Preclinical data have supported the use of fludarabine and cyclophosphamide (FC) in combination for the treatment of indolent lymphoid malignancies. Previously reported schedules were highly effective, but were complicated by significant myelotoxicity and infectious complications. In the current study, the authors analyzed their experience with an attenuated dose regimen to determine whether equivalent efficacy could be achieved with reduced toxicity.
Sixty-four patients with indolent lymphoid malignancies were treated with intravenous fludarabine at a dose of 25 mg/m2 and cyclophosphamide at a dose of 250 mg/m2, each given on Days 1–3 for a median of 4 cycles. The median age of the patients was 60 years. Nineteen percent of the patients were previously untreated, and 45% had refractory disease; the patients had received a median of 2 prior therapies. With regard to histology, 41% of the patients had chronic lymphocytic leukemia or its variants, whereas the remainder of patients had low-grade non-Hodgkin lymphoma, predominantly follicule center cell lymphoma.
A total of 237 cycles were delivered. The principal toxicities reported were neutropenia (NCI CTC Grade 4 in 17% of cycles) and infection (Grade ≥ 3 in 6% of cycles). The overall response rate and complete response rate were 86% and 29%, respectively. No significant difference could be discerned with regard to response rates for patients with untreated, recurrent, or refractory disease.
Fludarabine is one of the most active agents in the treatment of patients with chronic lymphocytic leukemia (CLL) and indolent non-Hodgkin lymphoma (NHL).1, 2 However, the overall response (OR) and complete response (CR) rates remain modest, particularly in the setting of recurrent disease. In addition, three Phase III trials of fludarabine in CLL have failed to demonstrate a survival advantage over alkylating agent-based therapy, although fludarabine did result in longer disease remissions in previously untreated patients.3–5 The goal of durable disease control first requires a high rate of CR, providing the rationale for the investigation of fludarabine-based combination regimens.
Early fludarabine combination regimens explored the addition of anthracyclines or related topoisomerase-II inhibitors, with or without corticosteroids. The fludarabine, mitoxantrone, and dexamethasone (FND) regimen was highly effective, but was complicated by a 12% incidence of Pneumocystis carinii pneumonia (PCP).6 The subsequent omission of corticosteroids from this regimen resulted in the elimination of PCP risk, but may have decreased its efficacy.7 The relatively modest activity of anthracyclines in patients with CLL8 and indolent NHL9 suggested that other cytotoxic classes may be even more active in combination, leading to the development of the fludarabine and cyclophosphamide (FC) regimen.
The rationale for combining fludarabine with cyclophosphamide is well supported by preclinical data. DNA interstrand cross-links induced in CLL lymphocytes after exposure to 4-hydroperoxycyclophosphamide (4-HC; activated cyclophosphamide) were quickly repaired and became undetectable after 6–8 hours. However, the addition of fludarabine markedly inhibited this repair process, such that the majority of cross-links were still detectable 6 hours later.10 Further in vitro studies have demonstrated increased incorporation of F-ara-A (the nucleoside of fludarabine) into lymphocyte DNA after exposure to 4-HC, with a greater than additive effect in apoptosis induction.11 Other studies have confirmed findings of impaired DNA repair and synergistic apoptosis induction with FC.12
Clinically, the in vitro synergistic cytotoxicity of FC is reported to translate into highly promising OR and CR rates for patients with indolent lymphoid malignancies.13–27 Early data from a Phase III trial of FC versus fludarabine alone support improved response rates and progression-free survival with FC.28 However, there remains considerable heterogeneity in the published schedules of FC, and significant uncertainty exists regarding which regimen is most effective and least toxic. Although some investigators added a single dose of cyclophosphamide to 5 days of fludarabine,18, 19 others advocated the daily coadministration of both drugs.22 The latter approach is designed to capitalize on the synergy demonstrated in preclinical studies, with one such regimen popularized by the M. D. Anderson Cancer Center (MDACC). There appears to be a direct correlation between the cyclophosphamide dose and increasing toxicity in the MDACC regimen.15, 22 To establish whether a lower dose of cyclophosphamide may achieve equivalent results with less toxicity, we analyzed our observed results using an attenuated-dose MDACC regimen in a population of predominantly heavily pretreated patients with advanced indolent lymphoid malignancies.
MATERIALS AND METHODS
Between October 1996 and April 2003, FC was the recommended treatment for indolent lymphoid malignancies in the study institution. To be suitable for this treatment, patients were required to have a histologic diagnosis of either “low-grade NHL” (according to the International Working Formulation),29 CLL (according to the criteria of the National Cancer Institute [NCI]Working Group),30 or prolymphocytic leukemia (PLL) or prolymphocytoid transformation of CLL.31 Patients with the World Health Organization (WHO) classification of mantle cell lymphoma (MCL)32 also were treated. Other eligibility requirements included a serum creatinine level < 0.20 mmol/L, serum bilirubin < 35 μmol/L, and an Eastern Cooperative Oncology Group (ECOG) performance status ≤ 3. Pretreatment staging investigations included computed tomography scans of the chest, abdomen, and pelvis; a complete blood count; biochemistry panel; determination of lactate dehydrogenase (LDH) and β2-microglobulin (β2m) levels; and bone marrow aspiration and biopsy. Functional staging with 18fluorodeoxyglucose (18FDG)-positron emission tomography was performed in some patients.33
Patients received fludarabine at a dose of 25 mg/m2 followed by cyclophosphamide at a dose of 250 mg/m2 (both given intravenously over 15–30 minutes) daily on Days 1–3. Corticosteroids were prohibited as antiemetics during initial therapy but could be added at the discretion of the investigator to improve antiemetic control with subsequent cycles. Treatment was repeated at 4-week intervals provided that hematologic recovery (a neutrophil count > 1.5 × 109/L and a platelet count > 100 × 109/L) had occurred. In patients with thrombocytopenia at the beginning of therapy, courses were delayed only if the platelet count had not returned to baseline after 4 weeks. Dose reduction (to 25 mg/m2 × 2 days for fludarabine and 250 mg/m2 × 2 days for cyclophosphamide) was recommended in the event of prolonged cytopenias, severe sepsis associated with neutropenia, or other nonhematologic toxicities of ≥ National Cancer Institute Common Toxicity Criteria (NCI CTC; version 3) Grade 3. Hematopoietic growth factors were not used routinely. Patients did not receive PCP prophylaxis in the absence of significant corticosteroid exposure or a prior episode of proven PCP. The duration of treatment was planned for a total of four to six courses, or until the maximum response was attained.
Response Criteria and Data Analysis
Responses were categorized according to published criteria for NHL,34 and required the normalization of all radiologic studies and the resolution of all manifestations of disease for a CR and a ≥ 50% reduction in the sum of the products of the perpendicular dimensions of all measurable disease for a partial response (PR). The response criteria for patients with CLL and its variants were taken from those of the 1996 NCI Working Group35 and the response criteria for patients with Waldenström macroglobulinemia (WM) were those established by Dimopoulos et al.36 The International Prognostic Index (IPI) score was calculated using published criteria.37 Overall survival was calculated from the date of treatment initiation, and disease remission duration was calculated from the date that disease remission first was established, with patients receiving further therapy during ongoing disease remission censored at that date. These parameters were analyzed using the method of Kaplan and Meier. Comparisons were made using the log-rank test. Categoric data were compared using the chi-square test or Fisher exact test and ordinal data were compared using the Wilcoxon two-sample test or the Kruskall–Wallis test, as appropriate. All reported P values were two-sided.
Comparison with Historic Cohort
The actuarial disease remission and survival duration of previously treated patients in the FC cohort were compared with those of a historic cohort of patients who were treated with fludarabine and mitoxantrone (FM) at the study institution.7
A total of 64 patients were treated (Table 1). Greater than 80% of the patients had recurrent or refractory disease; of these patients, the majority were heavily pretreated (median number of prior therapies, 2; range, 1–10 prior therapies), approximately one-third had disease that was refractory to alkylating agents, and approximately one-third had previous exposure to fludarabine. The median time from diagnosis was 4 years. Histologic subtypes included CLL or small lymphocytic lymphoma (SLL) (n = 21), PLL or CLL/PLL (n = 5), follicle center cell lymphoma (FCL) (n = 22), MCL (n = 3), marginal zone lymphoma (MZL) (n = 4), and WM (n = 9). Advanced stage disease (Ann Arbor Stage III/IV) was found to be present in 92% of patients and the median IPI score was 2.
Table 1. Baseline Characteristics of the FC and FM Cohorts. The Values Shown Are the Median Unless Specified Otherwise
FC cohort (n = 64)
FM cohort (n = 29)
FC: fludarabine and cyclophosphamide; FM: fludarabine and mitoxantrone; CLL: chronic lymphocytic leukemia; SLL: small lymphocytic lymphoma; PLL: prolymphocytic leukemia; ECOG: Eastern Cooperative Oncology Group; LDH: lactate dehydrogenase; β2m: β2-microglobulin.
Either chronic lymphocytic leukemia with 15–55% peripheral blood prolymphocytes (two cases) or primary prolymphocytic leukemia (three cases).
Where data required for the calculation of the International Prognostic Index score were not known (e.g., lactate dehydrogenase), this was assumed to be normal.
A comparison with a historic cohort of patients who were treated with FM (Table 1) demonstrated that the baseline variables were similar, with a comparable distribution of histologic subtypes in both cohorts.
One patient who withdrew from treatment because of osteomyelitis complicating his only course of FC did not undergo restaging investigations, making him inevaluable for response. Of the 63 patients who were assessable for response (Table 2), the OR rate was 86% and CR rate was 29%. Among previously untreated patients, the OR and CR rates were 75% and 42%, respectively; these rates were not found to be significantly different from those of previously treated patients (an OR of 88% and a CR of 25%, respectively). Among previously treated patients, no significant difference in response rates was observed between patients with recurrent disease and patients with refractory disease.
In the 21 patients with CLL or SLL, the OR and CR rates were 90% and 19%, respectively. It is interesting to note that 2 of 3 patients with previously untreated CLL (66%) attained a CR. The OR and CR rates for other histologic tumor types were 80% and 60%, respectively, for CLL/PLL or PLL; 81% and 48%, respectively, for FCL; 67% and 33%, respectively, for MCL; 75% and 0%, respectively, for MZL; and 100% and 0%, respectively, for WM.
Advanced patient age was found to be associated with an inferior OR rate (94% for patients age < 60 years vs. 77% for patients age > 60 years; P = 0.05), but no correlation was observed between response rates, poor performance status, high LDH or β2m levels, or IPI score.
Delivery of Therapy
A total of 237 cycles of FC were administered (median of 4 cycles per patient; range, 1–6 cycles). The median intercycle interval was 28 days with 29 cycles (17%) delayed by > 7 days. There was no evidence of cumulative myelotoxicity, with 14% of Cycles 2–3 and 23% of Cycles 4–6 delayed by > 7 days (P = 0.24). There were few treatment modifications; 21 cycles (9%) were dose reduced, and 19 cycles (8%) had granulocyte–colony-stimulating factor (G-CSF) added. PCP prophylaxis was administered in 18 patients (28%).
Restricting the analysis to those cycles in which there were no dose modifications or the addition of G-CSF, a total of 131 cycles were evaluable for hematologic toxicity (Table 3). Neutropenia was the predominant hematologic toxicity, with a neutrophil count < 1.0 × 109/L and < 0.5 × 109/L reported in 35% and 17% of cycles, respectively. Thrombocytopenia was appreciably less common, with ≥ Grade 3 thrombocytopenia (< 50 × 109) reported to complicate only 11% of cycles. There were no reported episodes of clinical bleeding.
Table 3. Hematololgic Toxicity in FC Analysis Restricted to Full-Dose, Nongrowth Factor-Supported Cycles
No. of cases
% of cycles
FC: fludarabine and cyclophospha-mide.
Grade 3 or 4
22/131 (17/64 patients)
17% (27% patients)
Grade 3 or 4
Infections complicated 13% of cycles, affecting 38% of patients. The majority were mild, with Grade 3 or 4 infections reported to complicate 6% of cycles in 10 patients (16% of patients). We explored the incidence of ≥ Grade 3 infection according to the extent of past therapy (untreated vs. pretreated, 2% vs. 7%; P = 0.22), advanced patient age (age ≤ 60 years vs. age > 60 years, 4% vs. 9%; P = 0.10), and poor baseline performance status (0–1 vs. ≥ 2, 5% vs. 10%; P = 0.27). Specific severe infections included five episodes of culture-negative febrile neutropenia, three episodes of bacterial pneumonia, and one episode each of osteomyelitis, disseminated listeriosis, disseminated varicella zoster sepsis, respiratory syncytial virus pneumonia,38aspergillus fumigatus pneumonia, and hepatic candidiasis. One additional patient died of Pseudomonas aeruginosa sepsis in association with prolonged neutropenia, 6 weeks after his final cycle of FC.
Nonhematologic toxicities were uncommon. Nausea complicated 32% of cycles but was rarely severe, with ≥ Grade 3 nausea reported to occur in 3% of cycles. Hepatotoxicity reportedly complicated 13% of cycles but no cases of ≥ Grade 3 toxicity were noted. One patient with prolonged pancytopenia after four cycles of FC was found to have morphologic and cytogenetic evidence of myelodysplasia. This patient had received six prior therapies including alkylating agents and irradiation, and although cytogenetics were not available from a pretreatment bone marrow examination, there was no morphologic evidence of myelodysplasia at that time. No autoimmune phenomena were observed.
The median follow-up for surviving patients was 31.5 months (range, 4.5–70.5 months). Of the 54 patients who achieved a CR or PR, 28 had developed recurrent disease, 5 had undergone histologic transformation, 1 had died of sepsis from prolonged aplasia, and 7 were censored for additional therapy in ongoing disease remission. Histologic transformation occurred at a median of 11 months (range, 2–30 months) after FC therapy; there was no correlation noted between the number of FC cycles delivered and the risk of histologic transformation. The actuarial median duration of disease remission was 13.1 months (Fig. 1).
There were 23 deaths reported among the study patients. One previously mentioned death from sepsis in a patient with prolonged aplasia was considered to be related to treatment. Twenty-one patients died of complications related to refractory or recurrent disease. One patient who was not assessable for response died of metastatic skin carcinoma 21 months after the cessation of FC therapy. At the time of last follow-up, the median survival had not been reached, with a 5-year actuarial overall survival rate of 55% ± 8% (Fig. 2).
Because all responders had achieved the maximum response within 6 months of the initiation of therapy with FC, a landmark analysis of survival by response was performed at that time point. The actuarial 5-year survival rate for all responders from 6 months after the administration of FC therapy was 65% ± 9%, compared with an actuarial 4-year survival rate of 17% ± 15% for nonresponders (P = 0.003).
Comparison with Historic Cohort
When restricting the analysis to patients who had received prior therapy, Kaplan–Meier plots were constructed to estimate the median disease remission of the FC cohort compared with a historic cohort of patients who were treated with FM (Fig. 3). There was a trend toward a longer duration of disease remission among the FC patients, with more patients reported to be in ongoing disease remission at 2 years (37% ± 8% vs. 6% ± 6%; P = 0.07).
Peripheral Blood Progenitor Cell Mobilization
Nineteen patients underwent 21 episodes of peripheral blood progenitor cell (PBPC) mobilization, and in 12 patients (63%) an adequate number of stem cells (defined as > 2 × 106/kg CD34+ cells) was collected. There was no apparent correlation between the amount or timing of the fludarabine exposure (as measured by total fludarabine exposure and time between the last administration of FC and the collection attempt) and the adequacy of stem cell collection. However, there was a trend for patients who had attained a CR to have a higher rate of successful PBSC collection (86% vs. 50%; P = 0.12), suggesting that adequacy of disease clearance was important for stem cell mobilization.
Early reports of FC emerged in the late 1990s, when response rates of 70–100% were reported in the salvage treatment of patients with recurrent, low-grade lymphoid malignancies, predominantly FCL or CLL.13–17 Later reports evaluated the activity of FC in untreated patients,18, 19, 21, 22, 25, 26, 28, 39 as well as in the individual histologic subtypes of CLL,19–22, 25, 28 FCL,18, 19, 27 MCL,19, 39 PLL,40 and WM23 (Table 4).
Table 4. Published Response Rates of FC by Histology. Only Studies with ≥ 10 Patients per Histologic-Group were Included
The OR and CR rates of 86% and 29%, respectively, reported herein are similar to those of earlier reports of FC in predominantly pretreated patients with mixed histologies.13, 14, 16, 24, 27 However, with emerging evidence that individual histologies differ in their response to FC, it is more significant to analyze response rates by histology.
To our knowledge, CLL is the single most studied histology in reports of FC. In what to our knowledge, is the largest study published to date of FC as the first-line treatment of CLL, the German CLL Study Group reported interim OR and CR rates of 94% and 20%, respectively.28 High response rates to FC also were observed among patients with recurrent/refractory CLL; in pretreated patients who were not resistant to fludarabine, O'Brien et al. reported OR and CR rates of 82% and 14%, respectively.22 It is important to note that response rates were significantly lower among fludarabine-resistant patients, and such patients were nearly three times more likely to develop significant infections during FC therapy.22 The current study OR and CR rates of 90% and 19%, respectively, in a cohort of predominantly pretreated patients were similar to those of other reports.
Reports of FC efficacy in patients with FCL are limited by their inclusion in studies in which they are intermixed with other NHL patients.13–17 Nevertheless, the few studies in which individual response rates for FCL are available suggest that FCL is highly sensitive to FC (Table 4). Two studies in which 5 days of fludarabine and 1 day of cyclophosphamide treatment were used in untreated FCL patients reported excellent OR and CR rates of 90–100% and 60–89%, respectively, but were limited by significant toxicity.18, 19 Even in the setting of recurrent/refractory disease, FCL remains highly sensitive to FC, and the current study OR and CR rates of 83% and 44%, respectively, are comparable to those reported by Santini et al.27
To our knowledge, reports of FC in other histologic tumor subtypes are limited. Among patients with MCL, the current study experience with three pretreated patients resulting in one CR and one PR is similar to that of prior reports.19, 39 Of the five patients with PLL, three CRs and one PR were achieved; the results of the current study compare favorably with the experience of Herold et al.40 and are highly promising in a disease histology that is notorious for treatment resistance.41 Similarly, the current study cohort of nine patients with WM who were treated with FC resulted in an OR rate of 100%, which is higher than that reported by Dimopoulos et al. in a similar patient population.23
A surprising finding of the current study was the lack of a significant effect of pretreatment on response rates. In part, this is because of the relatively low number of untreated patients (n = 12) in the current series and the high response rates achieved, even among pretreated patients (OR of 88% and CR of 25%), resulting in insufficient power with which to detect small differences in the response rates. Our experience with high OR rates in pretreated patients is reflected in the literature, in which comparisons between published studies revealed little difference with regard to the OR rate between previously untreated and pretreated patients for most tumor histologies (Table 4). In CLL, three studies comparing the results between previously untreated and pretreated patients found no significant differences with regard to the OR rate,21, 22, 25 although in what to our knowledge is the largest study published to date, significantly fewer CRs were observed in the pretreated cohort.22 Another important observation is that patients with high LDH and/or β2m levels appeared to have response rates that were equal or superior to patients with no such elevated parameters in the current study (data not shown). Therefore, FC appears to be effective in settings in which inferior responses can be predicted with the use of alkylator-based therapy or single-agent fludarabine alone.
The major reported toxicities of FC are myelosuppression and infections. The degree of toxicity appears to correlate with the schedule and dosage of cyclophosphamide employed. Using the schedule of fludarabine given at a dose of 25 mg/m2 × 5 days and a single dose of cyclophosphamide (600–1000 mg/m2) given on Day 1, Hochster et al. reported severe neutropenia complicating 43% of cycles,18 even in a relatively low-risk population of patients with previously untreated NHL. In addition, severe infections complicated therapy in 11% of patients in the Hochster study, despite the use of prophylaxis against PCP and Herpes simplex. The addition of G-CSF was reported to reduce the frequency of neutropenia, but appeared to have no impact on the reduction of severe infections.19 Subsequent expansion of this single-dose cyclophosphamide schedule in a Phase III trial was abandoned because of unacceptable toxicity.42
In the pivotal MDACC study of FC using a daily coadministration schedule of fludarabine at a dose of 30 mg/m2 on Days 1–3 and cyclophosphamide at a dose of 300–500 mg/m2 on Days 1–3,22 dosages of cyclophosphamide > 300 mg/m2/day were associated with prohibitive myelosuppression and a high frequency of severe infections, an observation supported by earlier studies.15, 16 However, even at the recommended cyclophosphamide dosage of 300 mg/m2, toxicity remained significant with severe neutropenia (< 0.5 × 106/L) reported to occur in approximately 33–50% of patients not receiving growth factors, and severe infection reported to affect approximately one-quarter of treated patients.22, 25
Our reduced dose schedule of fludarabine (25 mg/m2 × 3) and cyclophosphamide (250 mg/m2 × 3) resulted in acceptable degrees of toxicity in a high-risk population of predominantly elderly patients with pretreated disease, with 1 in 6 unsupported cycles reported to be complicated by Grade 4 neutropenia and 1 in 17 cycles reportedly complicated by severe infection. Overall, 27% of treated patients experienced at least 1 episode of Grade 4 neutropenia and 16% experienced at least 1 episode of severe infection; these findings compare favorably with the results of other studies in which higher cyclophosphamide dosages were used without growth factor support.22, 25 Despite the majority of patients in the current study not receiving PCP prophylaxis, no case of PCP was encountered, thereby reconfirming our previous finding that PCP prophylaxis was not required in fludarabine combination therapy in the absence of steroid usage.7
Compared with a historic cohort of similar patients treated with FM, treatment with FC resulted in significant improvement in the duration of disease remission without additional toxicity.43 We believe that the improved outcome achieved with FC was significant and confirmed both preclinical and clinical findings of synergy between fludarabine and cyclophosphamide.
FC as used in the current study was found to be highly effective, with acceptable toxicity despite the omission of routine growth factor support and antimicrobial prophylaxis. The addition of highly active but nonmyelosuppressive agents may increase CR rates in FC therapy, especially in patients with previously treated CLL, in whom CR rates are reported to remain inadequate. Based on promising in vitro evidence concerning rituximab (anti-CD20 monoclonal antibody) in the sensitization of tumor cell lines to the effects of cytotoxic drugs,44, 45 we have initiated a Phase II study of the combination of fludarabine, cyclophosphamide, and rituximab in the treatment of patients with indolent lymphoid malignancies.