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Related Collections Neoplasia Transplantation Brief Reports Social Bookmarking                   What's this?  Previous Article  |  Table of Contents  |  Next Article  Blood, 15 September 2001, Vol. 98, No. 6, pp. 1982-1985 BRIEF REPORT The significance of graft-versus-host disease and pretransplantation minimal residual disease status to outcome after allogeneic stem cell transplantation in patients with acute lymphoblastic leukemia Mehmet Uzunel, Jonas Mattsson, Marie Jaksch, Mats Remberger, and Olle Ringdén From the Department of Clinical Immunology and Center for Allogeneic Stem Cell Transplantation, Karolinska Institute at Huddinge University Hospital, Huddinge, Sweden.     Abstract Top Abstract Introduction Study design Results and discussion References Relapse is the major cause of treatment failure after allogeneic stem cell transplantation (SCT) in patients with acute lymphoblastic leukemia. Minimal residual disease (MRD) was analyzed before SCT in 30 patients with acute lymphoblastic leukemia. The aim was to determine whether the level of MRD before transplantation was correlated with outcome. Fifteen patients were found to have high-level MRD (102 to 103), 10 had low-level MRD (< 103), and 5 were MRD. Among MRD patients the probability of relapse was 0 in 5, which was less than in MRD+ patients (13 of 25) (P = .05). No major difference was found between the high- and low-level MRD+ groups. Among the MRD+ patients, only 2 of 11 with acute and chronic graft-versus-host disease had a relapse, versus 11 of 14 without (P = .005). In conclusion, for patients entering transplantation while they have residual disease, a combination of acute and chronic graft-versus-host disease may be needed to decrease the risk of relapse after SCT. (Blood. 2001;98:1982-1985) © 2001 by The American Society of Hematology.     Introduction Top Abstract Introduction Study design Results and discussion References Relapse still remains an obstacle to successful allogeneic stem cell transplantation (SCT) for patients with acute lymphoblastic leukemia (ALL).1,2 Graft-versus-host (GVH) disease, however, has been shown to protect against relapse.1,3 In particular, the combination of acute and chronic GVH disease seems to have the best antitumor effect.3-5 Analysis of antigen receptor (immunoglobulin [Ig] and T-cell receptor [TcR]) gene rearrangements to assess minimal residual disease (MRD) has started to become a part of routine laboratory work, and standardized protocols for monitoring MRD have therefore been developed.6 Using the polymerase chain reaction, MRD techniques are now sensitive enough to detect 1 leukemic cell among 104 to 106 normal cells.7,8 MRD studies after SCT have found a strong correlation between the presence of MRD and relapse.9,10 Increasing MRD levels have usually preceded a hematologic relapse. The importance of a pretransplant tumor burden to transplantation outcome has been shown by a better outcome in patients receiving transplants in complete remission (CR) than in those receiving transplants during relapse or with high MRD levels.11,12 However, many patients receiving transplants in remission still relapse after allogeneic SCT. The existence of residual disease not detected with morphologic analysis may therefore have an effect on outcome. In this study of 30 patients with ALL, we quantified the leukemic cell burden before SCT to determine whether the level of MRD was correlated with outcome.     Study design Top Abstract Introduction Study design Results and discussion References Patients Between May 1989 and February 1999, 91 ALL patients received transplants at the Center for Allogeneic Stem Cell Transplantation, Huddinge University Hospital. Of these 91, 80 received transplants in remission. Nine patients were excluded from the study because of transplant-related mortality before day 100 after SCT, and another 37 were excluded for whom no samples were available at diagnosis or before SCT. Antigen receptor rearrangement was not found in 4 of the remaining 34 patients. Table 1 summarizes patient and donor characteristics of the remaining 30 patients in relation to the MRD results.                                View this table: [in this window] [in a new window]   Table 1. Patient characteristics, MRD results, and relapse incidence Details regarding the transplantation procedure and supportive care have been published elsewhere.13,14 Remission and relapse Patients with regenerating peripheral blood values were considered in clinical remission when fewer than 5% blast cells among at least 200 nucleated cells were found in a bone marrow (BM) sample as defined by morphology. Clinical relapse was defined as when at least 30% blast cells were found in BM or when leukemic cells were detected extramedullary. DNA samples and MRD analysis All DNA material used in the MRD analysis was extracted from archival slides from BM aspirates. A salting-out procedure was performed as described by others.15 For MRD detection, the junctional regions of Ig and TcR gene rearrangements were amplified, cloned, and sequenced, and "patient-specific" primers were constructed for each patient. The methodology, polymerase chain reaction protocols, and primers for IgH, TcR, TcR, and Ig (Kde) gene rearrangements are described in detail elsewhere.6,16,17 Quantification was performed by parallel amplification of 1 µg pre-SCT DNA with a 10-fold serial dilution of leukemic cell DNA in mononuclear cell DNA from 5 healthy donors. The percentage of leukemic cells in the diagnosis sample was known from morphology and immunophenotype analysis done on the same day as the preparation of the slides. MRD levels were defined as high (102 to 103), low (104 to 105), or negative. Samples The diagnosis or relapse samples from which the patient-specific primers were generated were taken at a median of 4 (range, 2-13) months before SCT. The pre-SCT samples, analyzed for the presence of MRD, were taken at a median of 9 (range, 0-30) days before SCT. Statistical analysis The probability of relapse was calculated according to the Kaplan-Meier method. Differences in the incidence of GVH disease and relapse were compared with the Fisher exact test. The logistic regression model was used for multivariate analysis, which included risk factors such as sex, age, CR status, and acute and chronic GVH disease.     Results and discussion Top Abstract Introduction Study design Results and discussion References Antigen receptor rearrangements and primer sensitivity To avoid the problem of false negative results due to continuing rearrangements, usually observed in IgH rearrangements, several gene targets were used to identify clone-specific rearrangements.18,19 Twenty-seven patients were analyzed with primers reaching a sensitivity of 104 (n = 17) or 105 (n = 10). A target sensitivity of 103 was observed in 3 patients. All 3 patients, however, had an MRD level of more than 103 in the pre-SCT sample. Patients Thirteen patients died at a median of 10 (range, 2-22) months after SCT. Causes of death were BM relapse in 12 cases (median 8 [range, 2-22] months) and multiorgan failure in 1. Sixteen patients are alive and without relapse with a median follow-up of 39 (range, 13-119) months. One is alive with relapse. MRD results and outcome Fifteen patients had high-level MRD (102 to 103), 10 low-level MRD (104 to 105), and 5 were MRD. There were 8, 5, and 0 relapses in the 3 groups, respectively, with a higher incidence among patients who were MRD+ (13 of 25) than in those who were MRD (0 of 5) (P = .05). We found no significant difference in relapse rates between the high- and low-level MRD groups (Figure 1). This result does not accord with the findings of Knechtli et al,12 who had a relapse incidence of 100% in patients with high-level MRD, about 50% in those with low-level MRD, and about 20% in MRD patients. This may be because most of their patients received a T-cell-depleted graft, which is associated with an increased risk of relapse.5,20,21 View larger version (15K): [in this window] [in a new window]   Figure 1. Time to and cumulative incidence of relapse among different MRD groups. MRD high: 102 to 103; MRD low: less than 103; and MRD neg: MRD before transplantation. Tick marks indicate patients without leukemic relapse. The importance of an alloreaction was seen when GVH disease was analyzed in the 3 MRD-level groups (Table 1). Among patients with both acute and chronic GVH disease, only 2 of 15 patients relapsed, compared with 11 of 15 in patients without or only acute or chronic GVH disease (P = .003). In the 25 patients with detectable MRD before SCT, 2 of 11 patients with both acute and chronic GVH disease developed a hematologic relapse, as compared with 11 of 14 patients with no GVH disease or only acute or chronic GVH disease (P = .005). In multivariate analysis, the combination of acute and chronic GVH disease was significantly associated with lower risk of relapse (odds ratio 0.07; 95% confidence interval, 0.01-0.52; P = .014). The incidence of relapse was also higher in patients receiving transplants in second or later remission than in those receiving transplants in first remission (P = .077). MRD could not be included in the multivariate analysis because there was no relapse in the MRD group. Although the present study is retrospective and includes a small number of patients, it indicates that patients with persistent disease are more likely to relapse than those in molecular remission. Patients at higher risk of relapse should therefore be followed more frequently after SCT, and those with persistent or increasing MRD levels may be given additional antitumor therapy, such as withdrawal of immunosuppression and/or donor lymphocyte infusions.22 It may be desirable to induce acute as well as chronic GVH disease to achieve the best antileukemic effect, as shown by several other studies.4,5,23 Our data may support this in patients with ALL who are MRD+ at the time of transplantation. Knowledge of the MRD status before SCT therefore may permit us to individually design the posttransplantation immunosuppressive strategy to decrease the risk of a threatening relapse.24     Acknowledgments We are indebted to Inger Buskas and Anita Lindström for their help with the patient material. We thank the staff at the Center for Allogeneic Stem Cell Transplantation, Department of Hematology and Pediatrics, for compassionate and competent patient care.     Footnotes Submitted May 15, 2000; accepted May 11, 2001. Supported by grants from the Swedish Cancer Foundation (0070-B95-09XCC), the Children's Cancer Foundation (1995-035), the Swedish Medical Research Council (B96-16X-05971-16C), the FRF Foundation, the Tobias Foundation, and the Ellen Bachrach Foundation. The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734. Reprints: Mehmet Uzunel, Dept of Clinical Immunology, Huddinge University Hospital, SE- 141 86 Stockholm, Sweden; e-mail: mehmet.uzunel{at}impi.ki.se.     References Top Abstract Introduction Study design Results and discussion References 1. Weiden PL, Flournoy N, Thomas ED, et al. Antileukemic effect of graft-versus-host disease in human recipients of allogeneic-marrow grafts. N Engl J Med. 1979;300:1068-1073[Abstract]. 2. Weisdorf DJ, Nesbit ME, Ramsay NK, et al. Allogeneic bone marrow transplantation for acute lymphoblastic leukemia in remission: prolonged survival associated with acute graft-versus-host disease. J Clin Oncol. 1987;5:1348-1355[Abstract/Free Full Text]. 3. Ringden O, Hermans J, Labopin M, Apperley J, Gorin NC, Gratwohl A. The highest leukaemia-free survival after allogeneic bone marrow transplantation is seen in patients with grade I acute graft-versus-host disease. Acute and Chronic Leukaemia Working Parties of the European Group for Blood and Marrow Transplantation (EBMT). Leuk Lymphoma. 1996;24:71-79[Medline] [Order article via Infotrieve]. 4. Sullivan KM, Weiden PL, Storb R, et al. Influence of acute and chronic graft-versus-host disease on relapse and survival after bone marrow transplantation from HLA-identical siblings as treatment of acute and chronic leukemia [published erratum appears in Blood. 1989;74:1180]. Blood. 1989;73:1720-1728[Abstract/Free Full Text]. 5. Horowitz MM, Gale RP, Sondel PM, et al. Graft-versus-leukemia reactions after bone marrow transplantation. Blood. 1990;75:555-562[Abstract/Free Full Text]. 6. Pongers-Willemse MJ, Seriu T, Stolz F, et al. Primers and protocols for standardized detection of minimal residual disease in acute lymphoblastic leukemia using immunoglobulin and T cell receptor gene rearrangements and TAL1 deletions as PCR targets: report of the Biomed-1 Concerted Action: investigation of minimal residual disease in acute leukemia. Leukemia. 1999;13:110-118[CrossRef][Medline] [Order article via Infotrieve]. 7. Campana D, Pui CH. Detection of minimal residual disease in acute leukemia: methodologic advances and clinical significance. Blood. 1995;85:1416-1434[Free Full Text]. 8. Foroni L, Harrison CJ, Hoffbrand AV, Potter MN. Investigation of minimal residual disease in childhood and adult acute lymphoblastic leukaemia by molecular analysis. Br J Haematol. 1999;105:7-24[Medline] [Order article via Infotrieve]. 9. Knechtli CJ, Goulden NJ, Hancock JP, et al. Minimal residual disease status as a predictor of relapse after allogeneic bone marrow transplantation for children with acute lymphoblastic leukaemia. Br J Haematol. 1998;102:860-871[CrossRef][Medline] [Order article via Infotrieve]. 10. Radich J, Ladne P, Gooley T. Polymerase chain reaction-based detection of minimal residual disease in acute lymphoblastic leukemia predicts relapse after allogeneic BMT. Biol Blood Marrow Transplant. 1995;1:24-31[Medline] [Order article via Infotrieve]. 11. Barrett AJ, Horowitz MM, Gale RP, et al. Marrow transplantation for acute lymphoblastic leukemia: factors affecting relapse and survival. Blood. 1989;74:862-871[Abstract/Free Full Text]. 12. Knechtli CJ, Goulden NJ, Hancock JP, et al. Minimal residual disease status before allogeneic bone marrow transplantation is an important determinant of successful outcome for children and adolescents with acute lymphoblastic leukemia. Blood. 1998;92:4072-4079[Abstract/Free Full Text]. 13. Ringden O, Pihlstedt P, Markling L, et al. Prevention of graft-versus-host disease with T cell depletion or cyclosporin and methotrexate: a randomized trial in adult leukemic marrow recipients. Bone Marrow Transplant. 1991;7:221-226[Medline] [Order article via Infotrieve]. 14. Ringden O, Remberger M, Persson U, et al. Similar incidence of graft-versus-host disease using HLA-A, -B and -DR identical unrelated bone marrow donors as with HLA-identical siblings. Bone Marrow Transplant. 1995;15:619-625[Medline] [Order article via Infotrieve]. 15. Vince A, Poljak M, Seme K. DNA extraction from archival Giemsa-stained bone-marrow slides: comparison of six rapid methods. Br J Haematol. 1998;101:349-351[CrossRef][Medline] [Order article via Infotrieve]. 16. Zetterquist H, Mattsson J, Uzunel M, et al. Mixed chimerism in the B-cell lineage is a rapid and sensitive indicator of minimal residual disease in bone marrow transplant recipients with pre-B-cell acute lymphoblastic leukemia. Bone Marrow Transplant. 2000;25:843-851[CrossRef][Medline] [Order article via Infotrieve]. 17. Mattsson J, Uzunel M, Remberger M, et al. Minimal residual disease is common after allogeneic stem cell transplantation in patients with B cell chronic lymphocytic leukemia and may be controlled by graft-versus-host disease. Leukemia. 2000;14:247-254[CrossRef][Medline] [Order article via Infotrieve]. 18. Beishuizen A, Hahlen K, Hagemeijer A, et al. Multiple rearranged immunoglobulin genes in childhood acute lymphoblastic leukemia of precursor B-cell origin. Leukemia. 1991;5:657-667[Medline] [Order article via Infotrieve]. 19. Wasserman R, Yamada M, Ito Y, et al. VH gene rearrangement events can modify the immunoglobulin heavy chain during progression of B-lineage acute lymphoblastic leukemia. Blood. 1992;79:223-228[Abstract/Free Full Text]. 20. Marmont AM, Horowitz MM, Gale RP, et al. T-cell depletion of HLA-identical transplants in leukemia. Blood. 1991;78:2120-2130[Abstract/Free Full Text]. 21. Goldman JM, Gale RP, Horowitz MM, et al. Bone marrow transplantation for chronic myelogenous leukemia in chronic phase: increased risk for relapse associated with T-cell depletion. Ann Intern Med. 1988;108:806-814. 22. Kolb HJ, Mittermuller J, Clemm C, et al. Donor leukocyte transfusions for treatment of recurrent chronic myelogenous leukemia in marrow transplant patients. Blood. 1990;76:2462-2465[Abstract/Free Full Text]. 23. Ringden O, Labopin M, Gluckman E, et al. Graft-versus-leukemia effect in allogeneic marrow transplant recipients with acute leukemia is maintained using cyclosporin A combined with methotrexate as prophylaxis. Acute Leukemia Working Party of the European Group for Blood and Marrow Transplantation. Bone Marrow Transplant. 1996;18:921-929[Medline] [Order article via Infotrieve]. 24. Carlens S, Aschan J, Remberger M, Dilber M, Ringden O. Low-dose cyclosporine of short duration increases the risk of mild and moderate GVHD and reduces the risk of relapse in HLA-identical sibling marrow transplant recipients with leukaemia. Bone Marrow Transplant. 1999;24:629-635[CrossRef][Medline] [Order article via Infotrieve]. © 2001 by The American Society of Hematology.   CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this? This article has been cited by other articles: P. Bader, H. Kreyenberg, G. H.R. Henze, C. Eckert, M. Reising, A. Willasch, A. Barth, A. Borkhardt, C. Peters, R. Handgretinger, et al. 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