Use of peripheral blood stem cells for autologous transplantation in ...

Bone Marrow Transplantation, (1999) 24, 467–472  1999 Stockton Press All rights reserved 0268–3369/99 $15.00 http://www.stockton-press.co.uk/bmt

Use of peripheral blood stem cells for autologous transplantation in acute myeloid leukemia patients allows faster engraftment and equivalent disease-free survival compared with bone marrow cells G Visani, RM Lemoli, P Tosi, G Martinelli, N Testoni, P Ricci, MR Motta, F Gherlinzoni, G Leopardi, R Pastano, S Rizzi, PP Piccaluga, A Isidori and S Tura Institute of Haematology and Medical Oncology, ‘Seragnoli’, Bologna University, Bologna, Italy

Summary: We compared the feasibility and efficacy of autologous bone marrow (ABMT) and peripheral blood progenitor cell transplantation (PBSCT) performed after an identical induction/consolidation in adults with acute myeloid leukemia (AML). From January 1993 to June 1996 91 consecutive AML patients were enrolled in a program consisting of anthracycline-based induction and highdose cytarabine consolidation (NOVIA). Until May 1994 ABMT was performed; from June 1994, if PBSC collection was adequate, PBSCT was performed. Out of 88 evaluable patients, 73 obtained a complete remission (CR) and 15 were resistant. Allogeneic bone marrow transplantation was performed in 16 patients. Fortyfour (50%) were given autologous stem cell transplantation. ABMT was performed in 21 cases; twenty-nine patients were given G-CSF mobilization after NOVIA administration. An adequate number of PBSC was obtained in 23/29 (79%) cases, which were then reinfused. Median times to both neutrophil and platelet recovery from transplant were significantly shorter for the PBSC group (17 vs 36 days to 500 PMN/␮l, P ⬍ 0.01; 20 vs 150 days to 20 000 platelets/␮l, P ⬍ 0.02; 37 vs 279 days to 50 000 platelets/␮l, P ⬍ 0.03), as were days of hospitalization after the reinfusion (18 vs 33, P ⬍ 0.03) and median days to transfusion independence. Toxicity was not significant in either group. After a minimum follow-up for live patients of 24 months (longer than the mean time for relapse observed for the PBSC series – 14 months) the percentage of relapses was similar: 11 of 21 (52.4%) and 12 of 23 (52.1%) in the ABMT and PBSC groups, respectively. Our results indicate that autologous PBSC transplantation, performed after an intensive chemotherapy regimen, is not inferior to ABMT in terms of disease-free survival and allows faster recovery times and reduced need for tranfusion support. Keywords: leukemia; myeloid; autologous; transplantation; stem cells

Correspondence: Dr G Visani, Institute of Hematology and Medical Oncology, ‘Seragnoli’, Policlinico S Orsola, Via Massarenti, 9, 40138 Bologna, Italy Received 27 November 1998; accepted 23 March 1999

Standard induction/consolidation chemotherapy in acute myeloid leukemia (AML) patients results in a median remission duration of 12–18 months, with less than 30% free of disease at 5 years.1–3 When an HLA matched donor is not available, many centers currently program autologous bone marrow transplantation.4,5 In AML, ABMT generally leads to slow and often incomplete hematological recovery.6–12 The use of peripheral blood stem cells (PBSC), in addition to or instead of bone marrow for ABMT performed after classical induction regimens, has resulted in rapid neutrophil and platelet recovery.13–19 Despite these encouraging results, relapse remains the most frequent cause of treatment failure for AML patients undergoing autologous PBSC transplantation. While there is reasonable evidence for the safety and efficacy of PBSC for autologous transplantation in most malignant diseases, it has yet to be demonstrated in AML. Studies20–22 have failed to detect tumor cells in the peripheral blood of leukemic subjects during the early recovery phase from induction/consolidation chemotherapy. However, the high relapse rate previously described13,15,23,24 suggests that the much larger number of cells infused with PBSC may result in reinfusion of more residual malignant cells. The issue of leukemia-free autografting has recently been underlined by gene marking studies which demonstrated the contribution of residual contaminating tumor cells to disease relapse.25 In addition, exposure to growth factors could increase susceptibility to relapse.24 In order to minimize these possible risks, intensive induction/consolidation regimens capable of determining an in vivo purging superior to standard treatments have been proposed.17,26 The feasibility of PBSC collection after double induction and high-dose cytarabine-based consolidation has recently been described.27 Here, we report two groups of AML patients in first CR after an intensive induction/consolidation chemotherapy who were sequentially given ABMT with bone marrow cells or PBSC and analyze the effects of the program in terms of harvest efficacy, treatment-related toxicity, disease-free survival and overall survival.

PBSC transplantation vs ABMT in AML G Visani et al

468

Patients and methods Patients From January 1993 to June 1996, 91 adult patients with newly diagnosed AML were consecutively enrolled in the ICE/NOVIA protocol and treated in our Institution. The study design is shown in Table 1. The diagnosis of AML was made according to French–American–British (FAB) criteria.28 De novo AML was defined if no documented hematologic abnormality was identified more than 2 months before diagnosis. All the patients received a remission induction cycle (ICE) consisting of idarubicin 10 mg/m2 i.v. bolus per day on days 1, 3, 5; cytosine arabinoside 100 mg/m2 in continuous infusion (preceded at the start of infusion by 100 mg bolus injection) on days 1–10; etoposide 100 mg/m2/day i.v. on days 1–5. Complete remission (CR) was defined as the complete normalization of the morphologic,29 immunophenotypic and, when applicable, cytogenetic and molecular marrow picture, lasting for at least 1 month. Resistant patients were considered off study. Patients achieving CR received high-dose NOVIA consolidation: cytarabine 500 mg/m2/twice a day on days 1–6; mitoxantrone 12 mg/m2/day on days 4–6. From January 1993 to May 1994 patients underwent bone marrow harvest. From May 1994, all CR patients received G-CSF (10 ␮g/kg/day), starting at day +13 after NOVIA administration and continuing until the completion of stem cell leukapheresis in order to mobilize peripheral blood progenitor cells, provided that cytological and, when applicable, cytogenetic and molecular analysis had previously confirmed the achievement of CR after ICE therapy. When mobilization was absent or insufficient, bone marrow was subsequently harvested. Bone marrow collection and processing Median time from CR to harvest was 3 months (range 2– 6); bone marrow was harvested from the bilateral posterior iliac crest under general anesthesia; at least 1 x 108 harvested mononuclear cells per kg body weight (bw) were required for ABMT; no patient needed two harvests. Marrow buffy-coat was cryopreserved at ⫺1°C/min in 10% dimethylsulfoxide (DMSO) and 30% autologous plasma and stored at ⫺196°C in liquid nitrogen. Thawing was performed rapidly in a 37°C waterbath without removing DMSO, and the marrow was immediately infused intravenously. Leukapheresis procedures and PBSC processing Progenitor cell apheresis was begun after an absolute count of 10/␮l CD34+ cells was achieved during the recovery phase from consolidation therapy. As already reported,30 circulating stem cells were collected by using either a Fenwal CS3000 continuous flow blood cell separator (Baxter, Rome, Italy) with the modified procedure N.1 program or a Cobe Spectra separator (Cobe BCT, Lakewood, CO, USA) to obtain a minimum yield of 2 x 106 CD 34+ cells/kg. Non-mobilizing patients underwent bone marrow har-

vest, as previously described. Continuous flow leukapheresis was performed daily until the collection of a minimum number of 2 x 106 CD34+ cells/kg. The final apheresis product was concentrated and cryopreserved after addition of DMSO 10%. When applicable, cytogenetic and molecular analyses were performed on the product in order to confirm CR.22 Conditioning regimen and PBSCT/ABMT The preparative regimen consisted of busulfan 4 mg/kg/day for 4 days and cyclophosphamide (Cy) 60 mg/kg/day given for 2 days associated with Uromitexan (Mesna; Asta Medica, Milan, Italy). At the time of stem cell reinfusion (day 0), each bag was thawed and infused via a central line. Supportive care and hematological recovery All patients were nursed in reverse isolation single rooms until hospital discharge and received antimicrobial prophylaxis that consisted of oral nystatin and ciprofloxacin. Broad-spectrum intravenous antibiotics were promptly instituted in case of fever ⬎38.5°C during neutropenia. Platelet support was given when the platelet count was ⬍10 x 109/l. Packed red cells were administered at a hemoglobin level ⬍8 g/dl. Hemopoietic growth factors were not administered. Statistical analysis Age of patients and follow-up were expressed as mean ⫾ standard deviation; number of cells infused and time for hematological recovery were expressed as median (range). Events were relapse or death. The time of analysis was June 1998. Both overall survival and disease-free survival (DFS) were calculated according to the Kaplan–Meier estimate.31 Comparison between the DFS curves was conducted by the log-rank test according to Peto et al.32 Analysis was performed using the BMDP Statistical Software 1990 Edition. Two-sided P values were used throughout. P values were considered significant when ⬍0.05.

Results Outcome of induction/consolidation The outcomes are summarized in Table 1. Three patients died after induction therapy due to infections. Seventy-three out of the remaining 88 (83%) obtained CR after ICE; 15 (17%) were resistant, and were considered off study. Three out of 73 CR patients were removed because of toxicity. Seventy cases were then submitted to consolidation. Afterwards, 16 patients who had an HLA-matched identical sibling donor received allogeneic bone marrow transplantation. Of the remaining 54 cases, three refused further therapy; seven experienced an early relapse. Forty-four patients (50% of those evaluable) proceeded to the autologous transplantation procedure. Their characteristics are shown in Table 2.


1.00

1.00

0.90

0.90 0.80

0.80

0.70

0.70

ABMT

0.60

%

0.60

%

469

0.50

PBSCT

0.40

0.40

ABMT

0.30

PBSCT

0.50 0.30 0.20

0.20

0.10

0.10

0.00

0.00

0 0

12

24

36

48

12

60

Table 1 therapy

Design of the study and outcome of induction/consolidation

36

48

60

Months

Months Figure 1 Comparison of disease-free survival (ABMT vs PBSCT).

24

Figure 2 Relapse risk after autotransplant (ABMT vs PBSCT). Table 2 tation

Characteristics of patients submitted to autologous transplan-

INDUCTION Patients Sex (M/F) Age (⫾ s.d)

PBSCT

ABMT

P

23 12/11 42.7 (⫾10.5)

21 10/11 43.9 (⫾10.4)

NS

5 6 6 4 1 1

4 7 6 4 — —

11 2 2 1 2 2 2 1

11 3 2 — 1 2 1 1

10.0 (1.4–329.0)

5.4 (1.7–65.4)

ICE 91

3 Death during induction

88 Further therapy

15 Resistant

OFF PROTOCOL

7 Relapsed

73 CR

3 Out for toxicity

70 NOVIA CONSOLIDATION

60 BMT

NO HLA-ID DONOR

3 No further therapy

HLA-ID DONOR

FAB M1 M2 M4 M5 M6 Granulocytic tumor Karyotype Normal Inv(16) t(8;21) ⫹8 11q23 Complex Other ND WBC (⫻103/␮l)

44 Autologous BMT

23 PBSC

NS

16 Allogeneic BMT

21 ABMT

Marrow harvesting and/or PBSC mobilization Bone marrow was harvested from 21 patients from 2 to 6 months after CR; ABMT followed at a median of 6 months (range 4–8 months) from CR. In 29 cases, G-CSF was administered after consolidation therapy to mobilize PBSC. An adequate number of PBSC was obtained in 23 patients, with a mean of two collections. In the six non-mobilizing cases, marrow was successfully harvested and the patients underwent ABMT.

PBSC group (see Table 3). Median times necessary to recover 500 PMN/␮l and 20 000 and 50 000 platelets/␮l were significantly shorter in the PBSC than in the ABMT group (days to 500 PMN/␮l: 17 vs 36: P ⬍ 0.01; days to 20 000 platelets/␮l: 20 vs 150: P ⬍ 0.01; days to 50 000 platelets/␮l: 37 vs 279; P ⬍ 0.02). Transplant toxicity and disease status Mild or moderate mucositis was the most frequent form of toxicity. There was no significant difference in the other parameters (days of fever, i.v. antibiotic therapy, FUO, documented infection, extrahematological toxicity). DFS and clinical status after hospital discharge

Autologous transplantation and hematopoietic recovery The reinfusion parameters (nucleated cells, as well as CFUGM and CD34+ cells) were significantly higher for the

Patients in the ABMT group became independent of red blood cell transfusions at a median of 130 days (range 18– 196) which was significantly longer than for the PBSC


470

Table 3

Autologous transplantation: reinfusion parameters and hematological recovery PBSC

Time (CR-SCT) (months) (range) Nucleated cells reinfused (⫻108/kg) (range) CFU-GM (⫻104/kg) (range) CD34⫹ cells (⫻106/kg) (range) Median days (range) to ANC ⬎500/␮l Platelets ⬎20 000/␮l Platelets ⬎50 000/␮l Hospital discharge

Table 4

ABMT

4.8 8.3 33 6.9

(2–8) (2.4–23) (0.3–155) (2.5–14.6)

17 20 37 19

(11–37) (10–95) (11–141) (11–38)

6.1 1.8 1.6 0.9

(5–8) (1.1–5.2) (0.1–11.8) (0.3–43.6)

36 (14–133) 150 (14–607) 279 (17–840) 34 (17–42)

P

0.001 0.001 0.001 0.01 0.01 0.02 0.001

Follow-up of patients receiving autologous transplantation

Patients Median days to transfusion independence (range) RBC Platelets Late complications CMV infection Bacterial pneumonitis FUO Mean follow-up (months ⫾ s.d.) Complete remission duration (range) Relapses

group (median 22 days, range 0–80, P ⬍ 0.01). ABMT patients received their last platelet transfusion at a median time of 119 days (range 18–550) after the reinfusion, which was again significantly longer than for the PBSC group (median 34 days, range 6–110, P ⬍ 0.02). As regards late complications (Table 4), no significant differences have been observed between the two groups. The actuarial probabilities of DFS and relapse are shown in Figures 1 and 2. The curves did not differ when we considered the six patients who did not mobilize in the PBSC group (intention to treat analysis) or in the ABMT group. Until now, with a minimum follow-up for live patients of 24 months, no significant differences are observable as to DFS, or overall survival. Eleven out of 21 (52.4%) and 12/23 (52.1%) patients have relapsed in the ABMT and PBSC transplantation groups, respectively (see Figure 2). Discussion ABMT has been shown to be effective in AML, provided that an adequate induction/consolidation treatment has previously resulted in effective in vivo purging.1,5,9,21 Nevertheless delayed hematological recovery occurs in a substantial proportion of patients after ABMT, causing significant morbidity and mortality.6–12 To overcome these pitfalls, a therapeutic strategy based on a feasible, intensive induction/consolidation therapy coupled with a tolerable post-consolidation treatment is necessary to improve results. Thus, we and others have addressed the question of whether the use of PBSC might result in a more rapid

PBSC

ABMT

P

23

21

22 (0–80) 34 (6–10)

130 (18–196) 119 (18–550)

0.01 0.02

1 2 2 27 (⫾10.5) 22 (7–47) 52.1% (12/23)

1 2 2 42 (⫾18.5) 34.5 (6–65) 52.4% (11/21)

— — — NS

engraftment in AML patients.13–19,21 This is true after standard induction/consolidation therapies, whereas the feasibility of collecting PBSC after intensive consolidation has only recently been reported.19,27 However, the much larger number of cells infused in comparison to ABMT may result in the reinfusion of more residual malignant cells, especially after standard-dose consolidation chemotherapy. In this study we report the results of a program based on an intensive induction/consolidation followed by a comparison between ABMT vs PBSC transplantation, performed in two sequential cohorts of patients after an identical conditioning regimen. The objectives of the study were: to determine the overall feasibility, expressed in terms of the percentage of cases able to complete the program; to determine the efficacy of stem cell transplantation performed in I CR, with a comparison of PBSC transplantation and ABMT in terms of hematopoietic recovery and antileukemic activity. Sixty out of 88 evaluable patients were able to complete the program (68%), indicating that the majority of AML patients submitted to this intensive induction/consolidation regimen can be succesfully allo- or autotransplanted. The two cohorts of patients subsequently submitted to autotransplantation were comparable for major prognostic factors at diagnosis (age, karyotype, WBC number). Mobilization with G-CSF after consolidation therapy was effective, requiring a mean of two collections. Twenty-three out of 29 patients (79%) were mobilized; the non-mobilizers were successfully rescued by harvesting bone marrow. When the latter patients were taken into consideration in the ABMT group no significant differences in survival were observable between the two groups. The


present intensive-dose study confirmed the advantages of PBSC autotransplantation, previously described after classical (standard-dose) regimens, in terms of reduction of recovery times, early and late support and hospitalization. Most importantly, DFS among PBSC patients was not significantly different as compared to the ABMT group. It has to be underlined that follow-up is significantly longer for the ABMT group, due to the sequential design of the study. However, the minimum follow-up for live patients (24 months) is longer than the mean time to relapse in the PBSC series (14 months). Thus, our results do not reveal any increased likelihood of relapse using PBSC obtained after chemotherapy and G-CSF mobilization, although the antileukemic effect of both arms could possibly be further increased with more intensive in vivo purging strategies, as recently suggested,5 or by in vitro purification.22 In conclusion, our study indicates that PBSC transplantation is feasible and provides an adequate hemopoietic support after intensive chemotherapeutic regimens. The risk of leukemia recurrence does not appear to be greater than that observed with ABMT performed after the same intensive program. These results suggest that PBSC transplantation has real advantages which await confirmation from further randomized studies.

Acknowledgements

9 10

11 12

13

14

15

This work was supported in part by MURST.

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