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. 2020 Feb 24;11(2):149. doi: 10.1038/s41419-020-2335-1

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — a KHOS/NP and SJSA-1 cells were grown in specific spheroid media in low adherent plates and treated with 0–25 μM of pimozide. After 5 days, the spheroids were photographed and counted. Scale bars represent 100 μm. b Primary spheroids were collected and separated into single cells and re-plated. Pimozide treatment significantly inhibited osteosphere formation (*p < 0.05). c and d Western blot analyses of lysates from 3D cultures showed significant increase of DCLK1 short form in both KHOS/NP and SJSA-1 cells when compared to 2D cultures. Pimozide inhibits the expression of the DCLK1 isoforms. Western blot analyses of lysates from 3D cultures showed significant increase of CD44, CD133, Oct-4, and ABCG2 levels in both KHOS/NP and SJSA-1 cells when compared to 2D cultures which was suppressed by pimozide. e Sorting of anti-DCLK1 antibody-tagged phycoerythrin in KHOS/NP and SJAS-1 cells by flow cytometry. After 24 h, pimozide treatment resulted in a significant reduction in the number of DCLK1 positive cells.