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Article: cMPL-Based Purification and Depletion of Human Hematopoietic Stem Cells: Implications for Pre-Transplant Conditioning.

Araki, Daisuke / Hong, Sogun / Linde, Nathaniel / Fisk, Bryan / Redekar, Neelam / Salisbury-Ruf, Christi / Krouse, Allen / Engels, Theresa / Golomb, Justin / Dagur, Pradeep / Magnani, Diogo M / Wang, Zhirui / Larochelle, Andre

bioRxiv : the preprint server for biology

2024

Abstract: The transplantation of gene-modified autologous hematopoietic stem and progenitor cells (HSPCs) offers a promising therapeutic approach for hematological and immunological disorders. However, this strategy is often limited by the toxicities associated ... ...

Abstract	The transplantation of gene-modified autologous hematopoietic stem and progenitor cells (HSPCs) offers a promising therapeutic approach for hematological and immunological disorders. However, this strategy is often limited by the toxicities associated with traditional conditioning regimens. Antibody-based conditioning strategies targeting cKIT and CD45 antigens have shown potential in mitigating these toxicities, but their long-term safety and efficacy in clinical settings require further validation. In this study, we investigate the thrombopoietin (TPO) receptor, cMPL, as a novel target for conditioning protocols. We demonstrate that high surface expression of cMPL is a hallmark feature of long-term repopulating hematopoietic stem cells (LT-HSCs) within the adult human CD34+ HSPC subset. Targeting the cMPL receptor facilitates the separation of human LT-HSCs from mature progenitors, a delineation not achievable with cKIT. Leveraging this finding, we developed a cMPL-targeting immunotoxin, demonstrating its ability to selectively deplete host cMPL
Language	English
Publishing date	2024-02-28
Publishing country	United States
Document type	Preprint
DOI	10.1101/2024.02.24.581887
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Article ; Online: Antibody-mediated depletion of select leukocyte subsets in blood and tissue of nonhuman primates.

Sutton, Matthew S / Bucsan, Allison N / Lehman, Chelsea C / Kamath, Megha / Pokkali, Supriya / Magnani, Diogo M / Seder, Robert / Darrah, Patricia A / Roederer, Mario

Frontiers in immunology

2024 Volume 15, Page(s) 1359679

Abstract: Understanding the immunological control of pathogens requires a detailed evaluation of the mechanistic contributions of individual cell types within the immune system. While knockout mouse models that lack certain cell types have been used to help define ...

Abstract	Understanding the immunological control of pathogens requires a detailed evaluation of the mechanistic contributions of individual cell types within the immune system. While knockout mouse models that lack certain cell types have been used to help define the role of those cells, the biological and physiological characteristics of mice do not necessarily recapitulate that of a human. To overcome some of these differences, studies often look towards nonhuman primates (NHPs) due to their close phylogenetic relationship to humans. To evaluate the immunological role of select cell types, the NHP model provides distinct advantages since NHP more closely mirror the disease manifestations and immunological characteristics of humans. However, many of the experimental manipulations routinely used in mice (e.g., gene knock-out) cannot be used with the NHP model. As an alternative, the
MeSH term(s)	Animals ; Humans ; Mice ; Phylogeny ; Antibodies, Monoclonal/pharmacology ; T-Lymphocytes ; Primates
Chemical Substances	Antibodies, Monoclonal
Language	English
Publishing date	2024-03-11
Publishing country	Switzerland
Document type	Journal Article ; Research Support, N.I.H., Intramural
ZDB-ID	2606827-8
ISSN	1664-3224 ; 1664-3224
ISSN (online)	1664-3224
ISSN	1664-3224
DOI	10.3389/fimmu.2024.1359679
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Article: Do organisms need an impact factor? Citations of key biological resources including model organisms reveal usage patterns and impact.

Piekniewska, Agata / Anderson, Nathan / Roelandse, Martijn / Lloyd, K C Kent / Korf, Ian / Voss, S Randal / de Castro, Giovanni / Magnani, Diogo M / Varga, Zoltan / James-Zorn, Christina / Horb, Marko / Grethe, Jeffery S / Bandrowski, Anita

bioRxiv : the preprint server for biology

2024

Abstract: Research resources like transgenic animals and antibodies are the workhorses of biomedicine, enabling investigators to relatively easily study specific disease conditions. As key biological resources, transgenic animals and antibodies are often validated, ...

Abstract	Research resources like transgenic animals and antibodies are the workhorses of biomedicine, enabling investigators to relatively easily study specific disease conditions. As key biological resources, transgenic animals and antibodies are often validated, maintained, and distributed from university based stock centers. As these centers heavily rely largely on grant funding, it is critical that they are cited by investigators so that usage can be tracked. However, unlike systems for tracking the impact of papers, the conventions and systems for tracking key resource usage and impact lag behind. Previous studies have shown that about 50% of the resources are not findable, making the studies they are supporting irreproducible, but also makes tracking resources difficult. The RRID project is filling this gap by working with journals and resource providers to improve citation practices and to track the usage of these key resources. Here, we reviewed 10 years of citation practices for five university based stock centers, characterizing each reference into two broad categories: findable (authors could use the RRID, stock number, or full name) and not findable (authors could use a nickname or a common name that is not unique to the resource). The data revealed that when stock centers asked their communities to cite resources by RRID, in addition to helping stock centers more easily track resource usage by increasing the number of RRID papers, authors shifted from citing resources predominantly by nickname (~50% of the time) to citing them by one of the findable categories (~85%) in a matter of several years. In the case of one stock center, the MMRRC, the improvement in findability is also associated with improvements in the adherence to NIH rigor criteria, as determined by a significant increase in the Rigor and Transparency Index for studies using MMRRC mice. From this data, it was not possible to determine whether outreach to authors or changes to stock center websites drove better citation practices, but findability of research resources and rigor adherence was improved.
Language	English
Publishing date	2024-01-16
Publishing country	United States
Document type	Preprint
DOI	10.1101/2024.01.15.575636
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Article: Antibody-mediated depletion of select T cell subsets in blood and tissue of nonhuman primates.

Sutton, Matthew S / Bucsan, Allison N / Lehman, Chelsea C / Kamath, Megha / Pokkali, Supriya / Magnani, Diogo M / Seder, Robert / Darrah, Patricia A / Roederer, Mario

bioRxiv : the preprint server for biology

2023

Abstract	Understanding the immunological control of pathogens requires a detailed evaluation of the mechanistic contributions of individual cell types within the immune system. While knockout mouse models that lack certain cell types have been used to help define the role of those cells, the biological and physiological characteristics of mice do not necessarily recapitulate that of a human. To overcome some of these differences, studies often look towards nonhuman primates (NHPs) due to their close phylogenetic relationship to humans. To evaluate the immunological role of select cell types, the NHP model provides distinct advantages since NHP more closely mirror the disease manifestations and immunological characteristics of humans. However, many of the experimental manipulations routinely used in mice (e.g., gene knock-out) cannot be used with the NHP model. As an alternative, the
Language	English
Publishing date	2023-12-23
Publishing country	United States
Document type	Preprint
DOI	10.1101/2023.12.22.572898
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Article ; Online: The impact of IdeS (imlifidase) on allo-specific, xeno-reactive, and protective antibodies in a sensitized rhesus macaque model.

DeLaura, Isabel / Zikos, Joanna / Anwar, Imran J / Yoon, Janghoon / Ladowski, Joseph / Jackson, Annette / Van Rompay, Koen / Magnani, Diogo / Knechtle, Stuart J / Kwun, Jean

Xenotransplantation

2023 Volume 31, Issue 1, Page(s) e12833

Abstract: Background: Highly sensitized patients face many barriers to kidney transplantation, including higher rates of antibody-mediated rejection after HLA-incompatible transplant. IdeS, an endopeptidase that cleaves IgG nonspecifically, has been trialed as ... ...

Abstract	Background: Highly sensitized patients face many barriers to kidney transplantation, including higher rates of antibody-mediated rejection after HLA-incompatible transplant. IdeS, an endopeptidase that cleaves IgG nonspecifically, has been trialed as desensitization prior to kidney transplant, and successfully cleaves donor-specific antibody (DSA), albeit with rebound. Methods: IdeS was generated and tested (2 mg/kg, IV) in two naïve and four allosensitized nonhuman primates (NHP). Peripheral blood samples were collected at regular intervals following IdeS administration. Total IgG, total IgM, and anti-CMV antibodies were quantified with ELISA, and donor-specific antibody (DSA) and anti-pig antibodies were evaluated using flow cytometric crossmatch. B cell populations were assessed using flow cytometry. Results: IdeS successfully cleaved rhesus IgG in vitro. In allosensitized NHP, robust reduction of total, DSA, anti-pig, and anti-CMV IgG was observed within one day following IdeS administration. Rapid rebound of all IgG antibody populations was observed, with antibody levels returning to baseline around day 14 post-infusion. Total IgM level was not affected by IdeS. Interestingly, a comparable reduction in antibody populations was observed after the second dose of IdeS. However, we have not observed any significant modulation of B cell subpopulations after IdeS. Conclusions: This study evaluated efficacy of IdeS in the allosensitized NHP in IgG with various specificities, mirroring antibody kinetics in human patients. The efficacy of IdeS on preexisting anti-pig antibodies may be useful in clinical xenotransplantation. However, given the limitation of IdeS on its durability as a monotherapy, optimization of IdeS with other agents targeting the humoral response is further needed.
MeSH term(s)	Animals ; Humans ; Macaca mulatta ; Isoantibodies ; Graft Rejection/prevention & control ; Transplantation, Heterologous ; Immunosuppressive Agents/therapeutic use ; Immunoglobulin G ; Immunoglobulin M ; HLA Antigens
Chemical Substances	Isoantibodies ; Immunosuppressive Agents ; Immunoglobulin G ; Immunoglobulin M ; HLA Antigens
Language	English
Publishing date	2023-10-21
Publishing country	Denmark
Document type	Journal Article
ZDB-ID	1236298-0
ISSN	1399-3089 ; 0908-665X
ISSN (online)	1399-3089
ISSN	0908-665X
DOI	10.1111/xen.12833
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Article ; Online: SOSIP Trimer-Specific Antibodies Isolated from a Simian-Human Immunodeficiency Virus-Infected Monkey with versus without a Pre-blocking Step with gp41.

Duggan, Natasha N / Weisgrau, Kim L / Magnani, Diogo M / Rakasz, Eva G / Desrosiers, Ronald C / Martinez-Navio, Jose M

Journal of virology

2021 Volume 96, Issue 2, Page(s) e0158221

Abstract: BG505 SOSIP.664 (hereafter referred to as SOSIP), a stabilized trimeric mimic of the HIV-1 envelope spike resembling the native viral spike, is a useful tool for isolating anti-HIV-1 neutralizing antibodies. We screened long-term SHIV-AD8 infected rhesus ...

Abstract	BG505 SOSIP.664 (hereafter referred to as SOSIP), a stabilized trimeric mimic of the HIV-1 envelope spike resembling the native viral spike, is a useful tool for isolating anti-HIV-1 neutralizing antibodies. We screened long-term SHIV-AD8 infected rhesus monkeys for potency and breadth of serum neutralizing activity against autologous and heterologous viruses: SHIV-AD8, HIV-1 YU2, HIV-1 JR-CSF, and HIV-1 NL4-3. Monkey rh2436 neutralized all viruses tested and showed strong reactivity to the SOSIP trimer, suggesting this was a promising candidate for attempts at monoclonal antibody (MAb) isolation. MAbs were isolated by performing single B-cell sorts from peripheral blood mononuclear cells (PBMC) by FACS using the SOSIP trimer as a probe. An initial round of sorted cells revealed the majority of isolated MAbs were directed to the gp41 external domain portion of the SOSIP trimer and were mostly non-neutralizing against tested isolates. A second sort was performed, introducing a gp41 blocking step prior to PBMC staining and FACS sorting. These isolated MAbs bound SOSIP trimer but were no longer directed to the gp41 external domain portion. A significantly higher proportion of MAbs with neutralizing activity were obtained with this strategy. Our data show this pre-blocking step with gp41 greatly increases the yield of non-gp41-reactive, SOSIP-specific MAbs and increases the likelihood of isolating MAbs with neutralizing activity.
MeSH term(s)	Animals ; Antibodies, Monoclonal/genetics ; Antibodies, Monoclonal/immunology ; Antibodies, Monoclonal/isolation & purification ; Antibody Specificity ; Broadly Neutralizing Antibodies/genetics ; Broadly Neutralizing Antibodies/immunology ; Broadly Neutralizing Antibodies/isolation & purification ; HIV Antibodies/genetics ; HIV Antibodies/immunology ; HIV Antibodies/isolation & purification ; HIV Envelope Protein gp41/immunology ; HIV-1/immunology ; Immunoglobulin Variable Region/genetics ; Macaca mulatta ; Recombinant Proteins/immunology ; Simian Immunodeficiency Virus/immunology ; env Gene Products, Human Immunodeficiency Virus/immunology
Chemical Substances	Antibodies, Monoclonal ; Broadly Neutralizing Antibodies ; HIV Antibodies ; HIV Envelope Protein gp41 ; Immunoglobulin Variable Region ; Recombinant Proteins ; env Gene Products, Human Immunodeficiency Virus
Language	English
Publishing date	2021-11-03
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	80174-4
ISSN	1098-5514 ; 0022-538X
ISSN (online)	1098-5514
ISSN	0022-538X
DOI	10.1128/JVI.01582-21
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Article ; Online: A comparative study of human-and rhesus-specific antithymocyte globulins in Rhesus macaques.

Shaw, Brian I / Schmitz, Robin / Flores, Walter J / Magnani, Diogo M / Li, Jie / Song, Mingqing / Kirk, Allan D

Clinical transplantation

2021 Volume 35, Issue 8, Page(s) e14369

Abstract: Rabbit antithymocyte globulin (RATG) preparations are widely used in transplantation. They are developed in vivo against thymocytes and contain polyclonal antibodies specific for myriad cellular targets. The rhesus monkey is commonly used as a ... ...

Abstract	Rabbit antithymocyte globulin (RATG) preparations are widely used in transplantation. They are developed in vivo against thymocytes and contain polyclonal antibodies specific for myriad cellular targets. The rhesus monkey is commonly used as a preclinical transplant model, but the fidelity of commercially available human-specific RATGs to anticipate the effects of RATGs in rhesus has not been established. We therefore developed two rhesus-specific ATGs (rhATG) and compared them to human-specific RATG (huATG, Thymoglobulin
MeSH term(s)	Animals ; Antilymphocyte Serum ; Flow Cytometry ; Humans ; Macaca mulatta
Chemical Substances	Antilymphocyte Serum
Language	English
Publishing date	2021-05-31
Publishing country	Denmark
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	639001-8
ISSN	1399-0012 ; 0902-0063
ISSN (online)	1399-0012
ISSN	0902-0063
DOI	10.1111/ctr.14369
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Article ; Online: Genetic modifications designed for xenotransplantation attenuate sialoadhesin-dependent binding of human erythrocytes to porcine macrophages.

Petitpas, Kaitlyn / Habibabady, Zahra / Ritchie, Veronica / Connolly, Margaret R / Burdorf, Lars / Qin, Wenning / Kan, Yinan / Layer, Jacob V / Crabtree, Juliet N / Youd, Michele E / Westlin, William F / Magnani, Diogo M / Pierson, Richard N / Azimzadeh, Agnes M

Xenotransplantation

2022 Volume 29, Issue 6, Page(s) e12780

Abstract: The phenomenon of diminishing hematocrit after in vivo liver and lung xenotransplantation and during ex vivo liver xenoperfusion has largely been attributed to action by resident liver porcine macrophages, which bind and destroy human erythrocytes. ... ...

Abstract	The phenomenon of diminishing hematocrit after in vivo liver and lung xenotransplantation and during ex vivo liver xenoperfusion has largely been attributed to action by resident liver porcine macrophages, which bind and destroy human erythrocytes. Porcine sialoadhesin (siglec-1) was implicated previously in this interaction. This study examines the effect of porcine genetic modifications, including knockout of the CMAH gene responsible for expression of Neu5Gc sialic acid, on the adhesion of human red blood cells (RBCs) to porcine macrophages. Wild-type (WT) porcine macrophages and macrophages from several strains of genetically engineered pigs, including CMAH gene knockout and several human transgenes (TKO+hTg), were incubated with human RBCs and "rosettes" (≥3 erythrocytes bound to one macrophage) were quantified by microscopy. Our results show that TKO+hTg genetic modifications significantly reduced rosette formation. The monoclonal antibody 1F1, which blocks porcine sialoadhesin, significantly reduced rosette formation by WT and TKO+hTg macrophages compared with an isotype control antibody. Further, desialation of human RBCs with neuraminidase before addition to WT or TKO+hTg macrophages resulted in near-complete abrogation of rosette formation, to a level not significantly different from porcine RBC rosette formation on porcine macrophages. These observations are consistent with rosette formation being mediated by binding of sialic acid on human RBCs to sialoadhesin on porcine macrophages. In conclusion, the data predict that TKO+hTg genetic modifications, coupled with targeting of porcine sialoadhesin by the 1F1 mAb, will attenuate erythrocyte sequestration and anemia during ex vivo xenoperfusion and following in vivo liver, lung, and potentially other organ xenotransplantation.
MeSH term(s)	Humans ; Swine ; Animals ; Sialic Acid Binding Ig-like Lectin 1/genetics ; Transplantation, Heterologous/methods ; N-Acetylneuraminic Acid/metabolism ; Macrophages ; Erythrocytes/metabolism
Chemical Substances	Sialic Acid Binding Ig-like Lectin 1 ; N-Acetylneuraminic Acid (GZP2782OP0)
Language	English
Publishing date	2022-09-20
Publishing country	Denmark
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	1236298-0
ISSN	1399-3089 ; 0908-665X
ISSN (online)	1399-3089
ISSN	0908-665X
DOI	10.1111/xen.12780
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Article: SIV clearance from neonatal macaques following transient CCR5 depletion.

bioRxiv : the preprint server for biology

2023

Abstract: Treatment of people with HIV (PWH) with antiretroviral therapy (ART) results in sustained suppression of viremia, but HIV persists indefinitely as integrated provirus in CD4-expressing cells. Intact persistent provirus, the "rebound competent viral ... ...

Abstract	Treatment of people with HIV (PWH) with antiretroviral therapy (ART) results in sustained suppression of viremia, but HIV persists indefinitely as integrated provirus in CD4-expressing cells. Intact persistent provirus, the "rebound competent viral reservoir" (RCVR), is the primary obstacle to achieving a cure. Most variants of HIV enter CD4
Language	English
Publishing date	2023-05-01
Publishing country	United States
Document type	Preprint
DOI	10.1101/2023.05.01.533682
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Article ; Online: Measuring the Impact of Targeting FcRn-Mediated IgG Recycling on Donor-Specific Alloantibodies in a Sensitized NHP Model.

Manook, Miriam / Flores, Walter J / Schmitz, Robin / Fitch, Zachary / Yoon, Janghoon / Bae, Yeeun / Shaw, Brian / Kirk, Allan / Harnois, Melissa / Permar, Sallie / Farris, Alton B / Magnani, Diogo M / Kwun, Jean / Knechtle, Stuart

Frontiers in immunology

2021 Volume 12, Page(s) 660900

Abstract: Background: In transplantation, plasmapheresis and IVIg provide the mainstay of treatment directed at reducing or removing circulating donor-specific antibody (DSA), yet both have limitations. We sought to test the efficacy of targeting the IgG ... ...

Abstract	Background: In transplantation, plasmapheresis and IVIg provide the mainstay of treatment directed at reducing or removing circulating donor-specific antibody (DSA), yet both have limitations. We sought to test the efficacy of targeting the IgG recycling mechanism of the neonatal Fc receptor (FcRn) using anti-FcRn mAb therapy in a sensitized non-human primate (NHP) model, as a pharmacological means of lowering DSA. Methods: Six (6) rhesus macaque monkeys, previously sensitized by skin transplantation, received a single dose of 30mg/kg anti-RhFcRn IV, and effects on total IgG, as well as DSA IgG, were measured, in addition to IgM and protective immunity. Subsequently, 60mg/kg IV was given in the setting of kidney transplantation from skin graft donors. Kidney transplant recipients received RhATG, and tacrolimus, MMF, and steroid for maintenance immunosuppression. Results: Circulating total IgG was reduced from a baseline 100% on D0 to 32.0% (mean, SD ± 10.6) on d4 post infusion (p<0.05), while using a DSA assay. T-cell flow cross match (TFXM) was reduced to 40.6±12.5% of baseline, and B-cell FXCM to 52.2±19.3%. Circulating total IgM and DSA IgM were unaffected by treatment. Pathogen-specific antibodies (anti-gB and anti-tetanus toxin IgG) were significantly reduced for 14d post infusion. Post-transplant, circulating IgG responded to anti-FcRn mAb treatment, but DSA increased rapidly. Conclusion: Targeting the FcRn-mediated recycling of IgG is an effective means of lowering circulating donor-specific IgG in the sensitized recipient, although in the setting of organ transplantation mechanisms of rapid antibody rise post-transplant remains unaffected.
MeSH term(s)	Animals ; Antibodies, Monoclonal/therapeutic use ; Graft Rejection/immunology ; Graft Rejection/prevention & control ; Graft Survival ; Histocompatibility Antigens Class I/immunology ; Histocompatibility Testing ; Immunoglobulin G/blood ; Immunoglobulin G/immunology ; Immunoglobulins, Intravenous/therapeutic use ; Immunosuppression Therapy ; Immunosuppressive Agents/administration & dosage ; Isoantibodies/immunology ; Kidney Transplantation ; Macaca mulatta ; Male ; Models, Animal ; Receptors, Fc/immunology ; Tissue Donors
Chemical Substances	Antibodies, Monoclonal ; Histocompatibility Antigens Class I ; Immunoglobulin G ; Immunoglobulins, Intravenous ; Immunosuppressive Agents ; Isoantibodies ; Receptors, Fc ; Fc receptor, neonatal (TW3XAW0RCY)
Language	English
Publishing date	2021-06-02
Publishing country	Switzerland
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	2606827-8
ISSN	1664-3224 ; 1664-3224
ISSN (online)	1664-3224
ISSN	1664-3224
DOI	10.3389/fimmu.2021.660900
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