Extended Rituximab (anti-CD20 monoclonal antibody) therapy for relapsed or refractory low-grade or follicular non-Hodgkin's lymphoma

Annals of Oncology 10 655-661, 1999. © 1999 Khmer Academic Publishers. Primed in the Netherlands.

Original article Extended Rituximab (anti-CD20 monoclonal antibody) therapy for relapsed or refractory low-grade or follicular non-Hodgkin's lymphoma L. D. Piro,1 C. A. White,3 A. J. Grillo-Lopez,3 N. Janakiraman, 4 A. Saven,2 T. M. Beck,5 C.Varns,3 S. Shuey,3 M. Czuczman,6 J.W. Lynch,7 J. E. Kolitz8 & V. Jain 9 'Cedars Sinai Hospital and UCLA School of Medicine, Los Angeles; 2Scripps Clinic. La Jolla; 3IDEC Pharmaceuticals Corp., San Diego. CA; Henry Ford Hospital, Detroit, Ml; iMountain States Tumor Institute. Boise, ID: 6Roswell Park Cancer Institute, Buffalo, NY; ' University of Florida. Gainesville, FL; 8North Shore University Hospital, Manhasset, NY, 9Texas Oncology. PA, Dallas, TX, USA

A

Summary Background: Rituximab is a chimeric monoclonal antibody directed against the B-cell CD20 antigen which has been utilized for therapy of B-cell non-Hodgkin's lymphoma (NHL). A previous clinical trial demonstrated that treatment with four weekly doses of 375 mg/rrr of Rituximab in patients with relapsed or refractory low-grade or follicular B-cell non-Hodgkin's lymphoma was well tolerated and had significant clinical activity. Patients and methods: To assess the safety and efficacy of Rituximab treatment, an open-label, single-arm, multi-center, phase II study of eight consecutive weekly infusions of 375 mg/m 2 Rituximab in patients with low-grade or follicular B-cell NHL who had relapsed or had failed primary therapy was conducted. Thirty-seven patients with a median age of 55 years were treated. Results: Grade 1 or 2 adverse events were the majority of reported toxicities and occurred most frequently with the first infusion, decreasing with subsequent infusions. No patients developed a host antibody response (HACA) to Rituximab.

Introduction

Attempts to treat B-cell malignancies with immunotherapy, using monoclonal antibodies reactive against B-cell antigens, began over 15 years ago [1]. Since antibody treatments specifically target a subset of cells, the extent of adverse events associated with treatment is less than with chemotherapy [2]. Rituximab is a highly specific mouse/human chimeric antibody engineered by grafting the variable regions targeting the CD20 antigen from murine antibody genes onto human IgGl (kappa) heavy-chain and light-chain constant regions. Relative to murine antibodies, chimeric antibodies have the clinical advantages of reduced immunogenicity [3-5], extended half-lives in humans [3 4, 6], and enhanced interaction with human effector cells [7-10]. The CD20 antigen is a 35 kDa phosphoprotein present exclusively on B cells, including those in most

The mean serum immunoglobulin levels for IgG, IgA, and IgM stayed within the normal range throughout the study. The majority of patients who were bcl-2 positive at baseline in peripheral blood became bcl-2 negative during treatment and remained negative at the time of B-cell recovery. In the 37 intent-to-treat patients, 5 (14%) had a complete response and 16 (43%) had a partial response for an overall response rate of 57%. Of 35 evaluable patients, 21 (60%) responded to treatment (14% CR and 46% PR). In responders, the median time to progression (TTP) and the median response duration have not been reached after 19.4+ months and 13.4+ months, respectively. Conclusions: The safety profile and efficacy achieved in this pilot study of extended treatment with Rituximab compares favorably with those seen with four weekly doses. Further studies are warranted to investigate whether this or other extended Rituximab schedules will result in increased efficacy in all or in certain subgroups of patients with low-grade or follicular NHL. Key words: bcl-2, CD20 antigen, follicular NHL, Rituximab

B-cell lymphomas [11]. The CD20 antigen is an appealing target for treatment of B-cell lymphomas because it does not circulate in the plasma as free protein [8], shed from the surface of CD20+ cells after antibody binding [9], or appear to be internalized, and subsequently downregulated, upon antibody binding [9]. Additionally, the CD20 antigen is present on mature B cells but not on normal plasma cells. Prior studies with Rituximab given as four consecutive weekly infusions of 375 mg/m2 in patients with low-grade NHL resulted in a 48%-50% response rate [2, 12]. Here we report the results from a phase II study of the safety and efficacy of extended Rituximab treatment administered as eight weekly infusions of 375 mg/m2 in patients with low-grade or follicular NHL.

656 Patients and methods Study design This was an open-label, single-arm, multi-center, phase II safety and clinical efficacy study of eight-weekly infusions of 375 mg/m~ of Rituximab in patients with relapsed or refractory low-grade or follicular NHL Study objectives included the following' evaluation of the clinical efficacy (overall response rate, overall complete response rate, time to progression, progressive disease-free interval, and bcl-2 tumor specific marker status), qualitative and quantitative characterization of the safety (adverse events), and determination of the pharmacokinetics.

(six months) after the last dose of Rituximab. The serum was serially diluted and added to microtiter plates coated with polyclonal goat antiRituximab antibody Bound Rituximab was detected using labeled goat antihuman IgG and compared with a standard curve. Patients were monitored at baseline and at specified timepoints during followup for the development of human anti-chimeric antibody (HACA). HACA was detected using one-site (sandwich) ELISA [2, 12]. IgG, IgA. and IgM immunoglobulin levels were to be measured to assess any changes that may have occurred as a result of B-cell depletion. Flow cytometry studies were used to monitor lymphocyte cell surface antigens. Detection of the tumor-specific marker, the bcl-2 translocation t(14:18). in peripheral blood was determined as described in previous methodology [14]. Patient monitoring

Antibody The production of the antibody was as has been described previously [2, 12, 13]. The vialed product was formulated as 10 mg/ml Rituximab (Rituxan*, Mabthera*1), 25 mM sodium citrate. pH 6 5. with 0.07% polysorbate 80 in 0.15 M saline. The product was to be diluted with normal saline to allow a maximum concentration of 1 mg/ml and was to be administered as a single intravenous infusion through a 0.22 micron in-line filter. Infusions were to be performed on an outpatient basis, using either a peripheral or central intravenous (i.v.) line. The initial dose rate of the first Rituximab infusion was to be 50 mg/hour for the first hour. If no signs and symptoms of toxicity were observed, the dose rate was allowed to be escalated gradually (50 mg/hour increments at 30-minute intervals) to a maximum of 300 mg/hour. Patient population and sample size Patients with histologically confirmed low-grade or follicular B-cell lymphoma (IWF Types A - D , small lymphocytic lymphoma. follicular center lymphoma, follicular grades I, 2, and 3) who had relapsed disease or had failed primary therapy and had progressive disease requiring further treatment were enrolled provided they had the following: bi-dimensionally measurable disease, no more than four relapses following standard therapies including chemotherapy, radiotherapy, immunotherapy, and/or autologous bone marrow transplantation, a demonstrable monoclonal CD20+ B-cell population in lymph nodes or bone marrow, and a prestudy performance status of 0, 1, or 2. according to the WHO scale. In addition, patients had to be at least 18 years old, not be pregnant or lactating, following accepted birth control methods, and have a life expectancy of 5 4 months. Within two weeks prior to receiving the first infusion, patients were required to have a hemoglobin level of 3= 8.0 g/dl, absolute granulocyte count of 5 1.5 x 103/mm 3 . a platelet count of ^ 75 x 103/mm3, serum creatinine <2.0 mg/dl (177 umol/l). total bilirubin <2.0 mg/dl (35 umol/l), alkaline phosphatase < 2 x normal, and AST (SGOT) < 2 x normal. The study was approved by the Institutional Review Boards at each study site and all patients were required to give written informed consent. Study sites The following nine sites enrolled patients in this trial: Henry Ford Hospital (Detroit. MI). University of Florida (Gainesville. Fl). Kenneth Norris Cancer Center (Los Angeles, CA). North Shore University Hospital (Manhasset, NY). Scripps Clinic (La Jolla, CA). Mountain States Tumor Institute (Boise. ID), Texas Oncology. PA (Dallas, TX), Roswell Park Cancer Institute (Buffalo, NY), and University of Rochester Cancer Center (Rochester. NY). Study sites were monitored by clinical trials associates of I DEC Pharmaceuticals. The study was initiated on 5 October 1995 and the date of last treatment of the final patient was 7 August 1996. Pharmacokinelic sampling and laboratory analyses Serum samples were obtained for all patients prior to and immediately following each infusion, and at weeks 1. 2. 3. 4. 8 (two months), and 26

Toxicity was evaluated using the National Cancer Institute's Adult Toxicity Criteria (Recommendations of representatives of the National Cancer Institute's Clinical Cooperative Groups and the Cancer Treatment Evaluation Program, February 1988). Peripheral blood was analyzed by flow cytometry at baseline, weeks 2, 4, 8, and 12, and follow-up months 2, 6, 9 and 12. Chest X-ray was performed at baseline and B-symploms were assessed at baseline, weeks 4, 8, and 12, and follow-up months 2. 6, 9, and 12. Computed tomography scans of the neck, chest, abdomen, and pelvis, as well as physical examination and measurement of palpable tumor masses or lymph nodes was conducted at baseline, week 9, and follow-up months 2, 6, 9, and 12 Subsequent to one year of followup, the following were assessed every three months for four years post-treatment: a physical examination and performance status. CBC. and lesion measurements. Serum chemistries and immunoglobulin levels were assessed at 15, 18, 21, and 24 months post-treatment. In addition, a CT-scan of the neck, chest, abdomen, and pelvis, as well as peripheral blood flow cytometry were performed at 18 months post-treatment and then again at months 24, 30. 36.42, and 48. Response criteria Response was determined by the reduction in the overall size of measured lesions and by the duration of lesion reduction. A patient was considered evaluable for efficacy if the patient completed eight full infusions of Rituximab, satisfied all prestudy entry criteria, and met criteria for evaluation of response. Responses were defined as follows, complete response (CR) - no disease is apparent; defined as all lymph nodes visible on CTscan are less than 1 cm x I cm in size, any node that was palpable on physical examination (and considered to be involved in lymphoma) must no longer be palpable or must be negative for lymphoma on biopsy or FNA, the bone marrow (if initally positive at baseline) must be histologically negative for lymphoma, and the liver and spleen (if abnormal at baseline) should return to normal; partial response (PR) - 50% or greater decrease from baseline in the sum of the products of the greatest perpendicular diameters of all the measured lesions and no simultaneous increase in size of any other lesion or no new lesions; stable disease (SD) - patients who do not exhibit at least a 50% decrease or increase in the sum of the products of the greatest perpendicular diameters of all measured lesions: and progressive disease (PD) - any single observation of a 50% increase in the sum of the products of the greatest perpendicular diameters of all the measured lesions or the appearance of a new lesion. Response classifications of PR and CR were confirmed by reassessment 28 days following the original determination of response. Time to progression and response duration were measured from the first infusion and the observation of response, respectively, until progression. Statistical methods The Kaplan- Meier method was used to analyze the time to progression and duration of response. Kaplan-Meier curves were generated using PROC LIFETEST. The Wilcoxon ranksum test was used for the comparison of serum concentration data by clinical response. Clinical adverse event data were assigned preferred terms using COSTART [15]

657 Table I. Patient characteristics. Characteristic Age (in years) Median Range Sex Female Male Race Caucasian Hispanic Asian Histologic gradea Low A B C Intermediate D Other" Disease stage I A II A III A B IV A B N/A Unknown A B N/A a b

Number of patients (%)

55 35-74 15(41) 22(59) 35(95) 1(3) 1(3) 33(89) 7(19) 18(49) 8(22) 4(11) 3(8) 1(3) 3(8) 3(8) 2(5) 2(5) 7(19) 6(16) 1(3) 18(49) 14(38) 3(8) 1(3) 7(19) 1(3) 1(3) 5(14)

Based on the International Working Formulation. Other. IWF type E with some mantle cells.

and were analyzed by calculating the number and percent of patients and events If the same adverse event was reported on consecutive days, then it was to be recorded as a single event. The most severe grading among the individual events was used to characterize this unified event.

Results Patient characteristics Thirty-seven patients were enrolled in the study. The demographics and the disease status of the study population are summarized in Table 1. Adverse events Adverse events were classified as having occurred during the treatment period (time interval between first Rituximab infusion and 30 days after the eighth infusion) or the long-term follow-up period (time interval between 31 days after the eighth infusion and one year after the first infusion). Any adverse event reported as probably or possibly related or of unknown relationship

to the study treatment was considered a related adverse event. The most frequent related adverse events reported during the treatment period are shown in Table 2. The most common events observed during the treatment period were infusion related, consisting of transient fever (38% of patients), chills (35%), asthenia (16%), nausea (11%), and headache (8%). These events were most frequent with the first infusion, with the frequency of events decreasing dramatically with subsequent infusions. Infusion-related adverse events were managed by temporarily halting the infusion and by the administration of acetaminophen or diphenhydramine. Twenty-four related grade 3 events occurred in 10 patients, two of whom withdrew from the study as a result. These events included a severe rash in one patient and shortness of breath, nausea, chills, rigor, urticaria, and hypertension during infusion in another patient. One grade 3 hypotensive event (83/55 mmHg) was reported during the first infusion; the infusion was interrupted and the patient recovered (BP of 96/64 mmHg) after receiving i.v. saline (baseline BP of 98/75 mmHg). One patient who had a CD20+ plasmacytoid lymphoma but no baseline flow cytometry performed to confirm a lack of T-cell antigens, developed a T-cell lymphoma at 19 months post-therapy. Concomitant medications included antimicrobials, antianxiety and antidepressant medications, dietary supplements, cardiovascular drugs, anesthetics, and estrogen hormones. No patients received steroid medications during the study. Infections Patients were monitored for infections for a year following their first infusion, throughout both the treatment and follow-up periods. A conservative stance was taken in recording the number of infections such that patients treated with antibacterial agents in whom no organism was isolated were presumed to have bacterial infection. One patient was hospitalized due to infection. The patient developed both of the reported grade 3 infections, a simultaneous herpes simplex and a varicella zoster infection on study day 125. The patient was hospitalized for three days and received intravenous acyclovir and morphine; both infections resolved. Analysis ofperipheral blood B cells and immunoglobulin levels Flow cytometry was used to determine the lymphocyte populations of all patients before, during, and after treatment with Rituximab. B-cells in peripheral blood were identified as those expressing the CD19 (pan-B) cell-surface antigen. The median absolute B-cell count in peripheral blood at baseline, 46 cells/ul (range 5-4272 cells/ul) was within the normal range (32-341 cells/ul). Treatment with Rituximab resulted in a rapid depletion of peripheral blood B cells, as shown in Figure I. Most patients had depleted B cells by study day 4. B-cell

658 Table 2. Most frequent" related*1 adverse events.

Any adverse event Body as a whole Fever Chills Asthenia Headache Back pain Cardiovascular system Hypertension Hypotension Digestive system Nausea Diarrhea Neutropenia Leukopema Metabolic and nutritional disorders Angioedema Peripheral edema Musculo-skeletal system Myalgia Nervous system Dizziness Skin and appendages Rash Urticaria Pruritus

Grade 1

Grade 2

Grade 3

Grade 4

Total

n

Events

n

Events

n

Events

n

Events

n (%)

Events

10

136

16

35

7

14

1

1

34 (92)

188(53)

12 7 4 1 2

17 9 9 1 2

2 4 2 1 1

3 4 2 2 1

0 2 0 1 0

0 2 0 1 0

0 0 0 0 0

0 0 0 0 0

14(38) 13(35) 6(16) 3(8) 3(8)

20(6) 15(4) 11(3) 4(1) 3(1)

2 1

2 1

0 1

0 1

1 1

1 1

0 0

0 0

3(8) 3(8)

3(1) 3(1)

4 2 3 1

7 2 6 2

0 1 3 4

0 1 3 7

0 0 1 2

0 0 1 4

0 0 1 0

0 0 1 0

4(11) 3(8) 8(22) 7(19)

7(2) 3(1) 11(3) 13(4)

6 3

6 4

1 0

1 0

0 0

0 0

0 0

0 0

7(19) 3(8)

7(2) 4(1)

3

5

1

1

0

0

0

0

4(11)

6(2)

4

6

1

1

0

0

0

0

5(14)

7(2)

5 3 3

5 3 3

0 0 1

0 0 1

1 2 0

1 2 0

0 0 0

0 0 0

6(16) 5(14) 4(11)

6(2) 5(1) 4(1)

Abbreviations: n - number of patients; Events - number of events. Included are events with an incidence of > 5.4%. b Probably or possibly related to study treatment or of unknown relationship. Note: Patient is counted only under worst grade experienced.

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Figure I. Median CD19+ lymphocytes count. The graph of the median CDI9+ lymphocyte levels measured over time demonstrates the changes in B cell levels in the study population during the study. The number of CD19+ lymphocytes was determined by FACS analysis (see 'Patients and methods"). The squares represent individual patient absolute CD19+ lymphocyte counts.

recovery started between six and nine months after treatment and counts returned to the lower limits of normal. Treatment did not alter the absolute T-cell counts, as measured by FACS analysis using CD3, CD4, and CD8 markers, nor the absolute natural killer (NK) cell which remained relatively stable throughout the study.

Serum immunoglobulin levels (IgG, IgA, and IgM) were measured at baseline and at specified intervals for one year during the study. The mean serum immunoglobulin levels for IgG, IgA, and IgM stayed within the normal range throughout the study. A greater than 50% decline from baseline in IgG, IgM, or IgA levels was observed in two. one and four patients respectively.

659 Table 3 Peripheral blood bcl-1 status and B-cell levels. Patient

Study day

Bcl-2 status 3

Absolute B cellsb

006

-8 105 362 -7 108 416 -6 115 311 -1 83 322 0 122 126 -37 -4d 102 2327235" 410 -13 105 322 601 -4 94 344 0 112 210 322

positive negative negative positive negative negative positive negative negative positive positive negative positive negative negative positive negative negative positive positive negative negative negative positive positive negative positive negative negative negative

318 0 111 23 0 27 10 0 15 27 0 36 636 0 736 33 0 56 230 201 0 90 147 41 0 68 69 0 0 29

029

034

030

025

022

008

033

026

converted from bcl-2 positive to bcl-2 negative after treatment and remained bcl-2 negative at the time of B-cell recovery between 6 and 9 months post-treatment as shown in Table 3. Eight of the nine patients who converted were classified as responders; seven of these eight were either complete responders or partial responders with residual lymph nodes of <1.5 x 1.5 cm. BM bcl-2 was measured post-treatment for only a limited number of patients; therefore, an analysis of this data was not performed. Hematologic and chemistry laboratory effects Most hematologic effects from treatment were mild and transient. Four patients had clinically significant reductions in hemoglobin levels, three of whom recovered to > 8 gm/dl by the end of the study. Ten related grade 3 neutropenic or leukopenic events occurred in four patients, all of whom recovered. There were two grade 4 events, both of which were neutropenia. One patient recovered and the other patient, who was neutropenic at study entry, continued to be neutropenic (ANC count < 1000/mm 3 ) at the end of the trial. A grade 3 platelet value of 38,000/mm 3 was reported in one patient which increased to 65,000/mm 3 by the end of the study. One grade 3 toxicity in clinical chemistry laboratory values (hyperglycemia) was noted in one patient (3%) on study day 126; the patient recovered within one day. No hepatic or renal toxicity was noted.

" Bcl-2 marker status was determined by PCR (see 'Patients and methods'). b B-cell levels were determined by measurement of CD 19+ cells (see 'Patients and methods'). c The bcl-2 tests were performed on this study day d The CD 19+ count was performed on this study day.

Table 4. Summary of clinical response. Patient group

n

CR (%)

PR (%)

CR and PR (%)

95% CI (CR and PR)

Intent to treat Evaluable

37 35

5(14) 5(14)

16(43) 16(46)

21 (57) 21 (60)

41%-73% 44%-76%

Abbreviation: n - number of patients.

Levels recovered in all but one patient who continued to have significantly low levels of IgG at the end by the study. No patients developed a detectable anti-chimeric titer (HACA) during the treatment period or during follow-up. Determination of bcl-2 status Bone marrow (BM) and peripheral blood (PB) samples were examined for the presence of cells bearing the bcl-2 genetic marker (corresponding to the t(14:18) translocation associated with the overexpression of the bcl-2 oncogene protein). In peripheral blood, eighteen patients were bcl-2 positive at baseline; 9 of the 18 patients

Clinical response Two patients who did not complete the eight infusions due to adverse events (infusion-related reaction and skin rash) were inevaluable for efficacy. In evaluable patients, 14% (5 of 35) of patients had a complete response and 46% (16 of 35) had a partial response, for an overall response rate of 60%, as displayed in Table 4. In the 37 intent-to-treat patients, 5 (14%) had a complete response and 16 (43%>) had a partial response for an overall response rate of 57%. Thirty-five of thirty-seven patients were considered evaluable for efficacy analysis. There was no apparent site-specific clustering of responses (Table 4). In all patients the average maximum lesion change was 62% reduction. The average maximum lesion change was 100% reduction in patients achieving a CR and 79% reduction in patients achieving a PR. Evaluable patients with SD had an average maximum lesion size reduction of 42%. The mean SPD continued to decline in responders until stabilizing approximately five months after treatment. The median duration of response for the 21 evaluable responders has not been reached after 13.4+ months (range 2.5-32.4+ months) of followup. At 19.4+ months of observation (range 5.3-34.5+ months) the median time to progression for responders has not been reached. For all patients, the median time to progression has not been reached at 7.7 months of observation (range 1.8-

660 ioo -: 1 = Evaluable Responders (N = 21) 2 = Evaluable Patients (N = 35)

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Months to Progression Figure 2. Time to progression. A Kaplan-Meier graph of the time to progression in the 21 responders and in the 35 evaluable patients. Time to progression was the time period between the first infusion and the date of progression; duration of response was the time period between the date of response and date of progression.

34.5+ months). A Kaplan-Meier curve of TTP in evaluable responders is presented in Figure 2. In responders, all B symptoms were resolved by the end of the study. Additionally, most nonresponders had temporary or permanent relief from one or more of their B symptoms and/or other disease-related symptoms. Patients with IWF type A histology had a statistically significant lower response rate compared with patients of IWF types B, C, and D. The response rate in IWF type A patients was 14% (1 of 7) compared with a response rate of 69% (20 of 29) in IWF type B, C, and D patients. Patients with bulky disease did respond to treatment. Sixty-eight percent of patients with lesions < 5 cm responded, 45% of patients with lesions between 5 and 7 cm responded, and 25% of patients with lesions > 7 cm responded. These differences were not found to be statistically significant. Antibody serum levels The median preinfusion serum concentration of Rituximab increased with each subsequent infusion. However, median postinfusion serum levels plateaued after the sixth infusion (range 518.1-558.1 ug/ml). Preinfusion antibody levels varied from 0 ug/ml (dose 1) to 898.2 Ug/ml (dose 7) and postinfusion levels varied from 581.9 ug/ml (dose 1) to 1177 ug/ml (dose 8). Serum concentrations of Rituximab, both during and post-treatment, were strongly correlated with response. The median Rituximab serum concentration was higher for responders as compared to nonresponders at all timepoints. This difference reached statistical significance at all time

points except after the first and sixth infusions. By the fifth infusion, IWF type A patients had significantly lower serum antibody levels than IWF types B, C, and D patients. Discussion Extended Rituximab therapy (eight-weekly treatments) for patients with recurrent or relapsed low-grade or follicular NHL resulted in a favorable 60% response rate in evaluable patients. Remissions appear durable as the time to progression in responders has not been reached after 19.4+ months. These results of extended therapy compare favorably with a previous phase III trial of four weekly courses of Rituximab therapy which showed a 50% response rate in 150 evaluable patients. Furthermore, in this times eight study, patients with stable diseases demonstrated a mean lesion reduction of 42%, indicating biologic sensitivity of the tumor to Rituximab. Thus, the increased response rate and the significant tumor shrinkage in stable disease patients with extended therapy may suggest that other schedules, in certain patient populations, may result in even better response rates. However, randomized trials would be necessary to confirm whether extended therapy results in increased response rates. Most adverse events associated with this outpatient treatment were mild and were similar to those seen in previous trials. Furthermore, extending Rituximab dosing to eight weekly doses did not delay B cell recovery, stimulate an increased incidence of HACA. nor result in

661 an increased incidence of infections compared with shorter Rituximab treatment regimen. The presence of a bcl-2 translocation was used as an additional efficacy parameter. The bcl-2 translocation [t(14; 18)] has been noted in B cells from NHL patients but not in normal B cells; the eradication of the translocation has been used to assess molecular remission [16-19]. In this trial, as was also seen in previous Rituximab studies, most patients who were bcl-2 marker positive at baseline in peripheral blood became bcl-2 negative during the treatment period as their B cells were depleted. Although bcl-2 negativity in the peripheral blood of some patients who did not achieve an objective response suggests that bcl-2 negativity was secondary to B cell depletion, persistence of bcl-2 negativity in nine patients who had recovered peripheral B cells at the time of this publication suggests that, at least in some patients, bcl-2 negativity may be more indicative of molecular remission. Analyses of responses in certain patient subgroups revealed responses in patients who typically have a poor prognosis. Elderly patients, patients with bulky disease, and patients who were resistant to previous chemotherapy do respond to Rituximab treatment. As seen in the four dose trial, serum levels and response were significantly lower in IWF type A patients compared with levels in IWF type B, C, or D patients. The question still remains whether an increase in Rituximab serum levels in IWF type A patients, or in bulky disease patients as well, would result in a higher response. Extended Rituximab treatment, given weekly for eight weeks, is effective therapy for low-grade NHL with acceptable toxicity. Given the limited toxicity and the ease of outpatient administration, further investigation of extended Rituximab dosing regimens is warranted.

Acknowledgement

4.

5.

6.

7.

8.

9.

10.

11.

12.

13.

14

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16.

17 18.

Supported by clinical grants from I DEC Pharmaceuticals Corporation, San Diego, CA. 19.

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Received 25 January 1999; accepted 22 April 1999. Correspondence to. L. D. Piro, MD Cedars Sinai Hospital and UCLA School of Medicine 8201 Beverly Boulevard Los Angeles, CA 90048-4520 USA