Evidence Of Graft-versus-host Disease Research Articles

Post Transplant Cyclophosphamide (PTCY) in Peripheral Blood Stem Cell Transplant Results in Successful Engraftment and Substantially Low Frequency of Chronic Graft Versus Host Disease (GvHD), without Impaired Cellular Immune Reconstitution

The Post-Transplant Cyclophosphamide (PTCY) approach was first studied in the haploidentical transplant setting, and it resulted in high percentages of engraftment and reduced frequency of acute and mainly chronic GvHD as well as non-relapsed mortality. PTCY is now increasingly being used in HLA-partially and fully matched unrelated, as well as matched related donors. The aim of this study was to investigate the impact of PTCY on the incidence of chronic GvHD, disease relapse and infections because of possible cellular immune impairment. Fifty-one consecutive patients who underwent allogeneic hematopoietic stem cell transplant from an unrelated donor were studied. PTCΥ was administered on days +3 and +4 post-transplant at a dose of 50 mg/kg/day. Additional GvHD prophylaxis included tacrolimus and mycophenolate mofetil starting on day +5. In patients with no evidence of GvHD, immunosuppression was gradually tapered, and eventually stopped by day +90. We evaluated the safety and efficacy of PTCY, focusing on toxicity, engraftment, aGvHD and cGvHD frequency, relapse, non-relapse mortality, survival and immune reconstitution. We performed 51 unrelated donor transplants with PTCY from January 2021 to May 2023 in patients with a median age of 52 (18-69) years. The indication for transplant was AML/MDS (n=36), NHL/CLL, (n=2), pMF (n=2), CML (n=2) και ALL (n=9). Disease risk index (DRI) was intermediate for most patients (34/51), 7 had low and 10 had high DRI. Thirty-seven patients received myeloablative conditioning and 14 reduced-intensity. Overall, three patients were transplanted from a matched sibling, 36 from an 8/8 matched unrelated donor and 12 from an 7/8 HLA-matched unrelated donor. All patients received a peripheral blood graft with a median CD34+ cell dose of 6.97×10e6/kg (3.27-15.82). The most common toxicities observed were mucositis (n=34), diarrhea (n=40), vomit (n=12), liver toxicity (n=3) and acute kidney injury (n=4) - which in all cases were grade Ι-ΙΙ. Engraftment, defined as neutrophils >500/μl, was achieved in 46 cases in a median of 16 (13-27) days. Three patients died before day +28 due to NRM and two additional patients with myelofibrosis and MDS respectively had primary graft failure. The cumulative incidence of aGvHD, grade ΙΙ-IV at 100 days was 46.7% (95%CI: 34.1-63.8). However, aGvHD, grade ΙΙΙ-IV was only 3.9% (95%CI: 1.0-15.5). The cumulative incidence of cGvHD at 18 months was 13,4% (95%CI: 5.8-31.0), with only one case of extensive cGvHD. In a median follow-up of 14.5 (1-28) months, disease free (DFS) was 76,8% (95%CI: 65.5-90.0) at 18 months and overall survival (OS) was 76.3% (95%CI: 64.9-89.8) and 71.6% (95%CI: 58.6-88.3) respectively. There was no statistically significant difference in survival rates for patients who received an HLA mismatched and those who received an HLA-matched unrelated-donor transplant. cGvHD-Relapse-Free Survival was 63.5% (95%CI: 50.3-80.1). Two patients died of their disease and non-relapse mortality was 19.1%. Specifically, 6 patients died of Infections, 1 due to primary graft failure, 1 due to heart failure, and 1 due to Idiopathic Pulmonary Syndrome. Immune reconstitution at month 6 was 420 cells/ul (20-2225) and 179 (13-594) for CD8 and CD4 respectively. At month 12 CD8 and CD4 were 552 (55-3002) and 364 (60-787). 31 viral infections were documented in 28 patients (Respiratory viruses n=13, BKV n=11, CMV n=7). In regard with opportunistic infections, 2 patients had fungal infections and 1 patient atypical mycobacterium. We used PTCY for GvHD prophylaxis in mismatched, full-matched unrelated and matched sibling donor transplants with peripheral blood stem cell graft. This approach was well tolerated. Engraftment was documented in most patients, with a low incidence of grade III-IV aGvHD. Importantly cGvHD, which is the most important barrier in unrelated donor transplant, was substantially low with this method. Cellular immune reconstitution was adequate for CD8 but somewhat impaired for CD4. However, the incidence of viral and opportunistic infections was not increased. Overall, there are implications that PTCY may revert the negative impact of HLA mismatch on the outcomes of unrelated donor transplantation, while it is also a viable option for MUD and MSD transplant.

P583: PATIENTS AT HIGH RISK OF RELAPSE POST-TRANSPLANT: A PHASE 1 STUDY DESIGN WITH A NOVEL TREATMENT STRATEGY USING THE ESTIMAND FRAMEWORK

Background: The risk of disease relapse in patients (pts) allografted for acute myeloid leukemia (AML) is high and remains the main cause of transplant failure. The great majority of pts destined to relapse post-allogeneic stem cell transplant (SCT) will do so within the first 12 months and survival after early relapse is dismal. Therefore, strategies to prevent relapse are urgently required. A potent graft-versus-leukemia (GvL) effect is exerted after SCT, and strategies that may augment a GVL effect have the potential to reduce relapse risk post-SCT. Siremadlin (HDM201) is a novel investigational MDM2 inhibitor with single-agent (SA) anti-AML activity. MDM2 inhibitors, including siremadlin, possess potent immunomodulatory effects (IME) in murine solid tumor and AML models. The post-allogeneic SCT setting is ideal for investigating the IME of MDM2 inhibition on enhancing GvL effect. Here we present a phase Ib/II proof-of-concept study designed to assess safety and preliminary efficacy of a treatment strategy with siremadlin as monotherapy and, subsequently, in combination with donor lymphocyte infusions (DLIs) for AML in post-SCT setting using the estimand framework (ICH E9 (R1) addendum). Aims: The objective of the planned treatment strategy is to enhance GvL post-SCT by siremadlin as monotherapy and in combination with DLIs. Due to the risk of Graft-vs-Host Disease (GvHD) with DLI early after SCT, siremadlin will be given as priming monotherapy to prevent early relapse prior to starting siremadlin/DLI combination therapy. SA siremadlin maintenance after the combination therapy is intended to prolong and enhance the GvL reaction to eradicate residual leukemic blasts. Methods: The estimand framework mandates a careful definition of the experimental intervention according to the clinical question of interest. In this trial, the experimental intervention follows a treatment strategy: Patients (pts) with AML in complete remission (CR) post-SCT but at high risk for relapse based on pre-SCT risk factors will start with siremadlin priming monotherapy. Pts who tolerate the siremadlin dose with no evidence of GvHD are eligible to start siremadlin/DLI combination therapy. After combination therapy, pts may continue with SA siremadlin maintenance. Depending on the treatment journey, pts may receive only a part of the intervention’s sequence (e.g., pts in siremadlin priming monotherapy who are not eligible for DLI combination will continue the priming part). The clinical questions addressed in this trial are to determine the recommended dose (RD) of siremadlin across all parts of the treatment strategy and to assess preliminary efficacy regardless of the parts of the treatment strategy pts received. The doses of interest were preselected based on SA studies and in pts who received siremadlin after SCT. Results: The proposed approach is tailored to investigate safety and efficacy of the treatment strategy. The RD of siremadlin will be determined over 2 parts: A Bayesian logistic regression model will guide dose finding of siremadlin during priming (Cycle 1) and a Bayesian time-to-DLT model will assess the siremadlin dose for combination. Efficacy will be assessed by proportion of pts remaining in CR for at least 6 months. Image:Summary/Conclusion: In clinical practice, treatment decisions follow a strategy. The estimand framework guided the design of a clinical study in AML post SCT to conceptualize the assessment of a treatment strategy. Accordingly, safety and efficacy of the treatment strategy as a whole will be assessed taking into account different patient journeys.

Combining CD34+ stem cell selection with prophylactic pathogen and leukemia directed T-cell immunotherapy to simultaneously reduce graft versus host disease, infection, and leukemia recurrence after allogeneic stem cell transplant.

We designed a trial to simultaneously address the problems of graft versus host disease (GVHD), infection, and recurrence of malignancy after allogeneic stem cell transplantation. CD34+ stem cell isolation was used to minimize the development of acute and chronic GVHD. Two prophylactic infusions, one combining donor-derived cytomegalovirus, Epstein-Barr virus, and Aspergillus fumigatus specific T-cells and the other comprising donor-derived CD19 directed chimeric antigen receptor (CAR) bearing T-cells, were given 21-28 days after transplant. Two patients were transplanted for acute lymphoblastic leukemia from HLA identical siblings using standard doses of cyclophosphamide and total body irradiation without antilymphocyte globulin. Patients received no post-transplant immune suppression and were given no pre-CAR T-cell lymphodepletion. Neutrophil and platelet engraftment was prompt. Following adoptive T-cell infusions, there was rapid appearance of antigen-experienced CD8+ and to a lesser extent CD4+ T-cells. Tetramer-positive T-cells targeting CMV and EBV appeared rapidly after T-cell infusion and persisted for at least 1 year. CAR T-cell expansion occurred and persisted for up to 3months. T-cell receptor tracking confirmed the presence of product-derived T-cell clones in blood targeting all three pathogens. Both patients are alive over 3 years post-transplant without evidence of GVHD or disease recurrence. Combining robust donor T-cell depletion with directed T-cell adoptive immunotherapy targeting infectious and malignant antigens permits independent modulation of GVHD, infection, and disease recurrence. The combination may separate GVHD from the graft versus tumor effect, accelerate immune reconstitution, and improve transplant tolerability.

KEYNOTE-B79 phase 1b trial to evaluate the allogeneic CAR T-cells CYAD-101 and pembrolizumab in refractory metastatic colorectal cancer patients.

TPS227 Background: The allogeneic chimeric antigen receptor (CAR) T-cell treatment CYAD-101 utilizes the NKG2D receptor that targets eight ligands expressed on tumor cells and also on stromal and immunosuppressive immune cells of the tumor microenvironment. CYAD-101 co-expresses a T-cell receptor (TCR) inhibitory peptide with the aim to eliminate the potential of graft versus host disease (GvHD), the main safety risk associated with engineered cells of allogeneic origin. In the previous alloSHRINK trial (NCT03692429), CYAD-101 was administered post FOLFOX preconditioning chemotherapy to 15 patients with progressive metastatic colorectal cancer (mCRC) who were previously treated with FOLFOX. Overall, the treatment was well tolerated with no evidence of GvHD. The disease control rate was 73.3% with two patients presenting a confirmed partial response per RECIST’s criteria (4 months and 8 months of duration from first CYAD-101 infusion) and nine had stable disease with a median duration of 4.6 months. Evidence of increase in the TCR repertoire and modulation of the cytokine profile post-treatment with CYAD-101 were observed implying that CYAD-101 may also be modulating the immune suppressive environment in patients mirroring what was demonstrated in pre-clinical models. We considered that a sequential therapy with the anti-PD1 monoclonal antibody pembrolizumab to further release the anti-tumor potential of this expanded T-cell population may drive deeper, more durable and new clinical responses beyond that currently demonstrated with the CAR T-cells alone. Methods: The phase 1b KEYNOTE-B79 trial (NCT04991948) will evaluate, according to a Simon’s two stage study design, the safety and clinical activity of three consecutive infusions of CYAD-101 (1x109 cells per infusion) post FOLFOX preconditioning chemotherapy with two-week interval between cycles, followed by pembrolizumab treatment (200 mg administered every three weeks for up to two years) in microsatellite stable/mismatch-repair proficient mCRC patients with recurrent/progressing disease after at least one metastatic line of therapy, which must include FOLFOX chemotherapy. The pembrolizumab treatment will be initiated 3 weeks after the last CYAD-101 infusion to fall outside the classical time window of potential CAR T-cell toxicities (e.g., cytokine release syndrome). The co-primary endpoints of the trial are the occurrence of dose-limiting toxicities (DLTs) at any time from the first FOLFOX preconditioning treatment up to 3 weeks after the first pembrolizumab treatment and the objective response rate (ORR) at the tumor assessment planned 6 weeks after the first pembrolizumab treatment administration. The KEYNOTE-B79 study will be initiated in Q4-2021 in five sites in USA and Europe. Clinical trial information: NCT04991948.

Breath Analysis in Gastrointestinal Graft-Versus-Host Disease after Allogeneic Hematopoietic Cell Transplantation (HCT)

Volatile organic compounds (VOCs) are generated during pathologic processes giving unique characteristics to breath. Breath analysis has thus become an important method to diagnose and monitor a variety of diseases. There is increasing data to support a critical role for the intestinal microbiome in graft-versus-host disease (GVHD). Various microorganisms are known to produce characteristic volatile metabolites, and may alter VOCs expelled in breath. We hypothesized that patients with gastrointestinal (GI) GVHD will have a unique breath signature compared to transplant recipients without GVHD and performed a pilot study of breath analysis in HCT recipients.Exhaled breath samples were obtained from 19 patients with clinical grade 2-4 acute GI GVHD and 10 patients with no evidence of GVHD by day 100; and analyzed using mass spectrometry technology. Baseline characteristics and VOCs were compared between patients with and without GI GVHD. 22 VOCs were included in stepwise linear discriminant analysis to identify the subset that could best discriminate between GI GVHD and no GVHD.Median age at transplant was 58 years (range 13-71). The majority of patients (93%) were White, and 66% male. 55% of patients underwent myeloablative conditioning and 45% reduced intensity, with 52% receiving peripheral blood stem cells, 41% bone marrow, and 7% cord blood grafts. 45% patients had a matched unrelated donor, 34% matched sibling donor, 14% haploidentical, and 7% cord blood transplants. GVHD prophylaxis primarily consisted of a calcinuerin inhibitor in combination with mycophenolate mofetil or methotrexate. Among recipients who developed GI GVHD, median time to onset of GVHD was 43 days from transplant. 9 (47%) patients had grade 2, 8 (42%) grade 3, and 2 (11%) grade 4 GVHD. 26% had only upper GI GVHD, 21% lower GI GVHD and 53% had both upper and lower GI manifestations.Stepwise canonical discriminant analysis identified 5 VOCs that distinguished between patients with and without GI GVHD: 2-propanolol, acetaldehyde, dimethylsulfide, isoprene, and 1-decene (Wilks' Lambda 0.43, F statistic 6.08, P=0.001). The model correctly classified 89% (17/19) and 90% (9/10) of patients with and without GI GVHD, respectively (Figure 1). We found no significant correlation between VOCs and GVHD severity. However, 2 VOCs, pentane and ammonia, distinguished patients who achieved a complete response (CR) to therapy by day 28 and those who did not (Wilks' Lambda 0.57, F statistic 6.05, P=0.011). The model correctly classified 90% (9/10) patients with CR and 78% (7/9) of patients with less than CR (Figure 2). An additional VOC, acetone, distinguished patients with at least a partial response versus no response. Patients with higher levels of acetone were less likely to have any response, odds ratio 0.22, 95% confidence interval 0.06-0.82, P=0.024.Breath analysis is a feasible and promising non-invasive method to diagnose and monitor acute GI GVHD. Further study of serial breath analysis and the gut microbiome in a larger cohort are needed to validate these findings. [Display omitted] DisclosuresGerds:CTI BioPharma: Consultancy; Incyte: Consultancy. Majhail:Sanofi: Honoraria; Anthem, Inc.: Consultancy.

Targeting T Cell Bioenergetics By Modulating P-Glycoprotein Selectively Depletes Alloreactive T Cells to Prevent Graft-Versus-Host Disease While Maintaining Antitumor Immunity

Immune effector cell therapy is a potent form of cancer treatment with a high curative potential for certain human malignancies. Yet, the success of current approaches is offset by associated toxicities, including acute graft-versus-host-disease (GVHD). In the allogeneic setting, GVHD is caused by an attack on the recipient's tissues by donor T cells. Ex vivo application of phototherapy to selectively deplete GVHD-causing cells prior to transplant may prevent GVHD. However, none of the photosensitive agents in use today have demonstrated selectivity without significant toxicity occurring in resting cells. We have designed the first-in-class photosensitizer 2-Se-Cl with the ability to accumulate in alloreactive T cells in proportion to degree of oxidative phosphorylation. Unique to 2-Se-Cl is the ability to potently stimulate P-glycoprotein (P-pg). Enhanced P-gp activity promotes the efficient removal of photosensitizer not sequestered in mitochondria, and protects resting lymphocytes essential for antipathogen and antitumor responses. We have previous demonstrated the ex vivo photodepletion of alloreactive T cell using 2-Se-Cl successfully prevented GVHD in a lethal model of the disease with full retention of antipathogen immunity and 3rd party responses. To evaluate for the retention of antitumor immunity after photodepletion with 2-Se-Cl, a well-established complete MHC antigen-mismatched murine model of hematopoietic stem cell transplant (HSCT) was employed. To prepare the cells for photodepletion, donor C57BL/6 splenocytes were cocultured for 4 days with irradiated (20 Gy) Balb/c splenocytes, and then photodepleted (PD) with 2-Se-Cl. On the day of HSCT, recipient Balb/c mice were irradiated with 8.5Gy, and received 7.5 x106 T cell-depleted (TCD) C57BL/6-derived bone marrow cells accompanied by 2.5 x106 PD - treated cells, or 105 recipient-derived (A20, Balb/c-derived) leukemia cells (engineered to express firefly luciferase), or both. To evaluate for the retention of 3rd party responses, C3H/HeJ were irradiated (9.5 Gy) and received a combination of 7.5 x106 TCD C57BL/6-derived bone marrow cells accompanied by 2.5 x106 PD - treated cells. All mice were monitored for signs of GVHD according to a well-established mouse GVHD grading system. Three animals underwent HSCT in each group in 3 independent experiments. Recipient Balb/c mice that received PD-treated splenocytes survived > 45 days without evidence of GVHD or leukemia, equivalent to controls (recipients of TCD bone marrow cells only). In contrast, mice that were inoculated with leukemia at time of HSCT without the addition of PD-treated splenocytes died of leukemia progression (see figure). Additionally, all third-party C3H/HeJ mice that received the same number of PD-treated cells died of lethal GVHD, consistent with the selective depletion of the alloimmune response towards BalB/c by 2-Se-Cl. These results demonstrate that ex vivo application of 2-Se-Cl protects against lethal GVHD while maintaining antitumor immunity. The high selectivity of this novel photosensitizer may have broad applications, including the rapid production of immune effector cell products with potent antitumor activity and a low potential for toxicity. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Preliminary Results of the First-in-Human Study of Nexi-001, a Multi-Antigen Specific CD8+ T Cell Product, in Acute Myeloid Leukemia (AML) Patients with Relapsed Disease after Allogeneic Hematopoietic Cell Transplantation (Allo-HSCT) Demonstrate Early Signs of Safety, Tolerability and Robust Immune Responses

Preliminary Results of the First-in-Human Study of Nexi-001, a Multi-Antigen Specific CD8+ T Cell Product, in Acute Myeloid Leukemia (AML) Patients with Relapsed Disease after Allogeneic Hematopoietic Cell Transplantation (Allo-HSCT) Demonstrate Early Signs of Safety, Tolerability and Robust Immune Responses

A Prospective Haploidentical Peripheral Blood Stem Cell Transplant Study Using a Pre-Defined Conditioning Regimen Intensity Based on Age and the Hematopoietic Cell Transplantation Comorbidity Index- Anzhit 1: Encouraging Preliminary Survival Outcomes at One Year Follow up

Background HLA-matched sibling and unrelated donors are not always available for patients that are in need of an allogeneic haematopoietic stem cell transplant. Partially HLA-mismatched related (haploidentical) donors can be identified for the vast majority of patients but these transplants were historically complicated by excessive graft versus host disease (GVHD), nonrelapse mortality (NRM), and poor overall survival (OS). Haploidentical haematopoietic stem cell transplantation (Haplo HSCT) has become increasingly common over the last ten years, predominately due to the successful use of post-transplant cyclophosphamide (PTCy) in mitigating graft versus host disease (GVHD). Most studies using PTCy have been retrospective with various conditioning regimens and stem cell sources making interpretation of results difficult. ANZHIT-1 is a prospective Australian multi-centre phase II study of Haplo HSCT with defined conditioning and GVHD prophylaxis regimens. Methods: This is an ethics approved Phase II study (ACTRN: 12617000151336) conducted between 2015-2020 at six Australian allogeneic HSCT centres. All patients received peripheral blood Haplo HSCT from haploidentical family members. The reduced intensity regimen (RIC) was fludarabine, cyclophosphamide and 200cGy TBI and the myeloablative regimen (MAC) was fludarabine with IV busulfan 3.2mg/kg x 4. PTCy 50mg/kg D+3, D+4 was used with MMF (to D35) and a calcineurin inhibitor (CNI) as GVHD prophylaxis. CNIs were weaned and ceased by D+120 in patients who did not have evidence of GVHD. Choice of conditioning regimen was based on age and hematopoietic cell transplantation comorbidity index (HCT-CI) scores. Patients who had an HCTCI >3 or age >50yrs received RIC. Data collection was through the Australian Bone Marrow Transplant Recipient Registry. Survival was analysed using the Kaplan-Meier method. Results: Seventy eight patients were included in the study with a median follow up of 402.5 days (28-1259). There was a male predominance (66%). The median age was 53 years (range 18-69) and the median HCT-CI score was 1 (range 0-5). Thirty nine patients had acute myeloid leukaemia, 12 acute lymphoblastic Leukaemia, 14 Non Hodgkin lymphoma or Hodgkin lymphoma, 4 myelodysplastic syndrome and 9 other diagnoses. The majority received RIC (59%). Median neutrophil and platelet engraftment times were 18 days (range: 12-92) and 28 days (range: 13-262) respectively. Acute GVHD grade III-IV occurred in 14.1% of patients whilst chronic GVHD occurred in 20.5%. Moderate to severe cGVHD according to NIH consensus criteria occurred in 10.3% of patients. Overall survival probability at one year was 74.1% (95% CI: 52.9-86.8) for MAC recipients and 77.8% (95% CI: 61.6-87.8) for RIC recipients (Figure 1). Non relapse mortality (NRM) at D100 and 1 year were 2.2% and 8.7% respectively in the RIC group compared to 6.3% and 18.8% respectively in the MAC group. Causes of NRM included infection (n=5), GVHD (n=3) and VOD, respiratory failure and multi-organ failure in one patient each. Haemorrhagic cystitis and poor graft function were relatively common post-transplant morbidities but rarely contributed to NRM. Despite the use of peripheral blood stem cells and early weaning of CNIs, only 3/11 NRM deaths were due to GVHD. Relapse was the most common cause of death in the RIC group with 17.4% of patients relapsing at a median of 114 days (range 36-564) compared to only 6.3% at a median of 90.5 days (range 64-154) in the MAC group. Conclusion: This prospective Haplo HSCT trial utilising PTCY demonstrates encouraging overall survival rates. The RIC regimen had a promising toxicity profile but early relapse remains a concern despite early weaning of CNIs. In contrast, the MAC regimen is associated with a low relapse rate but NRM at one year remains significant. Further refinement of the conditioning and GVHD prophylaxis regimens will be incorporated in the follow-on study, ANZHIT-2, which will begin accrual in late 2020. Disclosures Bajel: Pfizer: Honoraria; Amgen: Honoraria, Speakers Bureau; Novartis: Honoraria; Astellas: Honoraria; Abbvie: Honoraria. Greenwood:Servier: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; MSD: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Hamad:Novartis: Honoraria; Abbvie: Honoraria.

Pseudo-Allogeneic CAR-T Therapy after Allogeneic Stem Cell Transplantation in Relapsed/Refractory B-Cell NHL

Introduction Chimeric antigen receptor T-cell (CAR-T) therapy has emerged as a powerful therapy for relapsed and refractory (R/R) B-cell malignancies. A growing body of research suggests that CAR-T therapy can be safe and effective in patients who have previously received allogeneic hematopoietic stem cell transplantation (alloHSCT). "Pseudo-allogeneic" CAR-T therapy refers to manufacture of engineered T-cells collected from an alloHSCT recipient exhibiting 100% donor chimerism. This CAR-T approach poses theoretical risks due to alloreactivity and increased CAR-T toxicity. Most studies to date in this area have enrolled patients with leukemia. We present here a single-center experience with pseudo-allogeneic CAR-T following alloHSCT in 6 patients with large B-cell lymphoma (LBCL). Patients and Methods Patients with R/R LBCL who underwent treatment with axicabtagene ciloleucel (axi-cel) CAR-T after prior alloHSCT were identified. All patients underwent lymphodepletion with cyclophosphamide and fludarabine prior to CAR-T infusion. CAR-T toxicities, namely cytokine release syndrome (CRS) and immune effector-cell associated neurotoxicity syndrome (ICANS) were graded in accordance with the 2019 American Society for Transplantation and Cellular Therapy (ASTCT) consensus grading. Results Six patients who underwent axi-cel CAR-T due to post-alloHSCT R/R LBCL were identified. All patients had stage ≥II disease. At the time of CAR-T apheresis, all patients had ≥95% donor chimerism and were off immunosuppression, without evidence of GVHD. Manufacturing failure occurred in only one patient (case 2) due to low viability and upon second apheresis, manufacture was successful. All 6 pseudo-allogeneic CAR-T therapy patients tolerated infusion and did not show increased incidence or severity of CRS or ICANS compared to our center's experience with standard CAR-T patients, with only one patient experiencing greater than grade 2 toxicity. Three of six patients developed graft-versus-host disease (GVHD); however, one case was in the setting of PD-1 blockade and two were isolated cutaneous manifestations. There was no incidence of severe GVHD. All three cases of GVHD underwent CAR-T within ten months after alloHSCT, whereas the three patients without GVHD were >1 year post-HSCT at time of CAR-T. Two cases who had donor lymphocyte infusions prior to CAR-T did not develop GVHD. In terms of efficacy, three patients achieved a complete remission. There was no perceptible relationship between the development of GVHD and the response to CAR-T. Four of six had cytopenias at day 30 after CAR-T, with only two exhibiting persistent cytopenias at day 90. Lastly, in this small sample size, no differences were seen in incidence of GVHD or treatment response between haploidentical, matched-related, and matched-unrelated alloHSCT donors. Conclusion In our experience, pseudo-allogeneic CAR-T therapy following alloHSCT appears to be a safe and well-tolerated treatment modality for R/R B-cell lymphoma. The efficacy of this modality will need to be borne out in further studies. Disclosures No relevant conflicts of interest to declare.

Safe & Effective Reduced Intensity Conditioning Regimen in Patients with Dyskeratosis Congenita

Background: Dyskeratosis congenita (DC) is a rare inherited bone marrow failure syndrome, caused by telomerase dysfunction leading to shortened telomeres.1 Recent literature shows only a 23% estimated 10-year survival, secondary to death from pulmonary complications.2 Currently, hematopoietic cell transplantation (HCT) is the only curative therapy for marrow failure but has been limited by mortality related to infection, sinusoidal obstructive syndrome (SOS), graft-versus-host disease (GVHD) and respiratory failure.3,4 The ideal conditioning regimen that optimizes engraftment while maximizing long-term survival is yet to be determined. Methods: We describe three consecutive patients with DC who underwent matched unrelated donor (MUD) allogeneic HCT following a reduced-intensity conditioning regimen similar to protocol NCT03330795 that uses a reduced intensity conditioning (RIC) while maximizing the benefits of shielding vulnerable organs in this populations. These three patients received a RIC regimen of rituximab, alemtuzumab, hydroxyurea, thiotepa, fludarabine, and 250 cGy total body irradiation; patients 1 and 2 received 85% lung shielding and 50% liver shielding. Results: Patient 1 is a 17-year-old male with reduced diffusing capacity of the lungs for carbon monoxide (DLCO), splenomegaly, splenic varices and nodular regenerative hyperplasia (NRH) who presented with pancytopenia. Testing revealed telomeres below the 1st percentile in all WBC subtypes, a PARN c.1257dup heterozygous mutation and a TERC c.17_31 mutation. He underwent a 12/12 MUD HCT and engrafted on Day +18. His course was complicated by poor adherence to medications and visits, and development of skin and GI GVHD, which responded to steroids. He was weaned off of immunosuppression by 11 months post-transplant. Twenty-eight months post-transplant, he remains 100% donor engrafted with no chronic GVHD or organ toxicity, including stable pulmonary function tests (PFT). See figure 1 and 2. Patient 2 is a 16-year-old male with reduced DLCO, splenomegaly and NRH who was diagnosed after his brother (Patient 1). He was found to have very short telomeres in all testable WBC subtypes and the same mutations as his brother. After marrow failure, he underwent a 12/12 MUD HCT, with neutrophil engraftment on Day +16. His course was complicated by sudden noncompliance with tacrolimus at day +100, after which he suffered GI GVHD which responded to steroids. He then developed disseminated adenovirus infection which responded to an adenovirus-specific T-cell infusion. He is now Day +356 after transplant and has been off all immunosuppression for 4 months. He remains >98% donor-engrafted without evidence of GVHD or organ toxicity, and his PFTs have remained stable. Patient 3 is a 16-year-old male with a history of antibody-negative autoimmune hepatitis who later developed pancytopenia. Telomere testing revealed very short telomeres in 4 of the 5 evaluable WBC subtypes. His genetic testing did not uncover a mutation yet identified as pathogenic in DC. He received a 12/12 MUD HCT and engrafted on Day +12. He had skin GVHD which responded to ruxolitinib. He remains 100% donor-engrafted, most recently at 9 months post-transplant without organ toxicity. Conclusions: All patients had timely neutrophil engraftment without SOS or other organ toxicity, despite being at increased risk for organ morbidity due to preexisting liver and lung disease. GVHD was noted, although two of the cases were related to poor adherence, and all responded to treatment. There is sustained donor-engraftment at a median of 637 days (range 356-1186 days) post-transplant.

Photodepletion with 2-Se-Cl prevents lethal graft-versus-host disease while preserving antitumor immunity.

Acute graft-versus-host-disease (GVHD), limits the use of hematopoietic cell transplant (HCT) to treat a variety of malignancies. Any new therapeutic approach must satisfy three requirements: 1) Prevent GVHD, 2) Maintain anti-pathogen immunity, and 3) Maintain anti-tumor immunity. In prior studies we have shown that the selective photosensitizer 2-Se-Cl eliminates highly alloreactive lymphocytes from the graft prior to HCT preventing GVHD and that antiviral immune responses were preserved following incubation with 2-Se-Cl. In this report, we investigated whether 2-Se-Cl treatment preserves antitumor immunity, and then used high dimensional flow cytometry to identify the determinants of successful immune reconstitution. Donor C57BL/6 splenocytes were cocultured for 4 days with irradiated BALB/c splenocytes and then exposed to 2-Se-Cl. Photodepletion (PD)-treated splenocytes were then infused into lethally irradiated BALB/c mice inoculated with A20 leukemia/lymphoma cells. Recipient mice that received PD-treated splenocytes survived > 100 days without evidence of GVHD or leukemia. In contrast, mice that did not receive PD-treated cells at time of HCT died of leukemia progression. Multiparameter flow cytometry of cytokines and surface markers on peripheral blood samples 15 days after HCT demonstrated unique patterns of immune reconstitution. We found that before clinical disease onset GVHD was marked by functionally exhausted T cells, while tumor clearance and long-term survival were associated with an expansion of polyfunctional T cells, monocytes, and DCs early after transplantation. Taken together these results demonstrate that 2-Se-Cl photodepletion is a new treatment that can facilitate HCT by preventing GVHD while preserving antiviral and anti-tumor immunity.

Incidence of graft versus host disease in peri transplant hospitalization after clostridium difficile infection.

7542 Background: Hyperacute graft versus host disease (GVHD) after allogenic stem cell transplantation (SCT) has adverse outcomes with increased rates of chronic GVHD and relapse. GVHD risks include mismatched related or matched unrelated donors, myeloablative conditioning, heavy pretreatment, and donor-recipient sex mismatch. Clostridium difficile infection (CDI) is a leading cause of diarrhea in immunocompromised patients. Proposed microbiome effect on immunity and GVHD in allogenic SCT recipients prompts concern of microbiome modulation from CDI and antibiotics inciting GVDH. Methods: National Inpatient Sample database 2014 for hospitalizations with allogeneic SCT in patients ≥18yo. Characteristics (age, sex, race, insurance, graft source, hospital type, region, comorbidities) were compared for hospitalizations with and without CDI. Primary outcome was the difference in the incidence of GVHD during the transplant hospitalization between the 2 groups. Other outcomes were mortality, length of stay and hospital charges. Chi-square, t-test, and multivariate logistic regression utilized. Results: Of 6210 patients with allogenic SCT, 745 (12%) had CDI during the transplant hospitalization. In transplanted patients without CDI the average age was 55yo, 43.9% female, 69.5% Caucasian (C), 7.1% African American (AA), 8.6% Hispanic (H), 32.1% had Medicare/Medicaid, 61.8% private insurance, 5.7% uninsured, 44.7% had hypertension, 13.7% had diabetes, graft source was 84.3% PBSC (peripheral blood stem cells), 11.3% bone marrow, and 4.4% cord blood. CDI group the average age was 52.5yo, 45.3% female, 73% C, 4.7% AA, 8.1% H, 25.7% had Medicare/Medicaid, 66.2% private insurance, 8.1% uninsured, 41.9% had hypertension, 10.1% had diabetes, graft source was 83.8% PBSC, 10.8% bone marrow, and 5.4% cord blood. 25.7% of patients with CDI developed GVHD during that hospitalization while 14.2% of patients without CDI developed GVHD during the hospital stay (OR 2.1, p < 0.001 multivariate analysis). GVHD during the hospitalization had no difference in length of stay (p = 0.32), total cost of stay (p = 0.50) or same hospitalization mortality (p = 0.94). Conclusions: Allogeneic SCT patients with CDI develop GVHD on the same hospitalization at significantly higher rates than patients without CDI. This is true after controlling for age, sex, race, insurance, comorbidities, graft source, hospital location, and type of institution. Despite known associations of early evidence of GVHD on relapse, overall mortality was not different between the two groups.

Proof of Concept for Repair of Sickle Cell Disease By Non-Myeloablative MHC Disparate T Cell Depleted HSCT Combined with Donor-Derived Veto Cells

Although life extending medical treatments are available for sickle cell disease (SCD), allogenic hematopoietic stem cell transplantation (HSCT) is considered a treatment of choice (Ozdogu et al., Bone Marrow Transplant 2018; 53(7): 880-890). However, HSCT is associated with several limitations, including conditioning-related toxicity and graft-versus-host disease (GvHD), especially when using MHC disparate transplants. Thus, the development of safe transplantation protocols for MHC disparate HSCT in sickle cell disease is vital. While the risk of GvHD and conditioning toxicity can be effectively reduced by the use of T-cell-depleted HSCT (TD-HSCT) under reduced intensity conditioning (RIC), rejection of TD-HSCT remains a challenge. We previously demonstrated in wild type mice that this barrier can be overcome using donor-derived veto cells. Here, we demonstrate the safety and efficacy of this approach in a well-defined murine model for sickle cells disease. Veto activity is based on the ability of certain cells to attack host CTL-precursors (CTLp) which are directed against antigens expressed on the veto cells themselves, sparing cells that are not targeted against the veto cells including T cells needed for defense against pathogens. Central memory CD8 T cells exhibit the most robust veto activity upon transplantation; however, these cells are also endowed with marked GvH activity. We overcame this issue by expanding naïve or memory CD8 T cells against 3rd party MHC or viral antigens, respectively, under culture conditions favoring expression of central memory phenotype. Such anti-3rd party central memoryCD8 T cells (Tcm), which are endowed with marked veto activity, also exhibit reduced risk for GvHD in fully mis-matched recipients (Reviewed in Reisner Y, Or-Geva N. Semin Hematol. 2019; 56(3): 173-182.) To generate Tcm veto cells, splenocytes obtained from Balb/c donors (H2d) were cultured against irradiated third-party splenocytes (FVB; H2q) under cytokine deprivation. The selective expansion of CD8 mouse T cells against 3rd party stimulators leads to selective 'death by neglect' of bystander anti-host T cell clones potentialy mediating GvHD, and these are further diluted out by subsequent expansion of anti-3rd party T clones during continued culture in the presence of IL-15. Apart from selective loss of GvH reactive T cells, these culture conditions induce a central memory phenotype shown to be important for attaining robust veto activity in vivo (Ophir et al., Blood. 2013; 121(7): 1220-8). We first calibrated the optimal irradiation dose for sickle mice (Berkeley model, H2b), comparing 4.5 Gy versus 5 Gy TBI in a conditioning protocol also including short term rapmycin treatment (Scheme 1). Higher levels of engraftment and chimerism were found in the group receiving 5Gy TBI (Fig. 1). All mice of both treatment groups survived >140 days with no evidence of GvHD. To further evaluate this treatment approach, 8 week old sickle mice (N=7) were given bone marrow transplants using the protocol described in Scheme 1, including conditioning with 5 Gy TBI (day -1), rapamycin treatment (day -1 to day 4), and transplantation of NuBM (day 0) plus veto cells (day 7; Scheme 1). Notably, at 44 days post-transplant, 6 out of 7 mice receiving NuBM + TCM + Rapa (85.7%) showed donor chimerism in the peripheral blood, ranging between 77-94% (Fig.2A-B), while no chimerism was detected in mice receiving conditioning alone, or conditioning and transplantation with only NuBM or only veto cells . All mice in all groups survived (N=26) , and no GvHD was detected with a follow up of 77 days, even in the transplanted group which exhibited high donor-derived chimerism. Furthermore, reversal of sickle disease symptoms was observed, including reticulocyte levels (p=0.001;Fig.2C) and expression of wild type hemoglobin ( Fig.2D) in all engrafted mice. Our results offer a proof of concept for the treatment of sickle disease by MHC disparate non-myeloablative T cell depleted HSCT in conjunction with anti-3rd party central memory veto CD8 T cells. A clinical trial testing the safety and efficacy of anti-3rd party veto cells in the context of low toxicity non-myeloablative TD-HSCT in hematological malignancies is currently ongoing at MD Anderson Cancer Center. If successful, our present results support further evaluation of this platform for sickle cell disease. Disclosures Lustig: Yeda Ltd.: Patents & Royalties. Reisner:Cell Source, Inc.: Consultancy, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.

Membranous nephropathy succeeding autologous hematopoietic stem cell transplant: a case report

BackgroundMembranous nephropathy (MN), the leading cause of nephrotic syndrome in adults, is characterized by the deposition of subepithelial immune deposits. Most of the cases are primary, while only approximately 25% of the cases are secondary to some known diseases. Recently, MN has been considered to be a possible presentation of chronic graft-versus-host disease (GVHD) of the kidney in allogeneic hematopoietic stem cell transplantation (HSCT) patients. In autologous HSCT populations, there have been scarce reports of associated MN, as a result of immune dysregulation leading to systemic autoimmunity and miming chronic GVHD.Case presentationWe report an exceptional case of MN associated to an acute renal failure occurring within days following an autologous HSCT indicated by multiple myeloma. There was no evidence of GVHD or myeloma relapse. A complete remission of nephrotic syndrome with normalization of renal function were rapidly obtained by corticosteroid therapy.ConclusionThis is the first published case of acute renal failure due to MN occurring in the acute phase of an autologous HSCT. These findings support the antibodymediated autoimmune glomerular disease.

CGRA4: VENO‐OCCLUSIVE DISEASE WITH IMMUNODEFICIENCY (VODI) TREATED WITH HAPLO‐IDENTICAL BONE MARROW TRANSPLANT

Veno-Occlusive disease with Immunodeficiency (VODI) is a rare combined immunodeficiency. Mutations in SP110, have been described in the Lebanese population with VODI. Untreated, reported outcomes for VODI are 100% mortality at one year of life. Conservative management (PJP prophylaxis and Immunoglobulin) result in mortality of 75%. Bone marrow transplant has been demonstrated as the only definitive treatment, but has a high rate of complications. Haploidentical HSCT provides an opportunity for Stem Cell Transplantation in those where a matched donor is not available. Historically, Haploidentical HSCT has been limited by high rates of graft rejection, Graft-Versus-Host Disease (GVHD), Transplant-related mortality and poor immune reconstitution. We present the first case of VODI that has been managed with a TCR alpha/beta, CD19 depleted Haplo-identical HSCT. This patient, the first child to consanguineous Lebanese parents, presented at 4 months of age with a febrile illness, hepatomegaly and abdominal ascites. Immunoglobulins were undetectable and the patient had normal lymphocyte subsets. SP110 mutation was confirmed. Infections included CMV, confirmed on Whole Blood PCR, and Pneumocystis Jerovicii Pneumoniae (PJP) confirmed on Brochoalveolar Lavage PCR. The latter resulted in acute deterioration, for which the patient required High Frequency Oscillatory Ventilation prior to improvement with high-dose Bactrim. CMV infection was treated with Gancyclovir. Haploidentical HSCT was performed using a parent donor. Conditioning consisted of Treosulfan and Fludarabine. The patient engrafted on day 11. MMF was discontinued at day +35. Day +30 donor chimerism was 99%. The post-transplant course has been complicated by deranged liver function, presumed secondary to CMV as liver biopsy has demonstrated resolution of Veno-Occlusive Disease and no evidence of GVHD. This case highlights successful use of Haploidentical HSCT in treatment of VODI. Further evaluation is required with subsequent cases and long-term outcomes.

Targeting T Cell Bioenergetics by Modulating P-Glycoprotein Selectively Depletes Alloreactive T Cells To Prevent Graft-versus-Host Disease.

T lymphocytes play a central role in many human immunologic disorders, including autoimmune and alloimmune diseases. In hematopoietic stem cell transplantation, acute graft-versus-host-disease (GVHD) is caused by an attack on the recipient's tissues from donor allogeneic T cells. Selectively depleting GVHD-causing cells prior to transplant may prevent GVHD. In this study, we evaluated 24 chalcogenorhodamine photosensitizers for their ability to selectively deplete reactive T lymphocytes and identified the photosensitizer 2-Se-Cl, which accumulates in stimulated T cells in proportion to oxidative phosphorylation. The photosensitizer is also a potent stimulator of P-glycoprotein (P-gp). Enhanced P-gp activity promotes the efficient removal of photosensitizer not sequestered in mitochondria and protects resting lymphocytes that are essential for antipathogen and antitumor responses. To evaluate the selective depletion of alloimmune responses, donor C57BL/6 splenocytes were cocultured for 5 d with irradiated BALB/c splenocytes and then photodepleted (PD). PD-treated splenocytes were infused into lethally irradiated BALB/c (same-party) or C3H/HeJ (third-party) mice. Same-party mice that received PD-treated splenocytes at the time of transplant lived 100 d without evidence of GVHD. In contrast, all mice that received untreated primed splenocytes and third-party mice that received PD-treated splenocytes died of lethal GVHD. To evaluate the preservation of antiviral immune responses, acute lymphocytic choriomeningitis virus infection was used. After photodepletion, expansion of Ag-specific naive CD8(+) T cells and viral clearance remained fully intact. The high selectivity of this novel photosensitizer may have broad applications and provide alternative treatment options for patients with T lymphocyte-mediated diseases.

Diagnostic phrasing is independently correlated with the decision to treat for graft-versus-host disease: retrospective review of colon biopsies with rare apoptosis.

The risks of immunosuppression and the non-specific nature of rare crypt apoptosis has led to debate over the lower threshold for histological diagnosis of colonic graft-versus-host disease (GVHD). A recent study proposed the diagnostic category of indeterminate for GVHD (iGVHD) for cases with six or fewer apoptotic bodies per 10 crypts. Our aim was to assess colon biopsies with iGVHD histology to determine whether the diagnosis was retrospectively predictive of the decision to treat, and to correlate these findings with endoscopic and clinical findings. A retrospective search was performed for colonic biopsies taken to evaluate for GVHD from 2008 to 2014. Biopsies were blindly reviewed for the maximum number of apoptotic bodies per 10 contiguous crypts, evidence of crypt dropout, and ulceration. Clinical information was collected through chart review. One hundred and twenty-two biopsies from 84 transplant patients were included. Forty-seven cases met the histological criteria for iGVHD. Patients with an original diagnosis of iGVHD were more likely to be managed conservatively than those with a diagnosis of grade 1 GVHD (25% versus 0%). Eight symptomatic patients reclassified as iGVHD had resolution of symptoms without increased immunosuppression. A clinicopathologically similar group of 10 patients with iGVHD histology, normal or subtle endoscopic findings and no evidence of GVHD at other organ sites were treated with increased immunosuppression. On multivariate analysis, the original diagnostic category was the most significant predictor of the decision to treat. The use of the diagnostic category iGVHD alerts clinicians to the presence of minimal crypt apoptosis, and allows treatment based on clinical judgement.

Stepwise Endoscopy Based on Sigmoidoscopy in Evaluating Pediatric Graft-versus-Host Disease

PurposeThe aim of our study was to establish a safe and convenient diagnostic method for acute gastrointestinal (GI) graft-versus-host disease (GVHD) in children by determining the sensitivity and negative predictive values of upper and lower endoscopic biopsies for children suspected of GI GVHD.MethodsPatients suspected of GI GVHD who received endoscopic evaluation within 100 days after stem cell transplantation and endoscopies between January 2012 and March 2014 in Seoul National University Children's Hospital were included in our study.ResultsFifteen patients with a total of 20 endoscopic procedures were included in our study. Sensitivity at the esophagus, stomach, and duodenum were 22.2%, 30.0%, and 80.0%, respectively. Negative predictive values at the esophagus, stomach, and duodenum were 22.2%, 30.0%, and 60.0%, respectively. Overall sensitivity and negative predictive values of upper endoscopic biopsy for GVHD were 77.8% and 50.0%, respectively. Overall sensitivity and negative predictive values of lower endoscopic biopsy for GVHD were 88.9% and 66.7%, respectively.ConclusionWe recommend flexible sigmoidoscopy as a safe and accurate diagnostic tool for GVHD, similar to other studies reported previously. However, if there is no evidence of GVHD on sigmoidoscopy with high index of suspicion of GI bleeding, full colonoscopy and upper endoscopy should be considered.

P-Glycoprotein Modulation Facilitates the Selective Inhibition of Oxidative Phosphorylation in Alloreactive T Cells to Prevent Graft-Versus-Host Disease after Hematopoietic Stem Cell Transplant

T lymphocytes play a central role in many human immunologic disorders including both autoimmune and alloimmune diseases. In hematopoietic stem cell transplant (HSCT), acute graft-versus-host-disease (GVHD) is caused by an attack on the recipient's tissues from donor allogeneic T cells. Ex vivo application of phototherapy to selectively deplete GVHD-causing cells prior to transplant may prevent GVHD. However, none of the photosensitive agents in use today have demonstrated selectivity without significant toxicity occurring in resting cells. We have designed the first-in-class, novel photosensitizer 2-Se-Cl with the ability to accumulate in pathogenic T cells in proportion to degree of oxidative phosphorylation (OXPHOS). Unique to 2-Se-Cl is the ability to potently stimulate P-glycoprotein (P-pg). Enhanced P-gp activity promotes the efficient removal of photosensitizer not sequestered in mitochondria, and protects resting lymphocytes essential for antipathogen and antitumor responses. To confirm that 2-Se-Cl selectively depletes antigen-specific immune responses, human lymphocytes were cultured with staphylococcal enterotoxin B (SEB) for 72 hours. Cells were then pulsed with 2-Se-Cl and exposed to 5 J/cm2 of light, which resulted in the impedance of OXPHOS and induction of apoptosis in stimulated T cells. After photodepletion (PD), resting cells remained intact with the ability to respond to stimulation by Toxic Shock Syndrome Toxin-1 (TSST-1) but not SEB. To evaluate the selective depletion of alloimmune responses, a well-established complete mHC antigen-mismatched murine model of HSCT was employed. To prepare the PD - treated primed splenocytes, donor C57BL/6 splenocytes were cocultured for 5 days with irradiated Balb/c splenocytes, and then photodepleted. In vitro analysis of PD - treated primed splenocytes demonstrated selective depletion of antihost immune responses by CFSE dilution, with complete retention of third-party responses (C3H/HeJ mice). On the day of HSCT, recipient Balb/c mice were irradiated with 900 cGy, and received 5x106 T cell-depleted bone marrow cells accompanied by 5x106 PD - treated (treatment group) or untreated (control group) primed splenocytes, and then monitored for signs of GVHD according to an established mouse GVHD grading system. Five animals underwent HSCT in each group in 3 independent experiments. All mice died of acute GVHD after the addition of 5x106 untreated primed C57BL/6 cells at a median of 25 days after HSCT. In contrast, > 90% of mice that received the same number of PD - treated cells survived > 60 days without evidence of GVHD (see figure). Additionally, third-party C3H/Hej mice that received the same number of PD-treated cells died of GVHD, confirming the selective depletion of the alloimmune response for BalB/c by 2-Se-Cl. The high selectivity of this novel photosensitizer may have broad applications, including targeting the bioenergetics in alloimmune, autoimmune, and malignant T cells to provide alternative treatment options and improve clinical outcomes for patients with diseases such as GVHD, systemic lupus erythematosus, or peripheral T cell lymphoma. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Human Bone Marrow Stromal Cells Differentiate Into Corneal Tissue and Prevent Ocular Graft-Versus-Host Disease in Mice.

Clinical trials have assessed the use of human bone marrow stromal cells (hBMSCs) for the treatment of immune-related disorders such as graft-versus-host disease (GVHD). In the current study, we show that GFP(+)-transduced hBMSCs generated from bone marrow migrate and differentiate into corneal tissue after subconjunctival injection in mice. Interestingly, these hBMSCs display morphological features of epithelial, stromal, and endothelial cells and appear at different layers and with different morphologies depending on their position within the epithelium. Furthermore, these cells display ultrastructural properties, such as bundles of intermediate filaments, interdigitations, and desmosomes with GFP(-) cells, which confirms their differentiation into corneal tissues. GFP(+)-transduced hBMSCs were injected at different time points into the right eye of lethally irradiated mice undergoing bone marrow transplantation, which developed ocular GVHD (oGVHD). Remarkably, hBMSCs massively migrate to corneal tissues after subconjunctival injection. Both macroscopic and histopathological examination showed minimal or no evidence of GVHD in the right eye, while the left eye, where no hBMSCs were injected, displayed features of GVHD. Thus, in the current study, we confirm that hBMSCs may induce their therapeutic effect at least in part by differentiation and regeneration of damaged tissues in the host. Our results provide experimental evidence that hBMSCs represent a potential cellular therapy to attenuate oGVHD.