Farid Boulad

Infusions of Donor Leukocytes to Treat Epstein-Barr Virus-Associated Lymphoproliferative Disorders after Allogeneic Bone Marrow Transplantation

BACKGROUND Lymphoma associated with Epstein-Barr virus (EBV) is a complication of bone marrow transplantation that responds poorly to standard forms of therapy. The lymphoma is usually of donor origin. We hypothesized that treatment with infusions of donor leukocytes, which contain cytotoxic T cells presensitized to EBV, might be an effective treatment. METHODS We studied five patients in whom EBV-associated lymphoproliferative disorders developed after they received a T-cell-depleted allogeneic bone marrow transplant. Biopsy specimens were immunophenotyped, subjected to the polymerase chain reaction to determine the origin of the lymphoma (donor or host) and to detect the presence of EBV, and analyzed by Southern blotting for the presence of the clonal EBV genome and immunoglobulin-gene rearrangement. Patients were treated with infusions of unirradiated donor leukocytes at doses calculated to provide approximately 1.0 x 10(6) CD3+ T cells per kilogram of body weight. RESULTS Histopathological examination of biopsy specimens from all five patients demonstrated monomorphic, malignant lymphomas of B-cell origin. Each of the four specimens that could be evaluated was of donor-cell origin. Evidence of clonality was found in two of the three samples adequate for study. EBV DNA was detected by the polymerase chain reaction in all five samples. In all five patients there were complete pathological or clinical responses. The responses were first documented histologically within 8 to 21 days after infusion. Clinical remissions were achieved within 14 to 30 days after the infusions and were sustained without further therapy in the three surviving patients for 10, 16, and 16 months. CONCLUSIONS In a small number of patients, infusions of unirradiated donor leukocytes were an effective treatment for EBV-associated lymphoproliferative disease that arose after allogeneic bone marrow transplantation.

Sequence and Structural Convergence of Broad and Potent HIV Antibodies That Mimic CD4 Binding

Anti-HIV broadly neutralizing antibodies with similar specificities and modes of binding were found in multiple HIV-infected individuals. Passive transfer of broadly neutralizing HIV antibodies can prevent infection, which suggests that vaccines that elicit such antibodies would be protective. Thus far, however, few broadly neutralizing HIV antibodies that occur naturally have been characterized. To determine whether these antibodies are part of a larger group of related molecules, we cloned 576 new HIV antibodies from four unrelated individuals. All four individuals produced expanded clones of potent broadly neutralizing CD4-binding-site antibodies that mimic binding to CD4. Despite extensive hypermutation, the new antibodies shared a consensus sequence of 68 immunoglobulin H (IgH) chain amino acids and arise independently from two related IgH genes. Comparison of the crystal structure of one of the antibodies to the broadly neutralizing antibody VRC01 revealed conservation of the contacts to the HIV spike.

Adoptive immunotherapy with unselected or EBV-specific T cells for biopsy-proven EBV+ lymphomas after allogeneic hematopoietic cell transplantation.

We evaluated HLA-compatible donor leukocyte infusions (DLIs) and HLA-compatible or HLA-disparate EBV-specific T cells (EBV-CTLs) in 49 hematopoietic cell transplantation recipients with biopsy-proven EBV-lymphoproliferative disease (EBV-LPD). DLIs and EBV-CTLs each induced durable complete or partial remissions in 73% and 68% of treated patients including 74% and 72% of patients surviving ≥ 8 days after infusion, respectively. Reversible acute GVHD occurred in recipients of DLIs (17%) but not EBV-CTLs. The probability of complete response was significantly lower among patients with multiorgan involvement. In responders, DLIs and EBV-CTLs regularly induced exponential increases in EBV-specific CTL precursor (EBV-CTLp) frequencies within 7-14 days, with subsequent clearance of EBV viremia and resolution of disease. In nonresponders, EBV-CTLps did not increase and EBV viremia persisted. Treatment failures were correlated with impaired T-cell recognition of tumor targets. Either donor-derived EBV-CTLs that had been sensitized with autologous BLCLs transformed by EBV strain B95.8 could not lyse spontaneous donor-derived EBV-transformed BLCLs expanded from the patients blood or biopsied tumor or they failed to lyse their targets because they were selectively restricted by HLA alleles not shared by the EBV-LPD. Therefore, either unselected DLIs or EBV-specific CTLs can eradicate both untreated and Rituxan-resistant lymphomatous EBV-LPD, with failures ascribable to impaired T-cell recognition of tumor-associated viral antigens or their presenting HLA alleles.

Gonadal function after bone marrow transplantation for acute leukemia during childhood

OBJECTIVE To examine the impact of bone marrow transplantation (BMT), using high-dose chemotherapy and hyperfractionated total body irradiation, on gonadal function in survivors of acute leukemia treated during childhood. STUDY DESIGN We conducted a retrospective study of 33 subjects (17 boys) who underwent a BMT for acute leukemia (acute lymphoblastic leukemia, n = 20; acute myelogenous leukemia, n = 13) at a single institution. All patients were prepubertal at the time of BMT (median age, 7.1 years (3.7 to 11.6 years)); at the time of their last examination the boys were a median of 14 years (10.4 to 17.1 years) of age and the girls were a median of 16.9 years (9.5 to 21.9 years) of age. RESULTS Of 17 boys, 14 (82%) entered puberty spontaneously and 13 demonstrated age-appropriate plasma concentrations of testosterone. Two boys (aged 10.5 and 11 years) remain clinically and hormonally prepubertal, and one boy has overt Leydig cell failure requiring androgen replacement therapy. Thirty-six percent of pubertal boys have elevated plasma concentrations of luteinizing hormone and 64% have raised levels of follicle-stimulating hormone. Boys with increased levels of luteinizing hormone were significantly younger at BMT (5.4 +/- 0.8 vs 7.8 +/- 0.8 years; p = 0.024). Of 16 girls, 9 (56%) had spontaneous puberty with onset of menarche at a median age of 13 years (9.5 to 15.8 years). Though six (67%) of these nine girls have had increased plasma concentrations of luteinizing and follicle-stimulating hormones, normalization has occurred in two during a period of 4 to 7 years. The remaining seven subjects required hormone replacement because of clinical and biochemical evidence of ovarian failure. One of these subjects has recovered ovarian function after 5 1/2 years. Female patients with ovarian failure were significantly older at BMT compared with female patients with spontaneous puberty/menarche (8.6 +/- 23 years vs 6.1 +/- 1.8; p = 0.03). CONCLUSION Our results indicate that most prepubertal boys undergoing BMT with chemotherapy and hyperfractionated total body irradiation can expect to enter and progress normally through puberty. For prepubertal girls treated with these regimens, at least 50% retain adequate ovarian function to enter puberty and menstruate regularly.

Respiratory syncytial virus infection following hematopoietic stem cell transplantation

Respiratory syncytial virus, one of the most common causes of respiratory infections in immunocompetent individuals, is frequently spread to recipients of HSCT by family members, other patients, and health care workers. In immunosuppressed individuals, progression from upper respiratory tract disease to pneumonia is common, and usually fatal if left untreated. We performed a retrospective analysis of RSV infections in recipients of autologous or allogeneic transplants. The incidence of RSV following allogeneic or autologous HSCT was 5.7% and 1.5%, respectively. Of the 58 patients with an RSV infection, 16 of 21 patients identified within the first post-transplant month, developed pneumonia. Seventy-two percent of patients received aerosolized ribavirin and/or RSV-IGIV, including 23 of 25 patients diagnosed with RSV pneumonia. In this aggressively treated patient population, three patients died of RSV disease, each following an unrelated HSCT.Bone Marrow Transplantation (2002) 29, 321–327. doi:10.1038/sj.bmt.1703365

Allogeneic bone marrow transplantation versus chemotherapy for the treatment of childhood acute lymphoblastic leukemia in second remission : A single-institution study

PURPOSE A retrospective analysis of the treatment of childhood acute lymphoblastic leukemia (ALL) in second remission (CR2) was undertaken at our institution to compare the outcome and prognostic factors of patients treated with chemotherapy or allogeneic bone marrow transplantation (BMT). PATIENTS AND METHODS Seventy-five children who suffered a medullary relapse and achieved a second remission were treated with either an unmodified allogeneic HLA-matched sibling BMT after hyperfractionated total body irradiation (TBI) and cyclophosphamide (n = 38) or chemotherapy according to institutional chemotherapy protocols (n = 37). To avoid the bias of survival from the attainment of second remission in favor of BMT, the final comparative statistical analysis used the landmark approach and comprised 37 and 29 patients from the BMT and chemotherapy groups, respectively RESULTS The disease-free survival (DFS) rate was 62% and 26% at 5 years, respectively, for the BMT and the chemotherapy groups (P = .03), with relapse rates of 19% and 67%, respectively, for these two groups (P = .01). There was an overall advantage for the BMT therapeutic approach, as compared with chemotherapy, for patients with ALL in CR2 (1) for patients with a WBC count (at diagnosis) of 20 x 10(9)/L or higher (DFS, 40% v 0%) and those with a WBC count of less than 20 x 10(9)/L (DFS, 73% v35%), (2) for patients whose duration of CR1 was less than 24 months (DFS 48% v 9%) and for patients whose duration of CR1 was 24 months or longer (DFS, 81% v 37%) and (3) for patients who were initially treated with intensive regimens incorporating more than five chemotherapy agents (DFS, 57% v 20%) and for patients treated with five agents or fewer (DFS, 72% v 32%). CONCLUSION In our single-institution series, unmodified HLA-matched allogeneic sibling transplants using hyperfractionated TBI and cyclophosphamide for patients with ALL in CR2 have resulted in superior outcome with a significantly improved probability of DFS and a lower relapse rate, as compared with those for patients treated with chemotherapy, regardless of the duration of first remission, the disease characteristics at diagnosis, or the intensity of prior treatment during first remission.

Fludarabine-based cytoreductive regimen and T-cell-depleted grafts from alternative donors for the treatment of high-risk patients with Fanconi anaemia

Eighteen consecutive patients aged 5·5–24 years with Fanconi anaemia and diagnoses of aplastic anaemia (n = 8), myelodysplastic syndrome (n = 4), acute myeloid leukaemia (n = 6), received allogeneic haematopoietic stem cell transplants from alternative donors. All patients had been transfused, 13 had previously been treated with androgens and 14 had a history of infection. Donors were related human leucocyte antigen (HLA) mismatched for eight patients, unrelated HLA mismatched for seven patients and unrelated HLA matched for three patients. Cytoreduction included single dose total body irradiation (450 cGy), fludarabine (150 mg/m2) and cyclophosphamide (40 mg/kg). Immunosuppression included antithymocyte globulin and tacrolimus. Grafts were granulocyte colony‐stimulating factor‐mobilized, CD34+ T‐cell‐depleted peripheral blood stem cells in 15 patients and T‐cell‐depleted marrows in three. All 18 patients engrafted with 100% donor chimaerism; only one patient developed graft‐versus‐host disease (GVHD). With a median follow‐up of 4·2 years, 13/18 patients were alive, 12 of these were disease‐free. Five‐year overall survival and disease‐free survival were 72·2% and 66·6% respectively. Immune reconstitution was achieved at approximately 6 months post‐transplant for most patients. These are encouraging results of T‐cell‐depleted transplants from alternative donors using fludarabine‐based cytoreduction in 18 high‐risk patients with Fanconi anaemia, with no evidence of rejection and minimal GVHD.

Long-term follow-up of patients treated with daclizumab for steroid-refractory acute graft-vs-host disease.

Daclizumab has been shown to have activity in acute GVHD, but appears to be associated with an increased risk of infection. To investigate further the long-term effects of daclizumab, we performed a retrospective review of 57 patients who underwent an allogeneic hematopoietic stem cell transplant from January 1993 through June 2000 and were treated with daclizumab for steroid-refractory acute GVHD. The median number of daclizumab doses given was 5 (range 1–22). GVHD was assessed at baseline, days 15, 29 and 43. By day 43, 54% patients had an improvement in their overall GVHD score, including 76% patients aged ⩽18. Opportunistic infections developed in 95% patients. Forty-three patients (75%) died following treatment with daclizumab. The causes of death included active GVHD and infection (79%), active GVHD (5%), chronic GVHD (2%) and relapse (14%). Patients with grade 3–4 GVHD had a significantly shorter median survival than patients with grade 1–2 GVHD (2.0 vs 5.1 months, P=0.001). Daclizumab has no infusion-related toxicity, is active in steroid-refractory GVHD, especially among pediatric patients, but is associated with significant morbidity and mortality due to infectious complications. Careful patient selection and aggressive prophylaxis against viral and fungal infections are recommended.

Graft-versus-Host Disease after Double-Unit Cord Blood Transplantation Has Unique Features and an Association with Engrafting Unit-to-Recipient HLA Match

Manifestations of and risk factors for graft-versus-host disease (GVHD) after double-unit cord blood transplantation (DCBT) are not firmly established. We evaluated 115 DCBT recipients (median age, 37 years) who underwent transplantation for hematologic malignancies with myeloablative or nonmyeloablative conditioning and calcineurin inhibitor/mycophenolate mofetil immunosuppression. Incidence of day 180 grades II to IV and III to IV acute GVHD (aGVHD) were 53% (95% confidence interval, 44 to 62) and 23% (95% confidence interval, 15 to 31), respectively, with a median onset of 40 days (range, 14 to 169). Eighty percent of patients with grades II to IV aGVHD had gut involvement, and 79% and 85% had day 28 treatment responses to systemic corticosteroids or budesonide, respectively. Of 89 engrafted patients cancer-free at day 100, 54% subsequently had active GVHD, with 79% of those affected having persistent or recurrent aGVHD or overlap syndrome. Late GVHD in the form of classic chronic GVHD was uncommon. Notably, grades III to IV aGVHD incidence was lower if the engrafting unit human leukocyte antigen (HLA)-A, -B, -DRB1 allele match was >4/6 to the recipient (hazard ratio, 0.385; P = .031), whereas engrafting unit infused nucleated cell dose and unit-to-unit HLA match were not significant. GVHD after DCBT was common in our study, predominantly affected the gut, and had a high therapy response, and late GVHD frequently had acute features. Our findings support the consideration of HLA- A,-B,-DRB1 allele donor-recipient (but not unit-unit) HLA match in unit selection, a practice change in the field. Moreover, new prophylaxis strategies that target the gastrointestinal tract are needed.

Transplantation in Remission Improves the Disease-Free Survival of Patients with Advanced Myelodysplastic Syndromes Treated with Myeloablative T Cell-Depleted Stem Cell Transplants from HLA-Identical Siblings

From 1985 to 2004, 49 patients with advanced myelodysplastic syndromes (MDS) (> or =5% blasts) or acute myeloid leukemia (AML) transformed from MDS underwent T cell depleted bone marrow or peripheral blood hematopoietic stem cell transplantation (HSCT) from HLA-identical siblings following conditioning with a myeloablative regimen that included total body irradiation (44 patients) or busulfan (5 patients). Thirty-six patients received chemotherapy (3 low dose and 33 induction doses) before conditioning, and 13 patients did not receive any chemotherapy. Prior to transplantation, 22 of the 36 treated patients were in hematologic remission; 4 were in a second refractory cytopenia phase (26 responders); 8 had failed to achieve remission; and 2 of the responders had progression or relapse of their MDS (10 failures). No post-transplantation pharmacologic prophylaxis for graft-versus-host disease (GVHD) was given. The median age was 48 yrs (range 13-61). Forty-five of the 49 patients engrafted; 2 had primary graft failure; and 2 died before engraftment. Only 3 patients developed acute GVHD (aGVHD) (grades I and III) and 1 chronic GVHD (cGVHD). At 3 yrs post-transplantation, the overall survival (OS) was 54% in the responders; 31% in the untreated group; and 0% in the failure group (P=.0004). The disease free survival (DFS) was 50%, 15% and 0% in each group respectively (P=.0008). In multivariate analysis, disease status before cytoreduction remained highly correlated with DFS (P<.001). The cumulative incidence (CI) of relapse at 2-yrs post-transplantation for the responders was 23%; for the untreated group was 38%; and for the failures was 50%. The CI of non-relapse mortality at 2-yrs post-transplantation, for the responders was 23%; for the untreated group was 38%; and for the failures was 40%. All survivors achieved a Karnofsky Performance Status (KPS) of > or =90. These results indicate that patients with advanced MDS who achieve and remain in remission or a second refractory cytopenia phase with chemotherapy before conditioning can achieve successful long-term remissions following a myeloablative T cell depleted allogeneic HSCT.