Costimulation Blockade Research Articles

T Cell Costimulation Blockade with Abatacept Nearly Eliminates Early Severe Acute Graft Versus Host Disease after HLA-Mismatched (7/8 HLA Matched) Unrelated Donor Transplant, with a Favorable Impact on Disease-Free and Overall Survival

HLA mismatched unrelated donor (MMURD) HCT plays a critical role in the treatment of high-risk hematologic malignancies in ethnic minorities and others lacking a fully matched donor. However, the high incidence of severe (Grade III-IV) acute GVHD (AGVHD) and resultant mortality remain critical barriers to success. T cell costimulation blockade with CTLA4-Ig has been demonstrated to potently inhibit T cell activation, and is FDA-approved for rheumatoid arthritis and renal transplant. We previously completed a pilot study adding 4 doses of the ‘abatacept’ formulation of CTLA4-Ig to calcineurin inhibitor (CNI) + methotrexate (MTX) GVHD prophylaxis in URD HCT (BBMT 19:1638-49, 2013). Based on robust immune reconstitution and a low rate of AGVHD, we initiated a multicenter phase II trial of abatacept (10 mg/kg/dose, Days -1, +5, +14, +28) plus CNI/MTX (‘ABA2‘) in patients undergoing URD HCT (Clinicaltrials.gov # NCT01743131).ABA2 has 2 cohorts: (1) 8/8 URD (a randomized double-blind stratum vs CNI/MTX, current accrual =133 of 140 patients) and (2) 7/8 MMURD (a single-arm stratum with 43 patients, enrollment completed in Nov, 2016). The 7/8 stratum was non-randomized due to ethical concerns related to the high risk of death from severe AGVHD in 7/8 URD HCT in the absence of adjunctive immunosuppression. The 7/8 stratum thus underwent a pre-specified comparison to CIBMTR matched controls. Here we report the results from the 7/8 stratum with a follow-up of 227-1544 days (median 559).The primary outcome was the incidence of severe (Gr III-IV) AGVHD prior to day +100. Key secondary outcomes were Gr II-IV AGVHD, chronic GVHD (CGVHD), graft failure, engraftment, transplant-related mortality (TRM), relapse, disease-free survival (DFS) and overall survival (OS). Key inclusion criteria were age >6, myeloablative conditioning and a high-risk heme malignancy. Key exclusion criteria were the use of ATG, alemtuzumab, or other T cell depletion, or a cord blood graft. Stopping rules were in place for TRM and PTLD, neither of which were met. We compared ABA2 to two CIBMTR matched control cohorts: (1) 7/8 URD HCT using CNI + MTX (n =127) or (2) CNI/MTX plus anti-thymocyte globulin (‘+ATG’, n = 128). Matching was based on age, performance score and disease stage, with similar baseline characteristics (Table 1).42 of 43 ABA2 patients received all 4 abatacept doses, and all 43 successfully engrafted donor cells, with no delay in median neutrophil (17 days) or platelet (18 days) engraftment. ABA2 was associated with a marked reduction in the cumulative incidence of grade III-IV AGVHD at day 100, with only 2.3% Gr III-IV AGVHD in ABA2 vs 30% in CNI+MTX and 22% in +ATG (p =0.0070 and 0.025, Fig. 1A, 2A). Grade II-IV AGVHD was similar in ABA2 vs controls (44.9% in ABA2 vs 53% in CNI+MTX and 45% in +ATG, p= 0.08 and 0.36). CGVHD was also similar: 40% at 12 months in ABA2 vs 41% in CNI+MTX and 37% in +ATG (p =ns). Importantly, the reduction in severe AGVHD in ABA2 was achieved without significant impact on patient safety, resulting in striking improvement in 12 month TRM (12.5% vs 34% in CNI+MTX and 27% in +ATG (p = 0.002 and 0.04, Fig. 1B, 2B). Also noteworthy, there was no increase in relapse in ABA2, with 7.5% relapse at 12 months compared to 15% in CNI+MTX and 11% in +ATG (p=0.04 and 0.26). Detailed analyses of viral reactivation and other infectious outcomes are ongoing, but uncontrolled infections were not a major problem. This led to very promising rates of DFS and OS in ABA2: 81% 12 month DFS, which was better than both CNI+MTX (50%) and +ATG (63%), p=0.0003 and 0.016, Fig. 1C, 2C). The same was true for OS: 85.4% OS at 12 months in ABA2, vs CNI+MTX (57%) and +ATG (68%, p=0.0025 and 0.048 respectively, Fig. 1D, 2D). These survival advantages are stable thus-far through 2 years.Our results strongly suggest that the administration of 4 doses of abatacept for GVHD prophylaxis is safe and highly effective at preventing severe AGVHD, lessens the risk for TRM and improves survival in MMURD HCT for heme malignancies. Our results also suggest that the administration of 4 doses of abatacept prior to day +30 may not be sufficient to impact CGVHD, thus trials to assess the effect of longer courses of abatacept are warranted. These results are the first to demonstrate significant efficacy of in vivo T cell costimulation blockade in enhancing outcomes after MMURD HCT, which could significantly expand the donor pool and improve outcomes for patients requiring these high-risk transplants. [Display omitted] DisclosuresNorkin:Celgene: Honoraria, Research Funding. Pulsipher:Jazz Pharmaceuticals: Other: advisory board, education; CSL Behring: Other: advisory board Feb 2017; Chimerix: Other: Advisory board Dec 2015; Novartis Pharmaceuticals Corporation: Consultancy, Other: Phase II steering committee; Adaptive: Other: Advisory board 6/17. Kean:Juno: Research Funding; Regeneron: Research Funding; Kymab Ltd: Research Funding; Bristol Myers Squibb: Consultancy.

Pancreas transplantation.

To define recent changes and future directions in the practice of pancreas transplantation (PT). Two major events have occurred in the past 18 months: COVID-19 pandemic, and the first world consensus conference on PT. Several innovative studies were published after the consensus conference. During COVID-19 pandemic PT activity decreased. COVID-19 in transplant recipients increases mortality rates, but data from kidney transplantation show that mortality might be higher in waitlisted patients.The world consensus conference provided 49 jury deliberations on the impact of PT on management of diabetic patients and 110 practice recommendations.Recent evidence demonstrates that PT alone is safe and effective, that results of simultaneous pancreas and kidney (SPK) remain excellent despite older recipient age and higher prevalence of type 2 diabetes, that use of hepatitis C virus (HCV)-positive donors into HCV-negative recipients is associated with good outcomes, and that use of sirolimus as primary immunosuppressant and costimulation blockade does not improve results of SPK. COVID-19 pandemic and the first world consensus conference on PT were major events. Although COVID-19 pandemic should not reduce PT activity in the future, a major positive impact on both volume and outcomes of PT is awaited from the proceedings of the world consensus conference.

Costimulatory blockade modifies germinal center B cell function in NZB/W F1 mice

Abstract Systemic lupus erythematosus (SLE) is a complex autoimmune disease with significant morbidity and mortality - demanding further examination of disease reducing treatments. Previously, we showed that a 2-week course of combined CTLA4Ig and anti-CD40L costimulatory blockade prevented loss of tolerance to dsDNA in NZB/W mice while maintaining immune function against exogenous antigen, explained by an altered selection of autoreactive B cells in germinal centers (GC). Here, we examined both B and T cells to clarify if restoration of tolerance is due to a change in GC B cell profile or a decrease in T cell help. 3 months after treatment, large germinal centers persisted in the treated group, but due to smaller spleen size total GC B cells and TFH Cells were decreased compared with controls. The metabolic profile of splenic B and T cells showed a decreased spare respiratory capacity in both subsets and a decrease in B cell glycolysis among the treated group. RNA-seq analysis of GC dark zone and light zone B cells from treated mice showed decreased expression of markers of oxidative phosphorylation and cell proliferation and increased expression of signaling molecules downstream of BCR, including PLCγ, compared with untreated controls. GC B cell stimulation assays showed a delay in phosphorylation of PLCγ in response to anti-IgG in treated mice. These data show a long-lasting effect of short-term costimulatory blockade on both B and T cells with metabolic remodeling of GC B cells and an alteration of BCR signaling. Since both a profound decrease in BCR signaling and an increase in B cell glycolysis have been reported to enhance loss of tolerance in lupus prone mice, our findings help to explain the restoration of tolerance observed after treatment.

Traumatic brain injury does not disrupt costimulatory blockade-induced immunological tolerance to glial-restricted progenitor allografts

BackgroundCell transplantation-based treatments for neurological disease are promising, yet graft rejection remains a major barrier to successful regenerative therapies. Our group and others have shown that long-lasting tolerance of transplanted stem cells can be achieved in the brain with systemic application of monoclonal antibodies blocking co-stimulation signaling. However, it is unknown if subsequent injury and the blood-brain barrier breach could expose the transplanted cells to systemic immune system spurring fulminant rejection and fatal encephalitis. Therefore, we investigated whether delayed traumatic brain injury (TBI) could trigger graft rejection.MethodsGlial-restricted precursor cells (GRPs) were intracerebroventricularly transplanted in immunocompetent neonatal mice and co-stimulation blockade (CoB) was applied 0, 2, 4, and 6 days post-grafting. Bioluminescence imaging (BLI) was performed to monitor the grafted cell survival. Mice were subjected to TBI 12 weeks post-transplantation. MRI and open-field test were performed to assess the brain damage and behavioral change, respectively. The animals were decapitated at week 16 post-transplantation, and the brains were harvested. The survival and distribution of grafted cells were verified from brain sections. Hematoxylin and eosin staining (HE) was performed to observe TBI-induced brain legion, and neuroinflammation was evaluated immunohistochemically.ResultsBLI showed that grafted GRPs were rejected within 4 weeks after transplantation without CoB, while CoB administration resulted in long-term survival of allografts. BLI signal had a steep rise following TBI and subsequently declined but remained higher than the preinjury level. Open-field test showed TBI-induced anxiety for all animals but neither CoB nor GRP transplantation intensified the symptom. HE and MRI demonstrated a reduction in TBI-induced lesion volume in GRP-transplanted mice compared with non-transplanted mice. Brain sections further validated the survival of grafted GRPs and showed more GRPs surrounding the injured tissue. Furthermore, the brains of post-TBI shiverer mice had increased activation of microglia and astrocytes compared to post-TBI wildtype mice, but infiltration of CD45+ leukocytes remained low.ConclusionsCoB induces sustained immunological tolerance towards allografted cerebral GRPs which is not disrupted following TBI, and unexpectedly TBI may enhance GRPs engraftment and contribute to post-injury brain tissue repair.

CMV Status Drives Distinct Trajectories of CD4+ T Cell Differentiation.

Cytomegalovirus (CMV) is one of the most commonly recognized opportunistic pathogens and remains the most influential known parameter in shaping an individual’s immune system. As such, T cells induced by CMV infection could have a long-term impact on subsequent immune responses. Accumulating evidence indicates that memory T cells developed during past bacterial and viral infection can cross-react with unrelated pathogens, including transplant antigens, and can alter responses to de novo infections, vaccines, cancers, or rejection. Therefore, careful examination of T cell responses elicited by CMV is warranted to understand their potentially beneficial or harmful roles in future major immune events. Our detailed exploration of the distribution, phenotype, TCR repertoire and transcriptome of CD4+ T cells within CMV seropositive healthy individuals using high-dimensional flow cytometry and single cell multi-omics sequencing reveals that CMV seropositivity has highly significant age-independent effects, leading to a reduction in CD4+ naïve T cells and an expansion of CD4+ effector memory T cells and CD45RA+ effector memory T cells. These induced CD4+ effector memory T cells undergo a specific differentiation trajectory resulting in a subpopulation of CD57+CD27-CD28-CD244+ CD4+ T cells with cytotoxic function and TCR oligoclonality for optimal controlled coexistence with cytomegalovirus. Through gene set enrichment analysis, we found that this subpopulation is similar to virus-specific CD8+ T cells and T cells that mediate acute rejection in patients using tacrolimus and belatacept, a selective costimulation blocker. Together, these data suggest that memory CD4+ T cells induced by cytomegalovirus are formed via a distinct differentiation program to acquire cytotoxic function and can be potentially detrimental to transplant patients adopting costimulation blockade immunosuppressive regimen.

Blockade of tumor necrosis factor superfamily members CD30 and OX40 abrogates disease activity in murine immune-mediated glomerulonephritis

Co-stimulation is a prerequisite for pathogenic activity in T cell-mediated diseases and has been demonstrated to achieve tolerance in organ-specific autoimmunity as a therapeutic target. Here, we evaluated the involvement of the tumor necrosis factor family members CD30 and OX40 in immune-complex mediated kidney disease. Invitro stimulation and proliferation studies were performed with CD4+ cells from wild type and CD30/OX40 double knock-out (CD30OX40-/-) mice. Invivo studies were performed by induction of nephrotoxic serum nephritis in wild type, CD30OX40- /- , CD30-/-, OX40-/-, reconstituted Rag1-/- and C57Bl/6J mice treated with αCD30L αOX40L antibodies. CD30, OX40 and their ligands were upregulated on various leukocytes in nephrotoxic serum nephritis. CD30OX40-/- mice, but not CD30-/- or OX40-/- mice were protected from nephrotoxic serum nephritis. Similar protection was found in Rag1-/- mice injected with CD4+ T cells from CD30OX40-/- mice compared to Rag1-/- mice injected with CD4+ T cells from wild type mice. Furthermore, CD4+ T cells deficient in CD30OX40-/- displayed decreased expression of CCR6 invivo. CD30OX40-/- cells were fully capable of differentiating into disease mediating T helper cell subsets, but showed significantly decreased levels of proliferation invivo and invitro compared to wild type cells. Blocking antibodies against CD30L and OX40L ameliorated nephrotoxic serum nephritis without affecting pan-effector or memory T cell populations. Thus, our results indicate disease promotion via CD30 and OX40 signaling due to facilitation of exaggerated T cell proliferation and migration of T helper 17 cells in nephrotoxic serum nephritis. Hence, co-stimulation blockade targeting the CD30 and OX40 signaling pathways may provide a novel therapeutic strategy in autoimmune kidney disease.

Low Dose Interleukin-2 Induces Exosomes with Tolerance Markers (PDL1, CD73) and Significantly Delays Development of Chronic Rejection Following Murine Heterotopic Cardiac Transplantation

Purpose Aim of the study is to determine the effect of low dose interleukin-2 (IL-2) therapy on the kinetics of induction of circulating exosomes with tolerance markers (PDL1, CD73) and their role in abrogating chronic rejection following murine heterotopic cardiac transplant (HTx). Methods Allogenic (BALB/c to C57BL/6) HTx was performed heterotopically in abdominal cavity. Costimulatory blockade consisting of MR1 (250 μg/ip day 0) and CTLA4-Ig (200 μg/ip day 2) was performed. Low dose IL-2 (2000IU/day) was administered from day 15 post-transplant for 3 weeks. Sera was collected on days 7-100 for kinetic analysis of exosome induction and characterization of PDL1, CD73, cardiac self-antigens (SAgs) (Myosin, Vimentin), class II MHC trans-activator (CIITA), and 20S proteasome by western blot using specific antibodies (Abs) and Abs to SAgs. Results BALB/c to C57BL/6 HTx rejected transplanted heart with median survival time of 8 (7-9) days. HTx done following costimulatory blockade developed chronic rejection with mean survival time 26 (7-47) days. Exosome and Ab development demonstrated induction of exosomes with Vimentin (2.2± 1.0 vs 0.6±0.1; p Conclusion Circulating exosomes containing cardiac SAgs and development of Abs to SAgs lead to chronic rejection in a murine model occurring after costimulation blockade following allogenic HTx. Low dose IL-2 abrogated chronic rejection and induced circulating exosome with tolerance markers. This result supports an important role for exosomes, inducing tolerance and significantly delaying development of chronic rejection in murine model of chronic cardiac allograft rejection.

Cardiac Allograft Tolerance Induction with Ox40L Costimulatory Blockade

<h3>Purpose</h3> Cardiac allograft tolerance in non-human primates (NHPs) has been achieved by our group via hematopoietic mixed chimerism with allogeneic bone marrow and kidney co-transplantation. This model has failed without the regulatory mechanisms associated with the donor kidney. We hypothesized that rapamycin and anti-OX40L mAb (KY1005, Kymab Ltd) could substitute for kidney co-transplantation and induce cardiac allograft tolerance by providing a favorable immunologic milieu for regulatory T cells (Tregs). <h3>Methods</h3> Fourteen cynomolgus NHPs underwent mixed chimerism conditioning (ATGAM, total body and thymic irradiation) then organ and bone marrow transplant. Heart Alone recipients received a heart transplant (n=5), Heart/Kidney recipients received a heart and kidney co-transplant (n=7). Heart/OX40L recipients received a heart transplant plus 40 days of anti-OX40L mAb (n=2). All groups underwent a 12 day course of aCD154 mAb. The Heart Alone and Heart Kidney groups received 28 days of cyclosporine while the Heart aOX40L group received 100 days of rapamycin. <h3>Results</h3> All Heart Alone recipients had cellular rejection (ISHLT 3R) by 200 days post bone marrow transplant (pBMT). No Heart/Kidney recipients showed early rejection. Similarly, no Heart/OX40L recipients showed early rejection and one is currently 258 days pBMT without clinical or pathologic evidence of rejection (Figure 1). The other recipient died of viral complications. Mean peak lymphoid chimerism was 10% for the Heart Alone group, 26% for the Heart/Kidney group, and 29% for the Heart/OX40L group. The anti-OX40L protocol also showed an expansion of the Treg population with a peak of 35% Tregs in the CD4+ population. <h3>Conclusion</h3> The OX40L protocol achieved robust lymphoid chimerism with concomitant expansion of Tregs. The surviving Heart/OX40L recipient achieved longer freedom from allograft rejection when compared to the Heart Alone group. OX40L costimulatory blockade may allow for heart allograft tolerance without kidney co-transplantation.

Recent Advances in Costimulatory Blockade to Induce Immune Tolerance in Liver Transplantation

Liver transplantation is an effective therapy for end-stage liver disease. However, most postoperative patients must take immunosuppressive drugs to prevent organ rejection. Interestingly, some transplant recipients have normal liver function and do not experience organ rejection after the withdrawal of immunosuppressive agents. This phenomenon, called immune tolerance, is the ultimate goal in clinical transplantation. Costimulatory molecules play important roles in T cell-mediated immune responses and the maintenance of T cell tolerance. Blocking costimulatory pathways can alter T cell responses and prolong graft survival. Better understanding of the roles of costimulatory molecules has facilitated the use of costimulatory blockade to effectively induce immune tolerance in animal transplantation models. In this article, we review the state of the art in costimulatory pathway blockade for the induction of immune tolerance in transplantation and its potential application prospects for liver transplantation.

Research Highlights.

Research Highlights.

Berberine Promotes Induction of Immunological Tolerance to an Allograft via Downregulating Memory CD8+ T-Cells Through Altering the Gut Microbiota.

Emerging evidence has linked the gut microbiota dysbiosis to transplant rejection while memory T-cells pose a threat to long-term transplant survival. However, it's unclear if the gut microbiome alters the formation and function of alloreactive memory T-cells. Here we studied the effects of berberine, a narrow-spectrum antibiotic that is barely absorbed when orally administered, on the gut microbiota, memory T-cells, and allograft survival. In this study, C57BL/6 mice transplanted with islets or a heart from BALB/c mice were treated orally with berberine. Allograft survival was observed, while spleen, and lymph node T-cells from recipient mice were analyzed using a flow cytometer. High-throughput sequencing and qPCR were performed to analyze the gut microbiota. CD8+ T-cells from recipients were cultured with the bacteria to determine potential T-cell memory cross-reactivity to a specific pathogen. We found that berberine suppressed islet allograft rejection, reduced effector CD8+CD44highCD62Llow and central memory CD8+CD44highCD62Lhigh T-cells (TCM), altered the gut microbiota composition and specifically lowered Bacillus cereus abundance. Further, berberine promoted long-term islet allograft survival induced by conventional costimulatory blockade and induced cardiac allograft tolerance as well. Re-colonization of B. cereus upregulated CD8+ TCM cells and reversed long-term islet allograft survival induced by berberine plus the conventional costimulatory blockade. Finally, alloantigen-experienced memory CD8+ T-cells from transplanted recipients rapidly responded to B. cereus in vitro. Thus, berberine prolonged allograft survival by repressing CD8+ TCM through regulating the gut microbiota. We have provided the first evidence that donor-specific memory T-cell generation is linked to a specific microbe and uncovered a novel mechanism underlying the therapeutic effects of berberine. This study may be implicated for suppressing human transplant rejection since berberine is already used in clinic to treat intestinal infections.

433 - T Cell Co-Stimulatory Blockade By CTLA4-Ig Does Not Appear to Impact the Clinical Efficacy of Viral Specific T Cell Therapy in Pediatric Patients after HSCT

433 - T Cell Co-Stimulatory Blockade By CTLA4-Ig Does Not Appear to Impact the Clinical Efficacy of Viral Specific T Cell Therapy in Pediatric Patients after HSCT

What Therapeutic Regimen Will Be Optimal for Initial Clinical Trials of Pig Organ Transplantation?

We discuss what therapeutic regimen might be acceptable/successful in the first clinical trial of genetically engineered pig kidney or heart transplantation. As regimens based on a calcineurin inhibitor or CTLA4-Ig have proved unsuccessful, the regimen we administer to baboons is based on induction therapy with antithymocyte globulin, an anti-CD20 mAb (Rituximab), and cobra venom factor, with maintenance therapy based on blockade of the CD40/CD154 costimulation pathway (with an anti-CD40 mAb), with rapamycin, and a corticosteroid. An anti-inflammatory agent (etanercept) is administered for the first 2 wk, and adjuvant therapy includes prophylaxis against thrombotic complications, anemia, cytomegalovirus, and pneumocystis. Using this regimen, although antibody-mediated rejection certainly can occur, we have documented no definite evidence of an adaptive immune response to the pig xenograft. This regimen could also form the basis for the first clinical trial, except that cobra venom factor will be replaced by a clinically approved agent, for example, a C1-esterase inhibitor. However, none of the agents that block the CD40/CD154 pathway are yet approved for clinical use, and so this hurdle remains to be overcome. The role of anti-inflammatory agents remains unproven. The major difference between this suggested regimen and those used in allotransplantation is the replacement of a calcineurin inhibitor with a costimulation blockade agent, but this does not appear to increase the complications of the regimen.

Pregnancy-induced humoral sensitization overrides T cell tolerance to fetus-matched allografts in mice.

Immunological tolerance to semiallogeneic fetuses is necessary to achieving successful first pregnancy and permitting subsequent pregnancies with the same father. Paradoxically, pregnancy is an important cause of sensitization, resulting in the accelerated rejection of offspring-matched allografts. The underlying basis for divergent outcomes following reencounter of the same alloantigens on transplanted organs versus fetuses in postpartum females is incompletely understood. Using a mouse model that allows concurrent tracking of endogenous fetus-specific T and B cell responses in a single recipient, we show that semiallogeneic pregnancies simultaneously induce fetus-specific T cell tolerance and humoral sensitization. Pregnancy-induced antibodies, but not B cells, impeded transplantation tolerance elicited by costimulation blockade to offspring-matched cardiac grafts. Remarkably, in B cell-deficient mice, allogeneic pregnancy enabled the spontaneous acceptance of fetus-matched allografts. The presence of pregnancy-sensitized B cells that cannot secrete antibodies at the time of heart transplantation was sufficient to precipitate rejection and override pregnancy-established T cell tolerance. Thus, while induction of memory B cells and alloantibodies by pregnancies establishes formidable barriers to transplant success for multigravid women, our observations raise the possibility that humoral desensitization will not only improve transplantation outcomes, but also reveal an unexpected propensity of multiparous recipients to achieve tolerance to offspring-matched allografts.

Engineering of a trispecific tumor-targeted immunotherapy incorporating 4-1BB co-stimulation and PD-L1 blockade

ABSTRACT Co-stimulatory 4–1BB receptors on tumor-infiltrating T cells are a compelling target for overcoming resistance to immune checkpoint inhibitors, but initial clinical studies of 4–1BB agonist mAbs were accompanied by liver toxicity. We sought to engineer a tri-specific antibody-based molecule that stimulates intratumoral 4–1BB and blocks PD-L1/PD-1 signaling without systemic toxicity and with clinically favorable pharmacokinetics. Recombinant fusion proteins were constructed using scMATCH3 technology and humanized antibody single-chain variable fragments against PD-L1, 4–1BB, and human serum albumin. Paratope affinities were optimized using single amino acid substitutions, leading to design of the drug candidate NM21-1480. Multiple in vitro experiments evaluated pharmacodynamic properties of NM21-1480, and syngeneic mouse tumor models assessed antitumor efficacy and safety of murine analogues. A GLP multiple-dose toxicology study evaluated its safety in non-human primates. NM21-1480 inhibited PD-L1/PD-1 signaling with a potency similar to avelumab, and it potently stimulated 4–1BB signaling only in the presence of PD-L1, while exhibiting an EC50 that was largely independent of PD-L1 density. NM21-1480 exhibited high efficacy for co-activation of pre-stimulated T cells and dendritic cells. In xenograft models in syngeneic mice, NM21-1480 induced tumor regression and tumor infiltration of T cells without causing systemic T-cell activation. A GLP toxicology study revealed no evidence of liver toxicity at doses up to 140 mg/kg, and pharmacokinetic studies in non-human primates suggested a plasma half-life in humans of up to 2 weeks. NM21-1480 has the potential to overcome checkpoint resistance by co-activating tumor-infiltrating lymphocytes without liver toxicity.

New concepts in chronic antibody-mediated kidney allograft rejection: prevention and treatment.

Chronic antibody-mediated rejection (AMR) is a cardinal cause of transplant failure, with currently no proven effective prevention or treatment. The present review will focus on new therapeutic concepts currently under clinical evaluation. One interesting treatment approach may be interference with interleukin-6 (IL-6) signaling to modulate B-cell immunity and donor-specific antibody (DSA) production. Currently, a large phase III randomized controlled trial is underway to clarify the safety and efficacy of clazakizumab, a high-affinity anti-IL-6 antibody, in chronic AMR. A prevention/treatment strategy may be costimulation blockade using belatacept to interfere with germinal center responses and DSA formation. In a recent uncontrolled study, belatacept conversion was shown to stabilize renal function and dampen AMR activity. Moreover, preliminary clinical results suggest efficacy of CD38 antibodies to deplete plasma and natural killer cells to treat AMR, with anecdotal reports demonstrating at least transient resolution of active rejection. There are promising concepts on the horizon for the prevention and treatment of chronic AMR. The design of adequately powered placebo-controlled trials to clarify the safety and efficacy of such new therapies, however, remains a big challenge, and will rely on the definition of precise surrogate endpoints predicting long-term allograft survival. Mapping the natural history of AMR would greatly help the understanding of who would derive benefits from treatment.

Predicting clinical response to costimulation blockade in autoimmunity.

Curbing unwanted T cell responses by costimulation blockade has been a recognised immunosuppressive strategy for the last 15 years. However, our understanding of how best to deploy this intervention is still evolving. A key challenge has been the heterogeneity in the clinical response to costimulation blockade, and an inability to predict which individuals are likely to benefit most. Here, we discuss our recent findings based on the use of costimulation blockade in people with type 1 diabetes (T1D) and place them in the context of the current literature. We discuss how profiling follicular helper T cells (Tfh) in pre-treatment blood samples may have value in predicting which individuals are likely to benefit from costimulation blockade drugs such as abatacept.

Abstract 16582: Costimulation Blockade With Anti Cd80 and Cd86 Monoclonal Antibodies Prolonged Graft Survival in Allogeneic Ips Cell-Derived Cardiomyocyte Patch Transplantation

Introduction: How to avoid immune rejection after transplantation should be elucidated for confirmed efficacy in clinical application of allogenic induced pluripotent stem cell derived cardiomyocyte patch. The blockade of CD28-CD80/86 costimulation is known to induce T cell anergy and immune tolerance by the recruitment of regulatory T cells (T reg) and prolong graft survival in organ transplantation. Hypothesis: We hypothesized that the blockade of CD28-CD80/CD86 costimulatory pathway by anti-CD80/CD86 monoclonal antibodies (mAbs) induces T cell anergy in vivo and suppresses immune rejection in allogeneic iPSC-CMs subcutaneous transplantation mice model. Methods: T cell anergy was induced in vitro by co-culture of Balb/c and 25Gy irradiated C57BL/6 splenocytes with anti-CD80/86 mAbs for 5 days. The inhibitory effect of anergic cells was evaluated by mixed lymphocyte reaction (MLR) in C57BL/6 and Balb/c splenocytes with anergic cells. IFN-g in MLR supernatants was measured by ELISAs to assess immune response. C57BL/6 iPSC-CMs expressing luciferase were subcutaneously transplanted into Balb/c. Anti-CD80/CD86 mAbs were injected intraperitoneally 250μg/dose on day 0, 1, and 2 (treated mice, n=6). In control mice (n=6), equivalent volume of saline was injected. To evaluate iPSC-CMs graft survival, photon counts of iPSC-CMs were measured by bioluminescence imaging system (BLI). Cells of harvested grafts were analyzed by immunofluorescence staining (IF). Results: On ELISAs, IFN-g in MLR supernatants co-cultured with anergic cells was significantly lower as compared with those without anergic cells (12.3±14.4 vs. 251±80.2 pg/ml, p=0.007). The ability to suppress the alloresponses was dose-dependent. BLI showed that photon counts on day 14 in treated mice were significantly higher than in control mice (7397±2651 vs. 2703±1271, p=0.003). IF showed a siginificantly higher ratio of CD4 and Foxp3 double positive cells to CD4-postive cells in the grafts on day 7 in treated mice as compared with in control mice (23±3% vs. 9±3%, p=0.009) Conclusions: The blockade of CD28-CD80/CD86 costimulatory pathway might suppress immune rejection in allogeneic iPSC-CMs transplantation mice model and T reg might involve this immune tolerance.

Potent Interaction between CMV Reactivation and Gvhd: Immunologic Evidence for Blunting of CMV-Driven Immune Reconstitution in the Setting of Gvhd

Allogeneic hematopoietic cell transplantation (HCT) may be curative for patients with marrow and immune disorders, but graft-vs-host-disease (aGVHD) and infections cause significant morbidity and non-relapse mortality. We have conducted a multicenter, double blind, placebo-controlled phase II trial of costimulation blockade with abatacept (Aba) combined with standard GVHD prophylaxis with a calcineurin inhibitor and methotrexate (CNI + MTX) following HLA matched unrelated donor transplant (n=142). In order to assess the effects of Aba on immune reconstitution, and to assess whether this reconstitution is influenced during CMV reactivation, we longitudinally evaluated post-transplant whole blood samples with multiparameter flow cytometry using markers for CD3, CD4, CD8, CD197 and CD45RA to measure reconstitution of CD4 and CD8 T cell populations and their respective memory subsets over time. Results: We observe that post-transplant CMV reactivation induces a marked expansion of CD8 effector memory (EM) cells, which is similar in magnitude for Aba vs placebo patients. We found that development of moderate (gr 2-4) or severe (gr 3-4) GVHD was not associated with an increased frequency of CMV reactivation, but patients with moderate GVHD showed a blunted expansion of CD8 EM cells compared to those without GVHD, and CD8 EM expansion was essentially absent among CMV reactivating patients with severe aGVHD. Clinical correlates will be presented. Conclusions: Our results suggest that adding abatacept to CNI/MTX does not materially affect reconstitution of T cell immunity in the presence or absence of CMV reactivation, but aGVHD remains a major driver of compromised immune recovery after HCT. Disclosures Watkins: Bristol Myers Squibb: Research Funding. Qayed:Mesoblast: Consultancy; Novartis: Consultancy. Horan:Bristol Myers Squib: Honoraria, Research Funding. Kean:gilead: Research Funding; bluebird bio: Research Funding; fortyseven: Consultancy; magenta: Research Funding; regeneron: Research Funding; hifibio: Consultancy; kymab: Consultancy; Bristol Meyers Squibb: Research Funding; novartis: Consultancy. Langston:Kadmon Corporation: Research Funding; Bristol Myers Squib: Research Funding; Incyte: Research Funding; Chimerix: Research Funding; Takeda: Research Funding; Astellas Pharmaceuticals: Research Funding; Jazz Pharmaceuticals: Research Funding.

Unraveling the Crucial Roles of FoxP3+ Regulatory T Cells in Vascularized Composite Allograft Tolerance Induction and Maintenance.

The role of regulatory T cells (Treg) in tolerance induction of vascularized composite allotransplantation (VCA) remains unclear. This study was designed to examine characteristics of Treg after VCA and their capacity to rescue allografts from rejection. Osteomyocutaneous allografts were transplanted from Balb/c to C57BL/6 mice. All mice received costimulatory blockade and a short course of rapamycin. To elucidate the role of Treg for tolerance induction, Treg depletion was performed at postoperative day (POD) 0, 30, or 90. To assess capacity of Treg to rescue allografts from rejection, an injection of 2 × 106 Treg isolated from tolerant mice was applied. Eighty percent of VCA recipient mice using costimulatory blockade and rapamycin regimen developed tolerance. The tolerant recipients had a higher ratio of circulating Treg to effector T cells and elevated interleukin-10 at POD 30. A significantly higher rejection rate was observed when Treg were depleted at POD 30. But Treg depletion at POD 90 had no effect on tolerance. Treg from tolerant recipients showed stronger suppressive potential and the ability to rescue allografts from rejection. Furthermore, transplanted Treg-containing skin grafts from tolerant mice delayed rejection elicited by adoptively transferred effector T cells to Rag2-/- mice. Circulating Treg are crucial for inducing VCA tolerance in the early posttransplant phase, and allograft-residing Treg may maintain tolerance. Treg may, therefore, serve as a potential cellular therapeutic to improve VCA outcomes.