Rituximab in Combination With Fludarabine Chemotherapy in Low-Grade or Follicular Lymphoma
http://www.100md.com
《临床肿瘤学》
the Roswell Park Cancer Institute, Buffalo, NY
Berlex Laboratories, Richmond
Genentech Inc, San Francisco
IDEC Pharmaceuticals, San Diego, CA
ABSTRACT
PATIENTS AND METHODS: This was an open-label, single-arm, single-center phase II study enrolling 40 patients. During the first week of the study, patients received two infusions of rituximab 375 mg/m2 administered 4 days apart. Seventy-two hours after the second infusion of rituximab, patients received the first of six cycles of fludarabine chemotherapy (25 mg/m2/d for 5 days on a 28-day cycle). Single infusions of rituximab were administered 72 hours before the second, fourth, and sixth cycles of fludarabine, and two infusions of rituximab were given 4 weeks after the last cycle of fludarabine. Treatment duration was 26 weeks.
RESULTS: An overall response rate of 90% (80% complete response rate) was achieved in the intent-to-treat population. Similar response rates were seen in treatment-nave and previously treated patients. The median duration of response has not been reached at 40+ months. The median follow-up time in this study is 44 months (range, 15 to 66 months). In patients positive for the 14;18 translocation in blood and/or marrow at enrollment, molecular remission was achieved in 88% of cases, with patients remaining negative for up to 4 years to date. Hematologic toxicity was manageable, and except for a 15% incidence of herpes simplex/zoster infections, infectious complications were rare. Nonhematologic toxicities were minimal.
CONCLUSION: Rituximab plus fludarabine was well tolerated and associated with an excellent complete response rate, including molecular remissions, in patients with low-grade or follicular lymphoma.
INTRODUCTION
Combination fludarabine, mitoxantrone, and dexamethasone (FND) has been evaluated in previously treated7,8 and untreated9 patients with indolent B-cell lymphoma. The phase II study of FND7 in recurrent or refractory indolent lymphoma demonstrated an overall response rate of 94% (47% rate of complete response [CR]), median overall survival of 34 months, and median failure-free survival (FFS) of 14 months. In the upfront setting, FND had an overall response rate of 97% (79% rate of CR), with an 84% 5-year survival rate and 41% 5-year FFS rate. Of note, 20% of upfront FND patients did not complete all of the assigned FND courses. Despite its excellent antitumor activity, the FND regimen has been associated with significant myelosuppression and an increased risk of infections, in particular opportunistic infections (eg, Pneumocystis carinii pneumonia [PCP]).
Rituximab is a chimeric anti-CD20 monoclonal antibody with significant single-agent antitumor activity against B-cell lymphoma and CLL.10-14 Rituximab's biologic activity is associated with antibody-dependent cellular cytotoxicity (ADCC), complement-mediated cytotoxicity, and direct apoptosis. Byrd et al15 have previously shown that caspase-3 and caspase-9 are activated in CLL cells after rituximab infusion and that this is associated with induction of apoptosis via the mitochondrial pathway. Fludarabine16 and other chemotherapeutic agents used in the therapy of CLL and NHL also activate similar apoptotic pathways, providing a rationale for combining fludarabine and rituximab.
In contrast to fludarabine plus chemotherapy combinations, a combination of fludarabine and rituximab (F+R) should have a reduced toxicity profile, because these agents have no significant overlapping toxicities. Fludarabine toxicity is mainly hematologic,17 resulting in cytopenias and an increased risk of infection, whereas rituximab shows predominantly infusion-related toxicities.18
Indeed, F+R demonstrates synergistic antitumor activity in vitro,19,20 and seems to act by non–cross-resistant mechanisms. Fludarabine inhibits RNA and DNA synthesis and DNA repair.3 Rituximab can deplete target B cells via activation of the host immune system and apoptotic signaling pathways.21 In addition, rituximab has been shown in vitro to sensitize drug-resistant lymphoma cell lines to cytotoxic chemotherapy.22
An early clinical trial using rituximab in combination with cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) seemed to add significant antitumor activity to CHOP alone and set a precedence for further rituximab plus chemotherapy studies. Historically, treatment of indolent lymphoma patients with CHOP alone yields overall response rates of approximately 65%.23,24 Results from the first phase II multi-institutional CHOP plus rituximab clinical trial demonstrated a 100% overall response rate with durable remissions.25-27
On the basis of single-agent activity, documented synergy, and fludarabine's better nonhematologic toxicity profile compared with CHOP, a phase II clinical trial of F+R was undertaken. The primary aims of this open-label, single-arm, single-center phase II pilot study were to evaluate the safety and efficacy of rituximab in combination with fludarabine in treatment-nave or patients who had experienced relapse with low-grade and/or follicular NHL. Secondary aims included evaluation of changes in peripheral-blood lymphocyte subsets (especially natural killer [NK] cells) and the ability to convert and maintain bcl-2 negativity (by qualitative polymerase chain reaction [PCR]) post-therapy.
PATIENTS AND METHODS
Study Design
This was a nonrandomized study. Dose and schedule of F+R is outlined in Figure 1. Fludarabine and rituximab were infused as per individual package insert recommendations. Treatment lasted up to 26 weeks.
In the event of significant treatment-associated nonhematologic toxicity (ie, National Cancer Institute Common Toxicity Criteria grade ≥ 2), treatment was delayed for 1-week increments up to a total of 3 weeks until the toxicity improved to grade ≤ 1. If postponement lasted longer than 3 weeks, treatment was discontinued. With respect to hematologic parameters, significant unexpected toxicities were seen in the initial 10 patients, leading to the following changes to the study design. First, in the event of ≥ grade 3 hematologic toxicity (unsupported with growth factors) that persists ≥ 2 consecutive weeks, the dose of fludarabine was to be reduced by 40% (25 mg/m2/d on days 1 through 3) for all subsequent cycles. Patients without hematologic recovery after a 3-week interruption/delay were to be discontinued. Second, prophylactic trimethoprim/sulfamethoxazole was discontinued because it was believed to have been contributing to therapy-associated cytopenias and, because concurrent corticosteroids were not used in the F+R regimen, the risk for PCP was considered low. Third, granulocyte colony-stimulating factor (G-CSF) support was to be limited during active therapy.
Permitted treatment support included transfusion of blood and blood products, antibiotics, antiemetics, and colony-stimulating factors (G-CSF and/or granulocyte macrophage colony-stimulating factor). No concurrent antineoplastic therapy (including radiation therapy) or treatment with corticosteroids (other than transient topical application) was permitted.
Patient Monitoring
The following evaluations and procedures were performed during the pretreatment screening period: medical history and baseline laboratory and imaging studies, serum pregnancy test (for women of childbearing potential), bilateral bone marrow aspirates and core biopsies for assessment of the extent of disease and bcl-2, and lymph node or bone marrow aspiration sample demonstrating CD20 expression of tumor cells. Evaluations performed during and after treatment included physical examination, CBC counts, serum chemistry, imaging studies for disease assessment, peripheral-blood four-color flow cytometry evaluation of total lymphocyte subpopulations, quantitative serum immunoglobulins, peripheral-blood and bone marrow aspirate samples for bcl-2 analysis by PCR assay in patients testing positive pretreatment. The PCR assay used was as previously described by Czuczman et al.26 The intent-to-treat (ITT) population included all enrolled patients.
Follow-Up
During the 4-year follow-up period, imaging studies for response assessment and full laboratory parameters were performed at midtreatment, at completion of therapy, every 4 months for the first year, and then every 6 months thereafter. Patients who are in continuous response after 4 years will continue to be monitored for recurrent disease on an every-6-months follow-up schedule.
Response Criteria and Data Analysis
Determination of CR, unconfirmed CR, partial response (PR), stable disease, and progressive disease were defined using a modification of the International Workshop NHL Response Criteria published by Cheson et al.28 The only modifications were that a response needed to be confirmed by repeat measurement (eg, computed tomography [CT] scans 28 or more days later) and that gallium-avid disease that converted to negative on subsequent whole-body gallium scanning using single-photon emission computed tomography was accepted as evidence of complete remission. Typically, changes in bidimensional disease was determined by perpendicular measurements of sentinel lesions on serial CT scans.
The primary measure of efficacy for this study was CR rate at 30 weeks (1 month after completion of study therapy). Secondary efficacy parameters included PR rate at 30 weeks, time to progression (TTP), duration of response, survival, depletion of CD20+ cells from peripheral blood, and molecular clearing of bcl-2 (14;18 chromosomal translocation)–positive cells from blood and/or marrow aspirate samples.
Statistical Methods
A total of 40 patients were treated on this protocol. The ITT population included all enrolled patients. As a result of unexpected hematologic toxicities seen in the first 10 patients enrolled onto the study (subgroup 1), treatment modifications were instituted by protocol amendment as described above in Study Design above. The subsequent 30 patients enrolled onto the study were designated as subgroup 2. Most of the subsequent statistical analysis is stratified to account for this change in study design. Overall survival was defined as the time from the date of first treatment to the date of last follow-up examination (censored) or the date of death (event) from any cause. TTP was defined as the date of first treatment to the date of documented disease progression or death from NHL (event) or the date of last follow-up examination (censored). Duration of response was defined as the date of first CR or PR to the date of disease progression (event) or the date of last follow-up examination (censored). The Kaplan-Meier method was used to derive survival probabilities. Follow-up time was calculated using the potential follow-up method.29 The log-rank test was used to compare differences in survival times between patient subgroups. The subgroup 1 patient sample did not have enough events for survival or TTP analyses. Cox proportional hazards regression analysis was used to model the association of prognostic factors with time to disease progression, using an ITT approach. Because of the small sample size, multivariate analysis was not feasible. Fisher's exact test was used to measure the degree of association between having received a growth factor (yes or no) and neutropenia grade and to measure the association of pretreatment bone marrow status with grade of toxicity. Fisher's exact test was also used to test the associations of disease stage, sex, disease type, and International Prognostic Index (IPI) score 0 or 1 versus ≥ 2 with patient treatment subgroup (the first 10 patients v the remaining 30 patients). The Wilcoxon rank sum test was used to compare toxicity grade between the patient subgroups and to test the association between the percentage of pretreatment bone marrow with degree of neutropenia. Patient age was also compared between the treatment subgroups using the rank-sum test. In all statistical tests, the level of significance was set at 5%.
In addition, evaluation of changes in peripheral-blood lymphocyte subsets (in particular, B cells, T cells, NK cells, and activated NK cells [ie, NK cells with CD122 coexpression]) in peripheral blood after F+R treatment were performed. All statistical calculations for this analysis were performed using procedures from SAS/STAT software (The SAS System 8.1; SAS Institute, Cary, NC). The distributions of percentage changes in cell counts were analyzed using both the sign test and the Wilcoxon signed ranks test. Both the Kolmogorov-Smirnov test and the Cramer-von Mises tests were used to test the maximum likelihood fit of the lognormal distribution to the observed distribution of pretreatment. Box plots of the observed distributions of cell counts stratified by time were prepared using a logarithmic scale on the vertical axis. Normal values for B cells, NK cells, and activated NK cells were determined from a population of 20 healthy individuals and represented by three numbers: the lower 10th percentile, the median, and the upper 90th percentile.
Demographics
Of 40 patients, 38 patients were assessable for response. Although achieving transient objective responses (ie, PR, unconfirmed CR) halfway through therapy, two patients in subgroup 2 were considered nonassessable for response because the next CT scan demonstrated evidence of progressive disease (ie, no confirmatory scans for response).
Patient baseline clinical characteristics are listed in Table 1. All patients had advanced stage (III, IV) disease, and the majority were previously untreated (68%). The majority of patients (65%) were IWFB. The median patient age was 53 years (range, 40 to 77 years). The most frequent extranodal site of disease was bone marrow (65%). Half the patients had IPI scores of 0 or 1, and the others had IPI scores of ≥ 2. Fewer patients in subgroup 1 had a higher score of ≥ 2 (20%) compared with subgroup 2, in which 60% had a higher score (P = .03). There were no significant associations between patient subgroup and any of the other baseline clinical characteristics. Clinical factors were balanced between the newly diagnosed patients and patients who experienced relapse (results not shown).
RESULTS
The probability of overall survival in an ITT basic for all 40 patients at 50 months was equal to 0.80 (not shown). The overall survival rate among the subset of patients who completed all therapy (n = 34) was slightly higher, being equal to 0.83.
There were no statistically significant differences in survival times between the newly diagnosed patients and the patients who experienced relapse. The median survival time was not achieved in either group. There were also no significant differences in clinical response between the previously treated versus previously untreated patients or follicular versus nonfollicular patients. Among the newly diagnosed patients, 22 (81%) of 27 patients achieved a CR, and in the previously treated patients, 10 (77%) of 13 patients achieved a CR. Two (7%) of 27 of the newly diagnosed patients had a PR, compared with two (15%) of 13 patients in the previously treated group. Three patients (11%) in the newly diagnosed group and one patient (8%) in the relapsed group demonstrated progressive disease during therapy.
The relationships of histology, IPI score, disease type, sex, and age (> 60 years v ≤ 60 years) versus TTP were tested in univariate Cox proportional hazards regression analysis, and results are shown in Table 3. Age was significantly related to TTP, whereby the older (> 60 years) patients had more than 2.5 times increased risk of disease progression compared with the younger patients (relative risk = 2.59; P = .04). Trends were seen in the relationships of disease type, IPI score, and histology to progressive disease. Patients with relapsed disease had a 68% reduced risk of disease progression compared with those who were newly diagnosed (relative risk = 0.32; P = .07). Those with an IPI score ≥ 2 had more than twice the increase in risk of disease progression compared with those with an IPI score ≤ 1 (relative risk = 2.37; P = .08). There was no association found between histologic subtype and time to disease progression in this study sample.
Toxicity
Most patients had either no or only grade 1 or 2 anemia or thrombocytopenia (Table 2). However, there were eight patients (80%) in subgroup 1 with grade 3/4 neutropenia, and 21 patients (72%) in subgroup 2 with grade 3 or 4 neutropenia.
Overall, grade 3 or 4 neutropenia was transient and reversible in subgroup 2 patients. Whereas 70% of patients in subgroup 1 required G-CSF support, only 24% of the patients in subgroup 2 received G-CSF support. In fact, two of the total seven patients in subgroup 2 who received transient G-CSF did so only after completion of all scheduled study drugs. In subgroup 2 patients, transient therapy delay was required in nine patients: starting with the third cycle of therapy in four patients, the fourth cycle in two patients, the fifth cycle in one patient, and the sixth cycle in two patients. Excluding the single patient removed from study at midtherapy secondary to prolonged cytopenia in subgroup 2, three patients required a 40% fludarabine dose reduction: one patient starting at cycle 4 and two patients with cycle 6 only.
Additional data, not documented in Table 2, included infectious complications and/or hospitalizations, which were limited to the following: staphylococcal or culture-negative Mediport infections (n = 3), neutropenic fever requiring hospitalization (n = 4), primary or secondary herpes simplex/zoster skin infections (n = 6), and recurrent urinary tract infection (n = 1). There were no differences in infectious complications noted between patients in subgroup 1 versus subgroup 2. Overall, infectious complications (especially no occurrence of PCP or other serious opportunistic infection) seen in the ITT group seemed to be similar to that expected in a similar population treated with fludarabine alone. However, acyclovir prophylaxis was subsequently prescribed to all treated patients for 6 to 12 months after completion of therapy because of the relatively high incidence of herpes infections (six of 40 patients; 15%) believed to be secondary to T-cell depletion from fludarabine. No patient receiving acyclovir prophylaxis developed herpes infection.
Typical first-dose rituximab infusional toxicities were seen in our study population, as has been previously described in other trials.11-14 Fludarabine was well tolerated, and no acute infusion reactions were seen. Intermittent grade 1 to 2 fludarabine-associated nausea/vomiting and liver function test abnormalities were seen in one patient, which resolved after completion of therapy. One case of interstitial pneumonitis believed to be secondary to fludarabine was seen in a single patient in subgroup 1 that necessitated taking them off of study after fludarabine cycle 5. This patient's symptoms resolved quickly with initiation of corticosteroids.
Quantitative serum immunoglobulins (IgG, IgA, and IgM) were obtained pretherapy and monitored during and after treatment completion. In the 34 patients who completed all therapy, 79% (27 of 34 patients) demonstrated stable (n = 24) to improved (n = 3) quantitative immunoglobulin levels.
Factors Associated With Grade 3 or 4 Neutropenia
There was a highly significant association of IPI score with neutropenia grade (Table 4), in which patients with an IPI score of ≥ 2 were 22 times more likely to have grade 3 or 4 neutropenia as compared with those with a low IPI score (P = .009). Histology and marrow involvement were not found to be associated with grade of neutropenia observed.
Molecular bcl-2 Monitoring by PCR
There were 16 patients who were positive for bcl-2 by PCR in peripheral blood ± bone marrow aspirate before therapy. Among these, 87% (13 of 15 patients) converted to bcl-2–negative after treatment in peripheral blood. Among the 16 patients who were bcl-2–positive pretreatment in bone marrow aspiration samples, 88% (14 of 16 patients) became negative for bcl-2 after therapy. In those patients converting to negative, repeat bcl-2 studies were performed on a yearly basis. Serial bcl-2 assays have remained negative for up to 4 years in patients to date.
Flow Cytometric Monitoring of Lymphocyte Subsets
Multiparameter flow cytometric evaluation of changes in peripheral-blood lymphocyte subsets were evaluated during and after F+R immunochemotherapy. Changes in T cells, B cells, NK cells, and activated NK cells during the post-treatment period are shown in Figures 4 through 7. The upper and lower part of each box bound the interquartile distance (ie, the 25% and the 75% quartile); the dark line in the box is the median; the "+" is the mean; and the thin lines (whiskers) emanating from the top and bottom of the box extend as far as the most extreme values if they are no more than 1.5 x the interquartile distance from the top (or bottom) of the box. Values more extreme than this are represented as individual boxes, and the number next to an extreme point is the patient's study enrollment number. A line connects the median values at each time. Note that the width of each box varies with the number of patients available at the corresponding time. The vertical axis (ie, the cell count) is in log coordinates. As previously described, follow-up visits 1, 2, and 3 are 4 months apart, whereas each follow-up visit after follow-up visit 3 are at 6-month intervals. More marked decreases in CD3+ T cell and CD19+ B-cell lymphocyte subsets were seen after F+R therapy, as compared with changes seen in NK and activated NK cell subsets (Figs 4 through 7).
DISCUSSION
The first rituximab combination immunochemotherapy trial demonstrated that rituximab can be safely given with standard CHOP chemotherapy,25 with a resultant 100% overall response rate (57% rate of CR), in 38 assessable patients with a durable remission duration and the ability to molecularly clear blood and/or marrow of bcl-2–positive cells by PCR testing. The initial rationale behind our decision to evaluate fludarabine and rituximab was the hope that this combination might achieve similar efficacy as R-CHOP, but with less toxicity. Furthermore, because of the significant number of older patients with comorbid medical problems, a less aggressive, non–anthracycline-containing combination immunochemotherapy such as F+R could prove to be a valuable addition to current therapeutic options. In addition, basic research has demonstrated that fludarabine and rituximab have synergistic antitumor activity.19,20
Although late onset of response has been a feature of some studies using rituximab monotherapy,32 F+R induced a relatively rapid rate of response. In the 34 patients who completed therapy, 82% (28 of 34 patients) demonstrated maximal or near maximal nodal responses by midtherapy. Although B-cell and T-cell subsets were overall depleted by this regimen, NK cells (including activated NK cells) were relatively preserved. The first report of the potential sparing effect of infusional fludarabine on human T-10 positive (ie, probably NK) cells determined by flow cytometric evaluation of peripheral-blood mononuclear cells postchemotherapy was conducted by Boldt et al in 1984.33 Other researchers not only have corroborated this finding of a relative sparing of NK cells, but have also found that fludarabine may actually stimulate NK-cell lytic activity.34-36 This relative sparing, and possible activation, of NK cells by fludarabine may significantly contribute to the excellent antitumor activity demonstrated here by the F+R combination. NK cells likely play an important role as a major effector cell used in the destruction of rituximab-bound tumor cells via ADCC.37 Theoretically, it is possible that the addition of other agents to F+R (eg, drugs that deplete NK cells, corticosteroids, and so on) that may decrease ADCC and complement-mediated cytotoxicity activity may have a potentially negative impact on overall antitumor activity. The relative sparing of NK cells, along with the preservation of mean quantitative immunoglobulin levels in the majority of our patients, may explain the low incidence of infectious complications seen in our study. However, acyclovir prophylaxis is recommended in patients receiving F+R combination therapy because of the relatively high incidence of herpes infections seen.
Discontinuations resulting from cytopenia or infectious complications were rare in this study, and dose reductions of fludarabine occurred typically late in the course of therapy. Furthermore, transient therapy delays were necessary in less than one third of the patients in subgroup 2. Review of several previously published reports of fludarabine monotherapy38-42 in patients with either previously treated or untreated CLL and/or indolent B-cell lymphoma documented that myelosuppression is the major adverse effect associated with this drug, dermatomal herpes/zoster infection is not uncommon, and that nonhematologic toxicities are typically mild. In an article by Keating et al,38 post-therapy CD4 and CD8 lymphocyte counts are substantially decreased and seem to slowly increase, but do not reach median pretreatment values at more than 24 months. Results from our F+R trial are consistent with these previously published observations. Two phase II trials of combination F+R in CLL have recently been published.43,44 In the German trial,43 31 patients (11 previously untreated) received four cycles of fludarabine and four infusions of rituximab. An overall response rate of 87% (33% rate of CR; 55% rate of PR) and a median duration of response of 75 weeks were reported. The other trial was a randomized phase II Cancer and Leukemia Group B study (CALGB 9712)44 of fludarabine with concurrent versus sequential rituximab in previously untreated B-CLL patients. Concurrent rituximab and fludarabine (six cycles) resulted in an overall response rate of 90% (47% rate of CR; 43% rate of PR). This overall and CR rate was lower in the sequential arm (overall response rate = 77%; CR = 28%). At a median follow-up time of 23 months, median response duration and survival had not been reached for either arm. Comparing both arms, the concurrent arm experienced an increased incidence of grade 3 or 4 neutropenia of uncertain etiology. Notably, this finding was not accompanied by any increased risk of infections in the concurrent arm. Factors that could potentially be associated with an increased risk of developing grade 3 or 4 neutropenia were evaluated in our F+R study population. Of interest, the presence of marrow involvement (and potential resultant decrease in marrow reserve) was not found to be associated with grade 3 or 4 neutropenia. However, patients with IPI ≥ 2 were 22-fold more likely to have grade 3 or 4 neutropenia compared with patients with IPI of ≤ 1 (P = .009). How this finding is correlated to an increased risk of neutropenia is not clear and will require further investigation.
Statistical analysis of clinical characteristics versus TTP were evaluated in our F+R patients. Older patients (ie, older than 60 years; n = 12 of 40 patients) demonstrated more than 2.5 times the risk of experiencing relapse earlier than younger patients (ie, ≤ 60 years; n = 28 of 40 patients; P = .04). Actual median TTP was 26.3 months (range, 10.6 to 54+ months) versus not reached after a median follow-up time of 44 months in the older versus younger group, respectively. When comparing these two groups, it was not surprising to find an IPI score of ≥ 2 in 100% of older compared with 29% of younger patients. Whether these TTP results represent true biologic differences in indolent B-cell NHL based on age and subsequent responsiveness to F+R versus a limitation of univariate analysis in a relatively small number of patients is unclear. This question may be answered by prospectively analyzing a larger number of patients undergoing an F+R–containing regimen in future trials and/or evaluation of gene expression profiles by microarray analysis between older and younger patients receiving F+R. Nevertheless, F+R seems to be at least comparable in antitumor activity to other fludarabine combination chemotherapy regimens (eg, FND, F+C), but potentially less toxic and better tolerated in an older patient population.
In conclusion, we have demonstrated that concurrent F+R immunochemotherapy resulted in excellent antitumor activity and is well tolerated in previously treated and untreated patients with indolent B-cell lymphoma (overall response rate = 90% in ITT group; 80% rate of CR, 10% rate of PR; 88% bcl-2 conversion rate by PCR), with median TTP not reached at 44+ months. Molecular remissions up to 4 years have been demonstrated to date. This is encouraging because other work has indicated that patients who achieve molecular remission during the first year of treatment have a significantly longer FFS at 4 years compared with those who do not (76% v 38%; P < .001).45 Although the majority of patients experienced transient, reversible grade 3 or 4 neutropenia, this was not associated with a higher-than-expected infection risk. The low infection rate in our study population is likely multifactorial: minimal to no mucosal membrane breakdown associated with F+R, preservation of serum immunoglobulin levels in approximately 80% of patients; and relative sparing and possible activation of NK cells related to fludarabine therapy.
Secondary to the documented hematologic toxicity and the finding that more than 80% of patients had experienced maximal or near maximal nodal response by midtherapy using the current F+R schedule, a trial of reduced fludarabine (eg, possibly day 1 rituximab plus days 3 through 5 of fludarabine every 28 days for six to eight cycles) is warranted and should potentially maintain excellent activity with less hematologic toxicity. Our data support use of an F+R arm in future phase III studies comparing rituximab chemotherapy regimens to each other in an attempt to define an optimal therapy for patients with indolent B-cell lymphoma.
Authors' Disclosures of Potential Conflicts of Interest
NOTES
Supported by Genentech Inc, IDEC Pharmaceuticals, and Berlex Laboratories.
Authors' disclosures of potential conflicts of interest are found at the end of this article.
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31. Solal-Celigny P, Brice P, Brousse N, et al: Phase II trial of fludarabine monophosphate as first-line treatment in patients with advanced follicular lymphoma: A multicenter study by the Groupe d'Etude des Lymphomes de l'Adulte. J Clin Oncol 14:514-519, 1996
32. Colombat P, Salles G, Brousse N, et al: Rituximab (anti-CD20 monoclonal antibody) as single first-line therapy for patients with follicular lymphoma with a low tumor burden: Clinical and molecular evaluation. Blood 97:101-106, 2001
33. Boldt DH, Von Hoff DD, Kuhn JG, et al: Effects on human peripheral lymphocytes of in vivo administration of 9-beta-D-arabinofuranosyl-2-fluoroadenine-5'-monophosphate (NSC 312887), a new purine antimetabolite. Cancer Res 44:4661-4666, 1984
34. Priebe T, Platsoucas CD, Nelson JA: Adenosine receptors and modulation of natural killer cell activity by purine nucleosides. Cancer Res 50:4328-4331, 1990
35. Priebe T, Ruiz L, Nelson JA: Role of natural killer cells in the modulation of primary antibody production by purine nucleosides and their analogs. Cell Immunol 130:513-519, 1990
36. Robertson LE, Denny AW, Huh YO, et al: Natural killer cell activity in chronic lymphocytic leukemia patients treated with fludarabine. Cancer Chemother Pharmacol 37:445-450, 1996
37. Janakiraman N, MP, White C, et al: Rituximab: Correlation between effector cells and clinical activity in NHL. Blood 92:337a, 1998 (abstr 1384)
38. Keating MJ, O'Brien S, Lerner S, et al: Long-term follow-up of patients with chronic lymphocytic leukemia (CLL) receiving fludarabine regimens as initial therapy. Blood 92:1165-1171, 1998
39. Zinzani PL, Lauria F, Rondelli D, et al: Fludarabine: An active agent in the treatment of previously-treated and untreated low-grade non-Hodgkin's lymphoma. Ann Oncol 4:575-578, 1993
40. Ross SR, McTavish D, Faulds D: Fludarabine: A review of its pharmacological properties and therapeutic potential in malignancy. Drugs 45:737-759, 1993
41. Pigaditou A, Rohatiner AZ, Whelan JS, et al: Fludarabine in low-grade lymphoma. Semin Oncol 20:24-27, 1993 (suppl 7)
42. Wijermans PW, Gerrits WB, Haak HL: Severe immunodeficiency in patients treated with fludarabine monophosphate. Eur J Haematol 50:292-296, 1993
43. Schulz H, Klein SK, Rehwald U, et al: Phase 2 study of a combined immunochemotherapy using rituximab and fludarabine in patients with chronic lymphocytic leukemia. Blood 100:3115-3120, 2002
44. Byrd JC, Peterson BL, Morrison VA, et al: Randomized phase 2 study of fludarabine with concurrent versus sequential treatment with rituximab in symptomatic, untreated patients with B-cell chronic lymphocytic leukemia: Results from Cancer and Leukemia Group B 9712 (CALGB 9712). Blood 101:6-14, 2003
45. Lopez-Guillermo A, Cabanillas F, McLaughlin P, et al: The clinical significance of molecular response in indolent follicular lymphomas. Blood 91:2955-2960, 1998(M.S. Czuczman, A. Koryzna)
Berlex Laboratories, Richmond
Genentech Inc, San Francisco
IDEC Pharmaceuticals, San Diego, CA
ABSTRACT
PATIENTS AND METHODS: This was an open-label, single-arm, single-center phase II study enrolling 40 patients. During the first week of the study, patients received two infusions of rituximab 375 mg/m2 administered 4 days apart. Seventy-two hours after the second infusion of rituximab, patients received the first of six cycles of fludarabine chemotherapy (25 mg/m2/d for 5 days on a 28-day cycle). Single infusions of rituximab were administered 72 hours before the second, fourth, and sixth cycles of fludarabine, and two infusions of rituximab were given 4 weeks after the last cycle of fludarabine. Treatment duration was 26 weeks.
RESULTS: An overall response rate of 90% (80% complete response rate) was achieved in the intent-to-treat population. Similar response rates were seen in treatment-nave and previously treated patients. The median duration of response has not been reached at 40+ months. The median follow-up time in this study is 44 months (range, 15 to 66 months). In patients positive for the 14;18 translocation in blood and/or marrow at enrollment, molecular remission was achieved in 88% of cases, with patients remaining negative for up to 4 years to date. Hematologic toxicity was manageable, and except for a 15% incidence of herpes simplex/zoster infections, infectious complications were rare. Nonhematologic toxicities were minimal.
CONCLUSION: Rituximab plus fludarabine was well tolerated and associated with an excellent complete response rate, including molecular remissions, in patients with low-grade or follicular lymphoma.
INTRODUCTION
Combination fludarabine, mitoxantrone, and dexamethasone (FND) has been evaluated in previously treated7,8 and untreated9 patients with indolent B-cell lymphoma. The phase II study of FND7 in recurrent or refractory indolent lymphoma demonstrated an overall response rate of 94% (47% rate of complete response [CR]), median overall survival of 34 months, and median failure-free survival (FFS) of 14 months. In the upfront setting, FND had an overall response rate of 97% (79% rate of CR), with an 84% 5-year survival rate and 41% 5-year FFS rate. Of note, 20% of upfront FND patients did not complete all of the assigned FND courses. Despite its excellent antitumor activity, the FND regimen has been associated with significant myelosuppression and an increased risk of infections, in particular opportunistic infections (eg, Pneumocystis carinii pneumonia [PCP]).
Rituximab is a chimeric anti-CD20 monoclonal antibody with significant single-agent antitumor activity against B-cell lymphoma and CLL.10-14 Rituximab's biologic activity is associated with antibody-dependent cellular cytotoxicity (ADCC), complement-mediated cytotoxicity, and direct apoptosis. Byrd et al15 have previously shown that caspase-3 and caspase-9 are activated in CLL cells after rituximab infusion and that this is associated with induction of apoptosis via the mitochondrial pathway. Fludarabine16 and other chemotherapeutic agents used in the therapy of CLL and NHL also activate similar apoptotic pathways, providing a rationale for combining fludarabine and rituximab.
In contrast to fludarabine plus chemotherapy combinations, a combination of fludarabine and rituximab (F+R) should have a reduced toxicity profile, because these agents have no significant overlapping toxicities. Fludarabine toxicity is mainly hematologic,17 resulting in cytopenias and an increased risk of infection, whereas rituximab shows predominantly infusion-related toxicities.18
Indeed, F+R demonstrates synergistic antitumor activity in vitro,19,20 and seems to act by non–cross-resistant mechanisms. Fludarabine inhibits RNA and DNA synthesis and DNA repair.3 Rituximab can deplete target B cells via activation of the host immune system and apoptotic signaling pathways.21 In addition, rituximab has been shown in vitro to sensitize drug-resistant lymphoma cell lines to cytotoxic chemotherapy.22
An early clinical trial using rituximab in combination with cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) seemed to add significant antitumor activity to CHOP alone and set a precedence for further rituximab plus chemotherapy studies. Historically, treatment of indolent lymphoma patients with CHOP alone yields overall response rates of approximately 65%.23,24 Results from the first phase II multi-institutional CHOP plus rituximab clinical trial demonstrated a 100% overall response rate with durable remissions.25-27
On the basis of single-agent activity, documented synergy, and fludarabine's better nonhematologic toxicity profile compared with CHOP, a phase II clinical trial of F+R was undertaken. The primary aims of this open-label, single-arm, single-center phase II pilot study were to evaluate the safety and efficacy of rituximab in combination with fludarabine in treatment-nave or patients who had experienced relapse with low-grade and/or follicular NHL. Secondary aims included evaluation of changes in peripheral-blood lymphocyte subsets (especially natural killer [NK] cells) and the ability to convert and maintain bcl-2 negativity (by qualitative polymerase chain reaction [PCR]) post-therapy.
PATIENTS AND METHODS
Study Design
This was a nonrandomized study. Dose and schedule of F+R is outlined in Figure 1. Fludarabine and rituximab were infused as per individual package insert recommendations. Treatment lasted up to 26 weeks.
In the event of significant treatment-associated nonhematologic toxicity (ie, National Cancer Institute Common Toxicity Criteria grade ≥ 2), treatment was delayed for 1-week increments up to a total of 3 weeks until the toxicity improved to grade ≤ 1. If postponement lasted longer than 3 weeks, treatment was discontinued. With respect to hematologic parameters, significant unexpected toxicities were seen in the initial 10 patients, leading to the following changes to the study design. First, in the event of ≥ grade 3 hematologic toxicity (unsupported with growth factors) that persists ≥ 2 consecutive weeks, the dose of fludarabine was to be reduced by 40% (25 mg/m2/d on days 1 through 3) for all subsequent cycles. Patients without hematologic recovery after a 3-week interruption/delay were to be discontinued. Second, prophylactic trimethoprim/sulfamethoxazole was discontinued because it was believed to have been contributing to therapy-associated cytopenias and, because concurrent corticosteroids were not used in the F+R regimen, the risk for PCP was considered low. Third, granulocyte colony-stimulating factor (G-CSF) support was to be limited during active therapy.
Permitted treatment support included transfusion of blood and blood products, antibiotics, antiemetics, and colony-stimulating factors (G-CSF and/or granulocyte macrophage colony-stimulating factor). No concurrent antineoplastic therapy (including radiation therapy) or treatment with corticosteroids (other than transient topical application) was permitted.
Patient Monitoring
The following evaluations and procedures were performed during the pretreatment screening period: medical history and baseline laboratory and imaging studies, serum pregnancy test (for women of childbearing potential), bilateral bone marrow aspirates and core biopsies for assessment of the extent of disease and bcl-2, and lymph node or bone marrow aspiration sample demonstrating CD20 expression of tumor cells. Evaluations performed during and after treatment included physical examination, CBC counts, serum chemistry, imaging studies for disease assessment, peripheral-blood four-color flow cytometry evaluation of total lymphocyte subpopulations, quantitative serum immunoglobulins, peripheral-blood and bone marrow aspirate samples for bcl-2 analysis by PCR assay in patients testing positive pretreatment. The PCR assay used was as previously described by Czuczman et al.26 The intent-to-treat (ITT) population included all enrolled patients.
Follow-Up
During the 4-year follow-up period, imaging studies for response assessment and full laboratory parameters were performed at midtreatment, at completion of therapy, every 4 months for the first year, and then every 6 months thereafter. Patients who are in continuous response after 4 years will continue to be monitored for recurrent disease on an every-6-months follow-up schedule.
Response Criteria and Data Analysis
Determination of CR, unconfirmed CR, partial response (PR), stable disease, and progressive disease were defined using a modification of the International Workshop NHL Response Criteria published by Cheson et al.28 The only modifications were that a response needed to be confirmed by repeat measurement (eg, computed tomography [CT] scans 28 or more days later) and that gallium-avid disease that converted to negative on subsequent whole-body gallium scanning using single-photon emission computed tomography was accepted as evidence of complete remission. Typically, changes in bidimensional disease was determined by perpendicular measurements of sentinel lesions on serial CT scans.
The primary measure of efficacy for this study was CR rate at 30 weeks (1 month after completion of study therapy). Secondary efficacy parameters included PR rate at 30 weeks, time to progression (TTP), duration of response, survival, depletion of CD20+ cells from peripheral blood, and molecular clearing of bcl-2 (14;18 chromosomal translocation)–positive cells from blood and/or marrow aspirate samples.
Statistical Methods
A total of 40 patients were treated on this protocol. The ITT population included all enrolled patients. As a result of unexpected hematologic toxicities seen in the first 10 patients enrolled onto the study (subgroup 1), treatment modifications were instituted by protocol amendment as described above in Study Design above. The subsequent 30 patients enrolled onto the study were designated as subgroup 2. Most of the subsequent statistical analysis is stratified to account for this change in study design. Overall survival was defined as the time from the date of first treatment to the date of last follow-up examination (censored) or the date of death (event) from any cause. TTP was defined as the date of first treatment to the date of documented disease progression or death from NHL (event) or the date of last follow-up examination (censored). Duration of response was defined as the date of first CR or PR to the date of disease progression (event) or the date of last follow-up examination (censored). The Kaplan-Meier method was used to derive survival probabilities. Follow-up time was calculated using the potential follow-up method.29 The log-rank test was used to compare differences in survival times between patient subgroups. The subgroup 1 patient sample did not have enough events for survival or TTP analyses. Cox proportional hazards regression analysis was used to model the association of prognostic factors with time to disease progression, using an ITT approach. Because of the small sample size, multivariate analysis was not feasible. Fisher's exact test was used to measure the degree of association between having received a growth factor (yes or no) and neutropenia grade and to measure the association of pretreatment bone marrow status with grade of toxicity. Fisher's exact test was also used to test the associations of disease stage, sex, disease type, and International Prognostic Index (IPI) score 0 or 1 versus ≥ 2 with patient treatment subgroup (the first 10 patients v the remaining 30 patients). The Wilcoxon rank sum test was used to compare toxicity grade between the patient subgroups and to test the association between the percentage of pretreatment bone marrow with degree of neutropenia. Patient age was also compared between the treatment subgroups using the rank-sum test. In all statistical tests, the level of significance was set at 5%.
In addition, evaluation of changes in peripheral-blood lymphocyte subsets (in particular, B cells, T cells, NK cells, and activated NK cells [ie, NK cells with CD122 coexpression]) in peripheral blood after F+R treatment were performed. All statistical calculations for this analysis were performed using procedures from SAS/STAT software (The SAS System 8.1; SAS Institute, Cary, NC). The distributions of percentage changes in cell counts were analyzed using both the sign test and the Wilcoxon signed ranks test. Both the Kolmogorov-Smirnov test and the Cramer-von Mises tests were used to test the maximum likelihood fit of the lognormal distribution to the observed distribution of pretreatment. Box plots of the observed distributions of cell counts stratified by time were prepared using a logarithmic scale on the vertical axis. Normal values for B cells, NK cells, and activated NK cells were determined from a population of 20 healthy individuals and represented by three numbers: the lower 10th percentile, the median, and the upper 90th percentile.
Demographics
Of 40 patients, 38 patients were assessable for response. Although achieving transient objective responses (ie, PR, unconfirmed CR) halfway through therapy, two patients in subgroup 2 were considered nonassessable for response because the next CT scan demonstrated evidence of progressive disease (ie, no confirmatory scans for response).
Patient baseline clinical characteristics are listed in Table 1. All patients had advanced stage (III, IV) disease, and the majority were previously untreated (68%). The majority of patients (65%) were IWFB. The median patient age was 53 years (range, 40 to 77 years). The most frequent extranodal site of disease was bone marrow (65%). Half the patients had IPI scores of 0 or 1, and the others had IPI scores of ≥ 2. Fewer patients in subgroup 1 had a higher score of ≥ 2 (20%) compared with subgroup 2, in which 60% had a higher score (P = .03). There were no significant associations between patient subgroup and any of the other baseline clinical characteristics. Clinical factors were balanced between the newly diagnosed patients and patients who experienced relapse (results not shown).
RESULTS
The probability of overall survival in an ITT basic for all 40 patients at 50 months was equal to 0.80 (not shown). The overall survival rate among the subset of patients who completed all therapy (n = 34) was slightly higher, being equal to 0.83.
There were no statistically significant differences in survival times between the newly diagnosed patients and the patients who experienced relapse. The median survival time was not achieved in either group. There were also no significant differences in clinical response between the previously treated versus previously untreated patients or follicular versus nonfollicular patients. Among the newly diagnosed patients, 22 (81%) of 27 patients achieved a CR, and in the previously treated patients, 10 (77%) of 13 patients achieved a CR. Two (7%) of 27 of the newly diagnosed patients had a PR, compared with two (15%) of 13 patients in the previously treated group. Three patients (11%) in the newly diagnosed group and one patient (8%) in the relapsed group demonstrated progressive disease during therapy.
The relationships of histology, IPI score, disease type, sex, and age (> 60 years v ≤ 60 years) versus TTP were tested in univariate Cox proportional hazards regression analysis, and results are shown in Table 3. Age was significantly related to TTP, whereby the older (> 60 years) patients had more than 2.5 times increased risk of disease progression compared with the younger patients (relative risk = 2.59; P = .04). Trends were seen in the relationships of disease type, IPI score, and histology to progressive disease. Patients with relapsed disease had a 68% reduced risk of disease progression compared with those who were newly diagnosed (relative risk = 0.32; P = .07). Those with an IPI score ≥ 2 had more than twice the increase in risk of disease progression compared with those with an IPI score ≤ 1 (relative risk = 2.37; P = .08). There was no association found between histologic subtype and time to disease progression in this study sample.
Toxicity
Most patients had either no or only grade 1 or 2 anemia or thrombocytopenia (Table 2). However, there were eight patients (80%) in subgroup 1 with grade 3/4 neutropenia, and 21 patients (72%) in subgroup 2 with grade 3 or 4 neutropenia.
Overall, grade 3 or 4 neutropenia was transient and reversible in subgroup 2 patients. Whereas 70% of patients in subgroup 1 required G-CSF support, only 24% of the patients in subgroup 2 received G-CSF support. In fact, two of the total seven patients in subgroup 2 who received transient G-CSF did so only after completion of all scheduled study drugs. In subgroup 2 patients, transient therapy delay was required in nine patients: starting with the third cycle of therapy in four patients, the fourth cycle in two patients, the fifth cycle in one patient, and the sixth cycle in two patients. Excluding the single patient removed from study at midtherapy secondary to prolonged cytopenia in subgroup 2, three patients required a 40% fludarabine dose reduction: one patient starting at cycle 4 and two patients with cycle 6 only.
Additional data, not documented in Table 2, included infectious complications and/or hospitalizations, which were limited to the following: staphylococcal or culture-negative Mediport infections (n = 3), neutropenic fever requiring hospitalization (n = 4), primary or secondary herpes simplex/zoster skin infections (n = 6), and recurrent urinary tract infection (n = 1). There were no differences in infectious complications noted between patients in subgroup 1 versus subgroup 2. Overall, infectious complications (especially no occurrence of PCP or other serious opportunistic infection) seen in the ITT group seemed to be similar to that expected in a similar population treated with fludarabine alone. However, acyclovir prophylaxis was subsequently prescribed to all treated patients for 6 to 12 months after completion of therapy because of the relatively high incidence of herpes infections (six of 40 patients; 15%) believed to be secondary to T-cell depletion from fludarabine. No patient receiving acyclovir prophylaxis developed herpes infection.
Typical first-dose rituximab infusional toxicities were seen in our study population, as has been previously described in other trials.11-14 Fludarabine was well tolerated, and no acute infusion reactions were seen. Intermittent grade 1 to 2 fludarabine-associated nausea/vomiting and liver function test abnormalities were seen in one patient, which resolved after completion of therapy. One case of interstitial pneumonitis believed to be secondary to fludarabine was seen in a single patient in subgroup 1 that necessitated taking them off of study after fludarabine cycle 5. This patient's symptoms resolved quickly with initiation of corticosteroids.
Quantitative serum immunoglobulins (IgG, IgA, and IgM) were obtained pretherapy and monitored during and after treatment completion. In the 34 patients who completed all therapy, 79% (27 of 34 patients) demonstrated stable (n = 24) to improved (n = 3) quantitative immunoglobulin levels.
Factors Associated With Grade 3 or 4 Neutropenia
There was a highly significant association of IPI score with neutropenia grade (Table 4), in which patients with an IPI score of ≥ 2 were 22 times more likely to have grade 3 or 4 neutropenia as compared with those with a low IPI score (P = .009). Histology and marrow involvement were not found to be associated with grade of neutropenia observed.
Molecular bcl-2 Monitoring by PCR
There were 16 patients who were positive for bcl-2 by PCR in peripheral blood ± bone marrow aspirate before therapy. Among these, 87% (13 of 15 patients) converted to bcl-2–negative after treatment in peripheral blood. Among the 16 patients who were bcl-2–positive pretreatment in bone marrow aspiration samples, 88% (14 of 16 patients) became negative for bcl-2 after therapy. In those patients converting to negative, repeat bcl-2 studies were performed on a yearly basis. Serial bcl-2 assays have remained negative for up to 4 years in patients to date.
Flow Cytometric Monitoring of Lymphocyte Subsets
Multiparameter flow cytometric evaluation of changes in peripheral-blood lymphocyte subsets were evaluated during and after F+R immunochemotherapy. Changes in T cells, B cells, NK cells, and activated NK cells during the post-treatment period are shown in Figures 4 through 7. The upper and lower part of each box bound the interquartile distance (ie, the 25% and the 75% quartile); the dark line in the box is the median; the "+" is the mean; and the thin lines (whiskers) emanating from the top and bottom of the box extend as far as the most extreme values if they are no more than 1.5 x the interquartile distance from the top (or bottom) of the box. Values more extreme than this are represented as individual boxes, and the number next to an extreme point is the patient's study enrollment number. A line connects the median values at each time. Note that the width of each box varies with the number of patients available at the corresponding time. The vertical axis (ie, the cell count) is in log coordinates. As previously described, follow-up visits 1, 2, and 3 are 4 months apart, whereas each follow-up visit after follow-up visit 3 are at 6-month intervals. More marked decreases in CD3+ T cell and CD19+ B-cell lymphocyte subsets were seen after F+R therapy, as compared with changes seen in NK and activated NK cell subsets (Figs 4 through 7).
DISCUSSION
The first rituximab combination immunochemotherapy trial demonstrated that rituximab can be safely given with standard CHOP chemotherapy,25 with a resultant 100% overall response rate (57% rate of CR), in 38 assessable patients with a durable remission duration and the ability to molecularly clear blood and/or marrow of bcl-2–positive cells by PCR testing. The initial rationale behind our decision to evaluate fludarabine and rituximab was the hope that this combination might achieve similar efficacy as R-CHOP, but with less toxicity. Furthermore, because of the significant number of older patients with comorbid medical problems, a less aggressive, non–anthracycline-containing combination immunochemotherapy such as F+R could prove to be a valuable addition to current therapeutic options. In addition, basic research has demonstrated that fludarabine and rituximab have synergistic antitumor activity.19,20
Although late onset of response has been a feature of some studies using rituximab monotherapy,32 F+R induced a relatively rapid rate of response. In the 34 patients who completed therapy, 82% (28 of 34 patients) demonstrated maximal or near maximal nodal responses by midtherapy. Although B-cell and T-cell subsets were overall depleted by this regimen, NK cells (including activated NK cells) were relatively preserved. The first report of the potential sparing effect of infusional fludarabine on human T-10 positive (ie, probably NK) cells determined by flow cytometric evaluation of peripheral-blood mononuclear cells postchemotherapy was conducted by Boldt et al in 1984.33 Other researchers not only have corroborated this finding of a relative sparing of NK cells, but have also found that fludarabine may actually stimulate NK-cell lytic activity.34-36 This relative sparing, and possible activation, of NK cells by fludarabine may significantly contribute to the excellent antitumor activity demonstrated here by the F+R combination. NK cells likely play an important role as a major effector cell used in the destruction of rituximab-bound tumor cells via ADCC.37 Theoretically, it is possible that the addition of other agents to F+R (eg, drugs that deplete NK cells, corticosteroids, and so on) that may decrease ADCC and complement-mediated cytotoxicity activity may have a potentially negative impact on overall antitumor activity. The relative sparing of NK cells, along with the preservation of mean quantitative immunoglobulin levels in the majority of our patients, may explain the low incidence of infectious complications seen in our study. However, acyclovir prophylaxis is recommended in patients receiving F+R combination therapy because of the relatively high incidence of herpes infections seen.
Discontinuations resulting from cytopenia or infectious complications were rare in this study, and dose reductions of fludarabine occurred typically late in the course of therapy. Furthermore, transient therapy delays were necessary in less than one third of the patients in subgroup 2. Review of several previously published reports of fludarabine monotherapy38-42 in patients with either previously treated or untreated CLL and/or indolent B-cell lymphoma documented that myelosuppression is the major adverse effect associated with this drug, dermatomal herpes/zoster infection is not uncommon, and that nonhematologic toxicities are typically mild. In an article by Keating et al,38 post-therapy CD4 and CD8 lymphocyte counts are substantially decreased and seem to slowly increase, but do not reach median pretreatment values at more than 24 months. Results from our F+R trial are consistent with these previously published observations. Two phase II trials of combination F+R in CLL have recently been published.43,44 In the German trial,43 31 patients (11 previously untreated) received four cycles of fludarabine and four infusions of rituximab. An overall response rate of 87% (33% rate of CR; 55% rate of PR) and a median duration of response of 75 weeks were reported. The other trial was a randomized phase II Cancer and Leukemia Group B study (CALGB 9712)44 of fludarabine with concurrent versus sequential rituximab in previously untreated B-CLL patients. Concurrent rituximab and fludarabine (six cycles) resulted in an overall response rate of 90% (47% rate of CR; 43% rate of PR). This overall and CR rate was lower in the sequential arm (overall response rate = 77%; CR = 28%). At a median follow-up time of 23 months, median response duration and survival had not been reached for either arm. Comparing both arms, the concurrent arm experienced an increased incidence of grade 3 or 4 neutropenia of uncertain etiology. Notably, this finding was not accompanied by any increased risk of infections in the concurrent arm. Factors that could potentially be associated with an increased risk of developing grade 3 or 4 neutropenia were evaluated in our F+R study population. Of interest, the presence of marrow involvement (and potential resultant decrease in marrow reserve) was not found to be associated with grade 3 or 4 neutropenia. However, patients with IPI ≥ 2 were 22-fold more likely to have grade 3 or 4 neutropenia compared with patients with IPI of ≤ 1 (P = .009). How this finding is correlated to an increased risk of neutropenia is not clear and will require further investigation.
Statistical analysis of clinical characteristics versus TTP were evaluated in our F+R patients. Older patients (ie, older than 60 years; n = 12 of 40 patients) demonstrated more than 2.5 times the risk of experiencing relapse earlier than younger patients (ie, ≤ 60 years; n = 28 of 40 patients; P = .04). Actual median TTP was 26.3 months (range, 10.6 to 54+ months) versus not reached after a median follow-up time of 44 months in the older versus younger group, respectively. When comparing these two groups, it was not surprising to find an IPI score of ≥ 2 in 100% of older compared with 29% of younger patients. Whether these TTP results represent true biologic differences in indolent B-cell NHL based on age and subsequent responsiveness to F+R versus a limitation of univariate analysis in a relatively small number of patients is unclear. This question may be answered by prospectively analyzing a larger number of patients undergoing an F+R–containing regimen in future trials and/or evaluation of gene expression profiles by microarray analysis between older and younger patients receiving F+R. Nevertheless, F+R seems to be at least comparable in antitumor activity to other fludarabine combination chemotherapy regimens (eg, FND, F+C), but potentially less toxic and better tolerated in an older patient population.
In conclusion, we have demonstrated that concurrent F+R immunochemotherapy resulted in excellent antitumor activity and is well tolerated in previously treated and untreated patients with indolent B-cell lymphoma (overall response rate = 90% in ITT group; 80% rate of CR, 10% rate of PR; 88% bcl-2 conversion rate by PCR), with median TTP not reached at 44+ months. Molecular remissions up to 4 years have been demonstrated to date. This is encouraging because other work has indicated that patients who achieve molecular remission during the first year of treatment have a significantly longer FFS at 4 years compared with those who do not (76% v 38%; P < .001).45 Although the majority of patients experienced transient, reversible grade 3 or 4 neutropenia, this was not associated with a higher-than-expected infection risk. The low infection rate in our study population is likely multifactorial: minimal to no mucosal membrane breakdown associated with F+R, preservation of serum immunoglobulin levels in approximately 80% of patients; and relative sparing and possible activation of NK cells related to fludarabine therapy.
Secondary to the documented hematologic toxicity and the finding that more than 80% of patients had experienced maximal or near maximal nodal response by midtherapy using the current F+R schedule, a trial of reduced fludarabine (eg, possibly day 1 rituximab plus days 3 through 5 of fludarabine every 28 days for six to eight cycles) is warranted and should potentially maintain excellent activity with less hematologic toxicity. Our data support use of an F+R arm in future phase III studies comparing rituximab chemotherapy regimens to each other in an attempt to define an optimal therapy for patients with indolent B-cell lymphoma.
Authors' Disclosures of Potential Conflicts of Interest
NOTES
Supported by Genentech Inc, IDEC Pharmaceuticals, and Berlex Laboratories.
Authors' disclosures of potential conflicts of interest are found at the end of this article.
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