Phenotype and functional evaluation of ex vivo generated antigen-specific immune effector cells with potential for therapeutic applications

Background: Antibody recruiting molecules (ARM) are novel, immunotherapeutic bifunctional molecules composed of two active termini connected by a linker. One of the termini binds to a target molecule on a cancer cell. The other terminus, called universal antibody binding terminus (uABT), recruit all endogenous IgG antibodies independent of their antigen binding specificity. As a result, the target cell is "opsonized" by antibodies which then bring the immune effector cells to eliminate the target through various antibody-dependent destruction mechanisms. Kleo Pharmaceuticals has developed a series of CD38-ARM mlecules which target human CD38 highly expressed by multiple myeloma cells. CD38-ARM compounds are able to mediate ADCC without depleting CD38 expressing immune effector cells like existing therapeutic antibodies such as Daratumumab. Methods: Cyclized peptides containing natural and non-natural amino-acid that selectively bind to human CD38 were identified using Peptidream Flexizyme-based, cell free Peptide Discovery Translation System. These peptides were linked to uABT antibody binder via a linker to generate the final CD38-ARM molecules Binding of CD38-ARM was tested by ternary complex formation between CD38 expressing cells, CD38-ARM and labelled human IgG1. To confirm the activity of CD38-ARM, surrogate CD16a binding and signaling assays were performed using the NFAT Promega system. Antibody dependent cellular cytotoxicity (ADCC) assays using purified NK cells from multiple donors with polymorphism variants (V/V, F/F, and V/F) of CD16a were performed to confirm activity. Live cell imaging was utilized to assess the dynamics of NK-RAJI cell interactions mediated by CD38-ARM +/- IgG. We evaluated the ability of compounds to mediate complement dependent cytotoxicity (CDC). We tested the effect of CD38-ARM on human immune cell populations within PBMC and whole bone marrow (WBM) by flow cytometry. Lastly, ex vivo samples from WBM of MM patients at diagnosis or relapse were used to evaluate CD38-ARM anti-tumor activity as well as off-target effects, without the addition of external source of IgG, through multiparametric flow-cytometry (CD45, CD19, CD38, CD138, CD56, CD27, CD8, CD117). Results: The CD38-ARM were shown to have the ability to bind to CD38 with a 7nM affinity and to human IgG1 and IgG2 with affinity of 15nM and 11nM by SPR. Activity of KP compounds was observed in all assays except for CDC. In ternary assay, KP-6 had an apparent EC50 of 16nM while KP-7's EC50 was 6nM. Both KP-6 and 7 had comparable EC50s in the single digit nanamolar range in the NFAT activation assay induced by CD16a binding was confirmed using human IgG to induce, while Daratumumab had an apparent EC50 of 0.04nM. In the ADCC assay, both KP-6 & KP-7 had EC50s of 7 & 6nM respectively, while Daratumumab had an EC50 of 0.1nM. In addition, no NK cell depletion was observed when PBMC were treated with KP compounds, whereas a profound reduction in both percentages and absolute numbers in this cell subset was observed with Daratumumab treatment. Increasing dose of CD38-ARM (range 0.1uM- 25uM) were tested in ex vivo WBM samples from MM patients together with a negative control and Daratumumab. At concentrations of 10uM and 25uM, CD38-ARM induced a significant reduction of MM cells achieving results comparable to those of Daratumumab activity (p >0.05 in both cases), while sparing all other CD38+ normal cells such as NK, T lymphocytes, monocytes and granulocytes, which are always reduced in the presence of Daratumumab. Conclusions: CD38-ARMs are able to kill MM cells by ADCC without depleting CD38 expressing immune cells contrary to existing antibodies such as Daratumumab. CD38-ARMs do not activate complement, which might be involved in the infusion reaction seen with Daratumumab. Most importantly, CD38-ARMs kill multiple myeloma cells ex vivo in patient bone marrow samples as well as plasma cell leukemia in patient blood. Combined with the in vivo efficacy data presented elsewhere, this data establishes the therapeutic potential of CD38-ARM. They also represent the first demonstration of the ARM platform ability to generate therapeutic agents tailored to a specific indication, by varying target binding moiety of the molecule. Disclosures Rossi: Kleo pharmaceuticals: Employment, Equity Ownership. Bunin:Kleo pharmaceuticals: Employment, Equity Ownership. Iben:Kleo Pharmaceuticals: Employment, Equity Ownership. Welsch:Kleo pharmaceuticals: Employment, Equity Ownership. Berbasova:Kleo Pharmaceuticals: Employment, Equity Ownership. Riillo:Kleo Pharmaceuticals: Research Funding. Ohuchi:Peptidream Inc.: Employment. Alvarez:Kleo pharmaceuticals: Employment, Equity Ownership. Kawakami:Peptidream Inc.: Employment. Nagasawa:Peptidream Inc.: Employment. Spiegel:Kleo pharmaceuticals: Equity Ownership. Rastelli:Kleo pharmaceuticals: Employment, Equity Ownership.

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The Role of Microglia in Sporadic Alzheimer’s Disease

Journal of Alzheimer s Disease ◽

10.3233/jad-201248 ◽

2021 ◽

Vol 79 (3) ◽

pp. 961-968

Author(s):

Wolfgang J. Streit ◽

Habibeh Khoshbouei ◽

Ingo Bechmann

Keyword(s):

Alzheimer’S Disease ◽

Alzheimer's Disease ◽

Late Onset ◽

Genetic Mutations ◽

Sporadic Alzheimer’S Disease ◽

Immune Effector ◽

Effector Cells ◽

Immune Effector Cells ◽

Amyloid Cascade

Microglia constitute the brain’s immune system and their involvement in Alzheimer’s disease has been discussed. Commonly, and in line with the amyloid/neuroinflammation cascade hypothesis, microglia have been portrayed as potentially dangerous immune effector cells thought to be overactivated by amyloid and producing neurotoxic inflammatory mediators that lead to neurofibrillary degeneration. We disagree with this theory and offer as an alternative the microglial dysfunction theory stating that microglia become impaired in their normally neuroprotective roles because of aging, i.e., they become senescent and aging neurons degenerate because they lack the needed microglial support for their survival. Thus, while the amyloid cascade theory relies primarily on genetic data, the dysfunction theory incorporates aging as a critical etiological factor. Aging is the greatest risk factor for the sporadic (late-onset) and most common form of Alzheimer’s disease, where fully penetrant genetic mutations are absent. In this review, we lay out and discuss the human evidence that supports senescent microglial dysfunction and conflicts with the amyloid/neuroinflammation idea.

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Next-generation cell therapies: the emerging role of CAR-NK cells

Hematology ◽

10.1182/hematology.2020002547 ◽

2020 ◽

Vol 2020 (1) ◽

pp. 570-578

Author(s):

Rafet Basar ◽

May Daher ◽

Katayoun Rezvani

Keyword(s):

T Cells ◽

Cell Therapy ◽

Natural Killer ◽

Γδ T Cells ◽

Antigen Receptors ◽

T Cell Therapy ◽

Cell Therapies ◽

Immune Effector ◽

Effector Cells ◽

Immune Effector Cells

Abstract T cells engineered with chimeric antigen receptors (CARs) have revolutionized the field of cell therapy and changed the paradigm of treatment for many patients with relapsed or refractory B-cell malignancies. Despite this progress, there are limitations to CAR-T cell therapy in both the autologous and allogeneic settings, including practical, logistical, and toxicity issues. Given these concerns, there is a rapidly growing interest in natural killer cells as alternative vehicles for CAR engineering, given their unique biological features and their established safety profile in the allogeneic setting. Other immune effector cells, such as invariant natural killer T cells, γδ T cells, and macrophages, are attracting interest as well and eventually may be added to the repertoire of engineered cell therapies against cancer. The pace of these developments will undoubtedly benefit from multiple innovative technologies, such as the CRISPR-Cas gene editing system, which offers great potential to enhance the natural ability of immune effector cells to eliminate refractory cancers.

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Soluble CD70: a novel immunotherapeutic agent for experimental glioblastoma

Journal of Neurosurgery ◽

10.3171/2009.11.jns09901 ◽

2010 ◽

Vol 113 (2) ◽

pp. 280-285 ◽

Cited By ~ 18

Author(s):

James Miller ◽

Guenter Eisele ◽

Ghazaleh Tabatabai ◽

Steffen Aulwurm ◽

Gabriele von Kürthy ◽

...

Keyword(s):

T Cells ◽

Immune Responses ◽

Malignant Gliomas ◽

Ectopic Expression ◽

Glioma Cells ◽

Interferon Γ ◽

Soluble Form ◽

Immune Effector ◽

Effector Cells ◽

Immune Effector Cells

Object Given the overall poor outcome with current treatment strategies in malignant gliomas, immunotherapy has been considered a promising experimental approach to glioblastoma for more than 2 decades. A cell surface molecule, CD70, may induce potent antitumor immune responses via activation of the costimulatory receptor CD27 expressed on immune effector cells. There is evidence that a soluble form of CD70 (sCD70) may exhibit biological activity, too. A soluble costimulatory ligand is attractive because it may facilitate immune activation and may achieve a superior tissue distribution. Methods To test the antiglioma effect of sCD70, the authors genetically modified SMA-560 mouse glioma cells to secrete the extracellular domain of CD70. They assessed the immunogenicity of the transfected cells in cocultures with immune effector cells by the determination of immune cell proliferation and the release of interferon-γ. Syngeneic VM/Dk mice were implanted orthotopically with control or sCD70-releasing glioma cells to determine a survival benefit mediated by sCD70. Depletion studies were performed to identify the cellular mediators of prolonged survival of sCD70-releasing glioma-bearing mice. Results The authors found that ectopic expression of sCD70 enhanced the proliferation and interferon-γ release of syngeneic splenocytes in vitro. More importantly, sCD70 prolonged the survival of syngeneic VM/Dk mice bearing intracranial SMA-560 gliomas. The survival rate at 60 days increased from 5 to 45%. Antibody-mediated depletion of CD8-positive T cells abrogates the survival advantage conferred by sCD70. Conclusions These data suggest that sCD70 is a potent stimulator of antiglioma immune responses that depend critically on CD8-positive T cells. Soluble CD70 could be a powerful adjuvant for future immunotherapy trials for glioblastoma.

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Depletion of immune effector cells induces myocardial damage in the acute experimental Trypanosoma cruzi infection: ultrastructural study in rats

Tissue and Cell ◽

10.1054/tice.1999.0040 ◽

1999 ◽

Vol 31 (3) ◽

pp. 281-290 ◽

Cited By ~ 13

Author(s):

R.C.N. Melo

Keyword(s):

Trypanosoma Cruzi ◽

Ultrastructural Study ◽

Myocardial Damage ◽

Immune Effector ◽

Effector Cells ◽

Immune Effector Cells ◽

Cruzi Infection

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Antibody-based therapy of leukaemia

Expert Reviews in Molecular Medicine ◽

10.1017/s1462399409001215 ◽

2009 ◽

Vol 11 ◽

Cited By ~ 25

Author(s):

John C. Morris ◽

Thomas A. Waldmann

Keyword(s):

Monoclonal Antibodies ◽

T Cell ◽

B Cell ◽

Gemtuzumab Ozogamicin ◽

Cell Leukaemia ◽

Target Cells ◽

Immune Effector ◽

Effector Cells ◽

Prolymphocytic Leukaemia ◽

Immune Effector Cells

Over the past decade, monoclonal antibodies have dramatically impacted the treatment of haematological malignancies, as evidenced by the effect of rituximab on the response rate and survival of patients with follicular and diffuse large B cell non-Hodgkin's lymphoma. Currently, only two monoclonal antibodies – the anti-CD33 immunotoxin gemtuzumab ozogamicin and the CD52-directed antibody alemtuzumab – are approved for treatment of relapsed acute myeloid leukaemia in older patients and B cell chronic lymphocytic leukaemia, respectively. Although not approved for such treatment, alemtuzumab is also active against T cell prolymphocytic leukaemia, cutaneous T cell lymphoma and Sézary syndrome, and adult T cell leukaemia and lymphoma. In addition, rituximab has demonstrated activity against B cell chronic lymphocytic and hairy cell leukaemia. Monoclonal antibodies targeting CD4, CD19, CD20, CD22, CD23, CD25, CD45, CD66 and CD122 are now being studied in the clinic for the treatment of leukaemia. Here, we discuss how these new antibodies have been engineered to reduce immunogenicity and improve antibody targeting and binding. Improved interactions with Fc receptors on immune effector cells can enhance destruction of target cells through antibody-dependent cellular cytotoxicity and complement-mediated cell lysis. The antibodies can also be armed with cellular toxins or radionuclides to enhance the destruction of leukaemia cells.

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The interactions between major immune effector cells and Hepatocellular Carcinoma: A systematic review

International Immunopharmacology ◽

10.1016/j.intimp.2021.108220 ◽

2021 ◽

Vol 101 ◽

pp. 108220

Author(s):

Markus Bo Schoenberg ◽

Xiaokang Li ◽

Xinyu Li ◽

Yongsheng Han ◽

Nikolaus Börner ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Systematic Review ◽

Immune Effector ◽

Effector Cells ◽

Immune Effector Cells

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

Frontiers in Oncology ◽

10.3389/fonc.2021.714550 ◽

2021 ◽

Vol 11 ◽

Author(s):

Cristina Saiz-Ladera ◽

Mariona Baliu-Piqué ◽

Francisco J. Cimas ◽

Aránzazu Manzano ◽

Vanesa García-Barberán ◽

...

Keyword(s):

Squamous Cell Carcinoma ◽

Immune System ◽

Cell Carcinoma ◽

Head And Neck ◽

Squamous Cell ◽

Immune Effector ◽

Effector Cells ◽

Favorable Prognosis ◽

Immune Effector Cells ◽

Tumor Types

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.

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The Advantages and Challenges of Anticancer Dendritic Cell Vaccines and NK Cells in Adoptive Cell Immunotherapy

Vaccines ◽

10.3390/vaccines9111363 ◽

2021 ◽

Vol 9 (11) ◽

pp. 1363

Author(s):

Elena V. Abakushina ◽

Liubov I. Popova ◽

Andrey A. Zamyatnin ◽

Jens Werner ◽

Nikolay V. Mikhailovsky ◽

...

Keyword(s):

Cytotoxic Activity ◽

Nk Cells ◽

Nk Cell ◽

Therapeutic Potential ◽

Ex Vivo ◽

Adoptive Cell Therapy ◽

Immune Effector ◽

Effector Cells ◽

Cell Immunotherapy

In the last decade, an impressive advance was achieved in adoptive cell therapy (ACT), which has improved therapeutic potential and significant value in promising cancer treatment for patients. The ACT is based on the cell transfer of dendritic cells (DCs) and/or immune effector cells. DCs are often used as vaccine carriers or antigen-presenting cells (APCs) to prime naive T cells ex vivo or in vivo. Cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are used as major tool effector cells for ACT. Despite the fact that NK cell immunotherapy is highly effective and promising against many cancer types, there are still some limitations, including insignificant infiltration, adverse conditions of the microenvironment, the immunosuppressive cellular populations, and the low cytotoxic activity in solid tumors. To overcome these difficulties, novel methods of NK cell isolation, expansion, and stimulation of cytotoxic activity should be designed. In this review, we discuss the basic characteristics of DC vaccines and NK cells as potential adoptive cell preparations in cancer therapy.

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