Dataset Information

Dynamic Analysis of Human Natural Killer Cell Response at Single-Cell Resolution in B-Cell Non-Hodgkin Lymphoma.

ABSTRACT: Natural killer (NK) cells are phenotypically and functionally diverse lymphocytes that recognize and kill cancer cells. The susceptibility of target cancer cells to NK cell-mediated cytotoxicity depends on the strength and balance of regulatory (activating/inhibitory) ligands expressed on target cell surface. We performed gene expression arrays to determine patterns of NK cell ligands associated with B-cell non-Hodgkin lymphoma (b-NHL). Microarray analyses revealed significant upregulation of a multitude of NK-activating and costimulatory ligands across varied b-NHL cell lines and primary lymphoma cells, including ULBP1, CD72, CD48, and SLAMF6. To correlate genetic signatures with functional anti-lymphoma activity, we developed a dynamic and quantitative cytotoxicity assay in an integrated microfluidic droplet generation and docking array. Individual NK cells and target lymphoma cells were co-encapsulated in picoliter-volume droplets to facilitate monitoring of transient cellular interactions and NK cell effector outcomes at single-cell level. We identified significant variability in NK-lymphoma cell contact duration, frequency, and subsequent cytolysis. Death of lymphoma cells undergoing single contact with NK cells occurred faster than cells that made multiple short contacts. NK cells also killed target cells in droplets via contact-independent mechanisms that partially relied on calcium-dependent processes and perforin secretion, but not on cytokines (interferon-? or tumor necrosis factor-?). We extended this technique to characterize functional heterogeneity in cytolysis of primary cells from b-NHL patients. Tumor cells from two diffuse large B-cell lymphoma patients showed similar contact durations with NK cells; primary Burkitt lymphoma cells made longer contacts and were lysed at later times. We also tested the cytotoxic efficacy of NK-92, a continuously growing NK cell line being investigated as an antitumor therapy, using our droplet-based bioassay. NK-92 cells were found to be more efficient in killing b-NHL cells compared with primary NK cells, requiring shorter contacts for faster killing activity. Taken together, our combined genetic and microfluidic analysis demonstrate b-NHL cell sensitivity to NK cell-based cytotoxicity, which was associated with significant heterogeneity in the dynamic interaction at single-cell level.

SUBMITTER: Sarkar S

PROVIDER: S-EPMC5735063 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Publications

Dynamic Analysis of Human Natural Killer Cell Response at Single-Cell Resolution in B-Cell Non-Hodgkin Lymphoma.

Sarkar Saheli S Sabhachandani Pooja P Ravi Dashnamoorthy D Potdar Sayalee S Purvey Sneha S Beheshti Afshin A Evens Andrew M AM Konry Tania T

Frontiers in immunology 20171214

Natural killer (NK) cells are phenotypically and functionally diverse lymphocytes that recognize and kill cancer cells. The susceptibility of target cancer cells to NK cell-mediated cytotoxicity depends on the strength and balance of regulatory (activating/inhibitory) ligands expressed on target cell surface. We performed gene expression arrays to determine patterns of NK cell ligands associated with B-cell non-Hodgkin lymphoma (b-NHL). Microarray analyses revealed significant upregulation of a ...[more]

PMID: 29312292

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Project description:Natural Killer (NK) cells are phenotypically and functionally diverse lymphocytes that recognize and kill cancer cells. The susceptibility of target cancer cells to NK cell-mediated cytotoxicity depends on the strength and balance of regulatory (activating/inhibitory) ligands expressed on target cell surface. We performed gene expression arrays to determine patterns of NK cell ligands associated with B-cell non-Hodgkin lymphoma (b-NHL). Microarray analyses revealed differential upregulation of a multitude of NK-activating and costimulatory ligands across varied lymphoma cell lines and b-NHL cells including ULBP1, CD72, CD48 and SLAMF6. To correlate genetic signatures with functional anti-lymphoma activity, we developed a dynamic and quantitative cytotoxicity assay in an integrated microfluidic droplet array. This assay facilitated the examination of transient interactions between NK cells and target lymphoma cells and also the functional heterogeneity in NK cell effector outcomes at single-cell level. Individual NK cells and target lymphoma cells were co-encapsulated in picoliter-volume droplets and imaged over time to quantify heterotypic cell-cell interaction. We identified significant variability in NK-lymphoma cell contact duration, frequency and subsequent cytolysis. The death of lymphoma cells undergoing single contact with NK cells occurred faster than cells that made multiple short contacts. NK cells also killed target cells in droplets via contact-independent mechanisms that partially relied on calcium-dependent processes and perforin secretion, but not on cytokines (IFN-γ or TNF-α). We extended this technique to characterize cytolysis of primary cells from b-NHL patients. Diffuse large B-cell lymphoma cells from two patients showed similar contact durations with NK cells; primary Burkitt lymphoma cells made longer contacts and were lysed at later times. We further tested the cytotoxic efficacy of NK-92, a continuously growing NK cell line being investigated as an anti-tumor therapy, using our droplet-based bioassay. NK-92 cells were found to be more efficient in killing b-NHL cells compared with primary/fresh NK cells, requiring shorter contacts for faster killing activity. Taken together, our combined genetic and microfluidic analysis demonstrate significant heterogeneity in the dynamic interaction and associated cytotoxicity between NK cellular therapy and b-NHL cells.

Project description:Extranodal natural killer/T-cell lymphoma, nasal type (ENKL), is a rare malignancy of Non-Hodgkin lymphoma characterized by an aggressive clinical course and poor prognosis. It shows strong association with Epstein-Barr virus infection and occurs more commonly in Asia and Latin America. Various genetic alterations have been identified in ENKL by gene expression profiling and sequencing techniques. The frequent deletion of chromosome 6q21 was reported to lead to the silence of several tumor suppressor genes. Also, there have been novel genetic mutations that were recently uncovered and were found to frequently activate several oncogenic pathways, including the JAK/STAT, NF-?B, and MAPK pathways. Besides, we believe that deregulated single genes and epigenetic dysregulation might be relevant to the mechanism of this disease and thus, may have the potential to shed lights on the development of new therapeutic strategies. The consensus on the standard treatment for ENKL has not yet been currently established. For localized ENKL patients, radiotherapy with concurrent chemotherapy and sequential patterns of chemotherapy and radiotherapy are recommended as first-line therapy. As for advanced or relapsed/refractory ENKL patients, the application of non-anthracycline-containing regimens have significantly improved the clinical outcome, contributing to higher response rate, longer overall survival and progression-free survival. Hematopoietic stem cell transplantation is widely recommended for consolidation after a complete remission or partial remission has been achieved. The anti-programmed death 1 antibody, an immune checkpoint inhibitor, has demonstrated favorable results in treating relapsed or refractory ENKL. Of the current ENKL treatment, researchers are still striving to validate how radiotherapy and chemotherapy should be optimally combined and which of the non-anthracycline-containing regimens is superior. In this review, we summarize the main genetic alterations frequently found in ENKL and their role in providing new insights into the therapeutic targets of this disease, and highlight the recent findings regarding new biologic markers, novel therapeutic strategies applied to this intriguing neoplasm.

Dataset Information

Dynamic Analysis of Human Natural Killer Cell Response at Single-Cell Resolution in B-Cell Non-Hodgkin Lymphoma.

Publications

Dynamic Analysis of Human Natural Killer Cell Response at Single-Cell Resolution in B-Cell Non-Hodgkin Lymphoma.

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