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摘要： Antibody-based therapies have emerged as a powerful strategy for the management of diverse cancers. Unfortunately, tumor-specific antigens remain challenging to identify and target. Recent work established that inhibitor-modified peptide adducts derived from KRAS G12C are competent for antigen presentation via MHC I and can be targeted by antibody-based therapeutics, offering a means to directly target an intracellular oncoprotein at the cell surface with combination therapies. Here, we validated antigen display of "haptenated" KRAS G12C peptide fragments on tumors in mouse models treated with the FDA-approved KRAS G12C covalent inhibitor Sotorasib using PET/CT imaging of an 89Zr-labeled P1B7 IgG antibody, which selectively binds Sotorasib-modified KRAS G12C MHC I complexes. Targeting this peptide-MHC I complex with radioligand therapy using 225Ac- or 177Lu-P1B7 IgG effectively inhibited tumor growth in combination with Sotorasib. Elucidation of the 3.1 Å cryo-EM structure of P1B7 bound to a haptenated KRAS G12C peptide-MHC I complex confirmed that the Sotorasib-modified KRAS G12C peptide is presented via a canonical binding pose and showed that P1B7 binds the complex in a T-cell receptor-like manner. Together, these findings demonstrate the potential value of targeting unique oncoprotein-derived, haptenated MHC I complexes with radioligand therapeutics and provide a structural framework for developing next generation antibodies.

摘要： The pathophysiology of sickle cell disease (SCD) is characterized by hemolytic anemia and vaso-occlusion, although its impact on the adaptive immune responses remains incompletely understood. To comprehensibly profile the humoral immune responses, we immunized SCD mice with T cell-independent (TI) and T cell-dependent (TD) antigens. Our study showed that SCD mice have significantly enhanced type 2 TI (TI-2) immune responses in a manner dependent on the level of type I IFN (IFN-I), while maintaining similar or decreased TD immune responses depending on the route of antigen administration. Consistent with the enhanced TI-2 immune responses in SCD mice, the frequencies of B-1b cells (B-1 cells in humans), a major cell type responding to TI-2 antigens, were significantly increased in both the peritoneal cavity and spleens of SCD mice and in blood of patients with SCD. In support of expanded B-1 cells, elevated levels of anti-red blood cell (RBC) autoantibodies were detected in both SCD mice and patients. Both the levels of TI-2 immune responses and anti-RBC autoantibodies were significantly reduced following IFNAR antibody blockades and in IFNAR-1 deficient SCD mice. Moreover, the alteration of B-1 cell subsets were reversed in IFNAR-1 deficient SCD mice, uncovering a critical role for IFN-I in the enhanced TI-2 immune responses and the increased production of anti-RBC autoantibodies by modulating the innate B-1 cell subsets in SCD. Overall, our study provides experimental evidence that the modulation of B-1 cells and IFN-I can regulate TI immune responses and the levels of anti-RBC autoantibodies in SCD.

关键词： neopeptide screening   multifocal lung adenocarcinoma   NANOGNB   immunogenicity neopeptide   artificial antigen-presenting cells  

摘要： Background Mutations commonly occur in cancer cells, arising neoantigen as potential targets for personalized immunotherapy of lung adenocarcinoma (LUAD). However, the substantial heterogeneity observed among individuals and distinct foci within the same patient presents significant challenges in formulating immunotherapy strategies. The aim of the work is to characterize the mutation pattern and identify neopeptides across different patients and diverse foci within the same patients with *** Seven lung adenocarcinoma samples and matched tissues/blood are collected from 4 patients with LUAD for whole exome sequencing, mutation signature analysis, HLA binding prediction and neoantigen screening. Dimeric HLA-A2 molecules were prepared by Bac-to-Bac baculovirus expression system to establish a T cell stimulation system based on HLA-A2-coated artificial antigen-presenting cells for the validation of immunogenic *** Similar mutation pattern with predominant missense mutation and high tumor mutation burden was observed across individuals with lung adenocarcinomas and between non-invasive and invasive foci. We screened and identified 3 consistent mutated genes among 100 top genes with highest mutation scores contributed across 4 patients, and 3 mutated peptides among 30 with highest HLA-A2 binding affinity distributed in at least 2 out of 4 foci in the same patient. Notably, LUAD-7-MT peptide encoded by NANOGNB demonstrated higher immunogenicity in promoting CD8+ T cells proliferation and IFN-gamma secretion than the corresponding wildtype *** This study provides an in-depth analysis of mutation characteristics of LUAD and establishes a neoantigen screening and validation system for identifying immunogenicity neopeptide across individual patients and diverse foci in the same patient with multifocal LUAD.

关键词： MT: Delivery Strategies   IL-12   mRNA   lipid nanoparticles   stereotactic body radiation therapy   antitumor immune response   pancreatic cancer   T cell immunity  

摘要： Pronounced T cell exhaustion characterizes immunosuppressive tumors, with the tumor microenvironment (TME) employing multiple mechanisms to elicit this suppression. Traditional immunotherapies, such as immune checkpoint blockade, often fail due to their focus primarily on T cells. To overcome this, we utilized a proinflammatory fl ammatory cytokine, interleukin (IL)-12, that re-wires the immunosuppressive TME by inducing T cell effector function while also repolarizing immunosuppressive myeloid cells. Due to toxicities observed with systemic administration of this cytokine, we utilized lipid nanoparticles encapsulating mRNA encoding IL-12 for intratumoral injection. This strategy has been proven safe and tolerable in early clinical trials for solid malignancies. We report an unprecedented loss of exhausted T cells and the emergence of an activated phenotype in murine pancreatic ductal adenocarcinoma (PDAC) treated with stereotactic body radiation therapy (SBRT) and IL-12mRNA. Our mechanistic fi ndings reveal that each treatment modality contributes to the T cell response differently, with SBRT expanding the T cell receptor repertoire and IL12mRNA promoting robust T cell proliferation and effector status. This distinctive T cell signature mediated marked growth reductions and long-term survival in local and metastatic PDAC models. This is the fi rst study of its kind combining SBRT with IL-12mRNA and provides a promising new approach for treating this aggressive malignancy.

关键词： Caspase-3   Caspase-8   GSDME   Ghrelin   High glucose   Pyroptosis   TNF-α  

摘要： Diabetic liver injury (DLI) refers to liver injury resulting from prolonged chronic hyperglycemia and represents a significant complication associated with diabetes, The specific pathogenic mechanism of DLI remains incompletely understood. Tumor necrosis factor α (TNF-α) has been demonstrated to play a crucial role in diabetic complications through intricate signalling pathways, including pyroptosis. However, it remains uncertain whether TNF-α mediates pyroptosis in DLI, we initially established an in vitro model of DLI and confirmed the presence of an inflammatory state characterized by TNF-α in DLI. Furthermore, evidence of gasdermin E (GSDME)-mediated pyroptosis and the activation of cysteinyl aspartate specific proteinase (caspase)-8 was observed in AML-12 cell exposed to high glucose concentrations. We subsequently demonstrated that TNF-α can trigger caspase-8 activation, leading to GSDME-mediated cellular pyroptosis. Furthermore, treatment with ghrelin effectively suppressed hepatic cell pyroptosis induced by high glucose concentrations and provided protection against liver injury. Therefore, we propose that the TNF-α/caspase-8/caspase-3/GSDME pathway represents a novel mechanism underlying pyrodeath in DLI cells and to explore the protective role and molecular mechanisms underlying the effects of ghrelin on DLI by this special pathway, These findings may present potential therapeutic implications for the management of DLI.

摘要： Long-term allograft survival is limited by humoral-associated chronic allograft rejection, suggesting inadequate constraint of humoral alloimmunity by contemporary immunosuppression. Heterogeneity in alloreactive B cells and the incomplete definition of which B cells participate in chronic rejection in immunosuppressed transplant recipients limits our ability to develop effective therapies. Using a double-fluorochrome single-HLA tetramer approach combined with single-cell culture, we investigated the B-cell receptor (BCR) repertoire characteristics, avidity, and phenotype of donor HLA-DQ reactive B cells in a transplant recipient with end-stage donor specific antibody (DSA)-associated cardiac allograft vasculopathy while receiving maintenance immunosuppression (tacrolimus, mycophenolate mofetil, prednisone). Donor DQB1*03:02/DQA1*03:01 (DQ8)-reactive IgG+ B cells were enriched for minimally mutated and germline encoded high avidity BCRs (median K 4.26×10 ) with an atypical, antigen-experienced and proliferative phenotype (CD27 CD21 CD71 CD11c ). These B cells coexisted with a smaller subset of more highly mutated, affinity matured IgG+CD27+ B cells. Circulating donor-reactive B cells and DSA remained detectable after rituximab, contrasting with the marked reduction in DSA after allograft explant and retransplant. Together, these findings define the persistence of germline high-avidity HLA-DQ alloreactive B cells and their co-existence with affinity matured clones that were both driven by the allograft despite conventional immunosuppression.

关键词： 创伤性脑损伤   骨痂   骨折愈合   炎症反应   细胞因子   激素   神经肽   基因   干细胞  

摘要： 背景：骨折延迟愈合和不愈合是常见的临床问题。在临床观察中，常见到合并创伤性脑损伤的四肢骨折患者的骨折愈合速度明显快于无脑损伤的患者。这一现象背后的可能机制已成为当前研究的重要焦点。近年来的研究表明，创伤性脑损伤通过调控细胞因子、激素、神经信号以及干细胞等机制，显著加速了骨痂形成和骨折愈合过程。目的：总结创伤性脑损伤促进骨痂形成及骨折愈合机制的最新研究进展，为临床应用提供理论依据。方法：文章第一作者通过检索中国知网、万方、维普、PubMed、Embase、Web of Science及Cochrane Library数据库2013年1月至2024年10月相关文献，个别文献追溯至20年前。检索词为“创伤性脑损伤，骨痂，骨折愈合，炎症反应，细胞因子，激素，神经肽，基因，干细胞”“traumatic brain injury,callus,fracture healing,inflammatory response,cytokines,hormones,neuropeptides,genes,stem cells”，最终选取符合纳入标准的文献共计83篇。结果与结论：创伤性脑损伤对骨痂形成及骨折愈合的促进作用机制极其复杂，涉及细胞因子、激素、神经系统和干细胞等多个调节环节。然而，目前关于创伤性脑损伤促进骨痂形成及骨折愈合的具体机制尚未得到全面、清晰的理解，仍需进一步深入研究。现有研究表明，创伤性脑损伤主要通过促进细胞因子(如胰岛素样生长因子1)和激素(如生长激素及瘦素)的释放，调控神经系统，促进干细胞的增殖与分化等机制来加速骨痂形成和骨组织再生。此外，创伤性脑损伤会引发一系列免疫反应，包括炎症因子的释放和免疫细胞的激活，这些反应能够调节骨折修复的过程。具体而言，免疫反应通过增强局部血流、促进细胞迁移和激活成纤维细胞，从而支持骨愈合的不同阶段。同时，创伤性脑损伤诱发的干细胞活化也在骨折修复中发挥了关键作用。活化的干细胞能够分化为骨细胞、软骨细胞和脂肪细胞，促进骨组织再生和修复。因此，创伤性脑损伤诱发的免疫反应与干细胞的活化共同作用，为骨折愈合提供了必要的支持。这些机制能够加速骨愈合过程，显著缩短愈合时间，提升骨折患者的预后。综上所述，创伤性脑损伤通过多重机制促进骨痂形成和骨折愈合，展现其在骨修复中的重要性。未来研究应聚焦创伤性脑损伤影响骨愈合的信号通路及调控因子，深入理解其作用机制，这将为开发靶向治疗、干细胞及神经调控疗法提供理论基础，具有缩短愈合时间、优化康复方案、改善预后的临床价值。因此，探索创伤性脑损伤引发的生物效应将为骨折治疗开辟新途径。

关键词： dECM   脱细胞化细胞外基质   巨噬细胞极化   ROS   炎症  

摘要： 目的：研究脱细胞化细胞外基质材料（dECM）浸提液对巨噬细胞存活、迁移作用、吞噬功能、促炎因子表达、ROS产生的影响；方法：通过光镜下观察、CCK-8法检测dECM材料浸提液对巨噬细胞有无细胞毒性；通过Transwell迁移实验检测dECM材料浸提液对巨噬细胞募集和趋化作用的影响；通过pHrodo染色检测dECM材料浸提液对巨噬细胞吞噬功能的影响；通过RT-qPCR和免疫荧光染色检测dECM材料浸提液对巨噬细胞促炎基因和促炎特异性功能分子表达的影响；通过DCFH-DA荧光探针方法检测dECM材料浸提液对巨噬细胞ROS产生的影响。结果：与DMEM完全培养基相比，①CCK-8实验结果显示dECM材料浸提液对巨噬细胞没有毒性，能促进巨噬细胞集落形成；②pHrodo染色实验结果显示dECM材料浸提液不影响巨噬细胞的吞噬功能；③Transwell迁移实验结果显示dECM材料浸提液不促进巨噬细胞发生迁移；④RT-qPCR结果显示dECM材料浸提液能够下调促炎细胞因子和特异性促炎功能分子的基因表达；⑤流式细胞术结果显示dECM材料浸提液能够降低巨噬细胞产生ROS；结果表明，dECM材料浸提液对于巨噬细胞具有良好的生物相容性。

关键词： antigen presentation   checkpoint blockade   ERAP   HLA-E   immunotherapy   NKG2A   peptide loading complex   Qa-1b   Qdm   VL9  

摘要： The aminopeptidase, endoplasmic reticulum aminopeptidase 1 (ERAP1), trims peptides for loading into major histocompatibility complex class I (MHC class I), and loss of this activity has broad effects on the MHC class I peptidome. Here, we investigated the impact of targeting ERAP1 in immune checkpoint blockade (ICB), as MHC class I interactions mediate both activating and inhibitory functions in antitumor immunity. Loss of ERAP sensitized mouse tumor models to ICB, and this sensitivity depended on CD8⁺ T cells and natural killer (NK) cells. In vivo suppression screens revealed that Erap1 deletion inactivated the inhibitory NKG2A-HLA-E checkpoint, which requires presentation of a restricted set of invariant epitopes (VL9) on HLA-E. Loss of ERAP altered the HLA-E peptidome, preventing NKG2A engagement. In humans, ERAP1 and ERAP2 showed functional redundancy for the processing and presentation of VL9, and loss of both inactivated the NKG2A checkpoint in cancer cells. Thus, loss of ERAP phenocopies the inhibition of the NKG2A-HLA-E pathway and represents an attractive approach to inhibit this critical checkpoint. © 2024 The Authors

摘要： The peripheral nervous system has been widely implicated in pathological conditions that exhibit distinct clinical presentations in men and women, most notably in chronic pain disorders. Here, we explored this sexual dimorphism at a molecular level. We expanded the available omics landscape in the PNS to include quantitative proteomics of the human dorsal root ganglia (hDRG) and nerve. Using data-independent acquisition mass spectrometry, we uncovered an extensive protein landscape, validated against tissue-specific differences between the nerve and hDRG. Using a combination of multi-omic analyses and functional support, we then examined sex-differences, highlighting TNFα signalling as a relevant sexually dimorphic pathway in males. These results support a functional sexually dimorphism in the periphery, which is of particular importance to sensory- and pain-related clinical translation.