ALBERT

Description: Although the quantification of MRD by PCR or flow cytometry (FC) is of proven prognostic value in ALL, both methods were never compared in adults. Since 3- to 4-color FC showed equal or inferior sensitivity to PCR in childhood ALL, we herein prospectively compared six-color FC (6C-FC) at initial diagnosis and during early follow-up in adult ALL to RQ-PCR as reference method. Diagnostic samples from 70 consecutive ALL patients (pts) (22 T-, 48 B-lineage; 18 to 71 yrs) were screened by 14 different multiplex immungene consensus PCRs as published (Bruggemann et al., Blood, 2006) and simultaneously by 8 different 6C-FC combinations which included a total of 29 different antibodies. At least one of the consensus PCRs revealed monoclonality in 64/70 pts (91 %), whereas immunophenotypic aberrations could distinguish ALL from benign hematopoiesis in 67/70 cases (96 %). 6C-FC was positive in all PCR negative samples, while PCR was successful in all samples lacking immunophenotypic aberrations. The standardized 6C-FC method revealed a median of 4 immunophenotypic aberrations per pt (range 1 to 8). The most frequently observed aberrations in B-lineage ALL included CD58++ (76 %), CD11a++(50%), CD22++ (44 %), low CD38 (50%), and KORSA+ (35 %). Tdt+cytoplasmatic(cy)CD3+ hallmarked 86% of T-lineage ALL cases which were additionally characterized by CD7+ (100%), surface(s) CD3- (90%), CD99+ (52 %), and CD1a+ (38 %). Up to now, RQ-PCR assays have been established for 38 pts (median sensitivity 10−4; range 10−5 to 10−2) thus allowing for comparisons of MRD assessments to 6C-FC in 155 follow-up samples. 110/155 (71 %) samples were collected during the first 4 therapy months. 6C-FC marker combinations for follow-up always included CD34/CD10/CD19/CD45 in B-lineage and TdT/cyCD3/sCD3/CD5 in T-lineage ALL. Stainings were individualized by adding 2 pt-specific markers each to up to 3 tubes. A total of 155 follow-up specimens comprised 13 FC-/RQ-PCR+ (8 %), 4 FC+/RQ-PCR- (3 %), 88 concordantly negative (57 %), and 50 concordantly positive (32 %) samples. Due to extremely low MRD, in 11/13 FC-/RQ-PCR+ samples (85 %) the results of RQ-PCR were qualitatively positive only, but not accurately quantifiable. The minimum MRD level detectable by 6C-FC was 3x10−5, as proven by a concordantly positive RQ-PCR result. MRD levels obtained by both methods correlated well (Spearman r = 0.85; p 〈 0.0001) in follow-up samples that were positive both by RQ-PCR and by 6C-FC. The median ratio between 6C-FC and RQ-PCR MRD results was 0.48 (range 0.012 to 113). In 11% of samples the ratio between MRD levels obtained by the two methods differed more than tenfold. Our novel standardized 6C-FC approach is highly sensitive for MRD assessments in adult ALL. Our results suggest the applicability of 6C-FC in a multicenter setting, an excellent specificity, a good quantitative correlation and a similar sensitivity when compared to RQ-PCR. Longer follow-up and the inclusion of more samples in particular from later time points are required to decide whether or not the two methods are of comparable clinical significance.

Description: Although levels of minimal residual disease (MRD) decrease below the detection limit in most adult patients with standard-risk acute lymphoblastic leukemia (ALL) after consolidation treatment, about 30% of these patients will ultimately relapse. To evaluate the power of MRD monitoring as an indicator of impending relapse, we prospectively analyzed postconsolidation samples of 105 patients enrolled in the German Multicenter ALL (GMALL) trial by real-time quantitative polymerase chain reaction (PCR) of clonal immune gene rearrangements. All patients were in hematologic remission, had completed first-year polychemotherapy, and tested MRD negative prior to study entry. Twenty-eight of 105 patients (27%) converted to MRD positivity thereafter, and 17 of 28 (61%) relapsed so far. Median time from molecular (MRD-positive) to clinical relapse was 9.5 months. In 15 of these patients, MRD within the quantitative range of PCR was measured in hematologic remission, and 13 of these patients (89%) relapsed after a median interval of 4.1 months. Of the 77 continuously MRD-negative patients, only 5 (6%) have relapsed. We conclude that conversion to MRD positivity during the early postconsolidation phase in adult standard-risk ALL patients is highly predictive of subsequent hematologic relapse. As a result of the study, as of spring 2006, salvage treatment in the ongoing GMALL trial is intended to be started at the time of recurrence of quantifiable MRD.

Description: With short intensive chemotherapy mainly based on HDMTX, fractionated alkylators and HDAC outcome of Burkitt’s NHL and mature B-ALL (B-ALL) in adults could be improved substantially to CR rates of 80% and overall survival (OS) of 50–70% (Hoelzer et al, Blood, 1996). Further intensification - namely increase of MTX dose - failed to improve these results. Therefore the German Multicenter Study Group for Adult ALL (GMALL) invented in 2002 a new protocol for mature B-ALL/Burkitt and other high-grade NHL, namely primary mediastinal (med) DLBCL, including 6x Rituximab® 375 mg/m2 before each chemo cycle and two R maintenance cycles. In addition 2 cycles based on HDAC 2 g /m2 were included. HDMTX was 1,5 g/m2 in the protocol for younger pts (55 yrs) received a dose reduced regimen without HDAC and with MTX at 500 mg/m2. 227 pts with Burkitt (27=Burkitt-like), B-ALL or med DLBCL aged between 16 and 78 enrolled between 09/02 and 12/06 were evaluable for response after the first two cycles. The median age was 36 yrs for Burkitt, 46 for B-ALL and 35 for med DLBCL; 18%, 41% and 12% were older than 55 yrs respectively. The subgroups were characterised as follows: 115 Burkitt (stage III–IV 52%, extranodal inv. 78%, aaIPI 〉1 47%), 70 B-ALL, 42 med DLBCL (stage III–IV 55%, extranodal inv. 71%, aaIPI 〉1 61%). The CR rate was 90% in Burkitt, 83% in B-ALL and 69% in med DLBCL; death under therapy occurred in 3%, 11% and 0% respectively. The overall survival at 3 yrs was 91% for Burkitt, 79% for B-ALL and 90% for med DLBCL in pts at the age of 15–55 yrs and 84%, 39% and 67% (N=5) respectively in pts 〉55 yrs. CNS relapses were observed in 3 out of 22 older CR patients with B-ALL whereas in younger pts the CNS relapse rate was 0. CNS relapses are among the reasons for inferior outcome in elderly B-ALL in contrast to elderly pts with Burkitt or med DLBCL. CNS relapse rate may hopefully be reduced by inclusion of an intermediate dose ARAC cycle in the elderly B-ALL. There was no difference in OS between pts with Burkitt (92%) vs Burkitt-like NHL (86%). Since no prognostic factors could be identified in younger pts, there was no need for SCT in CR1. Major grade III/IV toxicity was hematological (28–37%) and mucositis (36%, 37%, 28% in cycles A1, B1, C1 respectively). Compared to the previous GMLL trial B-NHL90 (without Rituximab) with 270 pts the OS of 272 patients (including LBL, LCAL, DLBCL) at 3 yrs improved significantly from 54% to 80% (p

Description: Abstract 170 The effect of Rituximab in conjunction with a chemo induction and consolidation therapy was studied in CD20+, Ph/BCR-ABL negative B-precursor ALL (Pre-B/Common) in the GMALL Study 07/2003. The rationale were encouraging results with combined intensive chemotherapy and Rituximab in CD20+ adult Burkitt lymphoma / leukemia. Furthermore that in previous GMALL studies, improvement of B-precursor ALL by intensification of chemotherapy was limited and the observation that patients with CD20+ cells (antigen expression 〉20%) had an inferior outcome in adult ALL (Thomas et al. Blood 2009. 113;6330). Aim: In standard risk (SR) patients the aim was to increase the rate of molecular remission (Mol. CR) thereby decreasing the relapse rate and in high risk (HR) patients to reduce the pre-transplant tumour-load and thereby reducing the relapse rate after SCT which was 30–40% in previous GMALL studies. Materials and Methods: Adult ALL patients (15 – 55 years) with standard risk B-precursor ALL being CD20 pos. received Rituximab 375 mg/m2 at day -1 before each induction course (phase I and II), the re-induction course and before each of the six consolidations for a total of 8 doses. High Risk patients, defined as WBC 〉 30.000 and/or late CR 〉 4 weeks, which are candidates for a stem cell transplantation in CR 1 after wk 16, received Rituximab three times (d -1 ind. I/II and Cons. I) before SCT. Patients receiving Rituximab were compared with earlier CD20+ patients in the GMALL study 07/2003 with identical chemo- and supportive therapy but no Rituximab. MRD method and chemo backbone was described earlier [Brüggemann, Blood 2006: 107;1116]. Results: A total of 263 CD20 pos. patients were analyzed in the GMALL study 07/2003; 196 were SR and 67 HR patients. 181 received Rituximab (R+ arm) and were compared to a cohort of 82 patients earlier recruited without Rituximab (R- arm). In the SR there was no difference in the results of induction therapy with a CR rate of 94 % and 91 % in the R+ vs. R- patients. There was also no difference in ED rate 5% vs. 3% or failure/PR 1% vs. 5%. However, MRD course differed substantially. Decrease in MRD load in the R+ vs. R- arm was faster with a Mol CR (MRD