Immunobiology and immunotherapeutic implication of syneneic/autologous graft-versus-host disease

Summary: Administration of the immunosuppressive drug cyclosporine (CsA) after syngeneic/autologous bone marrow transplantation (BMT) elicits an autoimmune syndrome with pathology virtually identical to graft-vs-host disease (GVHD). The induction of this syndrome, termed syngeneic/autologous GVHD, is a two-tiered process requiring both the active inhibition of thymic-dependent clonal deletion and the elimination of mature T cells that have an immunoregulatory effect. Eradication of the peripheral immunoregulatory compartment by the preparative regimen provides a permissive environment for the activation of the syngeneic/autologous GVHD effector T cells. Although the repertoire of autoreactive effector T lymphocytes is highly conserved, these T cells promiscuously recognize MHC class II determinants. This novel specificity of the autoreactive lymphocytes appears to he dependent on the peptide derived from the MHC class 11 invariant chain. Recent studies also suggest that these promiscuous autoreactive T cells can effectively target and eliminate MHC class Il-expressing tumor cells. Administration of cytokines that upregulate the target antigen or expand the effector population can potentiate the antitumor activity of syngeneic/autologous GVHD, Although the induction of syngeneic/autologous GVHD is an untoward effect of CsA immunosuppression, mobilization of these autoimmune mechanisms provides a promising immunotherapeutic approach for certain neoplastic diseases.     

Induction of autologous graft-versus-host disease: results of a randomized prospective clinical trial in patients with poor risk lymphoma.

The results of blood or marrow transplantation in patients with chemorefractory aggressive lymphoma, that is, those not responding to conventional-dose chemotherapy at the time of transplant, have been poor. The relapse rate has been high after autologous bone marrow transplant, whereas allogeneic transplantation has been associated with excessive transplant-related toxicity. Administration of cyclosporine after autologous transplantation can induce an autoreactive syndrome that resembles graft-versus-host disease (GVHD). This syndrome, named autologous graft-versus-host disease, has clear antitumor activity in animal models that can be enhanced by the addition of cytokines such as gamma-interferon and interleukin-2. A randomized, prospective study was conducted to evaluate the antitumor effect of autologous graft-versus-host disease induced with cyclosporine, and augmented by the administration of gamma-interferon and interleukin-2 in patients with chemorefractory Hodgkin and aggressive non-Hodgkin lymphomas. Fifty-one patients were randomized, 24 to the autologous GVHD induction arm, and 27 to the noninduction arm after autologous transplant using mobilized peripheral blood stem cell (PBSC) grafts. There were no differences in treatment-related mortality, overall and event-free survival (OS, EFS) between both groups; however, in the induction arm, GVHD developed only in 4 patients. The administration of oral cyclosporine followed by interleukin-2 and gamma-interferon is generally not well tolerated, and does not appear to be an effective method to induce autologous GVHD in patients receiving autologous PBSC grafts.     

Repifermin (keratinocyte growth factor-2) reduces the severity of graft-versus-host disease while preserving a graft-versus-leukemia effect.

Graft-versus-host disease (GVHD) is the principal complication after allogeneic bone marrow transplantation (BMT). Reductions in systemic GVHD are frequently associated with a corresponding diminishment of the graft-versus-leukemia (GVL) response. In this study, we tested the effects of a novel recombinant human keratinocyte growth factor, repifermin (keratinocyte growth factor-2), on the induction of GVHD in a well-defined murine BMT model (B6 --> B6D2F1). Administration of repifermin (5 mg/kg/d) to allogeneic BMT recipients resulted in a significant decrease in both systemic GVHD and target organ histopathology. Repifermin treatment also reduced serum levels of tumor necrosis factor alpha and lipopolysaccharide compared with control mice. In contrast, repifermin did not affect T-cell proliferation, cytokine production, or cytotoxic responses to host antigens. When 2000 host-derived P815 (H-2(d)) leukemia cells were added to the bone marrow inoculum, repifermin preserved GVL effects and resulted in significantly delayed mortality compared with control-treated allogeneic BMT recipients. Collectively, these data suggest that repifermin administration may represent a novel strategy to separate the toxicity of GVHD from the beneficial GVL effects after allogeneic BMT.     

Dose and timing of interleukin (IL)-12 and timing and type of total-body irradiation: effects on graft-vs.-host disease inhibition and toxicity of exogenous IL-12 in murine bone marrow transplant recipients.

Paradoxically, a single injection of recombinant murine interleukin (IL)-12 on the day of bone marrow transplantation (BMT) inhibits graft-vs.-host disease (GVHD) while preserving graft-vs.-leukemia (GVL) effects in lethally irradiated mice receiving fully MHC-mismatched bone marrow and spleen cells. These protective effects are mediated by interferon (IFN)-gamma, whose early secretion is induced by IL-12 treatment. We investigated the relationship of IL-12 dose and timing of administration, as well as timing and type of total-body irradiation (TBI), with the ability of IL-12 to inhibit GVHD or mediate toxicity. The results show that a relatively low dose of IL-12 (as little as 50 U in a single injection) can mediate significant GVHD protection. The timing of IL-12 administration, however, is a critical factor. IL-12 administered 1 hour before BMT was most protective, but protection was still observed when it was administered 1-12 hours after BMT. Delaying IL-12 administration to 36 hours post-BMT completely obviated its protective effect. Administration of a second IL-12 injection 6 days after BMT negated the protective effect of an initial injection at the time of BMT. While IL-12 protection was evident when TBI was administered by 137Cs-irradiator in one or two fractions on day -1 or day 0, the use of an X-irradiator to deliver TBI on day -1 was associated with marked IL-12 toxicity. Whereas the protective effect of IL-12 against GVHD depended on donor-derived IFN-gamma, toxicity depended on the ability of host cells to produce IFN-gamma. Careful studies are warranted to test the effects of IL-12 in the context of BMT with various conditioning regimens in large animal preclinical models before this novel approach to GVHD protection can be applied clinically.     

Differential graft-versus-leukaemia effect by CD28 and CD40 co-stimulatory blockade after graft-versus-host disease prophylaxis

Co-stimulatory blockade may be a promising strategy for tolerance induction in transplantation. In allogeneic bone marrow transplantation (BMT) for leukaemia treatment, however, preservation of the graft-versus-leukaemia (GVL) effect is another critical requirement for clinical application. In this study, we have compared the effect on GVL of using CD28 and CD40 co-stimulatory blockades as graft-versus-host disease (GVHD) prophylaxis in a murine allogeneic BMT model with simultaneous transfer of BCL1 leukaemia. Despite the relative improvement of GVHD as assessed by survival and body weight in both treatment regimes, treatment with anti-CD154 moAb clearly diminished the GVL effect, whereas treatment with anti-CD80 and CD86 MoAbs maintained this effect. Although T cell-mediated effector function at 14 days post-BMT assessed by IFNgamma expression and cytotoxicity against host alloantigen was comparable between both co-stimulatory blockades, IL-12 mRNA expression was preferentially reduced by CD40 blockade. Our results suggest the differential involvement of the CD28 and CD40 co-stimulatory pathways in the development of GVHD and GVL effects. CD28 blockade may be a favourable strategy for tolerance induction in leukaemia patients undergoing BMT.     

NK inhibitory-receptor blockade for purging of leukemia: effects on hematopoietic reconstitution.

One of the obstacles of BMT that limits its efficacy is failure to eradicate the original tumor. The incidence of tumor relapse is particularly high after autologous BMT. Natural killer (NK) cells comprise various subsets that can express inhibitory receptors for MHC class I determinants. We have recently demonstrated that blockade of NK-cell inhibitory receptors can augment antitumor effects in vitro and in vivo. However, breakdown of tolerance and autoreactivity may occur as a result of the inhibition of NK-cell inactivation to self MHC determinants. We have utilized F(ab')2 fragments of monoclonal antibody, 5E6, against Ly49C/I inhibitory receptors, which are expressed on 35% to 60% of NK cells in H2b strains of mice and are specific for H2Kb, to investigate the effect of inhibitory-receptor blockade on syngeneic bone marrow cell (BMC) and tumor cell growth. We show that treatment of interleukin 2-activated C57BL/6 (B6, H2b) SCID-mouse NK cells with 5E6 F(ab')2 fragments during 48-hour coculture resulted in autoreactivity against syngeneic BMCs as demonstrated by suppression of myeloid reconstitution on day 14 post-BMT. However, this suppressive effect was transient and normalized by day 21 post-BMT. In contrast, blockade of inhibitory receptors during 24-hour coculture had no adverse effects on myeloid reconstitution after BMT. Furthermore, under the same coculture conditions, NK cell-mediated purging of C1498 leukemia cells contaminating syngeneic BMCs was more effective with inhibitory-receptor blockade, leading to a significantly higher proportion of animals with long-term survival compared to the control recipients. These results demonstrate that short-term in vitro blockade of inhibitory receptors can augment antitumor activity without long-term inhibitory effects on BMCs and thus may be of potential use in the purging of contaminating tumor cells prior to autologous BMT.     

The anti-tumor effect of allogeneic bone marrow/stem cell transplant without graft vs. host disease toxicity and without a matched donor requirement?

The anti-tumor immune response that occurs in allogeneic bone marrow/stem cell transplant (BMT) settings is capable of eradicating tumors that are resistant to chemotherapy/radiation treatment. This anti-tumor immune response, known as the graft vs. tumor (GVT) effect, is the most effective immunotherapy treatment ever discovered. Unfortunately, the clinical application of GVT is severely limited due to the intimate association of GVT with the extremely toxic and often lethal side-effect known as graft vs. host disease (GVHD). It is a major research focus in the field of BMT to develop methods to separate the beneficial GVT effect from the detrimental GVHD toxicity. However, due to the intimate association of these effects, attempts to limit GVHD also have a tendency to limit the GVT effect. We propose a new concept for harnessing the power of the GVT effect without the toxicity of GVHD. Rather than trying to separate GVT from GVHD, we propose that these naturally coupled effects can 'mirrored' onto the host immune system and maintain their intimate association. The 'mirror' of GVHD is a host rejection of a graft (HVG). As rejection of an allograft would not be toxic, an HVG effect coupled to a host vs. tumor (HVT) effect, the 'mirror' of the GVT effect, would provide the anti-tumor effect of BMT without GVHD toxicity. In the 'mirror' setting, the HVT effect must occur against syngeneic tumors, while in the BMT setting the GVT effect occurs in the allogeneic setting. Previous attempts to elicit syngeneic anti-tumor immunity using therapeutic tumor vaccines have had disappointing results in the clinic due to the influence of tumor immunoavoidance mechanisms. We propose that the 'danger' signals that are released as a result of GVHD in the allogeneic BMT setting serve as an adjuvant to the GVT effect disabling tumor immunoavoidance. The chemotherapy/radiation conditioning prior to transplant is a required initiating event to the coupled GVT/GVHD effects. The conditioning releases 'danger' signals that mediate this adjuvant effect. To imitate this immunological event in immunocompetent, non-conditioned patients we propose that infusion of freshly activated, polyclonal CD4+ memory Th1 cells which express CD40L on the cell surface will stimulate a HVT/HVG 'mirror' effect, providing a non-toxic means to elicit the effective immune-mediated anti-tumor effect of BMT without the GVHD toxicity and without the requirement for a matched donor.     

Graft-versus-host disease and the Th1/Th2 paradigm

Graft-versus-host disease (GVHD) is the major complication after allogeneic bone marrow transplantation (BMT) and is initiated by alloreactive donor T cells recognizing foreign histocompatibility antigens of the host. There is now substantial experimental and clinical evidence to implicate a dysregulation of cytokine networks as a primary cause for the induction and maintenance of GVHD. In this article, current knowledge of the involvement of cytokines in GVHD is reviewed. The balance between type 1 cytokines (interleukin-2, interferon-γ) and type 2 cytokines (interleukin-4, interleukin-10) is hypothesized to govern the extent to which a cell-mediated immune response and a systemic inflammatory response develop after allogeneic BMT. Because type 2 cytokines can inhibit the production of the proinflammatory cytokines interleukin-1 and tumor necrosis factor-α, a type 1 to type 2 shift in the initial response of donor T cells to host alloantigens may interrupt the cytokine cascade after allogeneic BMT and may offer a new approach to the prevention and treatment of acute GVHD. Interventions to specifically eliminate or modify the response of donor T cells to alloantigens in order to reduce GVHD may obviate the need for T cell depletion in clinical BMT and thus avoid the increased risk of relapse of malignancy and impairment of donor cell engraftment.     

Regulatory dendritic cells protect mice from murine acute graft-versus-host disease and leukemia relapse

We have established a novel immunotherapeutic approach involving dendritic cells (DCs) with potent immunoregulatory property (designated as regulatory DCs [rDCs]) for acute graft-versus-host disease (GVHD) and leukemia relapse in allogeneic bone marrow (BM) transplantation (BMT) in mice bearing leukemia. rDCs displayed high levels of MHC molecules and extremely low levels of costimulatory molecules. A single injection of rDCs following allogeneic BMT controlled the ability of the transplanted T cells to induce acute GVHD and graft-versus-leukemia (GVL) effect in the recipients bearing leukemia, and that resulted in protection from the lethality caused by acute GVHD and tumor burden. Thus, the use of rDCs may be therapeutically useful for the treatment of acute GVHD and leukemia relapse in allogeneic BMT.     

Idiopathic pneumonia syndrome after bone marrow transplantation: the role of pre-transplant radiation conditioning and local cytokine dysregulation in promoting lung inflammation and fibrosis

Pulmonary complications and graft-vs.-host disease (GVHD) remain severe threats to survival after bone marrow transplantation (BMT). Idiopathic pneumonia syndrome (IPS) accounts for nearly 50% of all the cases of interstitial pneumonitis after BMT. IPS is characterized by an early inflammatory phase followed by chronic inflammation and fibrosis of lung tissue; however, the immunopathogenesis of this disease is not yet clearly understood. This biphasic syndrome has been reported to be associated with pre-transplant radiation conditioning in some studies while others have suggested that GVHD or autoimmune phenomena may be responsible for its development. The early post-BMT phase is characterized by the presence of inflammatory cytokines whose net effect is to promote lymphocyte influx into lungs with minimal fibrosis, that leads to an acute form of graft-vs.-host reaction-mediated pulmonary tissue damage. Gradual changes over time in leucocyte influx and activation lead to dysregulated wound repair mechanisms resulting from the shift in the balance of cytokines that promote fibrosis. Using data from new animal models of IPS and information from studies of human IPS, we hypothesize that cytokine-modulated immunological mechanisms which occur during the acute and chronic phases after bone marrow transplantation lead to the development of the progressive, inflammatory, and fibrotic lung disease typical of idiopathic pneumonia syndrome.     

Tumorigenesis in high-dose total body irradiated rhesus monkeys--a life span study

In the early sixties, studies have been performed at the TNO-Institutes for Health Research on acute effects of high dose total body irradiation (TBI) with X-rays and fission neutrons in Rhesus monkeys and the protective effect of autologous bone marrow transplantation (BMT). The surviving animals of this study were kept to investigate late radiation effects, ie, tumorigenesis. TBI in combination with chemotherapy, followed by rescue with BMT is increasingly used for the treatment of hematological malignancies and refractory autoimmune disease. The risk of radiation carcinogenesis after this treatment is of growing concern in man. Studies on tumor induction in nonhuman primates are of relevance in this context since the response of this species to radiation does not differ much from that in man. The group of long-term surviving monkeys comprised nine neutron irradiated animals (average total body dose 3A Gy, range 2.3-4.4 Gy) and 20 X-irradiated monkeys (average total body dose 7.1 Gy, range 2.8-8.6 Gy). A number of 21 age-matched nonirradiated Rhesus monkeys served as a control-group. All animals wereregularly screened for the occurrence of tumors. Complete necropsies were performed after natural death or euthanasia. At postirradiation intervals of 4-21 years an appreciable number of malignant tumors was observed. In the neutron irradiated group eight out of nine animals died with 1 or more malignant tumors. In the X-irradiated group this fraction was 10 out of 20. The tumors in the control group, in seven out of 21 animals, appeared at much older age compared with those in the irradiated cohorts. The histogenesis of the malignant tumors was diverse, as was the case for benign tumors. The observed shortening of latency periods and life span, as well as, the increase of mean number of tumors per tumor bearing animal for benign neoplasms parallels the trend observed for malignant tumors. The results of this study were compared to other radiation late effects after TBI followed by different BMT treatment modalities in Rhesus monkeys. The observation that the carcinogenic risk of TBI in the Rhesus monkeys is similar to that derived from the studies of the Japanese atomic bomb survivors and the increase of the risk by a factor of 8 emphasizes the need for regular screening for secondary radiation-induced tumors in long-term surviving patients after TBI followed by BMT.     

Bronchiolitis obliterans in chronic graft-versus-host disease: analysis of risk factors and treatment outcomes.

Bronchiolitis obliterans (BrOb), a late complication of bone marrow transplantation (BMT), is associated with chronic graft-versus-host disease (GVHD) and is frequently fatal. To identify the risk factors associated with BrOb, the factors affecting survival, treatment outcomes, and causes of death of patients with BrOb, we retrospectively analyzed 2859 BMT recipients. No cases of BrOb occurred among 1070 autologous BMT recipients. Among 1789 allogeneic BMT recipients, we identified 47 patients with BrOb. In multivariate analysis, older recipients or donors and acute GVHD were significantly associated with the development of BrOb. Among patients with BrOb, 5-year survival from the time of transplantation was only 10%, versus 40% among allogeneic BMT recipients without BrOb. The clinical course of BrOb had a significant effect on survival: 79% survived 5 years from the time of BrOb diagnosis if BrOb improved versus 13% if there was no improvement after the first-line therapy. Predictors of response included older donors and recipients, a previous diagnosis of chronic GVHD, and diagnosis of BrOb 6 months after transplantation; each of these significantly increased the likelihood of a favorable response to treatment. BrOb had high mortality rate of 55%, and pulmonary failure was the leading cause of death. More effective BrOb therapy is needed, especially for patients with unfavorable presentation.     

Sequential measurement of the murine acute-phase protein serum amyloid P component (SAP) as an indicator of graft-versus-host disease following allogeneic bone marrow transplantation in mice.

Murine models of bone marrow transplantation (BMT) are used commonly for studies of the pathogenesis and treatment of graft-versus-host disease (GVHD). We report here that the sequential measurement of the mouse acute-phase protein SAP can be used to provide a sensitive, quantitative index of the severity of GVHD. Thirty mice underwent allogeneic, and a further 30 syngeneic BMT. GVHD was assessed in vivo by clinical appearances and weight change, and post mortem by histology and calculation of splenic indices. Blood was obtained twice/week for SAP measurement and blood culture. In all mice an initial rise in SAP levels due to irradiation was followed by a return to baseline. Thereafter in syngeneic marrow recipients levels remained low. In contrast, after allogeneic BMT SAP levels rose progressively as mice developed GVHD, reaching a peak of 135 micrograms/ml prior to death, from a nadir at day 20 of 15 micrograms/ml. Mice with high splenic indices and histological evidence of severe GVHD had significantly higher SAP levels than mice with mild GVHD (P = 0.0002). Elevation in SAP levels occurred independently of bacteraemia. We conclude that in murine BMT sequential measurement of SAP provides an objective means of assessing GVHD in vivo.     

CMV antigenemia following bone marrow transplantation: risk factors and outcomes.

Cytomegalovirus (CMV) infection remains a major problem in blood and bone marrow transplant (BMT) recipients. Recent efforts have been directed at prevention, early diagnosis, and treatment of CMV disease following BMT. Assay for CMV early antigen pp65 on circulating leukocytes has been shown to be sensitive, and specific in detecting early CMV infection. We examined the frequency, risk factors, and outcomes of a positive CMV antigen assay in 118 consecutive BMT patients. Forty-three (36%) of the 118 patients developed CMV antigenemia a median of 26 days post-BMT (range, -6 to 209 days). The incidence of antigenemia in autologous, related donor, and unrelated donor BMT recipients was 15%, 50%, and 48%, respectively (P < .01) and was lower in CMV-seronegative patients (19% versus 51% in seropositive patients; P < .01). Patients with grade II to IV acute graft-versus-host disease (GVHD) had 2.2 times the risk of antigenemia of patients with no or only limited GVHD (P = .03). Age at transplantation, underlying disease, CMV prophylaxis regimen, and GVHD prophylaxis regimen did not affect the risk of CMV antigenemia. Ten of the 43 antigenemic patients, all CMV-seropositive allogeneic BMT (alloBMT) recipients, developed CMV organ disease a median of 101 days (range, 28-283 daya) post-BMT. These data suggest that CMV-seropositive alloBMT patients are at highest risk for CMV antigenemia and for organ disease as well. CMV disease may occur before antigenemia is detectable in leukopenic patients and may also develop late post-BMT, even in patients still receiving antiviral prophylaxis. In high-risk groups, intensive surveillance continuing for more than 6 months after BMT may be indicated.     

Lymphohematopoietic graft-vs.-host reactions can be induced without graft-vs.-host disease in murine mixed chimeras established with a cyclophosphamide-based nonmyeloablative conditioning regimen.

Mixed hematopoietic chimerism can be induced in mice receiving allogeneic bone marrow transplantation (BMT) after nonmyeloablative host conditioning with depletion T cells with of anti-T cell monoclonal antibodies (mAbs), low-dose (3 Gy) total-body irradiation (TBI), and local thymic irradiation (7 Gy). These mice are specifically tolerant to donor and host antigens. When nontolerant donor T cells are given to chimeras several months after BMT, full donor-type chimerism develops, but graft-vs.-host disease (GVHD) does not occur. The induction of such lymphohematopoietic GVH reactions without GVHD could provide an approach to separating graft-vs.-leukemia (GVL) from GVHD in patients with hematologic malignancies. To make the nonmyeloablative conditioning regimen described above more cytoreductive for such malignancies, we have now modified it by replacing TBI with cyclophosphamide (CP). Treatment with anti-CD4 and anti-CD8 mAbs on day -5, 200 mg/kg CP on day -1, and 7 Gy thymic irradiation on day 0 was only slightly myelosuppressive and allowed fully major histocompatibility complex (MHC)-mismatched (with or without multiple minor antigen disparities) allogeneic bone marrow to engraft and establish long-term mixed chimerism in 40 to 82% of recipients in three different strain combinations. The administration of nontolerant donor spleen cells at 5 weeks or at 5, 8, and 11 weeks posttransplant was capable of eliminating host hematopoietic cells, leading to full or nearly full donor chimerism in six of six and two of four chimeric animals in two different strain combinations. No clinical evidence of GVHD was observed in any recipients of these donor leukocyte infusions (DLI). These studies demonstrate that induction of mixed chimerism with nonmyeloablative conditioning followed at appropriate times by DLI might allow lymphohematopoietic GVH reactions, and hence GVL effects, to eliminate chronic hematologic malignancies without causing clinically significant GVHD.     

FTY720 enhances the activation-induced apoptosis of donor T cells and modulates graft-versus-host disease

FTY720 is a novel immunosuppressant that improves the outcomes after solid organ and bone marrow transplantation (BMT) due to the sequestration of T cells into LN. We tested the hypothesis that the sequestration of donor T cells in LN by FTY720 would enhance their interaction with host APC, thus causing a greater degree of activation-induced apoptosis of alloreactive T cells, and thereby resulting in a reduction of graft-vs.-host disease (GVHD). The short-term administration of FTY720 improved the recipient survival after allogeneic BMT. FTY720 treatment facilitated a rapid contraction of the donor T cell pool in association with an increased degree of apoptosis of donor T cells. The donor T cell reactivity to host alloantigens was diminished in host's LN and adoptive transfer of donor T cells isolated from LN of FTY720-treated recipients of allogeneic BMT induced less severe GVHD in secondary recipients than the transfer from controls. Caspase-dependent apoptosis was involved in this mechanism because FTY720-induced protection was abrogated when a pan-caspase inhibitor was administered. These findings thus demonstrate the presence of a novel mechanism by which FTY720 modulates the allogeneic T cell responses: namely, by the induction of activation-induced apoptosis of alloreactive T cells in LN.     

Early vaccination with tumor-lysate-pulsed dendritic cells after allogeneic bone marrow transplantation has antitumor effects.

Allogeneic bone marrow transplantation (BMT) remains the primary treatment for many hematologic malignancies but has had limited success against solid tumors. The antitumor activity of this treatment approach involves the tumoricidal activity of chemoradiation and the additive graft-versus-tumor activity of donor T cells. However, even with current protocols, some tumors develop resistance and become unresponsive to current therapeutic regimens. To address the problem of resistance and lack of solid tumor activity in allogeneic BMT, we undertook experiments to determine whether the graft-versus-tumor activity of donor T cells could be enhanced in the period immediately after allogeneic BMT with tumor lysate-pulsed dendritic cell (DC) vaccines. Using the B16 melanoma model, we found that the treatment of 6-day tumors with allogeneic BMT and 3 weekly vaccinations of tumor lysate-pulsed DCs starting 3 days after BMT had a significant effect on the growth of murine flank melanomas. This effect was tumor specific and occurred in the absence of full immune reconstitution as measured by donor T cell engraftment and cytotoxic T lymphocyte activity. In addition, DC vaccinations did not appear to exacerbate graft-versus-host disease. These experiments support the feasibility of DC vaccine strategies in the setting of allogeneic BMT.     

Selective elimination of alloreactive donor T cells attenuates graft-versus-host disease and enhances T-cell reconstitution.

Impaired T-cell immune reconstitution is a major complication after allogeneic bone marrow transplantation (BMT) and is particularly exacerbated in the setting of graft-versus-host disease (GVHD). Conventional approaches to reduce GVHD, such as T-cell depletion or pharmacologic immunosuppression, typically fail to enhance T-cell immunity and often further exacerbate this problem. An alternative strategy to mitigate GVHD severity is the selective elimination of graft-versus-host-reactive donor T cells by using an incorporated thymidine kinase suicide gene. This approach has been shown to effectively reduce GVHD, although the effect of this strategy on T-cell reconstitution is unresolved. We addressed this question in a murine BMT model (C57BL/6 [H-2(b)] --> AKR/J [H-2(k)]) in which donor and recipient differ at major and minor histocompatibility antigens. Lethally irradiated AKR recipients transplanted with T cell-depleted bone marrow plus thymidine kinase-positive T cells followed by post-BMT ganciclovir (GCV) administration had more prompt and complete normalization of the T-cell repertoire than phosphate-buffered saline-treated GVHD control animals. By 60 days after transplantation, mice administered GCV had T-cell repertoires that were virtually indistinguishable from those of mice that underwent transplantation with T cell-depleted bone marrow alone (no GVHD controls) when assayed by T-cell receptor (TCR) spectratyping. In contrast, phosphate-buffered saline-treated animals had persistent skewing in most Vbeta families. T cells obtained from GCV-treated mice also had significantly higher in vitro proliferative responses after posttransplantation inoculation with ovalbumin than GVHD animals, indicating that CD4(+) T-cell responses against a nominal antigen were better preserved in these chimeras. Finally, GCV-treated mice had augmented immune reconstitution in response to exogenous interleukin-7 administration, as evidenced by increased overall spleen cellularity and absolute numbers of T and B cells. This was in contrast to GVHD control animals, which had a blunted response to interleukin-7 administration. These data indicate that GVHD severity can be significantly reduced by selective elimination of alloreactive donor T cells without compromise of T-cell immunity. Moreover, in light of previous studies demonstrating that this strategy can reduce GVHD without loss of alloengraftment and antileukemia reactivity, further examination of this approach in humans seems warranted.     

Dynamic Change and Impact of Myeloid-Derived Suppressor Cells in Allogeneic Bone Marrow Transplantation in Mice

Myeloid-derived suppressor cells (MDSCs) are a group of myeloid cells composed of hematopoietic progenitor cells, immature macrophages, dendritic cells, and granulocytes, which accumulate in inflammatory diseases and various cancers. Here, we investigated the dynamic changes and effects of MDSCs in graft-versus-host disease (GVHD) development and/or tumor relapse after syngeneic and allogeneic bone marrow transplantation (BMT). We found that adding functional MDSCs in donor graft alleviated GVHD, whereas removal of MDSCs in vivo exacerbated GVHD. After T cell-deplete BMT, MDSCs transiently accumulated in the blood and spleen of recipients without GVHD. In contrast, after T cell-replete BMT, the levels of blood MDSCs were constantly elevated in recipients with GVHD. MDSC accumulation positively correlated with the severity of GVHD. Additionally, MDSC accumulation was further increased upon tumor relapse. Although MDSCs isolated from both syngeneic and allogeneic BMT recipients inhibited T cell proliferation in response to alloantigen stimulation ex vivo, MDSCs from the recipients with GVHD showed much higher suppressive potency compared with those from recipients without GVHD. These results indicate that MDSCs can regulate the immune response in acute GVHD, and possibly tumor relapse, subsequent to allogeneic BMT.