Antibodies最新文献

{"title":"Immune Cell Engagers: Advancing Precision Immunotherapy for Cancer Treatment.","authors":"Hyukmin In, Minkyoung Park, Hyeonsik Lee, Kyung Ho Han","doi":"10.3390/antib14010016","DOIUrl":"10.3390/antib14010016","url":null,"abstract":"<p><p>Immune cell engagers (ICEs) are an emerging class of immunotherapies designed to harness the immune system's anti-tumor potential through precise targeting and activation of immune effector cells. By engaging T cells, natural killer (NK) cells, and phagocytes, ICEs overcome challenges such as immune evasion and MHC downregulation, addressing critical barriers in cancer treatment. T-cell engagers (TCEs), led by bispecific T-cell engagers (BiTEs), dominate the field, with innovations such as half-life-extended BiTEs, trispecific antibodies, and checkpoint inhibitory T-cell engagers driving their application in hematologic and solid malignancies. NK cell engagers (NKCEs) and phagocyte cell engagers (PCEs) are rapidly progressing, drawing on NK cells' innate cytotoxicity and macrophages' phagocytic abilities to target tumors, particularly in immunosuppressive microenvironments. Since the FDA approval of Blinatumomab in 2014, ICEs have transformed the oncology landscape, with nine FDA-approved products and numerous candidates in clinical trials. Despite challenges such as toxicity, resistance, and limited efficacy in solid tumors, ongoing research into advanced platforms and combination therapies highlights the growing potential of ICEs to provide personalized, scalable, and effective cancer treatments. This review investigates the mechanisms, platforms, research trends, and clinical progress of ICEs, emphasizing their pivotal role in advancing precision immunotherapy and their promise as a cornerstone of next-generation cancer therapies.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-02-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843982/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466830","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Assessing the Influence of Selected Permeabilization Methods on Lymphocyte Single-Cell Multi-Omics.","authors":"Shifan Ding, Na Lu, Hassan Abolhassani","doi":"10.3390/antib14010015","DOIUrl":"10.3390/antib14010015","url":null,"abstract":"<p><p>(1) Background: Single-cell multi-omics is a powerful method for the dissection and detection of complicated immunologic functions and synapses. However, most currently available technologies merge datasets of different omics from separate portions of the same sample to generate combined multi-omics. This process is a source of bias, mainly in the field of immunology on cells originating from pluripotent hematopoietic stem cells with high flexibility during maturation. (2) Methods: Although new multi-omics approaches have been developed to use the advantages of cellular and molecular barcoding and next-generation sequencing to solve this issue, one of the main current challenges is intracellular proteomics, which should be combined with other omics data with high importance for immune system studies. We designed this study to evaluate previously recommended minimal permeabilization and fixation methods on the quality and quantity of transcriptomics and proteomics data generated by the BD Rhapsody™ Single-Cell Analysis System. (3) Results: Our findings showed that high-throughput sequencing with advanced quality and read-out is required for the combination of multi-omics outcomes from a permeabilized single cell. Therefore, the HiseqX platform was selected for further analysis. The effect of immune stimulation was observed clearly as the separated clusters of helper and cytotoxic T cells using unsupervised clustering. Importantly, fixation and permeabilization did not affect the general expression profile of unstimulated cells. However, fixation and permeabilization were proved to negatively impact the detection of the whole transcriptome for single-cell assay. Nevertheless, about 60% of the transcriptomic signature of the stimulation was detected. If the measurement of combined surface and intracellular markers is required to be achieved, the modified fixation and permeabilization method is recommended because of a lower transcriptomic loss and more precise proteomic fingerprint detected. (4) Conclusions: The findings of this study support the potential possibility for integrating intracellular proteomics, which needs to be optimized and tested with newly designed oligonucleotide-tagged antibodies targeting intracellular proteins.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-02-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843891/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466803","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Clinical Scaleup of Humanized AnnA1 Antibody Yielded Unexpected High Reticuloendothelial (RES) Uptake in Mice.","authors":"Lu Lucy Xu, Satyendra Kumar Singh, Chelsea Nayback, Abdullah Metebi, Dalen Agnew, Tim Buss, Jan Schnitzer, Kurt R Zinn","doi":"10.3390/antib14010014","DOIUrl":"10.3390/antib14010014","url":null,"abstract":"<p><strong>Background/objectives: </strong>A mouse antibody directed against truncated Annexin A1 showed high tumor retention in pre-clinical cancer models and was approved by the National Cancer Institute Experimental Therapeutics (NExT) program for humanization and large batch cGMP production for toxicology and clinical trials. In this process, a contractor for Leidos accidentally produced a mutated version of humanized AnnA1 (hAnnA1-mut) with a single nucleotide deletion in the terminal Fc coding region that increased the translated size by eight amino acids with random alterations in the final twenty-four amino acids. We investigated the tissue distribution of hAnnA1-mut, hAnnA1, mAnnA1, and isotope-matched human IgG1 under various injection and conjugation conditions with C57BL/6, FVB, and BALB/c nude mice strains.</p><p><strong>Methods: </strong>Biodistribution studies were performed 24 h after injection of Tc-99m-HYNIC radiolabeled antibodies (purity > 98%). Non-reducing gel electrophoresis studies were conducted with IR680 labeled antibodies incubated with various mouse sera.</p><p><strong>Results: </strong>Our results showed that Tc-99m-HYNIC-hAnnA1 had low spleen and liver retention not statistically different from Tc-99m-HYNIC-IgG1 and Tc-99m-HYNIC-mAnnA1, with corresponding higher blood levels; however, Tc-99m-HYNIC-hAnnA1-mut had high levels in the spleen and liver with differences identified among the mouse strains, radiolabeling conditions, and injection routes. Histopathology showed no morphological change in the liver or spleen from any conditions. Gel electrophoresis showed an upward shift of hAnnA1-mut, consistent with the binding of blood serum protein.</p><p><strong>Conclusions: </strong>The changes in the Fc region of hAnnA1-mut led to higher liver and spleen uptake, suggesting the antibody's recognition by the innate immune system (likely complement protein binding) and subsequent clearance. Future clinical translation using hAnnA1 and other antibodies needs to limit protein modifications that could drastically reduce blood clearance.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-02-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843838/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466810","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of SARS-CoV-2 Antibody Response Between Paired Fingerprick (HemaPEN^®) and Venepuncture Collected Samples in Children and Adults.","authors":"Nadia Mazarakis, Zheng Quan Toh, Jill Nguyen, Rachel A Higgins, James Rudge, Belinda Whittle, Nicholas J Woudberg, Justin Devine, Andrew Gooley, Florian Lapierre, Nigel W Crawford, Shidan Tosif, Paul V Licciardi","doi":"10.3390/antib14010013","DOIUrl":"10.3390/antib14010013","url":null,"abstract":"<p><p>Serological surveillance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies is important to monitor population COVID-19 immunity. Dried blood spots (DBS) are a valuable method for serosurveys, particularly in remote settings and in children. We compared the measurement of SARS-CoV-2 spike-specific IgG in paired blood samples collected using standard venepuncture (serum) and the hemaPEN^® microsampling DBS device from children and adults. A total of 83 participants (10 months to 65 years of age), comprising COVID-positive and -negative participants, were recruited. Paired serum and DBS samples were assayed for SARS-CoV-2 receptor-binding domain (RBD) and Spike (S1) antibodies using an established in-house ELISA. RBD and S1 IgG concentrations of paired hemaPEN DBS eluates and serum samples were compared using a non-parametric Wilcoxon matched-pairs signed ranked test. A Pearson's correlation was used for RBD and S1 IgG concentrations and the level of agreement between the hemaPEN DBS eluates and serum samples was assessed by Bland-Altman analysis. A total of N = 41 adults (36 COVID-positive and 5 COVID-negative), and N = 42 children (37 COVID-positive, and 5 COVID-negative) have paired serum and DBS assayed. We found moderate to strong correlations between paired hemaPEN DBS eluates and serum SARS-CoV-2 IgG antibodies for RBD (r = 0.9472, <i>p</i> < 0.0001) and S1 proteins (r = 0.6892, <i>p</i> < 0.0001). Similar results were observed in both adult and paediatric populations. No significant differences in S1-specific IgG levels were observed in hemaPEN DBS samples stored for up to 35 weeks at room temperature. Eluted hemaPEN samples showed high specificity and sensitivity (100% and 89.89%, respectively) compared with serum. The use of the microsampling hemaPEN device for DBS sample collection is a feasible approach for assessing SARS-CoV-2 antibodies for serosurveillance studies, particularly in remote settings and in children.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-02-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843976/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466815","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Efficient Identification of Monoclonal Antibodies Against Rift Valley Fever Virus Using High-Throughput Single Lymphocyte Transcriptomics of Immunized Mice.","authors":"Ronit Rosenfeld, Ron Alcalay, Yfat Yahalom-Ronen, Sharon Melamed, Avital Sarusi-Portuguez, Tal Noy-Porat, Ofir Israeli, Adi Beth-Din, Ronnie Blecher-Gonen, Theodor Chitlaru, Erez Bar-Haim, Tomer Israely, Anat Zvi, Efi Makdasi","doi":"10.3390/antib14010012","DOIUrl":"10.3390/antib14010012","url":null,"abstract":"<p><p><b>Background</b>: Rift Valley fever virus (RVFV) is a zoonotic virus that poses a significant threat to both livestock and human health and has caused outbreaks in endemic regions. In humans, most patients experience a febrile illness; however, in some patients, RVF disease may result in hemorrhagic fever, retinitis, or encephalitis. While several veterinary vaccines are being utilized in endemic countries, currently, there are no licensed RVF vaccines or therapeutics for human use. Neutralizing antibodies specifically targeting vulnerable pathogen epitopes are promising candidates for prophylactic and therapeutic interventions. In the case of RVFV, the surface glycoproteins Gc and Gn, which harbor neutralizing epitopes, represent the primary targets for vaccine and neutralizing antibody development. <b>Methods</b>: We report the implementation of advanced 10x Genomics technology, enabling high-throughput single-cell analysis for the identification of rare and potent antibodies against RVFV. Following the immunization of mice with live attenuated rMP-12-GFP virus and successive Gc/Gn boosts, memory B cell populations (both general and antigen-specific) were sorted from splenocytes by flow cytometry. Deep sequencing of the antibody repertoire at a single-cell resolution, together with bioinformatic analyses, was applied for BCR pair selection based on their abundance and specificity. <b>Results</b>: Twenty-three recombinant monoclonal antibodies (mAbs) were selected and expressed, and their antigen-binding capacities were characterized. About half of them demonstrated specific binding to their cognate antigen with relatively high binding affinities. <b>Conclusions</b>: These antibodies could be used for the future development of efficacious therapeutics, as well as for studying virus-neutralizing mechanisms. The current study, in which the single-cell sequencing approach was implemented for the development of antibodies targeting the RVFV surface proteins Gc and Gn, demonstrates the effective applicability of this technique for antibody discovery purposes.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-02-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843919/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466813","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Purification of a Fc-Fusion Protein with [Bathophenathroline:metal] Complexes.","authors":"Thisara Jayawickrama Withanage, Ron Alcalay, Olga Krichevsky, Ellen Wachtel, Ohad Mazor, Guy Patchornik","doi":"10.3390/antib14010011","DOIUrl":"10.3390/antib14010011","url":null,"abstract":"<p><p>In this study, we assess an alternative Fc-fusion protein purification method that does not rely on chromatographic media or ligands. Recombinant human acetylcholinesterase, fused to the Fc domain of human IgG1 (henceforth, AChE-Fc), was purified with precipitated aromatic complexes composed of the bathophenanthroline (henceforth, batho) chelator with either Zn²⁺ or Cu²⁺ ions (i.e., [(batho)₃:Zn²⁺] or [(batho)₂:Cu²⁺]) in the presence of polyethylene glycol 6000 (PEG-6000). In a three-step purification process conducted at pH 7, AChE-Fc was captured by the aromatic complexes (Step 1); unbound or weakly bound protein impurities were removed with 20 mM NaCl (Step 2); and AChE-Fc was then extracted at pH 7 (Step 3) using 100 mM Na citrate buffer in 250 mM NaCl. Purified AChE-Fc was not aggregated (as determined by dynamic light scattering (DLS) and Native PAGE). However, full enzymatic activity was only preserved with the [(batho)₃:Zn²⁺] complex. Interaction between AChE-Fc and [(batho)₃:Zn²⁺] led to ~83-88% overall protein yield. Thirty-fold process upscaling by volume required only proportional increase in the amounts of [(batho)₃:Zn²⁺] and PEG-6000. Efficient (95-97%) chelator recycling was achieved by recrystallization. Chelator leaching into purified AchE-Fc was estimated to be ~0.3% relative to the total amount used. Taken together, this novel procedure has the potential to provide an economical and practical avenue for the industrial purification of Fc-fusion proteins.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843901/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466839","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Influence of High <i>Eimeria tenella</i> Immunization Dosages on Total Oocyst Output and Specific Antibodies Recognition Response in Hybrid Pullets (<i>Gallus gallus</i>)-A Pilot Study.","authors":"Marco A Juarez-Estrada, Guillermo Tellez-Isaias, Víctor M Petrone-Garcia, Amanda Gayosso-Vazquez, Xochitl Hernandez-Velasco, Rogelio A Alonso-Morales","doi":"10.3390/antib14010009","DOIUrl":"10.3390/antib14010009","url":null,"abstract":"<p><strong>Background: </strong>Two high primary-immunization doses of a wild-type <i>E. tenella</i> strain were assessed in healthy pullets (5K <i>versus</i> 10K sporulated oocysts/bird) to understand the effects of coccidia infection.</p><p><strong>Methods: </strong>Acquired immunity was evaluated following primary immunization and two booster doses with the homologous strain. Total oocyst shedding, clinical signs, and viability of every bird/group after each immunization/booster were recorded. Indirect ELISA measured the time course of humoral responses from each immunization group against sporozoite and second-generation merozoite of <i>E. tenella</i>. Antigen pattern recognition on these two asexual zoite stages of <i>E. tenella</i> was analyzed using Western blotting with antibodies from each immunization program. Afterwards, antigen recognition of specific life-cycle stages was performed using individual pullet serums from the best immunization program.</p><p><strong>Results: </strong>A primary-immunization dose of 1 × 10⁴ oocysts/bird reduced the oocyst output; however, all pullets exhibited severe clinical signs and low specific antibodies titers, with decreased polypeptide recognition on both <i>E. tenella</i> asexual zoite stages. In contrast, immunization with 5 × 10³ oocysts/bird yielded the best outcomes regarding increased oocyst collection and early development of sterilizing immunity. After the first booster dosage, this group's antisera revealed a strong pattern of specific antigen recognition on the two assayed <i>E. tenella</i> life-cycle stages.</p><p><strong>Conclusions: </strong>The <i>E. tenella</i>-specific antibodies from the 5 × 10³ oocysts/bird immunization program can aid in passive immunization trials and further research to identify B-cell immunoprotective antigens, which could help in the development of a genetically modified anticoccidial vaccine.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843834/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466834","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Exploring Anticitrullinated Antibodies (ACPAs) and Serum-Derived Exosomes Cargoes.","authors":"Mohammed A Alghamdi, Sami M Bahlas, Sultan Abdulmughni Alamry, Ehab H Mattar, Elrashdy M Redwan","doi":"10.3390/antib14010010","DOIUrl":"10.3390/antib14010010","url":null,"abstract":"<p><strong>Background: </strong>Autoantibodies such as rheumatoid factor (RF) and anticitrullinated protein autoantibodies (ACPAs) are useful tools for rheumatoid arthritis (RA). The presence of ACPAs against citrullinated proteins (CPs), especially citrullinated fibrinogen (cFBG), seems to be a useful serological marker for diagnosing RA. RA patients' sera were found to be enriched in exosomes that can transmit many proteins. Exosomes have been found to express citrullinated protein such as cFBG.</p><p><strong>Objective: </strong>We conducted this study in two stages. In the first phase, we aimed to evaluate the association between autoantibodies and risk factors. In the next step, ACPA-positive serum samples from the first phase were subjected to exosomal studies to explore the presence of cFBG, which is a frequent target for ACPAs.</p><p><strong>Methods: </strong>We investigated the autoantibodies in one hundred and sixteen Saudi RA patients and correlated with host-related risk factors. Exosomes were extracted from patients' sera and examined for the presence of cFBG using monoclonal antibodies.</p><p><strong>Results: </strong>The study reported a high female-to-male ratio of 8:1, and seropositive RA (SPRA) was more frequent among included RA patients. The frequency and the levels of ACPAs were similar in both genders. Autoantibodies incidences have a direct correlations with patient age, while the average titers decreased as the age increased. Further, the highest incidence and levels of autoantibodies were reported in patients with RA duration between 5 and 10 years. Smoking and family history have no impact on autoantibody, except for ACPAs titers among smokers' RA. Our analysis of serum exosomes revealed that about 50% of SPRA patients expressed cFBG.</p><p><strong>Conclusions: </strong>The female-to-male ratio is 8:1, which is higher than the global ratio. We can conclude that patients' age and disease duration contribute to the autoantibodies, particularly RF and anti-MCV, whereas smoking and family history had no effects on autoantibodies. We detected cFBG in all exosomes from SPRA patients; thus, we suggest that the precise mechanism of exosomes in RA pathogenesis can be investigated to develop effective treatment strategies.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843936/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466828","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"In Vitro Functional Validation of an Anti-FREM2 Nanobody for Glioblastoma Cell Targeting.","authors":"Gloria Krapež, Neja Šamec, Alja Zottel, Mojca Katrašnik, Ana Kump, Jernej Šribar, Igor Križaj, Jurij Stojan, Rok Romih, Gregor Bajc, Matej Butala, Serge Muyldermans, Ivana Jovčevska","doi":"10.3390/antib14010008","DOIUrl":"10.3390/antib14010008","url":null,"abstract":"<p><p><b>Background/Objectives</b>: Glioblastomas are the most common brain malignancies. Despite the implementation of multimodal therapy, patient life expectancy after diagnosis is barely 12 to 18 months. Glioblastomas are highly heterogeneous at the genetic and epigenetic level and comprise multiple different cell subpopulations. Therefore, small molecules such as nanobodies, able to target membrane proteins specific to glioblastoma cells or specific cell types within the tumor are being investigated as novel tools to treat glioblastomas. <b>Methods</b>: Here, we describe the identification of such a nanobody and its in silico and in vitro validation. NB3F18, as we named it, is directed against the membrane-associated protein FREM2, overexpressed in glioblastoma stem cells. <b>Results</b>: Three dimensional in silico modeling indicated that NB3F18 and FREM2 form a stable complex. Surface plasmon resonance confirmed their interaction with moderate affinity. As we demonstrated by flow cytometry, NB3F18 binds to glioblastoma stem cells to a greater extent than to differentiated glioblastoma cells and astrocytes. Immunocytochemistry revealed surface localization of NB3F18 on glioblastoma stem cells, whereas cytoplasmic localization of NB3F18 was observed in other cell lines. NB3F18 was detected by transmission electron microscopy on the plasma membrane and in various compartments of the endocytic pathway, from endocytic vesicles to multivesicular bodies (endosomes) and lysosomes. Interestingly, NB3F18 was cytotoxic to glioblastoma stem cells. <b>Conclusions</b>: Collectively, NB3F18 has been qualified as an interesting tool to target glioblastoma cells and as a potential vehicle to deliver biological or pharmaceutical agents to these cells.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-01-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11843905/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143466833","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Strategy for Simultaneous Engineering of Interspecies Cross-Reactivity, Thermostability, and Expression of a Bispecific 5T4 x CD3 DART^® Molecule for Treatment of Solid Tumors.","authors":"Renhua R Huang, Michael Spliedt, Tom Kaufman, Sergey Gorlatov, Bhaswati Barat, Kalpana Shah, Jeffrey Gill, Kurt Stahl, Jennifer DiChiara, Qian Wang, Jonathan C Li, Ralph Alderson, Paul A Moore, Jennifer G Brown, James Tamura, Xiaoyu Zhang, Ezio Bonvini, Gundo Diedrich","doi":"10.3390/antib14010007","DOIUrl":"10.3390/antib14010007","url":null,"abstract":"<p><p><b>Background:</b> Bispecific antibodies represent a promising class of biologics for cancer treatment. However, their dual specificity and complex structure pose challenges in the engineering process, often resulting in molecules with good functional but poor physicochemical properties. <b>Method:</b> To overcome limitations in the properties of an anti-5T4 x anti-CD3 (α5T4 x αCD3) DART molecule, a phage-display method was developed, which succeeded in simultaneously engineering cross-reactivity to the cynomolgus 5T4 ortholog, improving thermostability and the elevating expression level. <b>Results:</b> This approach generated multiple DART molecules that exhibited significant improvements in all three properties. The lead DART molecule demonstrated potent in vitro and in vivo anti-tumor activity. Although its clearance in human FcRn-transgenic mice was comparable to that of the parental molecule, faster clearance was observed in cynomolgus monkeys. The lead α5T4 x αCD3 DART molecule displayed no evidence of off-target binding or polyspecificity, suggesting that the increased affinity for the target may account for its accelerated clearance in cynomolgus monkeys. <b>Conclusions:</b> This may reflect target-mediated drug disposition (TMDD), a potential limitation of targeting 5T4, despite its limited expression in healthy tissues.</p>","PeriodicalId":8188,"journal":{"name":"Antibodies","volume":"14 1","pages":""},"PeriodicalIF":3.0,"publicationDate":"2025-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11755548/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143021766","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}