Although immunotherapy has transformed the treatment landscape for many types of cancer, its therapeutic efficacy in glioblastoma (GBM) is limited by insufficient antigen presentation and the immunogenic cell exclusion in the tumor microenvironment. Here, we develop a candidate-based CRISPR activation (CRISPRa) functional screen to identify regulators of conventional dendritic cell (cDC)-fate specification. We determine that the transcription factors Zfp366/Znf366, Pu.1, Irf8, and Batf3 (ZPIB) are sufficient to convert GBM cells into cDC-like cells. ZPIB-mediated reprogramming results in global transcriptional and epigenetic remodeling in glioma cells. Single-cell RNA sequencing (scRNA-seq) profiling also reveals efficient and dynamic reprogramming of GBM cells to cDCs in vivo. Moreover, reprogrammed tumor cells remodel the microenvironment and elicit systemic tumor-eradicating and durable antitumor immunity in multiple mouse GBM models. Antitumor immunity elicited by ZPIB-DCs is synergistic with immune checkpoint inhibitors. Finally, we evaluate the clinical applicability of this approach by generating ZPIB-DCs from GBM patients within a humanized model. Our study represents a cellular reprogramming therapeutic strategy with broad implications for clinical immunotherapy.

Background The body-protective polypeptide BPC157 is a natural therapeutic agent that has demonstrated robust efficacy in promoting repair across multiple organ systems, indicating its significant clinical potential. However, its clinical translation has been substantially hindered by a lack of understanding of its precise molecular mechanisms of action. Given its prominent proangiogenic effects, deciphering the mechanistic basis of its vascular regenerative actions is critically important. Methods To investigate the molecular pathway underlying the proangiogenic effects of BPC157, we employed a combination of molecular, biochemical, and cellular approaches. These include techniques to identify protein‒protein interactions, assess ubiquitination status, and evaluate key cellular functions. The specific role of the proline residue at position 3 of BPC157 was also experimentally validated. Results We revealed that intracellular BPC157 engages the E3 ubiquitin ligase adaptor protein FBXO22 through its proline residue at position 3. This interaction results in the formation of a protein complex that effectively suppresses the ubiquitination and subsequent proteasomal degradation of the transcription factor BACH1, leading to significant stabilization of BACH1 protein levels. The consequent accumulation of BACH1 was shown to enhance critical processes in vascular regeneration, namely, the proliferation and tube-forming capacity of vascular endothelial cells. Conclusions This study establishes a novel and critical molecular mechanism for the pharmacological actions of BPC157, centered on the BPC157-FBXO22-BACH1 axis. These findings provide a solid mechanistic foundation for the future development of BPC157-based or sequential therapeutic agents targeted at enhancing vascular repair. Furthermore, this work offers robust scientific evidence that can inform improved strategies for the prevention and treatment of tissue injury.

BackgroundTriple-negative breast cancer (TNBC) is an aggressive subtype characterized by the absence of estrogen receptor, progesterone receptor, and HER2 expression, which limits the availability of targeted therapies and results in poor prognosis. Immune checkpoint blockade (ICB) therapies have emerged as promising treatments by enhancing anti-tumor immunity; however, a substantial proportion of patients with TNBC exhibit primary or acquired resistance. This resistance is largely influenced by the tumor microenvironment (TME). This study uses integrated single-cell and spatial transcriptomics to elucidate key cellular mechanisms of resistance, with particular emphasis on lipid-mediated stromal-immune interactions within the TNBC TME.MethodsThis investigation encompassed analysis of single-cell RNA sequencing (scRNA-seq) data from three TNBC datasets and spatial transcriptomic data from 43 TNBC samples. Spatial niches and cell-cell interactions were identified using the Multimodal Intersection Analysis (MIA) algorithm. Experimentally, adipose-derived mesenchymal stem cells (AD-SCs) were co-cultured with MDA-MB-231 TNBC cells to generate lipid-processing CAFs (lpCAFs) and subsequently co-cultured with THP-1 macrophages. Lipid metabolism and M2 polarization of macrophages were assessed using BODIPY staining, Oil Red O, qPCR, flow cytometry and Western blotting techniques.ResultsABCA8+ lpCAFs and APOE+ lipid-associated macrophages (LAMs) exhibited significant enrichment in ICB-resistant TNBC, with co-localization at the immune-stromal junction. lpCAFs facilitated M2 macrophage polarization through lipid metabolism reprogramming, establishing an immunosuppressive TME. High ABCA8 expression demonstrated correlation with enhanced M2 macrophage infiltration, decreased cytotoxic immune cells, and poorer prognosis. Experimental validation demonstrated that lpCAFs increased expression of lipid metabolism and M2 polarization marker in macrophages, substantiating their immunosuppressive function.ConclusionABCA8+ lpCAFs and APOE+ LAMs contribute to ICB resistance in TNBC through the establishment of an immunosuppressive TME via lipid metabolism reprogramming. Therapeutic intervention targeting the ABCA8-lipid axis presents a promising strategy to enhance ICB efficacy, potentially advancing TNBC treatment outcomes and improving patient survival.

BACKGROUND:Hepatocellular carcinoma (HCC) is a leading cause of cancer mortality with limited therapeutic options. Despite promising immunotherapy, response rates remain suboptimal. Tumour-associated macrophages (TAMs) constitute a pivotal component of the immunosuppressive HCC microenvironment, yet TAM heterogeneity and contributions to tumour progression and immunotherapy resistance remain poorly defined. OBJECTIVE:To identify and characterise critical TAM subsets in HCC and evaluate their potential as therapeutic targets. DESIGN:Integrated multiomics analysis of hepatocellular carcinoma (HCC) clinical specimens was performed and validated across independent cohorts. Single-cell RNA sequencing identified tumour-associated macrophage (TAM) subpopulations. Functional characterisation employed whole-body and macrophage-specific CD48 knockout mice, adoptive transfer experiments and co-culture systems. Mechanistic studies used immunoprecipitation-mass spectrometry, immunofluorescence colocalisation and pathway analysis. Therapeutic efficacy was evaluated using anti-CD48 monotherapy and combination with anti-programmed cell death protein 1 (PD1) in orthotopic HCC models. RESULTS:CD48+ TAMs were identified associating with accelerated tumour progression, immunotherapy resistance and poor clinical outcomes. These TAMs exhibited protumorous phenotypes, driving immunosuppression and promoting extracellular matrix remodelling. Genetic CD48 ablation attenuated HCC progression while promoting CD8+ T-cell function. Adoptive transfer of CD48-deficient macrophages validated tumour-suppressive effects. Mechanistically, matrix metalloproteinase-14 (MMP14) was identified as a novel cis-interacting partner for CD48, functioning independently of the canonical CD48-CD244 axis. This interaction activated RAP1 GTPase, triggering Yes-associated protein (YAP) nuclear translocation and YAP-signal transducer and activator of transcription 3 (STAT3) complex formation to upregulate immunosuppressive genes. Anti-CD48 antibodies effectively inhibited tumour progression and demonstrated synergistic effects with anti-PD1 therapy. CONCLUSION:CD48 represents a novel immune checkpoint on TAMs critical for HCC progression and immunotherapy resistance. Targeting CD48 may overcome immunosuppression and increase therapeutic efficacy in HCC.

Trastuzumab-based HER2-targeted therapy remains the cornerstone treatment for HER2-positive breast cancer. However, its clinical efficacy is significantly modulated by the tumor microenvironment (TME). Our single-cell sequencing analysis of clinical samples revealed that patients with poor radiologic response after trastuzumab-based neoadjuvant therapy presented significant enrichment of TIGIT+ NK cells with high immune checkpoint expression, exhausted CD8+ T cells, and immunosuppressive regulatory T cells (Tregs). Further analyses leveraging cell-cell communication, spatial transcriptomics, and multiplex immunofluorescence showed that SPP1+ tumor-associated macrophages (SPP1+ TAMs) enrichment was associated with dysfunctional NK- and T-cell states in tumors from patients with poor radiologic response. Functional validation studies revealed that SPP1+ TAMs actively induced exhaustion phenotypes in both NK cells and CD8+ T cells, thereby impairing trastuzumab-dependent antibody-dependent cellular cytotoxicity (ADCC) and adaptive immune responses. In vivo experiments using humanized NCG murine models further confirmed the SPP1+ TAMs-mediated suppression of NK cell function. Significantly, HER2-positive breast cancer patients with elevated SPP1⁺ TAMs levels experienced both reduced efficacy of trastuzumab neoadjuvant therapy and diminished long-term survival prospects. In summary, our findings provide the first systematic characterization of TME remodeling following trastuzumab therapy, identifying SPP1+ TAMs as a potential driver of trastuzumab resistance. This work advances our understanding of microenvironmental mechanisms underlying trastuzumab resistance and suggests new therapeutic strategies targeting TAM-mediated immunosuppression.

Targeted degradation of extracellular and membrane-associated proteins has emerged as a promising therapeutic modality. Here, we developed CD24-H7, a novel bispecific degrader that engages the transferrin receptor (TFRC) to mediate lysosomal degradation of CD24-an immunosuppressive protein commonly overexpressed in tumors. CD24-H7 consists of a TFRC-binding scFv and a CD24-specific scFv linked by a cathepsin-cleavable spacer, facilitating efficient internalization, lysosomal delivery, and subsequent recycling of TFRC. In vitro and in vivo experiments revealed potent and specific degradation of CD24, leading to marked suppression of tumor growth and enhanced antitumor immunity in humanized mouse glioblastoma (GBM) models. The degrader also exhibited a favorable safety profile with minimal on-target off-tumor toxicity. Moreover, combining CD24-H7 with anti-PD-1 antibodies synergistically promoted intratumoral CD8+ T cell infiltration and cytotoxicity while attenuating T cell exhaustion, resulting in significantly enhanced antitumor efficacy compared to monotherapy. These findings underscore the therapeutic potential of TFRC-recruiting degraders for selective targeting of membrane proteins and provide a compelling combinatorial approach to overcome immune evasion in oncology.

Glioblastoma (GBM) is the most common malignant brain tumor with a dismal prognosis (< 7

Checkpoint inhibitors have revolutionized hepatocellular carcinoma (HCC) treatment, yet their efficacy remains limited in advanced stages, with suboptimal objective response rates. Growth differentiation factor 15 (GDF15), a dual-functional cytokine implicated in tumor progression and immunosuppression, represents a promising therapeutic target. This study aims to develop a novel GDF15-targeted strategy to improve HCC management and synergize with PD-1 blockade. GFRAL-Fc fusion proteins were generated by fusing the extracellular domain of GFRAL with IgG1 Fc. The anti-tumor efficacy and the anti-cachexia ability of GFRAL-Fc was evaluated in a spontaneous HCC model on GDF15 humanized mice. Additionally, the half-life and drug safety were evaluated in mice. To investigate the underlying mechanisms, a CyTOF analysis was utilized to analysis the immunoregulation effects of GFRAL-Fc within HCC. Finally, the anti-tumor effects of GFRAL-Fc in combination with Programmed Death-1 (PD-1) inhibitors were assessed. GFRAL-Fc targets GDF15 to simultaneously prevent GDF15-CD48 interaction-driven ERK activation and block GDF15-GFRAL binding. Treatment with GFRAL-Fc achieved dual antitumor effects: reducing tumor progression and attenuating cancer-associated cachexia. Combination with PD-1 blockade further enhanced antitumor efficacy, resulting in a substantial decrease in tumor nodules. Mechanistic studies revealed that GFRAL-Fc reprograms the immunosuppressive tumor microenvironment by suppressing Treg activation while enhancing CD8+ T cell cytotoxicity. Our findings validate GDF15 targeting as a viable strategy to overcome checkpoint inhibitor resistance in HCC. The GFRAL-Fc fusion protein demonstrates multimodal therapeutic benefits through metabolic regulation and immune remodeling, providing a clinically translatable approach to optimize PD-1-based regimens. This study addresses critical gaps in current HCC management and warrants further clinical validation. Schematic diagram of GFRAL-Fc functional: After binding with GDF15, GFRAL-Fc inhibits the GDF15-GFRAL pathway to suppress cancer-associated cachexia, while also inhibiting the GDF15-CD48 pathway to reprograming the tumor immune microenvironment.

CDK4/6i, the first-line drug for treating ERα-positive breast cancer, significantly improves clinical outcomes. However, CDK4/6i resistance often develops and remains a major hurdle, and the underlying mechanisms remain challenging to fully investigate. Here, we used Genome-wide CRISPR/Cas9 library screening combined with single-cell sequencing to screen for molecules mediating CDK4/6i resistance and identified METTL14 as a determinant of CDK4/6i sensitivity. Clinical samples and datasets were analyzed and in vitro and in vivo experiments were performed to confirm the critical function of METTL14 in CDK4/6i resistance. Mechanistically, METTL14 can induce an increase in E2F1 expression in breast cancer cells via an m6A IGF2BP2-dependent mechanism and thus promote CDK4/6i resistance. Furthermore, through a small molecule screen, a novel METTL14 inhibitor named WKYMVM, which can restore sensitivity to CDK4/6i in CDK4/6i-resistant breast cancer cells, was identified. Treatment with folate-conjugated liposomes targeting breast cancer cells that contained both a CDK4/6i and WKYMVM revealed the synergistic effect of METTL14 inhibition with CDK4/6i therapy in a CDK4/6i-resistant PDX model. Together, our findings reveal the mechanism of CDK4/6i resistance and provide a strategy for overcoming CDK4/6i resistance via METTL14 inhibition.

In hepatocellular carcinoma (HCC), lenvatinib is a key first-line treatment that significantly improves survival in some patients with advanced stage. However, lenvatinib resistance presents a major clinical challenge. This study aims to identify key molecular factors driving lenvatinib resistance in HCC and propose intervention strategies to overcome this resistance, thereby enhancing therapeutic efficacy. A genome-wide CRISPR-Cas9 activation screen identified METTL8 as a crucial gene associated with lenvatinib resistance. Validation through in vitro and in vivo assays confirmed METTL8’s role in mediating lenvatinib resistance. Higher METTL8 expression was observed in lenvatinib-resistant HCC cells compared to parental cells. Immunohistochemical staining of tissue sections from HCC patients revealed a negative correlation between high METTL8 expression and lenvatinib sensitivity. To inhibit the function of METTL8 that mediate lenvatinib resistance, we conducted a screening using a natural compound library, virtual drug screening identified Rabdosiin as a potential METTL8 inhibitor, subsequent experiments demonstrated that Rabdosiin could effectively overcome METTL8-mediated lenvatinib resistance. In conclusion, this research highlights METTL8 as a novel target for mitigating lenvatinib resistance, proposing that targeting METTL8 could restore lenvatinib sensitivity in HCC, and underscores its value as a biomarker for lenvatinib application in clinical settings.

Leaky and structurally abnormal blood vessels and increased pressure in the tumor interstitium reduce the infiltration of CAR-T cells in solid tumors, including triple-negative breast cancer (TNBC). Furthermore, high burden of tumor cells may cause reduction of infiltrating CAR-T cells and their functional exhaustion. In this study, various effector-to-target (E:T) ratio experiments are established to model the treatment using CAR-T cells in leukemia (high E:T ratio) and solid tumor (low E:T ratio). It is found that the antitumor immune response is decreased in solid tumors with low E:T ratio. Furthermore, single cell sequencing is performed to investigate the functional exhaustion at a low ratio. It is revealed that the inhibition of mitophagy-mediated mitochondrial dysfunction diminished the antitumor efficacy of CAR-T-cell therapy. The mitophagy agonist BC1618 is screened via AI-deep learning and cytokine detection, in vivo and in vitro studies revealed that BC1618 significantly strengthened the antitumor response of CAR-T cells via improving mitophagy. Here, injection hydrogels are engineered for the controlled co-delivery of CAR-T cells and BC1618 that improves the treatment of TNBC. Local delivery of hydrogels creates an inflammatory and mitophagy-enhanced microenvironment at the tumor site, which stimulates the CAR-T cells proliferation, provides antitumor ability persistently, and improves the effect of treatment.

Despite the outstanding clinical success of immunotherapy, its therapeutic efficacy in glioblastoma (GBM) is still limited. To identify critical regulators of GBM immunity, we constructed a mouse single-guide RNA (sgRNA) library corresponding to all disease-related immune genes, and performed an in vivo CRISPR knockout (KO) screen in syngeneic GBM mouse models. We demonstrated that the deletion of GDF15 in GBM cells ameliorated the immunosuppressive tumor microenvironment (TME) and enhanced the antitumor efficacy of immune checkpoint blockade (ICB) response. Moreover, we designed unique nanoparticles for efficient encapsulation of CRISPR-Cas9, noninvasive brain delivery and tumor cell targeting, demonstrating an effective and safe strategy for GDF15 gene therapy. The CRISPR-Cas9 nanoparticles, known as ANPSS (Cas9/sgRNA), are easily created by enclosing a single Cas9/sgRNA complex in a polymer shell that is sensitive to glutathione. This shell also contains a dual-action ligand that aids in crossing the blood‒brain barrier, targeting tumor cells, and selectively releasing Cas9/sgRNA. Our encapsulating nanoparticles demonstrated promising GBM targeting, resulting in high GDF15 gene editing efficiency within brain tumors while showing minimal off-target gene editing in high-risk tissues. Treatment with ANPSS (Cas9/sgGDF15) effectively halted tumor growth, reversed immune suppression, and enhanced the efficacy of ICB therapy. These results emphasize the potential role of GDF15 in modulating the immune microenvironment and enhancing the effectiveness of current immunotherapy strategies for GBM. 1. In vivo CRISPR screens identify GDF15 as a critical driver of immune escape. 2. Synthesis of TME-responsive nanoparticles for GDF15 gene editing therapy. 3. GDF15 gene editing therapy enhances the antitumor efficacy of immune checkpoint blockade (ICB) response.

Cancer immunotherapy has demonstrated significant efficacy in various tumors, but its effectiveness in treating Hepatocellular Carcinoma (HCC) remains limited. Therefore, there is an urgent need to identify a new immunotherapy target and develop corresponding intervention strategies. Bioinformatics analysis has revealed that growth differentiation factor 15 (GDF15) is highly expressed in HCC and is closely related to poor prognosis of HCC patients. The previous study revealed that GDF15 can promote immunosuppression in the tumor microenvironment. Therefore, knocking out GDF15 through gene editing could potentially reverse the suppressive tumor immune microenvironment permanently. To deliver the CRISPR/Cas9 system specifically to HCC, nanocapsules (SNC) coated with HCC targeting peptides (SP94) on their surface is utilized. These nanocapsules incorporate disulfide bonds (SNCSS) that release their contents in the tumor microenvironment characterized by high levels of glutathione (GSH). In vivo, the SNCSS target HCC cells, exert a marked inhibitory effect on HCC progression, and promote HCC immunotherapy. Mechanistically, CyTOF analysis showed favorable changes in the immune microenvironment of HCC, immunocytes with killer function increased and immunocytes with inhibitive function decreased. These findings highlight the potential of the CRISPR-Cas9 gene editing system in modulating the immune microenvironment and improving the effectiveness of existing immunotherapy approaches for HCC.

Although dendritic cell (DC)-mediated immunotherapies are effective options for immunotherapy, traditional DC vaccines are hampered by a variety of drawbacks such as insufficient antigen delivery, weak lymph node homing, and the risk of living cell transfusion. To address the above-mentioned issues, we developed a personalized DC-mimicking nanovaccine (HybridDC) that enhances antigen presentation and elicits effective antitumor immunity. The biomimetic nanovaccine contains cell membranes derived from genetically engineered DCs, and several cellular components are simultaneously anchored onto these membranes, including CC-chemokine receptor 7 (CCR7), tumor-associated antigenic (TAA) peptide/tumor-derived exosome (TEX), and relevant costimulatory molecules. Compared with previous vaccines, the HybridDC vaccine showed an increased ability to target lymphoid tissues and reshape the immune landscape in the tumor milieu. HybridDC demonstrated significant therapeutic and prophylactic efficacy in poorly immunogenic, orthotopic models of glioma. Furthermore, the HybridDC vaccine potentiates the therapeutic efficacy of immune checkpoint blockade (ICB) therapy, providing a potential combination strategy to maximize the efficacy of ICB. Specifically, HybridDC can induce long-term protective immunity in memory T cells. Overall, the HybridDC vaccine is a promising platform for personalized cancer vaccines and may offer a combinational modality to improve current immunotherapy.

Cyclin-dependent kinase 4 (CDK4) and CDK6 inhibitors (CDK4/6i) have rapidly received Food and Drug Administration (FDA) approval as a new type of therapy for patients with advanced hormone receptor -positive breast cancer. However, with the widespread application of CDK4/6i, drug resistance has become a new challenge for clinical practice and has greatly limited the treatment effect. Here, the whole microenvironment landscape of ER+ breast cancer tumors was revealed through single -cell RNA sequencing, and a specific subset of cancer -associated fibroblasts (CD63+ CAFs) was identified as highly enriched in CDK4/6i resistant tumor tissues. Then, we found that CD63+ CAFs can distinctly promote resistance to CDK4/6i in breast cancer cells and tumor xenografts. In addition, it was discovered that miR-20 is markedly enriched in the CD63+ CAFs-derived exosomes, which are used to communicate with ER+ breast cancer cells, leading to CDK4/6i resistance. Furthermore, exosomal miR-20 could directly target the RB1 mRNA 3 ' UTR and negatively regulate RB1 expression to decrease CDK4/6i sensitivity in breast cancer cells. Most importantly, we designed and synthesized cRGD-miR-20 sponge nanoparticles and found that they can enhance the therapeutic effect of CDK4/6i in breast cancer. In summary, our findings reveal that CD63+ CAFs can promote CDK4/6i resistance via exosomal miR-20, which induces the downregulation of RB1 in breast cancer cells, and suggest that CD63+ CAFs may be a novel therapeutic target to enhance CDK4/6i sensitivity.

BACKGROUND & AIMS:The tumor microenvironment (TME) plays a crucial role in the limited efficacy of existing treatments for hepatocellular carcinoma (HCC), with tumor-associated endothelial cells (TECs) serving as fundamental TME components that substantially influence tumor progression and treatment efficacy. However, the precise roles and mechanisms of TECs in HCC remain inadequately understood. METHODS:We employed a multi-omics profiling strategy to investigate the single-cell and spatiotemporal evolution of TECs within the microenvironment of HCC tumors, showcasing varied responses to immunotherapy. Through an analysis of a clinical cohort of patients with HCC, we explored the correlation between TEC subpopulations and immunotherapy outcomes. The influence of TEC subsets on the immune microenvironment was confirmed through comprehensive in vitro and in vivo studies. To further explore the mechanisms of distinct TEC subpopulations in microenvironmental modulation and their impact on immunotherapy, we utilized TEC subset-specific knockout mouse models as well as humanized mouse models. RESULTS:In this study, we identified a new subset of CXCL12+ TECs that exert a crucial role in immune suppression within the HCC TME. Functionally, CXCL12+ TECs impede the differentiation of CD8+ naïve T cells into CD8+ cytotoxic T cells by secreting CXCL12. Furthermore, they attract myeloid-derived suppressor cells (MDSCs). A bispecific antibody was developed to target both CXCL12 and PD1 specifically, showing significant promise in bolstering anti-tumor immune responses and advancing HCC therapy. CONCLUSIONS:CXCL12+ TECs are pivotal in mediating immunosuppression within the HCC microenvironment and targeting CXCL12+ TECs presents a promising approach to augment the efficacy of immunotherapies in patients with HCC. IMPACT AND IMPLICATIONS:This investigation reveals a pivotal mechanism wherein CXCL12+ tumor-associated endothelial cells (TECs) emerge as crucial modulators of immune suppression in the tumor microenvironment of hepatocellular carcinoma (HCC). The discovery of CXCL12+ TECs as inhibitors of CD8+ naïve T cell activation and recruiters of myeloid-derived suppressor cells significantly advances our grasp of the dynamic between HCC and immune regulation. Moreover, the development and application of a bispecific antibody precisely targeting CXCL12 and PD1 has proven to enhance immune responses in a humanized mouse HCC model. This finding underscores a promising therapeutic direction for HCC, offering the potential to amplify the impact of current immunotherapies.

目的 观察敲低Geminin对人脑胶质瘤细胞放疗敏感性的影响.方法 利用GEPIA和CGGA数据库分析Geminin在脑胶质瘤组织中的表达及与生存期的相关性.实验分为两组:si-GL2组(对照组)和si-Gem组(干扰组),设计siRNA干扰序列(si-Gem)和阴性对照序列(si-GL2),分别转染脑胶质瘤细胞LN229和U87,qRT-PCR和Western blot检测转染效率及蛋白表达水平.流式细胞术检测碘化丙啶(PI)染色后DNA含量分布,进而分析敲低Geminin对DNA再复制的影响.通过Annexin V-FITC/PI法,用流式细胞术检测敲低Geminin对细胞凋亡的影响.CCK-8细胞增殖实验检测敲低Geminin对胶质瘤细胞LN229和U87放疗敏感性的影响.Western blot检测Geminin相关分子Cdt1和DNA相关损伤蛋白γ-H2AX的表达水平.结果脑胶质瘤组织中Geminin的表达水平高于癌旁正常组织,且Geminin表达越高,脑胶质瘤患者预后越差(P＜0.05).与si-GL2组相比,si-Gem组Geminin在LN229和U87细胞中的表达降低(P＜0.05).与si-GL2组相比,si-Gem组敲低Geminin可诱导脑胶质瘤细胞出现DNA再复制表型、细胞凋亡比例显著增加(P＜0.05),脑胶质瘤细胞对放疗敏感性增强(P＜0.05).与si-GL2组相比,si-Gem组Geminin相关分子Cdt1蛋白表达水平降低,γ-H2AX的蛋白表达水平升高.结论 敲低Geminin可以诱导脑胶质瘤细胞出现DNA再复制表型,造成DNA复制压力,并引起自发性细胞周期阻滞及细胞凋亡增多,进而增强脑胶质瘤细胞放疗敏感性,Geminin有望成为脑胶质瘤治疗的新靶点.

Although temozolomide (TMZ) provides significant clinical benefit for glioblastoma (GBM), responses are limited by the emergence of acquired resistance. Here, we demonstrate that exosomal circCABIN1 secreted from TMZ-resistant cells was packaged into exosomes and then disseminated TMZ resistance of receipt cells. CircCABIN1 could be cyclized by eukaryotic translation initiation factor 4A3 (EIF4A3) and is highly expressed in GBM tissues and glioma stem cells (GSCs). CircCABIN1 is required for the self-renewal maintenance of GSCs to initiate acquired resistance. Mechanistically, circCABIN1 regulated the expression of olfactomedin-like 3 (OLFML3) by sponging miR-637. Moreover, upregulation of OLFML3 activating the ErbB signaling pathway and ultimately contributing to stemness reprogramming and TMZ resistance. Treatment of GBM orthotopic mice xenografts with engineered exosomes targeting circCABIN1 and OLFML3 provided prominent targetability and had significantly improved antitumor activity of TMZ. In summary, our work proposed a novel mechanism for drug resistance transmission in GBM and provided evidence that engineered exosomes are a promising clinical tool for cancer prevention and therapy.

Body-protective compound (BPC) 157 demonstrates protective effects against damage to various organs and tissues. For future clinical applications, we had previously established a solid-phase synthesis process for BPC157, verified its biological activity in different wound models, and completed preclinical safety evaluations. This study aimed to investigate the pharmacokinetics, excretion, metabolism, and distribution profiles of BPC157. After a single intravenous (IV) administration, single intramuscular (IM) administrations at three doses in successive increments along with repeated IM administrations, the elimination half-life (t1/2) of prototype BPC157 was less than 30 min, and BPC157 showed linear pharmacokinetic characteristics in rats and beagle dogs at all doses. The mean absolute bioavailability of BPC157 following IM injection was approximately 14%–19% in rats and 45%–51% in beagle dogs. Using [3H]-labeled BPC157 and radioactivity examination, we proved that the main excretory pathways of BPC157 involved urine and bile. [3H]BPC157 was rapidly metabolized into a variety of small peptide fragments in vivo, thus forming single amino acids that entered normal amino acid metabolism and excretion pathways. In conclusion, this study provides the first analysis of the pharmacokinetics of BPC157, which will be helpful for its translation in the clinic.

Sorafenib is one of the few effective first-line drugs approved for the treatment of advanced hepatocellular carcinoma (HCC). However, the development of drug resistance is common among individuals with HCC. Recent evidence indicated that the anticancer activity of sorafenib mainly relies on the induction of ferroptosis. Furthermore, in our study, genes that suppress ferroptosis, especially GPX4 and DHODH, were enriched in sorafenib-resistant cells and primary tissues and were associated with poor prognosis of HCC patients who received sorafenib treatment. Therefore, a new ferroptosis inducer comprising a multiplex small interfering RNA (multi-siRNA) capable of simultaneously silencing GPX4 and DHODH was created. Then, exosomes with high multi-siRNA loading and HCC-specific targeting were established by fusing the SP94 peptide and the N-terminal RNA recognition motif (RRM) of U1-A with the exosomal membrane protein Lamp2b. The results from the in vitro and in vivo experiments indicate that this tumor-targeting nano-delivery system (ExoSP94-lamp2b-RRM-multi-siRNA) could enhance sorafenib-induced ferroptosis and overcome sorafenib resistance. Taken together, HCC-targeted exosomes (ExoSP94-Lamp2b-RRM) could specifically deliver multi-siRNA to HCC tissues, enhance sorafenib-induced ferroptosis by silencing GPX4 and DHODH expression and consequently increase HCC sensitivity to sorafenib, which opens a new avenue for clinically overcoming sorafenib resistance from the perspective of ferroptosis.