Type 2 immunity at mucosal surfaces is essential for helminth clearance, tissue repair, and barrier maintenance, processes governed by a robust feed-forward circuit between chemosensory tuft cells and group 2 innate lymphoid cells (ILC2s). However, the regulatory mechanisms that calibrate the magnitude and duration of this epithelial immune circuit remain incompletely defined. Here, we identify interleukin-18 (IL-18), a cytokine traditionally associated with type 1 immunity, as a key regulator of tuft cell-mediated type 2 responses during Nippostrongylus brasiliensis (N. brasiliensis) infection. IL-18-deficient (Il18-/-) mice exhibited reduced intestinal worm burden at day 7 post-infection, which correlated with an expansion of tuft cell lineage, increased expression of tuft cell and type 2 response associated genes, and enhanced ILC2 responses. Consistent with these findings, loss of IL-18 receptor (Il18r1-/-) further enhanced succinate-induced tuft cell hyperplasia. Within the intestinal epithelium, tuft cells were found to preferentially express IL-18Rα, suggesting potentially direct responsiveness to IL-18. Using small intestinal organoid models, we demonstrate that IL-18 directly attenuates IL-13-induced tuft cell differentiation, proliferation, and signature gene expression in an IL-18Rα-dependent manner. Mechanistically, pharmacological inhibition of p38 MAPK partially reversed the inhibitory effects of IL-18, identifying p38 signalling as a critical downstream mediator of this regulatory process. Together, these findings reveal that IL-18 functions as a negative feedback signal to constrain tuft cell-associated type 2 epithelial responses during helminth infection, thereby maintaining the balance between protective anti-helminth immunity and mucosal homeostasis.

Objective:To assess the self-management status of patients with SLE and identify the factors that influence it. Background:Systemic lupus erythematosus (SLE) is a chronic, relapsing autoimmune disease that requires lifelong management. Effective self-management is crucial for reducing disease activity, economic burden, and improving quality of life. However, research on SLE-specific self-management tools and influencing factors remains limited. Design:A cross-sectional study was conducted from March to September 2024, enrolling 370 SLE patients from three tertiary hospitals in Henan Province. Data were collected using validated scales, including self-management, self-efficacy, illness perception, coping styles, family support, and social support. Univariate analyses, correlation analyses, and multiple linear regression were performed. Methods:A cross-sectional study was conducted from March to September 2024, enrolling 370 SLE patients from three tertiary hospitals in Henan Province. Data were collected using validated scales, including self-management, self-efficacy, illness perception, coping styles, family support, and social support. Univariate analyses, correlation analyses, and multiple linear regression were performed. Multicollinearity was assessed using variance inflation factors (VIFs) and tolerance values. Results:Among the 370 SLE patients, the mean age was 38.16 ± 12.81 years; the mean duration of illness was 4.65 ± 3.18 years; Regarding disease activity, 134 SLE patients (36.2%) had no or minimal activity, 116 (31.4%) had mild activity, and 120 (32.4%) had moderate-to-high activity. The mean self-management score was 59.06 ± 16.75, with sub-scores for medication being 9.91 ± 3.45, disease surveillance (10.01 ± 3.56), daily life (29.40 ± 9.35), and reproductive health (9.74 ± 3.84). The key influencing factors included disease activity, education level, marital status, self-efficacy, illness perception, coping styles, family support, and social support, explaining 52.4% of variance. Conclusion:Tailored interventions should address SLE patients' unique needs by enhancing self-efficacy, optimizing illness perception, promoting positive coping, and strengthening social and family support. This approach can improve self-management, reduce disease burden, and enhance the quality of life.

Purpose:Disturbed sleep quantity and quality are associated with diabetes and have been recognized as significant risk factors for its development. However, it remains unclear whether sleep disturbances arise during the progression of diabetes and what underlying neurobiological mechanisms may be involved. We aimed to characterize the dynamic development of sleep-wake phenotype in diabetic db/db mice and to investigate the associated neural mechanisms. Methods:Polysomnographic recordings were conducted to characterize age-dependent changes in sleep-wake characteristics in diabetic db/db mice aged 2 to 6 months. Transcriptomic analyses of brain tissues were performed to investigate the underlying neural mechanisms associated with sleep alterations. Results:Although hyperglycemia was present in db/db mice at 2 months of age, significant fragmentation of non-rapid eye movement (NREM) sleep began at 3 months of age. The total amount of NREM sleep remained unchanged during light phase but was disrupted by an increased frequency of microarousal episodes lasting ≤ 8 s. The darkness-induced wakefulness response progressively declined from 3 months of age and was markedly attenuated by 6 months. Transcriptomic analyses revealed pronounced dysregulation of genes associated with sleep-wake regulation in subcortical regions at 3 months of age. Conclusion:These results indicate that diabetes contributes to fragmented sleep and impaired diurnal rhythmicity by influencing sleep-wake regulatory systems in subcortical brain regions.

Preeclampsia (PE) is a devastating hypertensive disorder affecting pregnant women worldwide. Disrupted metabolic reprogramming is recognized as a key feature of placental dysfunction in PE, yet the abnormal metabolic adaption and underlying mechanisms remain largely unknown. In this study, we perform targeted metabolomic profiling and identify placental serine deficiency as a hallmark metabolic alteration in PE, which favors PE occurrence. Serine-deficient chow exacerbates PE-like symptoms, such as hypertension and proteinuria, in mice. Mechanistically, serine deficiency attenuates SAM-dependent methylation, decreasing SP1 levels and impairing SP1-BNIP3-mediated mitophagy, thereby exacerbating oxidative stress to cause placental dysfunction. Notably, targeting serine to the placenta using mPEG5k-poly(D/L-serine) effectively relieves PE-like symptoms in the mouse model. Our findings elucidate an unknown serine-deficiency-mediated metabolic reprogramming in PE and suggest manipulating serine supplementation as a promising translational strategy for PE treatment.

Cancer-associated fibroblasts comprise diverse functionally distinct cellular subsets, with certain subpopulations exerting pivotal influence in shaping the pancreatic cancer immune microenvironment. Here we show that Lin28b+ cancer-associated fibroblasts contribute to establishing an immunologically cold tumor microenvironment in pancreatic ductal adenocarcinoma. Mechanistically, Lin28b directly binds to STING mRNA and promotes its degradation, thereby suppressing STING expression and downstream type I interferon signaling. Loss of Lin28b in cancer-associated fibroblasts activates the cGAS-STING-interferon signaling cascade, enhancing dendritic cell antigen presentation and CD8+ T cell cytotoxic function. Importantly, genetic inhibition of Lin28b in cancer-associated fibroblasts enhances sensitivity to anti-PD-L1 immune checkpoint blockade therapy. These findings reveal that targeting the Lin28b-STING axis represents a promising therapeutic strategy for overcoming the intrinsic resistance of pancreatic ductal adenocarcinoma to immunotherapy.

In a homozygous alanine-tRNA synthetase 2 R194C knock-in mouse model (mimicking the human R199C -pathogenic variant), this study demonstrates that a single mutation can cause ovarian insufficiency in vivo and identifies the lactylation-metabolism-mechanistic target of rapamycin axis as the molecular driver of follicle exhaustion. Abstract Premature ovarian insufficiency (POI) often arises from genetic causes, yet the pathogenic consequences of many variants remain undefined. The alanine-tRNA synthetase 2 (AARS2) R199C mutation has been repeatedly reported in patients, but its physiological effects are unknown. Here, we generated the first homozygous Aars2 R194C knock-in mouse to model this variant in vivo. Female knock-in mice showed irregular estrous cycles, reduced fecundity, altered endocrine profiles, and accelerated depletion of the primordial follicle pool, reproducing core features of POI. Mutant ovaries exhibited increased lysine lactylation of the metabolic enzymes pyruvate dehydrogenase alpha 1 and carnitine palmitoyltransferase 2, accompanied by reduced activity and impaired mitochondrial respiration in granulosa cells. These metabolic defects were associated with sustained activation of the mechanistic target of rapamycin complex 1 pathway and premature follicle activation. Loss of the mitochondrial de-lactylase sirtuin-3 mitigated these abnormalities, whereas pharmacological inhibition of pyruvate dehydrogenase and carnitine palmitoyltransferase in wild-type mice phenocopied key knock-in features. Together, these findings demonstrate that the Aars2 R194C/R199C mutation alone is sufficient to induce POI and establish a lactylation-driven metabolic mechanism underlying early follicle activation.

Isocitrate dehydrogenase 1 mutation IDH1R132H generates the oncometabolite (R)-2-hydroxyglutarate (2-HG). However, the mechanisms underlying the better clinical outcomes in patients with IDH1 mutation remain elusive. Here, we report that 2-HG sensitizes chemotherapy by destabilizing Fizzy And Cell Division Cycle 20 Related 1(CDH1) and reducing the number of quiescent cells (G0 phase cells). IDH1R132H-harboring glioblastoma tissues have decreased CDH1, which can be induced by IDH1R132H overexpression or 2-HG treatment in U87MG cells. Mechanistically, 2-HG inhibits Prolyl Hydroxylase Domain-containing protein 1 (PHD1), which hydroxylates proline 431 of CDH1 to maintain its stability. 2-HG induces hypo-hydroxylation of CDH1, rendering it susceptible to CDK1-mediated phosphorylation at serine 151/163, followed by β-TrCP-mediated ubiquitination and proteasomal degradation. Notably, in vitro and in vivo experiments demonstrate that 2-HG induces CDH1 downregulation and reduces quiescent cell populations not only in glioblastoma cells but also in lung cancer and colorectal cancer cells, and consistently sensitizes tumor cells to chemotherapy. Our findings shed light on why IDH1 mutations correlate with better prognosis and highlight the translational potential of 2-HG as a chemotherapy sensitizer. Cancer cells often use abnormal metabolism to grow and resist treatment, especially glioblastoma (a type of brain cancer). Our study focused on 2-hydroxyglutarate (2-HG), a molecule produced by cancer cells with an IDH1 gene mutation. We found 2-HG makes cancer cells more sensitive to chemotherapy. Here’s how: 2-HG blocks an enzyme called PHD1, which normally keeps the protein CDH1 stable. When PHD1 is blocked, CDH1 breaks down faster. This reduces the number of “dormant” cancer cells—quiet cells that are hard to kill with chemo. Our finding explains why glioblastoma patients with the IDH1 mutation often respond better to treatment. It also suggests 2-HG or PHD1 could be new targets to improve chemotherapy outcomes for brain, lung, and colorectal cancer patients.

Mapping the genomic locations of chromatin-associated proteins, such as transcription factors and histone modifications, is key to understanding the mechanisms of transcriptional regulation. ChIPmentation offers a simple and robust way of investigating the genomic binding sites of a protein using relatively low-input material. Here, we present a detailed protocol for the key steps that lead to a successful ChIPmentation experiment, as well as a quick analysis pipeline to examine the data.For complete details on the use and execution of this protocol, please refer to Schmidl et al.1 For example data produced by this protocol, please refer to Henriksson et al.2 and Zhang et al.3This protocol is an update to Xu et al.4

Aneuploidy is pervasive in cancers and contributes to chemoresistance; however, how aneuploidy-inducing stresses, such as infection and hypoxia, promote chemoresistance remains unclear. Here, we identify a prolyl hydroxylase domain protein 1 (PHD1)-E3 ubiquitin ligase TRIM21-cell division cycle protein 20 (CDC20) signaling axis that integrates infection- and PHD1-inhibitory signals to drive aneuploidy and chemoresistance. Analysis of clinical specimens revealed that HPV-positive cervical cancers exhibited reduced CDC20 expression and increased aneuploidy compared with HPV-negative tumors. Through proteomic screening, we found that CDC20 is targeted for degradation by TRIM21, which preferentially recognizes CDC20 when prolines 337 and 340 are non-hydroxylated. Hypoxia and α-ketoglutarate (α-KG) limitation impair the activity of the dioxygenase PHD1, thereby increasing the fraction of non-hydroxylated CDC20. In parallel, infection activates TRIM21. Thus, PHD1 inactivation and infection converge on CDC20 to reduce its abundance, leading to the accumulation of CDC20 substrates, including the separase inhibitor securin and the anti-apoptotic protein MCL1. Infection- and PHD1 inhibition-induced securin accumulation promotes aneuploidy, whereas MCL1 accumulation enhances chemoresistance. In cultured cancer cells and mouse xenograft models, stabilization of CDC20, either through TRIM21 inhibition or PHD1 activation, attenuates aneuploidy and restores chemosensitivity. Together, our study reveals a PHD1-TRIM21-CDC20 signaling axis that integrates hypoxic and infection-associated cues to regulate aneuploidy and chemoresistance, highlighting this pathway as a potential therapeutic target for overcoming chemoresistance.

The functional difference between the two catalytic subunits, α1 and α2, of AMP-activated protein kinase (AMPK) complexes remains elusive. Herein, we report that AMPKα2 specifically transduces amino acid insufficiency signals to protein synthesis. Low amino acid levels, high protein levels, and reduced phosphorylation of AMPKα threonine 172 (p-T172) are observed in blood samples in patients with Alzheimer's disease (AD) from a cohort of 1,000,000 Chinese individuals. Loss of α2, but not α1, recaptures these observations and induces AD-like cognitive dysfunction in mice. Mechanistically, low amino acid-activated general control nonderepressible 2 (GCN2) specifically phosphorylates α2 at T172 independent of AMP and fructose 1,6-bisphosphate to inhibit protein synthesis. α2-p-T172 loss renders protein over-synthesis and AD-pathologic protein aggregation in cells and in mouse brain. AMPK activators metformin and 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), as well as branched-chain amino acid (BCAA) or protein restriction, α2-p-T172-dependently prevent AD-like symptoms in mice. We identify AMPKα2 as a specific amino acid abundance detector for protein synthesis.

Fibrosis leads to structural damage and functional decline and is characterized by an accumulation of fibrous connective tissue and a reduction in parenchymal cells. Because of its extremely poor prognosis, organ fibrosis poses a significant economic burden. In order to prevent and treat fibrosis more effectively, potential mechanisms need to be investigated. A disintegrin and metalloprotease 17 (ADAM17) is a membrane-bound protein. It regulates intracellular signaling and membrane protein degradation. Fibrosis mediated by ADAM17 has been identified as an important contributor, although the specific relationship between its multiple regulatory functions and the pathogenesis is unclear. This article describes ADAM17 activation, function, and regulation, as well as the role of ADAM17 mediated fibrosis injury in kidney, liver, heart, lung, skin, endometrium, and retina. To develop new therapeutic approaches based on ADAM17 related signal pathways.

BACKGROUND:Protein palmitoylation is a reversible post-translational modification that increases protein hydrophobicity, which can affect protein localization, stability, and function. Although palmitoylation is frequently observed in various cancers, the specific mechanisms by which it influences clear cell renal cell carcinoma (ccRCC) are still not well understood. METHODS:This study used transcriptome expression profiles and clinical characteristics of clear cell renal cell carcinoma (ccRCC) obtained from The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO) databases. Kaplan-Meier (KM) survival analysis was performed to evaluate patient survival. Consensus clustering was applied to identify tumor palmitoylation patterns. A total of 101 different machine learning methods were used to develop predictive models. Functional enrichment analyses were conducted using Gene Ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and Gene Set Variation Analysis (GSVA). RESULTS:Of the 34 prognosis-related palmitoylation-related genes (PRGs), 29 were used to cluster patients in the TCGA-KIRC cohort, leading to the identification of four palmitoylation clusters. We developed a risk model and a nomogram based on palmitoylation scores to enhance risk classification. Functional analysis indicated that high-risk patients exhibited disrupted fatty acid metabolism. Correlation analysis identified ZDHHC18 as a potential hub gene associated with impaired fatty acid metabolism and cuproptosis. Finally, we validated the role of ZDHHC18 in ccRCC proliferation through in vitro experiments. CONCLUSION:Our research demonstrated that PRGs play a crucial role in the development of clear cell renal cell carcinoma (ccRCC). A nomogram based on palmitoylation scores may accurately predict the prognosis of ccRCC patients. Furthermore, the palmitoylation regulator ZDHHC18 affects cuproptosis in ccRCC, which in turn impacts patient survival.

In this study, fish oil (FO) and medium-chain triglyceride (MCT) was used to synthesize eicosapentaenoic acid (EPA) -rich medium- and long-chain triacylglycerol (MLCT). From the perspective of MLCT profile, the effects of MLCT (oil phase) on crystal structure, non-covalent interaction and in vitro digestion of monoglyceride oleogel were examined. The MML and LLM type triglycerides (MML structured triglycerides containing two medium-chain fatty acids and one long-chain fatty acid, LLM structured triglycerides containing one medium-chain fatty acid and two long-chain fatty acids) appeared in MLCT, but were not detected in FO and PM (physical mixture of MCT and FO). The TAG molecules in MLCT deeply participate in oleogel network construction through hydrogen bonding and van der Waals forces, which changed oleogel crystallization behavior. The G', hardness, melting point of oleogels were FO-G > PM-G > MLCT-G > MCT-G. Final free fatty acid released level of oleogels were MLCT-G (90.01%) > MCT-G (75.30%) > PM-G (69.97%) > FO-G (67.07%). The newly generated MML and LLM in MLCT weaken oleogel structure and facilitate the digestion of fatty acids.

Meiosis in mammalian oocytes is interrupted by a prolonged arrest at the germinal vesicle stage, during which oocytes have to repair DNA lesions to ensure genome integrity or otherwise undergo apoptosis. The FIRRM/FLIP-FIGNL1 complex dissociates RAD51 from the joint DNA molecules in both homologous recombination (HR) and DNA replication. However, as a type of non-meiotic, non-replicative cells, whether this RAD51-dismantling mechanism regulates genome integrity in oocytes remains elusive. Here, we show that FIRRM/FLIP is required for disassembly of RAD51-filaments and maintenance of genome integrity in oocytes. Deletion of FIRRM in oocytes leads to formation of massive nuclear RAD51 foci in oocytes of primordial follicles and activated follicles in mice. These RAD51 foci colocalize with the sites of DNA damage repair, as indicated by RPA2 and EdU, suggesting substantial DNA damage and extensive HR in oocytes. Especially in fully-grown FIRRM-deleted oocytes, RAD51 forms a net-like structure. As a consequence, FIRRM-deleted females are infertile due to aberrant homologous chromosome segregation at metaphase I and primordial follicle insufficiency at young adulthood. Hence, our study demonstrates the physiological importance of HR in maintaining genome integrity in oocytes.

Glutamine provides carbon and nitrogen to support the proliferation of cancer cells. However, the precise reason why cancer cells are particularly dependent on glutamine remains unclear. In this study, we report that glutamine modulates the tumor suppressor F-box and WD repeat domain-containing 7 (FBW7) to promote cancer cell proliferation and survival. Specifically, lysine 604 (K604) in the sixth of the 7 substrate-recruiting WD repeats of FBW7 undergoes glutaminylation (Gln-K604) by glutaminyl tRNA synthetase. Gln-K604 inhibits SCFFBW7-mediated degradation of c-Myc and Mcl-1, enhances glutamine utilization, and stimulates nucleotide and DNA biosynthesis through the activation of c-Myc. Additionally, Gln-K604 promotes resistance to apoptosis by activating Mcl-1. In contrast, SIRT1 deglutaminylates Gln-K604, thereby reversing its effects. Cancer cells lacking Gln-K604 exhibit overexpression of c-Myc and Mcl-1 and display resistance to chemotherapy-induced apoptosis. Silencing both c-MYC and MCL-1 in these cells sensitizes them to chemotherapy. These findings indicate that the glutamine-mediated signal via Gln-K604 is a key driver of cancer progression and suggest potential strategies for targeted cancer therapies based on varying Gln-K604 status.

Cell proliferation requires anabolic supports. How the cell cycle integrates anabolism remains poorly understood. Herein, it is identified that G1-phase regulator cell division cycle 20-like protein 1 (CDH1) coordinates anabolic events to ensure cell cycle initiation. CDH1 degrades Von Hippel-Lindau (VHL), concomitantly activates hypoxia-inducible factor 1α (HIF1α), which enhances angiogenesis and glucose metabolism, and activates mitochondrial lactyltransferase alanyl tRNA synthetase (AARS2), which lactylates and inactivates pyruvate dehydrogenase E1 subunit alpha 1 (PDHA1), thereby conserving anabolites. Among the CDH1-accumulated anabolites, ribose-5-phosphate (R5P) binds to transketolase-like-1 (TKTL1) to bridge CDH1 to cyclin-dependent kinase 2 (CDK2) and Skp1-Cullin-F-box and β-transducin repeat-containing protein (SCFβ -TRCP) complex, thereby facilitating CDH1 phosphorylation and degradation to promote cell cycle initiation. This CDH1-VHL-HIF1α/AARS2-R5P/TKTL1 circuit is supported by the observation that low R5P levels and high CDH1 expression correlate with proliferating cancer cells and tissues. Moreover, it is demonstrated that an artificial R5P signal, generated by ribose-5-sulfate (R5S), sensitizes cancer cells to apoptosis by initiating the cell cycle in the absence of sufficient anabolite supply. These suggest that cancer signatures, including the Warburg effect and angiogenesis, are intrinsically driven by CDH1.

Cecal appendix is a unique niche for commensal bacteria, and has been considered the primary site for immunoglobulin A production. Yet its immune function in anti-infection immunity has not been fully understood. In order to elucidate whether cecal patch (CeP), the murine version of appendix, would influence the immune response induced by Mycobacterium tuberculosis (M. tb) and the vaccine effect of Bacillus Calmette-Guérin (BCG), BALB/c mice at 4 weeks of age received appendectomy or sham operation and recovered for 2 weeks before intranasal infection with 2 × 107 CFU Mycobacterium tuberculosis H37Ra. Appendectomy of mice led to a reduction in lung macrophage numbers 7 days post infection (p. i.), and aggravated lung immunohistopathology 4 weeks p. i.. Appendectomized mice vaccinated with 5 × 106 CFU BCG exhibited attenuated BCG-specific serum IgG, reduced lung/splenic IFN-γ+ T response, and weakened T proliferation and cytotoxicity, and eventually worsened lung pathology compared to sham operated mice. Mechanistically, we found that appendectomized mice at a young age (4 weeks) had an attenuated maturation of mesenteric lymph node (MLN) conventional dendritic cells (cDCs), which accounted for the impaired systemic IFN-γ+ T response and cytotoxicity against M. tb. Our data suggest that intact appendix maintain intestinal DC maturation and systemic Th1 induction against M. tb and has an assistant role in increasing immune efficiency of BCG vaccine.

Lactate, a metabolite which is elevated in various developmental and pathological processes, exerts its signal through alanyl tRNA synthetases (AARS)-catalyzed protein lactylation. Herein, we report that elevated lactate and gain-of-function mitochondrial AARS (AARS2) mutations-induced hyper-lactylation promotes premature ovarian insufficiency (POI). Serum lactate is elevated in POI patients. POI-driving AARS2 mutations gain lactyltransferase activity. AARS2 lactylates and inactivates carnitine palmitoyl transferase 2 (CPT2), resulting in FFA accumulation that activates peroxisome proliferator-activated receptor γ (PPARγ), and potentiates follicle-stimulating hormone (FSH) to initiate follicle development. These, in synergy with the anabolites accumulation effects of AARS2, promoted lactylation-induced PDHA1 inactivation promote granular cell (GC) proliferation and primordial follicle development. GC-specific AARS2 overexpression does not affect primordial follicle number but speed up follicle depletion. AARS2 ablation or lactylation-inhibiting β-alanine treatments can prevent folliculogenesis and POI traits in mouse. These findings reveal that lactate signal drives follicle development, and inhibiting lactate signal could treat/prevent POI.

T cell engagers (TCEs) have been clinically validated but are mostly limited to hematological cancers. Developing BiTEs for solid tumors faces hurdles such as T cell penetration into tissue, suppressive nature of the tumor microenvironment and systemic toxicity, etc. This study aims to tackle these challenges by delivering an mRNA-encoded TCE targeting the hepatocellular carcinoma antigen Glypican-3 (GPC3) directly to liver tissue, thus enhancing the local TCE concentration to promote anti-tumor activity and exhibiting a favorable pharmacokinetics (PK) profile. The BiTE was engineered by fusing single-chain variable fragments (scFvs) targeting GPC3 and CD3, separated by a flexible peptide linker. The mRNA encoding the BiTE was engineered using a combinatorial approach to optimize codon usage and enhance translational efficiency and stability, balancing the Codon Adaptation Index (CAI) and Minimal Free Energy (MFE). The mRNA was encapsulated in a novel lipid nanoparticle (LNP) for efficient liver-tropic delivery, forming the complex named MTS105. BiTE’s cell-killing activity was tested using human GPC3-expressing cell lines and human PBMC. Anti-tumor efficacy was evaluated in liver orthotopic tumor-bearing mouse models. GLP Tox studies including PK and biodistribution were assessed in rodents and cynomolgus monkeys. Mechanistic studies demonstrated that MTS105-translated TCE mediated specific killing of GPC3-positive cells, T cell activation and proliferation in vitro and ex vivo. In the two liver orthotopic tumor-bearing mouse models, MTS105 achieved complete tumor regression in a dose-dependent manner, accompanied with intra-tumoral T cell activation. In mice, rats and cynomolgus monkeys, MTS105-translated TCE consistently exhibited higher exposure levels in the liver than the plasma. The sustained release of MTS105-translated TCE resulted in higher functional protein exposure in liver/tumor, while showing lower peripheral TCE Cmax (maximum concentration) and AUC (area under the curve) than the Fc-domain containing antibody-based TCE. In the cynomolgus monkeys, MTS105 displayed favorable and linear plasma PK profiles including mRNA, lipid and translated TCE in the up to 100 μg/kg following a single and 4-time repeat dosing weekly. The time to peak (Tmax) was 6 to 12 h in both periphery and targeting tissues, which correlated with the observation of fast Intra-tumoral T cell activity in mice. All monkeys tolerated well to a dose up to 100 μg/kg, with no sever adverse effects and gross pathology observed. The mRNA-encoded TCE demonstrated optimal tissue/tumor-specific PK, safety and potent anti-tumor activity in preclinical models. These preclinical studies enabled a first-in-human study to assess its safety and preliminary efficacy which is currently ongoing. Yan Huang, Shaoli Liu, Xiaoju Zhang, Bingxu Zhang, Hongya Han, Andong Liu, Xiaoyun Ma, Wei Xu. Preclinical development of an mRNA-encoded T cell engager targeting Glypican-3 with tissue- and tumor-specific enrichment in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3502.

Calcineurin is a calcium- and calmodulin-dependent serine/threonine protein phosphatase that connects the Ca2+-dependent signalling to multiple cellular responses. Calcineurin inhibitors (CNIs) have been widely used to suppress immune response in allograft patients. However, CNIs significantly increase cancer incidence in transplant recipients compared with the general population. Accumulating evidence suggests that CNIs may promote the malignant transformation of cancer cells in addition to its role in immunosuppression, but the underlying mechanisms remain poorly understood. Here, we show that calcineurin interacts with pyruvate dehydrogenase complex (PDC), a mitochondrial gatekeeper enzyme that connects two key metabolic pathways of cells, glycolysis and the tricarboxylic acid cycle. Mitochondrial-localized calcineurin dephosphorylates PDHA1 at Ser232, Ser293 and Ser300, and thus enhances PDC enzymatic activity, remodels cellular glycolysis and oxidative phosphorylation, and suppresses cancer cell proliferation. Hypoxia attenuates mitochondrial translocation of calcineurin to promote PDC inactivation. Moreover, CNIs promote metabolic remodelling and the Warburg effect by blocking calcineurin-mediated PDC activation in cancer cells. Our findings indicate that calcineurin is a critical regulator of mitochondrial metabolism and suggest that CNIs may promote tumorigenesis through inhibition of the calcineurin-PDC pathway.