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Chemical screening identifies novel small molecule activators of natural killer cell cytotoxicity against cancer cells

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Abstract

Natural killer (NK) cells are cytotoxic lymphocytes that play a major role in the innate immune system. NK cells exhibit potent cytotoxic activity against cancer cells and virally infected cells without antigen priming. These unique cytotoxic properties make NK cells a promising therapeutic against cancer. Limitations of NK cell therapy include deficiencies in high clinical efficacy often due to a need for a high NK cell to target cell ratio to achieve effective killing. In order to address the suboptimal efficacy of current adoptive NK cell therapy, a high throughput screen (HTS) was designed and performed to identify drug-like compounds that increase NK cytotoxic activity against tumor cells without affecting the normal cells. This screen was performed in a 384-well plate format utilizing an expanded primary NK cell product and ovarian cancer cells as a target cell (TC) line. Of the 8000 diverse small molecules screened, 16 hits were identified (0.2% hit rate) based on both a robust Z (RZ) score < -3 and a greater than 10% increase in NK cell killing. A validation screen had a confirmation rate of 70%. Select compounds were further validated and characterized by additional cytotoxicity assays including activity against multiple blood cancer and solid tumor cell lines, with no effect on primary human T cells. This work demonstrates that high-throughput screening can be reliably used to identify compounds that increase NK tumoricidal activity in vitro that can be further investigated and translated for potential clinical application. Précis: Our work led to the identification of promising compound that potently increases NK cell-mediated killing of a variety of different cancer cells, but no impact on the killing of normal cells. This compound demonstrates the utility of this assay.

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Availability of data and material

The datasets generated during and/or analyzed during the current study are available on PubChem.

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Acknowledgements

This research was supported by the following Case Comprehensive Cancer Center Shared Resources: Small Molecule Drug Development, Hematopoietic Biorepository and Cellular Therapy and Cytometry & Imaging Microscopy Shared Resource of the Case Comprehensive Cancer Center (P30CA043703). This work was also in part supported by an NIH-sponsored S10 grant (1S10OD018005-01 to B.A.P.) for the IN Cell Analyzer 6000 and R01CA259011. We thank Yong Han for performing LC/MS and HPLC analysis of compound 2.

Funding

P30CA043703, 1S10OD018005-01, R01CA259011. National Cancer Institute (US), P30CA043703,National Institutes of Health (US),1S10OD018005-01,Bruce Posner, National Cancer Institute,R01CA259011,David Wald

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Authors and Affiliations

  1. Department of Pathology, Case Western Reserve University, Cleveland, OH, USA

    Grace Lee, Sheela Karunanithi, Zachary Jackson & David Wald

  2. High-Throughput Screening Core, Department of Biochemistry, UT Southwestern Medical Center, Dallas, TX, USA

    Bruce Posner, Hanspeter Niederstrasser & Hong Cheng

  3. Small Molecule and Drug Discovery Facility, Case Western Reserve University, Cleveland, OH, USA

    Yuriy Federov

  4. Department of Medicine, University Hospitals Cleveland Medical Center, Cleveland, OH, USA

    Shivaprasad Manjappa

  5. Idaho College of Osteopathic Medicine, Meridian, ID, USA

    Karam Musaitif

  6. Department of Hematology, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, China

    Huaiyu Wang

  7. Department of Pathology, Louis Stokes Cleveland VA Medical Center, Cleveland, OH, USA

    David Wald

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  1. Grace Lee
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Contributions

G.L., K.M., H.W. and D.W. wrote and/or edited the manuscript. D.W., G.L., S.K., Y.F., and B.P. designed experiments. G.L., S.K., and H.N. performed the screening. D.W., Y.F., B.P., G.L., S.K., and H.C. analyzed the data. S.M. and Z.J. assisted with experiments.

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Correspondence to David Wald.

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Lee, G., Karunanithi, S., Posner, B. et al. Chemical screening identifies novel small molecule activators of natural killer cell cytotoxicity against cancer cells. Cancer Immunol Immunother 71, 1671–1680 (2022). https://doi.org/10.1007/s00262-021-03117-w

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