Ely Benaim

Cidofovir for the treatment of adenoviral infection in pediatric hematopoietic stem cell transplant patients

Background. Adenovirus (ADV) infections are associated with significant morbidity and mortality after hematopoietic stem cell transplantation (HSCT). The virus is endemic in the general pediatric population and frequently causes severe disease in immunocompromised patients, especially children. We report our experience with cidofovir (CDV) for treatment of ADV infection in 57 HSCT patients, median age 8 years (range 0.5–26). Methods. Peripheral blood was prospectively screened weekly on all patients for ADV by quantitative real-time PCR for the first 100 days post-HSCT or longer if clinically indicated. Cultures for viral pathogens were performed from other involved sites. Upon detection of ADV by PCR, culture or tissue histopathology, CDV was given intravenously at 5 mg/kg weekly for 2 consecutive weeks, then every 2 weeks until 3 consecutive ADV-negative samples were documented from all previously invoved sites. Results. The clinical manifestations of ADV infection were: diarrhea (53%), fever (21%), hemorrhagic cystitis (12%), and pneumonitis (11%). Eight patients (14%) presented with disseminated disease. CDV treatment resulted in complete resolution of clinical symptoms in 56 (98%) patients in whom the virus became undetectable by all methods. One patient died due to ADV pneumonitis. No cases of dose-limiting nephrotoxicity were observed. Conclusions. Cidofovir appeared safe and effective for the treatment of ADV infection in this predominantly pediatric HSCT population. Vigilant surveillance and early treatment with CDV can prevent the poor outcomes associated with ADV disease. A larger prospective study is needed to further determine the role of CDV in the treatment of ADV after HSCT.

Feasibility of Four Consecutive High-Dose Chemotherapy Cycles With Stem-Cell Rescue for Patients With Newly Diagnosed Medulloblastoma or Supratentorial Primitive Neuroectodermal Tumor After Craniospinal Radiotherapy: Results of a Collaborative Study

PURPOSE This study was designed to determine the feasibility and safety of delivering four consecutive cycles of high-dose cyclophosphamide, cisplatin, and vincristine, each followed by stem-cell rescue, every 4 weeks, after completion of risk-adapted craniospinal irradiation to children with newly diagnosed medulloblastoma or supratentorial primitive neuroectodermal tumor (PNET). PATIENTS AND METHODS Fifty-three patients, 19 with high-risk disease and 34 with average-risk disease, were enrolled onto this study. After surgical resection, high-risk patients were treated with topotecan in a 6-week phase II window followed by craniospinal radiation therapy and four cycles of high-dose cyclophosphamide (4,000 mg/m2 per cycle), with cisplatin (75 mg/m2 per cycle), and vincristine (two 1.5-mg/m2 doses per cycle). Support with peripheral blood stem cells or bone marrow and with granulocyte colony-stimulating factor was administered after each cycle of high-dose chemotherapy. Treatment of average-risk patients consisted of surgical resection and craniospinal irradiation, followed by the same chemotherapy given to patients with high-risk disease. The expected duration of the chemotherapy was 16 weeks, with a cumulative cyclophosphamide dose of 16,000 mg/m2 and a planned dose-intensity of 1,000 mg/m2/wk. RESULTS Fifty of the 53 patients commenced high-dose chemotherapy, and 49 patients completed all four cycles. The median length of chemotherapy cycles one through four was 28, 27, 29, and 28 days, respectively. Engraftment occurred at a median of 14 to 15 days after infusion of stem cells or autologous bone marrow. The intended dose-intensity of cyclophosphamide was 1,000 mg/m2/wk; the median delivered dose-intensity was 1,014, 1,023, 974, and 991 mg/m2/wk for cycles 1 through 4, respectively; associated median relative dose-intensity was 101%, 102%, 97%, and 99%. No deaths were attributable to the toxic effects of high-dose chemotherapy. Early outcome analysis indicates a 2-year progression-free survival of 93.6% +/- 4.7% for the average-risk patients. For the high-risk patients, the 2-year progression-free survival is 73.7% +/- 10.5% from the start of therapy and 84.2% +/- 8.6% from the start of radiation therapy. CONCLUSION Administering four consecutive cycles of high-dose chemotherapy with stem-cell support after surgical resection and craniospinal irradiation is feasible in newly diagnosed patients with medulloblastoma/supratentorial PNET with aggressive supportive care. The early outcome results of this approach are very encouraging.

T‐cell alloreactivity dominates natural killer cell alloreactivity in minimally T‐cell‐depleted HLA‐non‐identical paediatric bone marrow transplantation

Summary. Natural killer (NK) cell alloreactivity resulting from killer immunoglobulin‐like receptor (KIR) ligand incompatibility improves outcomes in patients receiving extensively T‐cell‐depleted bone marrow (BM) grafts. Patients with KIR ligand incompatibility are at risk for donor T‐cell alloreactivity. We investigated the relative significance of NK‐cell and T‐cell alloreactivity in 105 paediatric patients who received a minimally T‐cell‐depleted human leucocyte antigen‐non‐identical BM transplantation. Donor NK‐cell incompatibility did not improve patient outcome [engraftment, graft‐versus‐host disease (GVHD), relapse or overall survival]. In contrast, donor T‐cell incompatibility was a risk factor for acute GVHD, chronic GVHD and death. Thus, T‐cell alloreactivity dominated that of NK cells in minimally T‐cell‐depleted grafts.

Rapid immune reconstitution after a reduced‐intensity conditioning regimen and a CD3‐depleted haploidentical stem cell graft for paediatric refractory haematological malignancies

The main obstacles to successful haploidentical haematopoietic stem cell transplantation from a mismatched family member donor are delayed immune reconstitution, vulnerability to infections and severe graft‐versus‐host disease (GvHD). We designed a reduced‐intensity conditioning regimen that excluded total body irradiation and anti‐thymocyte globulin in order to expedite immune reconstitution after a CD3‐depleted haploidentical stem cell transplant. This protocol was used to treat 22 paediatric patients with refractory haematological malignancies. After transplantation, 91% of the patients achieved full donor chimaerism. They also showed rapid recovery of CD3+ T‐cells, T‐cell receptor (TCR) excision circle counts, TCRβ repertoire diversity and natural killer (NK)‐cells during the first 4 months post‐transplantation, compared with those results from a group of patients treated with a myeloablative conditioning regimen. The incidence and extent of viremia were limited and no lethal infection was seen. Only 9% of patients had grade 3 acute GvHD, while 27% patients had grade 1 and another 27% had grade 2 acute GvHD. This well‐tolerated regimen appears to accelerate immune recovery and shorten the duration of early post‐transplant immunodeficiency, thereby reducing susceptibility to viral infections. Rapid T‐cell reconstitution, retention of NK‐cells in the graft and induction of low grade GvHD may also enhance the potential anti‐cancer immune effect.

Respiratory Virus Infections in Pediatric Hematopoietic Stem Cell Transplantation

Respiratory virus infections (RVI) have become an increasingly appreciated problem in the hematopoietic stem cell transplant (HSCT) population. A retrospective analysis of 274 patients undergoing 281 HSCT at St. Jude Childrens Research Hospital from January 1994 through December 1997 was performed. Medical and clinical laboratory records were reviewed beginning at the onset of conditioning through the year following each HSCT, and the analysis was done for the first RVI only. Thirty-two (11%) of 281 HSCT cases developed a RVI during the first year post-HSCT. The most frequent cause of RVI was human parainfluenza virus type 3. Univariate analysis was performed to determine the association between risk factors and the cumulative incidence of RVI. Respiratory viruses are frequent causes of infections in the first year post-HSCT in the pediatric population. Only allogeneic transplant and the degree of acute or chronic graft versus host disease were found to be statistically significant risk factors for RVI.

Hemorrhagic Cystitis after Allogeneic Bone Marrow Transplantation in Children: Clinical Characteristics and Outcome

Hemorrhagic cystitis (HC) is a well-documented adverse event experienced by patients undergoing hematopoietic stem cell transplantation. When severe, HC causes significant morbidity, leads to renal complications, prolongs hospitalization, increases health-care costs, and occasionally contributes to death. We retrospectively studied the medical records of 245 children undergoing an initial allogeneic bone marrow transplantation for malignant disease at St. Jude Childrens Research Hospital between 1992 and 1999 to describe the clinical course of HC in all patients and to identify the risk factors for HC in this cohort. Conditioning regimens included cyclophosphamide, cytarabine, and total body irradiation. Grafts from unrelated or mismatched related donors were depleted of T lymphocytes, whereas matched sibling grafts were unmanipulated. All patients received cyclosporine as prophylaxis for graft-versus-host disease. Recipients of grafts from matched siblings also received pentoxifylline or short-course methotrexate. Severe HC developed in 27 patients (11.0%). The median duration of HC was 73 days (range, 5-619 days); 12 patients had ongoing HC at the time of death. In univariate analyses, patients were at increased risk of severe HC if they were male (P =.021) or had received T cell-depleted grafts (P =.017), grafts from unrelated donors (P =.021), a lower total nucleated cell dose (P =.032), or antithymocyte globulin (P =.0446). Multiple regression analysis revealed male sex (beta =.97; P =.027) and unrelated donor graft recipients (beta =.83; P =.039) to be significant factors.

Improved cerebrovascular patency following therapy in patients with sickle cell disease: Initial results in 4 patients who received HLA-identical hematopoietic stem cell allografts

To test whether magnetic resonance angiography can document the evolution of vasculopathy in patients with sickle cell disease, we reviewed records to identify all patients who underwent magnetic resonance angiography from 1993 to 1999. Of 512 angiographies performed, 105 were of sickle cell disease patients, and 24 sickle cell disease patients 7 years of age or older underwent baseline and follow‐up examinations. Films were paired by patient, blinded as to examination date and treatment, and quantitatively compared. Four patients who received allogeneic bone marrow transplantation were compared to 7 patients who received other therapy and to 13 untreated patients. Quantitative analysis revealed a 10% increase in the measured diameter of 64 vessels (p = 0.001) following any treatment. Patients who had undergone allogeneic bone marrow transplantation exhibited a 12% increase in the lumen of 22 vessels (p = 0.041), whereas patients treated with chronic transfusion or hydroxyurea exhibited an 8% increase in 42 vessels (p = 0.016). In 2 patients with severe stenosis, the artery normalized after transplantation, and the blood flow rate was reduced in all patients who underwent transplantation. In untreated patients, there was a trend for the size of the arterial lumen to decrease, which is consistent with disease progression. Results suggest that treatment can reverse progression of vasculopathy. Bone marrow transplantation may enable stenoses to heal and can substantially reduce cranial blood velocity, suggesting that allogeneic bone marrow transplantation may prevent infarction or brain damage. Ann Neurol 2001;49:222–229

Effective donor lymphohematopoietic reconstitution after haploidentical CD34+ -selected hematopoietic stem cell transplantation in children with refractory severe aplastic anemia

Summary:Peritransplant toxicity and a delay in effective immune reconstitution have limited the utility of alternate donor transplantation for children with refractory severe aplastic anemia. We have assessed the effectiveness of infusing large numbers of highly purified haploidentical CD34+ cells after immunoablative conditioning in three patients who had failed intensive immunosuppression, lacked unrelated donors, and had active or recent serious infections. One patient rejected the first infusion, but engrafted after a second infusion from the same donor. This patient died 4 months after hematopoietic stem cell transplantation with no evidence of lymphoid reconstitution. Two patients experienced mixed chimerism requiring treatment with antibodies and/or donor lymphocyte infusion. Both currently survive more than 1 year after transplantation with normal blood counts, 100% donor engraftment, effective lymphoid reconstitution, and no chronic graft-versus-host disease. We observed functional thymopoiesis as measured by lymphocyte immunophenotyping, T cell receptor excision circles and T cell receptor Vβ spectratyping complexity analysis. Further study is required to validate the initial promise of these preliminary observations.

Effects of Retroviral-Mediated MDR1 Expression on Hematopoietic Stem Cell Self-Renewal and Differentiation in Culturea

Abstract: Ex vivo expansion of hematopoietic stem cells would be useful for bone marrow transplantation and gene therapy applications. Toward this goal, we have investigated whether retrovirally‐transduced murine stem cells could be expanded in culture with hematopoietic cytokines. Bone marrow cells were transduced with retroviral vectors expressing either the human multidrug resistance 1 gene (HaMDR1), a variant of human dihydrofolate reductase (HaDHFR), or both MDR1 and DHFR in an internal ribosomal entry site (IRES)‐containing bicistronic vector (HaMID). Cells were then expanded for 15 days in cultures stimulated with interleukin (IL)‐3, IL‐6, and stem cell factor. When very low marrow volumes were injected into lethally irradiated recipient mice, long‐term reconstitution with 100% donor cells was seen in all mice injected with HaMDR1‐ or HaMID‐transduced cells. By contrast, engraftment with HaDHFR‐ or mock‐transduced cells ranged from partial to undetectable despite injection of significantly larger marrow volumes. In addition, mice transplanted with expanded HaMDR1‐ or HaMID‐transduced stem cells developed a myeloproliferative disorder that was characterized by an increase in abnormal peripheral blood leukocytes. These results show that MDR1‐transduced stem cells can be expanded in vitro with hematopoietic cytokines, but indicate that an increased stem cell division frequency can lead to stem cell damage.

Effect of HLA class I or class II incompatibility in pediatric marrow transplantation from unrelated and related donors

The degree of histoincompatibility that can be tolerated, and the relative importance of matching at individual HLA class I and class II locus in bone marrow transplantation (BMT) has not been established. We hypothesized that matching for HLA-DR may not be more important than matching for HLA-A or HLA-B in selection of a donor for successful BMT. We retrospectively analyzed the outcomes of 248 consecutive pediatric patients who received allogeneic BMT from related donors (RD, n = 119) or unrelated donors (URD, n = 129). HLA-A and HLA-B were serologically matched, and HLA-DRB1 were identical by DNA typing in 69% of donor-recipient pairs. Most patients (89%) had hematologic malignancies; the rest had aplastic anemia or a congenital disorder. One HLA-A antigen mismatch was associated with a decrease in survival (p = 0.003) and a delay in granulocyte engraftment (p = 0.02) in recipients of RD marrow; as well as a decrease in survival (p = 0.02) and the development of severe acute graft-versus-host disease (GVHD) (p = 0.03) in recipients of URD marrow. One HLA-B antigen mismatch was associated with a decrease in the survival (p = 0.05) and the development of severe GVHD (p = 0.0007) in recipients of RD marrow. One HLA-DRB1 allele mismatch was associated only with a decrease in the survival (p = 0.0003) of recipients of RD marrow. Results of this study suggest that disparity in HLA-A and HLA-B antigens may not be better tolerated than disparity in HLA-DR allele in allogeneic BMT. Further studies are warranted to confirm our results.