lineage, there are still cases that lack defining genetic markers, complicating risk stratification, and limiting targeted therapeutic options. Recent studies have shown that enhancer hijacking drives oncogene activation in approximately half of T-ALL cases, with the BCL11B (BCL11 transcription factor B) enhancer frequently involved. Here, we describe a subtype of leukemia with a distinct gene-expression signature, and immunophenotype characterized by positivity for immature (CD38), myeloid (CD13), T-lymphoid (cytoplasmic [c]CD3, CD7), and B-lymphoid markers (CD19, CD79a, CD10). This subtype is defined by the t(14;16)(q32;q24) translocation, which places the FOXF1 gene and its antisense long noncoding RNA gene FENDRR under the regulatory control of the BCL11B enhancer, leading to their ectopic transcriptional activation. Common concomitant genetic lesions are loss-of-function alterations of GATA3, CDKN2A/CDKN2B deletion and activating JAK/STAT and NOTCH1 pathway mutations. Patients were predominantly children and adolescents/young adults and experienced poor treatment outcome. High-throughput drug screening of 176 compounds demonstrated efficacy of combined BCL2 apoptosis regulator family proteins and JAK/STAT signaling inhibitors. Additionally, the clinical use of tyrosine kinase inhibitors in some of these patients showed therapeutic efficacy. Collectively, these findings identify BCL11B-enhancer-mediated deregulation of FOXF1/FENDRR as a hallmark of a subtype of high-risk lineage ambiguous leukemia that is potentially amenable to targeted therapeutic intervention.

Humans↗

PubMed2024

Exploring the transcriptional cooperation between RUNX2 and its associated elncRNA RAIN.

Recent insights into the mechanisms controlling gene expression identified enhancer-associated long non-coding RNAs (elncRNAs) as master players of transcription in cancers. RUNX2, a mammalian RUNT-related transcription factor, is increasingly recognized in cancer biology for its role in supporting survival and progression also in thyroid cancer (TC). We recently identified, within the RUNX2 locus, a novel elncRNA that we named RAIN (RUNX2 associated intergenic lncRNA). We showed that RAIN and RUNX2 expression correlate in TC, both in vitro and in vivo, and that RAIN promotes RUNX2 expression by interacting with and affecting the activity of the RUNX2 P2 promoter through two distinct mechanisms. Here, we took forward these observations to explore the genome-wide transcriptional function of RAIN and its contribution to the RUNX2-dependent gene expression program in TC. By combining multiple omics data, we demonstrated that RAIN functionally cooperates with RUNX2 to the regulation of a subset of functionally related genes involved in promoting matrix remodeling, migration, and loss of differentiation. We showed that RAIN interacts with RUNX2 and its expression is required for the efficient recruitment of this TF to its target regulatory regions. In addition, our data revealed that besides RUNX2, RAIN governs a hierarchically organized complex transcriptional program by controlling a core of cancer-associated TFs that, in turn, orchestrate the expression of downstream genes. This evidence indicates that the functional cooperation observed between RAIN and RUNX2 can be a diffuse work mechanism for this elncRNA.

Core Binding Factor Alpha 1 Subunit↗

PubMed1979

Adenosine 3',5'-monophosphate effects on cartilage ribonucleic acid synthesis.

We have previously shown that N6-monobutyryl-cAMP stimulates the incorporation of radiolabeled precursors into the macromolecules of embryonic chicken cartilage incubated in organ culture. The present study assessed the specific effects of N6-monobutyryl-cAMP on RNA synthesis in the cartilage organ culture system. The data show that treatment of cartilage with N6-monobutyryl-cAMP results in a coordinated stimulation of mRNA, rTNA, and tRNA synthesis. Incubation of cartilage with N6-monobutyryl-cAMP causes an increase of [5-3H]uridine incorporation into poly A-containing cartilage RNA and 28S, 18S, and 4S RNA as well. Increased incorporation of the radiolabeled nucleotide occurs without a measurable change in uridine triphosphate specific activity or in the rate of RNA degradation. Further, the isolated poly A-containing RNA directs protein synthesis in an in vitro cell-free system, and stimulation of cartilage mRNA by N6-monobutyryl-cAMP does not alter the size distribution of the mRNA species synthesized from that present in control tissue. These data indicate that cAMP enhances cartilage transcriptional activity. In addition, the increased synthesis of mRNA occurs antecedent to demonstrable effects on protein synthesis. Thus, the effects of cAMP on protein synthesis may be secondary to action at the nucleus.

Animals↗

PubMed2023

Genome-wide identification of transcriptional enhancers during human placental development and association with function, differentiation, and disease†.

The placenta is a dynamic organ that must perform a remarkable variety of functions during its relatively short existence in order to support a developing fetus. These functions include nutrient delivery, gas exchange, waste removal, hormone production, and immune barrier protection. Proper placenta development and function are critical for healthy pregnancy outcomes, but the underlying genomic regulatory events that control this process remain largely unknown. We hypothesized that mapping sites of transcriptional enhancer activity and associated changes in gene expression across gestation in human placenta tissue would identify genomic loci and predicted transcription factor activity related to critical placenta functions. We used a suite of genomic assays [i.e., RNA-sequencing (RNA-seq), Precision run-on-sequencing (PRO-seq), and Chromatin immunoprecipitation-sequencing (ChIP-seq)] and computational pipelines to identify a set of >20 000 enhancers that are active at various time points in gestation. Changes in the activity of these enhancers correlate with changes in gene expression. In addition, some of these enhancers encode risk for adverse pregnancy outcomes. We further show that integrating enhancer activity, transcription factor motif analysis, and transcription factor expression can identify distinct sets of transcription factors predicted to be more active either in early pregnancy or at term. Knockdown of selected identified transcription factors in a trophoblast stem cell culture model altered the expression of key placental marker genes. These observations provide a framework for future mechanistic studies of individual enhancer-transcription factor-target gene interactions and have the potential to inform genetic risk prediction for adverse pregnancy outcomes.

Humans↗

PubMed2026

Mediator at the Helm: Coordinating transcription and biomolecular condensates in hematopoiesis.

Hematopoiesis relies on precisely coordinated transcriptional programs that balance stem cell self-renewal, lineage commitment, and terminal differentiation. Central to this regulation is the Mediator complex, a large multi-subunit transcriptional co-regulator that integrates signals from transcription factors and chromatin regulators to control RNA polymerase Ⅱ (Pol Ⅱ) activity. The dynamic and modular composition of Mediator enables context-dependent transcriptional outputs, while individual subunits can exert specialized regulatory functions during hematopoietic lineage specification, thereby contributing to cell-fate-specific transcriptional outputs. Recent advances further reveal that transcriptional regulation is shaped by the spatial organization of regulatory machinery with biomolecular condensates formed through liquid-liquid phase separation (LLPS), particularly at super-enhancers. In this emerging framework, Mediator functions not only as a transcriptional integrator but also as a key coordinator of transcriptional machinery within condensates at cell-fate-related gene loci. In this chapter, we summarize how distinct Mediator subunits confer specific modes of transcriptional regulation and discuss how the interplay between Mediator and phase-separated condensates shapes transcriptional control during hematopoiesis. We highlight how specific subunits, including MED1 and MED26, participate in distinct regulatory modes in erythropoiesis, spanning super-enhancer-driven transcriptional activation, progenitor expansion, and condensate-associated mechanisms that influence Pol Ⅱ pausing and global transcription repression during terminal differentiation. Together, these findings support a model in which Mediator integrates transcriptional regulation with nuclear organization through condensate-mediated mechanisms, providing a conceptual framework for understanding hematopoietic cell fate decisions and transcriptional dysregulation in hematological diseases.

Hematopoiesis↗

PubMed2026

Allele-specific chromatin architecture shapes imprinted domains and coordinates a distal enhancer and antisense transcription at the mouse Mest-Copg2 domain.

Genomic imprinting results in parent-of-origin-dependent gene expression, but how three-dimensional genome organization contributes to imprinted gene regulation remains unclear. Using Capture Hi-C in mouse cortex and primary cortical neurons, we identified parental allele-specific chromatin architectures across multiple imprinted domains. These architectures largely originate from imprinting control regions and correlate with DNA methylation-sensitive CTCF binding. Active and inactive alleles of imprinted genes show distinct promoter interaction profiles and differential engagement with distal regulatory elements in both contact frequency and the epigenetic state of distal regions. A CRISPR interference screen identified a distal enhancer that regulates Mest-Copg2 imprinted expression through allele-specific chromatin interactions. In neurons, this enhancer activates Copg2 on the maternal allele, whereas on the paternal allele it drives Mest isoforms transcribed antisense to Copg2 and contributes to Copg2 repression. In summary, we show that allele-specific chromatin architecture coordinates maternal enhancer activity and paternal antisense transcription to control imprinted expression in neurons.

Animals↗

PubMed2025

Synthetic allopolyploidy unveils hybridization-driven transcriptional reprogramming underlying thermal adaptation in Cucumis.

Both heterosis (hybrid vigor) resulting from hybridization and genetic plasticity conferred by whole-genome duplication (WGD) are recognized as drivers of evolutionary success and ecological adaptation in plants. Allopolyploids, which combine both hybridization and WGD, are widespread in both natural and agricultural settings and often exhibit superior performance. However, the relative contributions of these two elements to the success of allopolyploids remain poorly understood. Here, we employed an experimentally reconstructed allotetraploid Cucumis species (C. × hytivus, 2n = 4x = 38) and its diploid interspecific hybrid progenitor (allodiploid, 2n = 2x = 19) to decouple and investigate the distinct and combined contributions of hybridization and whole-genome doubling to immediate genetic and phenotypic consequences of allopolyploid formation under environmental stress. Both C. × hytivus and the allodiploid exhibited superior heat tolerance compared with the parental species with significantly higher semi-lethal temperature and enhanced physiological acclimation capacity. While the allodiploid and allotetraploid retain transcriptomic features where differences persist (e.g., WGCNA modules), comparative analysis of the 15,680 homoeologous gene pairs in the allodiploid and allotetraploid under heat stress (45°C) versus control conditions (28°C) revealed conserved heat-responsive transcriptional plasticity, suggesting that enhanced thermotolerance in C. × hytivus is presented as consequences arising dominantly after interspecific hybridization. This study provides mechanistic insights into allopolyploid adaptation through experimental reconstruction of allopolyploid genomes, demonstrating that hybridization initiates key transcriptional and physiological advantages under stress, subsequent WGD stabilizes these adaptations and contributes to the full phenotypic realization. This work decouples the roles of interspecific hybridization and WGD and proposes a synthetic biology approach for developing climate-resilient crops.

Hybridization, Genetic↗

PubMed2025

SETD1B-mediated broad H3K4me3 controls proper temporal patterns of gene expression critical for spermatid development.

Epigenetic programming governs cell fate determination during development through intricately controlling sequential gene activation and repression. Although H3K4me3 is widely recognized as a hallmark of gene activation, its role in modulating transcription output and timing within a continuously developing system remains poorly understood. In this study, we provide a detailed characterization of the epigenomic landscapes in developing male germ cells. We identified thousands of spermatid-specific broad H3K4me3 domains regulated by the SETD1B-RFX2 axis, representing a previously underappreciated form of H3K4me3. These domains, overlapping with H3K27ac-marked enhancers and promoters, play critical roles in orchestrating robust transcription and accurate temporal control of gene expression. Mechanistically, these broad H3K4me3 compete effectively with regular H3K4me3 for transcriptional machinery, thereby ensuring robust levels and precise timing of master gene expression in mouse spermiogenesis. Disruption of this mechanism compromises the accuracy of transcription dosage and timing, ultimately impairing spermiogenesis. Additionally, we unveil remarkable changes in the distribution of heterochromatin marks, including H3K27me3 and H3K9me2, during the mitosis-to-meiosis transition and completion of meiotic recombination, which closely correlates with gene silencing. This work underscores the highly orchestrated epigenetic regulation in spermatogenesis, highlighting the previously unrecognized role of Setd1b in the formation of broad H3K4me3 domains and transcriptional control, and provides an invaluable resource for future studies toward the elucidation of spermatogenesis.

Male↗

PubMed2024

A mammalian tripartite enhancer cluster controls hypothalamic Pomc expression, food intake, and body weight.

Food intake and energy balance are tightly regulated by a group of hypothalamic arcuate neurons expressing the proopiomelanocortin (POMC) gene. In mammals, arcuate-specific POMC expression is driven by two cis-acting transcriptional enhancers known as nPE1 and nPE2. Because mutant mice lacking these two enhancers still showed hypothalamic Pomc mRNA, we searched for additional elements contributing to arcuate Pomc expression. By combining molecular evolution with reporter gene expression in transgenic zebrafish and mice, here, we identified a mammalian arcuate-specific Pomc enhancer that we named nPE3, carrying several binding sites also present in nPE1 and nPE2 for transcription factors known to activate neuronal Pomc expression, such as ISL1, NKX2.1, and ERα. We found that nPE3 originated in the lineage leading to placental mammals and remained under purifying selection in all mammalian orders, although it was lost in Simiiformes (monkeys, apes, and humans) following a unique segmental deletion event. Interestingly, ablation of nPE3 from the mouse genome led to a drastic reduction (>70%) in hypothalamic Pomc mRNA during development and only moderate (<33%) in adult mice. Comparison between double (nPE1 and nPE2) and triple (nPE1, nPE2, and nPE3) enhancer mutants revealed the relative contribution of nPE3 to hypothalamic Pomc expression and its importance in the control of food intake and adiposity in male and female mice. Altogether, these results demonstrate that nPE3 integrates a tripartite cluster of partially redundant enhancers that originated upon a triple convergent evolutionary process in mammals and that is critical for hypothalamic Pomc expression and body weight homeostasis.

Animals↗

PubMed2025

Hepatocyte-Enriched miRNA-193b-3p Promotes Hepatitis B Virus Replication by Dual Activation of Viral Core Promoter Activity and Autophagy Induction by Targeting IGF-1R.

Hepatitis B virus (HBV) infection is a principal cause of severe liver disease in humans and is associated with increased levels of specific serum or intracellular microRNAs (miRNAs). Among these, miR-193b-3p is a liver-enriched miRNA; however, its role in HBV replication remains unknown. This study aimed to investigate the influence of chronic HBV infection on miR-193b-3p levels in the peripheral blood and liver tissues of patients with chronic hepatitis B (CHB), evaluate the effect of miR-193b-3p on HBV replication both in vitro and in vivo, and elucidate the potential underlying mechanisms. We showed that hepatic miR-193b-3p levels in patients with CHB were significantly elevated compared with those in healthy controls. Ectopic expression of miR-193b-3p significantly enhanced HBV replication and transcription in different hepatoma cell lines. Furthermore, we identified IGF-1R as a direct target through which miR-193b-3p regulates HBV replication. Mechanistically, miR-193b-3p increased HBV core promoter activity via the IGF-1R/FXRα axis, thereby enhancing HBV transcription. Additionally, miR-193b-3p increased IGF-1R/Akt/MDM2/p53 signaling-mediated autophagy induction, which in turn facilitated increased HBV post-transcriptional activity. Collectively, hepatocyte-enriched miR-193b-3p exerts a proviral effect on HBV replication through dual synergistic mechanisms, offering novel insights into its role in HBV replication and potential therapeutic implications in CHB infection.

Humans↗

PubMed2026

Ecr positively regulates activity of the PhoQ/PhoP signalling system in Klebsiella pneumoniae.

BACKGROUND: The rising prevalence of polymyxin resistance in multidrug-resistant Klebsiella pneumoniae presents a critical situation with limited therapeutic options. METHODS: Methods Genomic sequencing of 15 clinical polymyxin-resistant K. pneumoniae strains with multidrug resistance revealed that MgrB inactivation, predominantly disrupted by insertion sequences (ISs) in the IS1, IS4, and IS5 families, was the leading cause of polymyxin resistance. Comparative transcriptomics of wild-type, ΔmgrB, and ΔmgrBΔphoP were performed to elucidate the MgrB-PhoPQ regulatory network. RESULTS: This study conducted a system-wide analysis of the regulatory network and identified a species-specific PhoPQ regulon in K. pneumoniae.Beyond the classical MgrB-PhoPQ-ArnBCADTEF pathway, we identified a previously unannotated PhoPQ-regulated gene, 144 bp LN739_RS09850, encoding an Ecr homologue from Enterobacter cloacae. This protein has been reported to confer colistin heteroresistance, with the underlying mechanism not yet functionally validated. This study revealed that overexpression of Ecr homologues decreased colistin susceptibility in both K. pneumoniae and E. cloacae, but this phenotype was abolished upon phoP deletion, confirming PhoP's essential role. Consistent with this dependency, comparative transcriptomics of Ecr-overexpressing K. pneumoniae vs. control revealed significant upregulation of mgrB, phoPQ, arnBCADTE, and pmrD. Two-hybrid bacterial assays further demonstrated direct Ecr-PhoQ interaction. Electrophoretic mobility shift assay confirmed that PhoP directly binds to the ecr promoter in vitro, and a β-galactosidase reporter assay demonstrated that PhoP enhanced ecr promoter activity, indicating that PhoP regulates ecr expression by directly controlling its transcription. CONCLUSION: Collectively, these findings suggest that PhoP may directly activate the transcription of Ecr, with Ecr feedback activating the PhoPQ system via interaction with PhoQ, leading to induction of the arn operon and consequent polymyxin resistance.

Klebsiella pneumoniae↗

PubMed1977

Stimulation of RNA polymerase I and II activities by 17 beta -estradiol receptor on chick liver chromatin.

The endogenous transcriptional capacity (RNA polymerase I and II activity) of liver chromatin from chicks treated with 17 beta-estradiol for 24 h (E 24) was double that of the controls. E 24 chromatin contained estradiol receptor activity while control chromatin did not. Its presence suggested an implication in the enhanced activities of RNA polymerases of E 24 chromatin. When semi-purified estradiol receptor was added to control chromatin, the endogenous transcriptional capacity of this chromatin was greatly increased. Studies with alpha-amanitin showed that both RNA polymerase I and II were stimulated by the estradiol receptor. This stimulation was observed as long as homology of the system was maintained. Solubilized homologous RNA polymerases were stimulated much less by the hormone complex in the presence of heterologous DNA than with homologous chromatin. Prokaryotic RNA polymerase could not be stimulated by chick liver estradiol receptor in the presence of heterologous DNA.

Animals↗

PubMed2026

Inducible flocculation in Komagataella phaffii enables enhanced biomass separation for biopharmaceutical production.

Biomass separation represents a critical bottleneck in Komagataella phaffii-based biopharmaceutical processes, as typically high cell densities of 40 - 50 % create significant operational, technical and economic challenges for harvest operations. Yeast cell aggregation (flocculation) provides a solution to accelerate cell sedimentation by increasing particle size, thus allowing to improve biomass-supernatant separation efficiency during both natural gravity settling and (continuous) centrifugation operations. This study demonstrates successful engineering of K. phaffii strains with an inducible flocculation phenotype using CRISPR/Cas9-based genome editing to integrate the Saccharomyces cerevisiae FLO1 (ScFLO1) gene under control of various regulatory elements, including methanol-inducible and derepressible promoters. Flocculation strength could be enhanced by implementing transcriptional positive feedback circuits based on the methanol-inducible AOX1 promoter. To address methanol-free production requirements, we developed alternative systems to retrofit PAOX1-based ScFLO1 expression and exploited the derepressible PDF promoter, offering broader compatibility with biopharmaceutical manufacturing facilities. Flocculating cells cultivated in a bioreactor demonstrated significantly improved sedimentation behavior, with considerably lower supernatant turbidity after short low-speed centrifugation or gravity sedimentation compared to non-flocculating controls. Crucially, cell flocculation had no negative impact on product amount and quality when expressing a multivalent NANOBODY® VHH molecule with pharmaceutical relevance. Thus, this work establishes the first genetically engineered flocculation system in K. phaffii compatible with recombinant protein production, providing the basis for an innovative approach to streamline harvest operations in biopharmaceutical processes.

Flocculation↗

PubMed2025

Modulating the PPARγ pathway upregulates NECTIN4 and enhances chimeric antigen receptor (CAR) T cell therapy in bladder cancer.

With the approval of the antibody-drug conjugate enfortumab vedotin (EV), NECTIN4 has emerged as a bona fide therapeutic target in urothelial carcinoma (UC). Here, we report the development of a NECTIN4-directed chimeric antigen receptor (CAR) T cell, which exhibits reactivity across cells expressing a range of endogenous NECTIN4, with enhanced activity in high expressors. We demonstrate that the PPARγ pathway, critical for luminal differentiation, transcriptionally controls NECTIN4, and that the PPARγ agonist rosiglitazone primes and augments NECTIN4 expression, thereby increasing sensitivity to NECTIN4-CAR T cell-mediated killing. NECTIN4-CAR T cells have potent anti-tumor activity even against EV resistant cells, which largely retain NECTIN4 expression, including in a post-EV biopsy cohort. Our results elucidate a therapeutically actionable mechanism that UC cells use to control NECTIN4 expression and suggest therapeutic approaches that leverage PPARγ agonists for rational combinations with NECTIN4-targeting agents in UC, as well as future potential treatment options for EV-refractory patients.

Humans↗