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Triple-negative breast cancer (TNBC) is an aggressive and clinically challenging subtype defined by the absence of estrogen receptor,progesterone receptor,and HER2 amplification,resulting in poor prognosis and limited therapeutic options. Targeting alternative molecular pathways is urgently needed to overcome resistance and improve patient outcomes. CD147 has emerged as surface marker associated with tumor progression and immune evasion. In this study,CD147 and MHC class I—a key inhibitory ligand for natural killer cells—were analyzed in breast cancer cell lines (MCF7,MDA-MB-453,MDA-MB-231,and HCC38) using flow cytometry. The therapeutic efficacy of a humanized anti-CD147 monoclonal antibody (HuM6-1B9) was evaluated for its capacity to potentiate antibody-dependent cellular cytotoxicity (ADCC). HuM6-1B9 demonstrated the strong binding to MDA-MB-231 (KD = 4.982 nM) and HCC38 (KD = 4.523 nM),which are representative TNBC cell lines. In 3D spheroid models,HuM6-1B9 significantly enhanced PBMC-mediated ADCC,leading to a marked reduction in TNBC spheroid viability. Co-culture of CFSE-labeled MDA-MB-231 and HCC38 cells with primary NK cells confirmed robust ADCC,achieving 50% and 70% cytotoxicity,respectively,despite high MHC class I expression. Live-cell imaging demonstrated caspase-3/7 activation consistent with apoptosis in NK-targeted tumor cells,while CD107a degranulation and IFN-γ secretion confirmed the functional contribution of HuM6-1B9 to ADCC enhancement. Importantly,HuM6-1B9 did not promote migration or invasion in MDA-MB-231 cells,supporting its safety profile regarding metastasis. Collectively,these findings establish HuM6-1B9 as a promising immunotherapeutic candidate that overcomes immune resistance and selectively eliminates TNBC cells through ADCC without enhancing metastatic potential. By integrating mechanistic assays of NK cytotoxicity,apoptosis,and 3D tumor spheroids,this study provides clinically relevant insights underscoring the translational potential of HuM6-1B9 in TNBC immunotherapy. View Publication

Proteomic analysis of human cerebral organoids may reveal how psychedelics regulate biological processes,shedding light on drug-induced changes in the brain. This study elucidates the proteomic alterations induced by lysergic acid diethylamide (LSD) in human cerebral organoids. By employing high-resolution mass spectrometry-based proteomics,we quantitatively analyzed the differential abundance of proteins in cerebral organoids exposed to LSD. Our findings indicate changes in proteostasis,energy metabolism,and neuroplasticity-related pathways. Specifically,LSD exposure led to alterations in protein synthesis,folding,autophagy,and proteasomal degradation,suggesting a complex interplay in the regulation of neural cell function. Additionally,we observed modulation in glycolysis and oxidative phosphorylation,crucial for cellular energy management and synaptic function. In support of the proteomic data,complementary experiments demonstrated LSD’s potential to enhance neurite outgrowth in vitro,confirming its impact on neuroplasticity. Collectively,our results provide a comprehensive insight into the molecular mechanisms through which LSD may affect neuroplasticity and potentially contribute to therapeutic effects for neuropsychiatric disorders. View Publication

SummaryTissue regeneration following an injury requires dynamic cell-state transitions that allow for establishing the cell identities required for the restoration of tissue homeostasis and function. Here,we present a biochemical intervention that induces an intermediate cell state mirroring a transition identified during normal differentiation of myoblasts and other multipotent and pluripotent cells to mature cells. When applied in somatic differentiated cells,the intervention,composed of one-carbon metabolites,reduces some dedifferentiation markers without losing the lineage identity,thus inducing limited reprogramming into a more flexible cell state. Moreover,the intervention enabled accelerated repair after muscle injury in young and aged mice. Overall,our study uncovers a conserved biochemical transitional phase that enhances cellular plasticity in vivo and hints at potential and scalable biochemical interventions of use in regenerative medicine and rejuvenation interventions that may be more tractable than genetic ones. Graphical abstract Highlights•Early cell transitions in differentiation include metabolites,supporting identity changes•Cell-transition biochemicals can be leveraged to induce plasticity•1C-metabolite supplementation streamlines cell-identity changes in vitro•1C-metabolite in vivo administration impacts acetylation genes,aiding muscle regeneration Hernandez-Benitez et al. identify a metabolomic wave conserved in the early transition of cells differentiating in vitro,and they leverage this finding to customize an in vivo supplementation that facilitates the transition of cell phenotypes when needed,like in regeneration after an injury. View Publication

IntroductionUpon infection,T cell-driven B cell responses in GC reactions induce memory B cells and antibody-secreting cells that secrete protective antibodies. How formation of specifically long-lived plasma cells is regulated via the interplay between specific B and CD4+ T cells is not well understood. Generally,antibody levels decline over time after clearance of the primary infection.MethodIn this study,convalescent individuals with stable RBD antibody levels (n=14,“sustainers”) were compared with donors (n=13) with the greatest antibody decline from a cohort of 132. To investigate the role of the cellular immune compartment in the maintenance of antibody levels,SARS-CoV-2-specific responses at 4 to 6 weeks post-mild COVID-19 infection were characterized using deep immune profiling.ResultsBoth groups had similar frequencies of total SARS-CoV-2-specific B and CD4+ T cells. Sustainers had fewer Spike-specific IgG+ memory B cells early after infection and increased neutralizing capacity of RBD antibodies over time,unlike the declining group. However,declining IgG titers correlated with lower frequency of Spike-specific CD4+ T cells.ConclusionThese data suggest that “sustainers” have unique dynamics of GC reactions,yield different outputs of terminally differentiating cells,and improve the quality of protective antibodies over time. This study helps identify factors controlling formation of long-lived PC and sustained antibody responses. View Publication

Immune evasion by cancer cells involves reshaping the tumor microenvironment (TME) via communication with non-malignant cells. However,resistance-promoting interactions during treatment remain lesser known. Here we examine the composition,communication,and phenotypes of tumor-associated cells in serial biopsies from stage II and III high-risk estrogen receptor positive (ER+ ) breast cancers of patients receiving endocrine therapy (letrozole) as single agent or in combination with ribociclib,a CDK4/6-targeting cell cycle inhibitor. Single-cell RNA sequencing analyses on longitudinally collected samples show that in tumors overcoming the growth suppressive effects of ribociclib,first cancer cells upregulate cytokines and growth factors that stimulate immune-suppressive myeloid differentiation,resulting in reduced myeloid cell- CD8 + T-cell crosstalk via IL-15/18 signaling. Subsequently,tumors growing during treatment show diminished T-cell activation and recruitment. In vitro,ribociclib does not only inhibit cancer cell growth but also T cell proliferation and activation upon co-culturing. Exogenous IL-15 improves CDK4/6 inhibitor efficacy by augmenting T-cell proliferation and cancer cell killing by T cells. In summary,response to ribociclib in stage II and III high-risk ER +  breast cancer depends on the composition,activation phenotypes and communication network of immune cells. The CDK4/6 inhibitor ribociclib holds promise in cancer therapy but how cell cycle inhibitory drugs affect the anti-tumor immune response remains a question. Here authors show that poor response of early-stage estrogen receptor positive breast cancers to ribociclib is caused by changes in the immune cell composition and cancer-cell-immune-cell communication in the tumors rather than intrinsic cancer cell resistance to cell cycle inhibition. View Publication

BACKGROUND Vascularization is important for the clinical application of tissue engineered products. Both adipose-derived stem cells (ASCs) and surgical prefabrication can be used to induce angiogenesis in scaffolds. Our aim was to compare the angiogenic potential of ASC-seeded scaffolds combined with scaffold prefabrication with that of non-seeded,non-prefabricated scaffolds. METHODS For prefabrication,functional blood vessels were introduced into the scaffold using a flow-through pedicle system. ASCs were isolated from rat fat deposits. Three-dimensional-printed cylindrical poly-$\epsilon$-caprolactone scaffolds were fabricated by fused deposition modelling. Three groups,each containing six rats,were investigated by using non-seeded,ASC-seeded,and osteogenic induced ASC-seeded scaffolds. In each group,one rat was implanted with two scaffolds in the inguinal region. On the right side,a scaffold was implanted subcutaneously around the inferior epigastric vessels (classic prefabrication group). On the left side,the inferior epigastric vessels were placed inside the prefabricated scaffold in the flow-through pedicle system (flow-through prefabrication group). The vessel density and vascular architecture were examined histopathologically and by $\mu$CT imaging,respectively,at 2 months after implantation. RESULTS The mean vessel densities were 10- and 5-fold higher in the ASC-seeded and osteogenic induced ASC-seeded scaffolds with flow-through prefabrication,respectively,than in the non-seeded classic prefabricated group (p {\textless} 0.001). $\mu$CT imaging revealed functional vessels within the scaffold. CONCLUSION ASC-seeded scaffolds with prefabrication showed significantly improved scaffold vasculogenesis and could be useful for application to tissue engineering products in the clinical settings. View Publication

Environmental factors contribute to Type 1 diabetes (T1D) susceptibility. The gut microbiome,which includes bacteria,viruses,and fungi,contributes to this environmental influence,and can induce immunological changes. The gut viral component of the microbiome,related to T1D has mostly focused on coxsackieviruses and rotavirus. The role of norovirus,another common enteric virus,in susceptibility to T1D was hitherto unknown. Norovirus is highly infectious and encountered by many children. We studied the mouse norovirus 4 (MNV4),related to human noroviruses,in the Non-obese diabetic (NOD) mouse model,to determine its role in influencing susceptibility to T1D. We infected MNV-free NOD mice with MNV4 by exposing the mice to MNV4-positive bedding from an endemically-infected mouse colony to mimic a natural infection. Control MNV-free NOD mice were exposed to MNV-free bedding from the same colony. Interestingly,MNV4 infection protected NOD mice from the development of T1D and was associated with an expansion of Tregs and reduced proinflammatory T cells. We also found MNV4 significantly modified the gut commensal bacteria composition,promoting increased $\alpha$-diversity and Firmicutes/Bacteroidetes ratio. To elucidate whether T1D protection was directly related to MNV4,or indirectly through modulating gut microbiota,we colonized germ-free (GF) NOD mice with the MNV4-containing or non-MNV4-containing viral filtrate,isolated from filtered fecal material. We found that MNV4 induced significant changes in mucosal immunity,including altered Tuft cell markers,cytokine secretion,antiviral immune signaling markers,and the concentration of mucosal antibodies. Systemically,MNV4-infection altered the immune cells including B cell subsets,macrophages and T cells,and especially induced an increase in Treg number and function. Furthermore,in vitro primary exposure of the norovirus filtrate to na{\{i}}ve splenocytes identified significant increases in the proportion of activated and CTLA4-expressing Tregs. Our data provide novel knowledge that norovirus can protect NOD mice from T1D development by inducing the expansion of Tregs and reducing inflammatory T cells. Our study also highlights the importance of distinguishing the mucosal immunity mediated by bacteria from that by enteric viruses." View Publication

IntroductionAngelman Syndrome (AS) is characterized in large part by the loss of functional UBE3A protein in mature neurons. A majority of AS etiologies is linked to deletion of the maternal copy of the UBE3A gene and epigenetic silencing of the paternal copy. A common therapeutic strategy is to unsilence the intact paternal copy thereby restoring UBE3A levels. Identifying novel therapies has been aided by a UBE3A-YFP reporter mouse model. This study presents an analogous fluorescent UBE3A reporter system in human cells.MethodsPreviously derived induced Pluripotent Stem Cells (iPSCs) with a Class II large deletion at the UBE3A locus are used in this study. mGL and eGFP are integrated downstream of the endogenous UBE3A using CRISPR/Cas9. These reporter iPSCs are differentiated into 2D and 3D neural cultures to monitor long-term neuronal maturation. Green fluorescence dynamics are analyzed by immunostaining and flow cytometry.ResultsThe reporter is successfully integrated into the genome and reports paternal UBE3A expression. Fluorescence expression gradually reduces with UBE3A silencing in neurons as they mature. Expression patterns also reflect expected responses to molecules known to reactivate paternal UBE3A.DiscussionThis human-cell-based model can be used to screen novel therapeutic candidates,facilitate tracking of UBE3A expression in time and space,and study human-specific responses. However,its ability to restore UBE3A function cannot be studied using this model. Further research in human cells is needed to engineer systems with functional UBE3A to fully capture the therapeutic capabilities of novel candidates. View Publication

Affinity maturation and differentiation of B cells in the germinal center (GC) are tightly controlled by epigenetically regulated transcription programs,but the underlying mechanisms are only partially understood. Here we show that Cfp1,an integral component of the histone methyltransferase complex Setd1A/B,is critically required for GC responses. Cfp1 deficiency in activated B cells greatly impairs GC formation with diminished proliferation,somatic hypermutation and affinity maturation. Mechanistically,Cfp1 deletion reduces H3K4me3 marks at a subset of cell cycle and GC-related genes and impairs their transcription. Importantly,Cfp1 promotes the expression of transcription factors MEF2B and OCA-B and the Bcl6 enhancer-promoter looping for its efficient induction. Accordingly,Cfp1-deficient GCB cells upregulate IRF4 and preferentially differentiate into plasmablasts. Furthermore,Cfp1 ablation upregulates a panel of pre-memory genes with elevated H3K4me3 and leads to markedly expanded memory B populations. In summary,our study reveals that Cfp1-safeguarded epigenetic regulation ensures proper dynamics of GCB cells for affinity maturation and prevents the pre-mature exit from GC as memory cells. Cellular differentiation decisions,such as fates of B cells following entry into the germinal centres,are governed by epigenetically and transcriptionally regulated paths for bifurcating cell fates. Here the authors show that CFP1 is a master epigenetic regulator of activated B cells and controls their hypermutation and affinity maturation via the histone methyltransferase complex Setd1A/B. View Publication

Sickle cell disease is a devastating blood disorder that originates from a single point mutation in the HBB gene coding for hemoglobin. Here,we develop a GMP-compatible TALEN-mediated gene editing process enabling efficient HBB correction via a DNA repair template while minimizing risks associated with HBB inactivation. Comparing viral versus non-viral DNA repair template delivery in hematopoietic stem and progenitor cells in vitro,both strategies achieve comparable HBB correction and result in over 50% expression of normal adult hemoglobin in red blood cells without inducing β-thalassemic phenotype. In an immunodeficient female mouse model,transplanted cells edited with the non-viral strategy exhibit higher engraftment and gene correction levels compared to those edited with the viral strategy. Transcriptomic analysis reveals that non-viral DNA repair template delivery mitigates P53-mediated toxicity and preserves high levels of long-term hematopoietic stem cells. This work paves the way for TALEN-based autologous gene therapy for sickle cell disease. Subject terms: Targeted gene repair,Sickle cell disease View Publication

Proliferating cell nuclear antigen (PCNA),a well-documented anticancer target,is critical for DNA synthesis,replication,and repair. AOH1996,a small-molecule PCNA inhibitor,is currently undergoing clinical trials for the treatment of advanced solid tumors. However,the therapeutic effect of AOH1996 on head and neck squamous cell carcinoma (HNSCC) remains unclear. The effects of AOH1996 on HNSCC biological behaviors and cancer stemness were tested in HNSCC cells and nude mice. The combination treatment of AOH1996 and anti-PD1 was performed in a 4-nitroquinoline N-oxide (4NQO)-induced HNSCC mouse model. RNA sequencing,Western Blotting,immunofluorescence staining,comet assays,and qRT‒PCR were conducted for mechanistic studies. Our results showed that AOH1996 effectively inhibited HNSCC proliferation and invasion both in vitro and in vivo. AOH1996 suppressed HNSCC stemness,development,and metastasis. Moreover,AOH1996 altered the tumor immune microenvironment into an inflamed state with increased CD8 + T-cell infiltration,rendering it a favorable partner for combination therapy with immune checkpoint inhibitors. Mechanistically,AOH1996 induced cellular DNA damage,suppressed cancer stemness through the upregulation of p-TBK1,and promoted the secretion of CD8 + T-cell-recruiting chemokines by stimulating IRF3-mediated transcription. Taken together,our results demonstrated that AOH1996 suppressed tumor growth,eliminated cancer stem cells (CSCs),and synergistically enhanced the efficacy of anti-PD1 immunotherapy in HNSCC. The online version contains supplementary material available at 10.1186/s13287-025-04607-9. View Publication

Expansion of Human Hematopoietic Stem Cells (HSCs) is a rapidly advancing field showing great promise for clinical applications. Recent evidence has implicated the nervous system and glial family ligands (GFLs) as potential drivers of hematopoietic survival and self-renewal in the bone marrow niche,but how to apply this to HSC maintenance and expansion is yet to be explored. We demonstrate a role for the GFL receptor,RET,at the cell surface of HSCs,in mediating sustained cellular growth,resistance to stress and improved cell survival throughout in vitro expansion. HSCs treated with the key RET ligand/co-receptor complex,GDNF/GFRa1,show improved progenitor function at primary transplantation and improved long-term HSC function at secondary transplantation. Finally,we demonstrate that RET drives a multi-faceted intracellular signalling pathway,including key signalling intermediates AKT,ERK1/2,NFkB and p53,responsible for a wide range of cellular and genetic responses which improve cell growth and survival under culture conditions. View Publication