Feasibility, safety, and efficacy of early prophylactic donor lymphocyte infusion after T cell-depleted allogeneic stem cell transplantation in acute leukemia patients.

Prophylactic donor lymphocyte infusion (DLI) starting at 6 months after T cell-depleted allogeneic stem cell transplantation (TCD-alloSCT) can introduce a graft-versus-leukemia (GvL) effects with low risk of severe graft-versus-host-disease (GvHD). We established a policy to apply low-dose early DLI at 3 months after alloSCT to prevent early relapse. This study analyzes this strategy retrospectively. Of 220 consecutive acute leukemia patients undergoing TCD-alloSCT, 83 were prospectively classified to have a high relapse risk and 43 were scheduled for early DLI. 95% of these patients received freshly harvested DLI within 2 weeks of the planned date. In patients transplanted with reduced intensity conditioning and an unrelated donor, we found an increased cumulative incidence of GvHD between 3 and 6 months after TCD-alloSCT for patients receiving DLI at 3 months compared to patients who did not receive this DLI (0.42 (95%Confidence Interval (95% CI): 0.14-0.70) vs 0). Treatment success was defined as being alive without relapse or need for systemic immunosuppressive GvHD treatment. The five-year treatment success in patients with acute lymphatic leukemia was comparable between high- and non-high-risk disease (0.55 (95% CI: 0.42-0.74) and 0.59 (95% CI: 0.42-0.84)). It remained lower in high-risk acute myeloid leukemia (AML) (0.29 (95% CI: 0.18-0.46)) than in non-high-risk AML (0.47 (95% CI: 0.42-0.84)) due to an increased relapse rate despite early DLI.

Predictors of short-term and long-term mortality in critically ill patients admitted to the intensive care unit following allogeneic stem cell transplantation.

Historically, the mortality of patients admitted to the ICU after allogeneic stem cell transplantation (alloSCT) is high. Advancements in transplantation procedures, infectious monitoring and supportive care may have improved the outcome. This study aimed to determine short-term and long-term mortality after ICU admission of patients after alloSCT and to identify prognostic clinical and transplantation-related determinants present at ICU admission for long-term outcome. A multicenter cohort study was performed to determine 30-day and 1-year mortality within 2 years following alloSCT. A total of 251 patients were included. The 30-day and 1-year mortality was 55% and 80%, respectively. Platelet count <25 × 109/L (OR: 2.26, CI: 1.02-5.01) and serum bilirubin >19 µmol/L (OR: 2.47 CI: 1.08-5.65) at admission, other donor than a HLA-matched-related or HLA-matched-unrelated donor (OR: 4.59, CI: 1.49-14.1) and vasoactive medication within 24 h (OR: 2.35, CI: 1.28-4.31) were associated with increased 30-day mortality. Other donor than a HLA-matched-related or HLA-matched-unrelated donor (OR: 1.9, CI: 1.13-3.19), serum bilirubin >77 (OR: 2.05, CI: 1.28-3.30) and vasoactive medication within 24 h (OR: 1.65, CI: 1.12-2.43) were associated with increased 1-year mortality. Neutropenia was associated with decreased 30-day and 1-year mortality (OR: 0.29, CI: 0.14-0.59 and OR: 0.70, CI: 0.48-0.98). Myeloablative conditioning and T cell-depleted transplantation were not associated with increased mortality.

CMV seronegative donors: Effect on clinical severity of CMV infection and reconstitution of CMV-specific immunity.

BACKGROUND: Cytomegalovirus (CMV)-specific T-cells are crucial to prevent CMV disease. CMV seropositive recipients transplanted with stem cells from a CMV seronegative allogeneic donor (R+D-) may be at risk for CMV disease due to absence of donor CMV-specific memory T-cells in the graft. METHODS: We analyzed the duration of CMV reactivations and the incidence of CMV disease in R+D- and R+D+ patients after alemtuzumab-based T-cell depleted allogeneic stem cell transplantation (TCD alloSCT). To determine the presence of donor-derived primary CMV-specific T-cell responses we analyzed the origin of CMV-specific T-cells in R+D- patients. RESULTS: The duration of CMV reactivations (54 versus 38 days, respectively, p = 0.048) and the incidence of CMV disease (0.14 versus 0.02, p = 0.003 at 1 year after alloSCT) were higher in R+D- patients compared to R+D+ patients. In R+D- patients, CMV-specific CD4+ and CD8+ T-cells were mainly of recipient origin. However, in 53% of R+D- patients donor-derived CMV-specific T-cells were detected within the first year. CONCLUSIONS: In R+D- patients, immunity against CMV was predominantly mediated by recipient T-cells. Nevertheless, donor CMV serostatus significantly influenced the clinical severity of CMV reactivations indicating the role of CMV-specific memory T-cells transferred with the graft, despite the ultimate formation of primary donor-derived CMV-specific T-cell responses in R+D- patients.

Microarray Gene Expression Analysis to Evaluate Cell Type Specific Expression of Targets Relevant for Immunotherapy of Hematological Malignancies.

Cellular immunotherapy has proven to be effective in the treatment of hematological cancers by donor lymphocyte infusion after allogeneic hematopoietic stem cell transplantation and more recently by targeted therapy with chimeric antigen or T-cell receptor-engineered T cells. However, dependent on the tissue distribution of the antigens that are targeted, anti-tumor responses can be accompanied by undesired side effects. Therefore, detailed tissue distribution analysis is essential to estimate potential efficacy and toxicity of candidate targets for immunotherapy of hematological malignancies. We performed microarray gene expression analysis of hematological malignancies of different origins, healthy hematopoietic cells and various non-hematopoietic cell types from organs that are often targeted in detrimental immune responses after allogeneic stem cell transplantation leading to graft-versus-host disease. Non-hematopoietic cells were also cultured in the presence of IFN-γ to analyze gene expression under inflammatory circumstances. Gene expression was investigated by Illumina HT12.0 microarrays and quality control analysis was performed to confirm the cell-type origin and exclude contamination of non-hematopoietic cell samples with peripheral blood cells. Microarray data were validated by quantitative RT-PCR showing strong correlations between both platforms. Detailed gene expression profiles were generated for various minor histocompatibility antigens and B-cell surface antigens to illustrate the value of the microarray dataset to estimate efficacy and toxicity of candidate targets for immunotherapy. In conclusion, our microarray database provides a relevant platform to analyze and select candidate antigens with hematopoietic (lineage)-restricted expression as potential targets for immunotherapy of hematological cancers.

Effectivity of a strategy in elderly AML patients to reach allogeneic stem cell transplantation using intensive chemotherapy: Long-term survival is dependent on complete remission after first induction therapy.

Intensive chemotherapy followed by allogeneic stem cell transplantation (alloSCT) can cure AML. Most studies on alloSCT in elderly AML report results of highly selected patient cohorts. Hardly any data exist on the effectiveness of prospective strategies intended to bring as many patients as possible to transplant. Between 2006 and 2011 we implemented a treatment algorithm for all newly diagnosed AML patients aged 61-75 years, consisting of intensive chemotherapy cycles to induce complete remission, followed by alloSCT. 44 of 60 (73%) newly diagnosed elderly AML patients started with chemotherapy. By meticulously following our algorithm in almost all patients, we could induce complete remission (CR) in 66% of patients starting with chemotherapy, and transplant 32% of these patients in continuous CR. Main reasons for failure were early relapse (16%), early death (14%), primary refractory disease (9%), and patient or physician decision to stop treatment (16%). Patients in continuous CR after first induction benefit most with 36% long-term survival. Patients not in CR after first induction benefit less; although additional chemotherapy induces CR in 45% of these patients, only 23% are transplanted and no long-term survival is observed, mainly due to relapse. Long-term survival in the group of 44 patients is 9% (median 4.5 years after alloSCT). Considering that 27% of patients do not start with chemotherapy and 64% of patients starting with chemotherapy do not reach alloSCT, the reasons for failure presented here should be used as a guide to develop new treatment algorithms to improve long-term survival in elderly AML patients.

Intradermal influenza vaccination in immunocompromized patients is immunogenic and feasible.

BACKGROUND: Many strategies, including intradermal vaccination, have been tested to augment antibody responses upon vaccination. This strategy has not been evaluated in different groups of immunocompromized patients. We conducted a prospective, randomized study to compare the humoral response upon standard intramuscular influenza vaccination with the response upon reduced-dose intradermal vaccination in patients treated with anti-tumor necrosis factor (TNF)-alpha, human immunodeficiency virus (HIV)-infected patients, hematologic stem cell transplantation (HSCT) patients, and healthy controls. METHODS: In total 156 immunocompromized patients and 41 healthy controls were randomized to receive either 0.5mL of the 2005/2006 trivalent influenza vaccine intramuscular or 0.1mL intradermal. Humoral responses, determined by hemagglutination inhibition assay, were measured before and 28 days postvaccination. Geometric mean titers (GMTs) and protection rates (PRs) are reported as primary outcomes, adverse events as a secondary outcome. RESULTS: Reduced-dose intradermal vaccination leads to similar GMTs and PRs, within all tested groups, compared to the standard intramuscular vaccination. Healthy controls yielded significantly better GMTs and PRs than immunocompromized patients. Local skin reactions after intradermal vaccination occurred less frequent and were milder in immunocompromized patients than in healthy subjects and were predictive for a positive vaccination outcome for individual subjects. CONCLUSIONS: Intradermal influenza vaccination is a feasible alternative for standard intramuscular vaccination in several groups of immunocompromized patients, including those treated with anti-TNF, HIV-infected patients and HSCT patients. The occurrence of a local skin reaction after intradermal vaccination is predictive of a response to at least one of the vaccine antigens.

Direct cloning of leukemia-reactive T cells from patients treated with donor lymphocyte infusion shows a relative dominance of hematopoiesis-restricted minor histocompatibility antigen HA-1 and HA-2 specific T cells.

Donor T cells recognizing hematopoiesis-restricted minor histocompatibility antigens (mHags) HA-1 and HA-2 on malignant cells play a role in the antileukemia effect of donor lymphocyte infusion (DLI) in patients with relapsed leukemia after allogeneic stem cell transplantation. We quantified the contribution of HA-1 and HA-2 specific T cells to the total number of leukemia-reactive T cells in three HA-2 and/or HA-1 positive patients responding to DLI from their mHag negative donors. Clinical responses occurring 5-7 weeks after DLI were accompanied by an increase in percentages HLA-DR expressing T cells within the CD8+ T cell population. To clonally analyze the leukemia-reactive immune response, T cells responding to the malignancy by secreting IFNgamma were isolated from peripheral blood, directly cloned, and expanded. Tetramer analysis and specific lysis of peptide-pulsed target cells showed that 3-35% of cytotoxic T lymphocyte (CTL) clones isolated were specific for HA-1 or HA-2. TCR VB analysis showed oligoclonal origin of the HA-1 and HA-2 specific CTL clones. The HA-1 and HA-2 specific CTL clones inhibited leukemic progenitor cell growth in vitro. The relatively high frequency of HA-1 and HA-2 specific T cells within the total number of tumor-reactive T cells illustrates relative immunodominance of mHags HA-1 and HA-2.

Minimal GVHD following in-vitro T cell-depleted allogeneic stem cell transplantation with reduced-intensity conditioning allowing subsequent infusions of donor lymphocytes in patients with hematological malignancies and solid tumors.

OBJECTIVE: Allogeneic stem cell transplantation (alloSCT) following reduced-intensity conditioning offers a relatively nontoxic regimen while preserving rapid and sustained engraftment. Acute and chronic graft-vs-host disease (GVHD) is, however, a significant cause of severe morbidity. To reduce the incidence of GVHD, we treated a group of high-risk patients with a reduced-intensity conditioning regimen followed by in vitro T-cell-depleted alloSCT using Campath 1-H incubation. PATIENTS AND METHODS: Eighteen patients were treated with fludarabine (6 x 30 mg/m(2)), busulphan (2 x 3.2 mg/kg), and ATG (4 x 10 mg/kg) followed by the infusion of high-dose T-cell-depleted peripheral stem cells from sibling donors. No posttransplant GVHD prophylaxis was administered. At 6 months after alloSCT, low-dose donor lymphocyte infusion (DLI) was administered. RESULTS: All patients had sustained engraftment of donor cells with a median of 95% donor cells at 3 months after alloSCT. Minimal acute and no chronic GVHD was observed after alloSCT. A high incidence of cytomegalovirus (CMV) reactivation but no CMV disease was observed. Eleven patients received DLI at a median of 6.5 months after alloSCT. Acute GVHD grade II-III developed in 6 patients. All patients showed improvement of donor chimerism after DLI. With a median follow-up of 211 days, 11 patients are alive. Particular in patients with chronic lymphocytic leukemia and acute myeloid leukemia, a significant graft-vs-tumor effect was observed. CONCLUSIONS: In vitro T-cell-depleted alloSCT following reduced-intensity conditioning leads to durable donor engraftment without GVHD. The high levels of donor chimerism allow the subsequent use of cellular immunotherapy to treat residual disease.

Minor histocompatibility antigens as targets of graft-versus-leukemia reactions.

The main advantage of allogeneic stem cell transplantation over autologous stem cell transplantation for hematologic malignancies is the ability to perform cellular immunotherapy using donor-derived immune effector cells after transplantation. In HLA-matched allogeneic stem cell transplantation, the beneficial graft-versus-leukemia effect of donor lymphocytes appears to be caused mainly by alloreactive T cells that are capable of recognizing minor histocompatibility antigens on the malignant cell population from the patient. The tissue distribution of minor histocompatibility antigens probably determines the clinical result of T-cell responses against these antigens. Whereas T cells recognizing broadly expressed antigens cause not only graft-versus-leukemia but also graft-versus-host disease, T cells recognizing minor histocompatibility antigens specifically expressed on hematopoietic cells may mainly eliminate hematopoietic cells from the recipient, including the malignant cells, without affecting donor hematopoiesis or normal nonhematopoietic tissues. Graft-versus-host disease may still occur because of the induction of inflammatory responses against hematopoietic cells in the tissues. Vaccination of patients after transplantation or vaccination of stem cell donors before transplantation using minor histocompatibility antigen-specific peptides, production of minor histocompatibility antigen-specific T cells, and redirection of T-cell specificity by gene transfer of T-cell receptors may be strategies to eradicate specifically the malignant cells after allogeneic stem cell transplantation.