allo-BMT Recipients Research Articles

Allogeneic blood or marrow transplantation using haploidentical grandchildren donors and post-transplant cyclophosphamide-based graft-versus-host disease prophylaxis.

High-dose post-transplant cyclophosphamide allows safe and effective use of allografts from haploidentical relatives (siblings, parents and children) in patients undergoing allogeneic blood or marrow transplant (alloBMT). More recently, second- and third-degree relatives have also been shown to be safe allograft donors. An increasing number of older patients undergoing alloBMT have been receiving allografts from haploidentical donors. However, older patients are more likely to have older siblings and children, and older donor age is associated with worse outcomes. In the current study, we report the safety and utility of grandchildren as haploidentical donors and compared with children as donors in patients undergoing alloBMT. We compared characteristics and outcomes of alloBMT patients aged 55 years and older with children older than 30 years as donors (C group; n = 276) and those with grandchildren as donors (GC group; n = 40). Because many important baseline characteristics predict outcomes after alloBMT, we performed propensity score matched analysis based on recipient age, alloBMT year, disease, graft source and haematopoietic cell transplantation comorbidity index (HCT-CI). The median age of recipients was 67 years (range 55-79) in the C group and 73 years (range 57-78) in the GC group. More than 70% of recipients in the GC group were older than 70 years, compared with 27% in the C group. The median donor age was 37 years (range 31-52) in the C group and 20 years (range 14-34) in the GC group. More patients in the GC group had HCT-CI scores ≥3 than in the C group (32.5% vs. 23%, p = 0.27). Two-year overall survival did not differ between the two groups (GC 62% vs. C 60%, hazard ratio [HR] 0.96, 95% confidence interval [CI] 0.53-1.75, p = 0.90) despite recipients of allografts from grandchildren being older. The 2-year RFS was 55% in the C group compared with 50% in the GC group (HR 1.05, 95% CI 0.62-1.77, p = 0.85). Non-relapse mortality subdistribution [SD] (SDHR 1.36, 95% 0.70-2.63, p = 0.36), relapse (SDHR 0.72, 95% CI 0.33-1.58, p = 0.42) or relapse-free survival (HR 1.05, 95% CI 0.62-1.77, p = 0.85). Propensity score matching analysis showed no significant differences in 2-year overall survival (GC 64% vs. C 53%; HR 0.77, 95% CI 0.42-1.42, p = 0.40), non-relapse mortality (SDHR 1.26, 95% 0.66-2.41, p = 0.48), relapse (SDHR 0.57, 95% CI 0.21-1.52, p = 0.26) or relapse-free survival (HR 0.94, 95% CI 0.57-1.54, p = 0.81). Our results indicate that outcomes of alloBMT patients with grandchild donors are similar to those with child donors, despite recipients' older age and higher comorbidities in the GC group. Grandchildren should be considered when selecting a donor for older alloBMT recipients.

Vasoactive Intestinal Polypeptide in the Enteric Neural Plexus Promotes Microbiome Diversity and Resistance to GvHD

Abstract Graft-versus-host disease (GvHD) in allo-transplant recipients is characterized by donor T cell- mediated damage to recipient organs and changes in the gut microbiome. Vasoactive intestinal peptide (VIP), an immunosuppressive neuropeptide, directs T cell responses towards a Th2 phenotype while inhibiting proinflammatory cytokine production. VIP-producing enteric neurons contribute to the mucosal barrier and maintain gut microbiome homeostasis. The impact of VIP production by host cells on GvHD remains unclear. We examined the effect of endogenous VIP synthesis in modulating GvHD in multiple murine allo-BMT models. VIP knock-out mice were exquisitely sensitive to GvHD, experiencing activation and expansion of Th1- and Th17-polarized T cells and early death. Transplanting radiation chimeric mice in which the VIP gene was knocked out in either hematopoietic or non-hematopoietic cells demonstrated that VIP production by non-hematopoietic tissues limited the GvHD activity of donor T cells. Immunofluorescence staining demonstrated VIP expression in enteric neurons in allo-BMT recipients, consistent with local effects of VIP on T cells and the gut microbiome. VIP- KO mice had lower gut pH, higher frequencies of Bacillota species, and less microbiome diversity post-transplantation. These data support the crucial role of VIP in maintaining the diversity of the gut microbiome with beneficial commensal species and in modulating the alloreactivity of donor T cells after allo-BMT.

Recipient clonal hematopoiesis in allogeneic bone marrow transplantation for lymphoid malignancies

AbstractAllogeneic blood and marrow transplantation (alloBMT) is increasingly being used in older patients with blood cancer. Aging is associated with an increasing incidence of clonal hematopoiesis (CH). Although the effects of donor CH on alloBMT has been reported, the impact of recipient CH on alloBMT outcomes is unknown. In this retrospective study, alloBMT recipients age 60 and older with lymphoid malignancies were included. Among 97 consecutive patients who received alloBMT between 2017 and 2022, CH was detected in 60 (62%; 95% confidence interval [CI], 51-72). CH was found in 45% (95% CI, 28-64) of patients aged 60 to 64, 64% (95% CI, 44-81) of patients aged 65% to 69%, and 73% (95% CI, 59-87) in those above 70. Pretransplant CH was associated with worse survival after alloBMT: 3-year overall survival (OS) was 78% (95% CI, 65-94) for patients without CH vs 47% (95% CI, 35-63) for those with CH, (unadjusted HR, 3.1; [95% CI, 1.4-6.8; P < .001]). Nonrelapse mortality (NRM) was higher in patients with CH; cumulative incidence of NRM at 1-year was 11% (95% CI, 1-22) vs 35% (95% CI, 23-48), (HR, 3.4; [95% CI, 1.4-8.5], P = .009]). Among CH patients, worse OS and NRM was associated with CH burden and number of mutations. Recipient CH had no effect on relapse. In conclusion, older patients with CH experience worse outcomes after alloBMT, almost exclusively attributable to increased NRM. CH is a strong, independent predictor of outcomes. Novel strategies to ameliorate the adverse impacts of patient CH on transplant outcomes are being evaluated.

Vasoactive Intestinal Peptide Signaling in Transplant Recipients Modulates Gut Microbiome Diversity and Regulates Graft-Versus-Host Disease

Introduction: Allogeneic bone marrow transplantation (allo-BMT) presents a significant challenge in the form of graft-versus-host disease (GvHD), characterized by donor T cell-mediated attacks on recipient organs, leading to considerable morbidity and mortality. Vasoactive intestinal peptide (VIP), an immunosuppressive neuropeptide, plays a crucial role in shaping T cell responses towards the Th2 phenotype while inhibiting proinflammatory cytokine production. Furthermore, VIP-producing enteric neurons contribute to enteric mucosal barrier homeostasis, maintaining optimal gut pH, and orchestrating the delicate balance between beneficial bacteria colonization and resistance against opportunistic pathogens. The potential impact of VIP production by host cells on GvHD regulation remains unclear. Our hypothesis proposes that the absence of VIP production by transplant recipients may exacerbate GvHD, either through increased T cell allo-reactivity, or via secondary modulation of the gut microbiome resulting from submucosal inflammation. Methods and Results: We investigated how endogenous production of VIP by host tissues on GvHD modulation in three murine allo-BMT models, where C57BL/6 wildtype (WT) and VIP-knockout (VIPKO) mice were subjected to 11Gy irradiation on day 0 and subsequently transplanted intravenously with 5x10 6 T cell depleted (TCD) bone marrow (BM) in combination with 0, 1x10 6, or 3x10 6 T cells from MHC mismatched B10.BR, Balb/c or FvB donors. In all three models, WT recipients had better survival than VIPKO recipients. Specifically, in B10BR B6, Balb/c B6 and FvB B6 model, the addition of 3 million donor T lymphocytes to the graft resulted in significantly more GvHD-related mortality in VIP-KO recipients compared with WT recipients, with 0% vs 82%, 18% vs 96%, and 0% vs 78% survival (p&amp;lt;0.01) among WT vs VIP-KO recipients. To elucidate the role of host cell-derived VIP production in T cell alloreactivity and GvHD exacerbation, we performed cytokine profile analysis using both serum and donor-derived lymphocytes from the spleen of transplant recipients. For serum cytokine analysis, VIP-KO recipients had significantly higher levels of IFNγ (p&amp;lt;0.05), TNFα (p&amp;lt;0.05) and IL-1beta (p&amp;lt;0.05) on day 7 post allo-BMT, and significantly higher levels of and IL-6 (p&amp;lt;0.01) and IL-10 (p&amp;lt;0.05) on day 14 post allo-BMT compared with WT recipients. For donor-derived lymphocytes cytokine analysis, we observed significantly increased Th1 and Th17 cytokines in VIP-KO recipient, with significantly higher frequencies of CD4+ IL2 (p&amp;lt;0.01) and CD4+ IFNγ (p&amp;lt;0.001) on day 4, significantly higher levels of CD4+ TNFα (p&amp;lt;0.05) and CD8+ TNFα (p&amp;lt;0.001) on day 19, and significantly increased CD8+ TNFα (p&amp;lt;0.05) on day 25 post allo-BMT compared to WT recipients. Immunofluorescence whole-mount imaging section staining demonstrated detectable VIP expression in enteric neurons innervating the entirety of the intestine in FvB VIP-GFP transplant recipients. Notably, VIP immunofluorescent signal was much higher in allo-BMT recipient intestines compared to syngeneic transplant recipient intestines, demonstrating immunomodulation of VIP-producing enteric neurons in response to local (GVHD-targeted organ) and systemic inflammation (Fig 1). To investigate the gut microbiome profile, we performed 16S ribosomal RNA sequencing and analyzed stool samples from three distinct groups: 1) naïve WT and VIP KO mice, 2) WT and VIPKO mice after co-housing, and 3) WT and VIPKO mice after co-housing followed by allo-BMT. Uniform Manifold Approximation and Projection high-dimensional cluster analysis revealed that VIP-KO mice exhibited a distinct gut microbiome compared to WT mice, regardless of their co-housing or allo-BMT conditioning (Fig 2A, 2B). The heatmap analysis demonstrated that VIP-KO mice had increased lactobacillus species (Fig 2C) and significantly decreased intestinal pH levels (p&amp;lt;0.01) compared to WT mice. Moreover, VIP-KO mice displayed significantly decreased gut microbiome diversity, indicating an inflammatory effect in the gut that influences changes in the microbiome composition. Conclusion: Overall, our findings highlight the crucial role of VIP in regulating immune responses and gut microbiome homeostasis, which could have implications for developing strategies to mitigate GvHD in allo-BMT recipients.

Cell-Free DNA (cfDNA)-Based Measurable Residual Disease (MRD) Detection As a Predictor of Relapse Post-Allogeneic Blood or Marrow Transplant (alloBMT) in Patients with Myeloid Malignancies

Background: AlloBMT is the only curative therapy for many patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). With the significant improvement in non-relapse mortality, disease relapse remains the main cause of therapeutic failure. The presence of leukemia-specific somatic mutations provides an attractive strategy for molecular assessment of measurable residual disease (MRD). The level of MRD at the time of complete remission (CR) has been an important predictor of relapse in patients undergoing alloBMT. Widely utilized molecular MRD assays require invasive bone marrow (BM) aspiration to ensure an adequate amount of residual leukemia cells. Circulating tumor DNA (ctDNA) released from malignant cells can be reliably detected in patients' plasma and has been widely used in diagnosis, early detection, and disease burden monitoring in solid tumors. This assay is not widely used in myeloid malignancies as the proper assessment of its technical and clinical utility as an MRD monitoring strategy has not been performed. In the current study, we aimed to determine the differential sensitivity between bone marrow and ctDNA-based MRD and its utility as a minimally invasive diagnostic and disease monitoring tool. Additionally, we evaluated the clinical utility of dynamic ctDNA-based MRD monitoring and its effect on post-alloBMT relapse by using plasma samples from 2 large prospective clinical trials (BMT CTN 0201 and 0402). Methods: The technical validation cohort included 20 consecutive patients with myeloid malignancies undergoing alloBMT at Johns Hopkins. A minimum of 10mL of plasma and bone marrow DNA was collected at the time of remission and at multiple time points after alloBMT. The clinical validation cohort included 90 patients participating in two prospective alloBMT trials (BMT CTN-0201 and 0402). The eligibility criteria included the diagnosis of myeloid malignancy (72 AML, 18 MDS) and the minimum of 2mL of plasma available from day 0 and/or day 90 after alloBMT. Ultra-deep error-corrected targeted sequencing was performed on plasma and bone marrow-derived DNA. The variant allele frequency (VAF) per patient was represented by the maximum VAF of all detected mutations. Results: To measure the exact contribution of hematopoietic tissue to plasma cell-free DNA (cfDNA) we analyzed cfDNA in alloBMT recipients. Using donor and recipient-specific polymorphisms we demonstrated that after full donor hematopoietic reconstitution by standard chimerism assays (day +180), 94.6% (range 93.9-95.3%) of cfDNA was of hematopoietic origin. To determine the differential sensitivity of the cfDNA vs. BM-based MRD we focused only on leukemia-specific mutations found in the diagnostic specimens. The minimum level of detection was 0.03%. Forty leukemia-specific mutations were detected in the technical validation cohort. Out of 31 mutations seen in the BM, only 3 were not detected in cfDNA (3/31, 9.7%). On the contrary, out of 37 mutations in cfDNA, 9 were not seen in BM (9/37, 25%). While we observed a strong positive correlation (rho = 0.8, p<0.0001) at any level of MRD between BM and cfDNA, cfDNA was superior at the lower MRD level (VAF cut-off < 0.26%). The clinical utility of cfDNA MRD was utilized in a separate validation cohort of 90 patients enrolled on 2 BMT CTN trials. The median (Q1, Q3) follow-up was 26 months (25 - 27 months). cfDNA-MRD positivity (VAF cutoff >0.25%) at day +90 was associated with significantly worse relapse-free survival (RFS) (46.2% vs 84.0%, p<0.001) and overall survival (OS) at two years (53.8% vs 87.2%, p=0.0013) after alloBMT. Conclusions: cfDNA-based MRD assessment appears to be a promising, minimally invasive test that provides important prognostic information in patients with myeloid malignancies undergoing alloBMT. The presence of cfDNA MRD at day 90 post alloBMT was associated with significantly worse RFS and OS in patients with myeloid malignancies. The dynamic assessment of MRD by cfDNA may not only provide important prognostic information but may also guide the therapeutic decision-making and timely implementation of post-alloBMT maintenance therapies to improve outcomes of patients with myeloid malignancies. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

BAFF promotes heightened BCR responsiveness and manifestations of chronic GVHD after allogeneic stem cell transplantation

Patients with chronic graft-versus-host disease (cGVHD) have increased B cell–activating factor (BAFF) levels, but whether BAFF promotes disease after allogeneic bone marrow transplantation (allo-BMT) remains unknown. In a major histocompatibility complex–mismatched model with cGVHD-like manifestations, we first examined B-lymphopenic μMT allo-BMT recipients and found that increased BAFF levels in cGVHD mice were not merely a reflection of B-cell number. Mice that later developed cGVHD had significantly increased numbers of recipient fibroblastic reticular cells with higher BAFF transcript levels. Increased BAFF production by donor cells also likely contributed to cGVHD, because BAFF transcript in CD4+ T cells from diseased mice and patients was increased. cGVHD manifestations in mice were associated with high BAFF/B-cell ratios and persistence of B-cell receptor (BCR)–activated B cells in peripheral blood and lesional tissue. By employing BAFF transgenic (Tg) mice donor cells, we addressed whether high BAFF contributed to BCR activation in cGVHD. BAFF increased NOTCH2 expression on B cells, augmenting BCR responsiveness to surrogate antigen and NOTCH ligand. BAFF Tg B cells had significantly increased protein levels of the proximal BCR signaling molecule SYK, and high SYK protein was maintained by BAFF after in vitro BCR activation or when alloantigen was present in vivo. Using T cell–depleted (BM only) BAFF Tg donors, we found that BAFF promoted cGVHD manifestations, circulating GL7+ B cells, and alloantibody production. We demonstrate that pathologic production of BAFF promotes an altered B-cell compartment and augments BCR responsiveness. Our findings compel studies of therapeutic targeting of BAFF and BCR pathways in patients with cGVHD.

Cytomegalovirus in Adult Allogeneic Blood and Marrow Transplant Patients Before or Around the Period of Neutrophil Recovery: A Single-Center, Retrospective, Descriptive Study.

BackgroundFew reports exist on pre-engraftment cytomegalovirus (CMV) DNAemia in allogeneic blood or marrow transplant (allo BMT) recipients. We describe this clinical entity, its management, and the potential effect of 3 different quantitative CMV deoxyribonucleic acid (DNA) tests used during the 6-year study period. MethodsWe performed a retrospective, single-center study of allo BMT recipients from 2010 to 2015 who developed CMV DNAemia before neutrophil recovery (absolute neutrophil count [ANC] <1000 cells/mm3, “pre-engraftment CMV”) or who became neutropenic concomitant with detectable CMV DNA (“peri-engraftment CMV”). Clinical data were collected from the electronic medical record.ResultsAmong 1151 adult allo BMT patients, 73 developed CMV DNAemia before engraftment or while neutropenic after initial engraftment. Most patients were eventually treated (valganciclovir or ganciclovir, N = 68; foscarnet, N = 1); 4 were not treated. First CMV detection occurred at median day +12 (range, 0–48), but treatment was not started until median day +33 (range, 4–105) at median ANC of 760 cells/mm3. Six patients had peak viral loads >5000 IU/mL; none had tissue-invasive disease. One developed ganciclovir resistance. No significant differences were observed upon stratification by quantitative CMV DNA test. ConclusionsCytomegalovirus DNA was detected in 6.3% of pre- and peri-engraftment allo-HSCT patients. Ganciclovir derivatives were commonly used for treatment despite risk of neutropenia. Treatment was typically deferred until CMV DNA and ANC rose. With rare exceptions, this treatment strategy did not appear to have adverse clinical consequences with respect to acute CMV. Different CMV DNA quantification tests used performed similarly from a clinical perspective despite different analytical performance characteristics.

Transplanted Vasoactive Intestinal Peptide-Secreting Donor Plasmacytoid Dendritic Cells Migrate to Lymphoid Organs and GvHD Target Organs

Introduction: Donor plasmacytoid dendritic cells (pDCs) have important roles in the pathogenesis of graft versus host disease (GvHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Vasoactive intestinal peptide (VIP), a potent anti-inflammatory neuropeptide with pleiotropic effects, has shown promise in managing GvHD through induction of regulatory dendritic cells (Chorny, Blood 2006) and regulatory T cells (Delgado, JLB 2005). Our previous data showed that transplantation of hematopoietic stem cells (HSC) lacking VIP expression in enhanced Th1 polarization and antiviral immunity (Li, JI 2011). Whether VIP produced by pDCs plays a key role in preventing GvHD is the focus of the current study. Method: A reductive allo-HSCT model was established by transplanting purified populations of 5 x 103 hematopoietic stem cells (HSC), 5 x 104 pDCs and 106 T cells from B6 (H-2Kb) to MHC mismatched B10.BR (H-2Kk) recipients. Mice received either VIP knockout pDCs (experimental group) or wild-type pDCs (control). Survival and clinical GvHD scores were monitored for at least 60 days. Serum pro-inflammatory cytokines were measured by luminex on day 3 and day 8. Additional recipients were transplanted for interim analyses of GvHD pathology in histological sections of small intestine and colon on day 8 and day 15. To investigate pDC migration and proliferation status, 5 x 104 luciferase-expressing pDCs, together with 5 x 103 WT B6 HSC, were transplanted to MHC mismatched Balb/c (H2kd) recipients. Bioluminescent imaging was performed on day 14. We used mice carrying an eGFP reporter linked to a VIP promoter to measure VIP production by donor cells in vivo. The construct has the eGFP reporter gene, followed by a polyadenylation sequence, inserted into BAC clone RP23-25A8 at the initiating ATG codon of the first coding exon of the VIP gene, so that eGFP expression is driven by the regulatory sequences of the BAC gene. The BAC is inserted randomly in the genome. We used serial breeding to establish the eGFP-VIP reporter in B6 mice. Bone marrow pDCs were sorted from transgenic eGFP-VIP B6 mice and used in transplant experiments, allowing VIP gene activity from donor-derived pDCs to be detected as GFP expression using confocal microscopy. Results: Compared to recipients of WT pDCs, recipients of VIP-KO pDCs have a significantly lower survival rate with increased GvHD clinical score (Figure 1; *p&lt;0.05). Transplant recipients of VIP-KO donor pDC had higher serum pro-inflammatory cytokine levels on day 3 and day 8 respectively. (Figure 2). At 2 weeks post-transplant donor pDCs migrated primarily to the spleen, with limited presence of donor pDCs in the gastrointestinal tract and lung (Figure 3). Compared to mice transplanted with WT pDCs, recipients of VIP-KO pDCs had increased donor T cell migration to the lung and paw, which are chronic GvHD target organs after two weeks. (Figure 4; *p&lt;0.05). When stimulated with IFN-g, eGFP-VIP transgenic pDCs have activation of the VIP gene promoter in vitro, as detected using eGFP (Figure 5). Confocal microscopy data showed that pDCs have activated the VIP promoter and secreted more VIP following stimulation by IFN-g, consistent with VIP up-regulation by the inflammatory mileu in allo-transplant recipients. Conclusions: These data show that lack of VIP secretion by donor pDCs resulted in lower survival rates and higher GvHD scores in allo-BMT recipients. Recipients of VIP KO donor pDC had increased donor T cell proliferation in GvHD target organs and increased GvHD clinical signs. The major site of donor pDCs migration in recipients is spleen, suggesting that pDCs may interact with other lymphoid cells, such as T cells, inducing a more tolerogenic environment. Donor pDCs were also found, to a lesser extent, in the lung and gastrointestinal tract, which are both GvHD target organs. Thus, pDCs may protect recipients from GvHD by limiting the activity of alloreactive T cells at these sites. The mechanism by which VIP-secreting donor pDCs regulate GvHD will be discussed. Disclosures Waller: Amgen: Consultancy; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Other: Travel expenses, Research Funding; Cerus Corporation: Other: Stock, Patents &amp; Royalties; Chimerix: Other: Stock; Cambium Oncology: Patents &amp; Royalties: Patents, royalties or other intellectual property ; Kalytera: Consultancy.

Solid Tumor-Induced Immune Regulation Alters the GvHD/GvT Paradigm after Allogenic Bone Marrow Transplantation.

Growth of solid tumors is often associated with the development of an immunosuppressive tumor microenvironment (TME). It has been suggested that the influence of the TME may extend beyond the local tumor and results in systemic immunosuppression. Here, we utilize two murine cancer models to explore the influence of solid tumors on the occurrence of alloreactivity-driven GvHD and graft-versus-solid tumor (GvT) effects following MHC-mismatched allogeneic bone marrow transplantation (allo-BMT). Melanoma- or colon carcinoma-bearing C57BL/6 mice did not develop GvHD after BMT even when the bone marrow inoculum was supplemented with donor-type splenocytes. This protection against GvHD required the presence of tumors because its resection prior to allo-BMT promptly resulted in development of GvHD. In addition, tumor-bearing mice given T-cell-depleted allo-BMT (allo-TCD-BMT) failed to develop GvHD and also showed significantly stronger GvT effects than mice given allo-BMT. The GvT effects in allo-TCD-BMT recipients were associated with profound changes in tumor-infiltrating cells compared with that in allo-BMT recipients, with significantly reduced donor-derived regulatory T cells (Treg), increased cytotoxic effector (IFNγhi) CD8 T cells, and increased M1 macrophages (iNOShi, arginaselo, and IL10lo); the use of macrophage-depleted bone marrow abrogated the GvT effects. Collectively, these results indicate that the presence of M1 macrophages may disrupt the generation of donor-type Treg cells so that the immunomodulatory effect of the TME can affect systemic immunity. SIGNIFICANCE: These findings show that cells such as T cells or macrophages in the bone marrow inoculum may interfere with the systemic and local immune reactivity against tumors.

Vasoactive intestinal polypeptide synthesis by somatic tissues in transplant recipients limits GVHD in allo-BMT

Abstract Both donor and recipient hematopoietic cells regulate GvHD in allo-BMT, but the role of somatic host tissues in the induction and maintenance of GvHD is less clear. Vasoactive Intestinal peptide (VIP) is a neuropeptide with potent immune-regulatory activity. Our goal was to examine the role of VIP synthesis by recipient tissues on donor T cell activation and GvHD in allo-BMT. We used MHC mis-matched allo-BMT models, B10.BR--&amp;gt;B6 and Balb/c--&amp;gt;B6, irradiating WT and VIP-KO B6 mice with 11Gy on day −1 and transplanting with 5 × 10E6 T cell depleted (TCD) bone marrow (BM) plus 0, 1 × 10E6, or 3 × 10E6 T cells from MHC mis-matched donors. B6--&amp;gt;B6 radiation chimeras lacking VIP in either hematopoietic or non-hematopoietic cells were created by performing B6 WT--&amp;gt;VIP-KO and VIP-KO--&amp;gt;WT syngeneic transplants. Production of VIP in transplant recipients was visualized with immune-fluorescent staining of tissues using an anti-VIP antibody. Transplanting TCD BM plus 3 × 10E6 donor T cells from B10.BR or Balb/c donors in VIP KIO mice led significantly increased GVHD-mortality compared with WT recipients. Donor-derived T cells in VIP-KO recipients had significantly higher Th1 and Th17 polarization. B10.BR--&amp;gt;B6 second allogeneic transplantation of radiation chimeras caused lethal GVHD mortality in VIP-KO-- &amp;gt;VIP-KO and WT--&amp;gt;VIP-KO mice, but not in WT--&amp;gt;WT or VIP-KO--&amp;gt;WT B6 mice, demonstrating the protective effect of VIP was due to synthesis by somatic recipient cells. Immunofluorescent imaging of allo-BMT recipients showed up-regulation of VIP in lungs post-transplant and high VIP production within neurons innervating the lungs. Short-term administration of VIP from D0 to D10 prevented GvHD-mortality in VIP-KO recipients transplanted BM &amp; T cells.

The Blood and Marrow Transplant Time out: Application of Universal Protocol As a Model to Improve Patient Selection-Suitability to Improve Allogeneic 1-Year Survivals

Topic Significance & Study Purpose/Background/Rationale Patients being considered for allogeneic Blood and Marrow Transplant (Allo-BMT) undergo rigorous evaluations of which can influence Allo-BMT plan and outcomes. The goal of this project was to improve the patient selection process prior to admission for Allo-BMT. The process was modeled after the Universal Protocol (UP) used prior to surgery. UP is a National Patient Safety Goal, supported by Joint Commission, Institute of Medicine, and World Health Organization. Our center aimed to standardize the approach to determine patient suitability and treatment plan with the goal of improving 1-year survival. A UP approach was employed prior to BMT to secure consensus on patient suitability and treatment plan. Methods, Intervention, & Analysis A 1-page comprehensive BMT-Time Out (BMT-TO) checklist was created with critical domains selected to provide programmatic workflow (Figure 1). All Allo-BMT recipients underwent systematic BMT-TO reviews through progressive handoffs beginning July 2016. Checklists were audited monthly for completeness and adherence to standard operating procedures. Blood Marrow Transplant Information Patient System (bmTIPS™) generated Allogeneic Transplant Intake Dashboard (ATID) reports, providing real-time numerical data on intake distribution patterns (Figure 2). The 2017 Allo-BMT 1-year survival data reported by Center for Blood and Marrow Transplant Research(CIBMTR) were used to set confidence levels and forecast CY18,19 projected 1-year survivals. Findings & Interpretation ATID for calendar years (CY)13,14, and 15, showed acceptance rates at presentation of 92%(n=98), 94%(n=105), and 77%(n=89), respectively. ATID for CY16, 17, 18 after implementation of BMT-TO showed lower acceptance rates: 83%(n=89), 86%(n=92), 85%(n=72), respectively. CIBMTR results showed improvement in our center's Allo-BMT 1-year survival rates with use of BMT-TO checklist (Figure 3): CY14(64%/reported); CY15=64%(n=45/reported); CY16=78%(n=28/reported); CY17=89%(n=37/predicted). Discussion & Implications Implementation of systematic review of patient selection-suitability data and BMT treatment plans using structured BMT-TO checklist before Allo-BMT, correlated with improved allogeneic 1-year survivals. Further evaluation and validation of the BMT-TO form is warranted.

Indoles derived from intestinal microbiota act via type I interferon signaling to limit graft-versus-host disease

AbstractThe intestinal microbiota in allogeneic bone marrow transplant (allo-BMT) recipients modulates graft-versus-host disease (GVHD), a systemic inflammatory state initiated by donor T cells that leads to colitis, a key determinant of GVHD severity. Indole or indole derivatives produced by tryptophan metabolism in the intestinal microbiota limit intestinal inflammation caused by diverse stressors, so we tested their capacity to protect against GVHD in murine major histocompatibility complex–mismatched models of allo-BMT. Indole effects were assessed by colonization of allo-BMT recipient mice with tryptophanase positive or negative strains of Escherichia coli, or, alternatively, by exogenous administration of indole-3-carboxaldehyde (ICA), an indole derivative. Treatment with ICA limited gut epithelial damage, reduced transepithelial bacterial translocation, and decreased inflammatory cytokine production, reducing GVHD pathology and GVHD mortality, but did not compromise donor T-cell-mediated graft-versus-leukemia responses. ICA treatment also led to recipient-strain-specific tolerance of engrafted T cells. Transcriptional profiling and gene ontology analysis indicated that ICA administration upregulated genes associated with the type I interferon (IFN1) response, which has been shown to protect against radiation-induced intestinal damage and reduce subsequent GVHD pathology. Accordingly, protective effects of ICA following radiation exposure were abrogated in mice lacking IFN1 signaling. Taken together, these data indicate that indole metabolites produced by the intestinal microbiota act via type I IFNs to limit intestinal inflammation and damage associated with myeloablative chemotherapy or radiation exposure and acute GVHD, but preserve antitumor responses, and may provide a therapeutic option for BMT patients at risk for GVHD.

A New Cellular Immunotherapy for the Treatment of Multiple Myeloma without Development of Graft-Versus-Host Disease

Multiple myeloma (MM) is a malignant plasma cell disorder that accounts for approximately 10% of all hematological cancers. Despite recent advances, long-term survival is rare after autologous bone marrow transplantation (auto-BMT) and/or treatment with recently introduced anti-myeloma agents and disease recurs in virtually all patients. Allogeneic bone marrow transplantation (allo-BMT) is an effective treatment that can provide partial or complete remission in patients with MM. The therapeutic potential of allo-BMT is attributed to the “graft-versus-myeloma” (GvM) effect that aims to destroy residual tumor cells that survived an induction protocol of chemotherapy/radiotherapy and to maintain immune surveillance to prevent relapse. However, allo-BMT remains a controversial treatment, since the donor T cells that mediate the GvM effect are also the source of the cells that react to other tissue alloantigens and induce graft versus-host disease (GvHD), a major cause of morbidity and mortality in allo-BMT recipients. Nonetheless, allo-BMT remains the only potentially curable treatment for MM. Recently TCR Vβ CDR3-size spectratype analyses in an animal model of MM identified T cells subfamilies involved in the anti-host and anti-tumor reactivity. We have now carried this work further and tested the potential of integrating auto-BMT with a donor lymphocyte infusion (DLI) composed only of anti-MM reactive Vβ 2, 3 and 8.3 T cell subfamilies. The results demonstrate that these T cell subsets are indeed involved in the generation of a potent GvM response in MM bearing mice and is associated with enhanced survival. Importantly, the GvM response was not accompanied by the development of GvHD. Nonetheless, the GvM response was not sufficient to completely inhibit relapse. We then pre-stimulated donor T cells with MM cells in vitro in the presence of co-stimulatory factors and found that our selective DLI protocol induced a vigorous and long-lasting GvM which translated into long-term survival, again in the complete absence of GvHD. Interestingly, quite similar results were obtained by treating MM-bearing mice with repeat doses of naïve donor Vβ 2, 3 and 8.3 T cell subfamilies. The treated mice showed significantly lower serum paraprotein levels and myeloma infiltration in bone marrow and spleen. Taken together, the results suggest that a transplantation protocol involving only tumor cell-reactive donor T cell subfamilies might be devised for MM patients that results in a potent GvM with enhanced survival but without symptoms of GvHD. DisclosuresNo relevant conflicts of interest to declare.

Vasoactive Intestinal Peptide Synthesis in Transplant Recipients Critically Regulate Donor T Cell Alloreactivity and Graft-Versus-Host Disease

Introduction: Vasoactive Intestinal peptide (VIP) is a neuropeptide expressed by neural and lymphoid cells with pleiotropic effects on brain, immune, pulmonary and GI systems. The potent immunomodulatory activity of VIP make it an attractive therapeutic target to suppresses immune responses in conditions of deleterious inflammation. In allogeneic bone marrow transplantation (allo-BMT), graft-versus-host disease (GvHD) is mediated by activated alloreactive T cells that also upregulate a variety of co-inhibitory pathway molecules, including VIP expression on T cells. It is unclear whether VIP production by host cells in the transplant recipient influence the severity of GvHD and donor T cell activation. We tested the hypothesis that lack of VIP production by recipient cells would enhance donor T cell activation and increase GvHD in allo-BMT recipients. Methods and Results: We examined the effect of endogenous VIP signaling in host tissues in modulating GvHD in a murine allo-BMT model, in which C57BL/6 wildtype (WT) and VIP-knock-out (VIP-KO) mice received 11Gy irradiation on day -1 and were transplanted i.v. on day 0 with 5x106 T cell depleted (TCD) bone marrow (BM) plus 0, 1x106 or 3x106 splenocytes (SP) from MHC mis-matched B10.BR donors. All three groups of WT recipients had better survival than VIP-KO recipients. VIP-KO mice transplanted with TCD BM alone had 33.3% survival at day 75 compared with 73.3% survival of WT recipients. Chimerism studies on day 20 post-BMT showed equivalent levels of donor chimerism within each group that received the same dose of donor SP. WT and VIP-KO recipients received BM only showed significantly lower donor chimerism compared to either WT or VIP-KO recipients received additional SP (p<0.001), indicating increased mortality in recipients transplanted with BM only was primarily due to early graft rejection. The addition of donor SP to the graft resulted in significantly more GvHD-related mortality in VIP-KO mice compared with WT recipients, with 80% vs 100% day 75 survival among recipients of 1x106 SP (p=NS) and 0% vs 40% survival (p<0.01) among recipients of 3x106 donor SP. WT and VIP-KO mice treated with lethal doses of irradiation without allo-BMT showed no survival difference, suggesting VIP-KO mice are not intrinsically more susceptible to irradiation. To determine how the absence of VIP synthesis by VIP-KO host cells modulates immune effector mechanisms that contributes to increased GvHD, we analyzed donor-derived lymphocytes in the spleen of transplant recipients received 5M TCD BM plus 3 × 106 SP from WT B10BR donors following a time course, with flow cytometry. WT recipients had significantly increased levels of Lag3+ CD4 (p<0.001) on D6, PD1+ CD8 (p<0.01) on D13, Tim3+ CD4 (p<0.05) on D19, and increased levels of Tim3+ CD4 (p<0.001), PD1+ CD4 (p<0.05), and PD1+ CD8 (p<0.001) on D25 post allo-BMT compared to VIP-KO recipients. In addition, VIP-KO recipients had significantly higher expression of selected Th1 and Th17 cytokines. We observed significantly higher levels of IFN-γ+ CD4 (p<0.001) and IL2+ CD4 (p<0.01) on D4, significantly higher levels of TNF-α+ CD4 (p<0.05) and TNF-α+ CD8 (p<0.001) on D19, and significantly increased TNF-α+ CD8 (p<0.05) on D25 post allo-BMT in VIP-KO compared to WT recipients. Consistent with the significant elevation of IL17 in CD4 and CD8 (p<0.001) donor T cells of VIP-KO mice on D19, increased transcriptional factor RORγt in VIP-KO mice were detected in both CD4 (p<0.0001) and CD8 (p<0.0001) donor T cells. Transplant experiments using radiation chimeric recipients in which either the hematopoietic or non-hematopoietic compartment lacked expression of VIP further demonstrated that VIP production in non-hematopoietic cells is the key factor in limiting the GvHD activity of donor T cells (Fig 1). Finally, immunofluorescent imaging of transgenic mice in which GFP is regulated by the VIP promoter showed VIP production is highly expressed in efferent neurons innervating the lungs, and that host expression of VIP in lung tissues continues for at least 15 days post-transplant (Fig 2). Conclusion: These data suggest local production of VIP in the lung may be a key factor in the control of alloreactive donor T cells and subsequent GVHD in epithelial target organs. Furthermore, local production of VIP by host neurons surrounding lung alveoli suggests a novel pathway for pharmacological modulation of allo-reactive T cells and control of GVHD. DisclosuresWaller:Pharmacyclics: Other: Travel Expenses, EHA, Research Funding; Kalytera: Consultancy; Novartis Pharmaceuticals Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celldex: Research Funding; Cambium Medical Technologies: Consultancy, Equity Ownership.

IL-10 gene-modified dendritic cells-induced type 1 T regulatory cells inhibit graft-versus-host disease while preserving graft-versus-leukemia effect

To explore the role of type 1 regulatory T (Tr1) cells in allogeneic bone marrow transplantation (allo-BMT), we generated Tr1 cells from naïve CD4+ T cells in donor mice that were subjected to stimulation of recipient interleukin (IL)-10 gene-modified dendritic cells (DCs). In animal models, the severity of graft-versus-host disease (GVHD) in the group co-infused with DC-induced Tr1 cells was markedly milder than that in the groups without infusion of Tr1 cells. Moreover, the co-infusion Tr1 cells significantly prolonged the leukemia-free survival in tumor-bearing mice. Besides, Tr1-induced cells mediated the shift from T helper (Th)1 to Th2 cytokines in allo-BMT recipients. In Conclusions, our findings suggest that IL-10 gene-modified DC-induced Tr1 cells could inhibit GVHD while maintaining Graft-versus-leukemia (GVL) through mediating the shift of Th1/Th2 cytokines in an allo-BMT mice model.

Therapeutic potential of tranilast for the treatment of chronic graft-versus-host disease in mice.

Chronic graft-versus-host disease (cGVHD) is a marked complication of hematopoietic stem cell transplantation, and multiple organs can be affected by cGVHD-induced inflammation and fibrosis. In clinical settings, immunosuppressive agents have been the last resort to treat cGVHD. However, it has been only partially effective for cGVHD. Hence, efficacious treatment of cGVHD is eagerly awaited. Our previous work suggested that oxidative stress was elevated in cGVHD-disordered lacrimal glands and that epithelial-to-mesenchymal transition (EMT) was implicated in fibrosis caused by ocular cGVHD. In addition, our recent article demonstrated that thioredoxin interaction protein (TXNIP) and transcription factor nuclear factor kappa-light-chain-enhancer of activated B cells (NF-𝛋B) were associated with the development of cGVHD. After our search for effective drugs, we chose tranilast to combat systemic cGVHD. Tranilast is known to (1) act as an inhibitor of the inflammatory molecules TXNIP and NF-κB and (2) exert anti-fibrotic, anti-EMT and anti-oxidative effects. To investigate the effectiveness of tranilast for cGVHD, we used an MHC-compatible, multiple minor histocompatibility antigen-mismatched murine model of cGVHD. Tranilast or a solvent-vehicle were orally given to the allogeneic bone marrow transplantation (allo-BMT) recipients from the day before allo-BMT (Day-1) to Day 27 after allo-BMT. Their cGVHD-vulnerable organs were collected Day 28 after allo-BMT and analyzed by using various methods such as histology, immunohistochemistry and immunoblotting. As indicated by our results, tranilast alleviated cGVHD-elicited inflammation and fibrosis by suppressing the expression and/or activation of TXNIP and NF-κB and preventing EMT. Taken together, although this strategy may not be a complete cure for cGVHD, tranilast could be a promising medication to ameliorate cGVHD-triggered disabling symptoms.

Oral health and dental morbidity in long-term allogeneic blood and marrow transplant survivors in Australia.

Oral and dental disease is a major cause of long-term morbidity following allogeneic blood and marrow transplantation (Allo-BMT). This study aimed to describe the extent and range of oral and dental complications in BMT recipients and to identify gaps in service provision provided to this high-risk group. Participants were Allo-BMT recipients, aged >18years, and received transplants between 2000 and 2012 in NSW. They completed seven surveys, the purpose-designed Sydney Post-BMT Study survey and six other validated instruments. Of 441 respondents, many reported dry mouth (45.1%), dental caries (36.7%), mouth ulcers (35.3%), oral GVHD (35.1%), gingivitis (16.2%), tooth abscess (6.1%) and oral cancer (1.5%). Regular dental visits were reported by 66.2% of survivors. Middle-high income, older age and geographic location showed a positive association with regular dental visits. Of those who did not visit the dentist regularly, 37% stated they did not feel it necessary, 36% reported cost and 20% stated it was not advised by the treating team. Despite oral complications commonly occurring after Allo-BMT, many survivors receive inadequate dental care. These results emphasize the need for improved oral health education, the importance of regular dental checks and improvement in the delivery of dental health services for BMT survivors.

Decline in Late Mortality after Allogeneic Blood or Marrow Transplantation (alloBMT) Performed in Childhood: a Report from the BMT Survivor Study-2 (BMTSS-2)

Background: AlloBMT is a curative option for children with malignant and non-malignant diseases. Nonetheless, the high intensity of therapeutic exposures at a young age increases the risk of organ compromise and may lead to premature death. However, there remains a gap in knowledge regarding cause-specific late mortality experienced by children undergoing alloBMT. Furthermore, it is not clear whether mortality rates have changed over the past 4 decades. We address these gaps by examining premature death (all-cause, relapse-related mortality [RRM] and non-relapse-related mortality [NRM]) in patients undergoing alloBMT between 1974 and 2010 at City of Hope, University of Minnesota or University of Alabama at Birmingham. Methods: To be included in this analysis, the patients had to have received alloBMT before age 22, and survived for ≥2y after transplantation. Vital status information was ascertained as of Dec 31, 2016, using medical records, the National Death Index (NDI), and Accurint databases. Information on cause of death was obtained from the NDI Plus program and medical records. Using Kaplan-Meier techniques, we describe overall survival conditional on surviving 2+ and 20+y from alloBMT. We describe cumulative incidence of RRM and NRM using competing risk methods. We also describe all-cause late mortality for patients transplanted over 3 time periods: <1990; 1990-1999; 2000-2010. Standardized mortality ratio (SMR) was used to compare the mortality experienced by this cohort to the age- sex-, and calendar-specific mortality of the US general population. Proportional subdistribution hazards model (Fine-Gray) for competing risks was used for identifying predictors of late mortality, including demographics, primary disease, conditioning agents, disease status at alloBMT and GvHD prophylaxis. Results: In this cohort of 1,411 recipients of alloBMT performed in childhood and surviving ≥2y, the most common primary diagnoses were acute lymphoblastic leukemia (ALL: 25%), acute myeloid leukemia/myelodysplastic syndrome (AML/MDS: 24%), inborn errors of metabolism (IEM: 14%) and severe aplastic anemia (SAA: 11%). The distribution of the cohort over the 3 time periods was: <1990 (23%); 1990-1999 (29%); 2000-2010 (48%). After a median follow-up of 14.9y, we observed 295 deaths in this 2y survivor cohort, yielding an overall survival rate of 79.6% at 20y from alloBMT. The leading causes of premature death were infection (32.0%), primary disease (24.6%) and subsequent malignant neoplasms (18.4%). Overall, the cohort was at a 15-fold increased risk for premature death (SMR 15.3, 95% CI 13.6-17.2), when compared with the general population. Conditional on surviving 20y, the 5y overall survival was 96.0% for the entire cohort, and 98.4% for SAA, 96.0% for AML/MDS, 90.3% for ALL and 85.8% for IEM. Relative mortality declined with time from alloBMT, but remained significantly elevated at 25y after transplantation (SMR 1.9, 95% CI 1.1-2.9). The cumulative incidence of NRM exceeded that due to RRM (fig 1); the 20y cumulative incidence of NRM and RRM were 13.0% and 4.4%, respectively. The all-cause 10y cumulative mortality rate declined over the 3 eras (<1990: 18.9%, 1990-1999: 12.5%; >2000: 10.7%, p=0.001) (Fig 2). Adjusting for demographic and clinical variables, as well as conditioning, the hazard ratio (HR) of all-cause death decreased with more recent treatment eras (<1990: HR=1.0; 1990-1999: HR=0.7, 95%CI, 0.5-0.9, p=0.01; 2000-2010: HR=0.6, 95%CI, 0.4-0.8, p=0.004). Conclusions: This study demonstrates that compared with the general population, 2y survivors of alloBMT performed in childhood remain at an elevated risk of premature death, even after 25y from alloBMT. While the risk of relapse-related mortality plateaus with time, non-relapse-related mortality continues to increase and remains the major cause of late death in this population. However, we make an encouraging observation i.e., there has been a significant decline in all-cause mortality experienced by children undergoing alloBMT over the past 4 decades. [Display omitted] DisclosuresArora:Takeda Oncology: Consultancy.

Integrating cancer survivorship care into allogeneic BMT recovery.

39 Background: AlloBMT patients are at risk for chronic morbidity and late effects. Prior to 2015, the Stanford BMT program conducted group classes at three months post-transplant to prepare alloBMT patients and their families for the transition to survivorship, but no formal cancer survivorship program existed. Cancer survivorship care is required by the Foundation for the Accreditation of Cellular Therapy ( FACT). In order to fill this gap, we developed a standardized pathway to survivorship care for alloBMT survivors. Aim: To implement an initial cancer survivorship care consultation with comprehensive symptom assessment, late effects monitoring, and development of a survivorship care plan (SCP) three to six months after alloBMT. Methods: Standard work was implemented including patient introduction to survivorship by the BMT nurses and Advanced Practice Providers [APP] and during group ‘moving home’ class; responsibility for referral to survivorship; metric tracking, and administration of patient survey. The survivorship visit included an evaluation by the patient’s BMT social worker, introduction to post-BMT nurse coordinator and consult with cancer survivorship APP. Eligible patients were alloBMT recipients receiving care in the outpatient setting. Results: From July 28, 2015 to January 25, 2016, 90% (66/72) of eligible alloBMT patients were referred to cancer survivorship by day 100, 66% (42/66) of referred patient s were seen in survivorship clinic by day 180, 33% (22/66) deferred the visit or were unreachable, 1% (2/66) died. Of the 42 patients who received a cancer survivorship visit, 38% (13/42) responded to a post-visit survey. Responses indicated the patients believed their health goals and needs were addressed, and a SCP in the context of a comprehensive visit was valuable. Conclusions: Integrating standardized cancer survivorship care into the alloBMT recovery period is feasible, satisfactory to patients, and adds a structured approach for long-term effects monitoring and treatment. Early experience and patient surveys demonstrate value in an individual visits 3-6 months after alloBMT with a focus on returning to life and goal oriented post-transplant care.

Tumor Antigen Vaccination of Donors to Augment Graft-Versus-Tumor Effects after Allogeneic Bone Marrow Transplantation with Post-Transplantation Cyclophosphamide

High dose, post-transplantation cyclophosphamide (PTCy) effectively prevents graft-versus-host disease after alloBMT by killing alloreactive T cells recently stimulated by recipient alloantigens. However, remotely primed (>4 days) T cells are resistant to cyclophosphamide-induced immunologic tolerance, raising the possibility that priming BMT donors with tumor antigens could be an effective method to augment anti-tumor immunity after alloBMT with PTCy without exacerbating GVHD.To test this hypothesis, we generated tumor antigen-specific T cells by vaccinating mice with chicken ovalbumin (OVA) expressing Listeria monocytogenes OVA (Lm-OVA) 7 or 30 days before syngeneic or allogeneic BMT, with or without PTCy of recipients bearing the EL4 thymic lymphoma expressing ovalbumin (EL4-mOVA). In a syngeneic BMT model, transferring vaccine primed day -7 or day -30 donor T cells into EL4-mOVA-Luci bearing mice prolonged survival but did not result in cure due to late relapses. To test whether this phenomenon can be found in an MHC matched, minor antigen mismatched alloBMT mouse model, we transferred vaccine primed or unprimed C3H.SW donor splenocytes into tumor bearing B6 recipients. Recipients of alloBMT without PTCy from vaccinated donors had a significant cure rate of 38% (day-30) and 23% (day -7) compared to 0% survival rate for mice receiving unprimed donor splenocytes. Interestingly, PTCy completely abrogated both GVHD and the anti-tumor benefit of donor vaccination, resulting in no prolongation of tumor-free survival of recipients of cells from primed donors compared to recipients of cells from unprimed donors.In light of the abrogation of anti-tumor immunity by PTCy, we tracked the fate of vaccine-primed T cells in alloBMT recipients. Donor cells primed on 7 or 30 days before T cell transfer showed significant numbers of OVA-specific CD8+ effector T cells on day 15 after transplantation in mice that did not receive PTCy. Although PTCy diminished vaccine primed tumor protection, we did detect increased percentages of tumor-specific effector CD8+ T cells in the blood as well as in other compartments on day 15 after transplantation.In an attempt to restore anti-tumor immunity we tested the effects of giving donor lymphocyte infusions (DLI) from tumor antigen-primed donors. DLI depleted of naïve T cells augmented anti-tumor immunity but animals died from graft-versus-host disease. In contrast, CD8-depleted DLI from vaccinated donors given after vaccine-primed BMT plus PTCy prolonged survival from EL4-mOVA without resulting in fatal GVHD. These results are consistent with the hypothesis that vaccine-primed CD8+T cells survive PTCy and their function can be elicited post-transplantation with help provided by tumor-specific CD4+ T cells administered as donor lymphocyte infusion. We plan to test this hypothesis in patients by vaccinating allogeneic BMT donors against a tumor antigen prior to allogeneic BMT + PTCy, with or without CD8-depleted DLI. DisclosuresNo relevant conflicts of interest to declare.