Lyle L. Sensenbrenner

Marrow Transplantation for Acute Nonlymphocytic Leukemia after Treatment with Busulfan and Cyclophosphamide

Fifty-one patients with acute nonlymphocytic leukemia (16 with end-stage disease, 17 in second or third remission or in early relapse, and 18 in first remission) were given infusions of HLA-identical sibling marrow after cytoreduction with high doses of busulfan and cyclophosphamide. Actuarial two-year survival rates were 0 per cent, 29 per cent, and 44 per cent, respectively. Twelve patients are still alive and in remission after 327 to 1488 days, with 10 surviving beyond two years. Acute graft-versus-host disease and viral pneumonia were the major causes of death. Leukemic cells failed to clear in one patient with end-stage disease, and a relapse with meningeal leukemia occurred in another. Only one other relapse was seen--in a patient given a transplant during a third remission. Survival was favorably affected by younger age and transplantation during first remission. We conclude that high-dose chemotherapy with busulfan and cyclophosphamide, followed by allogeneic-marrow transplantation, can produce long-term remission of acute leukemia. Chemotherapy with high-dose busulfan and cyclophosphamide before transplantation provides an effective alternative to cyclophosphamide and total-body irradiation before transplantation for the treatment of acute nonlymphocytic leukemia.

VENOOCCLUSIVE DISEASE OF THE LIVER FOLLOWING BONE MARROW TRANSPLANTATION

Review of 235 consecutive patients undergoing bone marrow transplantation was performed in order to define the clinical syndrome of venoocclusive disease of the liver (VOD) in these patients. Analysis of all patients with histologically proven VOD revealed a consistent clinical syndrome of liver dysfunction occurring within the first 3 weeks after marrow infusion. This was characterized by hyperbilirubinemia peaking at greater than or equal to 2 mg/dl with at least 2 of 3 other findings: hepatomegaly, ascites, and 5% or greater weight gain. VOD developed in 22% (52 of 235). A persistently elevated aspartate aminotransferase (SGOT) prior to transplant was associated with an increased risk of developing VOD by multivariate analysis (P = 0.0003), and acute leukemia in first remission was associated with a decreased risk (P = 0.02). Neither the preparative regimen (busulfan and cyclophosphamide versus cyclophosphamide and total body irradiation) nor the type of graft (allogeneic versus autologous) influenced the occurrence. Twenty-four of these 52 patients (47%) died with VOD (10% of the entire group). This makes VOD the third leading cause of death in our allogeneic graft recipients, and the second leading cause in our patients receiving autologous transplants. VOD is a common complication of bone marrow transplantation and has a specific clinical presentation, which usually allows diagnosis without the need of liver biopsy.

Durable Treatment-Free Remission after High-Dose Cyclophosphamide Therapy for Previously Untreated Severe Aplastic Anemia

Acquired severe aplastic anemia is a life-threatening bone marrow failure disorder characterized by hypocellular bone marrow and peripheral blood pancytopenia (1). Although most cases of severe aplastic anemia are idiopathic, bone marrow failure usually results from an immune-mediated attack on hematopoietic stem cells regardless of the inciting event (2, 3). In patients treated with antibiotics and blood transfusions alone, the mortality rate is 80% at 2 years (4). Transplantation of bone marrow from an HLA-identical sibling after administration of a high-dose cyclophosphamide (200 mg/kg of body weight) conditioning regimen cures up to 90% of children and young adults with aplastic anemia (5, 6). However, the outcome in older patients, especially those older than 40 years of age, is significantly worse because of the increased risk for graft-versus-host disease (7, 8). Another limitation of allogeneic bone marrow transplantation is that most patients lack an HLA-identical sibling donor. Antilymphocyte globulin or antithymocyte globulin combined with cyclosporine and high-dose methylprednisolone is the most widely used immunosuppressive regimen for patients with severe aplastic anemia who are not suitable candidates for allogeneic bone marrow transplantation (9, 10). Antithymocyte globulin and cyclosporine is effective therapy for aplastic anemia; it improves hematopoiesis in most patients (60% to 80%). However, in contrast to bone marrow transplantation, hematologic improvement is usually incomplete. Moreover, responding patients will frequently have relapse, be dependent on cyclosporine, or develop a secondary clonal hematopoietic disease (1115). Recently, we demonstrated that high-dose cyclophosphamide therapy without bone marrow transplantation can lead to durable complete remission in patients with severe aplastic anemia and other autoimmune disorders (1618). In our pilot study of 10 patients with severe aplastic anemia, normal hematopoiesis was restored in 7 patients; no surviving patient has had relapse or has developed a secondary clonal disorder in more than 10 years. Here, we report on a prospective multi-institutional study to further evaluate the efficacy and safety of high-dose cyclophosphamide therapy (200 mg/kg of body weight) without bone marrow transplantation in patients with newly diagnosed severe aplastic anemia. Methods Patients Since January 1996, 19 patients with previously untreated severe aplastic anemia referred to the Johns Hopkins Oncology Center or the University of Maryland (Baltimore, Maryland) or MCP Hahnemann University (Philadelphia, Pennsylvania) were enrolled in the protocol. Patients younger than 40 years of age with an HLA-identical sibling were prioritized to undergo allogeneic bone marrow transplantation; the priority treatment in older patients and patients without an HLA-identical sibling was high-dose cyclophosphamide therapy without bone marrow transplantation. Severe aplastic anemia was defined as bone marrow cellularity of less than 25% and decreased values for at least two of three blood counts (reticulocyte count < 60 109 cells/L, platelet count < 20 109 cells/L, and neutrophil count < 0.5 109 cells/L). Aplastic anemia was considered very severe if the patient met the criteria for severe aplastic anemia and had a neutrophil count less than 0.2 109 cells/L. All patients gave informed consent for study participation, and the study was approved by the institutional review boards of Johns Hopkins University, MCP Hahnemann University, and the University of Maryland. Exclusion criteria were age older than 70 years, serum creatinine concentration greater than 176.99 mol/L (2.0 mg/dL), a cardiac ejection fraction less than 0.45, or any underlying cancer. Pregnant women and women of childbearing age who were unwilling to take oral contraceptives were also excluded. Patients with active infections were not excluded. Patients with Fanconi anemia or other congenital forms of aplastic anemia were not treated on this protocol. Treatment Schedule Cyclophosphamide, 50 mg/kg, was administered over 1 hour through a Hickman catheter on days 1 through 4. The dose of cyclophosphamide was based on the ideal body weight as determined by the Metropolitan Life Table. Prophylaxis of hemorrhagic cystitis consisted of intravenous mesna (10 mg/kg) administered 30 minutes before and 3, 6, and 8 hours after cyclophosphamide administration. On day 10 (6 days after the last dose of cyclophosphamide), all patients received granulocyte colony-stimulating factor (5 g/kg daily) until the neutrophil count was 1.0 109 cells/L for 2 consecutive days. No patient received additional immunosuppressive therapy. Supportive Care Intravenous ondansetron, 32 mg, was administered to all patients 1 hour before each dose of cyclophosphamide. All patients received prophylactic antibiotic support consisting of fluconazole (400 mg/d), norfloxacin (400 mg/d), and valacyclovir (500 mg twice daily) beginning on the day after the last dose of cyclophosphamide and continuing until the neutrophil count exceeded 0.5 109 cells/L. Prophylaxis against Pneumocystis carinii pneumonia with sulfamethoxazole (800 mg) and trimethoprim (160 mg) was given every Monday and Tuesday for 6 months after therapy. Packed red blood cell transfusions were administered to maintain a hematocrit greater than 0.25. Platelet transfusions were administered for bleeding or to maintain a platelet count greater than 20 109 cells/L; after recovery of the neutrophil count to greater than 0.5 109 cells/L, the platelet transfusion threshold was reduced to 10 109 cells/L. Statistical Analysis Event times were measured in months from treatment with high-dose cyclophosphamide to time of independence from transfusion, complete remission, death, or last follow-up. Survival was defined as the time until death or last follow-up. Time to complete remission was defined as the time until independence from transfusion in association with normal blood counts for age and sex. Time to independence from transfusion was defined as the time until independence for more than 3 months and a neutrophil count greater than 0.5 109 cells/L without growth factor support. Nonresponders were patients who remained transfusion dependent or fulfilled criteria for severe aplastic anemia. The probabilities of survival and remission were estimated by using the KaplanMeier method. The 95% CIs for the probability estimates were calculated by using Greenwoods formula. Deaths were censored using the longest follow-up time for time to remission analyses. All analyses were performed by using Stata 6.0 (Stata Corp., College Station, Texas). Results Patient Characteristics From January 1996 to January 2000, 33 patients with untreated severe aplastic anemia were evaluated. Of these patients, 14 were not treated on this trial, including 11 patients who received an HLA-matched sibling allogeneic bone marrow transplant and 3 patients who elected to receive immunosuppressive therapy (2 at institutions outside of this study and 1 at Johns Hopkins Hospital). The other 19 patients were treated with high-dose cyclophosphamide: 13 were treated at the Johns Hopkins Oncology Center, 5 at MCP Hahnemann University, and 1 at the University of Maryland. No patients were excluded from analysis. Demographic and disease characteristics of the 19 patients treated with high-dose cyclophosphamide are shown in Table 1. Ten patients met criteria for severe aplastic anemia. Nine patients met criteria for very severe aplastic anemia, and 8 patients had active infections at the time of treatment with high-dose cyclophosphamide (2 had bacterial sinusitis, 1 had bacterial pneumonia, 1 had perirectal abscess, 1 had herpes zoster, 1 had cellulitis, and 2 had febrile neutropenia with unclear source). The median age was 47 years (range, 18 to 68 years). The cause of the aplastic anemia was classified as idiopathic in 15 patients. Drug-induced aplastic anemia was diagnosed in 2 patients, and 2 patients developed aplastic anemia in association with viral infections. Table 1. Patient Characteristics Toxicity All patients developed transient alopecia. Most patients had some nausea or vomiting that resolved a few days after completion of cyclophosphamide therapy. One patient developed transient hemorrhagic cystitis. Three patients died, two of presumed fungal infections on days 26 and 57 after cyclophosphamide therapy and one of an intracerebral hemorrhage on day 67 after cyclophosphamide therapy. These three patients were older than 50 years of age and had very severe disease and an active infection at the time of treatment. No other patient developed invasive fungal infections. The median number of hospitalized days was 30 (range, 1 to 102 days). Response and Survival The probability of survival at 24 months was 84% (95% CI, 59% to 95%); the median follow-up among survivors was 34 months (Figure 1). The probability of treatment-free remission (independence from transfusion) was 73% (95% CI, 51% to 91%) at 24 months (Figure 2), and the probability of complete remission was 65% (95% CI, 39% to 89%) at 50 months (Figure 2). All responding patients continue to have treatment-free remission with Karnofsky scores of 100. Hematopoietic reconstitution in these patients was gradual; it took a median of 49 days to achieve an absolute neutrophil count of 0.5 109 cells/L (Table 2) and a median of 36 months to achieve a complete remission. The median time to independence from transfusion of red blood cells and platelets was 11 months. Of the 13 patients who were followed for more than 1 year, 12 are in treatment-free remission (9 complete responses and 3 partial responses). No patient has had relapse; has required additional immunosuppressive therapy; or has developed paroxysmal nocturnal hemoglobinuria, myelodysplasia, or other malignant diseases. None of the 10 patients who had a neutrophil count greater than 0.2 109 cells/L before therapy

Dye-mediated photolysis of normal and neoplastic hematopoietic cells.

The purpose of this study was to determine the sensitivity to merocyanine 540 (MC 540)-mediated photolysis of normal human hematopoietic progenitor cells and four leukemia cell lines (Daudi, Raji, K562 and HL-60). Late erythroid progenitors were the most sensitive normal cells. Early erythroid progenitors were of intermediate sensitivity. Granulocyte/macrophage progenitors and multipotent progenitors were the least sensitive normal marrow cells. A combination of dye concentration, serum concentration, and illumination that eliminated 50% of multipotent progenitor cells reduced the concentration of leukemic cells by greater than or equal to 4.5 log. It is conceivable that this difference in photosensitivity can be exploited for the extracorporeal purging of autologous remission marrow grafts.

Antiviral activity of merocyanine 540.

Abstract Simultaneous exposure to the lipophilic dye merocyanine 540 (MC 540) and white light inactivates several enveloped viruses. The same treatment appears to have little or no effect on pluripotent hematopoietic stem cells, mature red cells, and mature leukocytes. At least some components of the clotting system are spared, too. The molecular basis of the virucidal effect of MC 540 and light is not yet completely understood. Based on what is known about the interactions of MC 540 with cells and artificial membranes, it seems likely that MC 540 binds to and damages the viral envelope. MC 540‐mediated photosensitization may have implications for the sterilization of bone marrow and blood products, the preparation of vaccines, and selected areas of antiviral therapy.

Sinus Disease in the Bone Marrow Transplant Population: Incidence, Risk Factors, and Complications

BACKGROUND Fever associated with sinus disease in the immunocompromised bone marrow transplant recipient requires prompt evaluation and therapy. Very little is known about the incidence, risk factors, and sequelae of nonsurgically treated sinus disease in this population. METHODS A retrospective review of 107 consecutive allogeneic and autologous bone marrow transplant recipients from August 1987 to July 1989 was performed to determine (1) the overall incidence of sinus disease; (2) factors that influence the development of sinus disease; and (3) the sequelae of sinus disease treated nonsurgically. RESULTS Overall 33 (31%) of 107 bone marrow transplant recipients had sinus disease defined as a radiographic abnormality with clinical symptoms. Eleven (10%) of 107 recipients had preexisting sinus disease. Sinus disease developed in 22 (21%) of 107 recipients after bone marrow transplantation. Sinus abnormalities were significantly higher among allografted bone marrow transplant recipients than among autografted recipients (p = 0.027). The diagnosis, stage of disease, cytoreductive regimen, or graft-vs.-host disease were not different between recipients in whom sinus disease did and did not develop. There were no deaths as a result of sinus complications. CONCLUSIONS Sinus disease developed in 21% of the studied population after bone marrow transplantation. Allogeneic recipients had a higher incidence of sinus disease than autologous recipients. There were no deaths attributed to sinus complications. All sinus disease in this bone marrow transplant population was treated medically. No patient required surgical intervention either before or after bone marrow transplantation.

A case-control study of aplastic anemia

A case-control interview study of aplastic anemia was conducted to evaluate suspected risk factors. Cases (N = 59) newly diagnosed during 1975-82 at 25 Baltimore area hospitals were compared with 59 individually matched (on age, sex and race) controls selected by random digit dialing. The average educational level was less for cases than controls. The major job-related findings were a significant excess for occupational exposure to paint (OR = 6.1; 95% C.I. = 1.2-29.7), further substantiated by a positive dose-response relationship, although painters were not at excess risk. An increased risk of occupational exposure to viruses (OR = 9.0; 95% C.I. = 0.8-105.6) was noted. Additional evidence implicating viral factors included a significant association with prior history of hepatitis (OR = 9.0; 95% C.I. = 1.0, 84.2) and an elevated risk for pre-diagnostic receipt of blood transfusions (OR = 7.1; 95% C.I. = 0.7-68.4). Risks were not increased for other occupational, residential, personal, or medical treatment exposures or for other viral infections, medical conditions, smoking or alcohol consumption prior to diagnosis. Because of the small number of subjects studied and the multiple comparisons examined, these findings should be interpreted cautiously and confirmation should be undertaken in larger, population-based studies.

Allogeneic, syngeneic and autologous marrow transplantation in the acute leukemias and lymphomas--Baltimore experiences.

Allogeneic bone marrow transplants (BMT) in acute lymphoblastic leukemia following cyclophosphamide (Cy) and total body irradiation (TBI) has resulted in disease-free survival (DFS) for CR1, CR2, and CR3 of 10/18 (56%), 16/30 (53%) and 4/17 (24%), respectively. Median follow-up of survivors was 25-43 months. One relapse was seen in CR1, 1 in CR2, and 6 in CR3. Allogeneic BMT in acute nonlymphoblastic leukemia (ANLL) following busulfan (Bu) resulted in DFS for CR1, CR2 + CR3 + early relapse of 21/47 (45%) and 13/41 (32%), respectively. Median DFS of survivors of all groups together was 36 months. DFS of all patients 20 years or less was 15/27 (56%) and 19/61 (31%) for ages 21-46. Three relapses of 47 (6%) were seen in CR1 and 6/41 (15%) in subsequent remission and early relapse. BMT of autologous BMT with 4-hydroperoxy-cyclophosphamide (4-HC) purged marrow in ANLL in CR1 (5), CR2 (32) and CR3 (9) following Bu + Cy resulted in DFS of 19/46 (41%) for 1-67 months (median 15 months). Twenty patients with prior risk non-Hodgkins lymphoma received 4-HC-purged marrow following Cy + TBI. DFS was 10/20 (50%) for 1.4-9.5 years (median 2.9 years).

Treatment of refractory non-Hodgkin’s lymphoma with intensive chemo-radiotherapy and autologous bone marrow transplantation

The observation that syngeneic marrow transplantation following supralethal cytoreductive therapy can produce relatively long-term disease-free survival in a significant fraction of patients with acute leukemia [1] and non-Hodgkin’s lymphoma [2, 3] suggests that cure of these diseases may be achieved with marrow transplantation without the hazards of graft-versus-host disease that accompany allogeneic marrow transplants.

SERUM INHIBITOR OF GRANULOCYTE COLONY FORMATION ASSOCIATED WITH NEUTROPENIA IN TYPE I DYSGAMMAGLOBULINEMIA

A sixteen year old white male with X-linked Type I Dysgammaglobulinemia developed persistent neutropenia (total neutrophils < 500/mm3) which was unresponsive to broad spectrum antibiotics, prednisone, or a single infusion of one liter of fresh frozen plasma. His plasma agglutinated neutrophils in a microagglutination assay. His plasma (or serum) inhibited colony formation in a dose dependent fashion in a granulocyte/macrophage colony forming assay using human bone marrow cells. Complete inhibition was obtained when the patients serum reached a final concentration of 5%. This effect was similar when either the patients or normal bone marrow cells were used. Heating the patients serum to 56°C for 30 minutes did not destroy the inhibitor. Bone marrow cells cultured in a medium containing 5% of the patients serum and 5% of normal serum were also completely inhibited. Seven total plasma exchange procedures were performed over three weeks during which time the peripheral neutrophil count returned to normal and no serum inhibitor could be demonstrated. Upon completion of the plasma exchange procedures, the peripheral neutrophil count fell within three days to less than 500/mm3 and the serum inhibitor of colony formation reappeared. Three weeks after the plasma exchange procedures, the neutrophil count spontaneously returned to normal while on broad spectrum antibiotic therapy.