Infiltration Of Effector Cells Research Articles

Identification of Cellular Senescence-Related Critical Genes and Molecular Classification and Revealing the Drug-Resistant Therapeutic Effect of IGFBP2 in Chronic Myeloid Leukemia.

The occurrence and development of hematologic tumors are closely linked to cellular senescence. However, the molecular characteristics associated with this phenomenon in chronic myeloid leukemia (CML) have not been thoroughly investigated. The cellular senescence score was calculated using gene set variation analysis. Consensus clustering algorithm was used to identify the molecular subtypes associated with cellular senescence. Clinical samples were collected for sequencing analysis to verify the expression of critical cellular senescence-related genes (CSRG). The effect of targeted inhibition of IGFBP2 on the malignant phenotype of CML-resistant cells was studied by cell experiments. The cellular senescence score in CML samples was significantly lower compared to normal samples. Higher expression of immune checkpoint markers correlated with increased cellular senescence scores. We identified two distinct molecular subtypes (C1 and C2) related to cellular senescence. The C1 subtype exhibited enhanced metabolic function and DNA damage repair capacity, while the C2 subtype showed higher infiltration of immune effector cells and activity in immune-related signaling pathways. We also discovered a group of drugs that displayed significant sensitivity differences between these two molecular subtypes, with the C2 subtype showing greater responsiveness to immunotherapy. Four critical cellular senescence-related genes (CSRGs), namely IGFBP2, IL7R, PLAU, and SUN1 demonstrated high diagnostic value for CML. We validated the expression levels of these four genes using clinical samples and confirmed through cell experiments that targeted inhibition of IGFBP2 effectively suppressed proliferation of resistant CML cells, promoted apoptosis, and enhanced therapeutic sensitivity to imatinib. Our study conducted a comprehensive analysis on CSRG expression characteristics in CML and explored potential correlations between cellular senescence and immune function. The identification of molecular subtypes provides valuable insights into assessing individual patients' biological characteristics for guiding clinical treatment decisions. Additionally, IGFBP2has emerged as a promising therapeutic target for therapy-resistant cases of CML.

Interleukin-33 Deficiency Protects the Skin From Ulcer Formation in an Ischemia-Reperfusion-Induced Decubitus Mouse Model.

Interleukin-33 (IL-33) is an alarmin released upon epithelial tissue damage. It functions as a nuclear factor for regulating gene expression. We hypothesised that IL-33 is involved in the formation of decubitus ulcers through damaged epidermis. Therefore, this study aimed to clarify the mechanism of IL-33 action in decubitus ulcer formation. IL-33 knockout (KO), soluble stimulation-2 (ST2) transgenic, and wild-type (WT) mice were used to construct an ischemia-reperfusion (I/R) injury as a decubitus model. The ulcer area was significantly reduced in IL-33 KO mice compared to WT mice but was not reduced in ST2 transgenic mice. Anti-IL-33 receptor (transmembrane ST2) antibodies effectively prevented ulcer formation; however, an anti-IL-33 neutralising antibody was ineffective. The number of infiltrating macrophages was higher, while that of neutrophils and mast cells was lower in IL-33 KO mice than in WT mice. The number of M2 macrophages increased in IL-33 KO mice. Characterisation of gene expression levels revealed significantly reduced interleukin-1 beta (IL-1β) and increased C-C motif chemokine ligand 17 expression in IL-33 KO mice. Macrophages isolated from ulcers in WT or IL-33 KO mice stimulated with exogenous IL-33 produced comparable amounts of IL-1β. In conclusion, our study indicates that IL-33 is released in response to I/R injury in the skin, contributing to inflammatory macrophage and mast cell infiltration and stimulation, resulting in IL-1β production and the massive infiltration of effector cells, including neutrophils, which finally induces decubitus ulcer formation. These results suggest that suppressing IL-33 expression could be beneficial for treating early-phase decubitus ulcers.

Abstract B045: Sensitizing the PDAC tumor microenvironment to immune checkpoint therapies: characterization of a PDAC 3D model to decipher immune infiltration

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal solid tumor with an unfavorable prognosis, often resistant to conventional treatments due to its dense stroma rich in Cancer- Associated Fibroblasts (CAFs). Limited T cell infiltration and prevalence of immunosuppressive cells within the tumor hinder the efficacy of immune checkpoint inhibitors (ICIs). The objective of our project is to modify immune landscape of PDAC tumor microenvironment (TME) to induce ICIs sensitization. We thus develop novel therapeutic strategies combining an immunomodulatory cytokine Interleukine-15 (IL15), with conventional chemotherapies (Gemcitabine or Folfirinox). To accurately replicate the complex cancer-stroma interactions within the pancreatic tumor microenvironment, we constructed a three-dimensional in vitro heterospheroid model composed of xenograft-derived tumor cells from PDAC patients, both primary or immortalized CAFs, and peripheral blood mononuclear cells (PBMCs) from healthy donors. Human 3D models were first set up and characterized by cytometry, imaging mass spectrometry and immunohistochemistry. We showed that, CAFs produce components of the extracellular matrix, promote tumor cell growth, improve resistance to chemotherapy and are able to down-regulate immune cell infiltration and modulate the nature of infiltrated immune cells. Spatial analysis of spheroids using imaging mass cytometry show that CAF influences the distribution of immune cells in our models. Interestingly, Interleukine 15 treatment prompted robust infiltration of PBMCs into heterospheroids. Immunophenotyping experiments revealed a substantial shift in the nature of immune infiltration, marked by a pronounced increase in CD4+ and CD8+ T lymphocytes, gDelta T cell and NK cell populations. When combined with Gemcitabine and IL15, immune effector cell infiltration is remarkably increased, resulting in antitumoral effects and control of tumor cell growth. Moreover, therapeutic combinations induce activation of infiltrated immune effector cells with an increase of activation, degranulation and cytotoxic markers, such as IFNγ, CD107a, and Granzyme B respectively. Thus, the heterotypic spheroids described in our study are a suitable in vitro model to both characterize the influence of CAF on therapeutic effects and the mechanisms that drives immune suppressive microenvironment. Moving forward, the next steps will consist to conduct in vivo experiments using a syngeneic PDAC orthotopic model to assess whether this combined therapeutic approach sensitize the PDAC microenvironment to anti-PD1 therapies. Citation Format: Thomas Bessede, Clara Freixinos, Jennifer Barrat, Véronique Garambois, Nadia Vie, Henri-Alexandre Michaud, Julien Faget, Nathalie Bonnefoy, Céline Gongora, Bruno Robert, Laurent Gros, Christel Larbouret. Sensitizing the PDAC tumor microenvironment to immune checkpoint therapies: characterization of a PDAC 3D model to decipher immune infiltration [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B045.

Abstract B046: Sensitizing the PDAC tumor microenvironment to immune checkpoint therapies: characterization of a PDAC 3D model to decipher immune infiltration

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal solid tumor with an unfavorable prognosis, often resistant to conventional treatments due to its dense stroma rich in Cancer- Associated Fibroblasts (CAFs). Limited T cell infiltration and prevalence of immunosuppressive cells within the tumor hinder the efficacy of immune checkpoint inhibitors (ICIs). The objective of our project is to modify immune landscape of PDAC tumor microenvironment (TME) to induce ICIs sensitization. We thus develop novel therapeutic strategies combining an immunomodulatory cytokine Interleukine-15 (IL15), with conventional chemotherapies (Gemcitabine or Folfirinox). To accurately replicate the complex cancer-stroma interactions within the pancreatic tumor microenvironment, we constructed a three-dimensional in vitro heterospheroid model composed of xenograft-derived tumor cells from PDAC patients, both primary or immortalized CAFs, and peripheral blood mononuclear cells (PBMCs) from healthy donors. Human 3D models were first set up and characterized by cytometry, imaging mass spectrometry and immunohistochemistry. We showed that, CAFs produce components of the extracellular matrix, promote tumor cell growth, improve resistance to chemotherapy and are able to down-regulate immune cell infiltration and modulate the nature of infiltrated immune cells. Spatial analysis of spheroids using imaging mass cytometry show that CAF influences the distribution of immune cells in our models. Interestingly, Interleukine 15 treatment prompted robust infiltration of PBMCs into heterospheroids. Immunophenotyping experiments revealed a substantial shift in the nature of immune infiltration, marked by a pronounced increase in CD4+ and CD8+ T lymphocytes, gDelta T cell and NK cell populations. When combined with Gemcitabine and IL15, immune effector cell infiltration is remarkably increased, resulting in antitumoral effects and control of tumor cell growth. Moreover, therapeutic combinations induce activation of infiltrated immune effector cells with an increase of activation, degranulation and cytotoxic markers, such as IFNγ, CD107a, and Granzyme B respectively. Thus, the heterotypic spheroids described in our study are a suitable in vitro model to both characterize the influence of CAF on therapeutic effects and the mechanisms that drives immune suppressive microenvironment. Moving forward, the next steps will consist to conduct in vivo experiments using a syngeneic PDAC orthotopic model to assess whether this combined therapeutic approach sensitize the PDAC microenvironment to anti-PD1 therapies. Citation Format: Thomas Bessede, Clara Freixinos, Jennifer Barrat, Véronique Garambois, Nadia Vie, Henri-Alexandre Michaud, Julien Faget, Nathalie Bonnefoy, Céline Gongora, Bruno Robert, Laurent Gros, Christel Larbouret. Sensitizing the PDAC tumor microenvironment to immune checkpoint therapies: characterization of a PDAC 3D model to decipher immune infiltration [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B046.

Serine Depletion Promotes Antitumor Immunity by Activating Mitochondrial DNA-Mediated cGAS-STING Signaling.

Serine is critical for supporting cancer metabolism, and depriving malignant cells of this nonessential amino acid exerts antineoplastic effects, in large part, through disrupting metabolic pathways. Given the intricate relationship between cancer metabolism and the immune system, the metabolic defects imposed by serine deprivation might impact tumor-targeting immunity. In this study, we demonstrated that restricting endogenous and exogenous sources of serine in colorectal cancer cells results in mitochondrial dysfunction, leading to mitochondrial DNA (mtDNA) accumulation in the cytosol and consequent cGAS-STING1-driven type I IFN secretion. Depleting mtDNA or blocking its release attenuated cGAS-STING1 activation during serine deprivation. In vivo studies revealed that serine deprivation limits tumor growth, accompanied by enhanced type I IFN signaling and intratumoral infiltration of immune effector cells. Notably, the tumor-suppressive and immune-enhancing effects of serine restriction were impaired by T-cell depletion and IFN receptor blockade. Moreover, disrupting cGAS-STING1 signaling in colorectal cancer cells limited the immunostimulatory and tumor-suppressive effects of serine deprivation. Lastly, serine depletion increased the sensitivity of tumors to an immune checkpoint inhibitor targeting PD-1. Taken together, these findings reveal a role for serine as a suppressor of antitumor immunity, suggesting that serine deprivation may be employed to enhance tumor immunogenicity and improve responsiveness to immune checkpoint inhibitors. Significance: Depriving cancer cells of serine provokes mitochondrial perturbations that induce cytosolic mitochondrial DNA accumulation and subsequent activation of cGAS-STING signaling, stimulating tumor-targeting immune responses that can be enhanced with PD-1 targeted therapy. See related commentary by Borges and Garg, p. 2569.

Immunogenic Material Vaccine for Cancer Immunotherapy by Structure-Dependent Immune Cell Trafficking and Modulation.

Inherently immunogenic materials offer enormous prospects in enhancing vaccine efficacy. However, the understanding and improving material adjuvanticity remain elusive. Herein how the structural presentation of immunopotentiators in a material governs the dynamic dialogue between innate and adaptive immunity for enhanced cancer vaccination is reported. The immunopotentiator manganese into six differing structures that resemble the architectures of two types of pathogens (spherical viruses or rod-like bacteria) is precisely manipulated. The results reveal that innate immune cells accurately sense and respond to the architectures, of which two outperformed material candidates (151nm hollow spheres and hollow microrods with an aspect ratio of 4.5) show higher competence in creating local proinflammatory environment with promoted innate immune cell influx and stimulation on dendritic cells (DCs). In combination with viral peptides, model proteins, or cell lysate antigens, the outperformed microrod material remarkably primes antigen-specific CD8 cytolytic T cells. In prophylactic and therapeutic regimens, the microrod adjuvanted vaccines display optimal aptitude in tumor suppression in four aggressive murine tumor models, by promoting the infiltration of heterogeneous cytolytic effector cells while decreasing suppressive immunoregulatory populations in tumors. This study demonstrates that a rationally selected architecture of immunogenic materials potentially advances the clinical reality of cancer vaccination.

Abstract 6707: IgG based B7-H3xCD3 bispecific antibody for treatment of gynecological tumors

Abstract Ovarian cancer is characterized by late stage disease and a poor prognosis. Endometrial cancer is a heterogeneous disease with different prognoses depending on the molecular subtype. Novel treatment strategies are needed to improve patient survival in both entities. T cell-based immunotherapy has significantly improved the treatment options for many malignancies. B7-H3 (CD276) is an interesting target for immunotherapeutic approaches because it is overexpressed on a wide range of tumor entities and has characteristics of immune checkpoint molecules. Its expression not only on tumor cells but also on the tumor vasculature allows improved infiltration of immune effector cells into the tumor to overcome the major limitation of T cell-based therapy of solid tumors, which is the access of immune cells to the tumor site. A novel bispecific antibody in an IgG-based format called CC-3 with B7-H3xCD3 specificity has already been shown to efficiently induce T cell responses against gastrointestinal malignancies. Here we show that B7-H3 is highly expressed on endometrial and ovarian cancer cell lines. Treatment with CC-3 induced T cell activation and degranulation as well as secretion of IL-2, IFNy and perforin. These results demonstrate CC-3 induced T cell reactivity against ovarian and endometrial cancer cells. In addition, CC-3 induced efficient T cell proliferation and formation of T cell memory subsets. This resulted in potent target cell lysis. Taken together, our results underscore the potential of CC-3, which is currently in GMP manufacturing, to enable clinical evaluation for the treatment of gynecologic malignancies. Citation Format: Sarah Maria Greiner, Jonas Mauermann, Martina Lutz, Ilona Hagelstein, Engler Tobias, André Koch, Hartkopf Andreas, Brucker Sara, Latifa Zekri, Melanie Märklin. IgG based B7-H3xCD3 bispecific antibody for treatment of gynecological tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6707.

Interleukin-2/anti-interleukin-2 immune complex attenuates cold ischemia-reperfusion injury after kidney transplantation by increasing renal regulatory T cells.

Cold ischemia-reperfusion injury (IRI) is an unavoidable complication of kidney transplantation. We investigated the role of regulatory T cells (Treg) in cold IRI and whether the interleukin (IL)-2/anti-IL-2 antibody complex (IL-2C) can ameliorate cold IRI. We developed a cold IRI mouse model using kidney transplantation and analyzed the IL-2C impact on cold IRI in acute, subacute and chronic phases. Treg transfer attenuated cold IRI, while Treg depletion aggravated cold IRI. Next, IL-2C administration prior to IRI mitigated acute renal function decline, renal tissue damage and apoptosis and inhibited infiltration of effector cells into kidneys and pro-inflammatory cytokine expression on day 1 after IRI. On day 7 after IRI, IL-2C promoted renal regeneration and reduced subacute renal damage. Furthermore, on day 28 following IRI, IL-2C inhibited chronic fibrosis. IL-2C decreased reactive oxygen species-mediated injury and improved antioxidant function. When IL-2C was administered following IRI, it also increased renal regeneration with Treg infiltration and suppressed renal fibrosis. In contrast, Treg depletion in the presence of IL-2C eliminated the positive effects of IL-2C on IRI. Tregs protect kidneys from cold IRI and IL-2C inhibited cold IRI by increasing the renal Tregs, suggesting a potential of IL-2C in treating cold IRI. Interleukin (IL)-2/anti-IL-2 antibody complex attenuated acute renal injury, facilitated subacute renal regeneration and suppressed chronic renal fibrosis after cold ischemia-reperfusion injury (IRI) by increasing the renal Tregs. IL-2/anti-IL-2 antibody complex decreased reactive oxygen species-mediated injury and improved antioxidant function. This study suggests the therapeutic potential of the IL-2/anti-IL-2 antibody complex in kidney transplantation-associated cold IR.

Biomimetic Responsive Nanoconverters with Immune Checkpoint Blockade Plus Antiangiogenesis for Advanced Hepatocellular Carcinoma Treatment.

The first-line treatment for advanced hepatocellular carcinoma (HCC) combines immune checkpoint inhibitors and antiangiogenesis agents to prolong patient survival. Nonetheless, this approach has several limitations, including stringent inclusion criteria and suboptimal response rates that stem from the severe off-tumor side effects and the unfavorable pharmacodynamics and pharmacokinetics of different drugs delivered systemically. Herein, we propose a single-agent smart nanomedicine-based approach that mimics the therapeutic schedule in a targeted and biocompatible manner to elicit robust antitumor immunity in advanced HCC. Our strategy employed pH-responsive carriers, poly(ethylene glycol)-poly(β-amino esters) amphiphilic block copolymer (PEG-PAEs), for delivering apatinib (an angiogenesis inhibitor), that were surface-coated with plasma membrane derived from engineered cells overexpressing PD-1 proteins (an immune checkpoint inhibitor to block PD-L1). In an advanced HCC mouse model with metastasis, these biomimetic responsive nanoconverters induced significant tumor regression (5/9), liver function recovery, and complete suppression of lung metastasis. Examination of the tumor microenvironment revealed an increased infiltration of immune effector cells (CD8+ and CD4+ T cells) and reduced immunosuppressive cells (myeloid-derived suppressor cells and T regulatory cells) in treated tumors. Importantly, our nanomedicine selectively accumulated in both small and large HCC occupying >50% of the liver volume to exert therapeutic effects with minimal systemic side effects. Overall, these findings highlight the potential of such multifunctional nanoconverters to effectively reshape the tumor microenvironment for advanced HCC treatment.

POLQ identifies a better response subset to immunotherapy in muscle-invasive bladder cancer with high PD-L1.

Though programmed cell death-ligand 1 (PD-L1) has been used in predicting the efficacy of immune checkpoint blockade (ICB), it is insufficient as a single biomarker. As a key effector of an intrinsically mutagenic microhomology-mediated end joining (MMEJ) pathway, DNA polymerase theta (POLQ) was overexpressed in various malignancies, whose expression might have an influence on genomic stability, therefore altering the sensitivity to chemotherapy and immunotherapy. A total of 1304 patients with muscle-invasive bladder cancer (MIBC) from six independent cohorts were included in this study. The Zhongshan Hospital (ZSHS) cohort (n = 134), The Cancer Genome Atlas (TCGA) cohort (n = 391), and the Neo-cohort (n = 148) were included for the investigation of chemotherapeutic response. The IMvigor210 cohort (n = 234) and the UNC-108 cohort (n = 89) were used for the assessment of immunotherapeutic response. In addition, the relationship between POLQ and the immune microenvironment was assessed, and GSE32894 (n = 308) was used only for the evaluation of the immune microenvironment. We identified POLQhigh PD-L1high patients could benefit more from immunotherapy and platinum-based chemotherapy. Further analysis revealed that high POLQ expression was linked to chromosome instability and higher tumor mutational burden (TMB), which might elicit the production of neoantigens. Further, high POLQ expression was associated with an active tumor immune microenvironment with abundant infiltration of immune effector cells and molecules. The study demonstrated that high POLQ expression was correlated with chromosome instability and antitumor immune microenvironment in MIBC, and the combination of POLQ and PD-L1 could be used as a superior companion biomarker for predicting the efficacy of immunotherapy.

A combination of PD-L1-targeted IL-15 mRNA nanotherapy and ultrasound-targeted microbubble destruction for tumor immunotherapy

PD-1/PD-L1-based immune checkpoint blockade therapy has shown limited benefits in tumor patients, partially attributed to the inadequate infiltration of immune effector cells within tumors. Here, we established a nanoplatform named DPPA/IL-15 NPs to target PD-L1 for the tumor delivery of IL-15 messenger RNA (mRNA). DPPA/IL-15 NPs were endowed with ultrasound responsiveness and contrast-enhanced ultrasound (CEUS) imaging performance. They effectively protected IL-15 mRNA from degradation and specifically transfected it into tumor cells through the utilization of ultrasound-targeted microbubble destruction (UTMD). This resulted in the activation of IL-15-related immune effector cells while blocking the PD-1/PD-L1 pathway. In addition, UTMD could generate reactive oxygen species (ROS) that induce endoplasmic reticulum (ER) stress-driven immunogenic cell death (ICD), initiating anti-tumor immunity. In vitro and in vivo studies revealed that this combination therapy could induce a robust systemic immune response and enhance anti-tumor efficacy. Thus, this combination therapy has the potential for clinical translation through enhanced immunotherapy and provides real-time ultrasound imaging guidance.

Abstract A048: Enhancing the immune response in locally advanced pancreatic cancer (LAPC) with intratumoral endoscopic ultrasound-guided fine needle injection of large surface area microparticle paclitaxel (LSAM-PTX)

Abstract Background: The complex tumor microenvironment (TME) of locally advanced pancreatic cancer (LAPC) is resistant to immune effector cell infiltration and drug bioavailability, limiting the effectiveness of current systemic therapies. Large Surface Area Microparticle Paclitaxel (LSAM-PTX) was developed for intratumoral (IT) injection to solid tumors to provide a paclitaxel depot at therapeutic levels with a prolonged residence time. We previously reported enhanced tumor kill in LAPC subjects receiving two monthly IT LSAM-PTX injections and demonstrated immunomodulation in the TME. Multiplex immunofluorescence (mIF) performed on pre-treatment biopsy tissue and post-LSAM-PTX resection showed increases in densities of effector T cells and NK cells and decreases in myeloid derived suppressor cells (MDSC). To further evaluate immunomodulation following IT LSAM-PTX therapy, blood samples were obtained for immunophenotyping using flow cytometry (NCT03077685). Methods: Pretreatment samples were obtained from subjects administered four injections (n=14) and at visits throughout the treatment period and up to six months post first LSAM-PTX injection. Flow cytometric analysis was performed using three panels to evaluate absolute counts and percentages of up to 45 immunophenotypes. Results: Treatment with four IT injections of LSAM-PTX (ongoing median overall survival = 12.6 months; follow-up = 12.2 months) was associated with increases in peripheral lymphocytes during the treatment period. An enhancement of immune subsets associated with anti-tumor effector functions was found, including significant increases in circulating CD3+ and CD4+ cells (p=0.0012 and 0.002 respectively) which is consistent with the 4-fold increase in CD4+helper T cells detected in the TME of resected tissue. A significant increase in LAG3+CD8+ T cells was seen following treatment initiation (p=0.048) marking early T cell activation. Increases in circulating CD8+ T cells, B cells, and NK cells were seen during the treatment period corresponding with effector T cells and NK cells increases (4.8-fold increase in density) within the TME. Six months following treatment initiation there was a significant decrease in circulating Treg cells (p=0.0183) and a decrease in peripheral immunosuppressive G-MDSC, consistent with the reduction in MDSC measured in the TME. Notably, systemic taxane toxicities, including immunosuppression, were not reported following IT LSAM-PTX and negligible amounts of paclitaxel were found in blood samples. Conclusion: Immunophenotyping of blood from LAPC subjects treated with IT LSAM-PTX demonstrates immunomodulation to a phenotype associated with anti-tumor immune effects, including favorable immunosurveillance, and is consistent with changes found in the TME in resected tissues. Immunosuppressive cell types typically associated with poor outcomes were reduced at six months. Anti-tumor immunomodulation without immunosuppression suggests that IT LSAM-PTX may be amenable to combination with immunotherapy. Citation Format: Andrew E. Hendifar, Max M. Wattenberg, Holly A. Maulhardt, Alyson M. Marin, Alison E. Wendt, Shelagh J. Verco, Gere S. diZerega. Enhancing the immune response in locally advanced pancreatic cancer (LAPC) with intratumoral endoscopic ultrasound-guided fine needle injection of large surface area microparticle paclitaxel (LSAM-PTX) [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A048.

Tissue factor overexpression promotes resistance to KRAS-G12C inhibition in non-small cell lung cancer

The recently approved KRASG12C mutation-specific inhibitors sotorasib and adagrasib (KRASG12C-I) represent a promising therapy for KRASG12C-driven non-small cell lung cancer (NSCLC). However, many eligible patients do not benefit due to intrinsic or acquired drug resistance. Tissue factor (TF) is overexpressed in KRAS-mutated (KRASmut) NSCLC and is the target of the FDA-approved ADC Tivdak. Here, we employed HuSC1-39, the parent antibody of a clinical stage TF-ADC (NCT04843709), to investigate the role of TF in KRASmut NSCLC. We found that patients with TF-overexpression had poor survival, elevated P-ERK/P-AKT activity levels and low immune effector cell infiltration in the tumor. In a panel of KRASG12C cell lines, KRASG12C-I response correlated with suppression of TF mRNA, which was not observed in resistant cells. In the drug resistant cells, TF-overexpression relied on an mTORC2-mediated and proteasome-dependent pathway. Combination treatment of HuSC1-39 or mTORC1/2 inhibitor MTI-31 with KRASG12C-I each produced synergistic antitumor efficacy in cell culture and in an orthotopic lung tumor model. TF-depletion in the resistant cells diminished epithelial mesenchymal transition, reduced tumor growth and greatly sensitized KRASG12C-I response. Moreover, employing immunohistochemistry and coculture studies, we demonstrated that HuSC1-39 or MTI-31 reset the tumor microenvironment and restore KRASG12C-I sensitivity by reshaping an M1-like macrophage profile with greatly enhanced phagocytic capacity toward tumor cell killing. Thus, we have identified the TF/mTORC2 axis as a critical new mechanism for triggering immunosuppression and KRASG12C-I resistance. We propose that targeting this axis with HuSC1-39 or MTI-31 will improve KRASG12C-I response in KRAS-driven NSCLC.

Identification of the prognostic immune subtype in copy-number high endometrial cancer.

The TCGA molecular subtype of endometrial cancer (EC) is widely applied, among which the copy-number high (CNH) subtype has the poorest prognosis. However, the heterogeneity of this subtype remains elusive. In this study, we aimed to identify heterogeneous immune subtypes in CNH EC and explore their prognostic significance. We collected 60 CNH EC cases in the TCGA database and performed unsupervised cluster analysis based on the enrichment scores of immune-related gene signatures to identify immune subtypes. We described their immune characteristics and prognoses and conducted differential gene analysis and lasso regression to identify a prognostic biomarker, GZMM. For experimental validation, we performed immunohistochemical staining of GZMM in 39 p53-positive EC surgical samples. We defined two immune subtypes, immune-hot (IH) and immune-cold (IC), which differed in immune cell infiltration, cytokine and chemokine expression and prognosis. The IH subtype has significantly stronger immune activation than the IC subtype, showing a significant infiltration of immune effector cells and high expression of relevant chemokines, with better prognosis. Moreover, the immunohistochemical staining of GZMM in a cohort of 39 p53-positive EC surgical samples confirmed GZMM as a unique prognostic biomarker, with high expression in both tumor cells and lymphocytes predicting a better prognosis. Our study revealed heterogeneous immune subtypes in CNH EC and identified GZMM as a prognostic biomarker. The stratified classification strategy combining molecular and immune subtypes provides valuable insights for future clinical practice.

Cold to Hot: Tumor Immunotherapy by Promoting Vascular Normalization Based on PDGFB Nanocomposites.

Immunotherapy is a promising cancer therapeutic strategy. However, the "cold" tumor immune microenvironment (TIME), characterized by insufficient immune cell infiltration and immunosuppressive status, limits the efficacy of immunotherapy. Tumor vascular abnormalities due to defective pericyte coverage aregradually recognized as a profound determinant in "cold" TIME establishment by hindering immune cell trafficking. Recently, several vascular normalization strategies by improving pericyte coverage have been reported, whereas have unsatisfactory efficacy and high rates of resistance. Herein, a combinatorial strategy to induce tumor vasculature-targeted pericyte recruitment and zinc ion-mediated immune activation with aplatelet-derived growth factor B (PDGFB)-loaded, cyclo (Arg-Gly-Asp-D-Phe-Lys)-modified zeolitic imidazolate framework 8 (PDGFB@ZIF8-RGD) nanoplatform is proposed. PDGFB@ZIF8-RGD effectively induced tumor vascular normalization, which facilitated trafficking and infiltration of immune effector cells, including natural killer (NK) cells, M1-like macrophages and CD8+ T cells, into tumor microenvironment. Simultaneously, vascular normalization promoted the accumulation of zinc ions inside tumors to trigger effector cell immune activation and effector molecule production. The synergy between these two effects endowed PDGFB@ZIF8-RGD with superior capabilities in reprogramming the "cold" TIME to a "hot" TIME, thereby initiating robust antitumor immunity and suppressing tumor growth. This combinatorial strategy for improving immune effector cell infiltration and activation is a promising paradigm for solid tumor immunotherapy.

Immunoediting of KEAP1-NRF2 mutant tumours is required to circumvent NRF2-mediated immune surveillance

In human cancer, activating mutations in the KEAP1-NRF2 pathway are frequently observed, and positively selected for, as they confer the cytoprotective functions of the transcription factor NRF2 on the cancer cells. This results in the development of aggressive tumours which are resistant to treatment with chemotherapeutic compounds. Recent clinical developments have also revealed that NRF2-activated cancers are similarly resistant to immune checkpoint inhibitor drugs. As the mechanism of action of these immune modulating therapies is tangential to the classical cytoprotective function of NRF2, it is unclear how aberrant NRF2 activity could impact the anti-cancer functionality of the immune system. In this context, we found that in human cancer, NRF2-activated cells are highly immunoedited, which allows the cancer cells to escape immune surveillance and develop into malignant tumours. This immunoediting takes the form of reduced antigen presentation by the MHC-I complex, coupled with reduced expression of activating ligands for NK cells. Together, these modifications to the immunogenicity of NRF2-activated cancers inhibit immune effector cell infiltration and engagement, and contribute to the formation of the immunologically cold tumour microenvironment which is a characteristic feature of NRF2-activated cancers.

Glioblastoma cellular MAP4K1 facilitates tumor growth and disrupts T effector cell infiltration.

MAP4K1 has been identified as a cancer immunotherapy target. Whether and how cancer cell-intrinsic MAP4K1 contributes to glioblastoma multiforme (GBM) progression remains unclear. We found that MAP4K1 was highly expressed in the glioma cells of human GBM specimens. High levels of MAP4K1 mRNA were prevalent in IDH-WT and 1p/19q non-codeletion gliomas and correlated with poor prognosis of patients. MAP4K1 silencing inhibited GBM cell proliferation and glioma growth. Transcriptome analysis of GBM cells and patient samples showed that MAP4K1 modulated cytokine‒cytokine receptor interactions and chemokine signaling pathway, including IL-18R and IL-6R Importantly, MAP4K1 loss down-regulated membrane-bound IL-18R/IL-6R by inhibiting the PI3K-AKT pathway, whereas MAP4K1 restoration rescued this phenotype and therefore GBM cell proliferation. MAP4K1 deficiency abolished GBM cell pro-proliferation responses to IL-18, suggesting an oncogenic role of MAP4K1 via the intrinsic IL-18/IL-18R pathway. In addition, GBM cell-derived MAP4K1 impaired T-cell migration and reduced CD8+ T-cell infiltration in mouse glioma models. Together, our findings provide novel insight into the pathological significance of GBM cell-intrinsic MAP4K1 in driving tumor growth and immune evasion by remodeling cytokine-chemokine networks.

Dual-Targeting Nanoliposome Improves Proinflammatory Immunomodulation of the Tumor Microenvironment.

Immunotherapies targeting immune checkpoints have revolutionized cancer treatment by normalizing the immunosuppressive microenvironment of tumors and reducing adverse effects on the immune system. Indoleamine 2,3-dioxygenase (IDO) inhibitors have garnered attention as a promising therapeutic agent for cancer. However, their application alone has shown limited clinical benefits. Cabozantinib, a multitarget tyrosine kinase inhibitor, holds immunomodulatory potential by promoting infiltration and activation of effector cells and inhibiting suppressive immune cells. Despite its potential, cabozantinib as a monotherapy has shown limited efficacy in terms of objective response rate. In this study, IDO-IN-7 and cabozantinib are coencapsulated into liposomes to enhance tumor accumulation and minimize adverse effects. The liposomal combination exhibits potent cytotoxicity and inhibits the function of IDO enzyme. Furthermore, the dual-targeted treatment effectively inhibits tumor development and reverses the suppressive tumor microenvironment by regulating both adaptive and innate branch of immune system. This is evidenced by pronounced infiltration of T cells and B cells, a decrease of regulatory T lymphocytes, a shift to a proinflammatory phenotype of tumor-associated macrophages, and increases levels of neutrophils. This is the first developed of a liposome-delivered combination of IDO inhibitors and cabozantinib, and holds great potential for future clinical application as a promising anticancer strategy.

Systemic administration of STING agonist promotes myeloid cells maturation and antitumor immunity through regulating hematopoietic stem and progenitor cell fate.

STING is a pivotal mediator of effective innate and adaptive anti-tumor immunity; however, intratumoral administration of STING agonists have shown limited therapeutic benefit in clinical trials. The systemic effect of the intravenous delivery of STING agonists in cancer is not well-defined. Here, we demonstrated that systemic administration of STING agonist inhibited melanoma growth, improved inflammatory effector cell infiltration, and induced bone marrow mobilization and extramedullary hematopoiesis, causing widespread changes in immune components in the peripheral blood. The systemically administered STING agonist promoted HSC expansion and influenced lineage fate commitment, which was manifested as the differentiation of HSPCs was skewed toward myeloid cells at the expense of B-cell lymphopoiesis and erythropoiesis. Transcriptome analysis revealed upregulation of myeloid lineage differentiation-related and type I interferon-related genes. This myeloid-biased differentiation promoted the production and maturation of myeloid cells toward an activated phenotype. Furthermore, depletion of Gr-1+ myeloid cells attenuated the anti-tumor immunity of STING agonist. Our findings reveal the anti-tumor mechanism of systemic administration of STING agonist that involves modulating HSPC differentiation and promoting myeloid cells maturation. Our study may help explain the limited clinical activity of STING agonists administered intratumorally.

Establishment and validation of a prognostic signature for pancreatic ductal adenocarcinoma based on lactate metabolism-related genes.

Background: Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal malignancy with poor prognosis. To improve patient outcomes, it is necessary to gain a better understanding of the oncogenesis and progression of this disease. Metabolic reprogramming, particularly the regulation of lactate metabolism, is known to have a significant impact on tumor microenvironment and could provide valuable insights for the management of PDAC patients. In this study, we aimed to investigate the prognostic potential of lactate metabolism-related genes (LMRGs). Methods: Transcriptomic data of patients with PDAC along with the clinical outcomes were retrieved from The Cancer Genome Atlas database, and the expression data in normal pancreas from Genotype-Tissue Expression dataset were adopted as the normal control. By using Cox and LASSO regression models, we identified key genes that are differentially expressed in cancerous tissues and related to prognosis. To determine the prognostic value of LMRGs in PDAC, we evaluated their clinical significance and model performance using both the area under the receiver operator characteristic curve (AUC) and calibration curves. In addition, we evaluated the drug sensitivity prediction and immune infiltration by using oncoPredict algorithm, single sample gene set enrichment analysis and Tumor Immune Estimation Resource. Results: A total of 123 LMRGs were identified through differential gene screening analysis, among which 7 LMRGs were identified to comprise a LMRGs signature that independently predict overall survival of these PDAC patient. The AUC values for the LMRGs signature were 0.786, 0.820, 0.837, and 0.816 for predicting 1-, 2-, 3- and 5-year overall survival respectively. Furthermore, this prognostic signature was used to stratify patients into high-risk and low-risk groups, with the former having worse clinical outcomes. This observation was further validated through analysis of the International Cancer Genome Consortium database. In addition, lower sensitivity to gemcitabine and infiltration of immune effector cells were observed in the cancer tissue of patients in the high-risk group. Conclusion: In conclusion, our data suggests that a genomic signature comprised of these LMRGs may be a novel predictor of overall clinical outcomes and present therapeutic potential for PDAC patients.