Promoting T-cell Infiltration Research Articles

ITPRIPL1 binds CD3ε to impede T cell activation and enable tumor immune evasion

Cancer immunotherapy has transformed treatment possibilities, but its effectiveness differs significantly among patients, indicating the presence of alternative pathways for immune evasion. Here, we show that ITPRIPL1 functions as an inhibitory ligand of CD3ε, and its expression inhibits Tcells in the tumor microenvironment. The binding of ITPRIPL1 extracellular domain to CD3ε on Tcells significantly decreased calcium influx and ZAP70 phosphorylation, impeding initial Tcell activation. Treatment with a neutralizing antibody against ITPRIPL1 restrained tumor growth and promoted Tcell infiltration in mouse models across various solid tumor types. The antibody targeting canine ITPRIPL1 exhibited notable therapeutic efficacy against naturally occurring tumors in pet clinics. These findings highlight the role of ITPRIPL1 (or CD3L1, CD3ε ligand 1) in impeding Tcell activation during the critical "signal one" phase. This discovery positions ITPRIPL1 as a promising therapeutic target against multiple tumor types.

The CXCL16-CXCR6 axis in glioblastoma modulates T-cell activity in a spatiotemporal context.

Glioblastoma multiforme (GBM) pathobiology is characterized by its significant induction of immunosuppression within the tumor microenvironment, predominantly mediated by immunosuppressive tumor-associated myeloid cells (TAMCs). Myeloid cells play a pivotal role in shaping the GBM microenvironment and influencing immune responses, with direct interactions with effector immune cells critically impacting these processes. Our study investigates the role of the CXCR6/CXCL16 axis in T-cell myeloid interactions within GBM tissues. We examined the surface expression of CXCL16, revealing its limitation to TAMCs, while microglia release CXCL16 as a cytokine. The study explores how these distinct expression patterns affect T-cell engagement, focusing on the consequences for T-cell function within the tumor environment. Additionally, we assessed the significance of CXCR6 expression in T-cell activation and the initial migration to tumor tissues. Our data demonstrates that CXCL16 surface expression on TAMCs results in predominant T-cell engagement with these cells, leading to impaired T-cell function within the tumor environment. Conversely, our findings highlight the essential role of CXCR6 expression in facilitating T-cell activation and initial migration to tumor tissues. The CXCL16-CXCR6 axis exhibits dualistic characteristics, facilitating the early stages of the T-cell immune response and promoting T-cell infiltration into tumors. However, once inside the tumor, this axis contributes to immunosuppression. The dual nature of the CXCL16-CXCR6 axis underscores its potential as a therapeutic target in GBM. However, our results emphasize the importance of carefully considering the timing and context of intervention. While targeting this axis holds promise in combating GBM, the complex interplay between TAMCs, microglia, and T cells suggests that intervention strategies need to be tailored to optimize the balance between promoting antitumor immunity and preventing immunosuppression within the dynamic tumor microenvironment.

Superantigen-fused T cell engagers for tumor antigen-mediated robust T cell activation and tumor cell killing

Inadequate Tcell activation has severely limited the success of Tcell engager (TCE) therapy, especially in solid tumors. Enhancing Tcell activity while maintaining the tumor specificity of TCEs is the key to improving their clinical efficacy. However, currently, there needs to be more effective strategies in clinical practice. Here, we design novel superantigen-fused TCEs that display robust tumor antigen-mediated Tcell activation effects. These innovative drugs are not only armed with the powerful Tcell activation ability of superantigens but also retain the dependence of TCEs on tumor antigens, realizing the ingenious combination of the advantages of two existing drugs. Superantigen-fused TCEs have been preliminarily proven to have good (>30-fold more potent) and specific (>25-fold more potent) antitumor activity invitro and invivo. Surprisingly, they can also induce the activation of Tcell chemotaxis signals, which may promote Tcell infiltration and further provide an additional guarantee for improving TCE efficacy in solid tumors. Overall, this proof-of-concept provides a potential strategy for improving the clinical efficacy of TCEs.

Systematic Pan-Cancer Analysis Reveals X-C Motif Chemokine Receptor 1 as a Prognostic and Immunological Biomarker.

Chemokines and their receptors play an important role in immune monitoring and immune defense during tumor growth and metastasis. However, their prognostic roles in pan-cancer have not been elucidated. In this work, we screened all chemokine receptors in pan-cancer and discovered X-C Motif Chemokine Receptor 1 (XCR1) as a reliable immunological and prognostic biomarker in pan-cancer using bioinformation. The TCGA database served as the foundation for the primary research database analysis in this work. XCR1 was downregulated in tumors. Patients with reduced XCR1 showed worse prognoses and a concomitant decrease in immune cell infiltration (DCs and CD8+ T cells). According to a gene enrichment study, XCR1 enhanced immune system performance by promoting T-cell infiltration through the C-X-C Motif Chemokine Ligand 9 (CXCL9)- C-X-C Motif Chemokine Receptor 3 (CXCR3) axis. In addition, XCR1 is mainly expressed in infiltrated DCs and some malignant cells in tumor tissues. Our data revealed the important role of XCR1 in remodeling the tumor microenvironment and predicting the survival prognosis, which could also be used as a sensitive biomarker for tumor immunotherapy.

Isorhamnetin and anti-PD-L1 antibody dual-functional mesoporous silica nanoparticles improve tumor immune microenvironment and inhibit YY1-mediated tumor progression

BackgroundThe immune checkpoint inhibitor (ICI) anti-PD-L1 monoclonal antibody can inhibit the progress of hepatocellular carcinoma (HCC). Epithelial–mesenchymal transformation (EMT) can promote tumor migration and the formation of immune-suppression microenvironment, which affects the therapeutic effect of ICI. Yin-yang-1 (YY1) is an important transcription factor regulating proliferation, migration and EMT of tumor cells. This work proposed a drug-development strategy that combined the regulation of YY1-mediated tumor progression with ICIs for the treatment of HCC.MethodsWe first studied the proteins that regulated YY1 expression by using pull-down, co-immunoprecipitation, and duo-link assay. The active compound regulating YY1 content was screened by virtual screening and cell-function assay. Isorhamnetin (ISO) and anti-PD-L1 antibody dual-functional mesoporous silica nanoparticles (HMSN-ISO@ProA-PD-L1 Ab) were prepared as an antitumor drug to play a synergistic anti-tumor role.ResultsYY1 can specifically bind with the deubiquitination enzyme USP7. USP7 can prevent YY1 from ubiquitin-dependent degradation and stabilize YY1 expression, which can promote the proliferation, migration and EMT of HCC cells. Isorhamnetin (ISO) were screened out, which can target USP7 and promote YY1 ubiquitin-dependent degradation. The cell experiments revealed that the HMSN-ISO@ProA-PD-L1 Ab nanoparticles can specifically target tumor cells and play a role in the controlled release of ISO. HMSN-ISO@ProA-PD-L1 Ab nanoparticles inhibited the growth of Hepa1-6 transplanted tumors and the effect was better than that of PD-L1 Ab treatment group and ISO treatment group. HMSN-ISO@ProA-PD-L1 Ab nanoparticles also exerted a promising effect on reducing MDSC content in the tumor microenvironment and promoting T-cell infiltration in tumors.ConclusionsThe isorhamnetin and anti-PD-L1 antibody dual-functional nanoparticles can improve tumor immune microenvironment and inhibit YY1-mediated tumor progression. This study demonstrated the possibility of HCC treatment strategies based on inhibiting USP7-mediated YY1 deubiquitination combined with anti-PD-L1 monoclonal Ab.

Analysis of immunogenic cell death in ascending thoracic aortic aneurysms based on single-cell sequencing data.

At present, research on immunogenic cell death (ICD) is mainly associated with cancer therapy. Little is known about the role of ICD in cardiovascular disease, especially in ascending thoracic aortic aneurysms (ATAA). ATAA single-cell RNA (scRNA) sequencing data were analyzed to identify the involved cell types and determine their transcriptomic characteristics. The chi-square test, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, Gene Set Enrichment Analysis (GSEA), and CellChat for cell-to-cell communication analysis from the Gene Expression Omnibus (GEO) database were used. A total of 10 cell types were identified, namely, monocytes, macrophages, CD4 T/NK (CD4+ T cells and natural killer T cells), mast cells, B/Plasma B cells, fibroblasts, endothelial cells, cytotoxic T cells (CD8+ T cells, CTLs), vascular smooth muscle cells (vSMCs), and mature dendritic cells (mDCs). A large number of inflammation-related pathways were present in the GSEA results. A large number of ICD-related pathways were found in the KEGG enrichment analysis of differentially expressed genes in endothelial cells. The number of mDCs and CTLs in the ATAA group was significantly different from that in the control group. A total of 44 pathway networks were obtained, of which 9 were associated with ICD in endothelial cells (CCL, CXCL, ANNEXIN, CD40, IL1, IL6, TNF, IFN-II, GALECTIN). The most important ligand-receptor pair by which endothelial cells act on CD4 T/NK cells, CTLs and mDCs is CXCL12-CXCR4. The most important ligand-receptor pair by which endothelial cells act on monocytes and macrophages is ANXA1-FPR1. The most important ligand-receptor pair by which CD4 T/NK cells and CTLs act on endothelial cells is CCL5-ACKR1. The most important ligand-receptor pair that myeloid cells (macrophages, monocytes and mDCs) act on endothelial cells is CXCL8-ACKR1. Moreover, vSMCs and fibroblasts mainly promote inflammatory responses through the MIF signaling pathway. ICD is present in ATAA and plays an important role in the development of ATAA. The target cells of ICD may be mainly endothelial cells, in which the aortic endothelial cell ACKR1 receptor can not only promote T-cell infiltration through the CCL5 ligand but also promote myeloid cell infiltration through the CXCL8 ligand. ACKR1 and CXCL12 may become target genes for ATAA drug therapy in the future.

Abstract 2336: Extra-virgin olive oil prevented tumor immune escape and intratumoral metabolic reprogramming in a mouse model of colorectal cancer via promoting T-cell infiltration, DNA repair, attenuation of overexpression of immune checkpoint proteins, glutamine dependency, and the Warburg metabolic phenotype

Abstract Colorectal cancer (CRC) is the third leading cause of cancer death worldwide. Tumor Immune escape via activation of immune checkpoint proteins, T-cell exhaustion, impaired DNA repair mechanism, and metabolic reprogramming are crucial for CRC initiation, promotion, and progression. Immune checkpoint proteins such as programmed cell death 1 (PD1), PD1 ligand 1 (PD-L1), and cytotoxic T lymphocyte antigen 4 (CTLA4) act as inhibitory immunoreceptors that prevent cytotoxic T-cells from infiltrating tumor cells. Also, metabolic pathways are actively reprogrammed during CRC development. Indeed, upregulation of DNA repair mechanisms, enhancement of tumor immunosurveillance via immune checkpoint blockage, and inhibition of intratumoral metabolic reprogramming (IMR) can revolutionize CRC treatment and improve patients’ survival. Epidemiological studies have reported the association between the intake of extra virgin olive oil (EVOO) and a reduced risk of CRC development. However, there is a lack of information on the effect of EVOO on DNA repair, tumor immunosurveillance, and IMR in CRC. Therefore, we investigated the effect of EVOO on DNA repair, tumor immunosurveillance, and IMR in mice model of CRC. Male BALB/c mice were divided into five groups of 10 mice each. Group 1 mice served as controls. Group 2 mice received a single dose of azoxymethane (AOM) 10mg/kg and, after one week, they received three cycles of dextran sulfate sodium (DSS) at 3% in their drinking water. Group 3 mice received EVOO (2ml/Kg) by oral gavage in combination with AOM and three cycles of 3% DSS (W/V) in drinking water. Group 4 received EVOO (2ml/Kg) only while Group 5 received 5-fluorouracil (10mg/kg) in combination with AOM and three cycles of 3% DSS (W/V) in drinking water. The experiment was terminated after confirmation of colorectal adenocarcinoma. A high tumor burden was observed to be decreased in CRC mice treated with EVOO. Furthermore, CRC mice treated with EVOO had increased expression of MSH6, PMS2, MLH1, MSH2, P53, CD4+, and CD8+ T-cells and decreased expression of PD1, PD-L1, and CTLA4 when compared to mice with CRC. Also, it decreases the colonic expression of glucose transporters 1 & 4, hypoxia-inducible factor-1 α, pyruvate kinase M2, hexokinase 1 and 2, lactate dehydrogenase, monocarboxylate transporter 4, fatty acid synthase, and glutaminase 1 in CRC mice when compared to the control group. The results obtained from this study showed that EVOO prevented tumor immune escape and IMR in a mouse model of colorectal cancer via promoting T-cell infiltration, DNA repair, and attenuation of overexpression of immune checkpoint proteins, glutamine dependency and Warburg metabolic phenotype. Citation Format: Babajide O. Ajayi, Anuoluwapo Adeshina. Extra-virgin olive oil prevented tumor immune escape and intratumoral metabolic reprogramming in a mouse model of colorectal cancer via promoting T-cell infiltration, DNA repair, attenuation of overexpression of immune checkpoint proteins, glutamine dependency, and the Warburg metabolic phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2336.

Abstract 3646: New functional role of tamoxifen in breast cancer immunomodulation: role of RACK7 in activation of type I interferon signaling and CEACAM1/TIM-3-dependent immunosuppression

Abstract Despite advancements of immunotherapy against various cancers, breast cancer still retains poor response to immune checkpoint blockade (ICB) therapy. Therefore, the identification of promising target or new strategy to enhance ICB therapy in breast cancer is crucial. Here, we uncover that the DNA-damaging potential of tamoxifen (TAM) can shape the unfavorable but ready-to-fire tumor immune microenvironment in breast cancer. We discover that a long-term TAM administration unexpectedly induces JAK/STAT signaling and Type I interferon (IFN)-stimulated gene expression to promote T-cell infiltration. Furthermore, TAM also induces the expression of CEACAM1, which acts as an alternative T-cell inhibitory ligand via binding to TIM-3, a “checkpoint” receptor expressed on CD4+ and CD8+ T cells. The chromatin ‘‘reader’’ RACK7/ZMYND8, which functions as a transcriptional repressor of IFN-stimulated gene (ISG) and a critical factor in DNA repair, is found to be downregulated upon tamoxifen exposure and involved in the tamoxifen-mediated cellular modulation. We demonstrate that TAM in conjunction with RACK7-knockdown (KD) triggers robust upregulation of ISGs and CEACAM1 in both estrogen receptor-positive (ER+) and triple negative breast cancer (TNBC) cells. This immunomodulatory effect lead by loss of RACK7 is specific to TAM treatment, and is not observed when combined with other endocrine therapeutics. TAM combined with RACK7-KD promotes mitochondrial DNA damage, which leads to accumulation of cytosolic DNA and subsequent activation of the cGAS/STING pathway. The murine breast orthotopic models with TS/A, EO771 and 4T1 cells further demonstrate that TAM-mediated immunomodulatory in conjunction with RACK7-KD evokes cytokine/chemokine secretion and further induces T-cell infiltration into tumor microenvironment. However, the tumor killing effect is limited due to promotion of T-cell exhaustion from CEACAM1-TIM-3 interaction between tumor and T cells. Thus, our study indicates that targeting CEACAM1-TIM-3 interaction is crucial for TAM-mediated tumor immune response. This brings promising therapeutic approach with TAM by combining RACK7-KD with blockage to CEACAM1-TIM-3 interaction in breast cancer. The resistance of tamoxifen treatment may be overcome, and RACK7 may serve as both a therapeutic target and a biomarker to enable ICB therapy. Citation Format: Marvin A. Aberin, Yu-Ling (Pony) Lee, Xin-Guo Hsu, Chen-Yang Shen, Yao-Ming Chang, Kun-Yuan Lin, Chandan Guha, Shu-Ping Wang. New functional role of tamoxifen in breast cancer immunomodulation: role of RACK7 in activation of type I interferon signaling and CEACAM1/TIM-3-dependent immunosuppression. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3646.

Nanodrug removes physical barrier to promote T-cell infiltration for enhanced cancer immunotherapy.

The dense extracellular matrix (ECM) is a key barrier to tumor infiltration of cytotoxic T lymphocytes (CTLs), which greatly compromises T cell-dependent immunotherapy of hepatocellular carcinoma (HCC). Herein, hyaluronidase (HAase), IL-12, and anti-PD-L1 antibody (αPD-L1) were co-delivered using a pH and MMP-2 dual-sensitive polymer/calcium phosphate (CaP) hybrid nanocarrier. The dissolution of CaP triggered by tumor acidity facilitated the release of IL-12 and HAase responsible for ECM digestion, enhancing the tumor infiltration and proliferation of CTLs. Furthermore, the in situ-released αPD-L1 inside tumor, as triggered by an overexpressed MMP-2, prevented the tumor cell from escaping the killing effects of CTLs. Such combination strategy induced a robust antitumor immunity for efficiently suppressing HCC growth in mice. Additionally, tumor acidity-sheddable polyethylene glycol (PEG) coating enhanced the tumor accumulation of nanocarrier and reduced the immune-related adverse events (irAEs) induced by on-target off-tumor αPD-L1. This dual-sensitive nanodrug demonstrates an effective immunotherapy paradigm for other dense ECM-characterized solid tumors.

MIP3α as an early prognostic predictor for patients with B-cell malignancies receiving CD19/CD22-redirected CAR-T cell cocktail therapy

PurposeIdentifying the temporal pattern of recurrence and prognostic biomarkers would further help improve the efficacy of chimeric antigen receptor (CAR) -T therapy.MethodsWe examined the prognoses of 119 patients after sequential infusion of anti-CD19 and anti-CD22, a cocktail of 2 single-target CAR (CAR19/22) T cells in an open-label, single-center clinical trial (ChiCTR-OPN-16008526). And we, from a 70-biomarker panel, identified candidate cytokines that might predict the treatment failure, including primary non-response (NR) and early relapse (ER).ResultsIn our study, 3 (11.5%) patients with B-cell acute lymphoblastic leukemia (B-ALL) and 9 (12.2%) cases of B-cell non-Hodgkin lymphoma (NHL) failed to respond to sequential CAR19/22 T-cell infusion (NR). A total of 11 (42.3%) B-ALL patients and 30 (52.7%) B-NHL patients had relapses during follow-up. Most recurrence events (67.5%) occurred within six months of sequential CAR T-cell infusion (ER). We found that macrophage inflammatory protein (MIP)-3α was a highly sensitive and specific prognostic predictor for patients with NR/ER and those attaining over-6-month remission. Patients who had higher MIP3α levels after sequential CAR19/22 T-cell infusion had significantly favorable progression-free survival (PFS) than their counterparts with relatively lower MIP3α expression. Our experiments demonstrated that MIP3α could enhance the therapeutic effect of CAR-T cells by promoting T-cell infiltration into and enriching memory-phenotype T cells in the tumor environment.ConclusionThis study showed that relapse occurred mainly within six months after sequential CAR19/22 T-cell infusion. Moreover, MIP3α could act as a valuable post-infusion biomarker for identifying patients with NR/ER.

Moringa oleifera leaf polysaccharides exert anti-lung cancer effects upon targeting TLR4 to reverse the tumor-associated macrophage phenotype and promote T-cell infiltration.

Tumor-associated macrophages (TAMs) participate in tumorigenesis, growth, invasion as well as metastasis by facilitating an immunosuppressive tumor microenvironment. Reversing the pro-tumoral M2 phenotype of TAMs has become a hot spot in advancing cancer immunotherapy. In the current study, the content of Moringa oleifera leaf polysaccharides (MOLP) was determined and characterized, along with the anti-cancer mechanism of MOLP studied in a Lewis lung cancer (LLC) tumor-bearing mouse model and bone marrow-derived macrophages. The monosaccharide composition and gel permeation chromatography analyses show that MOLP are mainly composed of galactose, glucose, and arabinose, with approximately 17.35 kDa average molecular weight (Mw). In vivo studies demonstrate that MOLP convert TAMs from the immunosuppressive M2 phenotype to the antitumor M1 phenotype, thus inducing CXCL9 and CXCL10 expression and increasing T-cell infiltration in the tumor. Furthermore, macrophage depletion and T cell suppression demonstrated that the tumor suppressive effect of MOLP was reliant on reprogramming macrophage polarization and T cell infiltration. In vitro studies revealed that MOLP could induce the phenotypic switch from M2 macrophages to M1 by targeting TLR4. The current study highlights that MOLP are promising anticancer plant-derived polysaccharides with potential in modulating the immune microenvironment and have a bright application prospect in the immunotherapy of lung cancer.

Lung tumor microphysiological system with 3D endothelium to evaluate modulators of T-cell migration.

Lung cancer is a leading cause of death worldwide, with only a fraction of patients responding to immunotherapy. The correlation between increased T-cell infiltration and positive patient outcomes has motivated the search for therapeutics promoting T-cell infiltration. While transwell and spheroid platforms have been employed, these models lack flow and endothelial barriers, and cannot faithfully model T-cell adhesion, extravasation, and migration through 3D tissue. Presented here is a 3D chemotaxis assay, in a lung tumor-on-chip model with 3D endothelium (LToC-Endo), to address this need. The described assay consists of a HUVEC-derived vascular tubule cultured under rocking flow, through which T-cells are added; a collagenous stromal barrier, through which T-cells migrate; and a chemoattractant/tumor (HCC0827 or NCI-H520) compartment. Here, activated T-cells extravasate and migrate in response to gradients of rhCXCL11 and rhCXCL12. Adopting a T-cell activation protocol with a rest period enables proliferative burst prior to introducing T-cells into chips and enhances assay sensitivity. In addition, incorporating this rest recovers endothelial activation in response to rhCXCL12. As a final control, we show that blocking ICAM-1 interferes with T-cell adhesion and chemotaxis. This microphysiological system, which mimics in vivo stromal and vascular barriers, can be used to evaluate potentiation of immune chemotaxis into tumors while probing for vascular responses to potential therapeutics. Finally, we propose translational strategies by which this assay could be linked to preclinical and clinical models to support human dose prediction, personalized medicine, and the reduction, refinement, and replacement of animal models.

A T-cell engaging bispecific antibody with a tumor-selective bivalent folate receptor alpha binding arm for the treatment of ovarian cancer

ABSTRACT The use of T-cell engagers (TCEs) to treat solid tumors is challenging, and several have been limited by narrow therapeutic windows due to substantial on-target, off-tumor toxicities due to the expression of low levels of target antigens on healthy tissues. Here, we describe TNB-928B, a fully human TCE that has a bivalent binding arm for folate receptor alpha (FRα) to selectively target FRα overexpressing tumor cells while avoiding the lysis of cells with low levels of FRα expression. The bivalent design of the FRα binding arm confers tumor selectivity due to low-affinity but high-avidity binding to high FRα antigen density cells. TNB-928B induces preferential effector T-cell activation, proliferation, and selective cytotoxic activity on high FRα expressing cells while sparing low FRα expressing cells. In addition, TNB-928B induces minimal cytokine release compared to a positive control TCE containing OKT3. Moreover, TNB-928B exhibits substantial ex vivo tumor cell lysis using endogenous T-cells and robust tumor clearance in vivo, promoting T-cell infiltration and antitumor activity in mouse models of ovarian cancer. TNB-928B exhibits pharmacokinetics similar to conventional antibodies, which are projected to enable favorable administration in humans. TNB-928B is a novel TCE with enhanced safety and specificity for the treatment of ovarian cancer.

Reversing chemokine/chemokine receptor mismatch to enhance the antitumor efficacy of CAR-T cells.

Currently, the antitumor efficacy of chimeric antigen receptor Tcells in solid tumors is modest. Both chemokines and their receptors play a key role in the proliferation of cancer cells, tumor angiogenesis, organ-selective metastasisand migration of immune cells to solid tumors. Unfortunately, frequent chemokine/chemokine receptor 'mismatch'between effector cells and the tumor microenvironment results in inefficient T-cell infiltration and antitumor efficacy. Thus, reversing the 'mismatch'of chemokines and chemokine receptors appears to be a promising method for promoting T-cell infiltration into the tumor and enhancing their antitumor efficacy. In this review, we discuss functions of the chemokine/chemokine receptor axis in cancer immunity and the current understanding, challengesand prospects for improving the effect of chimeric antigen receptor T cells by reversing the mismatch between chemokines and chemokine receptors.

A new PD-1-specific nanobody enhances the antitumor activity of T-cells in synergy with dendritic cell vaccine.

Despite the many successes and opportunities presented by PD-1 blockade in cancer therapies, anti-PD-1 monoclonal antibodies still face multiple challenges. Herein we report a strategy based on a nanobody (Nb) to circumvent these obstacles. A new PD-1-blocking Nb (PD-1 Nb20) in combination with tumor-specific dendritic cell (DC)/tumor-fusion cell (FC) vaccine that aims to improve the activation, proliferation, cytokine secretion, and tumor cell cytotoxicity of CD8+ T-cells. This combination was found to effectively enhance the in vitro cytotoxicity of CD8+ T-cells to kill human non-small cell lung cancer (NSCLC) HCC827cells, hepatocellular carcinoma (HCC) HepG2 cells, and tongue squamous cell carcinoma (TSCC) Tca8113cells. Moreover, CD8+ T-cells pre-treated with PD-1 Nb20 and tumor-specific DC/tumor-FCs significantly suppressed the growth of NSCLC-, HCC- and TSCC-derived xenograft tumors and prolonged the survival of tumor-bearing mice, through promoting T-cell infiltration to kill tumor cells and inhibiting tumor angiogenesis. These data demonstrate that PD-1 Nb20 in synergy with DC/tumor-FC vaccine augment the broad spectrum of antitumor activity of CD8+ T-cells, providing an alternative and promising immunotherapeutic strategy for tumor patients who are T-cell-dysfunctional or not sensitive to anti-PD-1 therapy.

Pharmacologic Activation of p53 Triggers Viral Mimicry Response Thereby Abolishing Tumor Immune Evasion and Promoting Antitumor Immunity.

We found that p53 activated by MDM2 inhibitors induced the expression of ERVs, in part via epigenetic factors LSD1 and DNMT1. Induction of IFN response caused by ERV derepression upon p53-targeting therapies provides a possibility to overcome resistance to immune checkpoint blockade and potentially transform "cold" tumors into "hot." This article is highlighted in the In This Issue feature, p. 2945.

T-cell responses and combined immunotherapy against human carbonic anhydrase 9-expressing mouse renal cell carcinoma.

Renal cell carcinoma (RCC) is known to respond to immune checkpoint blockade (ICB) therapy, whereas there has been limited analysis of T-cell responses to RCC. In this study, we utilized human carbonic anhydrase 9 (hCA9) as a model neoantigen of mouse RENCA RCC. hCA9-expressing RENCA RCC (RENCA/hCA9) cells were rejected in young mice but grew in aged mice. CD8+ T cells were the primary effector cells involved in rejection in young mice, whereas CD4+ T cells participated at the early stage. Screening of a panel of hCA9-derived peptides revealed that mouse CD8+ T cells responded to hCA9288-296 peptide. Mouse CD4+ T cells responded to lysates of RENCA/hCA9, but not RENCA cells, and showed reactivity to hCA9 276-290, which shares three amino acids with hCA9 288-296 peptide. Immunohistochemistry analysis revealed that few T cells infiltrated RENCA/hCA9 tissues in aged mice. ICB therapy of anti-PD-1/anti-CTLA-4 antibodies promoted T-cell infiltration into tumor tissues, whereas no definite antitumor effect was observed. However, additional combination with cyclophosphamide or axitinib, a vascular endothelial growth factor receptor inhibitor, induced complete regression in half of the RENCA/hCA9-bearing aged mice with increased expression of PD-L1 in tumor tissues. These results indicate that hCA9 can be a useful model neoantigen to investigate antitumor T-cell responses in mice with RCC, and that RENCA/hCA9 in aged mice can serve as a non-inflamed 'cold' tumor model facilitating the development of effective combined immunotherapies for RCC.

STING Activated Tumor-Intrinsic Type I Interferon Signaling Promotes CXCR3 Dependent Antitumor Immunity in Pancreatic Cancer.

Background & AimsPancreatic ductal adenocarcinoma (PDA) is a lethal chemoresistant cancer that exhibits early metastatic spread. The highly immunosuppressive PDA tumor microenvironment renders patients resistant to emerging immune-targeted therapies. Building from our prior work, we evaluated stimulator of interferon genes (STING) agonist activation of PDA cell interferon-α/β-receptor (IFNAR) signaling in systemic antitumor immune responses.MethodsPDA cells were implanted subcutaneously to wild-type, IFNAR-, or CXCR3-knockout mice. Tumor growth was monitored, and immune responses were comprehensively profiled.ResultsHuman and mouse STING agonist ADU-S100 reduced local and distal tumor burden and activated systemic antitumor immune responses in PDA-bearing mice. Effector T-cell infiltration and inflammatory cytokine and chemokine production, including IFN-dependent CXCR3-agonist chemokines, were elevated, whereas suppressive immune populations were decreased in treated tumors. Intratumoral STING agonist treatment also generated inflammation in distal noninjected tumors and peripheral immune tissues. STING agonist treatment of type I IFN–responsive PDA tumors engrafted to IFNAR-/- recipient mice was sufficient to contract tumors and stimulate local and systemic T-cell activation. Tumor regression and CD8+ T-cell infiltration were abolished in PDA engrafted to CXCR3-/- mice treated with STING agonist.ConclusionsThese data indicate that STING agonists promote T-cell infiltration and counteract immune suppression in locally treated and distant tumors. Tumor-intrinsic type I IFN signaling initiated systemic STING-mediated antitumor inflammation and required CXCR3 expression. STING-mediated induction of systemic immune responses provides an approach to harness the immune system to treat primary and disseminated pancreatic cancers.

LBA31 Primary results from IMagyn050/GOG 3015/ENGOT-OV39, a double-blind placebo (pbo)-controlled randomised phase III trial of bevacizumab (bev)-containing therapy +/- atezolizumab (atezo) for newly diagnosed stage III/IV ovarian cancer (OC)

Atezo, which targets PD-L1, is effective in several cancers. Blocking tumour-associated VEGF may promote T-cell infiltration into the tumour bed and boost anti-tumour immune response, justifying combination with bev. Dual VEGF-A and PD-L1 blockade is effective in lung and hepatocellular cancers. IMagyn050 (NCT03038100) enrolled patients (pts) with newly diagnosed untreated stage III/IV OC who underwent either primary cytoreductive surgery (PCS) with gross residual disease or neoadjuvant chemotherapy (NACT) and interval surgery. Eligible pts were randomised 1:1 to atezo 1200 mg or pbo cycles 1–22, with paclitaxel 175 mg/m2 + carboplatin AUC6 cycles 1–6 + bev 15 mg/kg cycles 2–22 (PCS pts), omitting peri-operative bev in NACT pts. Cycles were repeated q3w. Stratification factors were: stage (III vs IV), ECOG PS (0 vs 1/2), PD-L1 staining in immune cells (IC <1% vs ≥1% [PD-L1+]) and treatment strategy (PCS vs NACT). The co-primary endpoints were investigator-assessed progression-free survival (PFS; RECIST v1.1) and overall survival (OS) in the intent-to-treat (ITT) and PD-L1+ populations. PFS was tested in parallel in the two populations; OS was tested hierarchically. Of 1301 enrolled pts, ∼25% received NACT. At the data cut-off (30 Mar 2020) median follow-up was ∼20 months in both arms. There was no statistically significant PFS improvement in either the ITT population (HR 0.92 [95% CI 0.79–1.07]; median 18.4 months with pbo vs 19.5 months with atezo) or the PD-L1+ population (HR 0.80 [0.65–0.99], median 18.5 vs 20.8 months, respectively). Exploratory PFS analyses in the PD-L1 IC ≥5% subgroup showed a trend favouring atezo. Though immature, first interim OS results did not show significant benefit from atezo. Similar proportions discontinued any study treatment for AEs (22% pbo vs 26% atezo pts). The safety profile of atezo + bev + chemotherapy was consistent with expected AEs. Atezo did not significantly improve PFS in the ITT or PD-L1+ population. The combination was generally well tolerated with manageable AEs. Exploratory biomarker subgroup analyses are ongoing.

Abstract PR11: Tumor cell-intrinsic factors underlie the heterogeneity of immune infiltration and response to immunotherapy in pancreatic cancer

Abstract Intertumoral heterogeneity—the biologic and functional differences among different individual tumors—poses a challenge for immunotherapy. To understand the tumor cell-intrinsic factors underlying the heterogeneity of tumor immunity and sensitivity to immunotherapy, we established a new experimental system by generating a library of congenic pancreatic tumor cell clones from a genetic mouse model driven by mutant Kras and p53. These tumor cell clones robustly formed implanted tumors that recapitulated the T cell-inflamed and non-T cell-inflamed tumor microenvironments in human patients, associated with distinct patterns of infiltration by T cells and myeloid cells. We found that the non-T cell-inflamed phenotype was dominant over the T cell-inflamed phenotype in the local tumor microenvironment. Both quantitative and qualitative features, specifically expression of markers of prior TCR activation, of intratumoral CD8+ T cells predicted the response to immunotherapies. An integrated transcriptomic and epigenetic analysis revealed that tumor cell-intrinsic expression of the chemokine CXCL1 as a major determinant of the non-T cell-inflamed microenvironment, and ablation of tumor cell-intrinsic CXCL1, promoted T-cell infiltration and sensitivity to a combination of chemotherapies, CD40 agonist, and checkpoint blockades. These results demonstrated that heterogeneity of tumor immune phenotypes is driven by tumor cell-intrinsic factors that can be manipulated to influence the outcome of immunotherapies. The observation that non-T cell-inflamed phenotype is dominant emphasized the importance of targeting mechanisms driving T-cell low phenotype for improving immunotherapy response. This experimental system will provide opportunities for understanding other aspects of tumor heterogeneity as well. This abstract is also being presented as Poster A45. Citation Format: Jinyang Li, Katelyn T. Byrne, Fangxue Yan, Taiji Yamazoe, Zeyu Chen, Timour Baslan, Lee P. Richman, Jeffrey Lin, Yu H. Sun, Shannon M. Liudahl, John W. Tobias, Scott Lowe, Lisa M. Coussens, John E Wherry, Robert H. Vonderheide, Ben Z. Stanger. Tumor cell-intrinsic factors underlie the heterogeneity of immune infiltration and response to immunotherapy in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2018 Nov 27-30; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(4 Suppl):Abstract nr PR11.