Tumor Microenvironment Research Articles

Trem2-expressing multinucleated giant macrophages are a biomarker of good prognosis in head and neck squamous cell carcinoma.

Patients with head and neck squamous cell carcinomas (HNSCC) often have poor outcomes due to suboptimal risk-management and treatment strategies; yet integrating novel prognostic biomarkers into clinical practice is challenging. Here, we report the presence of multinucleated giant cells (MGC) - a type of macrophages - in tumors from patients with HNSCC, which are associated with a favorable prognosis in treatment-naive and preoperative-chemotherapy-treated patients. Importantly, MGC density increased in tumors following preoperative therapy, suggesting a role of these cells in the anti-tumoral response. To enable clinical translation of MGC density as a prognostic marker, we developed a deep-learning model to automate its quantification on routinely stained pathological whole slide images. Finally, we used spatial transcriptomic and proteomic approaches to describe the MGC-related tumor microenvironment and observed an increase in central memory CD4 T cells. We defined an MGC-specific signature resembling to TREM2-expressing mononuclear tumor associated macrophages, which co-localized in keratin tumor niches.

Roles of the tumor microenvironment in the resistance to programmed cell death protein 1 inhibitors in patients with gastric cancer

Despite the continuous developments and advancements in the treatment of gastric cancer (GC), which is one of the most prevalent types of cancer in China, the overall survival is still poor for most patients with advanced GC. In recent years, with the progress in tumor immunology research, attention has shifted toward immunotherapy as a therapeutic approach for GC. Programmed cell death protein 1 (PD-1) inhibitors, as novel immunosuppressive medications, have been widely utilized in the treatment of GC. However, many patients are still resistant to PD-1 inhibitors and experience recurrence in the advanced stages of PD-1 immunotherapy. To reduce the occurrence of drug resistance and recurrence in GC patients receiving PD-1 immunotherapy, to maximize the clinical activity of immunosuppressive drugs, and to elicit a lasting immune response, it is essential to research the tumor microenvironment mechanisms leading to PD-1 inhibitor resistance in GC patients. This article reviews the progress in studying the factors influencing the resistance to PD-1 inhibitors in the GC tumor microenvironment, aiming to provide insights and a basis for reducing resistance to PD-1 inhibitors for GC patients in the future.

Multitask machine learning-based tumor-associated collagen signatures predict peritoneal recurrence and disease-free survival in gastric cancer.

Accurate prediction of peritoneal recurrence for gastric cancer (GC) is crucial in clinic. The collagen alterations in tumor microenvironment affect the migration and treatment response of cancer cells. Herein, we proposed multitask machine learning-based tumor-associated collagen signatures (TACS), which are composed of quantitative collagen features derived from multiphoton imaging, to simultaneously predict peritoneal recurrence (TACSPR) and disease-free survival (TACSDFS). Among 713 consecutive patients, with 275 in training cohort, 222 patients in internal validation cohort, and 216 patients in external validation cohort, we developed and validated a multitask machine learning model for simultaneously predicting peritoneal recurrence (TACSPR) and disease-free survival (TACSDFS). The accuracy of the model for prediction of peritoneal recurrence and prognosis as well as its association with adjuvant chemotherapy were evaluated. The TACSPR and TACSDFS were independently associated with peritoneal recurrence and disease-free survival in three cohorts, respectively (all P < 0.001). The TACSPR demonstrated a favorable performance for peritoneal recurrence in all three cohorts. In addition, the TACSDFS also showed a satisfactory accuracy for disease-free survival among included patients. For stage II and III diseases, adjuvant chemotherapy improved the survival of patients with low TACSPR and low TACSDFS, or high TACSPR and low TACSDFS, or low TACSPR and high TACSDFS, but had no impact on patients with high TACSPR and high TACSDFS. The multitask machine learning model allows accurate prediction of peritoneal recurrence and survival for GC and could distinguish patients who might benefit from adjuvant chemotherapy.

Research on curcumin mediating immunotherapy of colorectal cancer by regulating cancer associated fibroblasts.

The objective was to investigate curcumin's (Cur) function and associated molecular mechanisms in regulating tumor immunity in colon cancer. Primary cancer-associated fibroblasts (CAFs) from mouse CT26 colon cancer tumors were isolated. Validation of primary CAFs using immunofluorescence assay was done. Cell Counting Kit-8 experiments, real-time quantitative PCR (qPCR), and enzyme linked immunosorbent assay experiments were conducted to investigate how curcumin affected the growth and cytokine secretion functions of CAFs. The effect of curcumin on regulating PD-L1 expression on CT26 cells through CAFs in vitro was explored through coculture of CAFs and tumor cells, qPCR, and western blot experiments. A mouse colon cancer cell model was established in Balb/c nude mice to explore the effect of curcumin on colon tumor cells. Changes in the tumor microenvironment were detected by flow cytometry to explore the synergistic effect of curcumin combined with anti-PD-1 monoclonal antibody in the treatment of mouse colon cancer. In vitro, curcumin prevented the growth and TGF-β secretion of CT26 cells. At the same time, curcumin inhibited the secretion of TGF-β by CAFs, thereby downregulating the PD-L1 expression of CT26 cells. In vivo, curcumin combined with anti-PD-1 antibodies can further enhance the inhibitory effect of PD-1 antibodies on tumors and increase the number of tumor-suppressing immune cells in the tumor microenvironment, such as M1 macrophages and CD8 T cells, thus inhibiting tumors. Immune M2 macrophages, regulatory T cells, and other cells were reduced. In conclusion, curcumin reduces the expression of PD-L1 in colon cancer cells and improves the tumor immune microenvironment by inhibiting the proliferation of CAFs and the secretion of TGF-β. Curcumin and anti-PD-1 treatment have synergistic inhibitory effects on colon cancer.

Inflammation mediated by gut microbiome alterations promotes lung cancer development and an immunosuppressed tumor microenvironment.

Accumulating evidence indicates that the gut microbiome influences cancer progression and therapy. We recently showed that progressive changes in gut microbial diversity and composition are closely associated with tobacco-associated lung adenocarcinoma (LUAD) in a human-relevant mouse model. Furthermore, we demonstrated that the loss of the antimicrobial protein Lcn2 in these mice, exacerbates pro-tumor inflammatory phenotypes while further reducing microbial diversity. Yet, how gut microbiome alterations impinge on LUAD development remains poorly understood. Here, we investigated the role of gut microbiome changes in LUAD development using fecal microbiota transfer and delineated a pathway by which gut microbiome alterations incurred by loss of Lcn2 fostered the proliferation of pro-inflammatory bacteria of the genus Alistipes, triggering gut inflammation. This inflammation propagated systemically, exerting immunosuppression within the tumor microenvironment, augmenting tumor growth through an IL-6-dependent mechanism and dampening response to immunotherapy. Corroborating our preclinical findings, we found that patients with LUAD with a higher relative abundance of Alistipes species in the gut showed diminished response to neoadjuvant immunotherapy. These insights reveal the role of microbiome-induced inflammation in LUAD and present new potential targets for interception and therapy.

Targeting GProtein-Coupled Receptors in Immuno-Oncological Therapies.

The advent of cancer immunotherapy based on PD-1 and CTLA-4 immune checkpoint blockade (ICB) has revolutionized cancer treatment. However, many cancers do not respond to ICB, highlighting the urgent need for additional approaches to achieve durable cancer remission. The large family of G protein-coupled receptors (GPCRs) is the target of more than 30% of all approved drugs, but GPCRs have been underexploited in cancer immunotherapy. In this review, we discuss the central role of GPCRs in immune cell migration and function and describe how single-cell transcriptomic studies are illuminating the complexity of the human tumor immune GPCRome. These receptors include multiple GPCRs expressed in CD8 Tcells that are activated by inflammatory mediators, protons, neurotransmitters, and metabolites that accumulate in the tumor microenvironment, thereby promoting Tcell dysfunction. We also discuss new opportunities to target GPCRs as a multimodal approach to enhance the response to ICB for a myriad of human malignancies.

A necroptosis-regulated model from single-cell analysis that predicts survival and identifies the Pivotal role of MAGEA6 in hepatocellular carcinoma

ObjectiveHepatocellular carcinoma (HCC) ranks as the third leading cause of cancer-related deaths, constituting 75%–85 % of all primary liver cancers. The objective of this study was to develop a necroptosis-related gene signature using single-cell and bulk RNA sequencing to predict HCC patient prognoses. MethodsA total of 25 key necroptosis regulators were identified from previous literature. We evaluated the necroptosis scores of different cell types using single-cell sequencing data from HCC and analyzed 168 necroptosis-related genes. The Cancer Genome Atlas Liver Hepatocellular Carcinoma (TCGA-LIHC) dataset served as the training set for establishing a novel necroptosis-related gene risk signature, employing univariate and multivariate Cox regression analyses. Additionally, the study examined the model's relevance in immunity and immunotherapy, and predicted chemosensitivity in HCC patients based on the gene signature. The key genes were validated by the biological experiments. ResultsCompared to other cell types, hepatoma cells displayed the lowest necroptosis scores. A new six-gene necroptosis-related signature (S100A11, MAGEC2, MAGEA6, CTP2C9, SOX4, AKR1B10) was developed using the TCGA database and validated in the ICGC database. Patients in the high-risk category had poorer prognoses, with the risk score serving as an independent prognostic indicator beyond other clinical factors. These high-risk patients also exhibited greater immune infiltration but demonstrated a weaker anti-tumor response due to elevated expression of immune checkpoints. Pathways involving hypoxia, glycolysis, and P53, as well as the frequency of P53 somatic mutations, were notably heightened in the high-risk group. Additionally, the six genes in the model showed significantly different mRNA expression in hepatoma cell lines compared to normal hepatocytes, with the role of MAGEA6 in liver cancer being elucidated through critical experiments. ConclusionsThis study successfully developed a six-gene necroptosis-related signature to predict prognoses in HCC patients. It further explored the roles of necroptosis in hepatoma cells and the tumor microenvironment.

The Dark Knight: Functional Reprogramming of Neutrophils in the Pathogenesis of Colitis-Associated Cancer.

Neutrophils are the primary myeloid cells that are recruited to inflamed tissues, and they are key players during colitis, being also present within the tumor microenvironment during the initiation and growth of colon cancer. Neutrophils fundamentally serve to protect the host against microorganism invasion, but during cancer development, they can become protumoral and lead to tumor initiation, growth, and eventually, metastasis-hence, playing a dichotomic role for the host. Protumoral neutrophils in cancer patients can be immunosuppressive and serve as markers for disease progression but their characteristics are not fully defined. In this review, we explore the current knowledge on how neutrophils in the gut fluctuate between an inflammatory or immunosuppressive state and how they contribute to tumor development. We describe neutrophils' antitumoral and protumoral effects during inflammatory bowel diseases and highlight their capacity to provoke the advent of inflammation-driven colorectal cancer. We present the functional ambivalence of the neutrophil populations within the colon tumor microenvironment, which can be potentially exploited to establish therapies that will prevent, or even reverse, inflammation-dependent colon cancer incidence in high-risk patients.

Focal adhesion kinase-mediated interaction between tumor and immune cells in the tumor microenvironment: implications for cancer-associated therapies and tumor progression.

Focal adhesion kinase (FAK) expression has been linked to tumor growth, immunosuppression, metastasis, angiogenesis, and therapeutic resistance through kinase-dependent and kinase scaffolding functions in the nucleus and cytoplasm. Hence, targeting FAK alone or with other agents has gained attention as a potential therapeutic strategy. Moreover, mounting evidence shows that FAK activity can influence the tumor immune microenvironment crosstalk to support tumor progression. Recently, tumor immune microenvironment interaction orchestrators have shown to be promising therapeutic agents for cancer immunotherapies. Therefore, this review highlights how FAK regulates the tumor immune microenvironment interplay to promote tumor immune evasive mechanisms and their potential for combination therapies with standard cancer treatments.

Androgen Deprivation Therapy Drives a Distinct Immune Phenotype in Localized Prostate Cancer.

Androgen deprivation therapy (ADT) remains the backbone of prostate cancer treatment. Beyond suppression of testosterone and tumor cell growth, emerging evidence suggests ADT also modulates the immune tumor microenvironment (TME). However, a more precise understanding of the timing and intricacies of these immunological shifts is needed. Here we analyzed 49 primary prostate cancers, comparing those surgically removed either without treatment or following treatment with degarelix at 4, 7, and 14 days pre-surgery. Utilizing next-generation DNA and RNA sequencing, and multiplexed immunofluorescence, we examined alterations in immune phenotypes in the presence or absence of ADT. Our findings reveal that ADT rapidly transforms the typically bland prostate TME into an inflamed environment within days. Notably, we observed an increase in activated CD8 T-cells along with an increase in suppressive regulatory T-cells (Tregs). We also found an expansion of the myeloid compartment, particularly pro-inflammatory M1-like tumor-associated macrophages. Intriguingly, discernable changes which have not previously been described also occurred in tumor cells, including upregulation of antigen presentation by MHC class I and II and, unexpectedly, a decrease in the "don't eat me" signal CD47. These observations underscore the critical role of timing and disease context in order to optimize the therapeutic efficacy of immune modulators combined with androgen ablation, for which the presurgical neoadjuvant setting may be ideal. Our findings warrant future prospective validation, which is currently underway.

Mitochondria-Targeting AIEgens as Pyroptosis Inducers for Boosting Type-I Photodynamic Therapy of Tongue Squamous Cell Carcinoma.

The development of a photosensitizer (PS) that induces pyroptosis could be a star for photodynamic therapy (PDT), particularly with type-I PSs that produce reactive oxygen species (ROS) in a hypoxic tumor microenvironment. Since pyroptosis is a recently characterized cell death pathway, it holds promise for advancing PDT in oncology, with PSs playing a critical role. Herein, we develop a PS named Th-M with aggregation-induced emission (AIE) characteristics for type-I PDT against tongue squamous cell carcinoma (TSCC). Th-M stands out for its exceptional mitochondrial-targeting ability, which triggers mitochondrial dysfunction and leads to Caspase-3 and Gasdermin E (GSDME) cleavage under white light irradiation, inducing pyroptosis in TSCC cells. Our studies verify the effectiveness of Th-M in destroying cancer cells in vitro and suppressing tumor growth in vivo while also demonstrating a favorable biosafety profile. This work pioneers the application of Th-M as a mitochondria-targeted, type-I PS that leverages the mechanism of pyroptosis, offering a potent approach for the treatment of TSSC with promising implications for future PDT of cancers.

Liquid-liquid phase separation-related genes associated with prognosis, tumor microenvironment characteristics, and tumor cell features in bladder cancer.

This study aimed to explore the Liquid-liquid phase separation (LLPS)-related genes associated with the prognosis of bladder cancer (BCa) and assess the potential application of LLPS-related prognostic signature for predicting prognosis in BCa patients. Clinical information and transcriptome data of BCa patients were extracted from the Cancer Genome Atlas-BLCA (TCGA-BLCA) database and the GSE13507 database. Furthermore, 108 BCa patients who received treatment at our institution were subjected to a retrospective analysis. The least absolute shrinkage and selection operator (LASSO) analysis was performed to develop an LLPS-related prognostic signature for BCa. The CCK8, wound healing and Transwell assays were performed. Based on 62 differentially expressed LLPS-related genes (DELRGs), three DELRGs were screened by LASSO analysis including kallikrein-related peptidase 5 (KLK5), monoacylglycerol O-acyltransferase 2 (MOGAT2) and S100 calcium-binding protein A7 (S100A7). Based on three DELRGs, a novel LLPS-related prognostic signature was constructed for individualized prognosis assessment. Kaplan-Meier curve analyses showed that LLPS-related prognostic signature was significantly correlated with overall survival (OS) of BCa. ROC analyses demonstrated the LLPS-related prognostic signature performed well in predicting the prognosis of BCa patients in the training group (the area under the curve (AUC) = 0.733), which was externally verified in the validation cohort 1 (AUC = 0.794) and validation cohort 2 (AUC = 0.766). Further experiments demonstrated that inhibiting KLK5 could affect the proliferation, migration, and invasion of BCa cells. In this study, a novel LLPS-related prognostic signature was successfully developed and validated, demonstrating strong performance in predicting the prognosis of BCa patients.

4-Hydroxynonenal Promotes Colorectal Cancer Progression through Regulating Cancer Stem Cell Fate.

Tumor microenvironment (TME) plays a crucial role in sustaining cancer stem cells (CSCs). 4-hydroxynonenal (4-HNE) is abundantly present in the TME of colorectal cancer (CRC). However, the contribution of 4-HNE to CSCs and cancer progression remains unclear. This study aimed to investigate the impact of 4-HNE on the regulation of CSC fate and tumor progression. Human CRC cells were exposed to 4-HNE, and CSC signaling was analyzed using quantitative real-time PCR, immunofluorescent staining, fluorescence-activated cell sorting, and bioinformatic analysis. Tumor-promoting role of 4-HNE was confirmed using a xenograft model. Exposure of CRC cells to 4-HNE activated non-canonical Hedgehog (HH) signaling and homologous recombination repair (HRR) pathways in LGR5+ CSCs. Furthermore, blocking HH signaling led to a significant increase in the expression of γH2AX, indicating that 4-HNE induces double-stranded DNA breaks (DSBs) and simultaneously activates HH signaling to protect CSCs from 4-HNE-induced damage via the HRR pathway. Additionally, 4-HNE treatment increased the population of LGR5+ CSCs and promoted asymmetric division in these cells, leading to enhanced self-renewal and differentiation. Notably, 4-HNE also promoted xenograft tumor growth and activated CSC signaling in vivo. These findings demonstrate that 4-HNE, as a signaling inducer in the TME, activates the non-canonical HH pathway to shield CSCs from oxidative damage, enhances the proliferation and asymmetric division of LGR5+ CSCs, and thereby facilitates tumor growth. These novel insights shed light on the regulation of CSC fate within the oxidative TME, offering potential implications for understanding and targeting CSCs for CRC therapy.

Mathematical modeling of 18F-Fluoromisonidazole (18F-FMISO) radiopharmaceutical transport in vascularized solid tumors

18F-Fluoromisonidazole (18F-FMISO) is a highly promising positron emission tomography radiopharmaceutical for identifying hypoxic regions in solid tumors. This research employs spatiotemporal multi-scale mathematical modeling to explore how different levels of angiogenesis influence the transport of radiopharmaceuticals within tumors. In this study, two tumor geometries with heterogeneous and uniform distributions of capillary networks were employed to incorporate varying degrees of microvascular density. The synthetic image of the heterogeneous and vascularized tumor was generated by simulating the angiogenesis process. The proposed multi-scale spatiotemporal model accounts for intricate physiological and biochemical factors within the tumor microenvironment, such as the transvascular transport of the radiopharmaceutical agent, its movement into the interstitial space by diffusion and convection mechanisms, and ultimately its uptake by tumor cells. Results showed that both quantitative and semi-quantitative metrics of 18F-FMISO uptake differ spatially and temporally at different stages during tumor growth. The presence of a high microvascular density in uniformly vascularized tumor increases cellular uptake, as it allows for more efficient release and rapid distribution of radiopharmaceutical molecules. This results in enhanced uptake compared to the heterogeneous vascularized tumor. In both heterogeneous and uniform distribution of microvessels in tumors, the diffusion transport mechanism has a more pronounced than convection. The findings of this study shed light on the transport phenomena behind 18F-FMISO radiopharmaceutical distribution and its delivery in the tumor microenvironment, aiding oncologists in their routine decision-making processes.

Optimizing the Therapeutic Index of sdAb-Based Radiopharmaceuticals Using Pretargeting.

Single-domain antibodies (sdAbs) demonstrate favorable pharmacokinetic profiles for molecular imaging applications. However, their renal excretion and retention are obstacles for applications in targeted radionuclide therapy (TRT). Methods: Using a click-chemistry-based pretargeting approach, we aimed to reduce kidney retention of a fibroblast activation protein α (FAP)-targeted sdAb, 4AH29, for 177Lu-TRT. Key pretargeting parameters (sdAb-injected mass and lag time) were optimized in healthy mice and U87MG (FAP+) xenografts. A TRT study in a pancreatic ductal adenocarcinoma (PDAC) patient-derived xenograft (PDX) model was performed as a pilot study for sdAb-based pretargeting applications. Results: Modification of 4AH29 with trans-cyclooctene (TCO) moieties did not modify the sdAb pharmacokinetic profile. A 200-µg injected mass of 4AH29-TCO and an 8-h lag time for the injection of [177Lu]Lu-DOTA-PEG7-tetrazine resulted in the highest kidney therapeutic index (2.0 ± 0.4), which was 5-fold higher than that of [177Lu]Lu-DOTA-4AH29 (0.4 ± 0.1). FAP expression in the tumor microenvironment was validated in a PDAC PDX model with both immunohistochemistry and PET/CT imaging. Mice treated with the pretargeting high-activity approach (4AH29-TCO + [177Lu]Lu-DOTA-PEG7-tetrazine; 3 × 88 MBq, 1 injection per week for 3 wk) demonstrated prolonged survival compared with the vehicle control and conventionally treated ([177Lu]Lu-DOTA-4AH29; 3 × 37 MBq, 1 injection per week for 3 wk) mice. Mesangial expansion was reported in 7 of 10 mice in the conventional cohort, suggesting treatment-related kidney morphologic changes, but was not observed in the pretargeting cohort. Conclusion: This study validates pretargeting to mitigate sdAbs' kidney retention with no observation of morphologic changes on therapy regimen at early time points. Clinical translation of click-chemistry-based pre-TRT is warranted on the basis of its ability to alleviate toxicities related to biovectors' intrinsic pharmacokinetic profiles. The absence of representative animal models with extensive stroma and high FAP expression on cancer-associated fibroblasts led to a low mean tumor-absorbed dose even with high injected activity and consequently to modest survival benefit in this PDAC PDX.

Photoferroptosis as a potent strategy for neuroblastoma treatment

Photodynamic therapy (PDT) and photothermal therapy (PTT) have been developed to treat tumors with potential of clinical applications due to their high spatiotemporal selectivity and non-invasiveness. Nevertheless, the hypoxia within the tumor microenvironment (TME) limits the efficacy of PDT. PTT has the risk of damaging surrounding normal tissues due to the high temperatures essential for killing tumor cells. Herein, we propose a new tumor treatment strategy based on photo-triggered ferroptosis of tumor cells, which is termed photoferroptosis therapy (PFT). The PFT agent (CuS&AIPH@PEG-PAE@PM) was synthesized by encapsulating a radical generator (2,2’-azobis[2-(2-imidazolin-2-yl)propane] dihydrochloride, AIPH) and a photothermal agent (copper sulfide, CuS) into an amphiphilic polymer (poly(ethylene glycol)-poly(β-amino ester), PEG-PAE) via self-assembly and a following coating with platelet membrane (PM). Under near-infrared (NIR) irradiation, the PFT agent CuS&AIPH@PEG-PAE@PM generates abundant alkyl radicals (R●) to trigger tumor cell ferroptosis in a moderate temperature and oxygen-independent manner. Meanwhile, the PFT agent also reduces the GSH level and thus suppresses GPX4 expression to promote ferroptosis, which further consolidates the antitumor effect of PFT. The PFT is expected to establish a promising phototherapy strategy against tumors, which has the potential to overcome the limitations of PDT and PTT.

Distinct tumor architectures and microenvironments for the initiation of breast cancer metastasis in the brain

Brain metastasis, a serious complication of cancer, hinges on the initial survival, microenvironment adaptation, and outgrowth of disseminated cancer cells. To understand the early stages of brain colonization, we investigated two prevalent sources of cerebral relapse, triple-negative (TNBC) and HER2+ (HER2BC) breast cancers. Using mouse models and human tissue samples, we found that these tumor types colonize the brain, with a preference for distinctive tumor architectures, stromal interfaces, and autocrine programs. TNBC models tend to form perivascular sheaths with diffusive contact with astrocytes and microglia. In contrast, HER2BC models tend to form compact spheroids driven by autonomous tenascin C production, segregating stromal cells to the periphery. Single-cell transcriptomics of the tumor microenvironment revealed that these architectures evoke differential Alzheimer's disease-associated microglia (DAM) responses and engagement of the GAS6 receptor AXL. The spatial features of the two modes of brain colonization have relevance for leveraging the stroma to treat brain metastasis.

Circumventing resistance within the Ewing sarcoma microenvironment by combinatorial innate immunotherapy

BackgroundPediatric patients with recurrent/metastatic Ewing sarcoma (ES) have a dismal 5-year survival. Novel therapeutic approaches are desperately needed. Natural killer (NK) cell number and function are low in ES patient...

Open-Source Throttling of CD8+ T Cells in Brain with Low-Intensity Focused Ultrasound-Guided Sequential Delivery of CXCL10, IL-2, and aPD-L1 for Glioblastoma Immunotherapy.

Improving clinical immunotherapy for glioblastoma (GBM) relies on addressing the immunosuppressive tumor microenvironment (TME). Enhancing CD8+ T cell infiltration and preventing its exhaustion holds promise for effective GBM immunotherapy. Here, a low-intensity focused ultrasound (LIFU)-guided sequential delivery strategy is developed to enhance CD8+ T cells infiltration and activity in the GBM region. The sequential delivery of CXC chemokine ligand 10 (CXCL10) to recruit CD8+ T cells and interleukin-2 (IL-2) to reduce their exhaustion is termed an "open-source throttling" strategy. Consequently, up to 3.39-fold of CD8+ T cells are observed with LIFU-guided sequential delivery of CXCL10, IL-2, and anti-programmed cell death 1 ligand 1 (aPD-L1), compared to the free aPD-L1 group. The immune checkpoint inhibitors (ICIs) therapeutic efficacy is substantially enhanced by the reversed immunosuppressive TME due to the expansion of CD8+ T cells, resulting in the elimination of tumor, prolonged survival time, and long-term immune memory establishment in orthotopic GBM mice. Overall, LIFU-guided sequential cytokine and ICIs delivery offers an "open-source throttling" strategy of CD8+ T cells, which may present a promising strategy for brain-tumor immunotherapy.

TAK1 Promotes an Immunosuppressive Tumor Microenvironment through Cancer-Associated Fibroblast Phenotypic Conversion in Pancreatic Ductal Adenocarcinoma.

We aim to clarify the precise function of Transformed growth factor-beta 1 activated kinase-1 (TAK1) in cancer-associated fibroblasts (CAFs) within human pancreatic ductal adenocarcinoma (PDAC) by investigating its role in cytokine-mediated signaling pathways. The expression of TAK1 in pancreatic cancer was confirmed by TCGA data and human pancreatic cancer specimens. CAFs from freshly resected PDAC specimens were cultured and used in a three-dimensional model for direct and indirect co-culture with PDAC tumors to investigate TAK1 function. Additionally, organoids from KPC (LSL-K-RasLSLG12D/+; LSL-p53R172H/+; Pdx1-Cre) mice were mixed with CAFs and injected subcutaneously into C57BL/6 mice to explore in vivo functional interactions of TAK1. TCGA data revealed significant upregulation of TAK1 in PDAC, associating with a positive correlation with the T-cell exhaustion signature. Knockdown of TAK1 in CAFs decreased the iCAF signature and increased the myCAF signature both in vitro and in vivo. The absence of TAK1 hindered CAF proliferation, blocked several inflammatory factors via multiple pathways associated with immunosuppression, and hindered EMT, outgrowth in vitro in spheroid co-cultures with PDAC cells. Additionally, TAK1 inhibitor restrained tumor growth, increased CD4+ and CD8+ T cell abundance, and reduced immunosuppressive cells present in vivo. Blocking the TAK1+CAF phenotype leads to the conversion of protumorigenic CAFs to antitumorigenic CAFs. This highlights TAK1 as a potential therapeutic target, particularly in CAFs, and represents a novel avenue for combined immunotherapy in PDAC.