{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,2]],"date-time":"2026-03-02T13:17:59Z","timestamp":1772457479420,"version":"3.50.1"},"reference-count":34,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2026,2,28]],"date-time":"2026-02-28T00:00:00Z","timestamp":1772236800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Heart, Lung, and Blood Institute, National Institutes of Health","award":["R41HL167303"],"award-info":[{"award-number":["R41HL167303"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Thrombosis remains a leading cause of cardiovascular morbidity and mortality. During thrombosis, activated platelets and endothelial cells expose phosphatidylserine (PS) on their outer membranes, creating a surface that accelerates clot formation. Current antithrombotic therapies, such as heparin and warfarin, carry significant bleeding risks, highlighting the need for safer alternatives. In response, we developed a PS-targeting liposomal formulation composed of Zn-dipicolylamine (DPA)-cyanine-3[22,22] and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (molar ratio 3:97). This DPA-harboring liposome (DPAL) binds selectively to PS-rich surfaces such as activated platelets and has demonstrated efficacy in reducing thrombosis in mouse models, with minimal bleeding. In the present study, we examined the interaction of DPAL with albumin, the most abundant plasma protein and a key transporter in the bloodstream, to assess the potential for harmful protein aggregation or structural disruption. Using dynamic light scattering and intrinsic protein fluorescence, we found that, unlike warfarin and heparin, DPAL does not induce any large protein aggregates or cause significant conformational changes near the tryptophan residue when mixed with human serum albumin, suggesting a favorable interaction profile. In addition, we used transwell permeability assays and CyQUANT cell proliferation assays to assess the cytotoxicity of DPAL in cultured human umbilical vein endothelial cells (HUVECs). Our results showed that DPAL does not compromise endothelial barrier integrity in HUVEC monolayers nor the cells\u2019 viability. Our current and previous findings together suggest that DPAL could offer a promising approach to modulate harmful coagulation pathways and provide a new targeted therapeutic strategy for managing thrombotic disorders.<\/jats:p>","DOI":"10.3390\/ijms27052299","type":"journal-article","created":{"date-parts":[[2026,3,2]],"date-time":"2026-03-02T12:39:56Z","timestamp":1772455196000},"page":"2299","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["In Vitro Studies of the Effects of Antithrombotic Zn-Dipicolylamine-Harboring Liposomes (DPALs) on Serum Albumin and Human Umbilical Vein Endothelial Cells"],"prefix":"10.3390","volume":"27","author":[{"ORCID":"https:\/\/orcid.org\/0009-0004-0032-6227","authenticated-orcid":false,"given":"Michelle","family":"Tanujaya","sequence":"first","affiliation":[{"name":"Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA 19140, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Gianna","family":"Cai","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA 19140, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jia","family":"Patel","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA 19140, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Zana","family":"Moldavsky","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA 19140, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yumna","family":"Ejaz","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA 19140, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Malia Mahazabin","family":"Ahmed","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA 19140, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"SangSang","family":"Duong","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA 19140, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0525-6008","authenticated-orcid":false,"given":"Lawrence E.","family":"Goldfinger","sequence":"additional","affiliation":[{"name":"Cardeza Foundation for Hematologic Research, Department of Medicine, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5440-7576","authenticated-orcid":false,"given":"Koon Y.","family":"Pak","sequence":"additional","affiliation":[{"name":"Molecular Targeting Technologies Inc., West Chester, PA 19380, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Brian D.","family":"Gray","sequence":"additional","affiliation":[{"name":"Molecular Targeting Technologies Inc., West Chester, PA 19380, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Parkson Lee-Gau","family":"Chong","sequence":"additional","affiliation":[{"name":"Department of Medical Genetics and Molecular Biochemistry, The Lewis Katz School of Medicine at Temple University, Philadelphia, PA 19140, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2026,2,28]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1016\/j.omtn.2017.10.019","article-title":"A Novel Therapeutic Strategy for Cancer Using Phosphatidylserine Targeting Stearylamine-Bearing Cationic Liposomes","volume":"10","author":"De","year":"2018","journal-title":"Mol. Ther. Nucleic Acids"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"100897","DOI":"10.1016\/j.tranon.2020.100897","article-title":"BPRDP056, a Novel Small Molecule Drug Conjugate Specifically Targeting Phosphatidylserine for Cancer Therapy","volume":"14","author":"Chen","year":"2021","journal-title":"Transl. Oncol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1479","DOI":"10.1016\/S0360-3016(02)03928-7","article-title":"Phosphatidylserine Is a Marker of Tumor Vasculature and a Potential Target for Cancer Imaging and Therapy","volume":"54","author":"Ran","year":"2002","journal-title":"Int. J. Radiat. Oncol. Biol. Phys."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"50","DOI":"10.1186\/s13075-015-0565-x","article-title":"Non-Invasive in vivo Imaging of Arthritis in a Collagen-Induced Murine Model with Phosphatidylserine-Binding Near-Infrared (NIR) Dye","volume":"17","author":"Chan","year":"2015","journal-title":"Arthritis Res. Ther."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"5645","DOI":"10.1039\/C4OB00924J","article-title":"Selective Recognition of Anionic Cell Membranes Using Targeted Liposomes Coated with Zinc(II)-bis(dipicolylamine) Affinity Units","volume":"12","author":"Turkyilmaz","year":"2014","journal-title":"Org. Biomol. Chem."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"147","DOI":"10.1021\/acs.molpharmaceut.6b00760","article-title":"Liposomes Containing Lipid-Soluble Zn(II)-bis-dipicolylamine Derivatives Show Potential to be Targeted to Phosphatidylserine on the Surface of Cancer Cells","volume":"14","author":"Ayesa","year":"2017","journal-title":"Mol. Pharm."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"108","DOI":"10.1016\/j.jtha.2024.10.007","article-title":"Phosphatidylserine-blocking Nanoparticles Inhibit Thrombosis without Increased Bleeding in Mice","volume":"23","author":"Wurtzel","year":"2025","journal-title":"J. Thromb. Haemost."},{"key":"ref_8","unstructured":"Chong, P.L.-G., Goldfinger, L., Gray, B.D., and Pak, C. (2021). Anti-thrombotic Agents and Methods of Use Thereof. (11,090,309), U.S. Patent."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1942","DOI":"10.1007\/s10953-018-0830-0","article-title":"Mathematical Methods in the Calculation of the Zeta Potential of BSA","volume":"47","author":"Bukackova","year":"2018","journal-title":"J. Solut. Chem."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"732","DOI":"10.1016\/j.ajpath.2019.07.021","article-title":"Nanomechanics of the Endothelial Glycocalyx: From Structure to Function","volume":"190","author":"Cosgun","year":"2020","journal-title":"Am. J. Pathol."},{"key":"ref_11","unstructured":"Renkin, E.M., and Michel, C.C. (1984). The Cardiovascular System, Microcirculation. The Interstitium and Microvascular Exchange Handbook of Physiology, American Physiological Society."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"4033","DOI":"10.1016\/j.febslet.2015.11.004","article-title":"New Insights into in vitro Amyloidogenic Properties of Human Serum Albumin Suggest Considerations for Therapeutic Precautions","volume":"589","author":"Sharma","year":"2015","journal-title":"FEBS Lett."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"2353","DOI":"10.1016\/j.bpj.2008.12.3901","article-title":"Existence of Different Structural Intermediates on the Fibrillation Pathway of Human Serum Albumin","volume":"96","author":"Juarez","year":"2009","journal-title":"Biophys. J."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1016\/j.aca.2010.07.016","article-title":"Thermal Aggregation of Bovine Serum Albumin Studied by Asymmetrical Flow Field-Flow Fractionation","volume":"675","author":"Yohannes","year":"2010","journal-title":"Anal. Chim. Acta"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1007\/BF00211402","article-title":"The Subdomain Structure of Human Serum Albumin in Solution under Different pH Conditions Studied by Small Angle X-ray Scattering","volume":"24","author":"Olivieril","year":"1995","journal-title":"Eur. Biophys. J."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"14894","DOI":"10.1021\/la5034959","article-title":"Concentration-Dependent Reversible Self-Oligomerization of Serum Albumins through Intermolecular \u03b2-Sheet Formation","volume":"30","author":"Bhattacharya","year":"2014","journal-title":"Langmuir"},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Radomska, K., and Wolszczak, M. (2022). Spontaneous and Ionizing Radiation-Induced Aggregation of Human Serum Albumin: Dityrosine as a Fluorescent Probe. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23158090"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"20733","DOI":"10.1021\/jp064861r","article-title":"Amyloid Fibril Formation and Other Aggregate Species Formed by Human Serum Albumin Association","volume":"110","author":"Taboada","year":"2006","journal-title":"J. Phys. Chem. B"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"717","DOI":"10.1007\/s00249-007-0196-5","article-title":"Thermal Aggregation of Bovine Serum Albumin at Different pH: Comparison with Human Serum Albumin","volume":"36","author":"Vetri","year":"2007","journal-title":"Eur. Biophys. J."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Maci\u0105\u017cek-Jurczyk, M., Janas, K., Po\u017cycka, J., Szkudlarek, A., Rog\u00f3\u017c, W., Owczarzy, A., and Kulig, K. (2020). Human Serum Albumin Aggregation\/Fibrillation and its Abilities to Drugs Binding. Molecules, 25.","DOI":"10.3390\/molecules25030618"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"106678","DOI":"10.1016\/j.bpc.2021.106678","article-title":"Amyloid-like Aggregation of Bovine Serum Albumin at Physiological Temperature Induced by Cross-Seeding Effect of HEWL Amyloid Aggregates","volume":"278","author":"Nirwal","year":"2021","journal-title":"Biophys. Chem."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"153","DOI":"10.1016\/S0065-3233(08)60640-3","article-title":"Structure of Serum Albumin","volume":"45","author":"Carter","year":"1994","journal-title":"Adv. Protein Chem."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1679","DOI":"10.1039\/D0RA09580J","article-title":"Binding Interactions and FRET between Bovine Serum Albumin and Various Phenothiazine-\/Anthracene-Based Dyes: A Structure\u2013Property Relationship","volume":"11","author":"Bhuin","year":"2021","journal-title":"RSC Adv."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"107","DOI":"10.1016\/j.jpba.2009.03.015","article-title":"Fluorescence Spectral Resolution of Tryptophan Residues in Bovine and Human Serum Albumins","volume":"50","author":"Tayeh","year":"2009","journal-title":"J. Pharm. Biomed. Anal."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"3830","DOI":"10.1021\/jp980486u","article-title":"Complex Formation between Bovine Serum Albumin and Strong Polyelectrolytes: Effect of Polymer Charge Density","volume":"102","author":"Mattison","year":"1998","journal-title":"J. Phys. Chem. B"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"143","DOI":"10.1021\/acs.nanolett.6b03686","article-title":"Fluorescence Self-Quenching from Reporter Dyes Informs on the Structural Properties of Amyloid Clusters Formed in vitro and in Cells","volume":"17","author":"Chen","year":"2017","journal-title":"Nano Lett."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"3032","DOI":"10.1016\/S0021-9258(19)85847-7","article-title":"The Effect of Albumin Conformation on the Binding of Warfarin to Human Serum Albumin. The Dependence of the Binding of Warfarin to Human Serum Albumin on the Hydrogen, Calcium, and Chloride Ion Concentrations as Studie by Circular Dichroism, Fluorescence, and Equilibrium Dialysis","volume":"255","author":"Wilting","year":"1980","journal-title":"J. Biol. Chem."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"129","DOI":"10.1111\/j.1440-1681.1975.tb01826.x","article-title":"Spectroscopic Techniques in the Study of Protein Binding. A Fluorescence Technique for the Evaluation of the Albumin Binding and Displacement of Warfarin and Warfarin-Alcohol","volume":"2","author":"Sudlow","year":"1975","journal-title":"Clin. Exp. Pharmacol. Physiol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"225","DOI":"10.5530\/ijper.49.3.8","article-title":"Warfarin Binding to Native and Structurally-Altered Human Serum Albumins","volume":"49","author":"Sakhaei","year":"2015","journal-title":"Indian J. Pharm. Educ. Res."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"834501","DOI":"10.1155\/2014\/834501","article-title":"Interaction of Warfarin with Human Serum Albumin and Effect of Ferulic Acid on the Binding","volume":"2014","author":"Li","year":"2014","journal-title":"J. Spectrosc."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"122797","DOI":"10.1016\/j.carbpol.2024.122797","article-title":"Characterization of the Mechanisms Governing Heparin-Bovine Serum Albumin Complex Interactions and Associated Anticoagulant Activity","volume":"348","author":"Zhang","year":"2025","journal-title":"Carbohydr. Polym."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"2328","DOI":"10.1021\/acsomega.0c05638","article-title":"Heparin Accelerates the Protein Aggregation via the Downhill Polymerization Mechanism: Multi-spectroscopic Studies to Delineate the Implications on Proteinopathies","volume":"6","author":"Ahanger","year":"2021","journal-title":"ACS Omega"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"87","DOI":"10.1023\/A:1006878421990","article-title":"The Measurement of Free Fatty Acid Concentration with the Fluorescent Probe ADIFAB: A Practical Guide for the Use of the ADIFAB Probe","volume":"192","author":"Richieri","year":"1999","journal-title":"Mol. Cell. Biochem."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"23495","DOI":"10.1016\/S0021-9258(18)35866-6","article-title":"A Fluorescently Labeled Intestinal Fatty Acid Binding Protein. Interactions with Fatty Acids and Its Use in Monitoring Free Fatty Acids","volume":"267","author":"Richieri","year":"1992","journal-title":"J. Biol. Chem."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/27\/5\/2299\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,3,2]],"date-time":"2026-03-02T12:43:25Z","timestamp":1772455405000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/27\/5\/2299"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,2,28]]},"references-count":34,"journal-issue":{"issue":"5","published-online":{"date-parts":[[2026,3]]}},"alternative-id":["ijms27052299"],"URL":"https:\/\/doi.org\/10.3390\/ijms27052299","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,2,28]]}}}