{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,10,23]],"date-time":"2026-10-23T21:18:40Z","timestamp":1792790320859,"version":"4.5.0"},"posted":{"date-parts":[[2026,10,2]]},"group-title":"Immunology","reference-count":48,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,10,2]],"date-time":"2026-10-02T00:00:00Z","timestamp":1790899200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"award":["P30AI045008"],"award-info":[{"award-number":["P30AI045008"]}],"id":[{"id":"https:\/\/ror.org\/001d55x84","id-type":"ROR","asserted-by":"publisher"}]},{"award":["P30CA016520"],"award-info":[{"award-number":["P30CA016520"]}],"id":[{"id":"https:\/\/ror.org\/001d55x84","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R33CA281702"],"award-info":[{"award-number":["R33CA281702"]}],"id":[{"id":"https:\/\/ror.org\/001d55x84","id-type":"ROR","asserted-by":"publisher"}]},{"award":["K99CA282989"],"award-info":[{"award-number":["K99CA282989"]}],"id":[{"id":"https:\/\/ror.org\/001d55x84","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R00CA282989"],"award-info":[{"award-number":["R00CA282989"]}],"id":[{"id":"https:\/\/ror.org\/001d55x84","id-type":"ROR","asserted-by":"publisher"}]},{"name":"United States Department of Defense","award":["W81XWH-21-1-0514"],"award-info":[{"award-number":["W81XWH-21-1-0514"]}],"id":[{"id":"https:\/\/ror.org\/0447fe631","id-type":"ROR","asserted-by":"crossref"}]},{"name":"United States Department of Defense","award":["HT94252310472"],"award-info":[{"award-number":["HT94252310472"]}],"id":[{"id":"https:\/\/ror.org\/0447fe631","id-type":"ROR","asserted-by":"crossref"}]},{"name":"Cancer Research Institute","award":["CRI4964"],"award-info":[{"award-number":["CRI4964"]}],"id":[{"id":"https:\/\/ror.org\/02f3xk561","id-type":"ROR","asserted-by":"crossref"}]},{"id":[{"id":"https:\/\/ror.org\/01pkpj777","id-type":"ROR","asserted-by":"publisher"}]},{"id":[{"id":"https:\/\/ror.org\/02zhe7k92","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,10,2]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Despite the success of immune checkpoint blockade in cancer therapy, overcoming the immunosuppressive tumor microenvironment remains a major barrier to durable responses. Here, we introduce cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4)-targeting chimeras (cTACs), a class of synthetic molecules that harness CTLA-4-mediated trans-endocytosis (TE) for targeted protein degradation across cellular boundaries. The lead candidate, cTAC4.0, is composed of the extracellular domain of CD80 fused to an anti-programmed death-ligand 1 (PD-L1) fragment via a human immunoglobulin G4 (IgG4) heavy chain. Unlike existing protein degraders that function within cancer cells via house-keeping pathways, cTAC4.0 operates\n                  <jats:italic>in trans<\/jats:italic>\n                  between immune and tumor cells. It redirects the natural CD80-CTLA-4 internalization to capture and degrade PD-L1 from tumor cells into T cells, while simultaneously converting PD-L1-mediated suppression into CD28-driven co-stimulation, depending on the relative availability of CTLA-4 or CD28. Mechanistically, we demonstrate that cTAC4.0 promotes CTLA-4-dependent internalization and lysosomal degradation of both membrane-bound and soluble PD-L1, reduces surface PD-L1 levels on target cells, and activates T cells in a CD28-dependent manner. Functionally, cTAC4.0 enhances the cytotoxicity of both bispecific T cell engagers and T cell receptor (TCR)-engineered T cells\n                  <jats:italic>in vitro,<\/jats:italic>\n                  and when combined with these engineered T cells, it suppresses tumor growth\n                  <jats:italic>in vivo<\/jats:italic>\n                  in a glioblastoma xenograft model. By coupling immune checkpoint degradation with T cell activation in an immune synapse-enriched manner, cTACs establish a paradigm for intercellular protein degradation systems that simultaneously deplete suppressive ligands and enhance T cell function to overcome resistance in cancer immunotherapy.\n                <\/jats:p>","DOI":"10.64898\/2026.09.29.755403","type":"posted-content","created":{"date-parts":[[2026,10,2]],"date-time":"2026-10-02T16:15:13Z","timestamp":1790957713000},"source":"Crossref","is-referenced-by-count":0,"title":["Engineered CTLA-4 targeting chimeras for targeted protein degradation and immune synapse reprogramming"],"prefix":"10.64898","author":[{"given":"Ping","family":"Ren","sequence":"first","affiliation":[{"name":"Department of Genetics, Yale University School of Medicine, New Haven, CT, USA"},{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shao-Yu","family":"Fang","sequence":"additional","affiliation":[{"name":"Department of Genetics, Yale University School of Medicine, New Haven, CT, USA"},{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shushu","family":"Zhao","sequence":"additional","affiliation":[{"name":"Vaccine and Immunotherapy Center, The Wistar Institute, Philadelphia, PA, USA"},{"name":"Ellen and Ronald Caplan Cancer Center, The Wistar Institute, Philadelphia, PA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hanbing","family":"Cao","sequence":"additional","affiliation":[{"name":"Department of Genetics, Yale University School of Medicine, New Haven, CT, USA"},{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kaiyuan","family":"Tang","sequence":"additional","affiliation":[{"name":"Department of Genetics, Yale University School of Medicine, New Haven, CT, USA"},{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"},{"name":"Molecular Cell Biology, Genetics, and Development Program, Yale University, New Haven, CT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xingbo","family":"Shang","sequence":"additional","affiliation":[{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"},{"name":"Department of Biomedical Engineering, Yale University, New Haven, CT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cole W.","family":"Christopher","sequence":"additional","affiliation":[{"name":"Vaccine and Immunotherapy Center, The Wistar Institute, Philadelphia, PA, USA"},{"name":"Ellen and Ronald Caplan Cancer Center, The Wistar Institute, Philadelphia, PA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yunfei","family":"Jiao","sequence":"additional","affiliation":[{"name":"Vaccine and Immunotherapy Center, The Wistar Institute, Philadelphia, PA, USA"},{"name":"Ellen and Ronald Caplan Cancer Center, The Wistar Institute, Philadelphia, PA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Medha","family":"Majety","sequence":"additional","affiliation":[{"name":"Department of Genetics, Yale University School of Medicine, New Haven, CT, USA"},{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"},{"name":"Yale College, New Haven, CT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andre","family":"Levchenko","sequence":"additional","affiliation":[{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"},{"name":"Department of Biomedical Engineering, Yale University, New Haven, CT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2232-8512","authenticated-orcid":false,"given":"David B.","family":"Weiner","sequence":"additional","affiliation":[{"name":"Vaccine and Immunotherapy Center, The Wistar Institute, Philadelphia, PA, USA"},{"name":"Ellen and Ronald Caplan Cancer Center, The Wistar Institute, Philadelphia, PA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3819-5005","authenticated-orcid":false,"given":"Sidi","family":"Chen","sequence":"additional","affiliation":[{"name":"Department of Genetics, Yale University School of Medicine, New Haven, CT, USA"},{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"},{"name":"Molecular Cell Biology, Genetics, and Development Program, Yale University, New Haven, CT, USA"},{"name":"Department of Immunobiology, Yale University, New Haven, CT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2304-4191","authenticated-orcid":false,"given":"Xiaoyu","family":"Zhou","sequence":"additional","affiliation":[{"name":"Department of Genetics, Yale University School of Medicine, New Haven, CT, USA"},{"name":"System Biology Institute, Yale University, West Haven, CT, USA"},{"name":"Center for Cancer Systems Biology, Yale University, West Haven, CT, USA"},{"name":"Vaccine and Immunotherapy Center, The Wistar Institute, Philadelphia, PA, USA"},{"name":"Ellen and Ronald Caplan Cancer Center, The Wistar Institute, Philadelphia, PA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026100704201215000_2026.09.29.755403v1.1","doi-asserted-by":"publisher","DOI":"10.1056\/NEJMoa1200690"},{"key":"2026100704201215000_2026.09.29.755403v1.2","doi-asserted-by":"publisher","DOI":"10.1056\/NEJMoa1200694"},{"key":"2026100704201215000_2026.09.29.755403v1.3","doi-asserted-by":"publisher","DOI":"10.1056\/NEJMoa1003466"},{"key":"2026100704201215000_2026.09.29.755403v1.4","doi-asserted-by":"publisher","DOI":"10.1056\/NEJMoa1104621"},{"key":"2026100704201215000_2026.09.29.755403v1.5","doi-asserted-by":"publisher","DOI":"10.1038\/nature14011"},{"key":"2026100704201215000_2026.09.29.755403v1.6","doi-asserted-by":"publisher","DOI":"10.1056\/NEJMoa1302369"},{"key":"2026100704201215000_2026.09.29.755403v1.7","doi-asserted-by":"publisher","DOI":"10.1158\/1078-0432.CCR-16-0698"},{"key":"2026100704201215000_2026.09.29.755403v1.8","doi-asserted-by":"publisher","DOI":"10.1038\/nature13954"},{"key":"2026100704201215000_2026.09.29.755403v1.9","doi-asserted-by":"crossref","unstructured":"Wang, Q. et al. Soluble Immune Checkpoint-Related Proteins in Blood Are Associated With Invasion and Progression in Non-Small Cell Lung Cancer. Front Immunol 13, 887916 (2022).","DOI":"10.3389\/fimmu.2022.887916"},{"key":"2026100704201215000_2026.09.29.755403v1.10","doi-asserted-by":"crossref","unstructured":"Oh, S.Y. et al. Soluble PD-L1 is a predictive and prognostic biomarker in advanced cancer patients who receive immune checkpoint blockade treatment. Sci Rep 11, 19712 (2021).","DOI":"10.1038\/s41598-021-99311-y"},{"key":"2026100704201215000_2026.09.29.755403v1.11","doi-asserted-by":"publisher","DOI":"10.1038\/nchembio.1858"},{"key":"2026100704201215000_2026.09.29.755403v1.12","doi-asserted-by":"crossref","unstructured":"Zhang, D. et al. Transferrin receptor targeting chimeras for membrane protein degradation. Nature (2024).","DOI":"10.1101\/2023.08.10.552782"},{"key":"2026100704201215000_2026.09.29.755403v1.13","doi-asserted-by":"crossref","unstructured":"Huang, B. et al. Designed endocytosis-inducing proteins degrade targets and amplify signals. Nature (2024).","DOI":"10.1038\/s41586-024-07948-2"},{"key":"2026100704201215000_2026.09.29.755403v1.14","doi-asserted-by":"publisher","DOI":"10.1126\/science.1202947"},{"key":"2026100704201215000_2026.09.29.755403v1.15","doi-asserted-by":"publisher","DOI":"10.1038\/nri3108"},{"key":"2026100704201215000_2026.09.29.755403v1.16","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.192461099"},{"key":"2026100704201215000_2026.09.29.755403v1.17","doi-asserted-by":"crossref","unstructured":"Lee, J.Y. et al. Structural basis of checkpoint blockade by monoclonal antibodies in cancer immunotherapy. Nat Commun 7, 13354 (2016).","DOI":"10.1038\/ncomms13354"},{"key":"2026100704201215000_2026.09.29.755403v1.18","doi-asserted-by":"crossref","first-page":"2435","DOI":"10.1111\/cas.13666","article-title":"High-affinity PD-1 molecules deliver improved interaction with PD-L1 and PD-L2","volume":"109","year":"2018","journal-title":"Cancer Sci"},{"key":"2026100704201215000_2026.09.29.755403v1.19","doi-asserted-by":"publisher","DOI":"10.1158\/2326-6066.CIR-15-0059"},{"key":"2026100704201215000_2026.09.29.755403v1.20","doi-asserted-by":"publisher","DOI":"10.1038\/s41590-022-01289-w"},{"key":"2026100704201215000_2026.09.29.755403v1.21","doi-asserted-by":"publisher","DOI":"10.1126\/science.aav7062"},{"key":"2026100704201215000_2026.09.29.755403v1.22","doi-asserted-by":"crossref","first-page":"1059","DOI":"10.1016\/j.immuni.2019.11.003","article-title":"PD-L1:CD80 Cis-Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways","volume":"51","year":"2019","journal-title":"Immunity"},{"key":"2026100704201215000_2026.09.29.755403v1.23","doi-asserted-by":"publisher","DOI":"10.1080\/15548627.2015.1066957"},{"key":"2026100704201215000_2026.09.29.755403v1.24","doi-asserted-by":"publisher","DOI":"10.1128\/MCB.25.21.9543-9553.2005"},{"key":"2026100704201215000_2026.09.29.755403v1.25","doi-asserted-by":"publisher","DOI":"10.1016\/j.it.2014.12.001"},{"key":"2026100704201215000_2026.09.29.755403v1.26","doi-asserted-by":"publisher","DOI":"10.1158\/2326-6066.CIR-16-0329"},{"key":"2026100704201215000_2026.09.29.755403v1.27","doi-asserted-by":"crossref","first-page":"421","DOI":"10.1007\/s00262-018-2282-1","article-title":"A secreted PD-L1 splice variant that covalently dimerizes and mediates immunosuppression","volume":"68","year":"2019","journal-title":"Cancer Immunol Immunother"},{"key":"2026100704201215000_2026.09.29.755403v1.28","doi-asserted-by":"publisher","DOI":"10.1038\/s41591-021-01621-1"},{"key":"2026100704201215000_2026.09.29.755403v1.29","doi-asserted-by":"publisher","DOI":"10.1038\/icb.2014.93"},{"key":"2026100704201215000_2026.09.29.755403v1.30","doi-asserted-by":"publisher","DOI":"10.1182\/blood-2013-04-495531"},{"key":"2026100704201215000_2026.09.29.755403v1.31","doi-asserted-by":"crossref","unstructured":"Shen, Y. et al. Cancer cell-intrinsic resistance to BiTE therapy is mediated by loss of CD58 costimulation and modulation of the extrinsic apoptotic pathway. J Immunother Cancer 10 (2022).","DOI":"10.1136\/jitc-2021-004348"},{"key":"2026100704201215000_2026.09.29.755403v1.32","doi-asserted-by":"crossref","unstructured":"Zhao, S. et al. Targeting IL13Ralpha2 in melanoma with a bispecific T-cell engager: expression profiling and preclinical evaluation. J Immunother Cancer 13 (2025).","DOI":"10.1136\/jitc-2024-011073"},{"key":"2026100704201215000_2026.09.29.755403v1.33","doi-asserted-by":"publisher","DOI":"10.1016\/j.coi.2015.10.009"},{"key":"2026100704201215000_2026.09.29.755403v1.34","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2017.01.017"},{"key":"2026100704201215000_2026.09.29.755403v1.35","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-022-05235-6"},{"key":"2026100704201215000_2026.09.29.755403v1.36","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-020-2545-9"},{"key":"2026100704201215000_2026.09.29.755403v1.37","doi-asserted-by":"crossref","unstructured":"Yang, R. et al. Galectin-9 interacts with PD-1 and TIM-3 to regulate T cell death and is a target for cancer immunotherapy. Nat Commun 12, 832 (2021).","DOI":"10.1038\/s41467-021-21099-2"},{"key":"2026100704201215000_2026.09.29.755403v1.38","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1121623109"},{"key":"2026100704201215000_2026.09.29.755403v1.39","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-025-09212-7"},{"key":"2026100704201215000_2026.09.29.755403v1.40","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2016.09.011"},{"key":"2026100704201215000_2026.09.29.755403v1.41","doi-asserted-by":"crossref","unstructured":"Ren, P. et al. Function and Cryo-EM structures of broadly potent bispecific antibodies against multiple SARS-CoV-2 Omicron sublineages. Signal Transduct Target Ther 8, 281 (2023).","DOI":"10.1038\/s41392-023-01509-1"},{"key":"2026100704201215000_2026.09.29.755403v1.42","doi-asserted-by":"crossref","unstructured":"Handlogten, M.W. et al. Prevention of Fab-arm exchange and antibody reduction via stabilization of the IgG4 hinge region. MAbs 12, 1779974 (2020).","DOI":"10.1080\/19420862.2020.1779974"},{"key":"2026100704201215000_2026.09.29.755403v1.43","doi-asserted-by":"publisher","DOI":"10.1038\/s41590-023-01571-5"},{"key":"2026100704201215000_2026.09.29.755403v1.44","doi-asserted-by":"publisher","DOI":"10.1016\/j.jim.2016.01.007"},{"key":"2026100704201215000_2026.09.29.755403v1.45","doi-asserted-by":"publisher","DOI":"10.1038\/s41587-019-0246-4"},{"key":"2026100704201215000_2026.09.29.755403v1.46","doi-asserted-by":"publisher","DOI":"10.1002\/cpns.103"},{"key":"2026100704201215000_2026.09.29.755403v1.47","first-page":"86","article-title":"Trispecific antibodies enhance the therapeutic efficacy of tumor-directed T cells through T cell receptor co-stimulation","volume":"1","year":"2020","journal-title":"Nat Cancer"},{"key":"2026100704201215000_2026.09.29.755403v1.48","doi-asserted-by":"crossref","first-page":"85","DOI":"10.1016\/j.chembiol.2022.12.005","article-title":"RAMIHM generates fully human monoclonal antibodies by rapid mRNA immunization of humanized mice and BCR-seq","volume":"30","year":"2023","journal-title":"Cell Chem Biol"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.64898\/2026.09.29.755403","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,10,7]],"date-time":"2026-10-07T11:20:25Z","timestamp":1791372025000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.64898\/2026.09.29.755403"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,10,2]]},"references-count":48,"URL":"https:\/\/doi.org\/10.64898\/2026.09.29.755403","relation":{},"subject":[],"published":{"date-parts":[[2026,10,2]]},"subtype":"preprint"}}