{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,10,16]],"date-time":"2026-10-16T01:25:58Z","timestamp":1792113958542,"version":"4.4.0"},"posted":{"date-parts":[[2025,10,10]]},"group-title":"Cell Biology","reference-count":60,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T00:00:00Z","timestamp":1760054400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"award":["5T32GM135122"],"award-info":[{"award-number":["5T32GM135122"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"id":[{"id":"https:\/\/ror.org\/019af4n30","id-type":"ROR","asserted-by":"publisher"}]},{"DOI":"10.13039\/100013061","name":"Ontario Genomics","doi-asserted-by":"crossref","id":[{"id":"10.13039\/100013061","id-type":"DOI","asserted-by":"crossref"}]},{"id":[{"id":"https:\/\/ror.org\/02ddkpn78","id-type":"ROR","asserted-by":"publisher"}]},{"id":[{"id":"https:\/\/ror.org\/000az4664","id-type":"ROR","asserted-by":"publisher"}]},{"DOI":"10.13039\/100009744","name":"Eshelman Institute for Innovation, University of North Carolina at Chapel Hill","doi-asserted-by":"crossref","id":[{"id":"10.13039\/100009744","id-type":"DOI","asserted-by":"crossref"}]},{"id":[{"id":"https:\/\/ror.org\/029s29983","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,1,21]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>A pivotal part of kinase chemical probe and drug development is assessment of the selectivity of a putative lead compound. While there is no consensus around the panel size or the type of assay(s) that are most appropriate, there is concurrence that gauging the number of on- and off-targets of a kinase inhibitor is essential. As pharmacology takes place in cells, we have compared profiling results for ten kinase inhibitors generated using the cell-free assays to those obtained when a panel of cellular target engagement NanoBRET assays is used to assess selectivity in intact cells. This is the first systematic comparison of these two approaches across a broad kinase panel. Comparison of the data sets demonstrates divergent results that can influence chemical probe prioritization. We identify unanticipated kinase interactions in cells for type II kinase inhibitors that are not observed in biochemical, cell-free systems. Furthermore, we characterize TPKI-39 as a DDR1, DDR2, and FLT1 chemical probe based on its in-cell selectivity profile.<\/jats:p>\n                <jats:sec>\n                  <jats:title>For Table of Contents Only<\/jats:title>\n                  <jats:fig id=\"ufig1\" position=\"float\" fig-type=\"figure\" orientation=\"portrait\">\n                    <jats:graphic xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" xlink:href=\"681452v2_ufig1\" position=\"float\" orientation=\"portrait\"\/>\n                  <\/jats:fig>\n                <\/jats:sec>","DOI":"10.1101\/2025.10.09.681452","type":"posted-content","created":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T02:05:17Z","timestamp":1760148317000},"source":"Crossref","is-referenced-by-count":2,"title":["Cellular Context Influences Kinase Inhibitor Selectivity"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0009-0006-6715-4073","authenticated-orcid":false,"given":"Matthew J.","family":"Binder","sequence":"first","affiliation":[{"name":"Structural Genomics Consortium, UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4241-9873","authenticated-orcid":false,"given":"Frances M.","family":"Bashore","sequence":"additional","affiliation":[{"name":"Structural Genomics Consortium, UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kaitlin K. Dunn","family":"Hoffman","sequence":"additional","affiliation":[{"name":"Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cameron","family":"Damgaard","sequence":"additional","affiliation":[{"name":"Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2803-0338","authenticated-orcid":false,"given":"Michael","family":"Slater","sequence":"additional","affiliation":[{"name":"Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5973-5798","authenticated-orcid":false,"given":"David H.","family":"Drewry","sequence":"additional","affiliation":[{"name":"Structural Genomics Consortium, UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA"},{"name":"UNC Lineberger Comprehensive Cancer Center, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8505-5227","authenticated-orcid":false,"given":"Matthew B.","family":"Robers","sequence":"additional","affiliation":[{"name":"Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4779-9932","authenticated-orcid":false,"given":"Alison D.","family":"Axtman","sequence":"additional","affiliation":[{"name":"Structural Genomics Consortium, UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026020608193433000_2025.10.09.681452v2.1","doi-asserted-by":"publisher","DOI":"10.1038\/nchembio.296"},{"key":"2026020608193433000_2025.10.09.681452v2.2","doi-asserted-by":"publisher","DOI":"10.1038\/nchembio.1867"},{"key":"2026020608193433000_2025.10.09.681452v2.3","doi-asserted-by":"publisher","DOI":"10.4155\/fmc-2019-0231"},{"key":"2026020608193433000_2025.10.09.681452v2.4","doi-asserted-by":"publisher","DOI":"10.1016\/j.chembiol.2010.05.013"},{"key":"2026020608193433000_2025.10.09.681452v2.5","doi-asserted-by":"publisher","DOI":"10.1016\/S0092-8674(00)81092-2"},{"key":"2026020608193433000_2025.10.09.681452v2.6","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbc.2022.102247"},{"key":"2026020608193433000_2025.10.09.681452v2.7","doi-asserted-by":"publisher","DOI":"10.1016\/j.phrs.2015.10.021"},{"key":"2026020608193433000_2025.10.09.681452v2.8","doi-asserted-by":"publisher","DOI":"10.1158\/0008-5472.CAN-05-4187"},{"issue":"15","key":"2026020608193433000_2025.10.09.681452v2.9","first-page":"4236","article-title":"Crystal structures of the kinase domain of c-Abl in complex with the small molecule inhibitors PD173955 and imatinib (STI-571)","volume":"62","year":"2002","journal-title":"Cancer Res"},{"key":"2026020608193433000_2025.10.09.681452v2.10","doi-asserted-by":"publisher","DOI":"10.1126\/science.1095920"},{"key":"2026020608193433000_2025.10.09.681452v2.11","doi-asserted-by":"publisher","DOI":"10.1021\/cb500129t"},{"key":"2026020608193433000_2025.10.09.681452v2.12","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1811158115"},{"key":"2026020608193433000_2025.10.09.681452v2.13","doi-asserted-by":"publisher","DOI":"10.1126\/science.1075762"},{"key":"2026020608193433000_2025.10.09.681452v2.14","doi-asserted-by":"publisher","DOI":"10.1038\/nbt.1990"},{"key":"2026020608193433000_2025.10.09.681452v2.15","doi-asserted-by":"publisher","DOI":"10.1016\/j.chembiol.2017.10.010"},{"key":"2026020608193433000_2025.10.09.681452v2.16","doi-asserted-by":"publisher","DOI":"10.1038\/ncomms10091"},{"key":"2026020608193433000_2025.10.09.681452v2.17","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-020-16559-0"},{"key":"2026020608193433000_2025.10.09.681452v2.18","doi-asserted-by":"publisher","DOI":"10.1016\/j.xpro.2021.100822"},{"key":"2026020608193433000_2025.10.09.681452v2.19","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-024-49896-5"},{"key":"2026020608193433000_2025.10.09.681452v2.20","doi-asserted-by":"publisher","DOI":"10.1021\/acschembio.5b01018"},{"key":"2026020608193433000_2025.10.09.681452v2.21","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkz1023"},{"key":"2026020608193433000_2025.10.09.681452v2.22","doi-asserted-by":"publisher","DOI":"10.1158\/1078-0432.CCR-20-2563"},{"key":"2026020608193433000_2025.10.09.681452v2.23","doi-asserted-by":"publisher","DOI":"10.1021\/jm500118j"},{"key":"2026020608193433000_2025.10.09.681452v2.24","doi-asserted-by":"publisher","DOI":"10.1016\/j.phrs.2024.107059"},{"key":"2026020608193433000_2025.10.09.681452v2.25","doi-asserted-by":"publisher","DOI":"10.1111\/j.1476-5381.2009.00604.x"},{"key":"2026020608193433000_2025.10.09.681452v2.26","doi-asserted-by":"publisher","DOI":"10.1021\/acs.jmedchem.4c02531"},{"key":"2026020608193433000_2025.10.09.681452v2.27","doi-asserted-by":"publisher","DOI":"10.3390\/molecules2917158"},{"key":"2026020608193433000_2025.10.09.681452v2.28","doi-asserted-by":"publisher","DOI":"10.1021\/acs.jmedchem.4c00962"},{"key":"2026020608193433000_2025.10.09.681452v2.29","doi-asserted-by":"publisher","DOI":"10.1016\/j.chembiol.2018.11.005"},{"key":"2026020608193433000_2025.10.09.681452v2.30","doi-asserted-by":"publisher","DOI":"10.1016\/j.str.2013.01.016"},{"key":"2026020608193433000_2025.10.09.681452v2.31","doi-asserted-by":"publisher","DOI":"10.1021\/acs.jmedchem.6b01751"},{"key":"2026020608193433000_2025.10.09.681452v2.32","doi-asserted-by":"publisher","DOI":"10.1038\/s41418-019-0347-0"},{"key":"2026020608193433000_2025.10.09.681452v2.33","doi-asserted-by":"publisher","DOI":"10.1039\/D5MD00317B"},{"key":"2026020608193433000_2025.10.09.681452v2.34","doi-asserted-by":"publisher","DOI":"10.1186\/s12967-014-0294-y"},{"key":"2026020608193433000_2025.10.09.681452v2.35","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1600934113"},{"key":"2026020608193433000_2025.10.09.681452v2.36","doi-asserted-by":"publisher","DOI":"10.1056\/NEJMoa2026982"},{"key":"2026020608193433000_2025.10.09.681452v2.37","doi-asserted-by":"publisher","DOI":"10.1007\/s00280-017-3240-x"},{"key":"2026020608193433000_2025.10.09.681452v2.38","doi-asserted-by":"publisher","DOI":"10.1182\/bloodadvances.2019000268"},{"key":"2026020608193433000_2025.10.09.681452v2.39","doi-asserted-by":"publisher","DOI":"10.1002\/cpdd.1558"},{"key":"2026020608193433000_2025.10.09.681452v2.40","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0181585"},{"key":"2026020608193433000_2025.10.09.681452v2.41","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-020-72869-9"},{"key":"2026020608193433000_2025.10.09.681452v2.42","doi-asserted-by":"publisher","DOI":"10.1038\/s41589-023-01459-3"},{"key":"2026020608193433000_2025.10.09.681452v2.43","doi-asserted-by":"publisher","DOI":"10.1016\/j.bmc.2012.10.004"},{"key":"2026020608193433000_2025.10.09.681452v2.44","doi-asserted-by":"publisher","DOI":"10.1016\/j.chembiol.2017.11.004"},{"key":"2026020608193433000_2025.10.09.681452v2.45","doi-asserted-by":"publisher","DOI":"10.1021\/acschembio.5b00655"},{"key":"2026020608193433000_2025.10.09.681452v2.46","doi-asserted-by":"publisher","DOI":"10.1021\/acs.jmedchem.1c00868"},{"key":"2026020608193433000_2025.10.09.681452v2.47","doi-asserted-by":"publisher","DOI":"10.1111\/jnc.13877"},{"key":"2026020608193433000_2025.10.09.681452v2.48","doi-asserted-by":"publisher","DOI":"10.1021\/cb5000949"},{"key":"2026020608193433000_2025.10.09.681452v2.49","doi-asserted-by":"publisher","DOI":"10.1021\/cb400430t"},{"key":"2026020608193433000_2025.10.09.681452v2.50","doi-asserted-by":"publisher","DOI":"10.1021\/acs.jmedchem.6b00140"},{"key":"2026020608193433000_2025.10.09.681452v2.51","doi-asserted-by":"publisher","DOI":"10.1016\/S1097-2765(00)80003-9"},{"key":"2026020608193433000_2025.10.09.681452v2.52","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.25716"},{"key":"2026020608193433000_2025.10.09.681452v2.53","doi-asserted-by":"publisher","DOI":"10.1016\/j.drudis.2014.09.025"},{"key":"2026020608193433000_2025.10.09.681452v2.54","doi-asserted-by":"publisher","DOI":"10.3389\/fcell.2022.836797"},{"key":"2026020608193433000_2025.10.09.681452v2.55","doi-asserted-by":"publisher","DOI":"10.1016\/j.jprot.2011.10.007"},{"key":"2026020608193433000_2025.10.09.681452v2.56","doi-asserted-by":"publisher","DOI":"10.1016\/j.chembiol.2020.12.013"},{"key":"2026020608193433000_2025.10.09.681452v2.57","doi-asserted-by":"publisher","DOI":"10.1038\/nbt.3374"},{"key":"2026020608193433000_2025.10.09.681452v2.58","doi-asserted-by":"publisher","DOI":"10.1016\/j.chembiol.2020.12.013"},{"key":"2026020608193433000_2025.10.09.681452v2.59","doi-asserted-by":"publisher","DOI":"10.1007\/978-1-0716-3397-7_8"},{"key":"2026020608193433000_2025.10.09.681452v2.60","doi-asserted-by":"publisher","DOI":"10.1016\/j.cels.2018.07.001"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1101\/2025.10.09.681452","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,24]],"date-time":"2026-06-24T05:06:18Z","timestamp":1782277578000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.1101\/2025.10.09.681452"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,10,10]]},"references-count":60,"URL":"https:\/\/doi.org\/10.1101\/2025.10.09.681452","relation":{"is-preprint-of":[{"id-type":"doi","id":"10.1021\/acs.jmedchem.5c02916","asserted-by":"subject"}]},"subject":[],"published":{"date-parts":[[2025,10,10]]},"subtype":"preprint"}}