Asha Pillai

High success rate of hematopoietic cell transplantation regardless of donor source in children with very high-risk leukemia

We evaluated 190 children with very high-risk leukemia, who underwent allogeneic hematopoietic cell transplantation in 2 sequential treatment eras, to determine whether those treated with contemporary protocols had a high risk of relapse or toxic death, and whether non-HLA-identical transplantations yielded poor outcomes. For the recent cohorts, the 5-year overall survival rates were 65% for the 37 patients with acute lymphoblastic leukemia and 74% for the 46 with acute myeloid leukemia; these rates compared favorably with those of earlier cohorts (28%, n = 57; and 34%, n = 50, respectively). Improvement in the recent cohorts was observed regardless of donor type (sibling, 70% vs 24%; unrelated, 61% vs 37%; and haploidentical, 88% vs 19%), attributable to less infection (hazard ratio [HR] = 0.12; P = .005), regimen-related toxicity (HR = 0.25; P = .002), and leukemia-related death (HR = 0.40; P = .01). Survival probability was dependent on leukemia status (first remission vs more advanced disease; HR = 0.63; P = .03) or minimal residual disease (positive vs negative; HR = 2.10; P = .01) at the time of transplantation. We concluded that transplantation has improved over time and should be considered for all children with very high-risk leukemia, regardless of matched donor availability.

Host natural killer T cells induce an interleukin-4-dependent expansion of donor CD4+CD25+Foxp3+ T regulatory cells that protects against graft-versus-host disease.

Although CD4(+)CD25(+) T cells (T regulatory cells [Tregs]) and natural killer T cells (NKT cells) each protect against graft-versus-host disease (GVHD), interactions between these 2 regulatory cell populations after allogeneic bone marrow transplantation (BMT) have not been studied. We show that host NKT cells can induce an in vivo expansion of donor Tregs that prevents lethal GVHD in mice after conditioning with fractionated lymphoid irradiation (TLI) and anti-T-cell antibodies, a regimen that models human GVHD-protective nonmyeloablative protocols using TLI and antithymocyte globulin (ATG), followed by allogeneic hematopoietic cell transplantation (HCT). GVHD protection was lost in NKT-cell-deficient Jalpha18(-/-) hosts and interleukin-4 (IL-4)(-/-) hosts, or when the donor transplant was Treg depleted. Add-back of donor Tregs or wild-type host NKT cells restored GVHD protection. Donor Treg proliferation was lost in IL-4(-/-) hosts or when IL-4(-/-) mice were used as the source of NKT cells for adoptive transfer, indicating that host NKT cell augmentation of donor Treg proliferation after TLI/antithymocyte serum is IL-4 dependent. Our results demonstrate that host NKT cells and donor Tregs can act synergistically after BMT, and provide a mechanism by which strategies designed to preserve host regulatory cells can augment in vivo donor Treg expansion to regulate GVHD after allogeneic HCT.

Detectable minimal residual disease before hematopoietic cell transplantation is prognostic but does not preclude cure for children with very-high-risk leukemia.

In patients with acute leukemia, detection of minimal residual disease (MRD) before allogeneic hematopoietic cell transplantation (HCT) correlates with risk of relapse. However, the level of MRD that is most likely to preclude cure by HCT is unclear, and the benefit of further chemotherapy to reduce MRD before HCT is unknown. In 122 children with very-high-risk acute lymphoblastic leukemia (ALL; n = 64) or acute myeloid leukemia (AML, n = 58), higher MRD levels at the time of HCT predicted a poorer survival after HCT (P = .0019); MRD was an independent prognostic factor in a multivariate analysis (P = .0035). However, the increase in risk of death associated with a similar increment of MRD was greater in ALL than in AML, suggesting that a pretransplantation reduction of leukemia burden would have a higher impact in ALL. At any given MRD level, survival rates were higher for patients treated in recent protocols: the 5-year overall survival for patients with ALL was 49% if MRD was detectable and 88% if it was not and the corresponding rates for patients with AML were 67% and 80%, respectively. Although MRD before HCT is a strong prognostic factor, its impact has diminished and should not be regarded as a contraindication for HCT.

Host NKT Cells Can Prevent Graft-versus-Host Disease and Permit Graft Antitumor Activity after Bone Marrow Transplantation

Allogeneic bone marrow transplantation is a curative treatment for leukemia and lymphoma, but graft-vs-host disease (GVHD) remains a major complication. Using a GVHD protective nonmyeloablative conditioning regimen of total lymphoid irradiation and antithymocyte serum (TLI/ATS) in mice that has been recently adapted to clinical studies, we show that regulatory host NKT cells prevent the expansion and tissue inflammation induced by donor T cells, but allow retention of the killing activity of donor T cells against the BCL1 B cell lymphoma. Whereas wild-type hosts given transplants from wild-type donors were protected against progressive tumor growth and lethal GVHD, NKT cell-deficient CD1d−/− and Jα-18−/− host mice given wild-type transplants cleared the tumor cells but died of GVHD. In contrast, wild-type hosts given transplants from CD8−/− or perforin−/− donors had progressive tumor growth without GVHD. Injection of host-type NKT cells into Jα-18−/− host mice conditioned with TLI/ATS markedly reduced the early expansion and colon injury induced by donor T cells. In conclusion, after TLI/ATS host conditioning and allogeneic bone marrow transplantation, host NKT cells can separate the proinflammatory and tumor cytolytic functions of donor T cells.

NKT cells, Treg, and their interactions in bone marrow transplantation

Bone marrow transplantation (BMT) is a potentially curative treatment for patients with leukemia and lymphoma. Tumor eradication is promoted by the anti‐tumor activity of donor T cells contained in the transplant; however, donor T cells also mediate the serious side effect of graft‐versus‐host disease (GVHD). Separation of GVHD from graft anti‐tumor activity is an important goal of research in improving transplant outcome. One approach is to take advantage of the immunomodulatory activity of regulatory NKT cells and CD4+CD25+ Treg of host and/or donor origin. Both host and donor NKT cells and donor Treg are able to prevent GVHD in murine models. In this review, we summarize the mechanisms of NKT cell‐ and Treg‐mediated protection against GVHD in mice while maintaining graft anti‐tumor activity. In addition, we also examine the interactions between NKT cells and Treg in the context of BMT, and integrate the data from murine experimental models with the observations made in humans.

Recipient Myeloid-Derived Immunomodulatory Cells Induce PD-1 Ligand–Dependent Donor CD4+Foxp3+ Regulatory T Cell Proliferation and Donor–Recipient Immune Tolerance after Murine Nonmyeloablative Bone Marrow Transplantation

We showed previously that nonmyeloablative total lymphoid irradiation/rabbit anti-thymocyte serum (TLI/ATS) conditioning facilitates potent donor–recipient immune tolerance following bone marrow transplantation (BMT) across MHC barriers via recipient invariant NKT (iNKT) cell-derived IL-4–dependent expansion of donor Foxp3+ naturally occurring regulatory T cells (nTregs). In this study, we report a more specific mechanism. Wild-type (WT) BALB/c (H-2d) hosts were administered TLI/ATS and BMT from WT or STAT6−/− C57BL/6 (H-2b) donors. Following STAT6−/− BMT, donor nTregs demonstrated no loss of proliferation in vivo, indicating that an IL-4–responsive population in the recipient, rather than the donor, drives donor nTreg proliferation. In graft-versus-host disease (GVHD) target organs, three recipient CD11b+ cell subsets (Gr-1highCD11c−, Gr-1intCD11c−, and Gr-1lowCD11c+) were enriched early after TLI/ATS + BMT versus total body irradiation/ATS + BMT. Gr-1lowCD11c+ cells induced potent H-2Kb+CD4+Foxp3+ nTreg proliferation in vitro in 72-h MLRs. Gr-1lowCD11c+ cells were reduced significantly in STAT6−/− and iNKT cell–deficient Jα18−/− BALB/c recipients after TLI/ATS + BMT. Depletion of CD11b+ cells resulted in severe acute GVHD, and adoptive transfer of WT Gr-1lowCD11c+ cells to Jα18−/− BALB/c recipients of TLI/ATS + BMT restored day-6 donor Foxp3+ nTreg proliferation and protection from CD8 effector T cell–mediated GVHD. Blockade of programmed death ligand 1 and 2, but not CD40, TGF-β signaling, arginase 1, or iNOS, inhibited nTreg proliferation in cocultures of recipient-derived Gr-1lowCD11c+ cells with donor nTregs. Through iNKT-dependent Th2 polarization, myeloid-derived immunomodulatory dendritic cells are expanded after nonmyeloablative TLI/ATS conditioning and allogeneic BMT, induce PD-1 ligand–dependent donor nTreg proliferation, and maintain potent graft-versus-host immune tolerance.

The Weaned Pig as a Model for Doxorubicin-Induced Mucositis

Background: Chemotherapy-induced mucositis (CIM) complicates cancer therapy and limits maximum tolerated doses and efficacy. Rodent models do not reproducibly mimic clinical CIM, so alternative models are needed. Methods: CIM severity was assessed after weaned pigs were treated with doxorubicin (5 and 3.75 mg/kg) using clinical observations, laboratory parameters and gastrointestinal structure and functions. Bovine colostrum was provided as an experimental intervention to the pigs treated receiving the 3.75 mg/kg dose. Results: Doxorubin at 3.75 mg/kg decreased food intake and weight gain (p < 0.05) and caused diarrhea and vomiting that coincided with damage to the small intestine mucosa based on histological scoring (p < 0.05). It resulted in higher serum TNF-α concentrations, increased chloride secretion and reduced brush border membrane disaccharidase activities and carrier-mediated glucose uptake (all p < 0.05). The gastrointestinal damage and dysfunction resemble the clinical and laboratory features of CIM in humans; these can be partially prevented by providing cow colostrum. Conclusion: The weaned pig is a relevant large animal for studying CIM and evaluating existing and experimental interventions for mucositis.

Successful Allogeneic Hematopoietic Cell Engraftment after a Minimal Conditioning Regimen in Children with Relapsed or Refractory Solid Tumors

Children with relapsed or refractory solid tumors face dismal prognoses, and novel therapies are desperately needed. Allogeneic hematopoietic cell transplantation (HCT) offers potential for cell-based therapy, but the toxicity of myeloablation limits this approach in heavily pretreated patients. We sought to determine the feasibility of HCT in a cohort of 24 children with incurable solid tumors using human leukocyte antigen-matched sibling or unrelated donors and a minimal conditioning regimen. Before stem cell infusion, all patients received 3 daily doses of 30 mg/m(2) fludarabine followed by 2 Gy of total body irradiation. Hematopoietic cell recovery was rapid and reliable. Median time to neutrophil engraftment was 13.5 days for sibling donors and 12 days for unrelated donors. Donor lymphocyte infusions were used safely in 4 patients, all of whom had either improved chimerism or apparent tumor response. Graft-versus-host disease was comparable across donor sources and did not affect survival. Relapse remains a substantial barrier, although objective graft-versus-tumor effect was observed in several patients. Four patients with detectable disease before HCT achieved a complete response for at least 30 days after HCT, and two remain long-term survivors. Three patients were in complete response before HCT and remained in remission for 3, 6, and 74 months after HCT. Early disease response was associated with improved survival. Allogeneic HCT using this conditioning regimen offers a potential platform for novel immunotherapies.

Effects of conditioning regimens and T cell depletion in hematopoietic cell transplantation for primary immune deficiency.

This study analyzes the hematopoietic cell transplantation experience in patients with immune deficiency at a single institution. The objective is to comprehensively evaluate the short-term and long-term outcomes with various preparative regimens, donor grafts, and ex vivo manipulations to identify transplantation approaches that most likely favor early donor immune competency without generating excessive toxicity. Clinical outcomes were evaluated in 52 consecutive patients with immune deficiencies. Thirty-seven of the 52 patients (71%) survived with attenuation of their underlying disease. The use of a melphalan-based reduced-intensity conditioning preparative regimen and immunomagnetic CD3(+) T cell depletion techniques (when T cell depletion was indicated) were associated with improved event-free survival. Survivors who received a preparative regimen other than a melphalan-based reduced-intensity regimen suffered from therapy-related morbidities or chronic/recurrent infections. Our findings indicate that melphalan-based reduced-intensity conditioning regimens and immunomagnetic CD3(+) T cell depletion limit therapy-related toxicity, and demonstrate promising results for the early establishment of donor immune competency.

Favorable preliminary results using TLI/ATG‐based immunomodulatory conditioning for matched unrelated donor allogeneic hematopoietic stem cell transplantation in pediatric severe aplastic anemia

Pillai A, Hartford C, Wang C, Pei D, Yang J, Srinivasan A, Triplett B, Dallas M, Leung W. Favorable preliminary results using TLI/ATG‐based immunomodulatory conditioning for matched unrelated donor allogeneic hematopoietic stem cell transplantation in pediatric severe aplastic anemia.
Pediatr Transplantation 2011: 15: 628–634.