There Is Strength in Numbers: Quantitation of Fc Gamma Receptors on Murine Tissue-Resident Macrophages

Many of the effector functions of antibodies rely on the binding of antibodies/immune complexes to cellular Fcγ receptors (FcγRs). Since the majority of innate immune effector cells express both activating and inhibitory Fc receptors, the outcome of the binding of immune complexes to cells of a given population is influenced by the relative affinities of the respective IgG subclasses to these receptors, as well as by the numbers of activating and inhibitory FcγRs on the cell surface. A group of immune cells that has come into focus more recently is the various subsets of tissue-resident macrophages. The central functions of FcγRs on tissue macrophages include the clearance of opsonized pathogens, the removal of small immune complexes from the circulation and the depletion of antibody-opsonized cells in the therapy of autoimmunity and cancer. Despite these essential functions of FcγRs on tissue-resident macrophages, an in-depth quantification of FcγRs is lacking. Thus, the aim of our current study was to quantify the various Fcγ receptors on macrophages in murine liver, lung, kidney, brain, skin and spleen. Our study identified a pronounced heterogeneity between FcγR expression patterns of the different tissue macrophages, which may reflect their specialized functions within their unique niches in different organ environments.

671 Biological impacts of standard of care chemotherapies on immune effector cells from AML patients

BackgroundAcute myeloid leukemia (AML) is a heterogeneous group of malignant bone marrow diseases, characterized by massive and uncontrolled proliferation of myeloid precursor cells, which alters normal blood cell ratios. This disease is common to older adults and collectively displays one of the lowest 5-year overall survival rates (<25%) among all cancers, currently representing the deadliest form of leukemia. Improved treatments are clearly needed, and immunotherapies are attractive candidate therapies to explore.There are currently several standard chemotherapeutic treatment schemes for AML, which could be divided into two major groups: (1) cytotoxic chemotherapy (“7+3” or daunorubicin-cytarabine) and (2) hypomethylating agents (HMAs). HMAs include both 5-azacytidine and decitabine, which are cytidine analogs that inhibit DNA methyltransferase, resulting in the hypomethylation of DNA and inducing expression of silenced gene loci. Currently, HMAs are routinely delivered in combination with the Bcl-2 inhibitor venetoclax.The goals of this study are to determine how these standard first line therapies can affect the frequency and functional integrity of effector immune cells in patients' blood and establish when the phenotype and function of immune cells are restored to identify time windows when second line immunotherapies could be most effective.MethodsMore than 100 blood samples were obtained from 33 previously untreated AML patients. More than 50 measurable biomarkers were analyzed using 14-color flow cytometry to assess immune phenotypes of T and NK cells in peripheral blood of AML patients prior to treatment and at up to four timepoints after initiation of treatment with HMA or chemotherapy.ResultsWe found several significant changes in immune cell phenotype and function that occur in response to these therapies. Treatment with HMAs was strikingly less impactful on immune cells in patients compared to previously published in vitro studies. Nevertheless, HMA treatment increased perforin levels in T and NK cells, inhibited IFN-gamma secretion by CD8+ T cells, and changed expression of several checkpoint molecules. While chemotherapy caused fewer phenotypic changes it dramatically decreased the total number of immune cells. We also determined viable, functional and phenotypical recovery periods for immune effector cells after the treatments.ConclusionsOur results are important for introducing new second line immunotherapies to these chemotherapeutic regimens for treating AML and to improve overall understanding of immune cell behavior under conditions of anti-tumor treatment.AcknowledgementsSupported by grants from Janssen and the U.S./Israel Binational Science Foundation.Ethics ApprovalThe study was approved by the Fox Chase Cancer Center Institutional Review Board, approval number 17-8010, and all patients provided informed consent before taking part in the study.

Re-Expression of Poly/Oligo- Sialylated Adhesion Molecules on the Surface of Tumor Cells Disrupts Their Interaction with Immune-Effector Cells and Contributes to Pathophysiological Immune Escape

Glycans linked to surface proteins are the most complex biological macromolecules that play an active role in various cellular mechanisms. This diversity is the basis of cell–cell interaction and communication, cell growth, cell migration, as well as co-stimulatory or inhibitory signaling. Our review describes the importance of neuraminic acid and its derivatives as recognition elements, which are located at the outermost positions of carbohydrate chains linked to specific glycoproteins or glycolipids. Tumor cells, especially from solid tumors, mask themselves by re-expression of hypersialylated neural cell adhesion molecule (NCAM), neuropilin-2 (NRP-2), or synaptic cell adhesion molecule 1 (SynCAM 1) in order to protect themselves against the cytotoxic attack of the also highly sialylated immune effector cells. More particularly, we focus on α-2,8-linked polysialic acid chains, which characterize carrier glycoproteins such as NCAM, NRP-2, or SynCam-1. This characteristic property correlates with an aggressive clinical phenotype and endows them with multiple roles in biological processes that underlie all steps of cancer progression, including regulation of cell–cell and/or cell–extracellular matrix interactions, as well as increased proliferation, migration, reduced apoptosis rate of tumor cells, angiogenesis, and metastasis. Specifically, re-expression of poly/oligo-sialylated adhesion molecules on the surface of tumor cells disrupts their interaction with immune-effector cells and contributes to pathophysiological immune escape. Further, sialylated glycoproteins induce immunoregulatory cytokines and growth factors through interactions with sialic acid-binding immunoglobulin-like lectins. We describe the processes, which modulate the interaction between sialylated carrier glycoproteins and their ligands, and illustrate that sialic acids could be targets of novel therapeutic strategies for treatment of cancer and immune diseases.

Transcriptomic Correlates of Immunologic Activation in Head and Neck and Cervical Cancer

Targeting the immune system has emerged as an effective therapeutic strategy for the treatment of various tumor types, including Head and Neck Squamous Cell Carcinoma (HNSCC) and Non-small-Cell Lung Cancer (NSCLC), and checkpoint inhibitors have shown to improve patient survival in these tumor types. Unfortunately, not all cancers respond to these agents, making it necessary to identify responsive tumors. Several biomarkers of response have been described and clinically tested. As of yet what seems to be clear is that a pre-activation state of the immune system is necessary for these agents to be efficient. In this study, using established transcriptomic signatures, we identified a group of gene combination associated with favorable outcome in HNSCC linked to a higher presence of immune effector cells. CD2, CD3D, CD3E, and CXCR6 combined gene expression is associated with improved outcome of HNSCC patients and an increase of infiltrating immune effector cells. This new signature also identifies a subset of cervical squamous cell carcinoma (CSCC) patients with favorable prognosis, who show an increased presence of immune effector cells in the tumor, which outcome shows similarities with the HP-positive HNSCC cohort of patients. In addition, CD2, CD3D, CD3E, and CXCR6 signature is able to predict the best favorable prognosis in terms of overall survival of CSSC patients. Of note, these findings were not reproduced in other squamous cell carcinomas like esophageal SCC or lung SCC. Prospective confirmatory studies should be employed to validate these findings.

Combination Therapy with EGFR Specific CAR_NK_92 Cells and Cabozantinib against Human Renal Cell Carcinoma

Adaptive therapy using immune effector cells engineered by means of chimeric antigen receptors (CAR) has risen as a hopeful cancer management option. Despite their unprecedented success in haematological malignancies, CAR-modified T cells have shown limited efficacy in solid tumours, as the tumor’s immune-suppressive microenvironment inhibits CAR-modified immune effector cells’ functionality by different pathways, counting checkpoint receptor ligands expression like PD-L1 & recruitment tregs like suppressive immune cells. Receptor of epidermal growth factor (EGFR) could be the target of a ll-generation Chimeric antigen receptor T cellshat was transduced to NK-92 cell. In our research, we examined the antitumor efficacy of EGFR specific NK-92 (CAR-NK-92) cells using a xenograft mice model & in conjunction with tyrosine kinase inhibitor cabozantinib. We discovered that EGFR positive renal carcinoma cells (RCC) 786-0 and ACHN may specifically detect and activate CAR_NK_92 cells. They also displayed particular cytotoxicity against RCC in in vitro & in vivo models. Furthermore, we discovered that cabozantinib improves RCC-specific cytotoxicity by enhancing the expression of EGFR while reducing PD-L1 expression in RCC. Our research shows that CAR_NK_92 cells possess anti cancer therapeutic potential for EGFR-positive tumour cells, and that cabozantinib can boost CAR_NK_92 cell cytotoxicity when treated together.

IgA potentiates NETosis in response to viral infection

IgA is the second most abundant antibody present in circulation and is enriched at mucosal surfaces. As such, IgA plays a key role in protection against a variety of mucosal pathogens including viruses. In addition to neutralizing viruses directly, IgA can also stimulate Fc-dependent effector functions via engagement of Fc alpha receptors (Fc-αRI) expressed on the surface of certain immune effector cells. Neutrophils are the most abundant leukocyte, express Fc-αRI, and are often the first to respond to sites of injury and infection. Here, we describe a function for IgA–virus immune complexes (ICs) during viral infections. We show that IgA–virus ICs potentiate NETosis—the programmed cell-death pathway through which neutrophils release neutrophil extracellular traps (NETs). Mechanistically, IgA–virus ICs potentiated a suicidal NETosis pathway via engagement of Fc-αRI on neutrophils through a toll-like receptor–independent, NADPH oxidase complex–dependent pathway. NETs also were capable of trapping and inactivating viruses, consistent with an antiviral function.

IL-31, itch and hematological malignancies

Pruritus is one of the most common symptoms experienced by neoplastic patients. The pathogenesis of neoplastic itch is complex and multifactorial and could be due to an unbalanced production of humoral mediators by altered immune effector cells. IL-31 is a pro-inflammatory cytokine produced by CD4 + T helper cells. The aim of this review was to evaluate the role of this Th2 cytokine and its receptor IL-31RA, in the onset of neoplastic pruritus. We analysed scientific literature looking for the most relevant original articles linking IL-31to itch in oncologic diseases. Interleukin-31 seems to be a main itch mediator in several hematologic disease such as Cutaneous T cells lymphomas. In these patients IL-31 was positively linked to itch level, and IL-31 matched with disease stage. IL-31 seems to play an important role in the signalling pathway involved in pruritus, but it is also suggested to play a proinflammatory and immunomodulatory role which could play a part in the progression of the neoplastic disease. Further studies will be fundamental in facing pruritus in oncologic patients, since this problem compromise their quality of life worsening an already critic picture.