scholarly journals Cell-permeable transgelin-2 as a potent therapeutic for dendritic cell-based cancer immunotherapy

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Hye-Ran Kim ◽  
Jeong-Su Park ◽  
Jin-Hwa Park ◽  
Fatima Yasmin ◽  
Chang-Hyun Kim ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cell ◽  
Tumor Growth ◽  
Cancer Immunotherapy ◽  
Actin Binding ◽  
B16f10 Melanoma ◽  
Wide Range ◽  
Viral Transduction ◽  
Tumor Growth And Metastasis

Abstract Background Transgelin-2 is a 22 kDa actin-binding protein that has been proposed to act as an oncogenic factor, capable of contributing to tumorigenesis in a wide range of human malignancies. However, little is known whether this tiny protein also plays an important role in immunity, thereby keeping body from the cancer development and metastasis. Here, we investigated the functions of transgelin-2 in dendritic cell (DC) immunity. Further, we investigated whether the non-viral transduction of cell-permeable transgelin-2 peptide potentially enhance DC-based cancer immunotherapy. Methods To understand the functions of transgelin-2 in DCs, we utilized bone marrow-derived DCs (BMDCs) purified from transgelin-2 knockout (Tagln2−/−) mice. To observe the dynamic cellular mechanism of transgelin-2, we utilized confocal microscopy and flow cytometry. To monitor DC migration and cognate T–DC interaction in vivo, we used intravital two-photon microscopy. For the solid and metastasis tumor models, OVA+ B16F10 melanoma were inoculated into the C57BL/6 mice via intravenously (i.v.) and subcutaneously (s.c.), respectively. OTI TCR T cells were used for the adoptive transfer experiments. Cell-permeable, de-ubiquitinated recombinant transgelin-2 was purified from Escherichia coli and applied for DC-based adoptive immunotherapy. Results We found that transgelin-2 is remarkably expressed in BMDCs during maturation and lipopolysaccharide activation, suggesting that this protein plays a role in DC-based immunity. Although Tagln2−/− BMDCs exhibited no changes in maturation, they showed significant defects in their abilities to home to draining lymph nodes (LNs) and prime T cells to produce antigen-specific T cell clones, and these changes were associated with a failure to suppress tumor growth and metastasis of OVA+ B16F10 melanoma cells in mice. Tagln2−/− BMDCs had defects in filopodia-like membrane protrusion and podosome formation due to the attenuation of the signals that modulate actin remodeling in vitro and formed short, unstable contacts with cognate CD4+ T cells in vivo. Strikingly, non-viral transduction of cell-permeable, de-ubiquitinated recombinant transgelin-2 potentiated DC functions to suppress tumor growth and metastasis. Conclusion This work demonstrates that transgelin-2 is an essential protein for both cancer and immunity. Therefore, transgelin-2 can act as a double-edged sword depending on how we apply this protein to cancer therapy. Engineering and clinical application of this protein may unveil a new era in DC-based cancer immunotherapy. Our findings indicate that cell-permeable transgelin-2 have a potential clinical value as a cancer immunotherapy based on DCs.

scholarly journals Endothelial Rap1B mediates T-cell exclusion to promote tumor growth -a novel mechanism underlying vascular immunosuppression

2021 ◽  
Author(s):  
Guru Prasad Sharma ◽  
Ramoji Kosuru ◽  
Sribalaji Lakshmikanthan ◽  
Shikan Zheng ◽  
Yao Chen ◽  
...  
Keyword(s):  
T Cells ◽  
Tumor Growth ◽  
Cancer Immunotherapy ◽  
Small Gtpase ◽  
Vascular Endothelial ◽  
Promote Tumor Growth ◽  
Tnf Α ◽  
Inflammatory Stimuli ◽  
Endothelial Growth Factor

Overcoming vascular immunosuppression: lack of endothelial cell (EC) responsiveness to inflammatory stimuli in the proangiogenic environment of tumors, is essential for successful cancer immunotherapy. The mechanisms through which Vascular Endothelial Growth Factor (VEGF) modulates tumor EC response to exclude T cells are not well understood. The goal was to determine the role of EC Rap1B, a small GTPase that positively regulates VEGFangiogenesis during development, in tumor growth in vivo. Using mouse models of Rap1B deficiency, Rap1B+/- and EC-specific Rap1B KO (Rap1BiΔEC) we demonstrate that EC Rap1B restricts tumor growth and angiogenesis. More importantly, EC-specific Rap1B deletion leads to an altered tumor microenvironment with increased recruitment of leukocytes and increased activity of tumor CD8+ T cells. We find that tumor growth, albeit not angiogenesis, is restored in Rap1BiΔEC mice by depleting CD8+ T cells. Mechanistically, global transcriptome analysis indicated upregulation of the tumor cytokine, TNF-α, -induced signaling and NFκB transcriptional activity in Rap1B-deficient ECs. Functionally, EC Rap1B deletion led to upregulation of NFκB activity and enhanced Cell Adhesion Molecules (CAMs) expression in TNF-α stimulated ECs. Importantly, CAM expression was upregulated also in tumor ECs from Rap1BiΔEC mice, vs. controls. Significantly, deletion of Rap1B abrogated VEGF immunosuppressive downregulation of CAM expression, demonstrating that Rap1B is essential for VEGF-suppressive signaling. Thus, our studies identify a novel endothelial-endogenous mechanism underlying VEGF-dependent desensitization of EC to pro-inflammatory stimuli. Significantly, they identify EC Rap1 as a potential novel vascular target in cancer immunotherapy.

Reinforcement of Cancer Immunotherapy by Adoptive Transfer of Cblb-Deficient Cytotoxic T Lymphocytes Combined with a Dendritic Cell Vaccine

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 957-957
Author(s):  
Christina Lutz-Nicoladoni ◽  
Patrizia Stoizner ◽  
Magdalena Pircher ◽  
Stephanie Wallner ◽  
Anna Maria Wolf ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Dendritic Cell ◽  
Cancer Immunotherapy ◽  
Cytotoxic T Lymphocytes ◽  
Adoptive Transfer ◽  
Ex Vivo ◽  
Dc Vaccine

Abstract Abstract 957 Introduction: Various approaches to induce immunological rejection of tumors including transfer of autologous tumor infiltrating lymhocytes (TIL) after ex vivo clonal expansion or application of ex vivo transduced antigen specific T cell (TCR) transgenic T cells have been elaborated. In general, adoptive T cell transfer (ATC) has been combined with lympho-depleting agents (e.g. cyclophosphamide). However, the therapeutic efficacy of these cancer immunotherapy approaches is limited due to insufficient in vivo activation, expansion and survival of transferred effector immune cells, which is mainly due to suppressive mileu signals and immune evasion mechanisms induced by TGF-β. The E3 ubiquitin ligase Cbl-b is a key regulator of T cell activation and is assumed to confer TGF-β resistance. Thus we performed a proof-of-concept study evaluating Cbl-b targeting as “intracellular adjuvant” strategy to improve ATC for cancer immunotherapy. Material and Methods: We first tested the in vitro sensitivity of CTL towards TGF-β mediated immuno-suppressive cues and then in vivo evaluated the anti-tumor reactivity of cblb-deficient cytotoxic T lymphocytes (CTL) in murine tumor models alone or in combination with a dendritic cell (DC) vaccine. Results: Cblb-deficient CTL are hyper-responsive to TCR/CD28-stimulation in vitro and protected from the negative cues induced by TGF-β as determined by quantification fo IFN-g secretion and quantification of their proliferative capacity. Unexpectedly, adoptive transfer of polyclonal, non TCR-transgenic cblb-deficient CD8+ CTL, however, is not sufficient to reject B16ova or EG7 tumors in vivo, which is in clear contrast to previous reports using lymphopenic animals receiving adoptively transferred TCR-transgenic T cells. Thus, we next evaluated in vivo re-activation of adoptively transferred cblb-deficient T cells by a DC vaccine (i.e. SIINFEKL-pulsed DC). In strict contrast to ATC monotherapy, this approach now markedly delays tumor outgrowth and significantly increase survival rates, which is paralleled by an increased CTL infiltration rate to the tumor site and an enrichment of ova-specific and IFN-g-secreting CTL in the draining lymph nodes. Moreover, compared to wild-type CTL, cblb-deficient mice vaccinated with the DC vaccine show an increased cytolytic activity in vivo. Conclusions: In summary, we provide experimental evidence that genetic inactivation of cblb in polyclonal, non-TCR transgenic adoptively transferred CTL might serve as a novel “adjuvant approach”, suitable to augment the effectiveness of anti-cancer immunotherapies using ATC in immune-competent recipients. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Perforin and Granzyme B Expressed by Murine Myeloid-Derived Suppressor Cells: A Study on Their Role in Outgrowth of Cancer Cells

Cancers ◽  
2019 ◽  
Vol 11 (6) ◽  
pp. 808 ◽  
Author(s):  
Inès Dufait ◽  
Julian Pardo ◽  
David Escors ◽  
Yannick De Vlaeminck ◽  
Heng Jiang ◽  
...  
Keyword(s):  
T Cells ◽  
Cd8 T Cells ◽  
Granzyme B ◽  
Suppressor Cell ◽  
Melanoma Cells ◽  
B16f10 Melanoma ◽  
Knock Out ◽  
Wide Range ◽  
B16f10 Melanoma Cells

A wide-range of myeloid-derived suppressor cell (MDSC)-mediated immune suppressive functions has previously been described. Nevertheless, potential novel mechanisms by which MDSCs aid tumor progression are, in all likelihood, still unrecognized. Next to its well-known expression in natural killer cells and cytotoxic T lymphocytes (CTLs), granzyme B (GzmB) expression has been found in different cell types. In an MDSC culture model, we demonstrated perforin and GzmB expression. Furthermore, similar observations were made in MDSCs isolated from tumor-bearing mice. Even in MDSCs from humans, GzmB expression was demonstrated. Of note, B16F10 melanoma cells co-cultured with perforin/GzmB knock out mice (KO) MDSCs displayed a remarkable decrease in invasive potential. B16F10 melanoma cells co-injected with KO MDSCs, displayed a significant slower growth curve compared to tumor cells co-injected with wild type (WT) MDSCs. In vivo absence of perforin/GzmB in MDSCs resulted in a higher number of CD8+ T-cells. Despite this change in favor of CD8+ T-cell infiltration, we observed low interferon-γ (IFN-γ) and high programmed death-ligand 1 (PD-L1) expression, suggesting that other immunosuppressive mechanisms render these CD8+ T-cells dysfunctional. Taken together, our results suggest that GzmB expression in MDSCs is another means to promote tumor growth and warrants further investigation to unravel the exact underlying mechanism.

scholarly journals Recombinant Costimulatory Fusion Proteins as Functional Immunomodulators Enhance Antitumor Activity in Murine B16F10 Melanoma

Vaccines ◽  
2020 ◽  
Vol 8 (2) ◽  
pp. 223
Author(s):  
Huaman Cai ◽  
Wenfang Wang ◽  
Zhibing Lin ◽  
Yan Zhang ◽  
Bing Wu ◽  
...  
Keyword(s):  
T Cells ◽  
Cancer Immunotherapy ◽  
Transmembrane Domain ◽  
In Vitro Study ◽  
B16f10 Melanoma ◽  
Antitumor Effects ◽  
Therapeutic Modalities ◽  
Inhibitory Signaling

Blocking inhibitory signaling and engaging stimulatory signaling have emerged as important therapeutic modalities for cancer immunotherapy. This study aimed to investigate immunomodulatory features of three recombinant costimulatory ligand proteins in a mouse model, which are extracellular domains of OX40-ligand (OX40L), 4-1BB-ligand (4-1BBL), or two domains in tandem, fused with the transmembrane domain of diphtheria toxin (DTT), named DTT-COS1, DTT-COS2, and DTT-COS12, respectively. In vitro study showed that DTT-COS1 and DTT-COS12 had immunological activity increasing the ratio of CD8/CD4 T cells. Treatments with DTT-COS1 and DTT-COS12 dramatically generated immune protection against the B16F10 tumor challenge in both prophylactic and therapeutic efficacy. Furthermore, regarding tumor microenvironment (TME) immunomodulation, DTT-COS1 treatment increased the proportion of CD4+ effector T cells (Teff) and decreased the expression of a suppressive cytokine. Meanwhile, DTT-COS12 reduced regulatory T cells (Treg) and improved the level of stimulatory cytokines. In addition, endogenous antibodies against OX40L/4-1BBL were generated, which may help with antitumor responses. Unexpectedly, DTT-COS2 lacked antitumor effects in vitro and in vivo. Importantly, serum analysis of liver-function associated factors and pro-inflammatory cytokines demonstrated that treatments were safe formulations in mice without signs of systemic toxicity. Remarkably, DTT-COS1 and DTT-COS12 are functional immunomodulators for mouse B16F10 melanoma, creating practical preclinical value in cancer immunotherapy.

scholarly journals Cytotoxic T-cells mediate exercise-induced reductions in tumor growth

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Helene Rundqvist ◽  
Pedro Veliça ◽  
Laura Barbieri ◽  
Paulo A Gameiro ◽  
David Bargiela ◽  
...  
Keyword(s):  
T Cells ◽  
Tumor Growth ◽  
Tumor Progression ◽  
Cd8 T Cells ◽  
Cytotoxic T Cells ◽  
Central Carbon Metabolism ◽  
Effector Cells ◽  
Wide Range ◽  
Exercise Induced

Exercise has a wide range of systemic effects. In animal models, repeated exertion reduces malignant tumor progression, and clinically, exercise can improve outcome for cancer patients. The etiology of the effects of exercise on tumor progression are unclear, as are the cellular actors involved. We show here that in mice, exercise-induced reduction in tumor growth is dependent on CD8+ T cells, and that metabolites produced in skeletal muscle and excreted into plasma at high levels during exertion in both mice and humans enhance the effector profile of CD8+ T-cells. We found that activated murine CD8+ T cells alter their central carbon metabolism in response to exertion in vivo, and that immune cells from trained mice are more potent antitumor effector cells when transferred into tumor-bearing untrained animals. These data demonstrate that CD8+ T cells are metabolically altered by exercise in a manner that acts to improve their antitumoral efficacy.

scholarly journals Cancer-associated fibroblast secretion of PDGFC promotes gastrointestinal stromal tumor growth and metastasis

Oncogene ◽  
2021 ◽  
Vol 40 (11) ◽  
pp. 1957-1973
Author(s):  
Hyunho Yoon ◽  
Chih-Min Tang ◽  
Sudeep Banerjee ◽  
Mayra Yebra ◽  
Sangkyu Noh ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Tumor Growth ◽  
Gastrointestinal Stromal Tumor ◽  
Single Agent ◽  
Stromal Tumor ◽  
Paracrine Signaling ◽  
Mesenchymal Transition ◽  
Tumor Growth And Metastasis ◽  
Factor C

AbstractTargeted therapies for gastrointestinal stromal tumor (GIST) are modestly effective, but GIST cannot be cured with single agent tyrosine kinase inhibitors. In this study, we sought to identify new therapeutic targets in GIST by investigating the tumor microenvironment. Here, we identified a paracrine signaling network by which cancer-associated fibroblasts (CAFs) drive GIST growth and metastasis. Specifically, CAFs isolated from human tumors were found to produce high levels of platelet-derived growth factor C (PDGFC), which activated PDGFC-PDGFRA signal transduction in GIST cells that regulated the expression of SLUG, an epithelial-mesenchymal transition (EMT) transcription factor and downstream target of PDGFRA signaling. Together, this paracrine induce signal transduction cascade promoted tumor growth and metastasis in vivo. Moreover, in metastatic GIST patients, SLUG expression positively correlated with tumor size and mitotic index. Given that CAF paracrine signaling modulated GIST biology, we directly targeted CAFs with a dual PI3K/mTOR inhibitor, which synergized with imatinib to increase tumor cell killing and in vivo disease response. Taken together, we identified a previously unappreciated cellular target for GIST therapy in order to improve disease control and cure rates.

627 The DLL3-targeted half-life extended bispecific T cell engager (HLE BiTE®) immune-oncology therapy AMG 757 has potent antitumor activity in neuroendocrine cancer

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A663-A663
Author(s):  
Keegan Cooke ◽  
Juan Estrada ◽  
Jinghui Zhan ◽  
Jonathan Werner ◽  
Fei Lee ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Growth ◽  
Mouse Models ◽  
T Cell Activation ◽  
Phase 1 ◽  
Phase 1 Study ◽  
Human T Cells

BackgroundNeuroendocrine tumors (NET), including small cell lung cancer (SCLC), have poor prognosis and limited therapeutic options. AMG 757 is an HLE BiTE® immune therapy designed to redirect T cell cytotoxicity to NET cells by binding to Delta-like ligand 3 (DLL3) expressed on the tumor cell surface and CD3 on T cells.MethodsWe evaluated activity of AMG 757 in NET cells in vitro and in mouse models of neuroendocrine cancer in vivo. In vitro, co-cultures of NET cells and human T cells were treated with AMG 757 in a concentration range and T cell activation, cytokine production, and tumor cell killing were assessed. In vivo, AMG 757 antitumor efficacy was evaluated in xenograft NET and in orthotopic models designed to mimic primary and metastatic SCLC lesions. NSG mice bearing established NET were administered human T cells and then treated once weekly with AMG 757 or control HLE BiTE molecule; tumor growth inhibition was assessed. Pharmacodynamic effects of AMG 757 in tumors were also evaluated in SCLC models following a single administration of human T cells and AMG 757 or control HLE BiTE molecule.ResultsAMG 757 induced T cell activation, cytokine production, and potent T cell redirected killing of DLL3-expressing SCLC, neuroendocrine prostate cancer, and other DLL3-expressing NET cell lines in vitro. AMG 757-mediated redirected lysis was specific for DLL3-expressing cells. In patient-derived xenograft and orthotopic models of SCLC, single-dose AMG 757 effectively engaged human T cells administered systemically, leading to a significant increase in the number of human CD4+ and CD8+ T cells in primary and metastatic tumor lesions. Weekly administration of AMG 757 induced significant tumor growth inhibition of SCLC (figure 1) and other NET, including complete regression of established tumors and clearance of metastatic lesions. These findings warranted evaluation of AMG 757 (NCT03319940); the phase 1 study includes dose exploration (monotherapy and in combination with pembrolizumab) and dose expansion (monotherapy) in patients with SCLC (figure 2). A study of AMG 757 in patients with neuroendocrine prostate cancer is under development based on emerging data from the ongoing phase 1 study.Abstract 627 Figure 1AMG 757 Significantly reduced tumor growth in orthotopic SCLC mouse modelsAbstract 627 Figure 2AMG 757 Phase 1 study designConclusionsAMG 757 engages and activates T cells to kill DLL3-expressing SCLC and other NET cells in vitro and induces significant antitumor activity against established xenograft tumors in mouse models. These preclinical data support evaluation of AMG 757 in clinical studies of patients with NET.Ethics ApprovalAll in vivo work was conducted under IACUC-approved protocol #2009-00046.

245 Human TLR8 knock-in mice potentiate immunotherapy responses of MC38 syngeneic tumors

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A264-A264
Author(s):  
Shanshan Qi ◽  
Hongjuan Zhang ◽  
Ruilin Sun ◽  
Annie An ◽  
Henry Li ◽  
...  
Keyword(s):  
T Cells ◽  
Tumor Growth ◽  
Cancer Immunotherapy ◽  
Tumor Regression ◽  
Tumor Growth Rate ◽  
Tumor Growth Inhibition ◽  
List Type ◽  
Rna Recognition ◽  
Ethical Guidelines ◽  
Ifn Γ

BackgroundToll-like receptors (TLRs) serve critical roles in mediating innate immune responses against many pathogens. However, they may also bind to endogenous ligands and lead to the pathogenesis of autoimmunity. Although TLR8 belongs to the same TLR family as TLR7, its role in inflammation and tumor progression is not yet fully understood due to the lack of suitable animal models. In humans, both TLR7 and TLR8 recognize single-stranded self-RNA, viral RNA, and synthetic small molecule agonists.1, 2 However, mouse Tlr8 is non-functional due to the absence of 5 amino acids necessary for RNA recognition. In order to create a mouse model with functional TLR8, we replaced exon 3 of mouse Tlr8 with human TLR8, therefore developing a hTLR8 knock-in (KI) model. Both heterozygous and homozygous hTLR8 KI mice are viable with inflammatory phenotypes, i.e. enlarged spleens and livers, and significantly higher IL-12 p40 levels under TLR8 agonist treatment. In this study, we evaluated the potential use of hTLR8 mice for cancer immunotherapy studies.MethodshTLR8 mice, together with naïve C57BL/6 mice, were inoculated with MC38 syngeneic tumor cells. Tumor bearing mice were grouped at a mean tumor volume of approximately 100 mm3 for treatment with PBS or 10 mg/kg anti-PD-1 (RMP1-14) antibody. At the efficacy endpoint, spleens and tumors were collected for flow cytometry profiling.ResultsAnti-PD-1 treatment of MC38 tumors in naïve C57BL/6 led to moderate tumor growth inhibition (TGI = 54%). Interestingly, anti-PD-1 treatment showed improved efficacy in hTLR8 mice (TGI = 79%), including 2/10 tumors with complete tumor regression. In comparison, non-treated MC38 tumor growth rate was slower in hTLR8 mice than in naïve mice. Anti-PD-1 treated hTLR8 mice also had significantly increased IFN-γ and TNF-a positive CD4+ T cells in the spleen, along with higher numbers of differentiated effector T cells. In addition, hTLR8 mice have activated dendritic cells and macrophages, acting as critical steps in initiation of the inflammatory process, with higher levels of pro-inflammatory cytokines, such as IL-6, IFN-γ, TNF-a, and IL-1β, which may promote Th1 priming and differentiation of T cells into IFN-γ or TNF-a producing cells.ConclusionshTLR8 mice offer a great tool to model cancer immunotherapy in an inflammatory/autoimmunity prone background. Moreover, hTLR8 mice can be effectively used to shift a ‘cold’ tumor phenotype to ‘hot’ tumors in a syngeneic setting.Ethics ApprovalAnimal experiments were conducted in accordance with animal welfare law, approved by local authorities, and in accordance with the ethical guidelines of CrownBio (Taicang).ReferencesKugelberg E. Making mice more human the TLR8 way. Nat Rev Immunol 2014;14:6.Guiducci C, Gong M, Cepika A-M, et al. RNA recognition by human TLR8 can lead to autoimmune inflammation. J Exp Med 2013;210:2903–2919.

scholarly journals 114 Preclinical development of a novel iPSC-derived CAR-MICA/B NK cell immunotherapy to overcome solid tumor escape from NKG2D-mediated mechanisms of recognition and killing

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A126-A126
Author(s):  
John Goulding ◽  
Mochtar Pribadi ◽  
Robert Blum ◽  
Wen-I Yeh ◽  
Yijia Pan ◽  
...  
Keyword(s):  
T Cells ◽  
Cancer Immunotherapy ◽  
Tumor Immunity ◽  
Immune Cell ◽  
Nk Cell ◽  
Tumor Escape ◽  
Car T Cells ◽  
Car T

BackgroundMHC class I related proteins A (MICA) and B (MICB) are induced by cellular stress and transformation, and their expression has been reported for many cancer types. NKG2D, an activating receptor expressed on natural killer (NK) and T cells, targets the membrane-distal domains of MICA/B, activating a potent cytotoxic response. However, advanced cancer cells frequently evade immune cell recognition by proteolytic shedding of the α1 and α2 domains of MICA/B, which can significantly reduce NKG2D function and the cytolytic activity.MethodsRecent publications have shown that therapeutic antibodies targeting the membrane-proximal α3 domain inhibited MICA/B shedding, resulting in a substantial increase in the cell surface density of MICA/B and restoration of immune cell-mediated tumor immunity.1 We have developed a novel chimeric antigen receptor (CAR) targeting the conserved α3 domain of MICA/B (CAR-MICA/B). Additionally, utilizing our proprietary induced pluripotent stem cell (iPSC) product platform, we have developed multiplexed engineered, iPSC-derived CAR-MICA/B NK (iNK) cells for off-the-shelf cancer immunotherapy.ResultsA screen of CAR spacer and ScFv orientations in primary T cells delineated MICA-specific in vitro activation and cytotoxicity as well as in vivo tumor control against MICA+ cancer cells. The novel CAR-MICA/B design was used to compare efficacy against NKG2D CAR T cells, an alternative MICA/B targeting strategy. CAR-MICA/B T cells showed superior cytotoxicity against melanoma, breast cancer, renal cell carcinoma, and lung cancer lines in vitro compared to primary NKG2D CAR T cells (p<0.01). Additionally, using an in vivo xenograft metastasis model, CAR-MICA/B T cells eliminated A2058 human melanoma metastases in the majority of the mice treated. In contrast, NKG2D CAR T cells were unable to control tumor growth or metastases. To translate CAR-MICA/B functionality into an off-the-shelf cancer immunotherapy, CAR-MICA/B was introduced into a clonal master engineered iPSC line to derive a multiplexed engineered, CAR-MICA/B iNK cell product candidate. Using a panel of tumor cell lines expressing MICA/B, CAR-MICA/B iNK cells displayed MICA specificity, resulting in enhanced cytokine production, degranulation, and cytotoxicity. Furthermore, in vivo NK cell cytotoxicity was evaluated using the B16-F10 melanoma cell line, engineered to express MICA. In this model, CAR-MICA/B iNK cells significantly reduced liver and lung metastases, compared to untreated controls, by 93% and 87% respectively.ConclusionsOngoing work is focused on extending these preclinical studies to further support the clinical translation of an off-the-shelf, CAR-MICA/B iNK cell cancer immunotherapy with the potential to overcome solid tumor escape from NKG2D-mediated mechanisms of recognition and killing.ReferenceFerrari de Andrade L, Tay RE, Pan D, Luoma AM, Ito Y, Badrinath S, Tsoucas D, Franz B, May KF Jr, Harvey CJ, Kobold S, Pyrdol JW, Yoon C, Yuan GC, Hodi FS, Dranoff G, Wucherpfennig KW. Antibody-mediated inhibition of MICA and MICB shedding promotes NK cell-driven tumor immunity. Science 2018 Mar 30;359(6383):1537–1542.

scholarly journals IMMU-31. QUANTITATIVE EVALUATION OF ROUTES OF ADMINISTRATION OF IMMUNOTHERAPEUTIC T CELLS TO ORTHOTOPIC GLIOMA

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii111-ii111
Author(s):  
Lan Hoang-Minh ◽  
Angelie Rivera-Rodriguez ◽  
Fernanda Pohl-Guimarães ◽  
Seth Currlin ◽  
Christina Von Roemeling ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Growth ◽  
Ex Vivo ◽  
Adoptive T Cell Therapy ◽  
Whole Body ◽  
T Cell Therapy ◽  
Clinical Responses

Abstract SIGNIFICANCE Adoptive T cell therapy (ACT) has emerged as the most effective treatment against advanced malignant melanoma, eliciting remarkable objective clinical responses in up to 75% of patients with refractory metastatic disease, including within the central nervous system. Immunologic surrogate endpoints correlating with treatment outcome have been identified in these patients, with clinical responses being dependent on the migration of transferred T cells to sites of tumor growth. OBJECTIVE We investigated the biodistribution of intravenously or intraventricularly administered T cells in a murine model of glioblastoma at whole body, organ, and cellular levels. METHODS gp100-specific T cells were isolated from the spleens of pmel DsRed transgenic C57BL/6 mice and injected intravenously or intraventricularly, after in vitro expansion and activation, in murine KR158B-Luc-gp100 glioma-bearing mice. To determine transferred T cell spatial distribution, the brain, lymph nodes, heart, lungs, spleen, liver, and kidneys of mice were processed for 3D imaging using light-sheet and multiphoton imaging. ACT T cell quantification in various organs was performed ex vivo using flow cytometry, 2D optical imaging (IVIS), and magnetic particle imaging (MPI) after ferucarbotran nanoparticle transfection of T cells. T cell biodistribution was also assessed in vivo using MPI. RESULTS Following T cell intravenous injection, the spleen, liver, and lungs accounted for more than 90% of transferred T cells; the proportion of DsRed T cells in the brains was found to be very low, hovering below 1%. In contrast, most ACT T cells persisted in the tumor-bearing brains following intraventricular injections. ACT T cells mostly concentrated at the periphery of tumor masses and in proximity to blood vessels. CONCLUSIONS The success of ACT immunotherapy for brain tumors requires optimization of delivery route, dosing regimen, and enhancement of tumor-specific lymphocyte trafficking and effector functions to achieve maximal penetration and persistence at sites of invasive tumor growth.

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