Potential Therapeutic Applications of Oblimersen in CLL

B-cell chronic lymphocytic leukemia (B-CLL) is a disorder characterized by the unremitting accumulation of small, slowly proliferating CD5⁺/CD19⁺/CD20⁺ (weak)/CD23⁺/sIg⁺ (weak) monoclonal B cells.[1] CLL is very rare in people younger than age 40. However, its incidence increases sharply after the fourth decade of life, to the point of being the predominant leukemic type in the elderly in the Western world, across race and sex differences (Figure 1). It is estimated that 7,300 new cases were diagnosed in the United States in 2003.[2] Statistics from the nine population-based cancer registries of the National Cancer Institute's Surveillance, Epidemiology and End Results (SEER) Program for 2000 showed significantly better survival differences for patients younger than age 65 (average presentation age: 64 years) than for those 65 years of age and older, although CLL shortens life expectancy considerably, even in younger patients (Table 1).[3] Prognostic Indicators Neither the Rai[4] nor the Binet[5] staging systems, which discriminate CLL by the sites of disease and/or the degree of cytopenias induced by leukemic marrow replacement, enable physicians to identify patients in the good-prognosis group who will eventually progress. However, over time, these systems have proved to be extremely useful for categorizing patients, particularly those participating in clinical trials. The original Rai system[4] consisted of five stages (0 to IV) but was modified to a three-stage scheme in 1987,[6] when analysis of survival data in a large number of patients established three distinct risk categories (low, intermediate, and high), which differ significantly in terms of median survival. Patients in the low-risk group (stage 0) have lymphocytosis with no other abnormality; patients in the intermediate-risk group have enlarged lymph nodes (stage I) and/or spleen (stage II) in addition to lymphocytosis; and patients in the high-risk group have anemia (hemoglobin value < 11.0 g/dL, stage III) and/or thrombocytopenia (platelet count of < 100 * 10⁹/L, stage IV) (Table 2). Serum levels of beta₂-microglobulin,[ 7] lactate dehydrogenase, soluble CD23,[8] and the cell-surface expression of CD38[9] can help predict disease activity. However, the presence of certain cytogenetic abnormalities in the leukemic B cells [10,11] and/or somatic mutations in the immunoglobulin heavy-chain genes seem to predict rapid disease progression and survival more accurately.[12,13] Genetic Components CLL B cells with mutated immunoglobulin heavy-chain genes are associated with the more favorable genetic defect deletion 13q14. Conversely, unmutated immunoglobulin heavy-chain genes are associated with trisomy 12 and the high-risk 17p and 11q genomic aberrations.[14] CLL B cells with unmutated immunoglobulin heavy-chain genes generally show the distinctive expression of the zetaassociated protein of 70 kD (ZAP-70), an intracellular tyrosine kinase, which has been found to be associated with enhanced immunoglobulin receptor signaling in CLL B cells. Mounting data indicate that expression of ZAP- 70 is a more reliable marker of the risk of early disease progression than the mutational status of the expressed immunoglobulin heavy-chain genes in CLL.[15,16] Disease Subtypes Based on these recent findings, it has been proposed that there are two types of CLL (Table 3). The first type arises from relatively less differentiated or "immunologically naive" pregerminal B cells with unmutated immunoglobulin heavy-chain genes, displays atypical CLL morphology, and has a poor prognosis. The other type evolves from more differentiated postgerminal "memory" B cells with somatically mutated immunoglobulin heavy-chain genes and has a good prognosis.[17,18] Treatment With Chemotherapy or Chemoimmunotherapy Approximately one-third of patients with CLL never require treatment. In another third, an initial indolent phase is followed by disease progression. The remaining third show aggressive disease at the outset and need immediate treatment. Ultimately, 50% of patients with CLL will require treatment. Deferring therapy until progression of CLL does not affect survival. Neither single-agent chlorambucil(Drug information on chlorambucil) (Leukeran) nor a variety of combination chemotherapy regimens (some including anthracyclines) have been shown to increase response rates or improve survival in clinical studies.[ 19-21] A meta-analysis of 2,048 patients with early-stage disease included in seven randomized trials comparing immediate or deferred treatment with chlorambucil demonstrated no benefit in either arm.[22] A summary of clinical experience with chemotherapy and chemoimmunotherapy in CLL is provided in Table 4.[23-30] Fludarabine (Fludara), a purine analog, yields better response rates than chlorambucil but causes more myelosuppression and greater reduction in CD4+ lymphocytes.[31] The results of three phase III randomized trials in previously untreated patients with CLL demonstrated the superiority of fludarabine over other alkylating agent-based therapy in terms of response, duration of remission, and disease progression-free survival (but not overall survival).[24,25,27] A phase II trial conducted by Flinn and colleagues[26] showed a complete response rate of 47% in 17 untreated patients with CLL who received fludarabine and cyclophosphamide(Drug information on cyclophosphamide) plus filgrastim(Drug information on filgrastim) (Neupogen) support. In a subsequent randomized phase II trial that was conducted for the Cancer and Leukemia Group B (CALGB 9712), Byrd et al[29] demonstrated that the administration of fludarabine concurrently with rituximab(Drug information on rituximab) (Rituxan) for 6 monthly courses followed by rituximab 2 months later for 4 weekly courses also produced a complete response rate of 47% in 51 untreated patients with B-CLL. These results compared favorably with results that were observed in 53 patients who received fludarabine alone for 6 months followed sequentially 2 months later by rituximab consolidation.[ 29] Of note, two studies[32,33] testing higher doses of single-agent rituximab in patients with pretreated and untreated B-CLL demonstrated a doseresponse relationship for this agent. In addition, investigators at the University of Texas M. D. Anderson Cancer Center recently reported a complete response rate of 67% in 135 patients with CLL receiving fludarabine (25 mg/m²/d) for 3 days, cyclophosphamide (250 mg/m²/d) for 3 days, and rituximab (375 to 500 mg/m²) on day 1 of each treatment cycle.[30] These responses included a high proportion of patients with molecular remissions, indicating the possibility to consolidate the response with autologous hematopoietic cell transplantation. Another treatment option available for patients with chronic lymphocytic leukemia is conventional allogeneic bone marrow transplantation, which may be curative in some cases. However, only 10% of patients are eligible for this treatment, which is associated with significant morbidity and mortality.[34] Allogeneic bone marrow transplantation in which the patient's marrow is not completely ablated by chemotherapy with or without low-dose radiotherapy (often termed a minitransplant) is another option now being evaluated for patients with CLL.[35] Treatment With Oblimersen Sodium Preclinical Experience
A variety of investigational agents that include an assortment of chemical compounds, monoclonal antibodies, and biologic approaches have been evaluated for the treatment of CLL and await confirmation of their clinical usefulness in comparative studies (Table 5). Notable among them is oblimersen sodium injection (Genasense, formerly known as G3139), an innovative treatment modality in the form of an antisense oligonucleotide targeted to the Bcl-2 molecule. Bcl-2 protein is upregulated in a wide variety of lymphoid malignancies, including CLL. The protein is thought to be responsible for maintaining the viability of malignant lymphoid cells and may contribute to chemotherapy and radiotherapy resistance.[36,37] Higher levels of Bcl-2 protein expression have been inversely correlated with survival in previously untreated patients with CLL.[38] In B-CLL cells, expression levels of Bcl-2 protein were significantly elevated over those of normal B cells and correlated inversely with those of the proapoptotic protein BAX.[39] In addition, quantitative expression analysis by reverse transcription polymerase chain reaction showed the relative ratio of Bcl-2 protein to BAX to be markedly increased in CLL and also in mantle cell lymphoma.[40] Previous studies have shown that reduction of Bcl-2 protein expression by antisense therapy sensitizes cells to chemotherapy- induced apoptosis.[41,42] The activity of oblimersen in CLL has been evaluated in vitro and in vivo. Auer et al[43] showed that in cells obtained by CD19 selection of peripheral blood samples from patients with CLL, Bcl-2 protein expression was significantly downregulated and markers of apoptosis upregulated by oblimersen in a sequence-specific manner, compared with sense and nonsense oligonucleotide controls. In this system, oblimersen (2 μM) was more active than either fludarabine (50 μM) or dexamethasone(Drug information on dexamethasone) (1 μM) as an inducer of apoptosis, and potentiation of this activity was noted when oblimersen was combined with either fludarabine or dexamethasone.[43] Furthermore, pretreatment with oblimersen sensitized CLL cells to the apoptotic effect of rituximab in a doseresponse relationship. With similar culture systems, synergism to induce apoptosis has been shown in vitro for oblimersen combined with the novel proteasome inhibitor bortezomib(Drug information on bortezomib) (PS-341, Velcade) [44] and with the humanized anti- CD52 monoclonal antibody alemtuzumab(Drug information on alemtuzumab) (Campath 1H).[45] In these studies, CD19 antibody-selected CLL cells from 12 patients were put in short-term culture with or without oblimersen (at a concentration of 5 μM) to reduce Bcl-2 protein levels. Control sense and nonsense oligonucleotides were also used. Alemtuzumab was added at concentrations ranging from 1 μg/mL to 10 μg/mL. All CLL samples showed some apoptosis with alemtuzumab alone; however, those cells pretreated with oblimersen showed an enhanced level of apoptosis. The addition of oblimersen greatly enhanced the effectiveness of alemtuzumab with similar cell kill at 20% of the dose compared with alemtuzumab alone.[45] Clinical Experience
Oblimersen was evaluated in a nonrandomized phase I/II trial as monotherapy for heavily pretreated patients with relapsed or refractory CLL to determine the maximum tolerated dose and evaluate single-agent activity.[46] Fourteen patients received oblimersen as a continuous intravenous (IV) infusion at a daily dose ranging from 3 to 7 mg/kg for 5 to 7 days every 3 weeks. Several patients exhibited antitumor effects, including tumor lysis, reduction in circulating CLL cells, and decreases in lymphadenopathy and splenomegaly. One patient with Richter's syndrome developed a stable partial response that was reported to last several years. In cycle 1, patients received one of four different oblimersen doses: 3, 4, 5, or 7 mg/kg/d. Six of the patients receiving higher dosages (3 receiving 7 mg/kg/d, 1 receiving 5 mg/kg/d, and 2 receiving 4 mg/kg/d) showed doselimiting toxicity requiring treatment discontinuation (high fever, severe hypotension, hypoglycemia, and back pain requiring opiate analgesics) despite reduction of leukocytosis. In cycle 2, two patients who were escalated from a lower dose to 7 mg/kg/d also showed dose-limiting toxicity. These findings clearly demonstrated that patients with CLL are more sensitive to the side effects of oblimersen than patients with solid tumors, in whom these doses are generally well tolerated. Either tumor lysis or direct oligonucleotide immunostimulation of the malignant B cells is thought to be responsible for this distinct toxicity pattern.[47,48] In a subsequent phase I/II trial in 26 patients with relapsed or refractory CLL who had previously received fludarabine, oblimersen was administered as a continuous IV infusion at a dosage of 3 mg/kg/d for 5 to 7 days every 3 to 4 weeks.[49] Six patients in phase I received the phase II dose of oblimersen (3-4 mg/kg/d). Patients had received a median of 3 (range: 1- 13) prior chemotherapy regimens for CLL. Thirteen, 7, and 4 patients had Rai stages II, III, and IV, respectively, and 2 patients had Richter's transformation. The median age was 61 years (range, 44-70). The evaluable population included 23 patients who received the phase II dose for ≥ 2 cycles (6 enrolled in phase I, and 17 enrolled in phase II). Two patients (9%) achieved partial response, 11 (48%) showed stable disease, and the remaining 10 patients (43%) had progressive disease. Reductions in circulating CLL cells of ≥ 50% from baseline were seen in 9 of 23 patients (39%). In addition ≥ 50% decrease in lymphadenopathy was seen in 8 of 19 patients (42%), and a ≥ 50% reduction of hepatomegaly or splenomegaly was seen in 8 of 16 patients (50%). Oblimersen (3 to 4 mg/kg/d) was well tolerated, with only rare grade 3 or higher toxicities reported. The most common symptoms were fatigue, night sweats, increased dyspnea, and pneumonia. Results confirmed that oblimersen has antitumor activity in pa- tients with CLL, even in the absence of other cytotoxic drugs, and that a daily dose of 3 mg/kg was well tolerated. Subsequently, a phase III trial evaluating fludarabine (25 mg/m²) and cyclophosphamide (250 mg/m²; days 1-3) with or without oblimersen (3 mg/kg/d continuous IV infusion; days 1-7) was initiated in patients with relapsed or refractory CLL. Stratification at recruitment grouped patients according to three criteria: 1) "responsiveness" or "refractoriness" to fludarabine; 2) number of previous chemotherapy regimens (1-2 vs ≥ 3); and 3) duration of response to prior therapy (≥ 6 vs < 6 months). The primary end point was response rate (complete response plus nodular partial response), and secondary end points were overall response (complete response plus nodular partial response plus partial response), overall survival, and time to disease progression. This trial recently completed accrual of 241 patients. Conclusion The rationale for evaluating oblimersen in CLL is that Bcl-2 protein is highly expressed and in all likelihood is a key survival factor in this disease. Oblimersen has shown limited single-agent activity in patients with previously treated CLL. In addition, patients with CLL appear to have considerably more sensitivity to the development of significant hypotension and fever than patients with solid tumors who receive oblimersen at similar dosages, who exhibit better tolerance. Oblimersen 3 mg/kg/d was well tolerated in phase II trials, and this dosage was used in the phase III trial. A trial of oblimersen combined with fludarabine and rituximab in patients with previously untreated or relapsed CLL has recently been initiated.