Matched Sibling Stem Cell Transplantation Research Articles

Non-myeloablative matched sibling stem cell transplantation with the optional reinforced stem cell infusion for patients with hemoglobinopathies.

The NIH protocol for non-myeloablative (NMA) conditioning allogeneic stem cell transplantation (alloSCT) with alemtuzumab and low-dose total body irradiation corrected the abnormal sickle cell disease (SCD) phenotype without the risk of graft-versus-host disease. However, alloSCT using NMA conditioning had been rarely applied to β-thalassemia major (β-TM) patients. To avoid prolonged immunosuppression, we developed a two-stage strategy. Mixed donor chimerism was initially achieved using the protocol developed by the NIH protocol. Thereafter, we facilitated donor chimerism using the optional reinforced stem cell (SC) infusion in cases requiring protracted immunosuppression or experiencing impending graft failure. In this study, β-TM (n=9) and SCD (n=4) patients were equally effectively treated with eradicating the abnormal hemoglobin phenotype. Five patients, including four β-TM, achieved stable mixed chimerism without receiving optional reinforced SC infusion. All patients that received optional reinforced infusion achieved complete (n=4) or mixed chimerism (n=1). The overall survival rate and event-free survival at 4years were 91.7% (95% CI; 53.9-98.8) in both groups, with a thalassemia-free survival rate in β-TM patients of 87.5% (95% CI; 38.7-98.1). This study is the first to report successful NMA conditioning alloSCT to achieve stable mixed chimerism correcting the abnormal hemoglobin phenotype in adult β-TM patients.

Post-transplant Hemophagocytic Lymphohistiocytosis in Benign Hematological Disorders: Experience of 4 Cases with Review of Literature.

Post transplant Hemophagocytic lymphohistiocytosis (HLH) is a form of secondary HLH, which can be either early onset or late onset and is associated with significant morbidity and mortality. With the increasing popularity of post transplant cyclophosphamide based haploidentical stem cell transplantation (SCT), post transplant HLH is becoming a significant complication especially in benign hematological disorders. Methods: We present 4 cases of post transplant HLH occurring in 2 cases of severe aplastic anemia (post haploidentical SCT) and 2 cases of thalassemia major (post matched sibling SCT). All 4 cases had early onset variety with dismal prognosis. Conclusion: Post-transplant HLH is an important entity in benign hematological disorders, which needs to be identified early and treated promptly with steroids, monoclonal agents or immunosuppressive therapy. Serum ferritin levels are an important biomarker and help in monitoring response.

HLA- Matched Sibling Stem Cell Transplantation in Children with Thalassemia Major with ATG As Part of Preperative Regimen: A Suitable Choice Even for High Risk Cases

▪Background:Myeloablative chemotherapy using busulfan(BU) & cyclophosphamide(CY) is a standard form of conditioning for thalassemia transplantation. But this is associated with substantial incidence of graft rejection. This is much more common in high risk patients .Methodology:This study retrospectively analyses the outcome of 50 (34 boys & 16 girls) consecutive children, who underwent HLA matched stem cell transplantation from family donors at our centre All patients were classified into 3 risk groups on the basis of inadequate chelation, hepatomegaly and the presence of portal fibrosis in the liver. Pesaro Class 1 patients had none of these, class 2 patients had one or two and class 3 had all three.30 patients received oral BU 1 mg/kg, Q6H for 4 days followed by CY 60 mg/kg/D for two days. & rabbit ATG (2.5 mg/kg/day from Day-5 to Day-1). Patients with advanced disease (class 3) received BU 0.8mg/Kg & CY 50 mg/kg. with same dose of ATG. All patients received stem cells harvested from G-mobilized bone marrow Standard MTX & Cyclosporin based GVHD prophylaxis , infection prophylaxis & supportive care as per standard protocols were given as per standard protocols. Chimerism was routinely monitored at 30, 60, 90, 120 and 365 days post transplant using PCR-based analysis of selected polymorphic STRs loci.Results :OS and thalassemia-free-survival were estimated according to the Kaplan-Meier method. Probabilities of rejection, GVHD and infections were calculated by using the cumulative incidence estimator. The impact of the following parameters on OS and TFS was examined: (1) age of the recipient (above vs below the median value) (2) serum ferritin (above vs, the median value) (3) Pesaro class, (4) total nucleated cells per kg (contained in the graft) (above vs below the median), (5) CD34+ cells contained in the graft (above vs below the median) and (6) BU (oral vs i.v. BU). Both univariate and multivariate analysis of prognostic factors were made according to the log-rank test and to a stepwise Cox proportional hazards regression model, respectively. With a median follow-up of 11 years (range 1—17 years), the overall survival (OS) and thalassemia free survival (TFS) rates were 94% and 90%, respectively. The estimated TRM was 4% while the cumulative incidence of rejection/failure were 2%. One patient ( Class III, & HIV infection) died of severe veno-occlusive disease. ,The patient who rejected the graft after 10 months, received II transplant & is transfusion free with 98% donor chimerism post second transplant. The cumulative incidences of acute GVHD grade II & grade III were 15% & 8 %, respectively. 1 patient developed acute GVHD grade IV. None of the patients developed chronic GVHD. The estimated OS and TFS for patients with Class 1 and 2 disease according to Pesaro criteria were 94% and 92%, whereas for patients with Class 3 disease they were 90% and 85%, respectively.Conclusions: Myeloablative( BU-CY) conditioning with ATG as conditioning from matched sibling donors with innovative modifications can be used to achieve excellent survival , with a low incidence of TRM and graft rejection in low as well as high risk patients. DisclosuresNo relevant conflicts of interest to declare.

The Comparison Of Clinical Outcomes Between Matched Sibling Stem Cell Transplantation and Immunosuppressive Treatment As a First-Line Treatment For Adult Patients With Severe Aplastic Anemia

▪ BackgroundAllogeneic stem cell transplantation (SCT) from matched-sibling donor (MSD) and immunosuppressive treatment (IST) are the most widely used first-line treatments for patients with severe aplastic anemia (SAA). Overall long-term survival rates are comparable between the two groups. However, patients with age of over 40 have not been generally considered as candidates of SCT from MSD (MSD-SCT) due to higher transplant-related mortality. Recent improvements in MSD-SCT such as less intensive fludarabine-based conditioning, and use of rabbit anti-thymocyte globuline (ATG) instead of horse ATG as first-line IST may change these results. Therefore, we compared the clinical outcomes between MSD-SCT with fludarabine-based conditioning and IST with rabbit ATG and cyclosporine A (CsA). MethodsWe analyzed the clinical results of 54 adult SAA patients who were treated with MSD-SCT and 93 with IST as a first-line treatment from March 2006 to May 2012 at Seoul St. Mary's Hospital, Seoul, Korea. The patients who were treated with MSD-SCT received conditioning with fludarabine (30 mg/m2/day × 6 days), cyclophosphamide (50 mg/kg/day × 2 days), and rabbit ATG (Thymoglobulin®, 2.5 mg/kg/day × 4 days). Those who were treated with IST received rabbit ATG (2.5 mg/kg/day × 5 days) with CsA. ResultsThe median ages were not significantly different between the MSD-SCT group and IST group (38.5 years vs. 43.0 years; P=0.103). Other baseline characteristics were comparable except the interval from diagnosis to treatment (100 days vs. 40 days; P=0.013), absolute lymphocyte count (0.68 × 109/L vs. 0.93 × 109/L; P=0.013), and platelet count (10.0 × 109/L vs. 11.0 × 109/L; P=0.035). In the IST group, overall response and complete response rates at 1 year were 44.1% (95% CI, 33.8-54.8) and 10.8% (95% CI, 5.3-18.9). Treatment failure developed in 55 (59.1%) patients due to non-response in 34 (36.6%), relapse in 5 (5.4%), clonal evolution in 3 (3.2%), and treatment-related mortality in 13 (14.0%) patients. After treatment failure, 17 (18.3%) patients received SCT from MSD or unrelated donor. In the MSD-SCT group, 10 (18.5%) patients experienced treatment failure due to secondary graft failure in 5 (9.3%), clonal evolution in 1 (1.9%), and treatment-related mortality in 4 (7.4%) patients. Among the patients who experienced secondary graft failure, 4 (7.4%) patients received secondary SCT, which resulted in sustained graft function. Consequently, overall survival (OS) at 3 years in the MSD-SCT group was not significantly different compared to that in the IST group (90.7% vs. 81.0%; P=0.139). However, the MSD-SCT group showed significantly higher failure-free survival (FFS) at 3 years compared to the IST group (80.2% vs. 46.6%; P<0.001). When we analyzed the patients with age of over 40 years, OS at 3 years in the MSD-SCT group was not significantly different compared to that in the IST group (87.5% vs. 74.7%; P=0.251), whereas FFS at 3 years in the MSD-SCT group was significantly higher compared to that in the IST group (84.0% vs. 43.0%; P=0.001). ConclusionsOur data suggest that MSD-SCT is more favorable than IST as a first-line treatment, considering the curative nature of MSD-SCT even in patients over 40 years of age. Disclosures:No relevant conflicts of interest to declare.

Low Thymic Natural Regulatory T Cell Output in the Donor Is Associated with An Increased Incidence of Extensive Chronic GvHD After T Cell Depleted Matched Sibling Stem Cell Transplantation (SCT)

Abstract 328Low CD4+ T cell counts early post-SCT are associated with a greater risk of developing chronic GVHD (cGVHD). However, the factors affecting post graft CD4+ T cell recovery are not known. Furthermore, the CD4 T cell subset correlating best with protection from cGvHD is not defined. We hypothesized that the donor immune repertoire determines CD4+ reconstitution after SCT. We studied 220 donor-recipient pairs undergoing a myeloablative matched-sibling T-cell depleted SCT for a variety of hematological malignancies including AML, ALL, CML, and MDS. Median donor and patient age was 36 years (range, 9 – 70). Median CD34+ cell dose was 5.56 ×106/kg (range, 2.3–14.5), with a fixed T cell dose per protocol of 1–5 ×104/kg. Ninety-four patients (43%) developed chronic GvHD (32 limited, 64 extensive). Prior to SCT a cryopreserved mononuclear fraction (PBMC) was obtained by leukopheresis on all donors. PBMC from 139 donors were stained with fluorescently conjugated antibodies against CD3, CD4, CD8, CD27, CD45RA, CD31, CD25, CD14, CD19, FOXP3, and Helios and acquired on a custom-built LSR Fortessa (BD) flow cytometer. The following T cell subsets were examined: naive, CD27+CD45RA+; central memory, CD27+CD45RA-; effector memory, CD27-CD45RA-; and effector T cells, CD27-CD45RA+. Regulatory T cells (Tregs) and Treg subsets were identified within the CD4+ T cell population using a combination of FOXP3 and Helios The combination of FOXP3 with Helios has recently been shown to identify natural, thymus-derived Tregs (nTregs), whereas CD4+FOXP3+ T cells lacking Helios represent a pool of induced Tregs (Thornton et al., 2010, J Immunol. 184:3433–3441). Combined expression of CD31 and CD45RA in the CD4+ and nTreg populations was used to identify recent thymic emigrants (RTE). Data were analyzed by FlowJo (Treestar, Ashland, OH, version 8.6.6), and both univariate and multivariate analysis analyses using the Cox regression models with right censored time-to-event variables were performed on the readouts (SPSS 15.0, S-Plus 8 and Prism 4 Software). All p-values were computed based on the log-rank test statistics. Absolute lymphocyte counts in 220 donors were 2.11/μl (range 0.89 – 4.12). A low absolute lymphocyte count was significantly correlated with a higher cumulative probability for time to incidence of extensive cGVHD (HR 2.38, p = 0.0008). This relationship between lymphocytes and cGVHD was retained in CD4+ T cell subsets: CD3+CD4+ (HR 2.48, p = 0.005); CD3+CD4+CD25+ < 30 cells/μL (HR 2.1, p = 0.02); nTregs < 8 cells/μL (HR 1.97, p = 0.043). RTE of total CD4 did not correlate with RTE nTregs, indicating that nTreg production varies among donors with similar levels of thymic activity. Furthermore recipients of donors with low RTE nTregs relative to overall thymic output had a significantly higher likelihood of developing extensive chronic GvHD (HR 2.78, p=0.0014). In multivariate analysis including patient age, acute GvHD (grade II – IV), graft direction (female donor to male recipient vs. other combinations), disease and status at transplant, and administration of post SCT donor lymphocyte infusion, only thymic output of nTregs (HR = 0.8, p = 0.037) and graft direction (HR = 2.1, p = 0.023) remained significant risk factors for extensive cGvHD. These data show that the thymic output of nTreg varies inherently among donors. Donors with low thymic output of nTreg may recreate a less tolerant immune system in the recipient leading to cGVHD of greater severity.(ZAM and JJM contributed equally to this work). [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Clonal cytogenetic abnormalities after tyrosine kinase inhibitor therapy in Ph+ all resolution after decitabine therapy

Mixed chimerism and presence of minimal residual disease after stem cell transplantation (SCT) usually predict leukemia recurrence. In Philadelphia chromosome positive (Ph+) leukemia tyrosine kinase inhibitors can restore remission. These agents can induce clonal cytogenetic abnormalities in the Philadelphia negative cell population (CCA/Ph-), which may rarely progress to acute myeloid leukemia. A child with Ph+ acute lymphoblastic leukemia showed mixed donor chimerism and persistent bcr-abl transcripts after a matched sibling SCT. There was no response to tyrosine kinase inhibitor (TKI) therapy, but she has remained hematologically normal for more than 5 years. CCA/Ph- was detected but resolved with hypomethylating therapy.

HLA-Matched Sibling Stem Cell Transplantation Using Conditioning Regimen with Reduced Dose of Cyclophosphamide, ATG and Fludarabine in Patients with Severe Aplastic Anemia

Background: We have reported the outcome of HLA-matched stem cell transplantation (SCT) using triple immunosuppressive agents with cyclophosphamide (CY, 200mg/kg), ATG and procarbazine for 113 adult patients with severe aplastic anemia (SAA) (Kim et al, BMT 31:79, 2003). However, high dose CY (200mg/kg) causes serious cardiac toxicity in some cases which may lead to death within few weeks. To avoid high dose CY-associated cardiac toxicity we underwent HLA-matched sibling SCT using increasing dose of ATG and Fludarabine instead of reduced CY to half dose.Method: Between March 2002 and December 2007, fifty patients with adult SAA (six patients were AA/PNH syndrome) received matched sibling SCT. The median age of patients was 39 (16~53) and median interval between Dx and SCT was 19 months (1~352). The median number of transfusion prior to SCT was 34 units (2~680). Nineteen patients (38%) had a history of IST before SCT. The conditioning regimen consisted of Fludarabine (30mg/m2/day, 6 days), CY (50 mg/kg/day, 2 days) and ATG (Thymoglobulin 2.5mg/kg/day, 4 days). Stem cell sources were BM plus CD34+-selected PBSC (n=12), BM (n=20), PBSC (n=4) and G-CSF-primed BM (n=14). All patients received cyclosporine and methotrexate as GVHD prophylaxis.Results: The median dose of CD34+ cells infused was 3.7x106/kg (1.2~11.9). All patients achieved successful primary engraftment, and the median time for ANC and platelet to reach 0.5x109/L and 20x109/L was 12 (10~19) and 17 (10~25) days, respectively. Three patients (6%) developed delayed graft failure whereas 14% (16 out of 113) developed both primary and secondary graft failure in our previous study. But all achieved successful engraftment after booster infusion (n=1) and second SCT (n=2). The incidence of acute GVHD (more than grade II) and chronic GVHD was 8% (n=4) and 4% (n=2; extensive type), respectively. The incidence of acute and chronic GVHD seems to be lower than those of previous conditioning regimen (11% and 12%, respectively) (BMT 31:79, 2003). The incidence of CMV infection requiring preemptive treatment was 54 % (n=27). Three patients died of reactivation of chronic hepatitis C with hepatic GVHD (n=1), CMV pneumonia (n=1), and invasive fungal infection (n=1). PNH clone monitored by flow cytometry disappeared posttransplant in all 6 PNH patients. With median follow up of 32 months (1~74), the estimated probability of survival at 3 years was 94 % compared with those of 89% in our previous report.Conclusions: These data demonstrate that the conditioning regimen used in this study is feasible for patients with SAA who received matched sibling SCT. Of note, the observation of successful engraftment as well as lesser acute and chronic GVHD compared with previous study suggest that increasing dose of ATG and the addition of Fludarabine has potent in vivo T cell depletion and immunomodulatory activity.

Gemtuzumab Ozogamicin (GO) and Continuous Infusion Cytarabine (ARA-C) for Relapsed/Refractory Acute Myeloid Leukemia (AML) Prior to Allogeneic Stem Cell Transplantation (SCT).

The question whether to re-induce AML patients (pts) in first or subsequent relapse to reduce leukemia disease burden prior to SCT is not well studied. Several groups reported a 28–30% disease-free survival rate at five years for matched related donor SCT in first relapse without re-induction. Finally, re-induction regimen choices prior to SCT are relatively limited. Specifically, GO-containing regimens have been suspected to be associated with increased incidence of liver sinusoidal occlusive syndrome (SOS) when used prior to SCT. We evaluated our institutional experience of using GO and ARA-C for relapsed/refractory CD33-positive AML pts prior to allogeneic SCT. A total of 18 pts (median age 45.4 years; range 28–66) with relapsed/refractory AML were treated with GO/ARA-C. The median initial complete remission duration was 4.4 (range 0.5–14.5) months. The median time from relapse to GO/ ARA-C was 57 (range 6–142) days. Pts received GO (6 mg/m2 on day 1, 4 mg/m2 on day 8) and continuous infusion ARA-C (100 mg/m2) daily for 7 days. Bone marrow aspiration and biopsy were performed at a median of 15 (range 11–39) days post induction (one pt had >20% peripheral blasts on day 14 and therefore did not undergo bone marrow aspiration and biopsy). Three (17%) pts developed infectious complications following re-induction and one pt had disease progression that prevented them from undergoing bone marrow aspiration and biopsy and continuing on to SCT. Marrow aplasia (defined as <5% blasts and <20% cellularity) was reached in six of 14 (43%) evaluable pts. Three pts (all without achieving marrow aplasia) developed infectious complications that precluded SCT. A total of 11 pts proceeded onto SCT at a median of 30 (range 21–73) days after re-induction, including six pts who achieved marrow aplasia and five who did not. Six pts underwent matched unrelated donor SCT (two were mismatched at one allele) and five received matched sibling SCT. Reduced intensity preparatory regimens (fludarabine/melphalan) were used in all but one pt who received a fully myeloablative regimen (busulfan/cyclophosphamide). Engraftment to absolute neutrophil count ≥0.5x109/L for three days was achieved in all 11 pts at a median of 15 (range 9–21) days. During SCT, all pts were treated with prophylactic enoxaparin 40 mg subcutaneously once daily to prevent SOS of the liver. The two pts with mismatched unrelated donor SCT died three and nine days after achieving count recovery from pulmonary infiltrates and severe graft versus host disease (GVHD), respectively. Acute GVHD grade II-IV developed in six of the pts with matched donors (three related and three unrelated donors). Chronic GVHD developed in four (one related and three unrelated donors) of the pts. None developed liver SOS. Four pts are alive from starting GO/ARA-C at a median of 7.2 (range 1.2–52.7) months; two of them with and two of them without GVHD. Marrow aplasia prior to SCT was achieved in two of them. In summary, the question of whether to re-induce pts with relapsed/refractory AML to reduce leukemia burden prior to allogeneic SCT remains controversial. However, if one chooses to proceed with re-induction, then the combination of GO/ARA-C represents a feasible re-induction strategy with acceptable toxicity, achievement of marrow aplasia in approximately half of evaluable pts, and no increased risk of liver SOS in the setting of enoxaparin prophylaxis and low intensity conditioning regimens.

Alternative Donor Options for Hematopoietic Stem Cell Transplantation

While human leukocyte antigen (HLA) matched sibling stem cell transplantation can eradicate chemotherapy-resistant leukemia, only 30% of patients are fortunate enough to have such a match. Therefore, studies began determining if HLA unrelated adult volunteer donors could provide an effective source of allogenic stem cells.

Characteristics and Outcomes of 38 Patients with Acute Myeloid Leukemia Evolving from Previous Well Characterized Myeloproliferative Disorder.

Antecedent hematological disorder (AHD) is a known adverse prognostic factor for patients with AML. AHD, however, encompasses a range of diverse hematological disorders. The characteristics and outcome of AML evolving from myeloproliferative disorders (MPD) without features of dysplasia remain poorly defined. Between 08/73 and 01/06, 76 patients were treated at the MD Anderson Cancer Center for AML evolving from a previous MPD. Of these, 38 were excluded from further analyses: inadequate pathological information for confident diagnosis of MPD (n=15), concomitant myelodysplastic features (n=11), chronic myelomonocytic leukemia (n=4), and less than twelve months between MPD diagnosis and AML occurrence (n=8). Baseline characteristics of remaining 38 patients with well-characterized MPD: median age 51 years (range 24–70); male 61%; cytogenetics - diploid 86%, del20q 14%; splenomegaly 27%. Diagnoses by WHO classification: polycythemia vera 47%; myelofibrosis 24%; essential thrombocythemia 18%; chronic MPD unclassifiable 11%. MPD therapy: hydroxyurea in 87% (median 3 years exposure), alkylating agent in 16%, and radioactive phosphorus in 8% of patients. AML occurred a median of 111 months (range 22 – 376) after MPD diagnosis. Median age at AML diagnosis was 65 years (range 40–81); cytogenetics were poor risk (−5, −7, 11q23, or ≥3 abnormalities) in 50% and intermediate risk in others (including diploid in 28%). Median survival was 23 weeks after AML diagnosis, and was not significantly different between patients who received AML therapy and those who received supportive therapy only (p=0.33) (figure 1), or between patients with intermediate and poor risk cytogenetics (p=0.12). Twenty-four patients received standard or high-dose Ara-C based induction chemotherapy: complete response (CR) rate was 38%, but responses were short-lived with all patients relapsing within 37 weeks in the absence of stem cell transplantation. Two patients underwent non-myeloablative matched sibling stem cell transplant in first CR: one remain leukemia free at 70 weeks, and the second relapsed 22 weeks post transplant but responded to re-induction chemotherapy, and remains alive at 38 weeks. AML evolving from a previous MPD carries an adverse prognosis, with very poor results from conventional induction chemotherapy. Exploration of novel agents in this patient population as a first line therapy is warranted. [Display omitted]

Donor leukocyte infusions after unrelated donor hematopoietic stem cell transplantation

Donor leukocyte infusions provide direct and potent graft-versus-tumor activity to treat relapse after allogeneic stem cell transplantation. Extensive data are available on the use of donor leukocyte infusion after matched-sibling stem cell transplantation, but reports are remarkably few on the use of donor leukocyte infusion after unrelated-donor stem cell transplantation. But the role for unrelated-donor leukocyte infusion is not well established. The dramatic success of donor leukocyte infusion to treat relapse after matched-sibling stem cell transplantation has led to the use of unrelated-donor leukocyte infusion in many patients. Several case studies suggest that unrelated-donor leukocyte infusion effectively induces direct graft-versus-tumor reactions with toxicity comparable to that of matched-sibling donor leukocyte infusion. Important issues include the relationship between dose and response/toxicity appropriate timing, dose, and schedule; and identification of the best tumor targets. In particular nonmyeloablative transplant strategies using unrelated donors are expanding rapidly, but relapse rates are high. There is a paucity of data on unrelated-donor leukocyte infusion in this setting. This review summarizes recent data on the use of unrelated-donor leukocyte infusion. We discuss anticipated outcomes and identify areas under active investigation in both ablative and nonmyeloablative unrelated-donor stem cell transplantation.

Enhancing Graft-vs-Tumor (GVT) Activity of Donor Leukocyte Infusions by Ex-Vivo Expansion and Activation with CD3/CD28 Co-Stimulation.

Donor leukocyte infusions (DLI) can induce a potent GVT effect for some patients who relapse after allogeneic stem cell transplantation (SCT). However, GVT effects are most pronounced for patients with early phase CML and less effective for acute leukemia or other hematologic malignancies. One hypothesis for disease resistance suggests that donor T cells are not appropriately activated in-vivo to induce an anti-tumor response. Ex-vivo co-stimulation of T cells via CD3 and CD28 might overcome disease-induced anergy and augment GVT activity. To explore the possibilities for this approach, we performed a phase I trial of DLI followed by escalating doses of ex-vivo co-stimulated donor T cells (activated, or aDLI) for patients with relapse after allogeneic SCT (excluding CP-CML). Activated donor T cells are produced by co-stimulation and expansion by exposure to Dynal-magnetic beads coated with anti-CD3 (OKT3) and anti-CD28. 18 patients (11 male) with a median age of 45 (range 12–57) were treated for relapse after matched sibling SCT for ALL (7), AML (4), NHL (2), CLL (1), CML-BC (1), myeloma (1), Hodgkin's disease (1) and lymphoblastic lymphoma (LL, 1). The median time from SCT to relapse was 11.5 mo (2–90) and relapse to DLI was 6 wks (2–31). All patients received standard unstimulated DLI (median 1.5 x 108 mononuclear cells/kg, range 0.9–3.5) followed 10 days later by ex-vivo co-stimulated donor T cells as aDLI. Patients with acute leukemia or BC-CML first received standard induction chemotherapy followed by DLI at the nadir. aDLI was dose escalated between every 3–4 patients from 1 x 106 CD3+ cells/kg to 1 x 108 CD3+ cells/kg in 5 dose levels. Ex-vivo co-stimulation resulted in a median 130-fold expansion of CD3+ cells (range 32–526) consisting of 56% CD4+ cells (range 30–84%) and 34% CD8+ cells (range 5–71%). The infusion of activated donor T cells was well tolerated with minor infusional toxicity at the highest dose level. Following aDLI, 7 patients developed acute GVHD (5 grade I-II skin only, and 2 grade III skin and liver). 2 patients have chronic GVHD limited to the oral mucosa and 2 have extensive chronic GVHD. No patient died of complications related to GVHD. 6/16 evaluable patients achieved CR (3/5 ALL, 1/4 AML, 1/1 CLL and 1/2 NHL) and 5 of 6 remain alive in CR a median of 25 mo after aDLI (range 8–43). 1 patient with ALL in CR relapsed 8 months after aDLI despite prior grade II aGVHD and died of disease progression. 1 patient with APML has residual disease detectable only by PCR 5 wks after aDLI and 4 other patients remain alive with disease (AML, ALL, HD, NHL) a median 4 mo after aDLI (range 3–5). 7 patients died of disease (3 ALL, CML-BC, MM, AML, LL) a median of 4 mo after aDLI (range 1–14). Functional assays show that immunity against pathogens such as EBV and CMV are retained after aDLI. These data demonstrate that adoptive transfer of co-stimulated allogeneic T cells is feasible, can induce a durable CR in a subset of patients, and does not result in excessive GVHD or other toxicity. In diseases where conventional DLI has been disappointing, response rates are impressive. Further studies to enhance GVT activity and tumor-specificity of aDLI are therefore warranted.

Unrelated Donor Leukocyte Infusions to Treat Relapse after Unrelated Donor Bone Marrow Transplantation

Allogeneic stem cell transplantation (SCT) may cure many patients with hematologic malignancies due to both the intensive conditioning therapy, and in many patients, the potent graft-vs-leukemia (GVL) effect of the donor graft. The GVL effect is mediated in large part by mature T-cells contained in the donor graft and has been defined in detail in animal models of transplantation. The GVL activity has been observed in the clinical setting after SCT from both matched siblings and unrelated donors. The best demonstration and most direct evidence of GVL activity in humans come from the use of donor leukocyte infusions (DLI). For patients who relapse with chronic myelogenous leukemia after matched sibling SCT, infusions of leukocytes collected from the original transplant donor will re-establish complete and durable remission in 60-80% of patients. DLI is less effective for more advanced phases of CML and for patients who relapse with diseases other than CML. DLI after matched sibling SCT is complicated primarily by graft-vs-host disease (GVHD), marrow aplasia, and unfortunately, relapse in some cases. There has been little information regarding the use of unrelated DLI (UDLI). Available data now shows that despite initial concerns that UDLI would result in excessive toxicity, it is an effective approach to relapse after unrelated donor marrow grafting. Response rates are similar to those seen after the use of matched sibling DLI, and many remissions remain durable. Graft-vs-host disease is a frequent complication after UDLI though the incidence and severity of GVHD is also similar to the use of matched sibling DLI. It is not clear that the GVL and GVHD effects can be separated, since the majority of responding patients also develop GVHD. The most effective cell dose for UDLI has not been established, though there does not appear to be either a dose-response or dose-toxicity relationship from UDLI. Although second unrelated donor bone marrow transplantation (BMT) may cure a small minority of patients, GVL induction with UDLI offers a safer and potentially more effective therapy for relapsed leukemia, and offers insights in methods to manipulate the human immune system for therapeutic benefit.

Cyclophosphamide and other new agents for the treatment of severe aplastic anemia

Severe aplastic anemia (SAA) has a poor prognosis in the absence of treatment. Current accepted therapeutic strategies include allogeneic stem-cell transplantation and immunosuppression, both resulting in long-term survival in the majority of patients. Although human leukocyte antigen (HLA)-matched sibling stem-cell transplantation is highly effective, the 25% probability of finding a suitable sibling donor within a family renders this approach available to only a minority of patients. Transplantation using HLA-matched, unrelated donors carries a high risk of treatment failure along with considerable toxicity. While combined immunosuppression with both antithymocyte globulin (ATG) and cyclosporine A (CSA) produces hematologic improvement in most patients, relapse is common. Late evolution of aplastic anemia to other serious hematologic disorders, including paroxysmal nocturnal hemoglobinuria (PNH), myelodysplasia, and acute leukemia, is also a significant problem following treatment with ATG/CSA. Recently, results of immunosuppression in SAA with another potent immunosuppressive agent, Cyclophosphamide, were reported in a small number of patients. The overall response rate was similar to that seen with ATG/CSA, but relapse and late clonal disease were not observed during a long period of follow-up. A larger randomized trial comparing sustained hematologic response rates to either conventional immunosuppression with ATG/CSA or high-dose cyclophosphamide and CSA is now underway; secondary end points include response duration, event-free survival, and overall survival. Additionally, a number of protocols designed to test the efficacy of alternative immunosuppressive or immunomodulatory agents are being developed.