Evaluation of antiviral drugs’ central nervous system distribution using in vitro blood-brain and blood- CSF barrier models

Evaluation of antiviral drugs’ central nervous system distribution using in vitro blood-brain and blood- CSF barrier models | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Evaluation of antiviral drugs’ central nervous system distribution using in vitro blood-brain and blood- CSF barrier models Johid R. Malik, Ukamaka O. Modebelu, Courtney V. Fletcher, Siddappa N. Byrareddy, and 8 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8788392/v1 This work is licensed under a CC BY 4.0 License Status: Under Revision Version 1 posted 11 You are reading this latest preprint version Abstract Background: The blood–brain barrier (BBB) and blood–cerebrospinal fluid barrier (BCSFB) are dynamic neurovascular interfaces that maintain CNS homeostasis by tightly regulating paracellular and transcellular molecular transport. Intercellular communication between adjacent cells is critical for maintaining barrier function. Also, serum protein binding is a key factor influencing central nervous system (CNS) penetration. Accurate in vitro BBB and BCSFB models that recapitulate these physiological interactions offer valuable alternatives to animal-based systems for screening CNS targeted drugs. Methods: Building upon our published human primary cell–based BBB model, we employed complementary BBB and BCSFB systems to evaluate the permeability of nine antiviral agents (AVs) under low (L) and high (H) serum conditions. Barrier integrity was confirmed by transepithelial electrical resistance (TEER) and FITC-dextran permeability assays. Results: In H serum, AVs like dolutegravir and efavirenz with extensive protein binding demonstrated significantly reduced permeability across both barriers. These in vitro permeability profiles were fairly consistent with reported higher clinical cerebrospinal fluid concentrations for drugs that exhibit low protein binding. Furthermore, astrocytes exposed to dolutegravir and emtricitabine indicated upregulation of ATP-binding cassette transporter multidrug resistance protein (MRP1) and P-glycoprotein, while lacking breast cancer resistant protein and MRP4 transporters, additional factors for low CNS concentration of AVs. Conclusions: Our validated in vitro BBB and BCSFB models reproduce clinically relevant patterns of CNS antiviral penetration. These systems provide reliable, human-relevant platforms for preclinical CNS drug screening and align with current efforts to reduce animal use in biomedical research. in vitro model blood-brain-barrier blood-CSF-barrier central nervous system human primary cells antivirals Full Text Additional Declarations No competing interests reported. Supplementary Files 9ARVsSupplementaryMethods.pdf Rwadatasupplementaryfiguresforwesternblot.pdf Cite Share Download PDF Status: Under Revision Version 1 posted Editorial decision: Revision requested 23 Mar, 2026 Reviews received at journal 17 Mar, 2026 Reviews received at journal 16 Mar, 2026 Reviews received at journal 03 Mar, 2026 Reviewers agreed at journal 25 Feb, 2026 Reviewers agreed at journal 23 Feb, 2026 Reviewers agreed at journal 20 Feb, 2026 Reviewers invited by journal 20 Feb, 2026 Editor assigned by journal 19 Feb, 2026 Submission checks completed at journal 19 Feb, 2026 First submitted to journal 17 Feb, 2026 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8788392","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":595521138,"identity":"1a02f6de-bde2-44fd-b424-bfdea9dec04f","order_by":0,"name":"Johid R. 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Intercellular communication between adjacent cells is critical for maintaining barrier function. Also, serum protein binding is a key factor influencing central nervous system (CNS) penetration. Accurate in vitro BBB and BCSFB models that recapitulate these physiological interactions offer valuable alternatives to animal-based systems for screening CNS targeted drugs.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods:\u003c/strong\u003e Building upon our published human primary cell–based BBB model, we employed complementary BBB and BCSFB systems to evaluate the permeability of nine antiviral agents (AVs) under low (L) and high (H) serum conditions. Barrier integrity was confirmed by transepithelial electrical resistance (TEER) and FITC-dextran permeability assays.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults: \u003c/strong\u003eIn H serum, AVs like dolutegravir and efavirenz with extensive protein binding demonstrated significantly reduced permeability across both barriers. These in vitro permeability profiles were fairly consistent with reported higher clinical cerebrospinal fluid concentrations for drugs that exhibit low protein binding. 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These systems provide reliable, human-relevant platforms for preclinical CNS drug screening and align with current efforts to reduce animal use in biomedical research.\u003c/p\u003e","manuscriptTitle":"Evaluation of antiviral drugs’ central nervous system distribution using in vitro blood-brain and blood- CSF barrier models","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-02-24 17:42:07","doi":"10.21203/rs.3.rs-8788392/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-03-23T04:03:17+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-17T18:17:55+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-16T13:31:46+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-03T09:37:36+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"98487112626531003891370112956795772714","date":"2026-02-25T20:57:57+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"130816479606799037074837762228995565775","date":"2026-02-23T09:34:50+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"271692532746014874914313772654279035280","date":"2026-02-20T11:21:20+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-02-20T05:06:19+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-02-19T16:06:03+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-02-19T14:36:33+00:00","index":"","fulltext":""},{"type":"submitted","content":"Cellular and Molecular Neurobiology","date":"2026-02-17T15:15:56+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"cellular-and-molecular-neurobiology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"cemn","sideBox":"Learn more about [Cellular and Molecular Neurobiology](https://www.springer.com/journal/10571)","snPcode":"10571","submissionUrl":"https://submission.nature.com/new-submission/10571/3","title":"Cellular and Molecular Neurobiology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"c8f4f81a-9fd9-4135-833c-3f342c4bfdbb","owner":[],"postedDate":"February 24th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"in-revision","subjectAreas":[],"tags":[],"updatedAt":"2026-03-23T04:08:57+00:00","versionOfRecord":[],"versionCreatedAt":"2026-02-24 17:42:07","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8788392","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8788392","identity":"rs-8788392","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}