Transplantation
Abstract
Lymph node (LN) fibroblastic reticular cells (FRCs) define LN niches and regulate lymphocyte homeostasis through producing diverse extracellular matrix (ECM) components. We examined the role of ECM laminin α4 (Lama4) using FRC-Lama4 conditional KO Pdgfrb-Cre–/– × Lama4fl/fl mice. Single-cell RNA-sequencing (scRNA-Seq) data showed the promoter gene Pdgfrb was exclusively expressed in FRCs. Depleting FRC-Lama4 reduced Tregs and dendritic cells, decreased high endothelial venules, impaired the conduit system, and downregulated T cell survival factors in LNs. FRC-Lama4 depletion impaired the homing of lymphocytes to LNs in homeostasis and after allografting. Alloantigen-specific T cells proliferated, were activated to greater degrees in LNs lacking FRC-Lama4, and were more prone to differentiate into effector phenotypes relative to the Treg phenotype. In murine cardiac transplantation, tolerogenic immunosuppression was not effective in FRC-Lama4 recipients, which produced more alloantibodies than WT. After lung transplantation, FRC-Lama4–KO mice had more severe graft rejection with fewer Tregs in their LNs. Overall, FRC-Lama4 critically contributes to a tolerogenic LN niche by supporting T cell migration, constraining T cell activation and proliferation, and promoting Treg differentiation. Hence, it serves as a therapeutic target for immunoengineering.
Authors
Lushen Li, Marina W. Shirkey, Tianshu Zhang, Wenji Piao, Xiaofei Li, Jing Zhao, Zhongcheng Mei, Yizhan Guo, Vikas Saxena, Allison Kensiski, Samuel J. Gavzy, Yang Song, Bing Ma, Jing Wu, Yanbao Xiong, Long Wu, Xiaoxuan Fan, Holly Roussey, Meng Li, Alexæander S. Krupnick, Reza Abdi, Jonathan S. Bromberg
Abstract
DNA methyltransferase 3a (DNMT3a) is an important part of the epigenetic machinery that stabilizes patterns of activated T-cell responses. We hypothesized that donor T-cell DNMT3a regulates alloreactivity after allogeneic blood and marrow transplantation (allo-BMT). T-cell conditional Dnmt3a knock-out (KO) animals were used as donors in murine allo-BMT models. Mice receiving allo-BMT from KO donors developed severe acute graft-versus-host disease (aGVHD), with increases in inflammatory cytokine levels and organ histopathology. KO T-cells migrated and proliferated in secondary lymphoid organs earlier and demonstrated a trafficking advantage to the small intestine. Donor T-cell subsets were purified post-BMT for whole genome bisulfite sequencing (WGBS) and RNA sequencing. KO T-cells had similar global methylation to wild-type (WT), with distinct, localized areas of hypomethylation. Using a highly sensitive computational method, we produced a comprehensive profile of the altered epigenome landscape. Hypomethylation corresponded with changes in gene expression in several pathways of T-cell signaling and differentiation. Additionally, Dnmt3a KO T-cells conveyed superior graft-versus-tumor activity. Our findings demonstrate a critical role for DNMT3a in regulating T-cell alloreactivity and illuminate pathways that control T-cell tolerance. These results also provide a platform to decipher clinical data that associate donor DNMT3a mutations with increased GVHD, decreased relapse, and improved survival.
Authors
Yiouli P. Ktena, Michael A. Koldobskiy, Michael I. Barbato, Han-Hsuan Fu, Leo Luznik, Nicolas J. Llosa, Azeb Haile, Orly R. Klein, Chen Liu, Christopher J. Gamper, Kenneth R. Cooke
IL-33 acts as a costimulatory signal to generate alloreactive Th1 cells in graft-versus-host disease
Abstract
Antigen-presenting cells (APC) integrate signals emanating from local pathology and program appropriate T cell responses. In allogeneic hematopoietic stem cell transplantation (alloHCT), recipient conditioning releases Damage-Associated Molecular Patterns (DAMPs) that generate pro-inflammatory APC that secrete IL-12, which is a driver of donor Type 1 T helper (Th1) responses causing graft vs. host disease (GVHD). Nevertheless, other mechanisms exist to initiate alloreactive T cells responses, as recipients with disrupted DAMP signaling or lacking IL-12 develop GVHD. We established that tissue damage signals are perceived directly by donor CD4+ T cells and promoted T cell expansion and differentiation. Specifically, the fibroblastic reticular cell-derived DAMP, IL-33, is increased by recipient conditioning and is critical for the initial activation, proliferation, and differentiation of alloreactive Th1 cells. IL-33-stimulation of CD4+ T cell was not required for lymphopenia-induced expansion, however. IL-33 promoted IL-12-independent expression of Tbet and generation of Th1 cells that infiltrated GVHD target tissues. Mechanistically, IL-33 augmented CD4+ T cell TCR-associated signaling pathways in response to alloantigen. This enhanced T cell expansion and Th1 polarization, but inhibited the expression of regulatory molecules like IL-10 and Foxp3. These data established an unappreciated role for IL-33 as a costimulatory signal for donor Th1 generation after alloHCT.
Authors
Gaelen K. Dwyer, Lisa R. Mathews, Jose A. Villegas, Anna Lucas, Anne Gonzalez de Peredo, Bruce R. Blazar, Jean-Philippe Girard, Amanda C. Poholek, Sanjiv A. Luther, Warren Shlomchik, Hēth R. Turnquist
Abstract
Once human photoreceptors die, they do not regenerate, thus photoreceptor transplantation has emerged as a potential treatment approach for blinding diseases. Improvements in transplant organization, donor cell maturation and synaptic connectivity to the host will be critical in advancing this technology to clinical practice. Unlike the unstructured grafts of prior cell suspension transplantations into end-stage degeneration models, we describe extensive incorporation of iPSC retinal organoid-derived human photoreceptors into mice with cone dysfunction. This incorporative phenotype was validated in both cone-only as well as pan-photoreceptor transplantations. Rather than forming a glial barrier, Müller cells extended throughout the graft, even forming a series of adherens junctions between mouse and human cells, reminiscent of an outer limiting membrane. Donor-host interaction appeared to promote polarisation as well as development of morphological features critical for light detection, namely formation of inner and well stacked outer segments oriented towards the retinal pigment epithelium. Putative synapse formation and graft function was evident both at a structural and electrophysiological level. Overall, these results show that human photoreceptors interact readily with a partially degenerated retina. Moreover, incorporation into the host retina appears to be beneficial to graft maturation, polarisation and function.
Authors
Sylvia J. Gasparini, Karen Tessmer, Miriam Reh, Stephanie Wieneke, Madalena Carido, Manuela Völkner, Oliver Borsch, Anka Swiersy, Marta Zuzic, Olivier Goureau, Thomas Kurth, Volker Busskamp, Günther Zeck, Mike O. Karl, Marius Ader
Abstract
BACKGROUND. Responses to conventional donor lymphocyte infusion (DLI) for post-allogeneic hematopoietic cell transplantation (HCT) relapse are typically poor. Natural killer (NK) cell-based therapy is a promising modality to treat post-HCT relapse. METHODS. We initiated this ongoing phase I trial of adoptively transferred cytokine induced memory-like (CIML) NK cells in patients with myeloid malignancies relapsed after haploidentical HCT. All patients received a donor-derived NK cell dose of 5–10 million cells/kg after lymphodepleting chemotherapy, followed by systemic IL-2 for 7 doses. High resolution profiling with mass cytometry and single cell RNA sequencing characterized the expanding and persistent NK cell subpopulations in a longitudinal manner after infusion. RESULTS. In the first 6 enrolled patients on the trial, infusion of CIML NK cells led to a rapid 10 to 50-fold in vivo expansion that was sustained over months. The infusion was well-tolerated, with fever and pancytopenia as the most common adverse events. Expansion of NK cells was distinct from IL-2 effects on endogenous post-HCT NK cells, and not dependent on CMV viremia. Immunophenotypic and transcriptional profiling revealed a dynamic evolution of the activated CIML NK cell phenotype, superimposed on the natural variation in donor NK cell repertoires. CONCLUSION. Given their rapid expansion and long-term persistence in an immune compatible environment, CIML NK cells serve as a promising platform for the treatment of post-transplant relapse of myeloid disease. Further characterization of their unique in vivo biology and interaction with both T cells and tumor targets will lead to improvements in cell-based immunotherapies. TRIAL REGISTRATION. NCT04024761 FUNDING. Supported by Dunkin Donuts Breakthrough Award, the NIH/National Cancer Institute R21 CA245413, the Leukemia and Lymphoma Society Scholar and TRP awards.
Authors
Roman M. Shapiro, Grace C. Birch, Guangan Hu, Juliana Vergara Cadavid, Sarah Nikiforow, Joanna Baginska, Alaa K. Ali, Mubin Tarannum, Michal Sheffer, Yasmin Z. Abdulhamid, Benedetta Rambaldi, Yohei Arihara, Carol Reynolds, Max S. Halpern, Scott J. Rodig, Nicole Cullen, Jacquelyn O. Wolff, Kathleen L. Pfaff, Andrew A. Lane, R. Coleman Lindsley, Corey S. Cutler, Joseph H. Antin, Vincent T. Ho, John Koreth, Mahasweta Gooptu, Haesook T. Kim, Karl-Johan Malmberg, Catherine J. Wu, Jianzhu Chen, Robert J. Soiffer, Jerome Ritz, Rizwan Romee
Abstract
Despite the curative potential of hematopoietic stem cell transplantation (HSCT), conditioning-associated toxicities preclude broader clinical application. Antibody-drug conjugates (ADC) provide an attractive approach to HSCT conditioning that minimizes toxicity while retaining efficacy. Initial studies of ADC conditioning have largely focused on syngeneic HSCT. However, to treat acute leukemias or induce tolerance for solid organ transplantation, this approach must be expanded to allogeneic HSCT (allo-HSCT). Using murine allo-HSCT models, we show that pharmacologic Janus kinase 1/2 (JAK1/2) inhibition combined with CD45- or cKit-targeted ADCs enables robust multilineage alloengraftment. Strikingly, myeloid lineage donor chimerism exceeding 99% was achievable in fully MHC-mismatched HSCT using this approach. Mechanistic studies using the JAK1/2 inhibitor baricitinib revealed marked impairment of T and NK cell survival, proliferation and effector function. NK cells were exquisitely sensitive to JAK1/2 inhibition due to interference with IL-15 signaling. Unlike irradiated mice, ADC-conditioned mice did not develop pathogenic graft-versus-host alloreactivity when challenged with mismatched T cells. Finally, the combination of ADCs and baricitinib balanced graft-versus-host disease and graft-versus-leukemia responses in delayed donor lymphocyte infusion models. Our allo-HSCT conditioning strategy exemplifies the promise of immunotherapy to improve the safety of HSCT for treating hematologic diseases.
Authors
Stephen P. Persaud, Julie K. Ritchey, Sena Kim, Sora Lim, Peter G. Ruminski, Matthew L. Cooper, Michael P. Rettig, Jaebok Choi, John F. DiPersio
Abstract
Apolipoprotein L1 (APOL1) risk-alleles in donor kidneys associate with graft loss but whether recipient risk-allele expression impacts transplant outcomes is unclear. To test whether recipient APOL1 risk-alleles independently correlate with transplant outcomes, we analyzed genome-wide SNP genotyping data of donors and recipients from two kidney transplant cohorts, Genomics of Chronic Allograft Rejection (GOCAR) and Clinical Trials in Organ Transplantation (CTOT1/17). We estimated genetic ancestry (quantified as proportion of African ancestry or pAFR) by ADMIXTURE and correlated APOL1 genotypes and pAFR with outcomes. In the GOCAR discovery set, we observed that the number of recipient APOL1 G1/G2 alleles (R-nAPOL1) associated with increased risk of death-censored allograft loss (DCAL), independent of ancestry (HR = 2.14; P = 0.006), and within the subgroup of African American and Hispanic (AA/H) recipients (HR = 2.36; P = 0.003). R-nAPOL1 also associated with increased risk of any T cell-mediated rejection (TCMR) event. These associations were validated in CTOT1/17. Ex vivo studies of peripheral blood mononuclear cells revealed unanticipated high APOL1 expression in activated CD4+/CD8+ T cells and natural killer cells. We detected enriched immune response gene pathways in risk-allele carriers vs. non-carriers on the kidney transplant waitlist and among healthy controls. Our findings demonstrate an immunomodulatory role for recipient APOL1 risk-alleles associating with TCMR and DCAL. This finding has broader implications for immune mediated injury to native kidneys.
Authors
Zhongyang Zhang, Zeguo Sun, Jia Fu, Qisheng Lin, Khadija Banu, Kinsuk Chauhan, Marina Planoutene, Chengguo Wei, Fadi Salem, Zhengzi Yi, Ruijie Liu, Paolo Cravedi, Haoxiang Cheng, Ke Hao, Philip J. O’Connell, Shuta Ishibe, Weijia Zhang, Steven G. Coca, Ian W. Gibson, Robert B. Colvin, John C. He, Peter S. Heeger, Barbara T. Murphy, Madhav C. Menon
Abstract
While direct allorecognition underpins both solid organ allograft rejection and tolerance induction, the specific molecular targets of most directly-alloreactive CD8+ T cells have not been defined. In this study, we used a combination of genetically-engineered MHC class I (MHC I) constructs, mice with a hepatocyte-specific mutation in the class I antigen-presentation pathway and immunopeptidomic analysis to provide definitive evidence for the contribution of the peptide cargo of allogeneic MHC I molecules to transplant tolerance induction. We established a systematic approach for the discovery of directly-recognised pMHC epitopes, and identified 17 strongly immunogenic H-2Kb-associated peptides recognised by CD8+ T cells from B10.BR (H-2k) mice, 13 of which were also recognised by BALB/c (H-2d) mice. As few as five different tetramers used together were able to identify a high proportion of alloreactive T cells within a polyclonal population, suggesting that there are immunodominant allogeneic MHC-peptide complexes that can account for a large component of the alloresponse.
Authors
Eric T. Son, Pouya Faridi, Moumita Paul-Heng, Mario L. Leong, Kieran English, Sri H. Ramarathinam, Asolina Braun, Nadine L. Dudek, Ian E. Alexander, Leszek Lisowski, Patrick Bertolino, David G. Bowen, Anthony W. Purcell, Nicole A. Mifsud, Alexandra F. Sharland
Abstract
Novel mRNA-based vaccines have been proven powerful tools to combat the global pandemic caused by SARS-CoV2 with BNT162b2 (trade name: Comirnaty) efficiently protecting individuals from COVID-19 across a broad age range. Still, it remains largely unknown how renal insufficiency and immunosuppressive medication affect development of vaccine induced immunity. We therefore comprehensively analyzed humoral and cellular responses in kidney transplant recipients after the standard second vaccination dose. As opposed to all healthy vaccinees and the majority of hemodialysis patients, only 4/39 and 1/39 transplanted individuals showed IgA and IgG seroconversion at day 8 ± 1 after booster immunization with minor changes until day 23 ± 5, respectively. Although most transplanted patients mounted spike-specific T helper cell responses, frequencies were significantly reduced compared to controls and dialysis patients, accompanied by a broad impairment in effector cytokine production, memory differentiation and activation-related signatures. Spike-specific CD8+ T cell responses were less abundant than their CD4+ counterparts in healthy controls and hemodialysis patients and almost undetectable in transplant patients. Promotion of anti-HLA antibodies or acute rejection was not detected after vaccination. In summary, our data strongly suggest revised vaccination approaches in immunosuppressed patients, including individual immune monitoring for protection of this vulnerable group at risk to develop severe COVID-19.
Authors
Arne Sattler, Eva Schrezenmeier, Ulrike A. Weber, Alexander Potekhin, Friederike Bachmann, Henriette Straub-Hohenbleicher, Klemens Budde, Elena Storz, Vanessa Proß, Yasmin Bergmann, Linda M.L. Thole, Caroline Tizian, Oliver Hölsken, Andreas Diefenbach, Hubert Schrezenmeier, Bernd Jahrsdörfer, Tomasz Zemojtel, Katharina Jechow, Christian Conrad, Sören Lukassen, Diana Stauch, Nils Lachmann, Mira Choi, Fabian Halleck, Katja Kotsch
Abstract
Adoptive transfer of Tregs has been shown to improve alloengraftment in animal models. However, it is technically challenging to expand Tregs ex vivo for the purpose of infusing large numbers of cells in the clinic. We demonstrate an innovative approach to engineering an orthogonal IL-2/IL-2 receptor (IL-2R) pair, the parts of which selectively interact with each other, transmitting native IL-2 signals, but do not interact with the natural IL-2 or IL-2R counterparts, thereby enabling selective stimulation of target cells in vivo. Here, we introduced this orthogonal IL-2R into Tregs. Upon adoptive transfer in a murine mixed hematopoietic chimerism model, orthogonal IL-2 injection significantly promoted orthogonal IL-2R+Foxp3GFP+CD4+ cell proliferation without increasing other T cell subsets and facilitated donor hematopoietic cell engraftment followed by acceptance of heart allografts. Our data indicate that selective target cell stimulation enabled by the engineered orthogonal cytokine receptor improves Treg potential for the induction of organ transplantation tolerance.
Authors
Toshihito Hirai, Teresa L. Ramos, Po-Yu Lin, Federico Simonetta, Leon L. Su, Lora K. Picton, Jeanette Baker, Jian-Xin Lin, Peng Li, Kinya Seo, Juliane K. Lohmeyer, Sara B. Wagers, Melissa Mavers, Warren J. Leonard, Bruce R. Blazar, K. Christopher Garcia, Robert S. Negrin
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ISSN: 0021-9738 (print), 1558-8238 (online)