{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T22:07:10Z","timestamp":1768514830869,"version":"3.49.0"},"posted":{"date-parts":[[2024,8,3]]},"group-title":"Microbiology","reference-count":38,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2024,8,3]],"date-time":"2024-08-03T00:00:00Z","timestamp":1722643200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nd\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2024,8,3]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>The rise of \u03b2-lactam resistance necessitates new strategies to combat bacterial infections. We purposefully engineered the \u03b2-lactam prodrug AcephPT to exploit \u03b2-lactamase activity to selectively suppress resistant bacteria producing extended-spectrum-\u03b2-lactamases (ESBLs). Selective targeting of resistant bacteria requires avoiding interaction with penicillin-binding proteins, the conventional targets of \u03b2-lactam antibiotics, while maintaining recognition by ESBLs to activate AcephPT only in resistant cells. Computational approaches provide a rationale for structural modifications to the prodrug to achieve this biased activity. We show AcephPT selectively suppresses gram-negative ESBL-producing bacteria in clonal populations and in mixed microbial cultures, with effective selectivity for both lab strains and clinical isolates expressing ESBLs. Time-course NMR experiments confirm hydrolytic activation of AcephPT exclusively by ESBL-producing bacteria. In mixed microbial cultures, AcephPT suppresses proliferation of ESBL-producing strains while sustaining growth of \u03b2-lactamase-non-producing bacteria, highlighting its potential to combat \u03b2-lactam resistance while promoting antimicrobial stewardship.<\/jats:p>\n                <jats:sec>\n                  <jats:title>Abstract Figure<\/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=\"606422v1_ufig1\" position=\"float\" orientation=\"portrait\"\/>\n                  <\/jats:fig>\n                <\/jats:sec>","DOI":"10.1101\/2024.08.02.606422","type":"posted-content","created":{"date-parts":[[2024,8,4]],"date-time":"2024-08-04T01:07:28Z","timestamp":1722733648000},"source":"Crossref","is-referenced-by-count":0,"title":["An engineered prodrug selectively suppresses \u03b2-lactam resistant bacteria in a mixed microbial setting"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-8116-4714","authenticated-orcid":false,"given":"Addison M.","family":"Duda","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Helena R.","family":"Ma","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"C\u00e9sar A.","family":"Villalobos","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sophia A.","family":"Kuhn","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Katherine","family":"He","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sarah R.","family":"Seay","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Abigail C.","family":"Jackson","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Christine M.","family":"Suh","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Elena A.","family":"Puccio","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Deverick J.","family":"Anderson","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"suffix":"Jr.","given":"Vance G.","family":"Fowler","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lingchong","family":"You","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9015-0998","authenticated-orcid":false,"given":"Katherine J.","family":"Franz","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2024080604000630000_2024.08.02.606422v1.1","doi-asserted-by":"publisher","DOI":"10.1101\/cshperspect.a025247"},{"key":"2024080604000630000_2024.08.02.606422v1.2","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.00047-19"},{"issue":"5","key":"2024080604000630000_2024.08.02.606422v1.3","first-page":"L643","article-title":"; for the Antibacterial Resistance Leadership Group","volume":"323","year":"2022","journal-title":"Confronting Antimicrobial Resistance Together. Am. J. Physiol. Lung Cell Mol"},{"key":"2024080604000630000_2024.08.02.606422v1.4","unstructured":"Centers for Disease Control and Prevention (U.S.). Antibiotic Resistance Threats in the United States, 2019; Centers for Disease Control and Prevention (U.S.), 2019."},{"issue":"10325","key":"2024080604000630000_2024.08.02.606422v1.5","doi-asserted-by":"crossref","first-page":"629","DOI":"10.1016\/S0140-6736(21)02724-0","article-title":"Global Burden of Bacterial Antimicrobial Resistance in 2019: A Systematic Analysis","volume":"399","year":"2022","journal-title":"The Lancet"},{"key":"2024080604000630000_2024.08.02.606422v1.6","unstructured":"Core Elements of Antibiotic Stewardship | Antibiotic Use | CDC. https:\/\/www.cdc.gov\/antibiotic-use\/core-elements\/index.html."},{"issue":"12","key":"2024080604000630000_2024.08.02.606422v1.7","first-page":"e01733","article-title":"Estimating the Size of the U.S. Market for New Antibiotics with Activity against Carbapenem-Resistant Enterobacteriaceae","volume":"63","year":"2019","journal-title":"Antimicrob Agents Chemother"},{"key":"2024080604000630000_2024.08.02.606422v1.8","doi-asserted-by":"publisher","DOI":"10.1038\/d41586-020-02418-x"},{"issue":"10","key":"2024080604000630000_2024.08.02.606422v1.9","doi-asserted-by":"crossref","first-page":"1992","DOI":"10.1021\/acsinfecdis.2c00315","article-title":"\u03b2-Lactamase-Mediated Fragmentation: Historical Perspectives and Recent Advances in Diagnostics, Imaging, and Antibacterial Design","volume":"8","year":"2022","journal-title":"ACS Infect. Dis"},{"issue":"13","key":"2024080604000630000_2024.08.02.606422v1.10","doi-asserted-by":"crossref","first-page":"9141","DOI":"10.1021\/acs.jmedchem.1c00362","article-title":"Novel Cephalosporin Conjugates Display Potent and Selective Inhibition of Imipenemase-Type Metallo-\u03b2-Lactamases","volume":"64","year":"2021","journal-title":"J. Med. Chem"},{"issue":"11","key":"2024080604000630000_2024.08.02.606422v1.11","doi-asserted-by":"crossref","first-page":"3806","DOI":"10.1002\/chem.202004694","article-title":"Cephalosporin Prodrug Inhibitors Overcome Metallo-\u03b2-Lactamase Driven Antibiotic Resistance","volume":"27","year":"2021","journal-title":"Chem. Eur. J"},{"issue":"8","key":"2024080604000630000_2024.08.02.606422v1.12","doi-asserted-by":"crossref","first-page":"1433","DOI":"10.1021\/acsinfecdis.9b00112","article-title":"Dual-Pharmacophore Pyrithione-Containing Cephalosporins Kill Both Replicating and Nonreplicating Mycobacterium Tuberculosis","volume":"5","year":"2019","journal-title":"ACS Infect. Dis"},{"key":"2024080604000630000_2024.08.02.606422v1.13","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.10.2.245"},{"issue":"5","key":"2024080604000630000_2024.08.02.606422v1.14","doi-asserted-by":"crossref","first-page":"1264","DOI":"10.1021\/acsinfecdis.0c00083","article-title":"A Cephalosporin Prochelator Inhibits New Delhi Metallo-\u03b2-Lactamase 1 without Removing Zinc","volume":"6","year":"2020","journal-title":"ACS Infect. Dis"},{"issue":"6","key":"2024080604000630000_2024.08.02.606422v1.15","doi-asserted-by":"crossref","first-page":"1019","DOI":"10.1021\/acsinfecdis.8b00037","article-title":"Copper Influences the Antibacterial Outcomes of a \u03b2-Lactamase-Activated Prochelator against Drug-Resistant Bacteria","volume":"4","year":"2018","journal-title":"ACS Infect. Dis"},{"issue":"8","key":"2024080604000630000_2024.08.02.606422v1.16","doi-asserted-by":"crossref","first-page":"1546","DOI":"10.1021\/acsinfecdis.3c00110","article-title":"Zidovudine-\u03b2-Lactam Pronucleoside Strategy for Selective Delivery into Gram-Negative Bacteria Triggered by \u03b2-Lactamases","volume":"9","year":"2023","journal-title":"ACS Infect. Dis"},{"issue":"1","key":"2024080604000630000_2024.08.02.606422v1.17","doi-asserted-by":"crossref","first-page":"214","DOI":"10.1021\/acschembio.0c00900","article-title":"Protein Folding Stability Changes Across the Proteome Reveal Targets of Cu Toxicity in E. Coli","volume":"16","year":"2021","journal-title":"ACS Chem. Biol"},{"key":"2024080604000630000_2024.08.02.606422v1.18","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0098042"},{"issue":"1","key":"2024080604000630000_2024.08.02.606422v1.19","first-page":"e01651","article-title":"Penicillin-Binding Protein 3 Is Essential for Growth of Pseudomonas Aeruginosa","volume":"61","year":"2017","journal-title":"Antimicrob Agents Chemother"},{"key":"2024080604000630000_2024.08.02.606422v1.20","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.00774-09"},{"key":"2024080604000630000_2024.08.02.606422v1.21","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-018-06839-1"},{"key":"2024080604000630000_2024.08.02.606422v1.22","doi-asserted-by":"publisher","DOI":"10.4103\/ijmr.IJMR_594_15"},{"key":"2024080604000630000_2024.08.02.606422v1.23","doi-asserted-by":"publisher","DOI":"10.1111\/j.1574-6976.2007.00095.x"},{"key":"2024080604000630000_2024.08.02.606422v1.24","doi-asserted-by":"publisher","DOI":"10.1111\/j.1574-6976.2008.00105.x"},{"key":"2024080604000630000_2024.08.02.606422v1.25","doi-asserted-by":"crossref","first-page":"180","DOI":"10.3389\/fmolb.2020.00180","article-title":"Application of Fragment-Based Drug Discovery to Versatile Targets","volume":"7","year":"2020","journal-title":"Front. Mol. Biosci"},{"key":"2024080604000630000_2024.08.02.606422v1.26","doi-asserted-by":"crossref","first-page":"93","DOI":"10.3389\/fchem.2020.00093","article-title":"In Silico Strategies to Support Fragment-to-Lead Optimization in Drug Discovery","volume":"8","year":"2020","journal-title":"Front. Chem"},{"key":"2024080604000630000_2024.08.02.606422v1.27","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0023606"},{"key":"2024080604000630000_2024.08.02.606422v1.28","doi-asserted-by":"publisher","DOI":"10.1016\/S1473-3099(11)70059-7"},{"key":"2024080604000630000_2024.08.02.606422v1.29","first-page":"110","article-title":"CTX-M Enzymes: Origin and Diffusion","volume":"3","year":"2012","journal-title":"Front. Microbio"},{"key":"2024080604000630000_2024.08.02.606422v1.30","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.44.4.891-897.2000"},{"issue":"2","key":"2024080604000630000_2024.08.02.606422v1.31","doi-asserted-by":"crossref","first-page":"236","DOI":"10.3390\/antibiotics11020236","article-title":"Comprehensive Analysis of Imipenemase (IMP)-Type Metallo-\u03b2-Lactamase: A Global Distribution Threatening Asia","volume":"11","year":"2022","journal-title":"Antibiotics"},{"key":"2024080604000630000_2024.08.02.606422v1.32","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.18.4.657-686.2005"},{"key":"2024080604000630000_2024.08.02.606422v1.33","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.00117-13"},{"key":"2024080604000630000_2024.08.02.606422v1.34","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M109.053819"},{"key":"2024080604000630000_2024.08.02.606422v1.35","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.01804-15"},{"key":"2024080604000630000_2024.08.02.606422v1.36","doi-asserted-by":"publisher","DOI":"10.1002\/anie.201308636"},{"key":"2024080604000630000_2024.08.02.606422v1.37","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1013092107"},{"issue":"11","key":"2024080604000630000_2024.08.02.606422v1.38","first-page":"e01579","article-title":"Active-Site Conformational Fluctuations Promote the Enzymatic Activity of NDM-1","volume":"62","year":"2018","journal-title":"Antimicrob Agents Chemother"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1101\/2024.08.02.606422","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T06:48:35Z","timestamp":1768459715000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.1101\/2024.08.02.606422"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,8,3]]},"references-count":38,"URL":"https:\/\/doi.org\/10.1101\/2024.08.02.606422","relation":{"is-preprint-of":[{"id-type":"doi","id":"10.1021\/acsinfecdis.5c00179","asserted-by":"subject"}]},"subject":[],"published":{"date-parts":[[2024,8,3]]},"subtype":"preprint"}}