Pre-transplant Minimal Residual Disease Research Articles

Clofarabine in Pediatric Patients with Refractory or Relapsed Leukemia: the GATLA experience.

AbstractAbstract 2605▪▪This icon denotes a clinically relevant abstract Introduction:The outcome in pediatric patients with relapse-refractory acute leukemia (acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML)) is very poor. Clofarabine is an active agent in this high risk group. Purpose:Report the GATLA experience in patients with relapse-refractory acute leukemia treated with clofarabine based regimens. Methods:Between June 07 and April 12, 66 patients from 15 GATLA centers were enrolled. 61 ALL (52 B/9 T): 23 first relapse (20 BM, 1 CNS, 2 combined), 33 second relapse (28 BM, 1 CNS, 4 combined), 5 with refractory disease. 5 AML (4 first relapse, 1 refractory disease). Median age 10,5 years (1–20 ys). Gender: 44 boys and 22 girls.60 patients received chemotherapy combined clofarabine-etoposide-cyclophosphamide (CLO 218), 4 patients received clofarabine plus ara-c therapy and 2 patients were treated with clofarabine alone (52mg/m2).63.7% (42/66) of the patients were treated with 2 therapeutic lines prior to clofarabine regimens, 22.7% (15/66) received only 1 prior treatment, 13.6% (9/66) got 3 or more therapies and 5 patients undergone prior HSCT (Hematopoietic Stem Cell Transplantation). Results:Complete remission (CR) was achieved in: 36/66 patients (54,5 %). Early toxic death rate was 7/66 (10,6%). No response occurred in 22/66 (33,3%) patients. 1 patient is on induction, not evaluable at the moment.Of the 36 responders, 19 patients (52,7 %) proceeded to HSCT: 6 (31,5 %) remain alive post-transplantation with a median follow up of 55 weeks ( range 6–156 w), HSCT related death: 7 (37 %), relapse after HSCT: 6 (31,5%) at a mean of 20 weeks( range 8–56 weeks).Among the 61 ALL patients, 18/23 (78%) in first relapse achieved CR, 12/18 (66,6%) needed only one course to achieved CR, of them, 3/12 (25%) relapsed at 8, 8 and 14 weeks and died for progressive disease. 1/12 (8,3%) died in CR at week 8. 2/12 (16,6%) patients are on treatment. 6/12 (50%) proceeded to HSCT: 1/6 relapsed 56 weeks after. 3/6 patients died in CR after HSCT at 12, 15 and 24 weeks. 2/6 remain in CR at 6 weeks follow up. 6/18 (33,3%) of the patients in first relapse needed 2 courses to achieved CR. 1/6 (16,7%) relapsed at week 8. 2/6(33,3%) died in CR at week 9 and 16. 3/6 ( 50%) proceeded to HSCT, 1 is alive at 36 weeks, 1 relapsed at week 26 and 1 died in CR at week 14.13/33 (39%) ALL patients in second relapse achieved CR and 8/13 (61,5%) needed only one course to CR. 3/8 (37,5%) relapsed at 6,8 and 16 weeks and died for progressive disease. 5/8(62,5%) proceeded to HSCT, of them 3 (60%) relapsed after HSCT at 8,8 and 12 weeks. 1/5 (20%) is alive and disease free at 6 weeks after HSCT. 1/5 (20%) died in CR. 5/13 (38%) needed 2 courses to achieved CR, 1/5 relapsed after 4 weeks. 1/5 died in CR at week 6, 1/5 is alive 40 weeks after CR. 2/5 proceeded to HSCT and died in CR at 10 and 22 weeks after HSCT.2/5 ALL patients with refractory disease achieved CR with 2 and 3 courses of clofarabine, one proceeded to HSCT and is alive with a follow up of 156 weeks, the other relapsed at 8 weeks. 3/5 did not response and died.AML patients: 3/5 received clofarabine as 3rd therapeutic line: 1 patient achieved CR after one course but died of toxicity, 1 patient did not response and 1 died during induction. 2/5 received clofarabine as 2ndline ( 1 with refractory disease and 1 who undergone prior HSCT), they achieved CR with 2 courses of chemotherapy and proceeded to HSCT, one relapsed after 16 weeks and the other is alive and disease free after 120 weeks. Conclusion:In our series, Clofarabine was well tolerated and shows significant antileukemic activity in heavily pretreated children. The response rate and the durability of remission observed is better when clofarabine regimens are used earlier (first relapse). Responses allowed several refractory patients to proceed to HSCT. Levels of pre transplant MRD (Minimal Residual Disease)should be taken into account in order to follow this procedure. Disclosures:No relevant conflicts of interest to declare.

Impact of pretransplant minimal residual disease after cord blood transplantation for childhood acute lymphoblastic leukemia in remission: an Eurocord, PDWP–EBMT analysis

To address the prognostic value of minimal residual disease (MRD) before unrelated cord blood transplantation (UCBT) in children with acute lymphoblastic leukemia (ALL), we analyzed 170 ALL children transplanted in complete remission (CR) after myeloablative conditioning regimen. In all, 72 (43%) were in first CR (CR1), 77 (45%) in second CR (CR2) and 21 (12%) in third CR (CR3). The median interval from MRD quantification to UCBT was 18 days. All patients received single-unit UCBT. Median follow-up was 4 years. Cumulative incidence (CI) of day-60 neutrophil engraftment was 85%. CI of 4 years relapse was 30%, incidence being lower in patients with negative MRD before UCBT (hazard ratio (HR)=0.4, P=0.01) and for those transplanted in CR1 and CR2 (HR=0.3, P=0.002). Probability of 4 years leukemia-free survival (LFS) was 44%, (56, 44 and 14% for patients transplanted in CR1, CR2 and CR3, respectively (P=0.0001)). Patients with negative MRD before UCBT had better LFS after UCBT compared with those with positive MRD (54% vs 29%; HR=2, P=0.003). MRD assessment before UCBT for children with ALL in remission allows identifying patients at higher risk of relapse after transplantation. Approaches that may decrease relapse incidence in children given UCBT with positive MRD should be investigated to improve final outcomes.

Unrelated Cord Blood Transplantation in Adult and Pediatric Acute Lymphoblastic Leukemia: Effect of Minimal Residual Disease on Relapse and Survival

Data on pretransplantation minimal residual disease (MRD) and outcomes of umbilical cord blood transplantation (UCBT) are limited. Out of the 143 patients with acute lymphoblastic leukemia (ALL) who underwent UCBT at the University of Minnesota between 2004 and 2010, we evaluated 86 patients with available MRD assessment data by 4- and 8-color flow cytometry analysis immediately before transplantation. Ten patients (11.6%) were MRD-positive, and 76 were MRD-negative (88.4%). Most of the patients (82%) received myeloablative conditioning. GVHD prophylaxis consisted of cyclosporine and mycophenolate mofetil. In multivariate analysis, age, disease status (complete remission [CR] 1 versus CR2/CR3), disease group (precursor B cell ALL versus Philadelphia chromosome-positive ALL versus T cell ALL), and time to transplantation had no impact on relapse. Patients with MRD before UCBT had a greater incidence of relapse at 2 years (relapse rate, 30%; 95% confidence interval [CI], 4%-56%) and lower 3-year disease-free survival (30%; 95% CI, 7%-58%) compared with those without MRD (relapse rate, 16%; 95% CI, 8%-25%; P = .05; disease-free survival, 55%; 95% CI, 43%-66%; P = .02). Our data suggest that in patients with ALL, achieving an MRD-negative state before UCBT improves outcomes.

Prognostic Significance of Detection of Minimal Residual Disease by Multiparametric Flow Cytometry Before Allogeneic Stem Cell Transplantation in Patients with Acute Myeloid Leukemia,

Abstract 4116The main prognostic determinant of the outcome of allogeneic stem cell transplantation (allo-SCT) for acute myeloid leukemia (AML) is disease phase at transplant while the relevance of the assessment of minimal residual disease (MRD) at transplant has not been elucidated. Although molecular markers are highly sensitive for the detection of MRD, they are available in only half of AML cases to date. On the other hand, multiparametric flow cytometry (MFC) offers the feasibility of MRD assessment in virtually all patients. We retrospectively studied the influence of pre-transplant MRD status by MFC on the outcomes of patients with AML in relation to other known prognostic factors.From 01/2003 to 12/2010, 102 AML patients (male/female; 58/44), aged 15–64 (median; 42) years, received allo-SCT in our center at first (CR1, N=69) or subsequent (CR≥2, N=33) morphologic complete remission. Donors were HLA-compatible siblings (N=45), matched unrelated (N=34) or alternative (haploidentical relatives or unrelated cord blood; N=6 and 17, respectively). The conditioning regimen was myeloablative in 85 (83.33%), and reduced-intensity in 17 (16.67%) cases. Data on MRD assessment by four- or five-color MFC before transplant were available in 92 patients. MRD was defined as any percentage of cells with an aberrant antigen expression pattern compared to normal or regenerating marrow. Transplant outcomes associated with MRD in univariate and multivariate analysis were overall (OS) and disease-free (DFS) survival, relapse incidence, and non-relapse-related mortality (NRM). Prognostic parameters included in the statistical model were the following: age and gender of recipient or donor, interval from diagnosis to transplant, de novo versus secondary AML, CR1 versus CR≥2 at transplant, cytogenetic risk group (according to Intergroup ECOG/SWOG definition), recipient and donor cytomegalovirus (CMV) serostatus, donor type, conditioning regimen (myeloablative versus reduced-intensity), degree of HLA match, method of graft-versus-host disease (GVHD) prophylaxis, and the modified European Blood and Marrow Transplantation Group (EBMT) risk score (Hemmati et al, 2011).With a median follow-up of 47 (range, 6–98) months, OS and DFS were 54.2% and 51.72%, respectively. The cumulative incidence of relapse was 24.45%, and of NRM 23.82%. Eighteen out of 92 (19.57%) patients had MRD by MFC before allo-SCT. According to univariate analysis, the presence of MRD showed a negative association with OS (29.63% versus 63.17% at 4 years in patients with or without MRD, respectively, P=0.053) and DFS (28.52% in MRD-positive versus 59.74% in MRD-negative cases at 4 years, P=0.025, Figure 1). Presence of MRD correlated with higher incidence of relapse compared to MRD negativity (52.7% versus 15.8%, respectively, P=0.001) (Figure 2), but did not influence NRM. In multivariate analysis (Cox proportional hazards, backward stepwise selection), the parameters that remained independent adverse risk factors in terms of specific outcomes were: a) For OS: age and male gender of patient, alternative donor, unfavorable karyotype, CR≥2, and secondary AML, b) For DFS: presence of MRD (HR=3.2, P=0.009), CR≥2 (ÇR=3.1, P=0.005), reduced-intensity conditioning (HR=2.6, P=0.034), and unfavorable karyotype (HR=2.06, P=0.074), c) For relapse: presence of MRD (ÇR=3.83, P=0.003), and unfavorable karyotype (HR=3.55, P=0.007), and d) For NRM: age of recipient, CR≥2, and alternative donor. [Display omitted] [Display omitted] In conclusion, the presence of MRD by MFC before allo-SCT for AML is associated with a significantly higher risk of relapse and an inferior DFS, despite morphologic complete remission at transplant. Moreover, pre-transplantation MRD status has independent prognostic significance in addition to the disease phase, cytogenetic risk group, and intensity of the conditioning regimen. Prospective studies are warranted to examine if the adverse impact of MRD on the outcomes of allo-SCT can be reversed by immunologic manipulations aiming at augmenting graft-versus-leukemia (GVL) effect. Disclosures:No relevant conflicts of interest to declare.

Pre-Transplant Minimal Residual Disease Detected by Multiparameter Flow Cytometric Analysis Predicts for Disease Relapse in Adult Patients with Acute Lymphoblastic Leukemia Post Allogeneic Hematopoietic Stem Cell transplantation

Abstract 3072Allogeneic hematopoietic stem cell transplantation (SCT) is the only effective treatment for high-risk or relapsed acute lymphoblastic leukemia (ALL). However, SCT carries high treatment-related mortality (TRM). There have been continuing efforts to refine criteria for patients who may benefit most from SCT. Minimal residual disease (MRD) at the end of induction or induction/consolidation therapy has emerged as the most significant risk factor for disease relapse in ALL patients, and has become an indicator for SCT or intensified chemotherapy. In addition, MRD prior to SCT is highly predictive for post-transplant relapse in the pediatric population. The significance of pre-SCT MRD in adult patients with ALL remains to be determined. Patients and methods: In this retrospective analysis, we examined the impact of MRD detected within one month prior to SCT on risk of disease progression in adult patients with ALL who were in complete remission, defined by less than 5% blasts in bone marrow by morphologic assessment and normal blood counts at time of SCT. MRD has been uniformly assessed in patients using multiparameter flow cytometry since 2004. Bone marrow aspirates were stained using a comprehensive panel of >=15 markers and 200, 000 cells were analyzed. MRD was scored as positive based on a distinct cluster of >=20 cells, showing a significant difference in the level of expression (>=3-fold) of >=2 antigens, in comparison to the known phenotype of benign immature B-cell precursors. Disease progression (relapse) was defined as leukemic blast count >= 5%. Therefore, limiting our data set to transplants performed during February 2004 through May 2010 at MD Anderson Cancer Center, we identified 98 patients with median age 34 years (range 19–68) treated with a median of 2 lines of chemotherapy (range 1–6) who received a matched sibling (n=51), matched unrelated (n=36), haplo-identical family (n=3), or cord blood donor (n=8) transplant for ALL of B-(n=86) or T-lineage (n=12) in CR1 (n=52), CR2 (n=39) or CR3 (n=7) using a myeloablative non-TBI based (n=62) or TBI-based (n=36) transplant conditioning regimen. GVHD prophylaxis was tacrolimus-based for all patients. Among 88 patients with available cytogenetic data at diagnosis, 59% were considered high risk defined by the presence of the t(9;22), t(4;11), hypodiploid or complex cytogenetics, 2% were classified as good risk by the presence of hyperdiploidy, and the remaining 34% were intermediate risk. Results: Ninety-two patients were evaluable for response and all had sustained CR following transplant; 6 had early death. With a median follow-up of 33 months among survivors (range 5.8–80.7), patients transplanted without detectable MRD at time of SCT had a significantly lower cumulative incidence of relapse than those transplanted with MRD, 30% versus 70%, p=.004 (Figure 1). There was also a trend for better 3-year PFS in the MRD negative group, 38% versus 13%, p=0.1. Additional factors, including disease stage at time of SCT, cytogenetic profile at diagnosis, donor relation, and development of GVHD were evaluated in a univariate analysis for relapse rate, PFS, and overall survival. Only the presence of MRD remained a significant predictor for relapse. Conclusion: This study demonstrates that the presence of MRD prior to SCT in adult ALL patients is a significant predictor for relapse. Improvements in our current therapies are urgently needed to improve the outcomes for MRD positive patients. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

The Level of Minimal Residual Disease in the Bone Marrow of Patients with Multiple Myeloma before High-Dose Therapy and Autologous Blood Stem Cell Transplantation Is an Independent Predictive Parameter

The prognostic relevance of minimal residual disease (MRD) in patients with multiple myeloma is still an open question. We measured MRD levels in bone marrow (BM) samples of 53 patients treated with high-dose therapy (HDT) and autologous peripheral blood stem cell transplantation using real-time quantitative (RQ)-IgH-PCR with allel-specific oligonucleotide probes. We identified a prognostically relevant threshold level of 0.2% 2IgH/β-actin ratio in the BM before HDT. Twenty-six patients with MRD levels below this value were termed as the "low-MRD group," whereas 27 patients with levels above this threshold were allocated to the "high-MRD group." Median event-free-survival (EFS) in the low-MRD group was significantly (P = .001) longer than in the high-MRD group with 35 versus 20 months, respectively. Overall survival (OS) within the low-MRD group was also significantly longer with 70 versus 45 months (P = .04). Using multivariate analysis, we found that the pretransplantation MRD level was an independent prognostic factor for EFS (P = .003) and OS (P = .05). Further, EFS of patients in the high-MRD could be improved (P = .005) when they achieved a low MRD level after HDT. In conclusion, measuring MRD is of prognostic relevance in patients with MM, and low MRD levels should be a goal of treatment.

Impact of Pretransplantation Minimal Residual Disease, As Detected by Multiparametric Flow Cytometry, on Outcome of Myeloablative Hematopoietic Cell Transplantation for Acute Myeloid Leukemia

Allogeneic hematopoietic cell transplantation (HCT) benefits many patients with acute myeloid leukemia (AML) in first remission. Hitherto, little attention has been given to the prognostic impact of pretransplantation minimal residual disease (MRD). We retrospectively studied 99 consecutive patients receiving myeloablative HCT for AML in first morphologic remission. Ten-color multiparametric flow cytometry (MFC) was performed on bone marrow aspirates before HCT. MRD was identified as a cell population showing deviation from normal antigen expression patterns compared with normal or regenerating marrow. Any level of residual disease was considered MRD positive. Before HCT, 88 patients met morphologic criteria for complete remission (CR), whereas 11 had CR with incomplete blood count recovery (CRi). Twenty-four had MRD before HCT as determined by MFC. Two-year estimates of overall survival were 30.2% (range, 13.1% to 49.3%) and 76.6% (range, 64.4% to 85.1%) for MRD-positive and MRD-negative patients; 2-year estimates of relapse were 64.9% (range, 42.0% to 80.6%) and 17.6% (range, 9.5% to 27.9%). After adjustment for all or a subset of cytogenetic risk, secondary disease, incomplete blood count recovery, and abnormal karyotype pre-HCT, MRD-positive HCT was associated with increased overall mortality (hazard ratio [HR], 4.05; 95% CI, 1.90 to 8.62; P < .001) and relapse (HR, 8.49; 95% CI, 3.67 to 19.65; P < .001) relative to MRD-negative HCT. These data suggest that pre-HCT MRD is associated with increased risk of relapse and death after myeloablative HCT for AML in first morphologic CR, even after controlling for other risk factors.

Impact of Pretransplantation Minimal Residual Disease, As Detected by Multiparametric Flow Cytometry, on Outcome of Myeloablative Hematopoietic Cell Transplantation for Acute Myeloid Leukemia

Impact of Pretransplantation Minimal Residual Disease, As Detected by Multiparametric Flow Cytometry, on Outcome of Myeloablative Hematopoietic Cell Transplantation for Acute Myeloid Leukemia

Minimal Residual Disease following Allogeneic Hematopoietic Stem Cell Transplantation

Minimal residual disease (MRD), both before and after transplantation, is a clinically important yet relatively poorly defined aspect of allogeneic hematopoietic stem cell transplantation (alloHSCT). The clinical relevance of MRD in the context of alloHSCT has been demonstrated by its association with the development of clinical relapse. However, with the possible exception of chronic myeloid leukemia (CML), the specific techniques, timing, frequency, and clinical utility, relative to improvement in patient outcomes, for monitoring MRD in the setting of alloHSCT has yet to be clearly defined. A concise overview of monitoring techniques for detecting MRD, as well as treatment strategies and biological and clinical research initiatives for MRD suggested by the National Cancer Institute First International Workshop on the Biology, Prevention, and Treatment of Relapse after Allogeneic Hematopoietic Stem Cell Transplantation, is covered in this article.

Relationship between minimal residual disease measured by multiparametric flow cytometry prior to allogeneic hematopoietic stem cell transplantation and outcome in children with acute lymphoblastic leukemia

The presence of minimal residual disease detected by polymerase chain reaction techniques prior to allogeneic hematopoietic stem cell transplantation has proven to be an independent prognostic factor for poor outcome in children with acute lymphoblastic leukemia. The aim of this study was to ascertain whether the presence of minimal residual disease detected by multiparametric flow cytometry prior to allogeneic hematopoietic stem cell transplantation is related to outcome in children acute lymphoblastic leukemia. Minimal residual disease was quantified by multiparametric flow cytometry at a median of 10 days prior to hematopoietic stem cell transplantation in 31 children (age range, 10 months to 16 years) with acute lymphoblastic leukemia. Thirteen patients were transplanted in first remission. Stem cell donors were HLA-identical siblings in 8 cases and matched unrelated donors in 23. Twenty-six children received a total body irradiation-containing conditioning regimen. According to the level of minimal residual disease, patients were divided into two groups: minimal residual disease-positive (>or=0.01%) (n=10) and minimal residual disease-negative (<0.01%) (n=21). Estimated event-free survival rates at 2 years for the minimal residual disease-negative and -positive subgroups were 74% and 20%, respectively (P=0.004) and overall survival rates were 80% and 20%, respectively (P=0.005). Bivariate analysis identified pre-transplant minimal residual disease as the only significant factor for relapse and also for death (P<0.01). The presence of minimal residual disease measured by multiparametric flow cytometry identified a group of patients with a 9.5-fold higher risk of relapse and a 3.2-fold higher risk of death than those without minimal residual disease. This study supports the strong relationship between pre-transplantation minimal residual disease measured by multiparametric flow cytometry and outcome following allogeneic hematopoietic stem cell transplantation and concur with the results of previous studies using polymerase chain reaction techniques.

Sub-Morphologic Evidence of Disease Prior to Stem Cell Transplantation Correlates with Inferior Post Transplant Outcome in Childhood Acute Myeloid Leukemia.

Abstract 328▪▪This icon denotes an abstract that is clinically relevant.Hematopoietic cell transplantation (HCT) is utilized in the treatment of a subset of children with acute myeloid leukemia (AML). Although presence of minimal residual disease (MRD) has been associated with a high relapse rate in children with AML, the clinical implications of MRD prior to HCT have not previously been defined. We explored the clinical outcomes associated with the presence of morphologic or sub-morphologic disease prior to HCT. In the period of 1995-2007, 89 patients age 1-18 years received myeloablative conditioning and allogeneic HCT from related (N=36) or unrelated (N=53) donors at Fred Hutchinson Cancer Research Center. At the time of HCT, 63 patients received transplants in complete remission (CR) and 26 had morphologic disease. Among those in morphologic CR, 43 were in 1st CR, 17 in 2nd CR and 3 were in 3rd CR. All patients had bone marrow evaluation for MRD by multi-dimensional flow cytometry (MDF) 2-4 weeks prior to HCT. MDF testing showed that 10 of 63 patients (16%) who received transplants in morphologic CR had MRD levels ranging from 0.3% to 12% (median 3.7%) while the remaining 53 patients had no detectable MRD. The median follow up for the 38 survivors is 6 years (range 1 – 12). Actuarial overall survival at 3 years from HCT for patients who received HCT with and without morphologic disease was 12% vs. 59%, (p<0.001) with corresponding relapse rates of 62% and 24%, (p<0.001), respectively. We subsequently evaluated clinical outcomes for patients who received HCT in morphologic CR with and without evidence of MRD by MDF. Actuarial overall survival for patients who received transplants in morphologic CR with MRD was 15% vs. 67% for those without MRD, p=0.001. The corresponding relapse rate for those with MRD was 65% vs. 17% for those without MRD (p<0.001). Among the 10 CR patients with MRD, 4 had high risk cytogenetics and 6 patients had standard risk cytogenetics. In 53 MRD-negative patients who received HCT in CR, 10 (19%) had high risk, 7 had favorable risk cytogenetics and the remaining 37 (69%) had standard risk cytogenetics. It is notable that the relapse and overall survival probabilities for patients who were received HCT in morphologic CR but with MRD appear similar to those for subjects in morphologic relapse. This data demonstrates that presence of sub-morphologic disease prior to HCT portends high relapse risk and worse post transplant survival. Larger studies are evaluating the full impact of pre-transplant MRD as well and the clinically significant threshold of MRD prior to stem cell transplantation in AML. Disclosures:No relevant conflicts of interest to declare.

Impact of Pre-Transplant Minimal Residual Disease Assessed by Flow Cytometry on Outcome Following Myeloablative Hematopoietic Cell Transplantation for Patients with AML-CR1.

Allogeneic hematopoietic cell transplantation (HCT) often offers the best and sometimes only chance for cure for acute myeloid leukemia (AML) patients. However, the decision of whether to transplant a patient with AML while in first remission remains a difficult one, and a great deal of attention has been given to attempting to identify factors that will predict HCT outcome. While many patient- and disease-specific factors have been studied, little attention has been given to the impact of the presence of MRD as determined by multiparametric flow cytometry (MFC) at the time of presentation for transplant on outcome. We have therefore evaluated outcome among patients transplanted for AML in CR1, comparing those with evidence of MRD as determined by MFC compared to those with no evidence of MRD prior to myeloablative HCT. Between 1992 and 2004, 140 patients greater than 18 years of age with AML-CR1 received a myeloablative HCT at our Center. Immediately prior to HCT, 23 CR1 patients had evidence of MRD (MRD+) whereas 117 CR1 patients had no evidence of MRD (MRD-) by flow cytometric analysis. Four (17%) MRD+ patients and 23 (20%) MRD- patients had an unrelated donor. Median age was 48.1 and 40.7 years for MRD+ and MRD- patients, respectively. Conditioning included use of total body irradiation (n=8 MRD+; n=37 MRD-), busulfan and cyclophosphamide (n=8 MRD+; n=42 MRD-), an anti-CD45 radiolabeled antibody (n=6 MRD+; n=34 MRD-), or other regimens (n=1 MRD+; n=4 MRD-). Most received cyclosporine and methotrexate for GvHD prophylaxis (96% MRD+; 97% MRD-). Source of stem cells was bone marrow in 39% MRD+ and 58% MRD- patients. Patients were further classified by cytogenetic risk as favorable (0% MRD+; 3% MRD-), intermediate (61% MRD+; 74% MRD-), or unfavorable (30% MRD+; 18% MRD-) (cytogenetics were missing for 2 MRD+ patients and 5 MRD- patients). At last follow-up, 16 (70%) MRD+ patients had died compared to 46 (39%) MRD- patients. Five-year estimates of overall survival were 35% and 64%, respectively. After adjusting for age, source of stem cells, donor, and unfavorable cytogenetics, the hazard ratio (HR) for mortality was 2.13 (95% CI 1.11–4.08, p=.02). Nine (39%) MRD+ patients relapsed after HCT compared to 25 (21%) MRD- patients and the 5-year estimates of relapse were 39% and 21%, respectively. The adjusted HR or relapse was 2.83 (1.19–6.73, p=.02). Thirty percent (n=7) of patients in the MRD+ group experienced transplant-related mortality (TRM) compared to 21% (n=25) in the MRD- group [adjusted HR=1.50 (0.54–4.19, p=.44)]. Among those who were MRD+, there was a positive correlation between the percentage of flow blasts and the risk of mortality and TRM. However, there was no suggestion of an association with relapse in this group. When modeling flow blasts as a continuous linear variable, each increase in flow blasts of 10% was associated with an increase in the hazard of mortality [HR=1.20 (1.00–1.45, p=.05)] and TRM [HR=1.39 (1.10–1.76, p=.006)], but not relapse [HR=0.77 (0.35–1.70, p=.51)]. While these data need to be confirmed in a larger number of patients, they suggest that any evidence of MRD+ at HCT by flow cytometry in AML-CR1 patients increases the risk of relapse and death after HCT relative to MRD- patients.

Molecular Level of Minimal Residual Disease in Bone Marrow Before High-Dose Therapy and Autologous Stem Cell Transplantation Is a Prognostic Parameter in Patients with Multiple Myeloma.

Background: Using real-time quantitative (RQ) PCR we recently (Haematologica 89, 2004) identified a prognostic cut-off level of residual clonotypic cells in the bone marrow of patients with multiple myeloma before high-dose therapy (HDT) and autologous peripheral blood stem cell transplantation (PBSCT). In this study we validate this report with a larger number of patients.Patients and Methods: Bone marrow samples of 68 patients with stage II/III multiple myeloma and heavy chain disease were obtained at the time of diagnosis and after induction therapy and stem cell collection but before single HDT and autologous PBSCT. Sequencing of the patient specific immunoglobulin heavy chain (IgH) locus was successful in 51 patients (75%). For 49 patients (72%) RQ-PCR using allele-specific oligonucleotide (ASO) Taqman probes together with LightCycler technology could be established with a sensitivity of 10−4 to 10−6 and linear amplification conditions. The proportion of clonotypic cells was assessed as IgH / 2 beta-actin ratio in percent. Patients were divided in two prognostic groups by a threshold level of 0.03% clonotypic cells.Results: The median level of residual tumor cells in bone marrow of all patients at the time before transplantation was 0.05% (range: 0–21%). Time to progression (TTP) from the time of diagnosis of patients in the ¨good¨ prognostic group (n = 21) was 51 months and significantly (p = 0.002) longer in comparison to 20 months of patients with a pre-transplantation minimal residual disease level of more than 0.03% in BM (n = 28). Overall survival (OS) of patients within the ¨good¨ prognostic group was also significantly prolonged (median OS: not reached versus 46 months, p = 0.03). Univariate analysis also revealed kind of maintenance / consolidation therapy (thalidomide, interferon, reduced intensity conditioning (RIC) allogeneic transplant) and cytogenetic banding analysis as prognostic markers for TTP. For OS kind of maintenance therapy, cytogenetic abnormalities, ISS stage, CRP and LDH levels were of prognostic relevance. In multivariat analysis grouping by pre-transplantation MRD level was an independent prognostic factor for either TTP and OS.Conclusion: Quantitative molecular assessment of pre-transplantation tumor level in the bone marrow is an independent prognostic parameter for TTP and OS of patients with multiple myeloma. This finding has two controversial implications. One conclusion could be, that induction therapy should be continued and intensified e.g. with novel agents until a low MRD level is achieved. An alternative conclusion is, that a low tumor burden after induction therapy may be a surrogate parameter for chemosensitive disease, which makes patients more susceptible for high-dose chemotherapy. Therefore, further MRD studies are needed to answer this important question.

Erratum: Pretransplant minimal residual disease level predicts clinical outcome in patients with acute myeloid leukemia receiving high-dose chemotherapy and autologous stem cell transplantation

Erratum: Pretransplant minimal residual disease level predicts clinical outcome in patients with acute myeloid leukemia receiving high-dose chemotherapy and autologous stem cell transplantation

Clinical relevance of MRD in children undergoing allogeneic stem cell transplantation for ALL

The toxicity and expense of allogeneic stem cell transplantation (ASCT) is such that it is currently reserved for the treatment of children deemed to be at unacceptable risk of relapse when treated with chemotherapy alone. Although ASCT carries at least 10% treatment-related mortality, the commonest cause of death in transplant recipients is relapse of the original disease. This chapter reviews the literature pertaining to the clinical significance of minimal residual disease (MRD) in ASCT. Particular emphasis is placed on the strong correlation between pre-transplant minimal residual disease burden and clinical outcome. Recommendations for the design of MRD-based therapeutic trials are given.

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.

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 (10(-2) to 10(-3)), 10 had low-level MRD (< 10(-3)), 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.

Autologous Bone Marrow Transplantation for Acute Promyelocytic Leukemia in Second Remission: Prognostic Relevance of Pretransplant Minimal Residual Disease Assessment by Reverse-Transcription Polymerase Chain Reaction of the PML/RARα Fusion Gene

Reverse-transcription polymerase chain reaction (RT-PCR) of the PML/RARα fusion gene may predict relapse in acute promyelocytic leukemia (APL) patients in hematologic complete remission (CR). We have prospectively studied by RT-PCR 15 PML/RARα+ APL patients undergoing autologous bone marrow transplantation (ABMT) in second CR. The median time of first CR duration was 12 months (range, 6 to 40). All patients were reinduced with all-trans retinoic acid (ATRA), followed in 12 of 15 cases by mitoxantrone and Ara-C as consolidation. Fourteen patients received the BAVC (BCNU, Ara-C, m-AMSA, and VP-16) schedule as conditioning regimen. Unpurged marrows were collected immediately before conditioning treatment, analyzed by RT-PCR, and reinfused at median of 2 months (range, 2 to 7) from the achievement of second CR. Seven patients were PCR+ and eight PCR− for PML/RARα in their pretransplant marrows. All seven patients of the former group remained PCR+ during the follow-up and relapsed at a median time of 5 months (range, 2 to 9) from ABMT and 9 months (range, 4 to 14) from second CR. Of the eight PCR− patients, all remained PCR− during the follow-up controls. One patient relapsed at 10 months from ABMT, one died of a secondary (PML/RARα−) leukemia, and six are in hematologic and molecular remission at a median time of 28 months (range, 15 to 60) after ABMT and 32 months (range, 17 to 62) from second CR. Our results indicate that, in APL patients in second CR, ABMT with PML/RARα− marrow cells is likely to result in prolonged clinical and molecular remissions. Conversely, patients who test PCR+ after reinduction necessitate the use of alternative aggressive approaches, including unrelated allogeneic transplant.

Autologous Bone Marrow Transplantation for Acute Promyelocytic Leukemia in Second Remission: Prognostic Relevance of Pretransplant Minimal Residual Disease Assessment by Reverse-Transcription Polymerase Chain Reaction of the PML/RARα Fusion Gene

AbstractReverse-transcription polymerase chain reaction (RT-PCR) of the PML/RARα fusion gene may predict relapse in acute promyelocytic leukemia (APL) patients in hematologic complete remission (CR). We have prospectively studied by RT-PCR 15 PML/RARα+ APL patients undergoing autologous bone marrow transplantation (ABMT) in second CR. The median time of first CR duration was 12 months (range, 6 to 40). All patients were reinduced with all-trans retinoic acid (ATRA), followed in 12 of 15 cases by mitoxantrone and Ara-C as consolidation. Fourteen patients received the BAVC (BCNU, Ara-C, m-AMSA, and VP-16) schedule as conditioning regimen. Unpurged marrows were collected immediately before conditioning treatment, analyzed by RT-PCR, and reinfused at median of 2 months (range, 2 to 7) from the achievement of second CR. Seven patients were PCR+ and eight PCR− for PML/RARα in their pretransplant marrows. All seven patients of the former group remained PCR+ during the follow-up and relapsed at a median time of 5 months (range, 2 to 9) from ABMT and 9 months (range, 4 to 14) from second CR. Of the eight PCR− patients, all remained PCR− during the follow-up controls. One patient relapsed at 10 months from ABMT, one died of a secondary (PML/RARα−) leukemia, and six are in hematologic and molecular remission at a median time of 28 months (range, 15 to 60) after ABMT and 32 months (range, 17 to 62) from second CR. Our results indicate that, in APL patients in second CR, ABMT with PML/RARα− marrow cells is likely to result in prolonged clinical and molecular remissions. Conversely, patients who test PCR+ after reinduction necessitate the use of alternative aggressive approaches, including unrelated allogeneic transplant.