557 Determining the efficacy of ADCC by the 3D-EX ex vivo platform utilizing tumoroids of fresh patient tumor samples with intact tumor microenvironment

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A591-A591
Author(s):  
Jenny Kreahling ◽  
Jared Ehrhart ◽  
Stephen Iwanowycz ◽  
Mibel Pabon ◽  
Tina Pastoor ◽  
...  
Keyword(s):  
Cell Death ◽  
Tumor Microenvironment ◽  
Tumor Cell ◽  
Immune Cells ◽  
Immune Cell ◽  
Ex Vivo ◽  
Cell Killing ◽  
Confocal Imaging ◽  
Tumor Cell Death ◽  
Tumor Cell Killing

BackgroundAntibody-dependent cell-mediated cytotoxicity (ADCC) is an effective tool where antibody-coated cells are targeted and killed by effector immune cells. The application of ADCC therapies has been expanded for both solid tumors as well as hematologic malignancies. However, the immunosuppressive mechanisms present in the immune tumor microenvironment (TME) pose a formidable challenge to immune cell efficacy in addition to hinderance of immune cell infiltration by tumor stromal elements. Hence, it is important to develop clinically relevant platforms to assess the efficacy of antibodies for ADCC. Here we utilized our 3D-EX platform using tumoroids of fresh patient tumor samples to assess ADCC-mediated tumor cell killing.MethodsAll human tumor samples were obtained with proper patient consent and IRB approval. Fresh patient tumor tissue of various histologic types including non-small cell lung cancer (NSCLC) and colorectal cancer (CRC) was processed to generate uniform sized live 3D tumoroids measuring 150 µm in size. Treatment groups included cetuximab alone or in combination with nivolumab and/or ipilimumab. Culture supernatants were collected for multiplex analysis of cytokine release in media. Multiplex flow cytometry was used to assess the activation profile of tumor resident immune cells in combination with high-content confocal imaging to determine extent of ADCC-mediated tumor cell death in the intact tumor extracellular matrix.ResultsUsing fresh patient-derived tumor organoids, we observed ADCC-dependent death of EGFR expressing tumor cells. Flow cytometric analysis of immune cell populations demonstrated treatment mediated activation of resident immune cells, which coincided with cytokine profiles determined by Luminex multiplex cytokine analysis. Additionally, tumor cell killing observed through high-content confocal imaging and quantitative image analysis showed tumor cell death with the 3D tumoroids.ConclusionsIn this comprehensive study we demonstrate that the 3D-EX ex vivo model is a robust system to assess the efficacy of ADCC and to develop novel therapeutic combinations with other immuno-oncology therapies. Furthermore, implementation of this platform in clinical studies may also allow for determination of the most effective combinatorial immuno-oncology therapy strategies for specialized individual patient care.Ethics ApprovalThe study was approved by Chesapeake IRB Pro00014313.

619 Evaluating the effectiveness of targeted ADC therapy in a patient-derived ex vivo tumoroid model, 3D-EX, for quantitative tumor cell killing

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A655-A655
Author(s):  
Jenny Kreahling ◽  
Jared Ehrhart ◽  
Mibel Pabon ◽  
Stephen Iwanowycz ◽  
Tina Pastoor ◽  
...  
Keyword(s):  
Cell Death ◽  
Tumor Microenvironment ◽  
Tumor Cell ◽  
Immune Cells ◽  
Immune Cell ◽  
Confocal Imaging ◽  
Tumor Cell Death ◽  
Drug Conjugates ◽  
Tumor Cell Killing ◽  
Antibody Drug

BackgroundAntibody drug conjugates (ADCs) are an effective tool for site directed delivery of cytotoxic agents to cancer cells. Tailoring of ADC-specificity to the uniqueness of a patient‘s tumor can aid in direct-targeting of tumor cells and potentially improve drug responsiveness. Here we evaluate the potential of using an ADC therapy for targeted tumor cell death and immune cell activation in combination with checkpoint inhibitors in 3D tumoroids.MethodsAll human tumor samples were obtained with proper patient consent and IRB approval. Fresh patient tumor tissue of various histologic types including CRC and NSCLC were processed to generate uniform sized live 3D tumoroids measuring 150 µm in size. Treatment groups included a conjugated ADC therapeutic antibody alone or in combination with PD-1/PD-L1 inhibitors. Culture supernatants were collected for multiplex analysis of cytokine release in media. Additionally, flow cytometry was used to assess the activation profile of resident immune cells in combination with high-content confocal imaging to determine extent of tumor cell death in the intact tumor extracellular matrix.ResultsUsing fresh patient-derived tumoroids, we observed ADC-mediated cell death and activation of immune cells within the tumor microenvironment. Production of pro-inflammatory cytokines correlated with increased activation of tumor infiltrating immune cell populations. The improved immune response led to increased tumor cell killing within the 3D tumor microenvironment observed by high-content confocal imaging.ConclusionsIn this study we demonstrate that our physiologically relevant 3D tumoroid model is an effective system to assess novel antibody drug conjugates and to develop rational drug combinations with other immuno-oncology agents. Furthermore, implementation of 3D-EX platform, in the clinical setting, may also allow for determination of the most effective combinatorial immuno-oncology treatment strategies for individualized patient care.Ethics ApprovalThe study was approved by Chesapeake IRB Pro00014313.

174 An ex vivo tumoroid model of fresh patient tumors (3D-ACT) to assess efficacy of cellular therapy in immuno-oncology

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A188-A188
Author(s):  
Stephen Iwanowycz ◽  
Jared Ehrhart ◽  
Mibel Pabon ◽  
Tina Pastoor ◽  
Jenny Kreahling ◽  
...  
Keyword(s):  
T Cells ◽  
Tumor Microenvironment ◽  
Tumor Cell ◽  
Ex Vivo ◽  
Cell Killing ◽  
Confocal Imaging ◽  
Flow Cytometry Analysis ◽  
Car T ◽  
Engineered T Cells ◽  
Tumor Cell Killing

BackgroundAdoptive T cell therapy (ACT) strategies have achieved substantial advances in the treatment of malignant tumors. Some of the unique challenges posed to ACT by solid tumors include locating target cells, as well as entering and surviving the complex tumor microenvironment. To develop better ACT applications and identify combination therapies to enhance tumor cell killing efficacy of ACT it is imperative to develop preclinical platforms that recapitulate the complexity of patient tumor microenvironment (TME). The goal of this study was to develop an integrated confocal-based high-throughput, high-content real time imaging platform to assess immunogenic tumor cell killing (TCK) activity of ACT applications such as CAR-T and TCR using fresh patient tumor samples.MethodsAll patient tumor samples were obtained with patient consent and relevant IRB approval. For the confocal imaging platform, unpropagated 3D tumoroids with intact TME measuring 150 micron in size were prepared from fresh tumor samples of renal cell carcinoma (RCC), colorectal carcinoma (CRC) and non-small cell lung cancer (NSCLC) using proprietary technology developed at Nilogen Oncosystems. Engineered T-cells were labeled with different fluorescent cell tracker dyes to monitor cell migration and locations within tumoroids by confocal analysis. Comprehensive flow cytometry analysis was performed to corroborate confocal imaging findings from TCK and multiplex cytokine release assays used to assess changes in the TME.ResultsOur studies demonstrated that the confocal-based high-content real time imaging platform described here, combined with a custom image analysis algorithm, allowed for monitoring of treatment-mediated tumor cell killing with structural and functional analysis of engineered T-cells in intact 3D tumoroids. The penetration rate of CAR-T and TCR cells into tumoroids as well as associated tumor cell death varied significantly between different tumor types. Flow cytometry analysis allowed for monitoring of the activation status and viability of engineered T-cells, and treatment-mediated changes in tumor resident immune cell populations.ConclusionsOur data indicated that the immunosuppressive tumor microenvironment may have implications for the application of ACT. Use of the ex vivo platform described here (3D-ACT) may aid in the validation of combinatorial therapies that block or deplete suppressive factors present within the TME, allowing these therapies to overcome mechanisms associated with dysfunction in CAR-T and TCR cell applications.Ethics ApprovalThe study was approved by Chesapeake IRB Pro00014313.

A precision medicine tool “Knowledge Frames” to explain poor results of STING agonist trials.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e15632-e15632
Author(s):  
Al Blunt ◽  
Gerald L. Messerschmidt ◽  
Steve Gyorffy
Keyword(s):  
Tumor Microenvironment ◽  
Tumor Cell ◽  
Precision Medicine ◽  
Immune Cells ◽  
Immune Cell ◽  
Human Organism ◽  
Interferon Production ◽  
Tumor Cell Death ◽  
Cell Homing ◽  
Public Data

e15632 Background: STING activating cyclic-di-nucleotides result in the production of interferons, activating immune cells. We show that “Precision Medicine Knowledge Frames” (PMKF) are analyses inherently involving 2 processes: Frame 1 – Bench Science - biochemical functions of the experimental product, and Frame 2 - Human Organism Level - actions from product biochemical effects to tumor cell killing. Methods: STING agonists meet this PMK frame1 and activate STING (step 1) effectively. Activated STING binds TBK1 (step 2), then phosphorylates IRF3 (step 3), enters the nucleus (step 4), dimerizes (step 5) and binds specific regions of the DNA (step 6) for interferon transcription (step 7), which are secreted (step 8). Frame 2, the Human Organism Level - final common pathway requires many additional actions: Interferon must interact with immune cells (step 9) [within tumor or peripheral locations]. Very warm or hot tumors [contain functional immune cells] actively allow trafficking (homing) to the tumor (step 10) and infiltrating the tumor microenvironment (step 11). An activated immune cell can kill the tumor cell(s) (step 12). Cold tumors may not allow homing (step 10) to the tumor and/or infiltration (step 11) of the microenvironment. Results: PMKF modeling was applied to STING administration public data. Lack of efficacy was predicted if one of the critical pathway steps to tumor cell death are dysfunctional. Early steps (Frame 1) to interferon production and excretion into the environment occur within normal cells (antigen presenting cells) predictably. However, in the setting of malignancy, Frame 2 steps are often aberrant at the Organism Level. Cold tumors do not allow immune cell homing (step 10) and inhibit tumor infiltration (Step 11), tumor cell localization and killing of the tumor cell (step 12). Conclusions: STING Agonist are cutting edge therapies that perform PMKF Frame 1 well and increase interferon production. Application of the PMKF - Frame 2 demonstrate poor clinical activity may be explained by poor immune cell homing, infiltration and functional killing within the cold tumor microenvironment (dysfunctional steps 10, 11 and 12). Population selection may improve these results.

scholarly journals Combined Targeting of Distinct c-Myc and JunB Transcriptional Programs Inducing Synergistic Anti-Myeloma Activity

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2644-2644
Author(s):  
Judith Lind ◽  
Sonia Vallet ◽  
Karoline Kollmann ◽  
Osman Aksoy ◽  
Vincent Sunder-Plassmann ◽  
...  
Keyword(s):  
Cell Death ◽  
Tumor Cell ◽  
Cell Lines ◽  
Ex Vivo ◽  
Mapk Signaling ◽  
Loss Of Function ◽  
Tumor Cell Death ◽  
Expression Levels ◽  
Protein Levels

Abstract INTRODUCTION Transcription factors (TFs) are convergence points of signaling cascades that coordinate cell differentiation, proliferation and survival and are commonly deregulated in cancer, including multiple myeloma (MM). They contribute to the initiation of MM and promote tumor cell growth and drug resistance. Both cMyc, a merging point of the PI3K-, and JunB, a merging point of the MEK/MAPK-signaling pathway, play pivotal roles in MM. Exciting novel approaches to inhibit TFs like proteolysis-targeting-chimera (PROTAC) promise to lead to selective tumor cell death with little/no consequence for normal cells. However, redundancy phenomena of transcriptional programs are likely to challenge their efficacy. Here, we report our final results on combined targeting of distinct c-Myc & JunB transcriptional programs for MM therapy. METHODS MM cell lines and patient MM cells were analyzed. Following CRISPR-loss-of-function screens for cMyc & JunB across MM cell lines and correlation analyses in MM patient datasets, the functional relevance of BRD4/c-Myc- and MEK/JunB-induced TF programs was delineated using genomic and chemical approaches in 2D and 3D models of the bone marrow (BM) microenvironment. Specifically, effects of single or combined targeting of cMyc- and JunB-induced TF-programs were analyzed by flow cytometry, western blot, RNAseq, qPCR and luciferase assays. In vitro and ex vivo results were finally verified in a MM xenograft mouse model. RESULTS While CRISPR loss-of-function screens across various MM cell lines confirmed their growth dependency on cMyc and JunB, we did not observe correlative expression levels among these TFs, neither in the publicly available GSE6477 nor in the CoMMpass dataset. In contrast, a significant positive correlation was observed between Brd4 and cMyc, and MEK and JunB expression levels, respectively. The existence of two distinct Brd4/cMyc and MEK/JunB transcriptional programs in MM cells was subsequently supported by a lack of changes in cMyc mRNA/protein levels and resultant transcriptional activity upon JunB knockdown, and vice versa. Likewise, MZ-1, a novel PROTAC which targets Brd4, resulted in the inhibition of BMSC/IL-6- induced cMyc- but not JunB- upregulation. Conversely, neither the MEK inhibitor trametinib nor doxycycline-induced knockdown of BMSC/IL-6- induced JunB upregulation in TetshJunB/MM.1S cells reduced Brd4/c-Myc mRNA/protein levels. Importantly, the activity of MZ-1 and trametinib was predicted by Brd4 and JunB expression levels using mathematical models, respectively. Further, combination of MZ-1 with trametinib or JunB knockdown synergistically inhibited tumor cell proliferation, and induced cell death in a 2D and a dynamic 3D model of the MM-BM milieu. Finally, our in vitro and ex vivo results were confirmed in vivo, utilizing BMSC:TetshJunB/MM.1S vs. BMSC:TetshSCR/MM.1S-carrying NSG mice treated with MZ-1 with/without doxycycline or trametinib. CONCLUSION In summary, our data demonstrate for the first time the existence of non-overlapping cMyc and JunB-regulated TF programs providing a rationale for combined cMyc:JunB targeting treatment strategies in MM. Disclosures Vallet: Pfizer: Honoraria; MSD: Honoraria; Roche Pharmaceuticals: Consultancy. Podar: Celgene: Consultancy, Honoraria; Roche Pharmaceuticals: Research Funding; Janssen Pharmaceuticals: Consultancy, Honoraria; Amgen Inc.: Consultancy, Honoraria.

scholarly journals Ionizing radiation improves RIG-I mediated immunotherapy through enhanced p53 activation in malignant melanoma

2021 ◽  
Author(s):  
Silke Lambing ◽  
Stefan Holdenrieder ◽  
Patrick Müller ◽  
Christian Hagen ◽  
Stephan Garbe ◽  
...  
Keyword(s):  
Cell Death ◽  
Malignant Melanoma ◽  
Tumor Cell ◽  
Low Dose ◽  
Immune Cell ◽  
Cytotoxic T Cell ◽  
Great Promise ◽  
Type I ◽  
Tumor Cell Death

The activation of the innate immune receptor RIG-I is a promising approach in immunooncology and currently under investigation in clinical trials. RIG-I agonists elicit a strong immune activation in both tumor and immune cells and induce both direct and indirect immune cell-mediated tumor cell death which involves tumor-specific cytotoxic T-cell response and type I interferon-driven innate cytotoxic immunity. Besides RIG-I, irradiation is known to induce cytotoxic DNA damage resulting in tumor debulking followed by the induction of tumor-specific immunity. To date, it is unclear whether the molecular antitumor effects of RIG-I and irradiation are additive or even synergize. Here, we investigated the combination of RIG-I activation with radiotherapy in melanoma. We found that low dose x-ray irradiation enhanced the extent and immunogenicity of RIG-I mediated tumor cell death in human and murine melanoma cell lines and in the murine B16 melanoma model in vivo. Pathway analysis of transcriptomic data revealed a central role for p53 downstream of the combined treatment, which was corroborated using p53-/- B16 cells. In vivo, the additional effect of irradiation on immune cell activation and inhibition of tumor growth was lost in mice carrying p53-knockout B16 tumors, while the response to RIG-I stimulation in those mice was maintained. Thus, our results identify p53 as pivotal for the synergy of RIG-I with irradiation, resulting in potent induction of immunogenic tumor cell death. Consequently, low dose radiotherapy holds great promise to further improve the efficacy or RIG-I ligands especially in patients with malignant melanoma or other tumors exhibiting a functional p53 pathway.

Abstract 2788: VB6-845d tumor cell killing elicits biologic features of immunogenic cell death

2018 ◽  
Author(s):  
Rachelle L. Dillon ◽  
Shilpa Chooniedass ◽  
Arjune Premsukh ◽  
Glen C. MacDonald ◽  
Jeannick Cizeau ◽  
...  
Keyword(s):  
Cell Death ◽  
Tumor Cell ◽  
Cell Killing ◽  
Immunogenic Cell Death ◽  
Tumor Cell Killing

scholarly journals EXTH-44. INHIBITION OF MerTK ACTIVATES GLIOBLASTOMA-ASSOCIATED MACROPHAGES AND INDUCES TUMOR CELL DEATH IN GLIOMA MICROENVIRONMENT

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi91-vi91
Author(s):  
Yu-Ting Su ◽  
Madison Butler ◽  
Lee Hwang ◽  
Dragan Maric ◽  
Shelton Earp ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Growth ◽  
Tumor Microenvironment ◽  
Tumor Cell ◽  
Colony Formation ◽  
Glioblastoma Cells ◽  
Tumor Cell Death ◽  
Glioma Microenvironment

Abstract BACKGROUND Glioblastoma-associated macrophages and microglia (GAMs) are the predominant immune cells in the tumor microenvironment. Activation of MerTK, a receptor tyrosine kinase, triggers efferocytosis and polarizes GAMs to an immunosuppressive phenotype, promoting glioma growth. Our previous findings showed that UNC2371, a small-molecule inhibitor of MerTK, induced a less immunosuppressive phenotype of GAMs. Here, we investigate the role of MerTK inhibition on glioblastoma cells in the tumor microenvironment in vitro and in vivo. METHODS Cytotoxicity of UNC2371 in glioblastoma cells was determined by cell viability and colony formation assays. The protein expression of MerTK, AKT, and Erk were quantified by Western blotting in UNC2371-treated glioblastoma cells. A syngeneic GL261 mouse orthotopic glioblastoma model was used to evaluate the survival benefit of UNC2371 treatment. Fluorescent multiplex immunohistochemistry (IHC) was used to evaluate the expression of CD206, an anti-inflammatory marker on GAMs in murine brain tumor tissues. RESULTS UNC2371 inhibited GBM cell growth with an EC50 < 100 nM in both human U251 and mouse GL261 glioma cells, but not in GAMs. UNC2371-induced cell death and decreased cell proliferation were demonstrated by colony formation assays. UNC2371 decreased protein expression of phosphorylated MerTK, AKT, and Erk, which are essential for cell survival signaling, in U251 and GL261 cells. Furthermore, UNC2371 treatment prolonged survival in the mouse orthotopic GL261 glioblastoma model, suggesting that UNC2371 induces glioma cell death. A decreased of CD206+ GAMs was found in mice glioma tissues by fluorescent multiplex IHC, consistent with our previous findings in the in vitro cell-based assays. These data suggest that in addition to alleviate immunosuppression in the glioma microenvironment, UNC2371 directly inhibits GBM cell growth in vitro and in vivo. CONCLUSION Our findings suggest that UNC2371 has a therapeutic benefit via promoting GAM polarization towards proinflammatory status in the glioblastoma microenvironment and unexpectedly, inducing tumor cell death.

scholarly journals In-Depth Immune-Oncology Studies of the Tumor Microenvironment in a Humanized Melanoma Mouse Model

2021 ◽  
Vol 22 (3) ◽  
pp. 1011
Author(s):  
Jonathan Schupp ◽  
Arne Christians ◽  
Niklas Zimmer ◽  
Lukas Gleue ◽  
Helmut Jonuleit ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Tumor Microenvironment ◽  
Immune Cells ◽  
Sequential Analysis ◽  
Spatial Information ◽  
Immune Cell ◽  
Ex Vivo ◽  
Response To Therapy ◽  
Mouse Melanoma Model ◽  
Staining Methods

The presence and interaction of immune cells in the tumor microenvironment is of significant importance and has a great impact on disease progression and response to therapy. Hence, their identification is of high interest for prognosis and treatment decisions. Besides detailed phenotypic analyses of immune, as well as tumor cells, spatial analyses is an important parameter in the complex interplay of neoplastic and immune cells—especially when moving into focus efforts to develop and validate new therapeutic strategies. Ex vivo analysis of tumor samples by immunohistochemistry staining methods conserves spatial information is restricted to single markers, while flow cytometry (disrupting tissue into single cell suspensions) provides access to markers in larger numbers. Nevertheless, this comes at the cost of scarifying morphological information regarding tissue localization and cell–cell contacts. Further detrimental effects incurred by, for example, tissue digestion include staining artifacts. Consequently, ongoing efforts are directed towards methods that preserve, completely or in part, spatial information, while increasing the number of markers that can potentially be interrogated to the level of conventional flow cytometric methods. Progression in multiplex immunohistochemistry in the last ten years overcame the limitation to 1–2 markers in classical staining methods using DAB with counter stains or even pure chemical staining methods. In this study, we compared the multiplex method Chipcytometry to flow cytometry and classical IHC-P using DAB and hematoxylin. Chipcytometry uses frozen or paraffin-embedded tissue sections stained with readily available commercial fluorophore-labeled antibodies in repetitive cycles of staining and bleaching. The iterative staining approach enables sequential analysis of a virtually unlimited number of markers on the same sample, thereby identifying immune cell subpopulations in the tumor microenvironment in the present study in a humanized mouse melanoma model.

scholarly journals 681 An ex vivo 3D tumoroid model of fresh patient tissue (3D-EXplore) to assess the phagocytic activity of tumor resident innate immune cells

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A709-A709
Author(s):  
Kelly Guzman ◽  
Olivia McIntosh ◽  
Brittany Bunch ◽  
Jacob Yarinsky ◽  
Jared Ehrhart ◽  
...  
Keyword(s):  
Tumor Cell ◽  
Tumor Cells ◽  
Phagocytic Activity ◽  
Ex Vivo ◽  
Cell Killing ◽  
Innate Immune ◽  
Immune Microenvironment ◽  
Tumor Immune Microenvironment ◽  
The Impact ◽  
Tumor Cell Killing

BackgroundCD47 is an innate immune checkpoint receptor that is overexpressed on tumor cells and contributes to immune evasion through engagement of a myeloid-lineage inhibitory protein SIRPα. Blockade of the CD47-SIRPα interaction is proved to enhance the phagocytosis of cancer cells and to induce effective antitumor immune response. Here we developed a novel ex vivo platform using fresh patient tumor samples with intact stromal components and tumor immune microenvironment to assess the therapeutic activity of immunotherapeutic drugs targeting CD47-SIRPα signaling axis in combination with the human IgG1 αPD-L1 antibody avelumab.MethodsAll tumor samples were obtained with patient consent and relevant IRB approval. Unpropagated 3D tumoroids with intact TME measuring 150 µm in size were prepared from fresh tumor samples of renal cell carcinoma using proprietary technology developed at Nilogen Oncosystems. Tumoroids prepared from each patient's tumor sample were pooled to represent the tumor heterogeneity and treated ex vivo with phrodo-labeled avelumab alone or in combination with anti-CD47 or anti-SIRPα therapeutics.ResultsMultiparameter flow analysis demonstrated tumor binding of avelumab confirming drug penetration into the intact tumor stroma that is further corroborated by high content confocal analysis. Using our confocal-based tumor cell killing assay we were able to quantify drug-induced tumor cell killing ex vivo. We further documented the impact of anti-CD47 and anti-SIRPα therapeutics on phagocytosis of dead tumor cells by tumor resident macrophages and activation of innate and adaptive effector cells by flow cytometry and confocal imaging. Additionally, pHrodo-labeled bioparticles were used to corroborate treatment-mediated changes in the phagocytic activity of tumor resident macrophages.ConclusionsIn this comprehensive study we demonstrate that the 3D-EXplore ex vivo platform can be used to assess the efficacy of therapeutic blockade of CD47/SIRPα axis on stimulation of phagocytic process within an intact tumor immune microenvironment.

scholarly journals O2 Directly linking single T cell phenotype and function to genotype

2020 ◽  
Vol 8 (Suppl 2) ◽  
pp. A4.1-A4
Author(s):  
Y Bronevetsky
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cell ◽  
Cell Killing ◽  
Cell Phenotype ◽  
Functional Assay ◽  
Functional Screening ◽  
Tumor Cell Death ◽  
And Function ◽  
Tumor Cell Killing

T cell therapies for cancer treatment are challenging to develop because of the complex mechanisms and cell interactions that underly T cell-mediated tumor killing. Current technologies rely on correlating phenotype, function, and gene expression based on experiments performed on different populations of T cells because no one platform is able to assess cell surface marker expression, cytokine secretion, and tumor cell killing activity of the same T cell and recover this cell for downstream genomic analysis. Here we share two use cases - CAR-T cell functional screening and TCR sequence recovery following functional assay - that demonstrate how the T Cell Analysis Suite on the LightningTM optofluidic platform can be used to directly link T cell phenotype and function (IFNγ secretion and tumor cell killing) to genotype (TCR sequence recovery) at a single-cell level and on the same T cell, enabling deeper and more thorough characterization of how T cells mediate tumor cell death and potentially the development of more efficacious therapies.Disclosure InformationY. Bronevetsky: A. Employment (full or part-time); Significant; Berkeley Lights Inc.

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