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@article{pmid41556618,

title = {Tetravalent antibodies are more potent and efficacious erythropoiesis-stimulating agents than erythropoietin in vivo},

author = {Jarrett J Adams and Levi L Blazer and Jacky Chung and Minoo Karimi and Taylor Davidson and Bailey Blair and Carlos Waddle and Craig A Hokanson and Heather A Bruce and Alexander U Singer and Eva-Maria Tombak and Kiira Gildemann and Nele Tamberg and Kaja Kiiver and Mart Ustav and Yue Ma and Luigi Colombo and Lily Jun-Shen Huang and Stephen W Michnick and Orson W Moe and Sachdev S Sidhu},

doi = {10.1002/pro.70462},

issn = {1469-896X},

year  = {2026},

date = {2026-02-01},

urldate = {2026-02-01},

journal = {Protein Sci},

volume = {35},

number = {2},

pages = {e70462},

abstract = {Recent studies have shown that tetravalent antibodies are potent and efficacious agonists of the erythropoietin (EPO) receptor (EPOR) both in vitro and in vivo. To identify antibody-based erythropoiesis-stimulating agents (ESAs) with therapeutic potential, we evaluated various tetravalent antibody formats for EPOR agonism and key biophysical properties necessary for biologic drug development. We identified two distinct tetravalent antibody formats that strongly stimulated the growth of UT7/Epo cells, which rely on EPOR signaling for proliferation. Moreover, one of these formats exhibited ideal biophysical characteristics for drug development. This format consisted of a diabody (Db) and two antigen-binding fragment (Fab) arms fused to the N- and C-termini of an Fc domain, respectively, to form a tetravalent Db-Fc-Fab (EPRA-0322). In a mouse model expressing the human EPOR, EPRA-0322 induced erythropoiesis with greater potency, efficacy, and duration than darbepoetin, a hyperglycosylated EPO currently used in clinical practice. These findings highlight tetravalent antibodies, and the Db-Fc-Fab format in particular, as promising next-generation ESAs suitable for large-scale production and clinical use.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid41542531,

title = {Cellular consequences, citrullination substrates, and antigenicity resulting from wild-type and targeted PAD4 on cell surfaces},

author = {Sophie Kong and Trenton M Peters-Clarke and Corleone S Delaveris and Paul Phojanakong and Veronica Steri and James A Wells},

doi = {10.64898/2026.01.05.696859},

issn = {2692-8205},

year  = {2026},

date = {2026-01-01},

urldate = {2026-01-01},

journal = {bioRxiv},

abstract = {Protein arginine deiminase-4 (PAD4) catalyzes hydrolysis of arginine to citrulline in proteins that promotes widespread changes in cellular phenotypes through transcriptional regulation that can induce innate immunity and promote cancer. Overexpression and hyperactivity of PAD4 leads to a form of cell death called NETosis that releases PAD4 to the extracellular space. In excess, release of PAD4 is believed to be a major cause of various autoimmune diseases through the generation of anti-citrulline protein antibodies (ACPAs). Little is known about the specific protein substrates that become citrullinated and lead to autoimmunity, but there is growing evidence that PAD4 can be localized to the cell surface in response to inflammation. Here, we further characterize the cellular consequences for exogenous treatment with PAD4 showing that it induces morphological changes that increase cell migration, a hallmark of cancer. We then devised a more simplified and robust proteomics approach to identify PAD4 substrates. We identified some 1000 endogenously citrullinated peptides from 500 proteins, and 3000 citrullinated peptides from 1300 proteins upon exogenous addition of PAD4 both inside and outside of cells. This extracellular set can be further augmented by targeting PAD4 to a cancer target, HER2, using a binding protein conjugate. Finally, we studied how citrullinated cells can induce a robust humoral response in a syngeneic vaccine model to produce ACPAs. We believe these studies further our understanding of cell phenotypic consequences of extracellular PAD4 and new PAD4 substrates both inside and outside of cells that are potential neoepitopes for generation of ACPAs.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid41481455,

title = {E2 variants for probing E3 ubiquitin ligase activities},

author = {Jiale Du and Gisele A Andree and Daniel Horn-Ghetko and Luca Stier and Jaspal Singh and Sebastian Kostrhon and Leo Kiss and Matthias Mann and Sachdev S Sidhu and Brenda A Schulman},

doi = {10.1073/pnas.2524899122},

issn = {1091-6490},

year  = {2026},

date = {2026-01-01},

urldate = {2026-01-01},

journal = {Proc Natl Acad Sci U S A},

volume = {123},

number = {1},

pages = {e2524899122},

abstract = {E3 ligases partner with E2 enzymes to regulate vast eukaryotic biology. The hierarchical nature of these pairings, with >600 E3s and ~40 E2s in humans, necessitates that E2s cofunction with numerous different E3s. Here, focusing on E3s in the RING-between-RING (RBR) family and their partner UBE2L3 and UBE2D-family E2s, we report an approach to interrogate selected pathways. We screened phage-displayed libraries of structure-based E2 variants (E2Vs) to discover enzymes with enhanced affinity and specificity toward half of all RBR E3 ligases (ARIH1, ARIH2, ANKIB1, CUL9, HOIL1, HOIP, and RNF14). Collectively, these E2Vs allowed distinguishing actions of different cofunctioning E3s, obtaining high-resolution cryogenic Electron Microscopy (cryo-EM) structures of an RBR E3 in the context of a substrate-bound multiprotein complex, and profiling an endogenous RBR E3 response to an extracellular stimulus. Overall, we anticipate that E2V technology will be a generalizable tool to enable in-depth mechanistic and structural analysis of E3 ligase functions, and mapping their activity states and protein partners in cellular signaling cascades.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}