FC Plus Rituximab Research Articles

Comparison of Overall Survival in High Risk Patients with Minimal Residual Disease after First-Line Treatment across Three Generations of Phase 3 Trials of the German CLL Study Group

Comparison of Overall Survival in High Risk Patients with Minimal Residual Disease after First-Line Treatment across Three Generations of Phase 3 Trials of the German CLL Study Group

High-Resolution Genomic Copy Number Analysis on Sequential Samples from the CLL8 Trial: Relation Between Clonal Evolution and Defects in DNA Damage Response?

Genomic abnormalities have strong prognostic impact in chronic lymphocytic leukemia (CLL). However, clonal evolution has been studied in a limited number of cases and not within the setting of current standard therapy. It was therefore our aim to study changes in the composition of copy number alterations (CNA) over time with and without the influence of chemo(immuno)therapy.Sequential samples of 92 patients enrolled on the CLL8 trial of the GCLLSG were analyzed by Affymetrix® 6.0 single nucleotide polymorphism (SNP) arrays. Since procurement of a relapse sample was a prerequisite for this study, the cohort was not representative for the CLL8 trial [21% CR (N=19), 63% PR (N=58), 13% non response (N=12), 3% missing response (N=3)]. 48 patients received Fludarabine / Cyclophosphamide (FC), 44 patients FC plus Rituximab (FCR). Samples were taken at 3 time points: pre-treatment [N=27], time of first treatment in CLL8 [N=92] and post treatment at relapse / progressive disease [N=74]. The median observation period between samples was 35 months [range: 6-127] for the pre-treatment vs. first treatment and 41 months [range: 5-87] for the first treatment vs. post treatment (relapse) comparisons.The majority of cases maintained genomic stability over time. This applied in particular for the comparison between pre-treatment and first treatment [N=21 of 27; 78%] but also for the comparison between first treatment and relapse [N=49 of 74; 66%].The cohort was characterized by a high proportion of high-risk genomic abnormalities [30% del(11)(q22.3), 22% TP53 loss and/or mutation in the pre-treatment cohort; 36% del(11)(q22.3), 24% TP53 loss and/or mutation in the post treatment cohort]. The acquisition of novel clonal CNAs was associated with these: In the pre-treatment to first treatment comparison [N=27], only 6 [22%] cases had novel CNAs emerging over time and all of them carried an ATM loss by del(11)(q22.3). Eight cases with ATM and/or TP53 alteration and all cases without high-risk genomic aberrations [N=13] showed no evidence of clonal evolution. In the first treatment to relapse comparison [N=74], 25 [34%] cases had newly acquired CNAs at relapse and 20 of them [80%] carried high-risk genomic abnormalities [44% del(11)(q22.3), 36% TP53 loss and/or mutation]. Only 5 cases lacking ATM or TP53 alteration had newly acquired CNAs. In contrast, genomic stability was observed in 25 cases with high-risk genomic abnormalities [15 cases with del(11)(q22.3), 10 with TP53 loss and/or mutation) and 24 cases without high-risk abnormalities. No statistically significant increased incidence of clonal evolution was observed in IGHV unmutated cases [N=66 of 92] [24 (80%) cases with clonal evolution vs. 42 (70%) cases without clonal evolution, p=0.3].Nine patients had samples available from all 3 time points. While mainly genomic stability could be observed prior to treatment, 3 cases acquired 3 novel CNAs each after therapy.Comparing both treatment arms in the post treatment cohort [N=74] revealed a higher incidence of clonal evolution after treatment with FCR [FCR: N=16 of 35, 46%; FC: N=9 of 39, 23%; p=0.04] at a similar median observation period [36 and 35 months, respectively]. Also, the mean number of newly acquired CNAs at relapse was higher in the FCR treated group [3.3 vs. 2.6]. With regard to response no statistically significant differences were observed between cases with and without clonal evolution [cases with clonal evolution: 6 (24%) CR, 17 (64%) PR, 2 (8%) non-response; without: 8 (17%) CR, 31 (66%) PR, 8 (17%) non-response, p=0.5].Loss of clonal lesions was rare, occurring only under the selection pressure of therapy: 16 CNAs in 11 cases were not observed anymore at relapse. 3 of these CNAs were subclonal at time of first treatment [10-20% allelic burden] and might not yet have re-emerged after relapse. 5 CNAs in 4 cases were lost at relapse. The remaining 8 CNAs were del(13q) [N=5] and del(11q) [N=3] that were lost for a probably more advantageous del(13q) / del(11q) clone. The appearance of a more advantageous del(13q) / del(11q) clone was linked to a larger deletion size [N=3] or a larger discontinuous deletion very likely resulting from chromothripsis [N=3].The results of this study support previous data of a high genomic stability in CLL cases lacking alterations of TP53 and/or ATM. However, application of chemo(immuno)therapy did slightly increase the number of cases acquiring novel clonal CNAs. DisclosuresStilgenbauer:Pharmacyclics, Janssen: Honoraria, Research Funding.

Fludarabine, cyclophosphamide and rituximab as first-line treatment in patients with newly diagnosed follicular lymphoma.

Fludarabine-based regimens are highly effective as first-line therapy in patients with follicular lymphoma. Nevertheless, noticeable haematological toxicity has been reported using fludarabine-based regimens. To analyse the combination of low-dose oral fludarabine and cyclophosphamide plus rituximab (FCR) as induction therapy, followed by rituximab as maintenance therapy. We retrospectively analysed 73 patients diagnosed with low-grade follicular lymphoma treated with two different schemes: attenuated oral (AO) and standard intravenous (SIV) FCR. Overall response rate (ORR) was 95% (complete response rate, CRR 79.5%, partial response, PR 15.4%). CRR was 84.6% in AO vs. 61.9% in SIV (P=0.058). 44.4% of patients underwent maintenance therapy. Grade 3-4 toxicities included neutropenia: 65.4%; anaemia: 39.7%; thrombocytopenia: five patients; infectious complications: six patients. There were no treatment-related deaths. 6.8% had a secondary malignancy. Progression-free survival (PFS) was 84.6% at 12yr. The following variables influenced PFS in multivariate analysis: Hb<12g/dL [HR 4.7 (95% CI 1.18-18.6)], response after induction [HR 4.9 (95% CI 1.01-24)] for PR vs. CR and [HR 21.27 (95% CI 4.33-104)] for SD/DP vs. CR. OS was 83.1% at 12yr. The following variables significantly influenced OS in multivariate analysis: not receiving rituximab as maintenance therapy (HR 10.7 (95% CI 1.4-82.5), increased levels of β2-microglobulin [HR 5.2 (95% CI 1.16-23.7)]. FCR allowed us to obtain a high response rate, which translated into promising progression free and overall survival with an acceptable and manageable toxicity profile, especially with the attenuated oral scheme.

Updated Follow-Up In The CLL2007FMP Trial Supports The Non Superiority Of The Association Fludarabine-Cyclophosphamide (FC) Plus Campath Compared To FC Plus Rituximab and Points Out The Importance Of Minimal Residual Disease, By 6-Color Flow Cytometry Technique, On Progression Free Survival

▪ IntroductionCLL2007FMP (fit medically patients) is a Randomized Phase-III Trial conducted by the French Cooperative Group on CLL and WM (FCGCLL/WM) and the “Groupe Ouest-Est d'Etude des Leucémies et Autres Maladies du Sang” (GOELAMS), comparing FC plus Rituximab (FCR) to FC plus Campath (FCCam) in previously untreated fit patients with chronic lymphocytic leukemia (CLL). Early results showed that the FCCam regimen was associated with an unfavourable safety profile limiting significantly its use in this indication (Blood 2012). We present here the results of the extended follow up of the CLLFMP2007 trial, with particular emphasis on survival data, minimal residual disease (MRD) and late adverse events. MethodsIn this trial, 178 younger (<65) fit patients (pts) (cumulative illness rating scale (CIRS) score of up to 6), were enrolled between November 2007 and January 2009. Cases with del(17p) were excluded. Pts were randomly assigned to receive 6 oral courses of FCR (n=83) or FCCam arm (n=82). The primary endpoint of the study was 3-year progression-free survival (PFS). Secondary endpoints were safety, response to treatment, overall survival (OS) and MRD. MRD evaluation was performed by 6-color flow cytometry in an oligocentric manner. MRD testing was scheduled before therapy initiation and at final evaluation, (i.e. 3 months after completion of immunochemotherapy) where it was to be assessed for all responding patients in both peripheral blood (PB) and bone marrow (BM). Recruitment was interrupted in January 2009 after 165 pts had been randomized due to an excess of mortality in the FCCam arm. ResultsPFS and OS were not significantly different between the two arms. With a median follow-up of 55.5 months (interquartile range, 50-60), 57 pts in the FCCam arm were free of disease progression compared with 50 in the FCR arm, with a 3-year estimated PFS at 81% in both arms (p=0.80). Fourteen pts died in the FCCam arm (7 from progression and 7 from toxicity) and 9 died in the FCR arm (all from progression), with a 3-year estimated survival at 90% vs. 88% (p=0.85). PFS was significantly impacted by IGHV mutational status (p=0.001), Binet stage (p=0.0002) and MRD level. At final evaluation, MRD was established using the result in PB samples (available for 120 patients) and was determined in 103 pts by combining the results from blood and BM samples. Interpretation was based firstly on the use of the classical 10-4 threshold as reference and secondly on the limit of detection of the technique (0.7x10-5). In MRD-positive patients, the median PFS was 44.7 months (PB) whereas it was not reached in the group with MRD lower than 10-4 (p<0.0001, figure 1) ; similar data were found in MRD-positive PB+BM patients with a median PFS of 46 months whereas it was not reached in the group with MRD lower than 10-4 (p=0.002). No significant difference was found regarding OS but follow-up is still short for this evaluation. Similar results were observed when considering the limit of detection of the MRD technique (data not shown). Late toxicities (occurring after the final evaluation at 3 months after the end of treatment or at the ninth month when treatment was prematurely stopped) included : 1 bile duct cancer, 1 myelodysplastic syndrome, 1 transient ischemic attack, 1 lung adenocarcinoma and one prostate cancer in the FCR arm and 3 febrile neutropenia, 3 pneumonia (1 due to legionella), 1 pneumococcal sepsis, 1 bronchitis, 1 toxoplasma eye infection, 1 pyelonephritis, 2 herpes zoster, 1 acrodermatitis, 1 subdural hematoma, 1 autoimmune thrombocytopenia, 1 agranulocytosis, 1 autoimmune haemolytic anaemia in the FCCam arm. [Display omitted] ConclusionResults of this extended follow-up of the CLL2007FMP trial confirm the absence of superiority of the FCCam regimen on OS and PFS. Interestingly, longer follow-up did not reveal a higher rate of late toxicity in FCCam arm, notably in terms of secondary malignancies; Similarly to early toxicity, late was adverse events were mainly infectious. Finally, MRD status determined by 6-color technique in PB and/or BM at post-treatment evaluation was predictive of PFS whatever the treatment arm. This finding is in line with recent reports in other studies pointing out to the powerful value of MRD as prognostic factor, supporting its use as PFS surrogate primary endpoint in clinical trials. Disclosures:Feugier:roche: Honoraria. Cazin:roche: meeting invitation Other, Membership on an entity's Board of Directors or advisory committees; GSK: meeting invitation, meeting invitation Other, Membership on an entity's Board of Directors or advisory committees.

Extended Follow up of the CLL8 Protocol, a Randomized Phase-III Trial of the German CLL Study Group (GCLLSG) Comparing Fludarabine and Cyclophosphamide (FC) to FC Plus Rituximab (FCR) for Previously Untreated Patients with Chronic Lymphocytic Leukemia (CLL): Results On Survival, Progression-Free Survival, Delayed Neutropenias and Secondary Malignancies Confirm Superiority of the FCR Regimen

AbstractAbstract ▪435▪This icon denotes a clinically relevant abstract IntroductionEarly results for the randomized phase-III CLL8 trial of the German CLL study group (GCLLSG) showed that the addition of a monoclonal antibody, rituximab (R), to a first-line chemotherapy composed of fludarabine and cyclophosphamide (FC) improved both the progression-free survival (PFS) and the overall survival (OS) of patients with CLL (Hallek M and Fischer K et al., Lancet 2010). On the basis of these findings FCR has become the standard therapy for physically fit patients with previously untreated CLL. In this report, we present the results of an extended follow up of the CLL8 trial, with particular emphasis on survival data and late adverse events such as secondary malignancies and late neutropenia following chemoimmunotherapy with FCR. MethodsIn this prospective, randomized, open-label phase-III trial, 817 treatment-na•ve patients with good physical fitness and CD20-positive CLL received six courses of either FCR or FC. The primary endpoint of the study was progression-free survival (PFS). Secondary endpoints were safety, response to treatment and overall survival (OS). ResultsAt a median follow up of 5.9 years, 38.0% of the patients in the FCR group were free of disease progression compared with 27.4% in the FC group (hazard ratio, 0.6 (CI 95%, 0.5 – 0.7), p<0.0001). At the same time-point, 69.4% of the patients were alive in the FCR group versus 62.3% in the FC group (hazard ratio 0.7 (95% CI, 0.5 – 0.9), p=0.001; figure 1). Median OS was not reached for patients in the FCR group, while median OS was 86.0 months (95% CI, 78.7 – 93.2 months) for the FC group (p = 0.001). Significant improvements in PFS were achieved in all Binet stages including patients with Binet stage C disease (median PFS, 42.5 months for patients in the FCR group versus 33.1 months for patients in the FC group (hazard ratio, 0.7 (95% CI, 0.5 – 0.9), p = 0.02; figure 2).All patients who had received at least one of the study drugs were included in the safety analysis (n=800). The frequency of secondary malignancies with a mean time to onset of 2.4 years after the end of treatment (range 2.0 days – 7.4 years) was independent of the treatment given (40 patients (9.9%) in the FCR group versus 48 patients (12.1%) in the FC group (p = 0.4)). Of these 88 patients, 46 patients (5.7%) had a solid tumor, 33 patients (4.1%) had a Richter transformation, and 12 patients (1.5%) had a myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML). These secondary malignancies did not occur with any difference when comparing the FCR with the FC arm. Late neutropenias as defined by the presence of grade 3 or 4 neutropenia 2 months after end of treatment, were noted more frequently in the FCR group (67 patients, 16.6%) compared to the FC group (35 patients, 8.8%) (p=0.007)). Late neutropenias occurred predominantly during the first year after end of FCR treatment and decreased one year after end of treatment (16 patients (3.9%) in the FCR group versus 15 patients (3.7%) in the FC group, p=0.7). ConclusionThe results of an extended follow up of the CLL8 protocol of the GCLLSG corroborate the concept of a superior outcome for patients that received FCR chemoimmunotherapy. Secondary malignancies occurred at 9.9% in the FCR arm and at 12.1% in the FC arm. FCR induced an increase in prolonged neutropenias during the first year after end of treatment but not thereafter. These prolonged neutropenias following FCR did not translate into an increased rate of MDS/AML thus far. [Display omitted] – FCR– FC [Display omitted] – FCR– FC Disclosures:Fischer:Roche: Honoraria, travel grants Other. Bahlo:Roche: Honoraria. Fink:Roche: travel grants Other. Böttcher:Roche: Honoraria, Research Funding. Mayer:Roche: Consultancy, Research Funding. Kneba:Roche: Membership on an entity’s Board of Directors or advisory committees, Research Funding. Eichhorst:Roche: Honoraria, Research Funding. Wendtner:Roche: Honoraria, Membership on an entity’s Board of Directors or advisory committees, Research Funding. Stilgenbauer:Roche: Consultancy, Honoraria, Research Funding. Hallek:Roche: Consultancy, Honoraria, Research Funding.

Making Advances in First-Line Chronic Lymphocytic Leukemia Treatment

Bendamustine was developed in East Germany as a potent alkylating agent. Bendamustine monotherapy was US Food and Drug Administration approved for both first-line and salvage treatment of patients with chronic lymphocytic leukemia (CLL) as a result of superior progression-free survival (PFS) compared with that seen with chlorambucil monotherapy in first-line treatment of CLL. Although CD20 monoclonal antibody (mAB) monotherapy has limited activity in CLL, the German CLL Study Group (GCLLSG) confirmed in their CLL8 trial that the addition of rituximab to chemotherapy significantly advanced treatment for CLL, with improved PFS and overall survival (OS). On the basis of these data and data from the REACH (Rituximab in the Study of Relapsed Chronic Lymphocytic Leukemia) trial, rituximab was US Food and Drug Administration approved in combination with fludarabine and cyclophosphamide (FC) for firstline and previously treated CLL. The GCLLSG reported results of a phase II trial of bendamustine combined with rituximab (BR) for relapsed and refractory CLL, and in the article that accompanies this editorial, Fischer et al, on behalf of the GCLLSG, report results of a first-line, multicenter phase II clinical trial of BR for CLL. They report an overall response rate of 88%, complete remission (CR) rate of 23%, and estimated median PFS of 34 months for 117 previously untreated patients with CLL. Grade 3 or 4 neutropenia, thrombocytopenia, and anemia occurred in 20%, 22%, and 20% of patients, respectively. Randomized first-line phase III clinical trials that evaluated chlorambucil versus fludarabine, then fludarabine versus FC, and most recently, FC versus FC plus rituximab (FCR), established FCR as a standard chemoimmunotherapy (CIT) regimen that provides excellent PFS and OS. Other agents have been evaluated in phase III trials as monotherapy compared with chlorambucil, such as alemtuzumab and bendamustine, and have garnered US Food and Drug Administration approval on the basis of superior PFS, but hardly represent significant therapeutic advances. Several phase II trials have aimed to improve outcomes by modifying FCR, but without obvious success. Examples include increasing the dose of rituximab, adding mitoxantrone, adding alemtuzumab, or adding a granulocytemacrophage colony-stimulating factor to the FCR regimen. These first-line phase II BR trial results can be compared with standard first-line CD20 mAb–containing CIT regimens like FCR, FR, or pentostatin, cyclophosphamide, and rituximab (Table 1). The difficulty in comparing outcomes between clinical trials is that

Minimal Residual Disease Quantification Is an Independent Predictor of Progression-Free and Overall Survival in Chronic Lymphocytic Leukemia: A Multivariate Analysis From the Randomized GCLLSG CLL8 Trial

To determine the clinical significance of flow cytometric minimal residual disease (MRD) quantification in chronic lymphocytic leukemia (CLL) in addition to pretherapeutic risk factors and to compare the prognostic impact of MRD between the arms of the German CLL Study Group CLL8 trial. MRD levels were prospectively quantified in 1,775 blood and bone marrow samples from 493 patients randomly assigned to receive fludarabine and cyclophosphamide (FC) or FC plus rituximab (FCR). Patients were categorized by MRD into low- (< 10(-4)), intermediate- (≥ 10(-4) to <10(-2)), and high-level (≥ 10(-2)) groups. Low MRD levels during and after therapy were associated with longer progression-free survival (PFS) and overall survival (OS; P < .0001). Median PFS is estimated at 68.7, 40.5, and 15.4 months for low, intermediate, and high MRD levels, respectively, when assessed 2 months after therapy. Compared with patients with low MRD, greater risks of disease progression were associated with intermediate and high MRD levels (hazard ratios, 2.49 and 14.7, respectively; both P < .0001). Median OS was 48.4 months in patients with high MRD and was not reached for lower MRD levels. MRD remained predictive for OS and PFS in multivariate analyses that included the most important pretherapeutic risk markers in CLL. PFS and OS did not differ between treatment arms within each MRD category. However, FCR induced low MRD levels more frequently than FC. MRD levels independently predict OS and PFS in CLL. Therefore, MRD quantification might serve as a surrogate marker to assess treatment efficacy in randomized trials before clinical end points can be evaluated.

Final Analysis of Induction Treatment with Fludarabine, Cyclophosphamide Plus Rituximab (FCR) Followed by Fludarabine Plus Rituximab (FR) and Remission Maintenance Therapy with Rituximab In Previously Untreated B-Chronic Lymphocytic Leukemia (B-CLL): The Chairos AGMT CLL4/Roche ML18434 Study

AbstractAbstract 1380 Introduction:Intensification of treatment in CLL, specifically with FCR, has yielded impressive remission rates across almost all risk groups, with the exception of disease with del17p. However, despite initial remission patients relapsed with a median PFS time of 51.8 months in the CLL8 German randomized FCR trial, making remission maintenance an interesting concept. Study design:Three cycles of FCR, used initially to break potential resistance, were followed by 3 cycles of FR, with the intent to limit toxicities. Maintenance treatment was rituximab 375mg/m2 in 3 monthly intervals for 2 years. Here we report final results from this phase II study in previously untreated patients with CLL. Primary objective was the analysis of CR rates, according to NCI-WG criteria. An interim analysis of the induction phase of the study had been presented at ASH 2007. Results:We present 43 patients as intention-to-treat (ITT) population. The median age was 60 years (36 to 81), Rai stages III and IV were present in 21%, median β2-MG levels were 3.35 mg/L and 60% had at least one high risk factor out of mutation state, CD38 or FISH. Median follow up is currently 32 months. All ITT patients are evaluable for toxicity and 33 patients are evaluable for response after induction. 10 patients failed to reach the staging after the end of induction: 4 for septicemia/neutropenia, 1 for rituximab intolerance, 2 for hemolysis and 3 due to unrelated adverse events. The currently presented response assessment includes stringent use of CT scan results and is thus corrected from the 2007 interim results, obtained using clinical staging only. The response rates at the end of induction treatment were 27% CR, 36% CRi, 6% CRu, 21% PR and 9% SD. The previously reported high rate of CRi with incomplete marrow recovery argues that following FCR by FR did not improve tolerability. Nevertheless 81% of ITT patients were able to receive 6 cycles of therapy. Overall response and toxicity results are comparable with previously presented large datasets on FCR. A completely novel experience, however, is presented regarding the rituximab maintenance phase. Of the 43 ITT patients, 31 (72%) received at least one dose of maintenance treatment (the maintenance population). Regarding the maintenance population 74% finished all 8 cycles. Reasons to stop maintenance were: infections in 13% (all of them grade 1 and 2, although one patient died from herpetic encephalitis, 4 months after maintenance treatment had been stopped for a grade 2 infection), leucopenia 3%, progressive disease in 3 % and patient wish in 6%. Surprisingly, only a maximal 10% drop in serum IgG or IgM levels was observed during 2 years of maintenance. Regarding response assessment 5/31 (16%) patients receiving maintenance treatment improved their responses and 7/7 patients in CRi after induction resolved their cytopenias during maintenance treatment. Progression-free survival (PFS) in the ITT population was 78% after 36 months and was not significantly influenced by age >60, mutation state, β2-MG > 3,5 mg/L, creatinine clearance <60ml/min or permitted comorbidities. However, positive DAT and best response to therapy as well as an early drop in measured MRD levels predicted PFS. Two patients with del17p had expected inferior PFS, while none of the 5 patients with del11q has relapsed yet. Considering only the maintenance population, the PFS at 36 months was 77% and an overall best response of CR/CRu during maintenance led to a PFS of 91% after 36 months. Conclusion:Induction with an FCR like-induction protocol followed by rituximab maintenance proved feasible. However, 28% of the ITT population did not start maintenance treatment and 18% (ITT) could not finish the maintenance treatment, although no severe toxicities were responsible for the latter. One toxic death was observed, which is in the range reported for FCR without maintenance. Noteworthy, we did not find an influence of mutation state or β2-MG on PFS as has been previously reported, making it interesting to speculate that maintenance may be able to overcome such risk factors at least in mid-term, while keeping in mind the limited size of our sample. We currently run a multi-national randomized trial comparing rituximab maintenance with observation after R-containing induction in 1st and 2nd line that may be able to more definitely answer these questions (NCT01118234). Disclosures:Egle:Roche: Honoraria, Research Funding, Speakers Bureau. Off Label Use: Rituximab maintenance therapy in CLL. Petzer:Roche: Honoraria. Fridrik:Roche: Honoraria, Speakers Bureau. Greil:Roche: Honoraria, Research Funding, Speakers Bureau.

Serum Factors Predict Therapeutic Outcome In Patients with Chronic Lymphocytic Leukemia Treated In the CLL8 Trial of the German CLL Study Group (GCLLSG)

AbstractAbstract 918 Introduction:Various factors are used to predict the clinical course of chronic lymphocytic leukemia (CLL). Here we evaluated the importance of the serum markers soluble CD23 (sCD23), β2-microglobulin (s-β2m) and thymidine kinase (s-TK) compared to other prognostic markers in patients (pts) receiving a first-line therapy with fludarabine and cyclophosphamide (FC) or FC plus rituximab (FCR) in the CLL8 trial of the GCLLSG. Methods:The analysis of the serum factors was centrally performed prior to treatment. Kaplan-Meier method and the log-rank test were used to compare progression-free survival (PFS) and overall survival (OS). A Cox regression model was applied for the multivariate analysis and included the following parameters: age, sex, disease stage, time from first diagnosis, ECOG performance status, presence of B-symptoms, white blood cell count, sCD23, s-TK and s-β2m genomic aberrations, IGHV mutational status, and type of therapy (FC versus FCR). Cut-off values for s-β2m and for s-TK were 3.5 mg/l and 10 U/l, respectively. Results:Since exploratory analyses suggested a particular role for sCD23, the contribution of sCD23 was examined in more detail. Data for sCD23 was available for 616 pts. This cohort was representative of the full trial population (n=817) with respect to baseline characteristics. Results for response rates, PFS, OS and sCD23 were available for 574 pts. Based on their sCD23 levels, pts were categorized by quartiles: 139 pts were assigned to group A (≤2807.5 U/mL), 145 to group B (>2807.5 U/mL -≤5507 U/mL), 146 to group C (>5507 U/mL-≤10920.5 U/mL) and 144 to group D (>10920.5 U/mL). Significant differences were observed with respect to complete remissions (CR): Pts of group A achieved a double fold CR rate (48.2%) compared to pts of group D (23.6%; p<0.001). Correspondingly, group D had the highest rate of non-responses (11.8%). PFS and OS were longest for pts with low serum levels for sCD23: The median PFS in group A was 51.9 months (mo) compared to 29.8 mo in group D (p<0.001). The median OS for group D was 62.5 mo, but was not reached in the other groups.Next multivariate analyses (Table1) were performed and showed that s-TK, 17p-deletion (del(17p)) and the type of therapy were independent predictors of PFS and OS. del(17p) was the strongest adverse factor for both PFS and OS. sCD23 and IGHV were independent prognostic factors for PFS but not for OS. s-ß2m, age and ECOG status independently predicted OS.Finally, we investigated whether combinations of different serum markers would define distinct risk groups. Using the three serum markers, four risk categories were defined: Low risk (LR) = no serum factor elevation, intermediate risk (IR) = either s-ß2m or s-TK high, but sCD23 low, high risk (HR) = high s-ß2m and s-TK, but low sCD23, and very high risk (VHR) = high sCD23, regardless of s-ß2m and s-TK. Median PFS was significantly different for the four groups, with 59.7, 49.3, 35 and 29.8 months for LR, IR, HR, and VHR patients respectively (p<0.001). However, the difference between HR and VHR was not statistically significant (p=0.2). Conclusion:Elevated serum levels of s-ß2m, s-TK, and sCD23 predict poor outcome after first-line therapy with FC or FCR in pts with CLL, independently of other features such as IGHV status or del(17p). Measurement of sCD23 in addition to s-TK and s-ß2m seems to allow a more precise prediction of PFS.Table 1Multivariate analysis of the effects of various prognostic parameters on PFS and OSPFS n=504OS n=515ParameterHazard ratio95% CIp-valueHazard ratio95% CIp valuesCD23 (>10920.5 U/mL)1.3401.020–1.7610.036–––s-TK ≥10.0 U/L1.4541.052–2.0090.0232.0071.085–3,7110.026s-β2m ≥ 3.5mg/L–––1.7261.123–2.6520.013del(17p)6.4304.154–9.952<0.0018.6604.841–15.490<0.001IGHV unmutated1.4111.035–1.9220.029–––Age–––0.6460.419–0.9950.047ECOG pre ≥ 1–––1.8681.239–2.8160.003FCR treatment0.5060.393–0.651<0.0010.6710.447–1.0080.05 Disclosures:Fink:Roche: . Pflug:Roche: Travel grants. Eichhorst:Roche: Research Funding, Travel grants. Wendtner:Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Travel grants. Mendila:Roche: Employment. Wenger:Roche: Employment. Stilgenbauer:Amgen: Research Funding; Bayer: Consultancy, Honoraria, Research Funding; Boehringer-Ingelheim: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Genzyme: Consultancy, Honoraria, Research Funding; GSK: Consultancy, Honoraria, Research Funding; Mundipharma: Consultancy, Honoraria, Research Funding; Roche: Consultancy, Honoraria, Research Funding; Sanofi Aventis: Research Funding. Hallek:Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Fischer:Roche: Research Funding, Travel grants.

TP53 Mutations and Outcome After Fludarabine and Cyclophosphamide (FC) or FC Plus Rituximab (FCR) in the CLL8 Trial of the GCLLSG.

Abstract 1267▪▪This icon denotes an abstract that is clinically relevant.Poster Board I-289The international multicenter randomized CLL8 trial evaluated 1st line treatment with FC or FCR in 817 CLL patients. Analysis of TP53 mutation status by a re-sequencing chip (Amplichip, Roche Molecular Systems) and confirmatory direct DNA sequencing were performed in a central reference laboratory. Samples were available for 628 (76.9%) patients and this cohort was representative of the full trial population regarding other baseline prognostic factors and demographics. Outcome was analyzed for subgroups defined by genetic parameters in univariate and multivariate analyses.The incidence of the TP53 mutations was 11.9% (71/628; 41 in FC arm, 30 in FCR arm). Forty-two of 51 patients (82.4%) with 17p deletion had a TP53 mutation. 5% of patients without 17p deletion (28/553) had a TP53 mutation.Patients with TP53 mutation showed lower complete response (CR) and overall response (OR) rates as compared to the group without TP53 mutation (6.9 vs. 36.4% and 62.1% vs. 95.3% (p<.01)). Lower response rates were observed for the TP53 mutation groups in both arms (FC and FCR): CR (3.2% and 11.1%), CR+PR (51.6% and 74.1%). Response rates for patients without TP53 mutation were 24.2% (CR (FC)) and 92.2% ORR (FC) vs. 47.8 (CR FCR arm) and 98.2% (ORR FCR).Median progression free survival (PFS) was significantly shorter for patients with TP53 mutations (12.4 months vs. 45 months) (HR: 4.4 (3.29-5.87) p<0.001). Median PFS was similar for the TP53 mutated subgroup in both treatment arms (FC 12.1 / FCR 15.4 months; HR 0.53 (0.31-0.9) p=0.19). PFS for the patients without TP53 mutations was 35.1 months in the FC arm and 51.9 months in the FCR arm (HR: 0.58 (0.45-0.75) p<0.001).Patients with TP53 mutation showed a median OS of 39.3 months whereas median OS was not reached in all other patients (HR 6.01 (4.08-8.87) p<0.001).Comparing OS separately for the treatment arms in the groups defined by presence or absence of TP53 mutation showed a slightly better outcome for the FCR arm for both subsets, although the comparison did not reach statistical significance (TP53 mutations HR: 0.64 (0.33-1.2) p=0.19; no TP53 mutation HR: 0.68 (0.43-1.2) p=0.09) (Fig. 1).Multivariate analysis was performed by Cox regression including age, stage, treatment arms, IGHV status, genomic aberrations and TP53 mutation. Regarding PFS (n=567), independent prognostic factors were 17p- (HR: 3.6; p<.001), TP53 mutation (HR: 2.2; p<.001), unmutated IGHV (HR: 1.7, p<.001), age (HR: 1.4; p<.001), and FCR treatment (HR: 0.52; p<.001). Regarding OS (n=580), 17p- (HR: 3.5; p<.001), TP53 mutation (HR: 2.6: p<.001), unmutated IGHV (HR: 1.6; p=.035), and FCR treatment (HR: 0.6; p=0.019) were identified as independent factors.In conclusion, 17p deletion and TP53 mutation remain powerful prognostic markers after 1st line FC and FCR treatment in CLL. When considering the genetic profile and treatment in a multivariate model, FCR improves PFS and OS compared to FC in CLL. TP53 mutations are associated with poor response and survival independent of 17p deletion. [Display omitted] DisclosuresZenz:Roche: Honoraria. Patten:Roche Molecular Systems: Employment. Truong:Roche Molecular Systems: Employment. Wu:Roche Molecular Systems: Employment. Fingerle-Rowson:Roche: Honoraria. Fischer:Roche: Travel reimbursment. Fink:Roche: Travel reimbursment. Jäger:Roche: Honoraria, Research Funding. Böttcher:Roche: Research Funding. Kneba:Roche: Membership on an entity's Board of Directors or advisory committees, Research Funding. Wenger:Roche: Employment. Mendila:Roche: Employment. Hallek:Roche: Consultancy, Research Funding. Döhner:Roche: Research Funding. Stilgenbauer:Roche: Consultancy, Honoraria, Research Funding.

Genomic Aberrations, VH Mutation Status and Outcome after Fludarabine and Cyclophosphamide (FC) or FC Plus Rituximab (FCR) in the CLL8 Trial

The international multicenter randomized CLL8 trial evaluated 1st line treatment with FC or FCR in 817 CLL patients. Analyses of genomic aberrations by FISH and VH mutation status by DNA sequencing were scheduled for a subset of countries in a central reference laboratory. Samples were available for 648 (79%) patients and this cohort was representative of the full trial population regarding other baseline prognostic factors and demographics.The incidences of the most common genomic aberrations were 13q- 56.7%, 13q- single 36.4%, 11q- 24.6%, +12 12.0%, and 17p- 8.2%. No aberration was found for these regions in 22.4%. VH was unmutated in 63.4% and V3-21 was rearranged in 4.9%. Distributions of genetic parameters were not significantly different between treatment arms. Outcome was analyzed for subgroups defined by genetic parameters in univariate analyses. Genomic aberrations according to the hierarchical model were correlated with differences in CR, CR+PR, PFS and OS in both treatment arms combined and individually (all p<.001). Particularly poor outcome was observed for 17p- in both arms (FC and FCR): CR (4.5% and 19.0%), CR+PR (45.5% and 71.4%), PFS (at 24 months: 0.0% and 29.6%), and OS (at 24 months: 41.0% and 53.3%). Unmutated VH status was correlated with shorter PFS in both arms combined and individually (all p<.001), shorter OS in the FC arm (p=.006), and a trend towards shorter OS in the FCR arm (p=.092).Treatment results of FCR and FC were compared in subgroups defined by genetic parameters to identify prognostic and predictive markers. While FCR in general improved outcome, this effect was different in specific genetic subgroups (Table).Table: Treatment effect of FCR (odds ratio (OR) and hazard ratio (HR) with respective significance level for improvement of outcome in genetic subgroups)VH mutVH unmut17p-11q-+12normal13q-singleCROR3.1963.360494110.0009.0001542.386p-value<.001<.001<.185<.001<.001.274.003CR+PPOR3.5372.7303.0001.3701.7741.03814,884p-value.083.008.124.7231.0001.000.001PFSHR.476.599.495.405.544.962.372p-value.008.001.032.001251.890.001OSHR.877.625.715.328.3701.678387p-value.800.073.420.047.373.339.104Multivariate analysis was performed by Cox regression with backward selection including age, sex, stage, treatment arms, VH status and genomic aberrations as parameters. Regarding PFS, independent prognostic factors were 17p- (HR 6.76, p<.001), unmutated VH (HR 1.97, p<.001), FCR (HR 0.51, p<.001) and +12 (HR 0.58, p=.020). Regarding OS, only 17p- (HR 7.47, p<.001) and unmutated VH (HR 2.09, p=.018) were identified as significant independent factors, while a trend was observed for FCR (HR 0.66, p=.085). In conclusion, genetic parameters remain powerful prognostic markers after 1st line FC and FCR treatment in CLL. The overall improvements by FCR result from specific treatment effects in distinct genetic subgroups and 11q- appears to benefit particularly. However, 17p- and unmutated VH status remain predictors for shorter PFS and OS independently of the overall improvement by FCR.

Standardized MRD Flow and ASO IGH RQ-PCR for MRD Quantification in CLL Patients after Rituximab-Containing Immunochemotherapy – a Comparative Analysis in 574 Samples from the Randomized GCLLSG CLL8 Trial

Combinations of purine analogues with the therapeutic anti-CD20 antibody Rituximab induce a high number of complete clinical remissions in CLL patients. Therefore, the detection of minimal residual disease (MRD) appears particularly warranted after such regimens. However, Rituximab prevents the assessment of CD20, one mainstay of flow cytometric MRD detection (MRD flow) in CLL. As no comparative analysis of MRD flow to ASO primer IGH real-time quantitative PCR (RQ-PCR) after Rituximab has been performed so far, doubts remained regarding the diagnostic utility of MRD flow in this clinical setting. We compared results obtained by RQ-PCR to 4-color 3-tube MRD flow in 574 samples from 69 patients randomized to receive fludarabine and cyclophosphamide (FC) or FC plus Rituximab (FCR). Thus we were able to investigate how treatment with FC and FCR impacts on the performance of DNA-based RQ-PCR and of antibody-dependent flow cytometric MRD detection. With 58.4 % positive and 27.0 % negative samples by both assays, we found a qualitative concordance between MRD flow and RQ-PCR of 85.4 % (490/574 samples). Only very few samples were exactly quantifiable with one method and negative by the other method (0.7% MRD flow pos/RQ-PCR neg and 1.4% MRD flow neg/RQ-PCR pos samples). Most discordant samples (12.5% of all samples) were MRD negative by flow but positive by RQ-PCR at low levels outside the quantitative range of MRD detection. We calculated a median quantitative range of 10−4 and a median sensitivity of 2×10−5 in those samples, demonstrating a higher sensitivity of PCR to detect low-level MRD. Considering a threshold of 10−4 for MRD positivity, 94% of all samples showed concordant results by both methods. The cases that were discordantly classified as to this threshold usually comprised samples with residual CLL cells close to 10−4 (3.7% RQ-PCR pos/MRD flow neg and 2.3% RQ-PCR neg/MRD flow pos samples). Quantitative MRD levels determined by both methods were closely correlated irrespective of therapy (Spearman r = 0.95 in FCR, r = 0.96 in FC). We next classified all samples according to the PCR result and compared MRD flow detection rates between the treatment arms. Regarding PCR positive samples within the quantitative range, MRD flow was positive in 96.8% and 98.0% of samples from the FCR and FC arms, respectively. Positive MRD flow results were recorded in 33.7% (FCR) and 40.0% (FC) of samples that concomitantly yielded PCR positive results outside the quantitative range. Within the PCR negative samples, MRD flow detected residual CLL cells in comparable proportions of samples (FCR 2.3% vs. FC 3.4%). We compared the MRD results obtained using combinations that include CD20 (CD20/CD5/CD19/CD43, CD79b/CD20/CD19/CD5) to the MRD results from the combination CD81/CD22/CD19/CD5 in individual samples. We found that CD20 is undetectable on residual CLL cells and therefore not informative for MRD flow in patients from the FCR arm up to 180 days after the last Rituximab infusion. For the same period of time benign B cells are significantly less frequent in patients after FCR than after FC (p<0.0001) and often account for less than 1 cell in 10,000 leukocytes after this therapy regimen. Thus the efficient reduction of background non-CLL B-cells by Rituximab likely explains the high specificity and sensitivity of MRD flow after FCR treatment. In summary, MRD assessment by flow and PCR are equally effective for MRD quantification in Rituximab treated CLL patients within the sensitivity range of 10−4, while PCR is more sensitive for detecting MRD below that level. The use of Rituximab does not influence MRD detection neither by flow nor by RQ-PCR.

Quantitative MRD Assessments Predict Progression Free Survival in CLL Patients Treated with Fludarabine and Cyclophosphamide with or without Rituximab – a Prospective Analysis in 471 Patients from the Randomized GCLLSG CLL8 Trial

Data from randomized clinical trials investigating the clinical significance of minimal residual disease (MRD) quantification during and immediately after standard therapy in CLL are still lacking. We therefore related 4-color MRD flow results to progression free survival (PFS) and achievement of complete clinical remission (CCR) in patients (pts) from the GCLLSG CLL8 trial who were randomized to receive 6 cycles of fludarabine and cyclophosphamide (FC) or FC plus Rituximab (FCR). MRD was assessed before therapy, after 3 cycles (interim staging, IS), 1 month (mo) after therapy (initial response assessment, IRA) and 2 mos after IRA (final restaging, FR). 1402 samples from 471 patients who had received at least 3 cycles of therapy were included in the analysis (1162 peripheral blood, PB, 240 bone marrow, BM samples). While initial disease levels were identical between the treatment arms, median PB MRD levels were significantly lower within the FCR arm at IS, IRA, and FR (IS 5.8×10−3 vs. 5.8×1−4, IRA 3.6×1−4 vs. 0, FR 4.4×1−4 vs. 0, all p < 0.0001). Likewise, median BM infiltration was higher after FC than after FCR at IRA (1.6×10−3 vs. 2.1×10−4, p = 0.05) and at FR (8.1×10−4 vs. 1.2×10−4, p<0.0001). The proportion of PB samples with MRD below 10−4 was significantly higher in the FCR arm than in the FC arm (33.6% vs. 6.4% at IS, 67.6% vs. 36.6% at IRA, 66.4% vs. 34.2% at FR, all p < 0.0001). BM samples with MRD levels below 10−4 at FR accounted for 47.6% and 27.3% of samples from the FCR and FC arms, respectively (p=0.005). MRD results were classified into 5 cohorts (<10−4, >=10−4 to < 10−3, >= 10−3 to <10−2, >= 10−2 to < 10−1, and >10−1) in order to assess the prognostic significance of MRD for PFS. Significant differences in median PFS were obtained when PB MRD levels at IS were grouped into <10−2 (70.2% of pts, PFS 44 mos), >= 10−2 to < 10−1 (19.7% of pts, PFS 27 mos) and >10−1 (10.1 % of pts, PFS 11 mos). The PFS of all pts who experienced an MRD reduction below 10−2 was similar, regardless of the exact level. Considering PB MRD levels at FR, clear cut differences in median PFS were observed between pts demonstrating levels < 10−4 (49.6 % of pts, PFS not reached), >= 10−4 and < 10−2 (36.8% of pts, 34 mos) and >= 10−2 (13.6% of pts, 15 mos). Further subdivisions within those 3 cohorts resulted in very similar Kaplan-Meier estimates. BM MRD levels at FR were best classified into two groups, comprising pts with MRD levels of at least 10−2 and below 10−2, respectively. Pts from the former group (11.9 % of pts) experienced a median PFS of 15 mos, compared to a median PFS of 43 mos in the latter group (88.1 %). When MRD levels and treatment regimens were analyzed simultaneously for prognostic significance, only MRD levels were identified as prognostic parameter for PFS using Cox regression in this model. This was tested for PB at IS as well as for PB and for BM at FR. Pts who received FCR achieved up to the FR significantly lower MRD levels and were more often MRD negative. However, once low-level MRD was achieved it had the same prognostic significance in both treatment arms. MRD levels at IS were also associated with the achievement of CCR. Thus, only 14.7% of patients with an MRD level of at least 10−2 achieved a CCR after therapy compared to 49.4% of pts with an MRD level below that threshold (p<0.0001). In summary higher MRD levels were generally associated with shorter PFS whereas lower MRD levels predict longer PFS, regardless of sampling time point, sample material and treatment regimen. Identical MRD levels are associated with similar PFS in both treatment arms. We show that the addition of Rituximab to FC leads to lower median MRD levels compared to FC alone, resulting in longer overall PFS in the FCR group.

Minimal Residual Disease (MRD) and T/NK Cell Dynamics during Fludarabine, Cyclophosphamide Plus Rituximab (FCR) Followed by Fludarabine Plus Rituximab (FR) and Remission Maintenance Therapy with Rituximab in Previously Untreated B-Chronic Lymphocytic Leukemia (B-CLL): Riskfactor Stratification in the Chairos Study

Based on the efficacy of Fludarabine, Cyclophosphamide plus Rituximab (FCR) as first-line therapy for chronic lymphocytic leukemia (CLL), we conducted a non-randomized multi-center phase II trial with early brief intensive induction with FCR for 3 cycles followed by fludarabine/rituximab for 3 cycles. In addition patients received a 2 year 3-monthly rituximab maintenance regimen. The trial recruited 42 patients. Clinical baseline data, clinical response results and toxicity of a planned first interim analysis were reported previously (Egle et al, ASH 2007). We now present the data from flow cytometric minimal residual disease (MRD) monitoring during the study in combination with the performed risk factor analyses and report on the findings of detailed analysis of T-cell and NK cell dynamics during the induction phase of the trial. From the previously reported high number of CRs (94% after complete 6 cycle induction) 64% were MRD negative by flow cytometric definition (less than 10−3 G/L CLL cells). All non CR patients were correctly called as MRD positive. Less than 0.1 G/L CLL cells after 3 cycles of FCR identified all patients achieving MDR negativity after the complete induction regimen. CD30 risk, as well as age, FISH cytogenetic analysis and analysis of IgVH mutation status were analysed for their impact on MRD dynamics. We could not find any difference in MRD responses between CD38 high and low risk groups or between age groups (> and <65a). Neither did abberations found in FISH cytogenetics predict the MRD responses and all CLL patients harboring an 11q deletion achieved MRD negativity. Interestingly, we found the only predictive factor to be the IgVH mutation status with 15 unmutated CLL patients significantly less likely to achieve MRD negativity after induction treatment (p=0,025). Especially, three patients with rearranged IgVH 3–21 did poorly, suggesting that these patients may need a different therapeutic approach. Six months of Rituximab maintenance therapy were able to convert 37% of patients from MRD positive state after remission induction to a MRD negative state. T and NK cell numbers and ratios were followed during therapy. In addition T cell subsets and their CD38 expression status were analysed. While absolute T and NK/NKT cell numbers decreased dramatically during treatment (CD4 and CD8 cell numbers decreased to medians of less than 100/μl after 6 cycles of induction treatment), a count of less than 200 CD4 cells/μl after 3 cycles of FCR was found to be predictive of MRD negativity. In addition, we observed an earlier recovery of NK and NKT cells after the end of induction treatment leading to an inverted NK(T) to T cell ratio early during maintenance treatment. This may have implications for NK-dependent ADCC during rituximab maintenance therapy. Regarding the CD4 and CD8 compartment we observed in addition to the reduction in absolute numbers a marked shift of T cell subsets. Naïve T cells were more completely depleted by FCR/FR treatment than T cells from the memory compartment, with central memory (CM) CD4 cells increasing from a median of 49% to 79%. This was less pronounced in the CD8 compartment. CD38 expression on T cells was monitored, because our group found that this was predictive of CLL course (Tinhofer Blood 2006). Interestingly we found a marked increase of the proportional representation of CD38+ CD8 and CD4 T cells in the T cell compartment with a more pronounced effect observed in the CD8 compartment. CD8 T cell counts were also quicker to recover during rituximab maintenance therapy. While these data point to essential caveats regarding the immunosuppressive toxicity of the treatment, they may also point to a window of opportunity regarding the modification of T cell responses during the phase of immune reconstitution after FCR/FR therapy. In conclusion we find that only IgVH mutation status was predictive of achievement of MRD negativity and we could show significant skewing of T cell and NK(T) cell subsets during chemoimmunotherapy. A better understanding of these T/NK cell dynamics may lead to the development of improved targeting of the immune recovery phase after chemotherapy using immunomodulating maintenance strategies.

Planned First Efficacy and Safety Analysis of De-Intensified Induction with Fludarabine, Cyclophosphamide Plus Rituximab (FCR) Followed by Fludarabine Plus Rituximab (FR) and Remission Maintenance Therapy with Rituximab in Previously Untreated B-Chronic Lymphocytic Leukemia (B-CLL): The Chairos Study.

Induction of long lasting complete remission is currently widely viewed as a potentially beneficial new goal in untreated CLL with indication for therapy. We devised a protocol for remission induction based on the FCR regimen, used for an initial 3 cycles to break potential resistance with an intensive regimen (phase I), followed by 3 cycles of FR (phase II), to limit toxicities of the induction phase. With the aim to maintain remissions longer we added a 2 year maintenance phase with rituximab in 3 monthly intervals. Here we report preliminary results from this non-randomized phase II study in previously untreated patients with CLL. Primary objective was the analysis of complete response rates, according to NCI-WG response criteria. Secondary goals included overall response rates, toxicity and remission quality using 4-color flow cytometric MRD analysis. Forty-three patients with a median age of 63 years (range 36 to 81) were recruited for the study. Median lymphocyte counts were 92 G/I and the median for bone marrow infiltration was greater than 80%. For this interim analysis 23 patients are evaluable for response after induction phase I and 16 patients for induction phase II. After 3 cycles of FCR we observed an 83% CR rate that was increased to 94% after addition of 3 cycles of FR. After 3 cycles of FCR 13/31 (42%) patients evaluable for MRD analysis showed less than 10−3 G/I CLL cells detectable (defined as a complete MRD response). After 6 cycles of therapy 10/17 (59%) were MRD negative. Of 12 patients evaluable after 6 months of maintenance 100% were in CR. However, in all stagings we observed about 25% of patients that clearly had no residual measurable disease (including CT scan and bone marrow biopsies) but had persistent cytopenias leading to their designation as CR with toxicity-associated cytopenias (CRtox). Scoring maximally observed toxicities across all cycles, we observed 28% grade 3 and 15% grade 4 hematotoxicities as main toxicity. Thirteen percent grade 3 infections were observed. One case of fatal herpes encephalitis was observed and occurred about one year after initiation of therapy. Two deaths were reported as not therapy related (malignant melanoma, one in the screening phase). Forty-one serious adverse events were reported, among which 10 led to a drop-out from the study. We present here a cohort of CLL patients in a multi-center setting receiving 3×FCR followed by 3×FR. We found excellent responses to initial treatment with close to 100% complete responses after 6 cycles of therapy. However, we also note an almost 25% drop-out rate, a not insignificant number of SAEs and a number of patients in CR that display long-lasting hematotoxic effects after the end of this treatment. This happened despite a de-escalation from the original FCR regimen, potentially indicating that such a dose reduction in late therapy may prevent additional toxicities without compromising efficacy. Still the efficacy of the induction regimen seems to generate a good starting point for the maintenance phase. Preliminary MRD analyses during the maintenance phase will be presented.