Helmut Renges

A randomized comparison of once versus twice daily recombinant human granulocyte colony-stimulating factor (filgrastim) for stem cell mobilization in healthy donors for allogeneic transplantation.

To evaluate the schedule dependency of granulocyte colony‐stimulating factor (G‐CSF) (filgrastim) for stem cell mobilization, we conducted a randomized comparison in 50 healthy donors, with one subcutaneous daily injection of 10 µg/kg G‐CSF (n = 25) compared with twice injections daily of 5 µg/kg G‐CSF (n = 25). The two groups were well balanced for age, body weight and sex. G‐CSF application was performed on an out‐patient basis and leukapheresis was started in all donors on day 5. The most frequent side‐effects of G‐CSF were mild to moderate bone pain (88%), mild headache (72%), mild fatigue (48–60%) and nausea (8%) without differences between the two groups. The CD34+ cell count in the first apheresis was 5·4 × 106/kg donor weight (range 2·8–13·3) in the 2 × 5 µg/kg group compared with 4·0 × 106/kg (range 0·4–8·8) in the 1 × 10µg/kg group (P = 0·007). The target of collecting > 3·0 × 106 CD34+ cells/kg donor weight with one apheresis procedure was achieved in 24/25 (96%) donors in the 2 × 5 µg/kg group and in 17/25 (68%) donors in the 1 × 10 µg/kg group. The target of collecting > 5·0 × 106 CD34+ cells/kg in the first apheresis was achieved in 64% in the 2 × 5µg/kg group, but in only 36% in the 1 × 10 µg/kg group. The progenitor cell assay for granulocyte–macrophage colony‐forming units (CFU‐GM) and erythroid burst‐forming units (BFU‐E) was higher in the 2 × 5 µg/kg group than in the 1 × 10 µg/kg group (7·0 vs. 3·5 × 105/kg, P = 0·01; 6·6 vs. 5·0 × 105/kg; P = 0·1). Administering G‐CSF (filgrastim) at a dosage of 5 µg/kg twice daily rather than 10 µg/kg once daily is recommended; this leads to a higher CD34+ cell yield and requires fewer apheresis procedures without increasing toxicity or cost.

Comparable results in patients with acute lymphoblastic leukemia after related and unrelated stem cell transplantation

We report the results of 84 patients with ALL after related (n=46) or unrelated (n=38) allogeneic SCT. Mean recipient age was 23 years (range: 1–60) and median follow-up was 18 months (range: 1–133). Forty-three patients were transplanted in CR1; 25 in CR2 or CR3; four were primary refractory; four in PR; eight in relapse. The conditioning regimen consisted of TBI/VP16/CY (n=76), TBI/VP16 (n=2), TBI/CY (n=2), Bu/VP16/CY (n=4). The OS at 3 years was 45% (44% unrelated, 46% related). Univariate analysis showed a significantly better OS for patients <18 years (P=0.03), mismatched sex-combination (P=0.03), both with a stronger effect on increasing OS after unrelated SCT. Factors decreasing TRM were patient age <18 years (P=0.004), patient CMV-seronegativity (P=0.014), female recipient (P=0.04). There was no significant difference in TRM and the relapse rate was similar in both donor type groups. Multivariate analysis showed that factors for increased OS which remained significant were mismatched sex-combination (RR: 0.70,95% CI: 0.51–0.93, P=0.015), patient age < 18 years (RR: 0.66, 95% CI: 0.47–0.93, P=0.016). A decreased TRM was found for female patients (RR: 0.56, 95% CI: 0.33–0.98, P=0.042), negative CMV status of the patient (RR: 0.57, 95% CI: 0.36–0.90, P=0.015). Unrelated stem cell transplantation for high-risk ALL patients with no HLA-compatible family donor is justifiable.

Patient cytomegalovirus seropositivity with or without reactivation is the most important prognostic factor for survival and treatment‐related mortality in stem cell transplantation from unrelated donors using pretransplant in vivo T‐cell depletion with anti‐thymocyte globulin

We evaluated the cytomegalovirus (CMV) serostatus as a risk factor for survival and treatment‐related mortality (TRM) in 125 patients allografted from an unrelated donor between 1994 and 1999. All patients received pretransplant in vivo T‐cell depletion using rabbit anti‐thymocyte globulin (ATG). Only one patient had primary graft failure and severe grade III/IV graft‐versus‐host disease occurred in 14% of the patients. The overall survival (OS) at 3 years was 70% for CMV‐negative patients (n = 76) and 29% in the seropositive cohort (n = 49) (P > 0·001). In multivariate analyses, CMV seropositivity remained an independent negative prognostic factor for OS (RR: 2·1; CI: 1·2–3·8; P = 0·014), apart from age > 20 years (RR: 2·74; CI: 1·2–3·8; P = 0·004) and late leucocyte engraftment (RR: 2·4; CI: 1·2–4·9; P = 0·015). The TRM for all patients was 27%. Despite monitoring for CMV antigenaemia and preemptive therapy with ganciclovir when reactivation occurred, seropositive patients had a three times higher risk of fatal treatment‐related complications than seronegative patients. In multivariate analyses, CMV seropositivity remained the strongest independent negative factor for TRM (RR: 5·3; CI: 1·9–14·6; P = 0·002), followed by age > 20 years (RR: 4·8; CI: 1·3–18·1; P = 0·02) and delayed leucocyte engraftment (RR: 3·6; CI: 1·2–11; P = 0·02). The TRM was identical in seropositive patients with (n = 27) or without (n = 22) CMV reactivation (44% versus 50%). We conclude that CMV seropositivity, despite preemptive ganciclovir therapy and even without reactivation, is a major negative prognostic factor for survival as well as for TRM in unrelated stem cell transplantation using pretransplant in vivo T‐cell depletion with ATG.

Reduced-toxicity conditioning with treosulfan, fludarabine and ATG as preparative regimen for allogeneic stem cell transplantation (alloSCT) in elderly patients with secondary acute myeloid leukemia (sAML) or myelodysplastic syndrome (MDS)

We investigated a dose-reduced conditioning regimen consisting of treosulfan and fludarabine followed by allogeneic stem cell transplantation (SCT) in 26 patients with secondary AML or MDS. Twenty patients were transplanted from matched or mismatched unrelated donors and six from HLA-identical sibling donors. The median age of the patients was 60 years (range, 44–70). None of the patients was eligible for a standard myeloablative preparative regimen. No graft-failure was observed, and leukocyte and platelet engraftment were observed after a median of 16 and 17 days, respectively. Acute graft-versus-host disease (GvHD) grade II–IV was seen in 23% and severe grade III GvHD in 12% of the patients. No patients experienced grade IV acute GvHD. Chronic GvHD was noted in 36% of the patients, which was extensive disease in 18%. The 2-year cumulative incidence of relapse was 21%. The relapse rate was higher in patients beyond CR1 or with intermediate two or high risk MDS (P=0.02). The treatment-related mortality at day 100 was 28%. The 2-year estimated overall and disease-free survival was 36–34%, respectively. No difference in survival was seen between unrelated and related SCT.

Real-Time Quantitative Y Chromosome-Specific PCR (QYCS-PCR) for Monitoring Hematopoietic Chimerism after Sex-Mismatched Allogeneic Stem Cell Transplantation

Y chromosome-specific sequences can be used to detect remaining male cells after sex-mismatched allogeneic blood stem cell transplantation (HSCT) involving a male patient and female donor, which represents approximately 25% of all cases. We developed a quantitative Y chromosome-specific PCR assay (QYCS-PCR) based on the DFFRY gene for the determination of hematopoietic donor chimerism. We analyzed blood and marrow samples from more than 40 patients at various time points after both standard and nonmyeloablative allogeneic HSCT. We found that real-time PCR combines extreme sensitivity, with a detection level of less than 1 male in 100,000 female cells (<0.001%), with very good reproducibility, especially in the important range of minor host chimerism. QYCS-PCR results were in close agreement with data from other techniques as bcr/abl-PCR and/or fluorescent in situ hybridization (FISH) analysis. In two relapsed patients, increasing numbers of Y-positive hematopoietic cells indicated recurrence of malignant disease prior to clinical confirmation. In conclusion, quantitative Y chromosome-specific PCR is a promising approach for monitoring the extent of chimerism in blood and other tissues after sex-mismatched hematopoietic stem cell transplantation (HSCT) or organ transplantation.

In vivo T cell depletion with pretransplant anti-thymocyte globulin reduces graft-versus-host disease without increasing relapse in good risk myeloid leukemia patients after stem cell transplantation from matched related donors

One-hundred and two patients with good risk myeloid leukemia (CML first chronic phase or AML first CR) were transplanted from HLA-related donors after conditioning with (n = 45) or without anti-thymocyte globulin (ATG) (n = 57). One graft failure was observed in the non-ATG and none in the ATG group. The median time to leukocyte engraftment (>1 × 109/l) was 16 (range 12–33) in the ATG group and 17 days (range 11–29) in the non-ATG group (NS) and for platelet engraftment (>20 × 109/l) 24 and 19 days (P = 0.002), respectively. Acute GVHD grade II–IV was observed in 47% of the non-ATG and in 20% of the ATG group (P = 0.004). Grade III/IV GVHD occurred in 7% of the ATG and in 32% of the non-ATG group (P = 0.002). Chronic GVHD was seen in 36% and 67% (P = 0.005), respectively. After a median follow-up of 48 months (range 2–128), the 5-year estimated OS is 66% (95% KI: 51–81%) for the ATG group and 59% (95% KI: 46–72%) for the non-ATG group (NS). The 5-year estimated DFS is 64% (95% KI: 50–78%) for ATG and 55% (95% KI: 43–67%) for the non-ATG regimen (NS). The 5-year probability of relapse was 5% in the ATG and 15% in the non-ATG group (NS). ATG as part of the conditioning regimen leads to a significant reduction in GVHD without increase of relapse in patients with myeloid leukemia after stem cell transplantation from HLA-related donors.

Use of a five-agent GVHD prevention regimen in recipients of unrelated donor marrow

A five-agent GVHD prophylaxis programme consisting of cyclosporin A, methotrexate, anti-thymocyte-globulin, pentaglobin and metronidazol was given to 48 recipients of unrelated donor marrow with chronic myelogenous leukemia, acute leukemia, myelodysplastic syndromes, and familiar lymphocytic hemophagocytosis of an average age of 33.5 (0.6–56) years. GVHD grades II–IV occurred in 18 patients (39%) and grades III–IV in five patients (11%). Chronic GVHD developed in nine patients (23%), three limited and six extensive. Fifteen patients died. Clinical relapse was detected in eight patients. Four patients died as a consequence of the underlying disease and subsequent treatment, 11 patients died of transplant-related causes. After a median follow-up of 19 months, the overall and disease-free survival are 67% and 62%, respectively. Survival by age is as follows: 0–19 years: 12/13 patients; 20–39 years: 14/25 patients; 40–59 years: 7/10 patients. The five-agent GVHD prophylaxis regimen is effective. Matched-unrelated donor transplants can be carried out safely in patients younger than 50 years of age. The results in patients younger than 20 years of age should encourage matched-unrelated donor transplants at earlier stages of the disease.

Dose‐reduced conditioning regimen followed by allogeneic stem cell transplantation in patients with myelofibrosis with myeloid metaplasia

Summary. Three patients with myelofibrosis received allogeneic stem cell transplantation after a dose‐reduced conditioning regimen of busulphan (8 mg/kg), fludarabine (180 mg/m2) and antithymocyte globulin (4 × 10 mg/kg). The median age at transplantation was 51 years (range 44–58). All patients engrafted with a leucocyte count > 1·0 × 109/l after a median of 18 d (range 16–20). Grade II acute skin graft‐versus‐host disease (GvHD) occurred in one patient. One limited and one extensive chronic GvHD was observed. All patients achieved complete haematological remission. In one patient the fibrosis resolved completely 180 d post transplant. All patients are alive 126, 466 and 764 d after transplantation.

A fludarabine-based dose-reduced conditioning regimen followed by allogeneic stem cell transplantation from related or unrelated donors in patients with myelodysplastic syndrome.

We investigated the feasibility and efficacy of a fludarabine-based dose-reduced conditioning regimen followed by stem cell transplantation from related (n = 5) or unrelated HLA-matched donors (n = 7) in 12 patients with high risk MDS, who were not eligible for a standard myeloablative conditioning regimen. The conditioning regimen consisted of fludarabine 30 mg/m2 daily for 6 days, busulfan 4 mg/kg daily for 2 days and anti-thymocyte globulin (ATG, rabbit) 10 mg/kg daily for 4 days in 11 patients, while one patient received fludarabine, ATG, cyclophosphamide and thiotepa. Graft-versus-host disease prophylaxis consisted of cyclosporine and a short course of methotrexate. The median age of the patients was 53 years (range 37–59). The median percentage of blasts in bone marrow aspirate at transplantation was 15% (range <5% to 35%). Diagnosis at transplant was RA (n = 1), RAEB (n = 5), RAEB-T (n = 5) and sAML (n = 1). A complex karyotype including monosomy 7 was noted in five patients. The reasons for using a dose-reduced conditioning regimen were prior autologous/syngeneic BMT (n = 4), active fungal infection (n = 2) or age/reduced performance status (n = 6). Engraftment was observed in all patients with complete donor chimerism. The incidence of acute GVHD (grade II–IV) was 33%. Eight patients died during follow-up due to relapse (n = 4), liver toxicity (n = 2), aspergillus (n = 1) or aGVHD grade IV (n = 1). After a median follow-up of 19 months, the 2-year estimated disease-free survival is 12% (95% CI: 2–23%) and the overall survival is 26% (95% CI: 4–52%). Fludarabine dose-reduced conditioning prior to allogeneic stem cell transplantation in high risk MDS patients, who were not eligible for standard transplantation, resulted in stable engraftment with complete chimerism, but the toxicity and relapse rate were considerable. Bone Marrow Transplantation (2001) 28, 643–647.

Stem cell mobilisation with 16 μg/kg vs 10 μg/kg of G-CSF for allogeneic transplantation in healthy donors

We compared two doses of recombinant human granulocyte-stimulating factor (G-CSF) for stem cell mobilisation in 90 healthy donors for allogeneic stem cell transplantation in a retrospective analysis. Group I (n = 46) received 10 μg/kg G-CSF (filgrastim) given as 5 μg/kg twice daily, and group II (n = 44) received 16 μg/kg, given as 8 μg/kg twice daily with a 12-h interval. The groups were well-balanced for age and body-weight. G-CSF application was performed on an out-patient basis, and leukapheresis was started in all donors on day 5. The most frequent side-effects of G-CSF were grade I/II, bone pain, headache and fatigue in both groups, whereas grade III of bone pain, headache and fatigue occurred in the 2 × 8 μg/kg group only. One serious non-fatal event with non-traumatic spleen rupture occurred in the 2 × 5 μg/kg group. The CD34+cell count in the first apheresis of all donors was 5.1 × 106/kg donor weight (range, 1.5–19.3). The CD34+ cell harvest was higher in the 2 × 8 μg/kg group than in the 2 × 5 μg/kg group (7.1 × 106/kg vs 4.9 × 106/kg; P = 0.09). The target of collecting >5.0 × 106 CD34+ cells/kg donor weight with one apheresis procedure was achieved in 45% of group I and in 61% of group II, respectively. Administering G-CSF at a dosage of 8 μg/kg twice daily leads to a higher CD34+ cell yield than a dosage of 2 × 5 μg/kg, but is associated with increased toxicity and higher cost.