InVivoMAb polyclonal Syrian hamster IgG

Catalog #BE0087
Product Citations:
39
Clone:
Polyclonal

$164.00 - $4,280.00

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Product Details

The polyclonal Syrian hamster IgG is purified from Syrian hamster serum. It is ideal for use as a non-reactive control IgG for Syrian hamster antibodies in most in vivo and in vitro applications.

Specifications

Isotype Syrian hamster IgG
Recommended Dilution Buffer InVivoPure pH 7.0 Dilution Buffer
Conjugation This product is unconjugated. Conjugation is available via our Antibody Conjugation Services.
Formulation PBS, pH 7.0
Contains no stabilizers or preservatives
Endotoxin <2EU/mg (<0.002EU/μg)
Determined by LAL gel clotting assay
Purity >95%
Determined by SDS-PAGE
Sterility 0.2 µm filtration
Purification Protein G
RRID AB_1107782
Molecular Weight 150 kDa
Murine Pathogen Tests Ectromelia/Mousepox Virus: Negative
Hantavirus: Negative
K Virus: Negative
Lactate Dehydrogenase-Elevating Virus: Negative
Lymphocytic Choriomeningitis virus: Negative
Mouse Adenovirus: Negative
Mouse Cytomegalovirus: Negative
Mouse Hepatitis Virus: Negative
Mouse Minute Virus: Negative
Mouse Norovirus: Negative
Mouse Parvovirus: Negative
Mouse Rotavirus: Negative
Mycoplasma Pulmonis: Negative
Pneumonia Virus of Mice: Negative
Polyoma Virus: Negative
Reovirus Screen: Negative
Sendai Virus: Negative
Theiler’s Murine Encephalomyelitis: Negative
Storage The antibody solution should be stored at the stock concentration at 4°C. Do not freeze.

Additional Formats

Bradley, T., et al. (2020). "Immune checkpoint modulation enhances HIV-1 antibody induction" Nat Commun 11(1): 948. PubMed

Eliciting protective titers of HIV-1 broadly neutralizing antibodies (bnAbs) is a goal of HIV-1 vaccine development, but current vaccine strategies have yet to induce bnAbs in humans. Many bnAbs isolated from HIV-1-infected individuals are encoded by immunoglobulin gene rearrangments with infrequent naive B cell precursors and with unusual genetic features that may be subject to host regulatory control. Here, we administer antibodies targeting immune cell regulatory receptors CTLA-4, PD-1 or OX40 along with HIV envelope (Env) vaccines to rhesus macaques and bnAb immunoglobulin knock-in (KI) mice expressing diverse precursors of CD4 binding site HIV-1 bnAbs. CTLA-4 blockade augments HIV-1 Env antibody responses in macaques, and in a bnAb-precursor mouse model, CTLA-4 blocking or OX40 agonist antibodies increase germinal center B and T follicular helper cells and plasma neutralizing antibodies. Thus, modulation of CTLA-4 or OX40 immune checkpoints during vaccination can promote germinal center activity and enhance HIV-1 Env antibody responses.

Klepsch, V., et al. (2020). "Targeting the orphan nuclear receptor NR2F6 in T cells primes tumors for immune checkpoint therapy" Cell Commun Signal 18(1): 8. PubMed

BACKGROUND: NR2F6 has been proposed as an alternative cancer immune checkpoint in the effector T cell compartment. However, a realistic assessment of the in vivo therapeutic potential of NR2F6 requires acute depletion. METHODS: Employing primary T cells isolated from Cas9-transgenic mice for electroporation of chemically synthesized sgRNA, we established a CRISPR/Cas9-mediated acute knockout protocol of Nr2f6 in primary mouse T cells. RESULTS: Analyzing these Nr2f6(CRISPR/Cas9 knockout) T cells, we reproducibly observed a hyper-reactive effector phenotype upon CD3/CD28 stimulation in vitro, highly reminiscent to Nr2f6(-/-) T cells. Importantly, CRISPR/Cas9-mediated Nr2f6 ablation prior to adoptive cell therapy (ACT) of autologous polyclonal T cells into wild-type tumor-bearing recipient mice in combination with PD-L1 or CTLA-4 tumor immune checkpoint blockade significantly delayed MC38 tumor progression and induced superior survival, thus further validating a T cell-inhibitory function of NR2F6 during tumor progression. CONCLUSIONS: These findings indicate that Nr2f6(CRISPR/Cas9 knockout) T cells are comparable to germline Nr2f6(-/-) T cells, a result providing an independent confirmation of the immune checkpoint function of lymphatic NR2F6. Taken together, CRISPR/Cas9-mediated acute Nr2f6 gene ablation in primary mouse T cells prior to ACT appeared feasible for potentiating established PD-L1 and CTLA-4 blockade therapies, thereby pioneering NR2F6 inhibition as a sensitizing target for augmented tumor regression. Video abstract.

Wang, Q., et al. (2019). "Single-cell profiling guided combinatorial immunotherapy for fast-evolving CDK4/6 inhibitor-resistant HER2-positive breast cancer" Nat Commun 10(1): 3817. PubMed

Acquired resistance to targeted cancer therapy is a significant clinical challenge. In parallel with clinical trials combining CDK4/6 inhibitors to treat HER2+ breast cancer, we sought to prospectively model tumor evolution in response to this regimen in vivo and identify a clinically actionable strategy to combat drug resistance. Despite a promising initial response, acquired resistance emerges rapidly to the combination of anti-HER2/neu antibody and CDK4/6 inhibitor Palbociclib. Using high-throughput single-cell profiling over the course of treatments, we reveal a distinct immunosuppressive immature myeloid cell (IMC) population to infiltrate the resistant tumors. Guided by single-cell transcriptome analysis, we demonstrate that combination of IMC-targeting tyrosine kinase inhibitor cabozantinib and immune checkpoint blockade enhances anti-tumor immunity, and overcomes the resistance. Furthermore, sequential combinatorial immunotherapy enables a sustained control of the fast-evolving CDK4/6 inhibitor-resistant tumors. Our study demonstrates a translational framework for treating rapidly evolving tumors through preclinical modeling and single-cell analyses.

Su, W., et al. (2019). "The Polycomb Repressor Complex 1 Drives Double-Negative Prostate Cancer Metastasis by Coordinating Stemness and Immune Suppression" Cancer Cell 36(2): 139-155.e110. PubMed

The mechanisms that enable immune evasion at metastatic sites are poorly understood. We show that the Polycomb Repressor Complex 1 (PRC1) drives colonization of the bones and visceral organs in double-negative prostate cancer (DNPC). In vivo genetic screening identifies CCL2 as the top prometastatic gene induced by PRC1. CCL2 governs self-renewal and induces the recruitment of M2-like tumor-associated macrophages and regulatory T cells, thus coordinating metastasis initiation with immune suppression and neoangiogenesis. A catalytic inhibitor of PRC1 cooperates with immune checkpoint therapy to reverse these processes and suppress metastasis in genetically engineered mouse transplantation models of DNPC. These results reveal that PRC1 coordinates stemness with immune evasion and neoangiogenesis and point to the potential clinical utility of targeting PRC1 in DNPC.

Binnewies, M., et al. (2019). "Unleashing Type-2 Dendritic Cells to Drive Protective Antitumor CD4(+) T Cell Immunity" Cell 177(3): 556-571.e516. PubMed

Differentiation of proinflammatory CD4(+) conventional T cells (T(conv)) is critical for productive antitumor responses yet their elicitation remains poorly understood. We comprehensively characterized myeloid cells in tumor draining lymph nodes (tdLN) of mice and identified two subsets of conventional type-2 dendritic cells (cDC2) that traffic from tumor to tdLN and present tumor-derived antigens to CD4(+) T(conv), but then fail to support antitumor CD4(+) T(conv) differentiation. Regulatory T cell (T(reg)) depletion enhanced their capacity to elicit strong CD4(+) T(conv) responses and ensuing antitumor protection. Analogous cDC2 populations were identified in patients, and as in mice, their abundance relative to T(reg) predicts protective ICOS(+) PD-1(lo) CD4(+) T(conv) phenotypes and survival. Further, in melanoma patients with low T(reg) abundance, intratumoral cDC2 density alone correlates with abundant CD4(+) T(conv) and with responsiveness to anti-PD-1 therapy. Together, this highlights a pathway that restrains cDC2 and whose reversal enhances CD4(+) T(conv) abundance and controls tumor growth.

    • Mus musculus (House mouse)
    • ,
    • Cancer Research
    • ,
    • Immunology and Microbiology
    CD4 T cell-activating neoantigens enhance personalized cancer vaccine efficacy.

    In JCI Insight on 8 December 2023 by Huff, A. L., Longway, G., et al.

    PubMed

    Personalized cancer vaccines aim to activate and expand cytotoxic antitumor CD8+ T cells to recognize and kill tumor cells. However, the role of CD4+ T cell activation in the clinical benefit of these vaccines is not well defined. We previously established a personalized neoantigen vaccine (PancVAX) for the pancreatic cancer cell line Panc02, which activates tumor-specific CD8+ T cells but required combinatorial checkpoint modulators to achieve therapeutic efficacy. To determine the effects of neoantigen-specific CD4+ T cell activation, we generated a vaccine (PancVAX2) targeting both major histocompatibility complex class I- (MHCI-) and MHCII-specific neoantigens. Tumor-bearing mice vaccinated with PancVAX2 had significantly improved control of tumor growth and long-term survival benefit without concurrent administration of checkpoint inhibitors. PancVAX2 significantly enhanced priming and recruitment of neoantigen-specific CD8+ T cells into the tumor with lower PD-1 expression after reactivation compared with the CD8+ vaccine alone. Vaccine-induced neoantigen-specific Th1 CD4+ T cells in the tumor were associated with decreased Tregs. Consistent with this, PancVAX2 was associated with more proimmune myeloid-derived suppressor cells and M1-like macrophages in the tumor, demonstrating a less immunosuppressive tumor microenvironment. This study demonstrates the biological importance of prioritizing and including CD4+ T cell-specific neoantigens for personalized cancer vaccine modalities.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    Differential requirements for CD4+ T cells in the efficacy of the anti-PD-1+LAG-3 and anti-PD-1+CTLA-4 combinations in melanoma flank and brain metastasis models.

    In Journal for Immunotherapy of Cancer on 6 December 2023 by Phadke, M. S., Li, J., et al.

    PubMed

    Although the anti-PD-1+LAG-3 and the anti-PD-1+CTLA-4 combinations are effective in advanced melanoma, it remains unclear whether their mechanisms of action overlap. We used single cell (sc) RNA-seq, flow cytometry and IHC analysis of responding SM1, D4M-UV2 and B16 melanoma flank tumors and SM1 brain metastases to explore the mechanism of action of the anti-PD-1+LAG-3 and the anti-PD-1+CTLA-4 combination. CD4+ and CD8+ T cell depletion, tetramer binding assays and ELISPOT assays were used to demonstrate the unique role of CD4+T cell help in the antitumor effects of the anti-PD-1+LAG-3 combination. The anti-PD-1+CTLA-4 combination was associated with the infiltration of FOXP3+regulatory CD4+ cells (Tregs), fewer activated CD4+T cells and the accumulation of a subset of IFNγ secreting cytotoxic CD8+T cells, whereas the anti-PD-1+LAG-3 combination led to the accumulation of CD4+T helper cells that expressed CXCR4, TNFSF8, IL21R and a subset of CD8+T cells with reduced expression of cytotoxic markers. T cell depletion studies showed a requirement for CD4+T cells for the anti-PD-1+LAG-3 combination, but not the PD-1-CTLA-4 combination at both flank and brain tumor sites. In anti-PD-1+LAG-3 treated tumors, CD4+T cell depletion was associated with fewer activated (CD69+) CD8+T cells and impaired IFNγ release but, conversely, increased numbers of activated CD8+T cells and IFNγ release in anti-PD-1+CTLA-4 treated tumors. Together these studies suggest that these two clinically relevant immune checkpoint inhibitor (ICI) combinations have differential effects on CD4+T cell polarization, which in turn, impacted cytotoxic CD8+T cell function. Further insights into the mechanisms of action/resistance of these clinically-relevant ICI combinations will allow therapy to be further personalized. © Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.

    • Mus musculus (House mouse)
    • ,
    • Immunology and Microbiology
    Local and distant response to intratumoral immunotherapy assessed by immunoPET in mice.

    In Journal for Immunotherapy of Cancer on 1 November 2023 by Meyblum, L., Chevaleyre, C., et al.

    PubMed

    Despite the promising efficacy of immune checkpoint blockers (ICB), tumor resistance and immune-related adverse events hinder their success in cancer treatment. To address these challenges, intratumoral delivery of immunotherapies has emerged as a potential solution, aiming to mitigate side effects through reduced systemic exposure while increasing effectiveness by enhancing local bioavailability. However, a comprehensive understanding of the local and systemic distribution of ICBs following intratumoral administration, as well as their impact on distant tumors, remains crucial for optimizing their therapeutic potential.To comprehensively investigate the distribution patterns following the intratumoral and intravenous administration of radiolabeled anti-cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and to assess its corresponding efficacy in both injected and non-injected tumors, we conducted an immunoPET imaging study. CT26 and MC38 syngeneic colorectal tumor cells were implanted subcutaneously on both flanks of Balb/c and C57Bl/6 mice, respectively. Hamster anti-mouse CTLA-4 antibody (9H10) labeled with zirconium-89 ([89Zr]9H10) was intratumorally or intravenously administered. Whole-body distribution of the antibody was monitored by immunoPET imaging (n=12 CT26 Balb/c mice, n=10 MC38 C57Bl/6 mice). Tumorous responses to injected doses (1-10 mg/kg) were correlated with specific uptake of [89Zr]9H10 (n=24). Impacts on the tumor microenvironment were assessed by immunofluorescence and flow cytometry. Half of the dose was cleared into the blood 1 hour after intratumoral administration. Despite this, 7 days post-injection, 6-8% of the dose remained in the intratumoral-injected tumors. CT26 tumors with prolonged ICB exposure demonstrated complete responses. Seven days post-injection, the contralateral non-injected tumor uptake of the ICB was comparable to the one achieved through intravenous administration (7.5±1.7% ID.cm-3 and 7.6±2.1% ID.cm-3, respectively) at the same dose in the CT26 model. This observation was confirmed in the MC38 model. Consistent intratumoral pharmacodynamic effects were observed in both intratumoral and intravenous treatment groups, as evidenced by a notable increase in CD8+T cells within the CT26 tumors following treatment. ImmunoPET-derived pharmacokinetics supports intratumoral injection of ICBs to decrease systemic exposure while maintaining efficacy compared with intravenous. Intratumoral-ICBs lead to high local drug exposure while maintaining significant therapeutic exposure in non-injected tumors. This immunoPET approach is applicable for clinical practice to support evidence-based drug development. © Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    The GPCR-Gαs-PKA signaling axis promotes T cell dysfunction and cancer immunotherapy failure.

    In Nature Immunology on 1 August 2023 by Wu, V. H., Yung, B. S., et al.

    PubMed

    Immune checkpoint blockade (ICB) targeting PD-1 and CTLA-4 has revolutionized cancer treatment. However, many cancers do not respond to ICB, prompting the search for additional strategies to achieve durable responses. G-protein-coupled receptors (GPCRs) are the most intensively studied drug targets but are underexplored in immuno-oncology. Here, we cross-integrated large singe-cell RNA-sequencing datasets from CD8+ T cells covering 19 distinct cancer types and identified an enrichment of Gαs-coupled GPCRs on exhausted CD8+ T cells. These include EP2, EP4, A2AR, β1AR and β2AR, all of which promote T cell dysfunction. We also developed transgenic mice expressing a chemogenetic CD8-restricted Gαs-DREADD to activate CD8-restricted Gαs signaling and show that a Gαs-PKA signaling axis promotes CD8+ T cell dysfunction and immunotherapy failure. These data indicate that Gαs-GPCRs are druggable immune checkpoints that might be targeted to enhance the response to ICB immunotherapies. © 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.

    • Cancer Research
    • ,
    • Genetics
    METTL1 promotes tumorigenesis through tRNA-derived fragment biogenesis in prostate cancer.

    In Molecular Cancer on 29 July 2023 by García-Vílchez, R., Añazco-Guenkova, A. M., et al.

    PubMed

    Newly growing evidence highlights the essential role that epitranscriptomic marks play in the development of many cancers; however, little is known about the role and implications of altered epitranscriptome deposition in prostate cancer. Here, we show that the transfer RNA N7-methylguanosine (m7G) transferase METTL1 is highly expressed in primary and advanced prostate tumours. Mechanistically, we find that METTL1 depletion causes the loss of m7G tRNA methylation and promotes the biogenesis of a novel class of small non-coding RNAs derived from 5'tRNA fragments. 5'tRNA-derived small RNAs steer translation control to favour the synthesis of key regulators of tumour growth suppression, interferon pathway, and immune effectors. Knockdown of Mettl1 in prostate cancer preclinical models increases intratumoural infiltration of pro-inflammatory immune cells and enhances responses to immunotherapy. Collectively, our findings reveal a therapeutically actionable role of METTL1-directed m7G tRNA methylation in cancer cell translation control and tumour biology. © 2023. The Author(s).

    • In Vivo
    • ,
    • Mus musculus (House mouse)
    • ,
    • Cancer Research
    Lymphocyte networks are dynamic cellular communities in the immunoregulatory landscape of lung adenocarcinoma.

    In Cancer Cell on 8 May 2023 by Gaglia, G., Burger, M. L., et al.

    PubMed

    Lymphocytes are key for immune surveillance of tumors, but our understanding of the spatial organization and physical interactions that facilitate lymphocyte anti-cancer functions is limited. We used multiplexed imaging, quantitative spatial analysis, and machine learning to create high-definition maps of lung tumors from a Kras/Trp53-mutant mouse model and human resections. Networks of interacting lymphocytes ("lymphonets") emerged as a distinctive feature of the anti-cancer immune response. Lymphonets nucleated from small T cell clusters and incorporated B cells with increasing size. CXCR3-mediated trafficking modulated lymphonet size and number, but T cell antigen expression directed intratumoral localization. Lymphonets preferentially harbored TCF1+ PD-1+ progenitor CD8+ T cells involved in responses to immune checkpoint blockade (ICB) therapy. Upon treatment of mice with ICB or an antigen-targeted vaccine, lymphonets retained progenitor and gained cytotoxic CD8+ T cell populations, likely via progenitor differentiation. These data show that lymphonets create a spatial environment supportive of CD8+ T cell anti-tumor responses. Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    Addressing Tumor Heterogeneity by Sensitizing Resistant Cancer Cells to T cell-Secreted Cytokines.

    In Cancer Discovery on 4 May 2023 by Ito, Y., Pan, D., et al.

    PubMed

    Tumor heterogeneity is a major barrier to cancer therapy, including immunotherapy. Activated T cells can efficiently kill tumor cells following recognition of MHC class I (MHC-I)-bound peptides, but this selection pressure favors outgrowth of MHC-I-deficient tumor cells. We performed a genome-scale screen to discover alternative pathways for T cell-mediated killing of MHC-I-deficient tumor cells. Autophagy and TNF signaling emerged as top pathways, and inactivation of Rnf31 (TNF signaling) and Atg5 (autophagy) sensitized MHC-I-deficient tumor cells to apoptosis by T cell-derived cytokines. Mechanistic studies demonstrated that inhibition of autophagy amplified proapoptotic effects of cytokines in tumor cells. Antigens from apoptotic MHC-I-deficient tumor cells were efficiently cross-presented by dendritic cells, resulting in heightened tumor infiltration by IFNγ-and TNFα-producing T cells. Tumors with a substantial population of MHC-I-deficient cancer cells could be controlled by T cells when both pathways were targeted using genetic or pharmacologic approaches. Tumor heterogeneity is a major barrier to immunotherapy. We show that MHC-I-deficient tumor cells are forced into apoptosis by T cell-derived cytokines when TNF signaling and autophagy pathways are targeted. This approach enables T cell-mediated elimination of tumors with a substantial population of resistant, MHC-I-deficient tumor cells. This article is highlighted in the In This Issue feature, p. 1027. ©2023 American Association for Cancer Research.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    Targeting CXCL16 and STAT1 augments immune checkpoint blockade therapy in triple-negative breast cancer.

    In Nature Communications on 13 April 2023 by Palakurthi, B., Fross, S. R., et al.

    PubMed

    Chemotherapy prior to immune checkpoint blockade (ICB) treatment appears to improve ICB efficacy but resistance to ICB remains a clinical challenge and is attributed to highly plastic myeloid cells associating with the tumor immune microenvironment (TIME). Here we show by CITE-seq single-cell transcriptomic and trajectory analyses that neoadjuvant low-dose metronomic chemotherapy (MCT) leads to a characteristic co-evolution of divergent myeloid cell subsets in female triple-negative breast cancer (TNBC). Specifically, we identify that the proportion of CXCL16 + myeloid cells increase and a high STAT1 regulon activity distinguishes Programmed Death Ligand 1 (PD-L1) expressing immature myeloid cells. Chemical inhibition of STAT1 signaling in MCT-primed breast cancer sensitizes TNBC to ICB treatment, which underscores the STAT1's role in modulating TIME. In summary, we leverage single-cell analyses to dissect the cellular dynamics in the tumor microenvironment (TME) following neoadjuvant chemotherapy and provide a pre-clinical rationale for modulating STAT1 in combination with anti-PD-1 for TNBC patients. © 2023. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.

    • Mus musculus (House mouse)
    • ,
    • Cancer Research
    • ,
    • Immunology and Microbiology
    Antigen-presenting type-I conventional dendritic cells facilitate curative checkpoint blockade immunotherapy in pancreatic cancer

    Preprint on BioRxiv : the Preprint Server for Biology on 6 March 2023 by Mahadevan, K. K., Dyevoich, A. M., et al.

    PubMed

    Inflammation and tissue damage associated with pancreatitis can precede or occur concurrently with pancreatic ductal adenocarcinoma (PDAC). We demonstrate that in PDAC coupled with pancreatitis (ptPDAC), antigen-presenting type-I conventional dendritic cells (cDC1s) are specifically activated. Immune checkpoint blockade therapy (iCBT) leads to cytotoxic CD8 + T cell activation and eradication of ptPDAC with restoration of lifespan even upon PDAC re-challenge. Such eradication of ptPDAC was reversed following specific depletion of dendritic cells. Employing PDAC antigen-loaded cDC1s as a vaccine, immunotherapy-resistant PDAC was rendered sensitive to iCBT with a curative outcome. Analysis of the T-cell receptor (TCR) sequences in the tumor infiltrating CD8 + T cells following cDC1 vaccination coupled with iCBT identified unique CDR3 sequences with potential therapeutic significance. Our findings identify a fundamental difference in the immune microenvironment and adaptive immune response in PDAC concurrent with, or without pancreatitis, and provides a rationale for combining cDC1 vaccination with iCBT as a potential treatment option.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    Oncogenic KrasG12Dspecific non-covalent inhibitor reprograms tumor microenvironment to prevent and reverse early pre-neoplastic pancreatic lesions and in combination with immunotherapy regresses advanced PDAC in a CD8

    Preprint on BioRxiv : the Preprint Server for Biology on 16 February 2023 by Mahadevan, K. K., McAndrews, K. M., et al.

    PubMed

    Pancreatic ductal adenocarcinoma (PDAC) is associated with mutations in Kras, a known oncogenic driver of PDAC; and the KRAS G12D mutation is present in nearly half of PDAC patients. Recently, a non-covalent small molecule inhibitor (MRTX1133) was identified with specificity to the Kras G12D mutant protein. Here we explore the impact of Kras G12D inhibition by MRTX1133 on advanced PDAC and its influence on the tumor microenvironment. Employing different orthotopic xenograft and syngeneic tumor models, eight different PDXs, and two different autochthonous genetic models, we demonstrate that MRTX1133 reverses early PDAC growth, increases intratumoral CD8 + effector T cells, decreases myeloid infiltration, and reprograms cancer associated fibroblasts. Autochthonous genetic mouse models treated with MRTX1133 leads to regression of both established PanINs and advanced PDAC. Regression of advanced PDAC requires CD8 + T cells and immune checkpoint blockade therapy (iCBT) synergizes with MRTX1133 to eradicate PDAC and prolong overall survival. Mechanistically, inhibition of mutant Kras in advanced PDAC and human patient derived organoids (PDOs) induces Fas expression in cancer cells and facilitates CD8 + T cell mediated death. These results demonstrate the efficacy of MRTX1133 in different mouse models of PDAC associated with reprogramming of stromal fibroblasts and a dependency on CD8 + T cell mediated tumor clearance. Collectively, this study provides a rationale for a synergistic combination of MRTX1133 with iCBT in clinical trials.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    Durable Responses to Anti-PD1 and Anti-CTLA4 in a Preclinical Model of Melanoma Displaying Key Immunotherapy Response Biomarkers.

    In Cancers on 3 October 2022 by Shklovskaya, E., Pedersen, B., et al.

    PubMed

    Immunotherapy has transformed the management of patients with advanced melanoma, with five-year overall survival rates reaching 52% for combination immunotherapies blocking the cytotoxic T-lymphocyte-associated antigen-4 (CTLA4) and programmed cell death-1 (PD1) immune axes. Yet, our understanding of local and systemic determinants of immunotherapy response and resistance is restrained by the paucity of preclinical models, particularly those for anti-PD1 monotherapy. We have therefore generated a novel murine model of melanoma by integrating key immunotherapy response biomarkers into the model development workflow. The resulting YUMM3.3UVRc34 (BrafV600E; Cdkn2a-/-) model demonstrated high mutation burden and response to interferon (IFN)γ, including induced expression of antigen-presenting molecule MHC-I and the principal PD1 ligand PD-L1, consistent with phenotypes of human melanoma biopsies from patients subsequently responding to anti-PD1 monotherapy. Syngeneic immunosufficient mice bearing YUMM3.3UVRc34 tumors demonstrated durable responses to anti-PD1, anti-CTLA4, or combined treatment. Immunotherapy responses were associated with early on-treatment changes in the tumor microenvironment and circulating T-cell subsets, and systemic immunological memory underlying protection from tumor recurrence. Local and systemic immunological landscapes associated with immunotherapy response in the YUMM3.3UVRc34 melanoma model recapitulate immunotherapy responses observed in melanoma patients and identify discrete immunological mechanisms underlying the durability of responses to anti-PD1 and anti-CTLA4 treatments.

    • Immunology and Microbiology
    Allelic polymorphism controls autoreactivity and vaccine elicitation of human broadly neutralizing antibodies against influenza virus.

    In Immunity on 13 September 2022 by Sangesland, M., Torrents de la Peña, A., et al.

    PubMed

    Human broadly neutralizing antibodies (bnAbs) targeting the hemagglutinin stalk of group 1 influenza A viruses (IAVs) are biased for IGHV1-69 alleles that use phenylalanine (F54) but not leucine (L54) within their CDRH2 loops. Despite this, we demonstrated that both alleles encode for human IAV bnAbs that employ structurally convergent modes of contact to the same epitope. To resolve differences in lineage expandability, we compared F54 versus L54 as substrate within humanized mice, where antibodies develop with human-like CDRH3 diversity but are restricted to single VH genes. While both alleles encoded for bnAb precursors, only F54 IGHV1-69 supported elicitation of heterosubtypic serum bnAbs following immunization with a stalk-only nanoparticle vaccine. L54 IGHV1-69 was unproductive, co-encoding for anergic B cells and autoreactive stalk antibodies that were cleared from B cell memory. Moreover, human stalk antibodies also demonstrated L54-dependent autoreactivity. Therefore, IGHV1-69 polymorphism, which is skewed ethnically, gates tolerance and vaccine expandability of influenza bnAbs. Copyright © 2022 Elsevier Inc. All rights reserved.

    • IHC
    • ,
    • Mus musculus (House mouse)
    • ,
    • Cancer Research
    • ,
    • Immunology and Microbiology
    Interleukin-6 blockade abrogates immunotherapy toxicity and promotes tumor immunity.

    In Cancer Cell on 9 May 2022 by Hailemichael, Y., Johnson, D. H., et al.

    PubMed

    Immune checkpoint blockade (ICB) therapy frequently induces immune-related adverse events. To elucidate the underlying immunobiology, we performed a deep immune analysis of intestinal, colitis, and tumor tissue from ICB-treated patients with parallel studies in preclinical models. Expression of interleukin-6 (IL-6), neutrophil, and chemotactic markers was higher in colitis than in normal intestinal tissue; T helper 17 (Th17) cells were more prevalent in immune-related enterocolitis (irEC) than T helper 1 (Th1). Anti-cytotoxic T-lymphocyte-associated antigen 4 (anti-CTLA-4) induced stronger Th17 memory in colitis than anti-program death 1 (anti-PD-1). In murine models, IL-6 blockade associated with improved tumor control and a higher density of CD4+/CD8+ effector T cells, with reduced Th17, macrophages, and myeloid cells. In an experimental autoimmune encephalomyelitis (EAE) model with tumors, combined IL-6 blockade and ICB enhanced tumor rejection while simultaneously mitigating EAE symptoms versus ICB alone. IL-6 blockade with ICB could de-couple autoimmunity from antitumor immunity. Copyright © 2022 Elsevier Inc. All rights reserved.

    • Mus musculus (House mouse)
    • ,
    • Cancer Research
    • ,
    • Immunology and Microbiology
    Nanoliposome C6-Ceramide in combination with anti-CTLA4 antibody improves anti-tumor immunity in hepatocellular cancer.

    In The FASEB Journal on 1 April 2022 by Qi, X., Wu, F., et al.

    PubMed

    Combination therapy represents an effective therapeutic approach to overcome hepatocellular cancer (HCC) resistance to immune checkpoint blockade (ICB). Based upon previous work demonstrating that nanoliposome C6-ceramide (LipC6) not only induces HCC apoptosis but also prevents HCC-induced immune tolerance, we now investigate the potential of LipC6 in combination with ICB in HCC treatment. We generated orthotopic HCC-bearing mice, which have typical features in common with human patients, and then treated them with LipC6 in combination with the antibodies (Abs) for programmed cell death protein 1 (PD-1) or cytotoxic T-lymphocyte antigen 4 (CTLA4). The tumor growth was monitored by magnetic resonance imaging (MRI) and the intrahepatic immune profiles were checked by flow cytometry in response to the treatments. Realtime PCR (qPCR) was used to detect the expression of target genes. The results show that LipC6 in combination with anti-CTLA4 Ab, but not anti-PD-1 Ab, significantly slowed tumor growth, enhanced tumor-infiltrating CD8+ T cells, and suppressed tumor-resident CD4+ CD25+ FoxP3+ Tregs. Further molecular investigation indicates that the combinational treatment suppressed transcriptional factor Krüppel-like Factor 2 (KLF2), forkhead box protein P3 (FoxP3), and CTLA4. Our studies suggest that LipC6 in combination with anti-CTLA4 Ab represents a novel therapeutic approach with significant potential in activating anti-HCC immune response and suppressing HCC growth. © 2022 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.

    • Mus musculus (House mouse)
    • ,
    • Cancer Research
    • ,
    • Immunology and Microbiology
    Maximizing the Anti-tumor Potential of Immune Checkpoint Blockade through Modulation of Myeloid-specific CXCL16 and STAT1 Signaling

    Preprint on BioRxiv : the Preprint Server for Biology on 28 March 2022 by Palakurthi, B., Guldner, I., et al.

    PubMed

    h4>ABSTRACT/h4> Sensitivity to immune checkpoint blockades (ICB) depends on the overall balance of immunogenic and immunosuppressive signals in the tumor immune microenvironment (TIME). Chemotherapy as an immunostimulatory strategy showed potential in improving ICB’s clinical efficacy. Yet, evolution of highly plastic tumor-associated myeloid cells hinders ICB’s potential to reach its full therapeutic potential. In this study, we leveraged single-cell transcriptomic and trajectory analyses to delineate TIME dynamics after chemotherapy priming. We found that metronomic chemotherapy (MCT) treatment led to an accelerated T cell exhaustion through CXCL16-mediated recruitment of peripheral immature myeloid cells and expansion of STAT1-driven PD-L1 expressing myeloid cells. Inhibiting STAT1 signaling in MCT-primed breast cancer relieved T cell exhaustion and significantly enhanced the efficacy of anti-PD-1 ICB treatment. Our study leveraged single-cell analyses to dissect the dynamics of breast cancer TIME and provides a pre-clinical rationale to translate the anti-STAT1 plus anti-PD-1 combinatorial immunotherapy regimen to maximize ICB’s efficacy. h4>Manuscript Summary/h4> Single-cell analyses on low dose chemotherapy primed breast tumor-associated immune cells demonstrates a parallel coexistence of immunogenic and immunosuppressive myeloid cell subsets. Modulating STAT1 signaling in the tumor microenvironment fine-tunes immunogenic and immunosuppressive balance and maximizes the anti-PD-1 immunotherapy efficacy in chemotherapy-primed breast cancer.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    • ,
    • FC/FACS
    • ,
    • Mus musculus (House mouse)
    Antigen dominance hierarchies shape TCF1+ progenitor CD8 T cell phenotypes in tumors.

    In Cell on 16 September 2021 by Burger, M. L., Cruz, A. M., et al.

    PubMed

    CD8 T cell responses against different tumor neoantigens occur simultaneously, yet little is known about the interplay between responses and its impact on T cell function and tumor control. In mouse lung adenocarcinoma, we found that immunodominance is established in tumors, wherein CD8 T cell expansion is predominantly driven by the antigen that most stably binds MHC. T cells responding to subdominant antigens were enriched for a TCF1+ progenitor phenotype correlated with response to immune checkpoint blockade (ICB) therapy. However, the subdominant T cell response did not preferentially benefit from ICB due to a dysfunctional subset of TCF1+ cells marked by CCR6 and Tc17 differentiation. Analysis of human samples and sequencing datasets revealed that CCR6+ TCF1+ cells exist across human cancers and are not correlated with ICB response. Vaccination eliminated CCR6+ TCF1+ cells and dramatically improved the subdominant response, highlighting a strategy to optimally engage concurrent neoantigen responses against tumors. Copyright © 2021 Elsevier Inc. All rights reserved.

    Chitinase 3-like-1 contributes to acetaminophen-induced liver injury by promoting hepatic platelet recruitment.

    In eLife on 10 June 2021 by Shan, Z., Li, L., et al.

    PubMed

    Hepatic platelet accumulation contributes to acetaminophen (APAP)-induced liver injury (AILI). However, little is known about the molecular pathways involved in platelet recruitment to the liver and whether targeting such pathways could attenuate AILI. Mice were fasted overnight before intraperitoneally (i.p.) injected with APAP at a dose of 210 mg/kg for male mice and 325 mg/kg for female mice. Platelets adherent to Kupffer cells were determined in both mice and patients overdosed with APAP. The impact of α-chitinase 3-like-1 (α-Chi3l1) on alleviation of AILI was determined in a therapeutic setting, and liver injury was analyzed. The present study unveiled a critical role of Chi3l1 in hepatic platelet recruitment during AILI. Increased Chi3l1 and platelets in the liver were observed in patients and mice overdosed with APAP. Compared to wild-type (WT) mice, Chil1-/- mice developed attenuated AILI with markedly reduced hepatic platelet accumulation. Mechanistic studies revealed that Chi3l1 signaled through CD44 on macrophages to induce podoplanin expression, which mediated platelet recruitment through C-type lectin-like receptor 2. Moreover, APAP treatment of Cd44-/- mice resulted in much lower numbers of hepatic platelets and liver injury than WT mice, a phenotype similar to that in Chil1-/- mice. Recombinant Chi3l1 could restore hepatic platelet accumulation and AILI in Chil1-/- mice, but not in Cd44-/- mice. Importantly, we generated anti-Chi3l1 monoclonal antibodies and demonstrated that they could effectively inhibit hepatic platelet accumulation and AILI. We uncovered the Chi3l1/CD44 axis as a critical pathway mediating APAP-induced hepatic platelet recruitment and tissue injury. We demonstrated the feasibility and potential of targeting Chi3l1 to treat AILI. ZS received funding from NSFC (32071129). FWL received funding from NIH (GM123261). ALFSG received funding from NIDDK (DK 058369). ZA received funding from CPRIT (RP150551 and RP190561) and the Welch Foundation (AU-0042-20030616). CJ received funding from NIH (DK122708, DK109574, DK121330, and DK122796) and support from a University of Texas System Translational STARs award. Portions of this work were supported with resources and the use of facilities of the Michael E. DeBakey VA Medical Center and funding from Department of Veterans Affairs I01 BX002551 (Equipment, Personnel, Supplies). The contents do not represent the views of the US Department of Veterans Affairs or the US Government.

    Chitinase 3-like-1 Contributes to Acetaminophen-induced Liver Injury by Promoting Hepatic Platelet Recruitment

    Preprint on BioRxiv : the Preprint Server for Biology on 9 April 2021 by Shan, Z., Li, L., et al.

    PubMed

    Hepatic platelet accumulation contributes to acetaminophen (APAP)-induced liver injury (AILI). However, little is known about the molecular pathways involved in platelet recruitment to the liver and whether targeting such pathways could attenuate AILI. The present study unveiled a critical role of chitinase 3-like-1 (Chi3l1) in hepatic platelet recruitment during AILI. Increased Chi3l1 and platelets in the liver were observed in patients and mice overdosed with APAP. Compared to wild-type (WT) mice, Chi3l1 -/- mice developed attenuated AILI with markedly reduced hepatic platelet accumulation. Mechanistic studies revealed that Chi3l1 signaled through CD44 on macrophages to induce podoplanin expression, which mediated platelet recruitment through C-type lectin-like receptor 2. Moreover, APAP treatment of CD44 -/- mice resulted in much lower numbers of hepatic platelets and liver injury than WT mice, a phenotype similar to that in Chi3l1 -/- mice. Recombinant Chi3l1 could restore hepatic platelet accumulation and AILI in Chi3l1 -/- mice, but not in CD44 -/- mice. Importantly, we generated anti-Chi3l1 monoclonal antibodies and demonstrated that they could effectively inhibit hepatic platelet accumulation and AILI. Overall, we uncovered the Chi3l1/CD44 axis as a critical pathway mediating APAP-induced hepatic platelet recruitment and tissue injury. We demonstrated the feasibility and potential of targeting Chi3l1 to treat AILI.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    Fascin inhibitor increases intratumoral dendritic cell activation and anti-cancer immunity.

    In Cell Reports on 6 April 2021 by Wang, Y., Song, M., et al.

    PubMed

    Fascin protein is the main actin-bundling protein in filopodia and invadopodia, which are critical for tumor cell migration, invasion, and metastasis. Small-molecule fascin inhibitors block tumor invasion and metastasis and increase the overall survival of tumor-bearing mice. Here, we report a finding that fascin blockade additionally reinvigorates anti-tumor immune response in syngeneic mouse models of various cancers. Fascin protein levels are increased in conventional dendritic cells (cDCs) in the tumor microenvironment. Mechanistically, fascin inhibitor NP-G2-044 increases the number of intratumoral-activated cDCs and enhances the antigen uptake by cDCs. Furthermore, together with PD-1 blocking antibody, NP-G2-044 markedly increases the number of activated CD8+ T cells in the otherwise anti-PD-1 refractory tumors. Reduction of fascin levels in cDCs, but not fascin gene knockout in tumor cells, mimics the anti-tumor immune effect of NP-G2-044. These data demonstrate that fascin inhibitor NP-G2-044 simultaneously limits tumor metastasis and reinvigorates anti-tumor immune responses. Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.

    • Cancer Research
    • ,
    • Immunology and Microbiology
    Combined BRAF, MEK, and CDK4/6 Inhibition Depletes Intratumoral Immune-Potentiating Myeloid Populations in Melanoma.

    In Cancer Immunology Research on 1 February 2021 by Lelliott, E. J., Mangiola, S., et al.

    PubMed

    Combined inhibition of BRAF, MEK, and CDK4/6 is currently under evaluation in clinical trials for patients with melanoma harboring a BRAFV600 mutation. While this triple therapy has potent tumor-intrinsic effects, the impact of this combination on antitumor immunity remains unexplored. Here, using a syngeneic BrafV600ECdkn2a-/-Pten-/- melanoma model, we demonstrated that triple therapy promoted durable tumor control through tumor-intrinsic mechanisms and promoted immunogenic cell death and T-cell infiltration. Despite this, tumors treated with triple therapy were unresponsive to immune checkpoint blockade (ICB). Flow cytometric and single-cell RNA sequencing analyses of tumor-infiltrating immune populations revealed that triple therapy markedly depleted proinflammatory macrophages and cross-priming CD103+ dendritic cells, the absence of which correlated with poor overall survival and clinical responses to ICB in patients with melanoma. Indeed, immune populations isolated from tumors of mice treated with triple therapy failed to stimulate T-cell responses ex vivo While combined BRAF, MEK, and CDK4/6 inhibition demonstrates favorable tumor-intrinsic activity, these data suggest that collateral effects on tumor-infiltrating myeloid populations may impact antitumor immunity. These findings have important implications for the design of combination strategies and clinical trials that incorporate BRAF, MEK, and CDK4/6 inhibition with immunotherapy for the treatment of patients with melanoma. ©2020 American Association for Cancer Research.

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