Non-Hodgkins Lymphoma Safety and feasibility of CHOP ... - Nature

Bone Marrow Transplantation (2003) 31, 775–782 & 2003 Nature Publishing Group All rights reserved 0268-3369/03 $25.00

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Non-Hodgkins Lymphoma Safety and feasibility of CHOP/rituximab induction treatment followed by high-dose chemo/radiotherapy and autologous PBSC-transplantation in patients with previously untreated mantle cell or indolent B-cell-non-Hodgkin’s lymphoma G He, T Flohr, C Huber, K Kolbe, H-G Derigs and T Fischer IIIrd Department of Medicine, Johannes Gutenberg-University, Mainz, Germany

Summary:

Introduction

Patients with no prior chemotherapy and with advanced and progressive follicular lymphoma (FCL) or mantle cell lymphoma (MCL) were enrolled into a treatment protocol combining CHOP/rituximab–CHOP therapy with subsequent consolidation high-dose therapy (HDT) to evaluate the safety and feasibility of this treatment. Overall, 15 patients were enrolled and 13 patients completed the entire treatment protocol without major toxicities or increased infectious complications. One patient withdrew consent after achieving complete remission (CR) prior to HDT. One patient was taken off study with signs of disease progression after induction treatment. All patients showed stable engraftment after HDT. Response rates appear to be favorable, indicating an additional effect of rituximab and HDT. Overall, 12 of 13 patients achieved CR/CRu and one patient partial remission. Follow-up of immune reconstitution displayed transient severe combined immunodeficiency with slow normalization of the cellular and humoral compartments without a significant increase of infectious complications. Taken together, high-dose chemotherapy can be safely given following treatment with CHOP+rituximab. Efficacy in this small cohort of patients was encouraging with sustained remissions in both FCL and MCL patients. Upfront HDT should be considered as a therapeutic option especially in young and/or high-risk patients. Bone Marrow Transplantation (2003) 31, 775–782. doi:10.1038/sj.bmt.1703925 Keywords: B-NHL; rituximab; high-dose therapy; autologous PBSC-transplantation; immune reconstitution

A watch-and-wait strategy is still considered as standard therapy for patients with indolent follicular lymphoma (FCL), until symptomatic disease requires therapy. Early treatment of patients with advanced disease with standard chemotherapy combinations induces high remission rates, but does not alter overall survival (OS) rates in comparison with symptom-oriented therapy. In contrast mantle cell lymphoma (MCL), although classified as an indolent lymphoma, is now known for its aggressive clinical course, requiring early intensified treatment. However, in spite of numerous efforts with conventional chemotherapy, cure has almost never been achieved in either patient group and optimum treatment strategies, especially for young patients, have still to be defined. Therefore, the introduction of new therapeutic approaches in first-line therapy may improve the prognosis for both patient groups. High-dose therapy (HDT) followed by autologous stem cell transplantation has been shown to be effective in patients with relapsed lymphoma and now represents the standard treatment for patients with chemotherapy-sensitive relapsed aggressive lymphoma.1–2 Uncontrolled trials also suggest a positive effect in progression free survival (PFS) for upfront HDT in indolent lymphoma, but the value of HDT is not yet proven, as an influence on OS has not been demonstrated so far.3–6 HDT effectively eliminates residual disease, which itself represents an important risk factor in the long-term prognosis of relapsed indolent lymphoma.7–9 Secondly, monoclonal antibody-based therapies have been shown to be effective in indolent and aggressive lymphoma in relapse and first-line therapy. Results of the first published series of patients with relapsed FCL showed an efficacy of 48% as a single agent, with a median response duration of 12 months.10 This result was confirmed in other studies.11,12 In patients with untreated FCL with a low tumor burden, four doses of rituximab given during a 4-week course induced high remission rates of 54% in a study by Hainsworth et al.13 and a 73% remission rate with a substantial proportion of molecular remissions in a French cooperative study.14 In advanced

Correspondence: Dr med. G He, IIIrd Department of Medicine, Johannes Gutenberg-University, Langenbeckstr. 1, 55131 Mainz, Germany Received 14 June 2002; accepted 26 November 2002

CHOP/rituximab plus high-dose consolidation therapy for indolent lymphoma G He et al

776

MCL, single-agent antibody treatment had only moderate activity.15 All studies demonstrated a comparably low toxicity of rituximab treatment. Combination therapies with conventional (CHOP-like) chemotherapy regimens and rituximab have been shown to be feasible and safe, with almost no added toxicity and demonstrating favorable remission rates.16–18 However, because of the prolonged natural course of indolent lymphoma, the superiority of OS in comparison to conventional therapies has so far not been seen in concurrent clinical trials. Recently, the updated data of the pivotal trial showed more than 50% of patients treated with CHOP/rituximab still in stable remission 5 years after completion of treatment.19 Supporting the evidence for synergistic effects of combination therapy, a benefit of introducing antibody treatment in standard chemotherapy was observed in a recent GELA-trial for patients with aggressive lymphoma.20 This proved to be statistically significant for both PFS and OS with a median follow-up of 24 months. The addition of rituximab in the salvage chemotherapy and conditioning regimen was feasible and safe and produced promising CR rates.21–25 Besides increasing potency of combination induction therapy, rituximab also presents a powerful in vivo agent for the clearance of stem cell products from residual disease.22,23,26,27 With these two new tools – monoclonal antibody treatment and HDT – available, we initiated a treatment protocol for evaluating the feasibility and safety of the combination of CHOP with rituximab followed by HDT with autologous stem cell support as consolidation treatment in first-line therapy. The first two cycles of CHOP were given without rituximab comedication to minimize the risk of serious tumor lysis syndrome and to identify patients refractory to CHOP therapy. Over the last few years, acute therapy-related morbidity and mortality of HDT followed by autologous stem cell support has markedly decreased, from up to 25% in the early series to less than 2% in most transplantation centers worldwide.28 Nevertheless, comparatively little is known about the toxicity and immunodeficiency caused by intensified treatment protocols in combination with antibody treatment. At the initiation of this study, the first results of our own study experience using of rituximab in HDT as salvage therapy were already available. They showed a delay in cellular and humoral immune response after chemoimmunotherapy.23 Therefore, in order to detect potentially harmful side effects, careful monitoring of immune reconstitution was included into this trial.

Patients and methods

Induction chemotherapy and rituximab treatment Treatment consisted of two cycles of CHOP (cyclophosphamide 750 mg/m2 i.v., day 1; doxorubicin i.v. 50 mg/m2, day 1; vincristine i.v., 1.4 mg/m2 (maximum 2 mg), day 1; prednisone 100 mg/m2 p.o., days 1–5) and 4 additional cycles of CHOP+Rituximab (375 mg/m2 i.v., day 1 in cycles 3–6) (R-CHOP). Treatment was scheduled every 21 days. Restaging was scheduled every two cycles and in the event of disease progression patients were taken off study.

Graft mobilization, graft collection and cell processing Patients with at least stable disease upon induction treatment received mobilization chemotherapy with DexaBEAM/G-CSF (dexamethasone 24 mg in three doses p.o., days 1–10; BCNU 60 mg/m2 i.v., day 2; etoposide 75 mg/m2 i.v., days 4-7; cytosine arabinoside 200 mg/m2 i.v., days 4–7 in two daily doses; melphalan 20 mg/m2 i.v., day 3) and GCSF (lenograstim) was given in a dose of 150 mg/m2 starting from day 11 until the completion of the mobilization procedure. Leukaphereses (LPs) were performed until a minimum of 2.5 106/kg bw CD34+ cells had been collected. LP products containing DMSO 7.5% and PBS/ HAS 4%, heparin 10 U/ml were cryopreserved using a programmed freezer (Cryoson, Scho¨llkrippen, Germany).

Conditioning regimen, PBSC reinfusion, supportive care HDT was scheduled within 8 weeks of mobilization and consisted of total body irradiation (TBI) plus cyclophosphamide (cy) (TBI 14.4 G in eight doses, days 7 to 4, cyclophosphamide 60 mg/kg bw i.v. days 3 to 2) or BEAC (BCNU 600 mg/m2 i.v., day 7; cytosine arabinoside 6 g/m2 in two doses i.v., day 6 to –3; etoposide 400 mg/m2 i.v., days 6 to –3; cyclophosphamide 90 mg/ kg bw i.v., day 2). At 48 h after completion of the conditioning regimen, LP products were reinfused. G-CSF was given starting day +4. Patients received prophylactic oral antibiotic therapy with ofloxacin. Standard antibiotic/antimycotic coverage was given. Supplements of packed red blood cells or platelets were administered for hemoglobin levels below 80 g/l or platelet counts below 10 GI/l. Intravenous immunoglobulin preparations were given to maintain IgG serum levels above 4 g/l until day +100.

Response evaluation Response evaluation referred to criteria of the International Workshop to standardize response criteria for lymphomas.29

Patient population Patients aged 18–60 years who had received no prior chemotherapy with proven CD20+ stage IIB to IV indolent NHL including MCL according to the REAL classification were enrolled into the treatment protocol given below. Prior local radiotherapy was allowed. The protocol had been approved by the local ethics committee and all patients gave informed consent. Bone Marrow Transplantation

Sample collection Samples for minimal residual disease analysis and assessment of immune reconstitution were obtained before each separate part of treatment and during follow-up with a schedule, which allowed a variation of up to 10% for the different time points.


Immunophenotyping Flow cytometry analysis for evaluation of CD34+cells, lymphoma cells, B cells, T cells and NK-cell subsets in whole-blood or LP products was performed by direct immunofluorescence according to the slightly modified ISHAGE guidelines for CD34+ cell enumeration and CD4 quantitation.30,31

Monitoring of MRD Nested PCR amplification for bcl-1 or -2 rearrangements was performed according to methods previously described.7,32 Serial samples of blood and bone marrow (BM) before, during and after treatment, or LP products were analyzed.

Definitions, statistical methods OS time was defined as the time between reinfusion of blood stem cells or end of treatment and death. PFS was considered as the time between reinfusion of blood stem cells or end of treatment and occurrence of relapse or disease progression. Survival curves were estimated as described by Kaplan and Meier.33

Results Patient characteristics Details are given in Table 1. A total of 15 patients were enrolled. According to REAL-classification diagnoses they included nine patients with Grade I or II FCL, five with MCL and one with a small lymphocytic lymphoma. Median age was 48 (range 31–60) years. One patient had stage IIB, five stage III and nine patients stage IV disease. Nine of 15 patients had BM-involvement, two presented with B-symptoms and in two patients elevated levels of LDH or 2-microglobulin were observed. Three patients

Table 1

Patient characteristics

Total Sex (f/m) Median age (range)

777

CHOP and CHOP+rituximab (R-CHOP) treatment CHOP and R-CHOP therapy was given according to the treatment schedule for all but one cycle, which was delayed because of fever of unknown origin in one patient. Overall, treatment-associated toxicity was low and no acute antibody-related side effects greater than Grade I occurred. There was no apparent additional toxicity from rituximab. All patients completed the CHOP/R-CHOP chemotherapy. One patient withdrew his consent before mobilization/high-dose chemotherapy after achieving complete remission (CR). One patient with MCL showed signs of disease progression after completing R-CHOP and was taken off the study to undergo salvage chemotherapy.

Blood stem cell mobilization and processing Sufficient numbers of peripheral blood stem cells (PBSC) were collected in all patients mobilized (13/13). One patient developed unstable angina pectoris during the mobilization procedure and underwent coronary arteriography with angioplasty. In this patient, mobilization was repeated without further complications. Adverse events of mobilization therapy were as follows: fever of unknown origin (FUO) (n ¼ 4), herpes simplex reactivation (n ¼ 2), pneumonitis (n ¼ 1), sinusitis (n ¼ 1) and thrush (n ¼ 2). All infectious complications resolved with appropriate antibiotic therapy. Median duration from the end of chemotherapy until start of LP was 9 (range 8–13) days, median number of LP procedures was 1 (range 1–3) and median CD34+ cells collected per kilogram was 9 (range 2.8–25) (Table 2). Measurement of residual B-cell contamination of the graft by cytometry revealed measurable B cells (39 cells/ml) in only one patient. Molecular analysis of LP-products from patients with initially informative molecular analysis (bcl-1 and bcl-2 rearrangements) showed negativity in all patients for the translocation products (5/5).

15 4/11 48 (31–60)

Histology (REAL-classification) Follicle center (I/II1) Mantle cell Small lymphocytic

9 5 1

Stage according to Ann-Arbor II III IV

1 5 9

BM involvement LDH/b2MG > UPN

9/15 2/15

Prior radiotherapy

1/15

UPN=upper normal value, b2MG=beta2-microglobulin.

showed extranodal involvement and three presented with bulky disease.

Table 2 Mobilization of peripheral blood stem cells and engraftment Total patients Median number of LP needed (range) Number of CD34 106/kg bw (range) Median days until start of LP (range) Hematopoietic recovery after HDT ANC >500 GI/l (days) Platelets >20 GI/l (days) Platelets >50 GI/l (days) Required transfusions Platelet transfusions Packed red cells

13 1 (1–3) 9 (2.8–25.4) 9 (8–13) Median (range) 10 (8–13) 11 (8–27) 18 (8–51)

3 (1–14) 2 (0–10)

kgbw=kilogram body weight, ANC=absolute neutrophil count. Bone Marrow Transplantation


778

Blood stem cell transplantation Median duration from LP to reinfusion of PBSC (day 0) was 61 (range 29–71) days. In all, 13 patients received HDT (TBI/Cy, n ¼ 11; BEAC/M, n ¼ 2). Chemo/radiotherapy was given with no unexpected complications. Mucositis occurred in all patients to a variable extent. During neutropenia all patients developed fever, including FUO (N ¼ 4), central line infections (N ¼ 3), pneumonia (N ¼ 2), suspected fungal pneumonia requiring amphotericin B treatment (N ¼ 2), viral reactivations (HSV) (N ¼ 2) and enteritis (N ¼ 2). No other toxicities greater than Grade II occurred (Table 3). Median time to recovery of an absolute neutrophil count (ANC) 4 500 GI/L was 10 (5–13) days. Median time to recovery of platelets 4 50 GI/l was 18 (8– 51) days. Patients required a median of two packed red cell transfusions and three thrombopheresis product transfusions during hospitalization (Table 2). During follow-up after hematopoietic recovery, the following adverse events were documented: gastric bleeding in one patient (not associated with thrombopenia), pneumonitis in four cases, aseptic necrosis of the femoral head in one patient and deep vein thrombosis in one patient. Another patient demonstrated increasing liver function values during follow-up. Liver biopsy revealed signs of hepatic cirrhosis, which was considered to be toxic and was associated with a history of alcohol abuse.

After a median follow up of 22.5 months (range 12–33 months) PFS and OS are 62 and 82% for all patients (Figure 1). For patients completing the entire treatment protocol PFS and OS rates are both 79%, with one patient dying at day +562 of invasive bladder cancer without signs of recurrence of lymphoma. One other patient, who initially presented with extensive disease, relapsed after HDT at day +180; he expired on day +246 because of disease progression. In one patient, relapse was suspected on day +180. However, after 2.5 years there have been no signs of disease progression and the patient is now considered to be in continuing CR. Both patients who did not complete the entire protocol relapsed. In five patients, bcl-1 or bcl-2 rearrangement was detected on PCR-analysis prior to therapy. After CHOP induction, three of four patients tested still showed detectable disease. After completion of the induction treatment, 4/4 patients analyzed demonstrated MRD negativity. LP products from the five informative patients showed no detectable MRD on molecular analysis. During follow up, 4/4 evaluable patients have remained PCR negative to date and none of these patients has any clinical signs of relapse.

Reconstitution of cellular and humoral immunity Immune reconstitution was monitored closely during the treatment course and post-HDT. One patient was excluded PFS and OS

Evaluation upon the first two cycles of CHOP (n ¼ 15) showed 10 patients achieving minor response (MR) or stable disease (SD), including all MCL patients. Three patients achieved partial remission (PR) and two CR. Restaging after completion of an additional four cycles of R-CHOP showed 10 patients to have achieved PR and four patients to be in CR. With MCL there was one CR and three PR. One patient with MCL had signs of progressive disease and was taken off study. Restaging at days +60 to +90 after HDT (n ¼ 13) revealed 11 CR, one unconfirmed CR (CRu) and 1 PR (Table 3). All patients with MCL completing HDT (4/4) achieved complete remission at the end of treatment. One patient with FCL achieving PR and initially presenting with retroperitoneal bulk disease received involved field radiotherapy as consolidation.

% surviving

Treatment efficacy 100 90 80 70 60 50 40 30 20 10 0

OS PFS

0

10

20 30 Months after HDT

40

Figure 1 OS and PFS post-HDT or end of therapy Kaplan–Meier estimation of OS and PFS for patients entering the study protocol (n ¼ 15). Median time post-HDT or end of therapy: 22.5 (12–32.9) months.

Therapy-related morbidity and response to therapy

Table 3 a

Morbidity (number of episodes) >1II toxicitya Infection Response evaluation oPR (%) PR (%) CR/CRu (%) a

CHOP and R-CHOP (n=90 cycles)

Mobilization (n=13)

HDC (n=13)

0 2

3 11

2 13

Two cycles of CHOP (n=15)

Four cycles of R-CHOP (n=15)

HDT (n=13)

67 20 13

7 66 26

0 8 92

Excludes hematological toxicity and mucositis. Evaluation of response after two cycles of CHOP, after completion of four cycles of R-CHOP and at day +60 after HDT. PR denotes partial remission, CR denotes complete remission, CRu denotes unconfirmed CR.

Bone Marrow Transplantation


from analysis, since he developed liver cirrhosis after autologous transplantation with polyclonal gammopathy. Mean hemoglobin-levels pretreatment (n ¼ 13), before HDT(n ¼ 13), 6 months (n ¼ 10) and 1 year after HDT (n ¼ 6) were 138, 110, 115 and 127 g/l, respectively. Thrombocyte counts reached normal levels by day +60 after HDT. Absolute numbers of leukocytes normalized within 30 days after HDT, with absolute numbers of granulocyte subsets not differing significantly during treatment and recovery (data not shown). Immune monitoring via flow cytometry was performed at study entry (n ¼ 10), after CHOP (n ¼ 9), after R-CHOP (n ¼ 8), before mobilization chemotherapy (n ¼ 8), before HDT (n ¼ 10), and at days +30, +60, +90, +180 (n ¼ 8) and on day +360 (n ¼ 6) (Figure 2). One patient, who developed liver cirrhosis, was not included in this analysis. Investigation of lymphocyte subsets showed reoccurrence of polyclonal CD 19 positive B cells at day +60 after HDT. With regard to rituximab treatment, B cells recovered 6 months after termination of antibody treatment (Figure 2a). Hematopoietic recovery was accompanied by an increase in absolute CD 8 cell numbers with maximum values measured at day +30 (mean: 655/ml) (Figure 2d). The number of CD 8 cells declined after this time point and reached pretreatment levels 1 year after HDT (mean at days

b

B-cells

1500 1250 1000 750

300

500

-200

-100

0

100 Time

200

300

0 -300

400

c 1500 1250

-100

0

100 Time

200

300

400

CD8 positive T-cells

1250

1000 Cells/µl

1000

750

750

500

500

250

250

0 -300

-200

d 1500

CD4 positive T-cells

Cells/µl

200

100

250

0 -300

779

NK-cells

400

cells/µl

Number cells/µl

a 1750

+180 and +360: 435/ml and 288/ml). NK-cell recovery showed a similar pattern as CD 8 T-cell kinetics reaching normal values at approximately day +180 (Figure 2b). In contrast, absolute numbers of CD 4 cells declined after HDT with the lowest numbers at around day +60 (mean 165/ml), thereafter slowly increasing, but not reaching baseline levels at day +180 (mean 221/ml) and +360 (mean 317/ml) (Figure 2c). Monitoring of immunoglobulin levels was undertaken during the entire treatment course and the first year of follow-up (study initiation (n ¼ 12), before cycle 3 (n ¼ 11), before cycle 5 (n ¼ 11), before mobilization (n ¼ 10), before HDT (n ¼ 11) and on day +30 (n ¼ 7), +60, +90 (each n ¼ 9), 180 (n ¼ 10) and +360 (n ¼ 7)). Analysis demonstrated a decrease in IgA, IgG and IgM levels, with lowest values between HDT and day +90. Mean numbers decreased below lower normal values for IgG and IgM, but a substantial portion of patients had in addition subnormal levels of IgA. A slow recovery of levels was seen for IgG levels. In contrast, IgM levels increased more rapidly shortly after HDT. However, on day +180, 5/10 patients had subnormal IgA and IgM levels and 7/10 patients had continuous subnormal IgG levels, respectively. On day +360, 2/7 patients had subnormal IgA levels, 1/7 had a subnormal IgM level and 3/7 still had not reached the lower normal level of IgG (Figure 3).

-200

-100

0

100 Time

200

300

400

0 -300

-200

-100

0

100

200

300

400

Time

Figure 2 Reconstitution of different lymphocyte subsets after HDT. Results of cytometry analysis of cellular subsets. Kinetics of mean absolute numbers of B cells (panel a), NK-cells (panel b), CD 4 T cells (panel c) and CD 8 T cells (panel d). Time 0 reflects reinfusion of stem cells after HDT. Error bars show standard error of mean. Left from day 0 the course during CHOP and R-CHOP is shown. Right to day 0 post-treatment follow-up until day +360 is demonstrated. Bone Marrow Transplantation


780 12.5

Immunglobulin levels during treatment

1.5

7.5 1.0 5.0 0.5

2.5 0.0 -300

-200 -100 0 100 200 Time, 0 = start ofHDC

300

IgA, IgM in g/l

IgG in g/l

10.0

2.0

0.0 400

Figure 3

Serum immunoglobulin levels before and post-HDT Mean levels of Ig are presented. Normal values: IgG: 7–16 g/l (dotted line), IgA: 0.7–4 g/l (dashed line), IgM: 0.4–2.3 g/l (solid line). Day 0 reflects the day of start of conditioning therapy for HDT.

Discussion New, promising agents are currently being investigated for the treatment of indolent lymphoma. Here, we present data on a patient population treated with an intensified treatment protocol including combination rituximab/ CHOP-chemotherapy followed by consolidation HDT. Overall, the combination therapy was well tolerated, with only a single episode of treatment delay caused by infection and no treatment-related mortality occurred. There was one case of unstable angina during mobilization chemotherapy, which was not considered to be related to treatment. Collection of a sufficient number of stem cells was feasible in all patients mobilized. This demonstrates sufficient stem cell reserve after pretreatment with CHOP/ R-CHOP.34 Underlining the in vivo purging potency of rituximab, analysis of LP products revealed complete clearance of contaminating B cells in all but one patients.9,23–25,35 However, this patient still is in clinical remission. Consolidation HDT had comparable side effects as seen in relapsed lymphoma patients, with hematopoietic recovery occurring promptly. We and others have shown a high rate of CMV-reactivation in patients undergoing HDT, especially with the use of purged stem cells. In this series, only one patient was CMV-seropositive and he did not show signs of reactivation. Nevertheless, careful monitoring for CMV-reactivation should be considered after combination chemo-immunotherapy.23,36–39 As knowledge about immune reconstitution after intensified treatment approaches is still limited, we carefully monitored immune reconstitution in our patients. A slow reconstitution of cellular subsets was observed. Absolute numbers of B cells were diminished up to days +120–180 after HDT, reflecting the effect of antibody treatment, as demonstrated in an earlier series of patients.23 There was a marked increase in the absolute number of CD8 T cells and NK-cells shortly after HDT, which can be related to infection during aplasia. CD4 T-cell numbers were diminished during the entire first year post-HDT. The T4/T8 ratio at day +360 was still markedly altered. Additionally, Bone Marrow Transplantation

immunoglobulin levels were reduced approximately 1 year after HDT and some patients still had not achieved normal values at this time point. Owing to immunoglobulin replacement therapy up to day +100, the true extent of the reduction in serum levels was masked. Together, these data show a disturbance in the immune system after this treatment modality, which might be in part related to rituximab.40 However, the number of infectious complications during follow-up has been low, comparable to our former experiences with HDT-related complications.23,41 Efficacy of isolated CHOP chemotherapy was low, with only two patients achieving CR and most of the patients achieving SD/MR. However, after additional R-CHOP therapy, the number of PRs and CRs increased markedly. Restaging after HDT revealed additional efficiency with a high rate of CRs. The value of consolidation HDT is underlined by the data of Howard et al. in which patients with newly diagnosed MCL were treated with R-CHOP. With an overall response rate of 96%, 28/40 patients relapsed or progressed with a median PFS of 16.6 months.42 In contrast, in our small series none of the four patients with MCL who completed HDT have relapsed to date, with a follow-up of up to 29.2 months. All patients, in whom a molecular marker could be identified, reverted to PCR negativity during treatment and 3/4 converted after introduction of rituximab, with no molecular relapse to date. This emphasizes the potential for inducing durable molecular responses.7–9,43 In this limited series of patients, demonstrating safety and feasibility of intensified treatment strategies, substantial therapy-related morbidity was seen. This has to be taken into account, when comparing OS and DFS reported in this study with standard care. Excellent results have been seen with watch-and-wait strategies, and outside of clinical studies the use of intensified treatment or upfront transplantation could not be recommended until now. However, as presented here, upfront autografting for NHL is feasible and with additional tools available should be considered for younger and/or high-risk patients. As the standard of four doses of rituximab was developed empirically, the value of extended cycles of rituximab in first line or maintenance therapy is not clear.44,45 However, the data of Ghielmini et al46 support the thesis of extended Rituximab use for the treatment of indolent lymphoma, demonstrating additional therapeutic efficacy and prolongation of PFS, especially in untreated patients, in a phase II clinical study. Additionally, whether the combination of rituximab in mobilization therapy after the achievement of MRD-negativity affects long-term outcome remains to be evaluated. Recently, data of combination therapy of rituximab and fludarabine have been published with encouraging results, reflecting the importance of defining optimal combination therapies.47 Currently, new drugs such as proteasome inhibitors, antisense oligonucleotides or vaccination strategies are being evaluated in phases I–III clinical studies.48,49 Combining these strategies after maximum intensive debulking chemotherapy seems to be an attractive approach towards controlling remaining MRD. Finally, radioimmunoconjugates have considerable potential in indolent lymphoma, especially with their ability to eliminate tumor cells even in


cell cycle arrest. Efforts are being made to identify the optimal use of these exciting modalities.

Acknowledgements We gratefully acknowledge study management by Gertrud Feldmann and the excellent technical assistance of Karola Schmidt. The Deutsche Krebshilfe supported this study, Grant Numbers 70-2427 and 70-2428.

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