Abstract 3090: Development and performance of the National Cancer Institute’s HTS384 NCI60 screen: A modernized platform to support drug discovery and development by the worldwide cancer research community
Authors: Coussens Nathan P. , Dexheimer Thomas S. , Jones Eric M. , Silvers Thomas , Britt John R. , Taylor Ronald C. , Kunkel Mark W. , Doroshow James H. , Teicher Beverly A. 
Published: March 2024

Abstract
The NCI60 screen was established in the 1980s and introduced in the 1990s as a screening service for the worldwide cancer research community. The screen features 60 human tumor cell lines that represent 9 cancer types including non-small cell lung, colon, central nervous system, ovarian, renal, prostate and breast, as well as leukemia and melanoma. On a weekly basis, synthetic molecules, natural products, and biologics are screened against the 60-cell line panel to evaluate their anticancer potential. On average, more than 13,000 compounds are screened annually for investigators throughout the world. Cell growth and cytotoxicity were assessed by the sulforhodamine B (SRB) assay, performed in a 96-well format, that measures cellular protein content with an absorbance readout and enabled unprecedented throughput in the 1980s. Although the SRB assay supported a robust screening platform for three decades, its sensitivity and throughput are limited. Recently the NCI’s DCTD introduced the HTS384 NCI60 as a modernized screening platform. While the same 60 cell lines are evaluated, the HTS384 NCI60 is fully automated and performed in a 384-well format to enable a higher throughput. Additionally, the 48-h test agent exposure period of the SRB assay was increased to 72 h, which may result in lower 50% growth inhibition values (GI50) for some agents. A highly sensitive CellTiter-Glo luminescence readout for cell viability, based on cellular ATP content, was selected to enable comparisons of the NCI60 monolayer data to 3D cell culture models where the same assay chemistry can be applied. The HTS384 NCI60 was evaluated by screening a library of 1,003 U.S. Food and Drug Administration-approved and investigational oncology agents at five concentrations. Assay performance of the 60 cell lines was assessed from the controls of 30 - 60 microplates per cell line. The average doubling time of the 60 cell lines during the 72-h exposure period ranged between 19 h for NCI-H460 (17 - 23 h, n = 57) to 117 h for MDA-MB-468 (43 - 272 h, n = 60). The average signal-to-background ratio of all 60 cell lines was 89 (3,413 microplates) and ranged from 6 for OVCAR-5 (3 - 13, n = 54) to 229 for SF-539 (141 - 566, n = 60). The average coefficient of variation for microplate vehicle controls (n = 14) of all 60 cell lines (n = 3,413) was 5.2% and ranged from 3.2% for SK-MEL-28 (1.4 - 7.7%, n = 60) to 11.1% for HL-60(TB) (5.2 - 20.2%, n = 59). The average Z’-factor value of all 60 cell lines (n = 3,413) was 0.82 and ranged between 0.66 for HL-60(TB) (0.39 - 0.84, n = 59) to 0.89 for MALME-3M (0.79 - 0.95, n = 59), SF-295 (0.49 - 0.97, n = 59), SK-MEL-28 (0.74 - 0.95, n = 60), and UO-31 (0.8 - 0.94, n = 54). The development and performance of the HTS384 NCI60 screen will be described. This project was funded in part with federal funds from the NCI, NIH, under contract no. HHSN261201500003I.
Citation Format: Nathan P. Coussens, Thomas S. Dexheimer, Eric M. Jones, Thomas Silvers, John R. Britt, Ronald C. Taylor, Mark W. Kunkel, James H. Doroshow, Beverly A. Teicher. Development and performance of the National Cancer Institute’s HTS384 NCI60 screen: A modernized platform to support drug discovery and development by the worldwide cancer research community [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3090.