Induction of ferroptosis is a potential strategy for treating cancer and improving the efficacy of immunotherapy. Ferroptosis is driven by excessive peroxidation of polyunsaturated fatty acid-containing phospholipids, suggesting that microenvironmental lipid metabolites may regulate ferroptotic sensitivity. By integrating single-cell and bulk transcriptomics from immunotherapy cohorts, we identified the prostaglandin (PG) pathway as closely associated with tumor ferroptosis and therapeutic efficacy. Further screening revealed PGF2α as a potent endogenous ferroptosis sensitizer. Mechanistically, microenvironmental PGF2α bound to ferroptosis suppressor protein 1 (FSP1) at alanine 295 (A295) and inhibited its enzymatic activity, leading to lipid peroxidation accumulation upon ferroptotic stimuli. Preclinically, PGF2α supplementation or FSP1 ablation enhanced tumoral ferroptosis, potentiated CD8+ T cell-mediated immunity, and suppressed tumor progression in immunocompetent mice. Moreover, PGF2α improved immunotherapy efficacy across multiple mouse models, including subcutaneous allografts, Braf/Pten-driven spontaneous melanoma, and humanized mice. Clinically, a high PGF2α activity-related transcriptomic signature correlated with elevated ferroptosis and improved patient survival. Collectively, these findings establish PGF2α as a pro-ferroptotic metabolite and propose that targeting the PGF2α/FSP1 axis may offer an effective cancer immunotherapeutic strategy.

Antibody-drug conjugates (ADCs) show clinical efficacy against several solid malignancies, inducing cytotoxic effects in malignant cells accompanied by changes in the composition of the tumor stroma. The therapeutic effects of some ADCs have been shown to be potentiated by combination with anti-PD(L)1 checkpoint inhibitors, highlighting the potential of combining ADCs and immunotherapy. Here, we investigated the potential of combining ADCs with agonist monoclonal antibodies targeting CD137 (4-1BB), a costimulatory receptor expressed on antigen-primed T lymphocytes and activated NK cells. The combination of MMAE- and deruxtecan-conjugated ADCs towards NECTIN-4 or HER2 with anti-CD137 agonists elicited synergistic effects in mouse tumor models. Treatment with the ADCs increased CD137 expression on intratumor CD8+ and CD4+ T lymphocytes. Moreover, the synergistic antitumor therapeutic efficacy was dependent on cDC1 dendritic cells and CD8+ T cells. ADCs induced features of immunogenic cell death in malignant cells, resulting in cross-presentation and maturation of cDC1 dendritic cells that cross-prime CD8+ T cells to become CD137-positive. Importantly, the anti-CD137 plus ADC synergistic combination attained efficacy against concomitant tumor lesions that failed to express the ADC target. Together, these findings demonstrate that anti-CD137 agonists hold promise for combination strategies with ADCs.

Esophageal squamous cell carcinoma (ESCC) exhibits heterogeneous responses to immune checkpoint blockade, highlighting the need to define tumor-intrinsic mechanisms driving resistance to PD-L1 inhibition. Here, we identified GTP-binding protein overexpressed in skeletal muscle (GEM) as a tumor cell-intrinsic regulator associated with poor response to anti-PD-L1 therapy in ESCC. GEM expression in tumor cells drove tumor-associated macrophage (TAM)-mediated immunosuppression, leading to reduced CD8[+] T cell infiltration and impaired cytotoxic function. Mechanistically, GEM induced SERPINE1 expression that was secreted and acted on macrophage LRP1 to drive alternative activation and suppress antitumor immunity. GEM enhanced SERPINE1 expression through a MKK3-RACK1-p38-MEF2A cascade. GEM signaling was reinforced by a metabolically coupled feedback mechanism. Macrophages exposed to GEM-high tumor cells promoted tumor cell glycolysis and lactate production, which drove AARS1-dependent lactylation of GEM and strengthened its interaction with RACK1 to amplify downstream signaling. Disruption of the SERPINE1-LRP1 axis restored CD8[+] T cell activity and enhanced response to PD-L1 blockade in vivo. Pharmacological perturbation of GEM-dependent signaling remodeled the tumor immune microenvironment and improved responses to PD-L1 blockade. Together, these findings define a GEM-SERPINE1-LRP1 signaling axis that drives macrophage-mediated immune suppression and suggest that targeting this pathway may enhance the efficacy of PD-L1 blockade in ESCC.

Chronic stress is increasingly recognized as a driver of cancer metabolism, highlighting the need to elucidate the mechanism linking chronic stress to metabolic reprogramming. Here, we performed untargeted metabolomics on serum samples from esophageal squamous cell carcinoma (ESCC) patients and integrated the results with clinical and stress-related assessments, revealing pentose phosphate pathway (PPP) enrichment accompanied by elevated epinephrine levels in patients with stress-associated features. β2-adrenergic receptor (ADRB2) activated by chronic stress stabilized MYCBP by competitively displacing VHL, thereby enhancing MYC transcriptional activity and upregulating key PPP enzymes, including G6PD and TKT. Unexpectedly, stress-associated stimulation not only activated ADRB2 signaling but also increased ADRB2 protein abundance. Mechanistically, stress exposure enhanced RNMT-dependent N7-methylguanosine (m7G) cap modification of ADRB2 mRNA, thereby increasing its translational efficiency. Together, these findings define an RNMT-ADRB2-MYCBP-PPP axis that integrates epitranscriptomic regulation with adrenergic signaling to promote metabolic reprogramming and malignant progression in ESCC. The stress-associated RNA modification-metabolism circuitry comprises potential therapeutic targets to overcome stress-associated ESCC progression.

High dimension spatial technologies are broadly being used to map the cellular and molecular organization of human cancers including tumor antigens and drug targets. This has revealed extensive spatial heterogeneity of target proteins, but does not quantify drug distribution itself. Exposing additional complexities for drug targeting and delivery, Lu and colleagues recently applied highly multiplexed spatial proteomics to tumors from patients treated with fluorescently labeled antibody therapeutics. Quantifying drug penetration and distribution on cells of specific phenotypes, this approach identified compounding challenges that would limit therapeutic success as both target and therapeutic distribution are highly variable between patients and within individual tumors. Macrophage uptake, stromal barriers, vascularization differences, and epithelial permeability were all associated with altered antibody distribution. As expected, highly desmoplastic pancreatic cancers had poor drug delivery, and similar fibroblast and extracellular matrix (ECM) rich microenvironments were also identified in head and neck squamous cell carcinomas (HNSCCs) as broadly prevalent barriers to drug delivery. Providing a measurement strategy for single cell drug distribution, this work points towards opportunities to predict these barriers in precision medicine and design drugs that overcome them as a clinically impactful frontier.

Histone lactylation is an epigenetic modification triggered by lactate produced during glycolysis. In cancer, histone lactylation can coordinate metabolic and epigenetic states to promote tumor development and progression. Here, we identified circ2891 as a driver of metabolic reprogramming and histone lactylation in colorectal cancer (CRC). In patients, circ2891 was aberrantly upregulated in tumors and correlated with poor outcomes. Elevated circ2891 potentiated aerobic glycolysis and lactate production in CRC cells, supporting enhanced cell proliferation and tumor growth. Mechanistically, circ2891 specifically interacted with the m6A reader FXR1 and facilitated its phase separation, which stabilized PGK1 and ENO1 mRNAs by enhancing the recognition of their m6A modification sites. Consequently, elevated PGK1 and ENO1 led to increased aerobic glycolysis and drove histone lactylation, thereby transcriptionally activating the RNA helicase DDX21 to promote CRC progression. Simultaneously targeting circ2891 and histone lactylation suppressed patient-derived organoid and xenograft tumor growth. Together, these findings show that circ2891 promotes CRC by stabilizing PGK1/ENO1 via phase separation of FXR1, boosting glycolysis and histone lactylation to activate expression of DDX21. Targeting circ2891 offers a promising therapeutic strategy enhancing clinical outcomes.

Targeted kinase inhibitors induce marked tumor regressions but are limited by the emergence of drug-tolerant residual disease and resistance. Although tumor-intrinsic and adaptive immune mechanisms have been extensively studied, a better understanding of the contribution of innate immunity to targeted therapy durability is needed to devise optimal treatment strategies. Here, we showed that natural killer (NK) cells constrain targeted therapy resistance and identified a regression-associated macrophage program that promotes NK cell recruitment. In immunocompetent melanoma models that recapitulate patient treatment trajectories, tumor regression was characterized by robust NK infiltration, which transitioned to an NK-excluded residual state preceding resistance. A regression-associated macrophage subset (F4/80hiCCL5[+]MHCII[+]CD63[+]) contributed to NK cell recruitment via CCR2/5 signaling. Genetic depletion of macrophages using LysM-cre;iDTR mice impaired NK cell infiltration. Pharmacologic inhibition of PTPN22, a negative regulator of immune activation, reprogrammed the residual tumor microenvironment, restored NK cell recruitment, and delayed resistance onset. Analysis of patient tumor datasets from melanoma and lung cancer revealed concordant NK dynamics during therapy, linking this innate immune program to clinical outcomes. These findings identify innate immune remodeling as a key contributor to targeted therapy durability and identify NK cell recruitment, promoted in part by regression-associated macrophages, as a therapeutically actionable node for extending durability of response across oncogene-driven cancers.

Primary and metastatic lesions share similar biological properties despite existing in different microenvironments. A better understanding of the mechanisms underlying the maintenance of phenotypic homogeneity in heterogeneous microenvironments could help identify strategies for suppressing metastasis. Here, we found that mechanical memory enabled tumor cells to resist biomechanical stress and sustain malignant traits. Matrix stiffness activated the RhoA-ROCK1 signaling pathway, leading to actin cytoskeletal remodeling and suppression of mitochondrial fission. The dysregulation of mitochondrial dynamics enhanced fatty acid β-oxidation, leading to the accumulation of acetyl-CoA and subsequent histone hyperacetylation. The stiffness-mediated epigenetic reprogramming was mitotically heritable, allowing progeny cells to retain a proliferative advantage after detachment from stiff environments. Inhibition of RhoA-ROCK1 disrupted mechanical memory, thereby reducing metastasis. These findings suggest that targeting mechanical memory may be a strategy for preventing metastasis.

Cancer-associated fibroblasts (CAFs) drive key aspects of tumor malignancy. However, the profound heterogeneity in cytokine secretion and cell-cell communication orchestrated by CAFs renders therapeutic strategies targeting these molecules largely ineffective against cancer. In contrast, exploring the metabolic reprogramming mechanisms of CAFs could be conducive to therapeutic intervention in colorectal cancer (CRC). Here, we conducted metabolomic and single-cell RNA-sequencing analyses that uncovered increased glycolysis, intracellular lactate, and histone lactylation in CAFs compared to normal fibroblasts (NFs). Elevated histone 3 lysine 18 lactylation (H3K18la) in CAFs promoted the transition of CAFs to myofibroblastic CAFs (myCAFs), increased collagen deposition, and enhanced organoid growth. Furthermore, fibroblast-specific conditional lactic dehydrogenase A (Ldha) knockout in a mouse model mitigated CRC tumorigenesis and progression in vivo. Cancer cell-derived TGF-β accelerated metabolic reprogramming and histone lactylation in CAFs. NPM1, a histone chaperone functioning as a cofactor of P300, enhanced the H3K18la level of the ROCK1 promoter and activated ROCK1 transcription. The RhoA/ROCK1/MLC2/MRTF-A pathway and extracellular matrix (ECM) remodeling were indispensable for the tumor-promoting effect of CAF histone lactylation in CRC. Furthermore, CAF histone lactylation limited CD8+ T cell infiltration and aggravated CD8+ T cell exhaustion via ECM remodeling. Stiripentol, a clinical drug that targets lactylation, potentiated the efficacy of immunotherapy in CRC models. Together, this study demonstrates that elevated lactate levels in CAFs reprogram the epigenetic landscape to promote tumor progression and immunosuppression, highlighting the potential of targeting CAF histone lactylation as a therapeutic strategy for CRC.

Metastasis remains a leading cause of mortality in esophageal squamous cell carcinoma (ESCC) patients, underscoring the urgent need to elucidate the molecular mechanisms driving disease progression. In this study, we delineated that ZBED2, a zinc finger protein, correlates with inferior survival outcomes and metastasis in ESCC patients. ZBED2 formed phase-separated nuclear condensates that functionally promote tumor metastasis, and integrin-linked kinase (ILK) was pinpointed as a critical downstream effector in mediating the pro-metastatic function of ZBED2. Mechanistically, ZBED2 enhanced the transcription of HSP90AA1, promoted the physical interaction between HSP90AA1 and ILK, and consequently stabilized ILK by suppressing its ubiquitination. Subsequently, ILK promoted PD-L1 transcription and CD8+ T cell exhaustion, thereby creating an immunosuppressive microenvironment that facilitates cancer metastasis. Collectively, these findings establish the pivotal role of ZBED2 in driving ESCC metastasis and immune evasion, thereby validating it as a promising therapeutic target for this aggressive malignancy.

Immune evasion remains a major barrier to effective immunotherapy in cancer. Coordinated activation of different immune cell types shapes immune responses within the tumor microenvironment (TME). In this study, we uncovered a spatiotemporal synergy between cytotoxic T lymphocytes (CTL) and natural killer (NK) cells in tumors with low T-cell infiltration. An integrative multiomics approach combining spatial transcriptomics, single-cell RNA sequencing, and high-dimensional flow cytometry demonstrated that vaccination, capable of activating both innate and adaptive immunity, rapidly reshapes the TME. Early infiltration of IRF8+KLRG1+ NK cells established a chemokine-rich niche together with myeloid cells that promoted the recruitment of antigen-primed CXCR3+ CTLs. The accompanying inflammatory monocyte recruitment and dendritic cell activation led to the coordinated structural and chemotactic remodeling of the TME. Functionally, NK-CTL clustering at the tumor margin was associated with enhanced cytotoxic activity and sustained immune engagement. These spatially organized immune interactions involved CCR5-CCL5 and CXCR3-CXCL9 signaling pathways that coordinate communication between innate and adaptive immune compartments. Together, these findings reveal a previously unrecognized NK-CTL cooperative program that promotes the transition of poorly infiltrated tumors toward an inflamed, immune-responsive state. More broadly, this study illustrates how spatially resolved multiomics approaches can uncover immune interactions and provides a conceptual framework for designing next-generation immunotherapies that mobilize coordinated innate and adaptive immunity.

Casein kinase 1⍺ (CK1⍺) is a multifunctional serine/threonine kinase, serving as an upstream regulator of the p53 pathway and a potential therapeutic target for acute myeloid leukemia (AML). Small-molecule degraders show preclinical promise in AML by selectively degrading target proteins. Here, we developed and evaluated PinA1, a molecular glue degrader targeting CK1⍺, in preclinical AML models. PinA1 preferentially degraded CK1⍺ at nanomolar concentrations, resulting in enhanced p53 expression, cell-cycle arrest, and apoptosis in TP53 wild-type AML cell lines but not in TP53 mutated cells. PinA1 also induced CK1⍺ degradation and p53-dependent apoptosis in primary AML cells with wild-type TP53. Notably, PinA1 had minimal effects on the viability of human peripheral blood and bone marrow mononuclear cells, including CD34+ cells, with limited activation of the p53 pathway. In both cell lines and primary AML cells, PinA1 enhanced apoptosis when combined with targeted agents, including FLT3, BCL-2, or MDM2 inhibitors. PinA1 demonstrated robust anti-leukemic efficacy as a monotherapy and exhibited synergistic effects when combined with targeted agents in xenografts derived from human AML cell lines and primary AML cells. In conclusion, PinA1 degrades CK1⍺, activates the p53 pathway, and induces cell-cycle arrest and apoptosis in TP53 wild-type AML cells. The robust anti-leukemic efficacy of PinA1, both as monotherapy and further in combination with targeted agents, along with its minimal toxicity to normal hematopoietic cells, underscores its potential for future clinical applications.

Intratumor transcriptional heterogeneity (ITTH), defined as the coexistence of diverse cell states within a single tumor, complicates cancer treatment and contributes to variable therapeutic responses. Although single-cell RNA sequencing (scRNA-seq) can resolve this complexity, its cost and technical demands limit large-scale use. Bulk RNA sequencing (bulk RNA-seq) provides a scalable alternative but requires computational methods to deconvolve bulk transcriptomes into distinct cell states. Existing supervised approaches rely on accurate reference data, which are lacking for many cancer types, while unsupervised methods are not tailored to capture heterogeneity within the malignant compartment. To address these limitations, we developed CDState, an unsupervised deconvolution method based on nonnegative matrix factorization with a sum-to-one constraint and a cosine-similarity-based optimization, which infers malignant cell states using bulk RNA-seq data. CDState demonstrated robustness using pseudobulk scRNA-seq datasets from five cancer types, outperforming existing unsupervised methods in estimating both state-specific gene expression and cell proportions. Applied to 33 cancer types from The Cancer Genome Atlas, CDState revealed recurrent gene programs, including epithelial-mesenchymal transition, MYC targets, and oxidative phosphorylation, as major contributors to malignant ITTH. The malignant state proportions were linked to clinical features, including patient survival and therapeutic response. Finally, mutations and copy number alterations in genes such as TP53, KRAS, PIK3CA, SOX2, and SATB1 were identified as potential genetic drivers of malignant cell ITTH across cancer types. This study demonstrates the utility of CDState for characterization of malignant cell states from bulk RNA-seq data, establishing a framework for investigating malignant cell ITTH in large-scale cancer atlases.

Small cell lung cancer (SCLC) remains one of the most lethal malignancies, with a five-year survival of <10% despite advances in treatment. Seizure-related gene 6 (SEZ6), a type I transmembrane protein preferentially expressed in neuroendocrine tumors, has recently emerged as a promising therapeutic target, supported by clinical activity of SEZ6-targeted antibody-drug conjugates in relapsed SCLC. Here, we developed and evaluated a humanized anti-SEZ6 monoclonal antibody for SEZ6-targeted positron emission tomography (PET) imaging and α-particle radioimmunotherapy. Immunohistochemistry of patient tissues confirmed strong, stage-independent SEZ6 expression. Cell-based studies demonstrated high SEZ6 receptor densities, supporting high-affinity antibody binding and rapid receptor-mediated internalization. [89Zr]Zr-DFO-SEZ6-Ab administered at low antibody mass exhibited rapid hepatosplenic clearance with minimal tumor uptake despite high intratumoral levels. This pattern was consistent with active ectodomain shedding and local accumulation of soluble antigen within the tumor microenvironment, creating an extracellular antigen sink and limiting effective target engagement. Increasing the antibody mass overcame this barrier, dramatically increasing tumor uptake in xenografts. Treatment with [225Ac]Ac-Macropa-SEZ6-Ab induced complete tumor regression, while co-administration of the ATR inhibitor berzosertib enhanced efficacy at lower radioactivity doses without added toxicity. Together, these findings identify tumor-localized SEZ6 shedding as a biological barrier to SEZ6-targeted therapy, define dosing strategies to overcome it, and demonstrate potent antitumor activity of SEZ6-directed α-particle radioimmunotherapy in SCLC.