Treatment Of cGVHD Research Articles

Ibrutinib as a Secondary Treatment for Steroid-Refractory or Steroid-Dependent Chronic Graft-Versus-Host Disease: A Case Series of 11 Patients During the COVID-19 Era.

Ibrutinib, a Bruton's tyrosine kinase inhibitor, has recently become available for treating chronic graft-versus-host disease (cGVHD). Because the pivotal clinical trials for this approach were conducted before the COVID-19 pandemic, evidence regarding ibrutinib treatment for cGVHD in the COVID-19 era is insufficient. We assessed the safety and efficacy of ibrutinib treatment in a real-world setting by retrospectively analyzing the outcomes of 11 patients with steroid-refractory and steroid-dependent cGVHD who were treated with ibrutinib between November 2021 and April 2024 at our hospital. The best overall response rate was 63.6%, and the steroid dose was successfully reduced in seven of the patients. Ibrutinib improved hemolytic anemia and serositis in some of the patients. The most common adverse events were infections and bleeding. Four of the patients contracted COVID-19 during the study period, of whom one discontinued the treatment because of severe COVID-19 pneumonia. Ibrutinib was effective for treating cGVHD, particularly when B cells were involved in its pathogenesis, and decreased the steroid dose needed for treatment in a real-world setting. Patients should be carefully monitored and treated for adverse events during this treatment, particularly bleeding and any complications associated with COVID-19 infection.

Healthcare Resource Utilization and Associated Costs in Patients With Chronic Graft-Versus-Host Disease Post-Allogeneic Hematopoietic Stem Cell Transplantation in England

Limited evidence suggests chronic graft-versus-host disease (cGvHD) following allogeneic hematopoietic stem cell transplantation (allo-HSCT) increases healthcare resource utilization (HCRU) and costs. However, this burden has not been well characterized in England. This study assesses secondary care HCRU and costs for patients following allo HSCT in England with cGvHD and patients who did not develop graft-versus-host disease (GvHD). Further stratification was performed among patients who did or did not subsequently receive high-cost therapies for the treatment of cGvHD. This descriptive, retrospective cohort study used Hospital Episode Statistics (HES) data from April 2017 to March 2022. HES data captures information on reimbursed diagnoses and procedures from all National Health Service (NHS) secondary care admissions and attendances in England. High-cost drugs as defined by NHS England are recorded in HES, these drugs and other procedures including plasma exchange, were used to identify patients with cGvHD who were in receipt of high-cost therapies. HCRU and costs were described for patients with cGvHD following allo-HSCT (n = 721) and were matched with patients with no evidence of GvHD following allo-HSCT (n = 718). HCRU and costs were also described for the subset of patients with cGvHD (n = 198) following receipt of high-cost therapies and patients with cGvHD prior to or without such therapies (n = 523). A higher proportion of patients with cGvHD had at least one inpatient or intensive care unit (ICU) admission or emergency care attendance than patients without GvHD (inpatient: 74.6% versus 66.6%; emergency care: 39.3% versus 30.5%; ICU: 7.4% versus 4.7%; respectively); whilst the proportion of patients with an outpatient attendance were similar for both groups (outpatient: 80.3% versus 84.1%; respectively). The cost across all secondary care settings was higher for patients with cGvHD than patients without GvHD, with a mean cost of inpatient admissions of £17,339 ppy for those with cGvHD versus £8548 ppy in patients without GvHD. A higher proportion of patients who received high-cost therapies for the treatment of cGvHD had at least one secondary care admission or attendance, than patients who did not (inpatient: 85.4% versus 66.4%; ICU: 7.1% versus 5.4%; outpatient: 87.9% versus 76.7%; emergency care: 44.4% versus 36.5%; respectively). Patients who were treated with high-cost therapies for the treatment of cGvHD had a greater mean number (14.6 versus 8.2 ppy, respectively) for all-cause inpatient admissions after treatment than patients who did not. In all secondary care settings, the total cost ppy was higher for patients who received high-cost therapies for the treatment of cGvHD, than for those who did not. Patients who were treated with high-cost therapies for the treatment of cGvHD had a greater mean cost (£21,137 versus £15,956 ppy, respectively) for all-cause inpatient admissions than patients who did not. This study demonstrates that cGvHD and the use of associated high-cost therapies impacts healthcare activity and costs across various secondary care settings in England more than patients without GvHD and patients with cGvHD who received no high-cost therapies.

Characterization of ibrutinib's effects on the morphology, proliferation, phenotype, viability, and anti-inflammatory potential of adipose-derived mesenchymal stromal cells

Ibrutinib (IB) is a tyrosine kinase inhibitor (TKI) that has immunomodulatory action and can be used as second-line therapy for steroid-refractory or steroid-resistant chronic Graft versus Host Disease (cGVHD). Mesenchymal stromal cells (MSCs) are distributed throughout the body and their infusion has also been explored as a second-line therapeutic alternative for the treatment of cGVHD. Considering the currently unknown effects of IB on endogenous MSCs, as well as the possible combined use of IB and MSCs for cGVHD, we investigated whether adipose tissue-derived MSCs present IB-targets, as well as the consequences of treating MSCs with this drug, regarding cell viability, proliferation, phenotype, and anti-inflammatory potential. Interestingly, we show for the first time that MSCs express several IB target genes. Also of note, the treatment of such cells with this TKI elevated the levels of CD90 and CD105 surface proteins, as well as VCAM-1. Furthermore, IB-treated MSCs presented increased mRNA expression of the anti-inflammatory genes PD-L1, TSG-6, and IL-10. However, continued exposure to IB, even at low doses, compromised the viability of MSCs. These data indicate that the use of IB can stimulate an anti-inflammatory profile in MSCs, but also that a continued exposure to IB can compromise MSC viability over time.

Updates in chronic graft-versus-host disease: novel treatments and best practices in the current era.

Chronic graft-versus-host disease (cGVHD) is a serious complication of allogeneic hematopoietic cell transplant. The development of cGVHD involves a complex, multistep process that is characterized by early inflammation and tissue injury, followed by chronic inflammation, aberrant tissue repair, and fibrosis. Systemic corticosteroids remain the first line of treatment for cGVHD. New treatments for patients with cGVHD for whom treatment has failed or who develop steroid-dependent cGVHD are now available; these include ibrutinib, ruxolitinib, and belumosudil. Treatment selection may be based on the patient's individual needs, graft-versus-host disease organ involvement, and comorbidities. However, as therapeutic options for patients without a treatment response or with only a partial response remain an unmet need, new agents are under investigation. Furthermore, patients with cGVHD can develop multiorgan involvement and frequently require specialized care. A multidisciplinary team approach that focuses on the individual's needs and quality of life is strongly encouraged.

Efficacy and safety of ruxolitinib in the treatment of chronic graft-versus-host disease: a retrospective analysis

Steroid-refractory chronic graft-versus-host disease (cGvHD) is associated with significant morbidity and mortality, with ruxolitinib being the first drug approved for its treatment. We retrospectively analyzed the safety and efficacy of ruxolitinib for treatment of cGvHD at our center between 07/2015 and 12/2022 and identified 48 patients receiving ruxolitinib as second (18/48) or advanced (30/48) treatment line. Ruxolitinib was started on median day 340 (range 119–595) after cGvHD onset; median duration of administration was 176 (range, 79–294) days with 16/48 patients continuing treatment at last follow-up. National Institutes of Health organ grading and the intensity of immunosuppression were assessed at the start of ruxolitinib treatment and repeated after 1, 3, 6, and 12 months. Response assessment was terminated at the start of any additional new immunosuppressant treatment. The median time of follow-up was 582 (range, 104–1161) days. At the primary analysis after six months on ruxolitinib treatment, the overall response rate was 33%, and failure-free survival was 58%. Infectious adverse events ≥ CTCAE grade III were observed in 10/48 patients. The response rate was not associated with the severity of cGvHD, number of previous treatment lines, or number of additional agents combined with ruxolitinib applying a univariate regression model. At the time of the 12-month follow-up, four patients experienced recurrence of the underlying malignancy and two patients had experienced non-relapse-related mortality. Overall, ruxolitinib was relatively well-tolerated and showed outcomes comparable to the REACH3 trial in a heavily pretreated patient population.

Treatment of newly diagnosed moderate or severe chronic graft-versus-host disease with prednisone and everolimus (PredEver first): a prospective multicenter phase IIA study

Although most patients with chronic graft-versus-host disease (cGVHD) show initial response to first-line therapy, long-term clinically meaningful success of first-line treatment remains rare. In a prospective multicentre phase II trial in 6 German centers, patients with newly diagnosed moderate or severe cGVHD received prednisone and everolimus for 12 months followed by a 1-year follow-up period. Primary endpoint was treatment success (TS) at 6 months defined as patient being alive, achieving PR or CR of cGVHD, having no relapse of underlying disease and requiring no secondary treatment for cGVHD. Of the 34 patients evaluable for efficacy, 19 (56%) had TS at 6 months with 22 and 52% of the patients in a CR and PR respectively. Overall 30 patients (88%) had a CR or PR as best response, nearly all responses (29/30) occurring within the first 6 weeks of treatment. The cumulative incidence of treatment failure at 1 year was 63%, corresponding to 37% TS. Predefined safety endpoint (thrombotic microangiopathy, pneumonitis, and avascular necrosis) were not observed in any patient. Addition of everolimus to prednisolone is well tolerated and may improve long-term treatment success. Larger studies are necessary to ascertain the possible role of everolimus in first-line treatment of cGVHD.

Cost-Effectiveness of Extracorporeal Photopheresis in Patients With Chronic Graft-vs-Host Disease

Background: The mainstay first-line therapy for chronic graft-vs-host disease (cGVHD) is corticosteroids; however, for steroid-refractory patients, there is a distinct lack of cost-effective or efficacious treatment. The aim of this study was to assess the cost-effectiveness of extracorporeal photopheresis (ECP) compared with standard-of-care therapies for the treatment of cGVHD in Australia. The study formed part of an application to the Australian Government to reimburse ECP for these patients. Methods: A cost-utility analysis was conducted comparing ECP to standard of care, which modeled the response to treatment and disease progression of cGVHD patients in Australia. Mycophenolate, tacrolimus, and cyclosporin comprised second-line standard of care based on a survey of Australian clinicians. Health states in the model included treatment response, disease progression, and death. Transition probabilities were obtained from Australian-specific registry data and randomized controlled evidence. Quality-of-life values were applied based on treatment response. The analysis considered costs of second-line treatment and disease management including immunosuppressants, hospitalizations and subsequent therapy. Disease-specific mortality was calculated for treatment response and progression. Results: Over a 10-year time horizon, ECP resulted in an average cost reduction of $23 999 and an incremental improvement of 1.10 quality-adjusted life-years per patient compared with standard of care. The sensitivity analysis demonstrated robustness over a range of plausible scenarios. Conclusion: This analysis demonstrates that ECP improves quality of life, minimizes the harms associated with immunosuppressant therapy, and is a highly cost-effective option for steroid-refractory cGVHD patients in Australia. Based in part on this analysis, ECP was listed on the Medicare Benefits Schedule for public reimbursement.

Cost-Effectiveness of Extracorporeal Photopheresis in Patients With Chronic Graft-vs-Host Disease.

Background: The mainstay first-line therapy for chronic graft-vs-host disease (cGVHD) is corticosteroids; however, for steroid-refractory patients, there is a distinct lack of cost-effective or efficacious treatment. The aim of this study was to assess the cost-effectiveness of extracorporeal photopheresis (ECP) compared with standard-of-care therapies for the treatment of cGVHD in Australia. The study formed part of an application to the Australian Government to reimburse ECP for these patients. Methods: A cost-utility analysis was conducted comparing ECP to standard of care, which modeled the response to treatment and disease progression of cGVHD patients in Australia. Mycophenolate, tacrolimus, and cyclosporin comprised second-line standard of care based on a survey of Australian clinicians. Health states in the model included treatment response, disease progression, and death. Transition probabilities were obtained from Australian-specific registry data and randomized controlled evidence. Quality-of-life values were applied based on treatment response. The analysis considered costs of second-line treatment and disease management including immunosuppressants, hospitalizations and subsequent therapy. Disease-specific mortality was calculated for treatment response and progression. Results: Over a 10-year time horizon, ECP resulted in an average cost reduction of $23 999 and an incremental improvement of 1.10 quality-adjusted life-years per patient compared with standard of care. The sensitivity analysis demonstrated robustness over a range of plausible scenarios. Conclusion: This analysis demonstrates that ECP improves quality of life, minimizes the harms associated with immunosuppressant therapy, and is a highly cost-effective option for steroid-refractory cGVHD patients in Australia. Based in part on this analysis, ECP was listed on the Medicare Benefits Schedule for public reimbursement.

Recent advances in prevention and treatment of chronic graft-versus-host disease after allogeneic hematopoietic stem cell transplantation

Chronic graft-versus-host disease (cGVHD) negatively impacts long-term survival and quality of life (QOL) after allogeneic hematopoietic cell transplantation. Corticosteroids are the first-line treatment for cGVHD, but approximately 30% to 70% of patients develop steroid-refractory cGVHD (SR-cGVHD), which has an extremely poor prognosis. The pathophysiology of cGVHD is more complicated than acute GVHD, but recent advances using murine models in conjunction with human studies indicate three major phases: 1) acute inflammation, 2) chronic inflammation with loss of immune tolerance, and 3) disrupted target organ homeostasis and fibrosis. Strategies that help prevent cGVHD include optimal donor selection and choice of conditioning regimen as well as pharmacologic and graft manipulation strategies. The key cellular mediators of SR-cGVHD are T cells, B cells, antigen-presenting cells, and fibroblasts. T cells and B cells are now targetable with the inhibitors ibrutinib and ruxolitinib, respectively. Recently, promising results have been obtained by modulating pathologic T cell responses with Rock2 inhibitors and targeting fibrosis with CSF-1R inhibitors. To optimize the use of these medications, a better understanding of the biological and target organ-specific mechanisms of cGVHD is needed. Here we review recent advances in cGVHD pathogenesis and discuss how best to implement recently approved biology-driven treatments for cGVHD.

CSF-1R inhibitor PLX3397 attenuates peripheral and brain chronic GVHD and improves functional outcomes in mice

Graft-versus-host disease (GVHD) is a serious complication of otherwise curative allogeneic haematopoietic stem cell transplants. Chronic GVHD induces pathological changes in peripheral organs as well as the brain and is a frequent cause of late morbidity and death after bone-marrow transplantation. In the periphery, bone-marrow-derived macrophages are key drivers of pathology, but recent evidence suggests that these cells also infiltrate into cGVHD-affected brains. Microglia are also persistently activated in the cGVHD-affected brain. To understand the involvement of these myeloid cell populations in the development and/or progression of cGVHD pathology, we here utilized the blood–brain-barrier permeable colony stimulating factor-1 receptor (CSF-1R) inhibitor PLX3397 (pexidartinib) at varying doses to pharmacologically deplete both cell types. We demonstrate that PLX3397 treatment during the development of cGVHD (i.e., 30 days post-transplant) improves disease symptoms, reducing both the clinical scores and histopathology of multiple cGVHD target organs, including the sequestration of T cells in cGVHD-affected skin tissue. Cognitive impairments associated with cGVHD and neuroinflammation were also attenuated by PLX3397 treatment. PLX3397 treatment prior to the onset of cGVHD (i.e., immediately post-transplant) did not change in clinical scores or histopathology. Overall, our data demonstrate significant benefits of using PLX3397 for the treatment of cGVHD and associated organ pathologies in both the periphery and brain, highlighting the therapeutic potential of pexidartinib for this condition.

Chronic GVHD: review advances in prevention, novel endpoints, and targeted strategies.

Allogeneic hematopoietic cell transplantation (allo-HCT) is a curative therapy for many malignant and non-malignant hematologic disorders. Chronic graft-versus-host (cGVHD) disease remains a significant hurdle for long-term survival in patients post allo-HCT, and it remains the leading cause of late non-relapse mortality. The risk factors for development of cGVHD include degree of human leukocyte antigen (HLA) disparity, increasing recipient age, use of peripheral blood stem cells as a source, myeloablative conditioning regimens, prior acute GVHD (aGVHD), and female donor to male recipient. Our biological understanding of cGVHD is mostly derived from transplantation mouse models and patient data. There are three distinct phases in the development of cGVHD. Approaches to prevent GVHD include pharmacologic strategies such as calcineurin inhibitors (cyclosporine, tacrolimus) combined with methotrexate or mTOR inhibitors (sirolimus), and IMP dehydrogenase inhibitors (mycophenolate mofetil). Increasingly, posttransplant cyclophosphamide is emerging as a promising strategy for GVCHD prevention especially in a setting of reduced intensity conditioning. Other approaches include serotherapy (ATG, Campath) and graft manipulation strategies. A significant obstacle to evaluating the response of novel GVHD-directed therapies has been standardized response assessments. This has functioned as a barrier to designing and interpreting clinical trials that are structured around the treatment of cGVHD. Novel endpoints including failure-free survival, Graft-versus-host disease-free, relapse-free survival (GRFS), and current GVHD-free, relapse-free survival (CGRFS) may create a clearer picture for post-HCT outcomes. Targeted therapies including Bruton's tyrosine kinase inhibition, JAK1/2 inhibition, and ROCK2 inhibitors have improved cGVHD therapy, especially in the steroid refractory setting. Continued improvement in prophylactic strategies for cGVHD, identification of accurate cGVHD treatment endpoints, and access to novel therapeutic agents are expected to improve cGVHD outcomes.

CSF1R inhibition promotes neuroinflammation and behavioral deficits during graft-versus-host disease in mice

AbstractChronic graft-versus-host disease (cGVHD) remains a significant complication of allogeneic hematopoietic stem cell transplantation. Central nervous system (CNS) involvement is becoming increasingly recognized, in which brain-infiltrating donor major histocompatibility complex (MHC) class II+ bone marrow–derived macrophages (BMDM) drive pathology. BMDM are also mediators of cutaneous and pulmonary cGVHD, and clinical trials assessing the efficacy of antibody blockade of colony-stimulating factor 1 receptor (CSF1R) to deplete macrophages are promising. We hypothesized that CSF1R antibody blockade may also be a useful strategy to prevent/treat CNS cGVHD. Increased blood-brain barrier permeability during acute GVHD (aGVHD) facilitated CNS antibody access and microglia depletion by anti-CSF1R treatment. However, CSF1R blockade early after transplant unexpectedly exacerbated aGVHD neuroinflammation. In established cGVHD, vascular changes and anti-CSF1R efficacy were more limited. Anti-CSF1R–treated mice retained donor BMDM, activated microglia, CD8+ and CD4+ T cells, and local cytokine expression in the brain. These findings were recapitulated in GVHD recipients, in which CSF1R was conditionally depleted in donor CX3CR1+ BMDM. Notably, inhibition of CSF1R signaling after transplant failed to reverse GVHD-induced behavioral changes. Moreover, we observed aberrant behavior in non-GVHD control recipients administered anti-CSF1R blocking antibody and naïve mice lacking CSF1R in CX3CR1+ cells, revealing a novel role for homeostatic microglia and indicating that ongoing clinical trials of CSF1R inhibition should assess neurological adverse events in patients. In contrast, transfer of Ifngr–/– grafts could reduce MHC class II+ BMDM infiltration, resulting in improved neurocognitive function. Our findings highlight unexpected neurological immune toxicity during CSF1R blockade and provide alternative targets for the treatment of cGVHD within the CNS.

Effect of HLA-DPB1 Mismatch on Incidence and Severity of Chronic Graft-Versus-Host Disease Following Haploidentical Transplantation

Chronic graft-versus-host disease (cGVHD) is an important cause of late morbidity and mortality after haploidentical donor transplantation (HIDT), occurring in up to one-third of patients. The treatment of cGVHD can continue for years and can significantly impact quality-of-life in long-term transplant survivors. Therefore, it is important to identify risk factors that are associated with the incidence and severity of cGVHD. To this end, we investigated 335 consecutive recipients of HIDT utilizing post-transplant cyclophosphamide (PTCy) from 2005-2021, with a particular focus on class II HLA mismatches given prior published associations of HLA-DR and -DP mismatches with HIDT outcomes. HIDT recipients had a median age of 50 (19, 80). PBSC was the graft source in 81%, and conditioning was myeloablative in 49%. Whereas the incidence and severity of cGVHD did not differ between transplants with or without an HLA-DRB1 or -DQB1 allele-level mismatch, the presence of an HLA-DPB1 mismatch was associated with higher 1-yr incidence of all-grade (32% vs. 22%, p=0.07) and moderate-to-severe (22% vs. 12%, p=0.011) cGVHD (see figure). The effect of HLA-DPB1 mismatch on cGVHD was independent of permissiveness by the T cell epitope (TCE) algorithm or the vector of the mismatch (bidirectional, GVH- or HVG-only). In multivariate analysis, an HLA-DPB1 mismatch was associated with an increased risk of moderate-to-severe cGVHD (HR 2.05, p=0.044), while controlling for other covariates (CMV status, year of transplant). Other class II loci (HLA-DRB1 and HLA-DQB1) mismatching were not associated with chronic GVHD incidence in MVA. In summary, the presence of an HLA-DPB1 mismatch is associated with an increased incidence of moderate-to-severe cGVHD following PTCy-based HIDT. This effect, of course, must be balanced by the protective effect of a TCE nonpermissive HLA-DPB1 mismatch on relapse and survival, when selecting the optimal haploidentical donor.

CSF1R Inhibition Promotes Neuroinflammation and Behavioural Deficits during Graft-Versus-Host Disease in Mice

Chronic graft-versus-host disease (cGVHD) remains a significant complication of allogeneic hematopoietic stem cell transplantation. Central nervous system (CNS) involvement is becoming increasingly recognised, where brain-infiltrating donor MHC class II + bone marrow-derived macrophages (BMDM) are implicated mediators of pathology. Colony-stimulating factor 1 receptor (CSF1R)-dependent BMDM are established mediators of cutaneous and pulmonary cGVHD, and clinical trials assessing the efficacy of CSF1R antibody blockade to deplete macrophages are promising. We hypothesized that CSF1R antibody blockade may also be a useful strategy to prevent/treat CNS cGVHD. We leveraged studies in acute GVHD (aGVHD) to determine the feasibility of an antibody-based approach for depleting brain microglia and macrophages. Moreover, we queried whether CSF1R blockade may be a viable therapy for CNS aGVHD, which has been shown to be mediated by host microglia. Increased blood-brain barrier permeability during aGVHD facilitated CNS antibody access. However, CSF1R blockade early post-transplant induced region-specific microglia depletion and was associated with unexpected exacerbation of aGVHD neuroinflammation, including increased T cell and inflammatory monocyte infiltration, and whole brain cytokine levels. In established cGVHD, vascular changes and anti-CSF1R efficacy was more limited. Anti-CSF1R-treated mice retained donor BMDM, activated microglia, CD8 + and CD4 + T cells, and local cytokine expression in the brain. These findings were recapitulated in GVHD recipients where CSF1R was conditionally depleted in donor CX3CR1 + BMDM. Notably, inhibition of CSF1R signalling post-transplant failed to reverse either acute or chronic GVHD-induced behavioral changes. Taken together, these data suggest that ongoing clinical trials of CSF1R inhibition should assess neurological adverse events in patients. In contrast to the lack of efficacy with CSF1R inhibition, transfer of Ifngr-/- grafts reduced MHC class II + BMDM infiltration, resulting in improved behaviour. Furthermore, single nuclei RNA sequencing of brains from mice receiving wild-type or Ifngr-/- allografts revealed a disease-associated interferon-inducible mature oligodendrocyte population in the cGVHD brain. This was markedly reduced in recipients of Ifngr-/- grafts, indicating that targeting this axis can promote brain repair beyond the myeloid compartment during cGVHD. In summary, our findings reveal unexpected neurological immune toxicity during CSF1R blockade and highlight the IFNGR pathway as an alternative target for the treatment of cGVHD within the CNS.

Axatilimab Ameliorates Inflammation and Fibrosis By Targeting Macrophages in a Preclinical Model of Chronic GVHD

Introduction: Chronic graft-versus-host disease (cGVHD) is an immune-mediated serious and life-threatening complication after allogeneic hematopoietic stem cell transplantation, occurring in 30%-70% of patients. The pathophysiology of cGVHD may involve inflammation, cell-mediated immunity, humoral immunity, and fibrosis. Axatilimab is a high-affinity humanized IgG4 (kappa light chain) monoclonal antibody targeting colony-stimulating factor 1 receptor (CSF-1R) that is under investigation for the treatment of cGVHD. CSF-1R is a member of the receptor protein tyrosine kinase family of growth factor receptors and is expressed on monocyte precursors, monocytes, and macrophages. Signaling by colony-stimulating factor 1 (CSF-1) or interleukin-34 (IL-34) through CSF-1R has been shown to be the primary regulatory pathway for determining development and differentiation of the mononuclear phagocytic cell lineage. Recent work has established a role for monocyte-derived macrophages as the pathogenic population of cells that establish the fibrotic niche and drive the fibrotic process. Upon entry of monocytes into tissues, the tissue microenvironment regulates their differentiation, resulting in a heterogeneous pool of cells ranging from pro-inflammatory (often referred to as M1 macrophages) to immune suppressive/wound healing (M2 macrophages). An imbalance in the number and activity of these macrophage populations is believed to result in the excess production of fibrotic growth factors, such as transforming growth factor-β and platelet-derived growth factor, leading to uncontrolled fibroblast activation and subsequent fibrosis. Blocking CSF-1R signaling with axatilimab is expected to reduce the circulating levels of pathogenic monocyte-derived macrophage precursors and inhibit their activation in tissues, thereby providing an opportunity to therapeutically intervene in diseases such as cGVHD. The results described herein support this assertion by demonstrating the direct impact of axatilimab exposure to skewing differentiating monocytes away from a pro-inflammatory and pro-fibrotic phenotype. Methods: Primary human monocytes were differentiated in the presence or absence of ascending doses of axatilimab (0.07-7 μM) into distinct phenotypes: activated (M0), pro-inflammatory (M1-like), and suppressive (M2-like) macrophages. Supernatants and cell pellets were then assessed for secreted protein (Luminex Corporation) and gene expression (NanoString Technologies) profiles, respectively. Data were analyzed in R (version 4.1.1). Results: Gene expression profiles of pro-inflammatory cytokines and chemokines (eg, IL-1β, chemokine CC motif ligand 2 [CCL2], CCL7, CCL24, and CCL22) from primary differentiated macrophage subsets were dramatically reduced in the presence of ascending concentrations of axatilimab. Further, macrophages exposed to axatilimab while differentiated under pro-inflammatory conditions showed increased levels of c-MYC expression compared with control cultures. c-MYC is a transcription factor associated with suppressive M2-like macrophages, which suggests that axatilimab interferes with pro-inflammatory macrophage differentiation. Consistent with this, expression of M2-associated cytokines, including IL-10, IL-5, and IL-4, was enhanced in axatilimab-treated macrophage cultures. Conclusions: Blocking CSF-1R signaling in primary, differentiated macrophages dramatically alters the secretomes and gene expression profiles of these cells. Macrophages are key inducers of fibroblast growth factors, which in turn promote fibrotic disease. These data confirm that production of inflammatory and pro-fibrotic factors by pro-inflammatory macrophages is inhibited in the presence of axatilimab, thereby supporting the notion that axatilimab provides a unique opportunity to therapeutically intervene or prevent fibrosis in cGVHD.

Effective Prevention of Steroid-Requiring Chronic Graft-Vs.-Host Disease with B Cell Depletion: A Randomized, Placebo-Controlled Trial

Introduction: The pathophysiology of chronic graft-vs.-host disease (cGVHD) is complex, with many cell types playing critical roles. B cells are involved in cGVHD pathobiology, and strategies that eliminate or inhibit B cells (rituximab, ibrutinib) can be therapeutic in advanced disease. We previously demonstrated effective prevention of cGVHD in a phase II trial of B cell depletion with rituximab. Obinutuzumab (Obin, Genentech) is a 2 nd generation monoclonal anti-CD20 antibody with enhanced B cell depletion activity over rituximab. Here we present the results of a randomized trial of Obin for the prevention of cGVHD. Methods: We performed a randomized double-blind, placebo-controlled trial of B cell depletion using 4 doses of Obin to determine if steroid-requiring cGVHD (SR-cGVHD) could be prevented 12 months from HSCT (NCT# 02867384). Eligible subjects previously underwent myeloablative or reduced intensity 7-8/8 HLA-matched, related or unrelated HSCT using peripheral blood stem cells (PBSC) and tacrolimus-based primary GVHD prophylaxis without T cell depletion. Subjects without evidence of cGVHD, active Gr II-IV acute GVHD, or malignancy were enrolled between 60-90 days following HSCT. Eligibility included adequate hematopoiesis (ANC > 1000/mL, plt > 50 000/mL) and stable donor chimerism (> 80%). All subjects provided informed consent. Obin (1000 mg iv) or placebo was administered at 3, 6, 9 and 12 months after HSCT (+/- 10 days) with acetaminophen and diphenhydramine prior to each study drug dose. The primary endpoint was the cumulative incidence (CI) of SR-cGVHD, which was felt to be a more objective measurement of cGVHD severity than NIH cGVHD stage alone. Results: 181 subjects were enrolled (92 Obin, 89 placebo). 167 subjects who have been followed for >1 year from HSCT (84 Obin, 83 placebo) are included in this analysis. Baseline clinical characteristics, including age, recipient/donor gender, donor type/match, primary malignancy/DRI, conditioning intensity and HCT-CI, and prior Gr II-IV GVHD were balanced. The median number of study drug doses was equivalent (median 2.5 vs. 2 for Obin and placebo groups). For the entire cohort, the median follow-up time among survivors was 24 months (range 4, 74). The primary endpoint, the CI of SR-cGVHD at 12 months from HSCT, was significantly reduced with Obin - 11% (95% CI 5.8-20) in comparison with placebo - 38% (95% CI 28-49) (p=0.008, Fig 1). The corresponding rates of NIH Moderate-Severe cGVHD at 12 months were 20% vs. 35% (p=0.069). The rate of SR-cGVHD by 2 years remained lower in the Obin group (28% vs. 44%). In multivariable modeling, only study arm (Obin vs. placebo) was associated with SR-cGVHD (p=0.01). Obin was safe and well tolerated with only 5 Gr 3-4 infusion reactions (placebo - 0). There was more Gr 3-4 neutropenia (32.1% vs. 4.8%) but only 1 case of neutropenic fever in the Obin group. Neutropenia responded to G-CSF, administered at investigator discretion. There were 9 non-relapse related deaths in the Obin arm (COVID 3, infection 2, pulmonary 2, acute GVHD 1, unknown 1) compared with 3 in the placebo arm (unknown 2, pulmonary 1) (p=NS). There was no difference in the CI of relapse at 2 years (20% vs. 28%, p=NS). The SR-cGVHD-free, relapse-free survival in the Obin group was superior (CRFS, 43% vs. 30%, p=0.009, Fig 2) without a change in overall survival 82% vs. 86%, p=NS). CD19 + B cell depletion was complete in the Obin arm through 1 year (median cell count 0) while B cell recovery was normal in the placebo arm (p<0.0001 at 3, 6, 9, 12 months) and remained lower at 24 and 36 months (36 month median 55.5 vs. 484.4 cells/mL, p=0.02). Similar findings were noted for CD19 +CD27 + memory B cells and CD19 +CD27 - naïve B cells. No differences in T cells were noted, but NK cells were reduced in the Obin arm at late time points. H-Y antibody studies in F to M donor/recipient pairs are underway. Conclusions: In subjects undergoing well-matched PBSC HSCT using tacrolimus-based GVHD prophylaxis, a strategy of B cell depletion with 4 doses of Obin significantly reduces the requirement for corticosteroids for treatment of cGVHD and significantly increases CRFS. Obin is well tolerated and is associated only with transient neutropenia. Further studies may determine whether prolonged B cell depletion may be of greater utility, however this double-blind, placebo-controlled randomized trial supports practice changing cGVHD prophylaxis in tacrolimus-based HSCT.

Safety and Efficacy of Extracorporeal Photopheresis for Acute and Chronic Graft-Versus-Host-Disease

Introduction Graft-versus-host disease(GVHD) is a severe complication of allogeneic hematopoietic cell transplantation(alloHCT), directly linked to increased morbidity and mortality. Despite novel biologic agents under study or commercial use in this field, corticosteroids and conventional intensified immunosuppressive therapy remain the cornerstone of treatment. Extracorporeal photopheresis (ECP) has been used as an alternative treatment. We studied the safety and efficacy of ECP in a large real-world cohort of GVHD patients. Methods We enrolled consecutive patients, of our JACIE-accredited center, that received ECP post alloHCT over the last 2 decades for cGVHD and over the last decade for steroid-dependent or refractory grade II-IV aGVHD. All patients with unrelated or haploidentical donors received thymoglobulin (ATG) 5mg/kg as prophylaxis. GVHD prophylaxis included cyclosporine-methotrexate in myeloablative and cyclosporine-MMF in reduced toxicity/intensity regimens. Before ruxolitinib or ibrutinib availability, MMF, cyclosporine or ATG were commenced as second line treatment in steroid-refractory patients, depending on previous prophylaxis. ECP was mostly administered as third line treatment for cGVHD and after assessment of response of 5 days of steroid treatment according to our protocol: 1 session/week for the 1 st month for cGVHD and 2sessions/week for aGVHD, 1 session/2 weeks for 3 months, evaluation of response and 1 session/month for 6 months. Before Ruxolitinib's availability, ATG was commenced simultaneously with ECP initiation in steroid-refractory aGVHD patients. Results We studied 112 patients with moderate or severe cGVHD. Only 13 received 4 or less ECP sessions because of severe GVHD-related morbidity and were excluded from further analysis. Median ECP sessions were 17(6-49). There were no ECP-related adverse events. 67 patients presented with cutaneous sclerosis manifestations, 73 mucocutaneous disease, 36 liver, 42 visceral and 27 lung involvement. ECP was commenced as second line in 35 patients. Ruxolitinib was administered in combination with ECP in 19 patients, while ibrutinib in 2. Bacterial infections were observed in 43 patients, viral in 38 and fungal in 11 patients. Only 19 patients did not show response. Significantly lower rates of response presented in patients with visceral involvement(p=0.037) and earlier post-transplant GVHD diagnosis (p=0.001). With a follow-up of 45.2 (5.6-345.1) months, 5-year CI of cGVHD-related mortality was 21.2% and was significantly reduced in patients with ECP response (p<0.001). 5-year OS was 65.3% and was independently associated with HLA matching (p=0.011), higher number of ECP sessions(p<0.001), later initiation of ECP (p=0.002), response to ECP (p=0.036) and no relapse(p=0.001). Regarding aGVHD, we studied 28 patients, aged 45 (18-67), with myeloablative (16), reduced toxicity (8) and intensity (4) conditioning, from sibling (4), matched (11) or one locus mismatched (12) volunteer unrelated and haploidentical (1) donors. Disease risk index was very high (1), high (11), intermediate (14) and low (2). aGVHD was observed at day +17(8-50). Skin, intestine and liver involvement was evident in 9 patients, skin and intestine in 13 and skin only in 6 patients. 13 patients were steroid-dependent and 15 steroid-refractory. ECP was commenced at day +18 (8-56) for 15 (4-20) sessions. The majority of patients (19/27) presented partial (7), very good (11) or complete (1) response. With 9.9 (1.7-113) months of follow-up, immunosuppression was reduced in 12/27 and ceased in 1 patient. Clinically significant bacterial infections were found in 19 patients, fungal in 3, CMV and EBV reactivation in 19 and 12 respectively and other viral in 6 patients. Cumulative incidence of aGVHD was 56.4 at 1-year.Five-year overall survival (OS) was 34%. Reduction of immunosuppression(p=0.026) and number of ECP sessions(p<0.001) were associated with improved OS, irrespectively of other factors. In particular, optimal OS was observed in patients that received more than 19 ECP sessions (Figure). Conclusion Our data confirm that ECP is safe and effective for GVHD. Even in the era of novel biologics, ECP should be considered early in the course of GVHD, before irreversible end organ damage has been established. Combination with other treatments and individualized treatment algorithms remain important unanswered questions.

Propensity Score Matching Analysis Comparing the Efficacy and Steroid Tapering Benefit of Extracorporeal Photopheresis to Best Available Therapy in Third-Line or Beyond Treatment for Chronic GvHD

Graft-versus host disease (GVHD) is one of the major limitations to allogeneic hematopoietic stem cell transplantation (HCT). Although corticosteroids with calcineurin inhibitors are established first line-therapy for chronic graft-versus-host disease (cGVHD), approximately one-half of cGVHD patients are refractory to corticosteroid therapy. The goal of the present study was to compare treatment outcomes of patients treated with extracorporeal photopheresis (ECP) and best available therapy (BAT) as third-line or beyond treatment for cGVHD. Using propensity score matching (PSM), treatment outcomes were compared between ECP-treated patients (n=74) and a historical cohort of cGVHD patients treated with BAT (n=132). By adjusting for unbalanced risk factors between the groups, including GVHD severity at the start of therapy, acute GVHD history, and baseline corticosteroid dose, 62 patients were balanced and selected for PSM. In the PSM cohort, the ECP group showed a 12-month failure-free survival (FFS) rate of 70.1% versus 32.5% in the BAT group (P < .0001; hazard rate [HR], .214), and 93.1% 12 months' overall survival (OS) rate of 93.1% versus 68.1% in the BAT group (P=.0249; HR, .3811); multivariate analysis confirmed ECP's superior FFS and OS compared with BAT. Generalized linear model analysis showed faster tapering of corticosteroids and higher rates of prednisone discontinuation in the ECP versus BAT PSM groups in the first 6 months. The ECP group also had a higher percentage of prednisone discontinuation, by 6% at month 0, by 14.9% at month 3, and by 22.5% at month 6. The current study demonstrates superior FFS, OS, and steroid tapering efficacy for ECP compared with BAT as third-line therapy or beyond in cGVHD patients.

Mesenchymal stromal cells ameliorate chronic GVHD by boosting thymic regeneration in a CCR9-dependent manner in mice.

Chronic graft-versus-host disease (cGVHD) is a major cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation. Mature donor T cells within the graft contribute to severe damage of thymic epithelial cells (TECs), which are known as key mediators in the continuum of acute GVHD (aGVHD) and cGVHD pathology. Mesenchymal stromal cells (MSCs) are reportedly effective in the prevention and treatment of cGVHD. In our previous pilot clinical trial in patients with refractory aGVHD, the incidence and severity of cGVHD were decreased, along with an increase in levels of blood signal joint T-cell receptor excision DNA circles after MSCs treatment, which indicated an improvement in thymus function of patients with GVHD, but the mechanisms leading to these effects remain unknown. Here, we show in a murine GVHD model that MSCs promoted the quantity and maturity of TECs as well as elevated the proportion of Aire-positive medullary TECs, improving both CD4+CD8+ double-positive thymocytes and thymic regulatory T cells, balancing the CD4:CD8 ratio in the blood. In addition, CCL25-CCR9 signaling axis was found to play an important role in guiding MSC homing to the thymus. These studies reveal mechanisms through which MSCs ameliorate cGVHD by boosting thymic regeneration and offer innovative strategies for improving thymus function in patients with GVHD.

Therapeutic Potential of a Senolytic Approach in a Murine Model of Chronic GVHD.

Graft-versus-host disease (GVHD) is a life-threatening systemic complication of allogeneic hematopoietic stem cell transplantation (HSCT) characterized by dysregulation of T and B cell activation and function, scleroderma-like features, and multi-organ pathology. The treatment of cGVHD is limited to the management of symptoms and long-term use of immunosuppressive therapy, which underscores the need for developing novel treatment approaches. Notably, there is a striking similarity between cytokines/chemokines responsible for multi-organ damage in cGVHD and pro-inflammatory factors, immune modulators, and growth factors secreted by senescent cells upon the acquisition of senescence-associated secretory phenotype (SASP). In this pilot study, we questioned the involvement of senescent cell-derived factors in the pathogenesis of cGVHD triggered upon allogeneic transplantation in an irradiated host. Using a murine model that recapitulates sclerodermatous cGVHD, we investigated the therapeutic efficacy of a senolytic combination of dasatinib and quercetin (DQ) administered after 10 days of allogeneic transplantation and given every 7 days for 35 days. Treatment with DQ resulted in a significant improvement in several physical and tissue-specific features, such as alopecia and earlobe thickness, associated with cGVHD pathogenesis in allograft recipients. DQ also mitigated cGVHD-associated changes in the peripheral T cell pool and serum levels of SASP-like cytokines, such as IL-4, IL-6 and IL-8Rα. Our results support the involvement of senescent cells in the pathogenesis of cGVHD and provide a rationale for the use of DQ, a clinically approved senolytic approach, as a potential therapeutic strategy.