Curr Hematol Malig Rep (2013) 8:7–13 DOI 10.1007/s11899-012-0146-x

CHRONIC LEUKEMIAS (S O’BRIEN, SECTION EDITOR)

Lenalidomide Alone and in Combination for Chronic Lymphocytic Leukemia Christine I. Chen

Published online: 20 December 2012 # Springer Science+Business Media New York 2012

Abstract Lenalidomide is a member of the immunomodulatory agents (IMiDs), and is currently approved for use in myelodysplastic syndromes and multiple myeloma. In chronic lymphocytic leukemia (CLL), lenalidomide has anti-tumor activity which appears distinct, both mechanistically and clinically, from that observed in the approved indications. Furthermore, lenalidomide leads to toxicities, such as tumor flare reaction and tumor lysis, even at low dosing, that is not anticipated with lenalidomide therapy in other disorders. This review will discuss the current understanding of the mechanisms of action of lenalidomide in CLL, lessons of administration learned from clinical trials in CLL to date, and the potential role of lenalidomide in CLL for the future. Keywords Lenalidomide . Immunomodulatory drugs . Thalidomide analogue . Tumor flare reaction . Tumor lysis syndrome . Chronic lymphocytic leukemia

Introduction Chronic lymphocytic leukemia (CLL) is one of the most common hematologic malignancies in adults. Although CLL patients at presentation may be asymptomatic and can be monitored with a watch and wait approach, the majority of patients will ultimately progress to active disease with cytopenias, symptomatic adenopathy, and/or constitutional symptoms. The current standard for frontline therapy is the chemoimmunotherapy combination of fludarabine, cyclophosphamide, and rituximab (FCR), based on C. I. Chen (*) Princess Margaret Hospital, 610 University Avenue, 5-220, Toronto, Ontario, Canada M5G 2M9 e-mail: [email protected]

results from the CLL8 trial [1]. Although FCR is highly active with response rates higher than 95 %, its use is limited by marked toxicities and it is not curative. For relapsed or refractory CLL, single agent therapies such as bendamustine, alemtuzumab, and ofatumumab have activity, and numerous combinations thereof have been tested. None, however, have emerged superior and again, all remain noncurative. Patients with fludarabine-refractory disease or those with the presence of a p53 deletion/mutation are particularly challenging, as durable remissions are not achievable with available approved agents when used in combination or as single agents. Hence, novel agents are needed. In recent years, the importance of the tumor microenvironment in supporting the growth and proliferation of CLL cells has become well recognized. This has led to the development of agents, such as the immunomodulatory agents (IMiDs), which do not appear to act primarily by direct cytotoxicity to CLL B cells, but rather by targeting the surrounding microenvironment and immune system [2]. Lenalidomide (Revlimid; Celgene Corporation, Summit, NJ) is a member of the IMiD family of agents, and is currently approved for use in myelodysplastic syndromes (MDS) and multiple myeloma. Lenalidomide’s activity in CLL, both mechanistically and clinically, appears distinct from that observed in these two approved indications. This review will discuss the current understanding of the mechanisms of action of lenalidomide in CLL, lessons learned from clinical trials in CLL to date, and the potential role of lenalidomide in CLL for the future.

Mechanisms of Anti-CLL Activity Lenalidomide is an analogue of thalidomide, an IMiD agent investigated initially in cancer for its anti-angiogenic activity

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mediated through downregulation of cytokines such as vascular endothelial growth factor (VEGF) [3]. However, inhibition of angiogenesis is but one of many potential anti-tumor mechanisms of lenalidomide. In del(5q) MDS, lenalidomide upregulates Activin A and the tumor suppressor gene SPARC (secreted protein acidic and rich in cysteine), leading to inhibition of erythroid progenitor proliferation and cell adhesion [4]. In multiple myeloma, lenalidomide has direct antiproliferative and anti-apoptotic effects on tumor cells [3], and indirect immunomodulatory effects, including the induction of immune effector T and NK cells [5], inhibition of prosurvival cytokines such as TNFα and IL-6 [6], and inhibition of cell adhesion in the tumor microenvironment [7]. Recently, E3 protein ligase complex cereblon (CRBN) was identified as the direct protein target for lenalidomide. Hence, CRBN expression is mandatory for lenalidomide antimyeloma activity, and likewise acquired loss or suppression of CRBN is associated with IMiD resistance [8]. In contrast to MDS and myeloma, lenalidomide does not appear directly cytotoxic in CLL [9], though induction of tumor cell apoptosis through decreased activation of pro-survival kinases in the Akt pathway has been demonstrated [2, 10]. In CLL, an inability to mount an adequate immune response to tumor B cells can lead to unchecked tumor growth and proliferation. This impairment of immunosurveillance can be due to defects in tumor cell antigen presentation via downregulation of adhesion/costimulatory molecules and expression of inhibitory molecules on CLL B cells [11•], as well as defects in effector cell responses, such as T cell migration and immune synapse dysfunction [12]. Lenalidomide upregulates costimulatory molecules such as CD80 and CD86 on CLL B cells [13], and downregulates inhibitory molecule expression (CD200, CD270, CD274, CD276), potentially enhancing antigen presentation. In addition, lenalidomide reverses the actin cytoskeletal defects in T-cells responsible for the defective T cell response [14] and enhances antibody-dependent cellular cytotoxicity (ADCC) of natural killer (NK) cells. Lenalidomide’s immunomodulatory effects therefore appear to be key to the drug’s anti-tumor activity, but these effects may also enhance global immunity. Hypogammaglobulinemia, reported in virtually all CLL patients, may be reversed by lenalidomide-induced upregulation of CD154 on CLL cells, which in turn costimulate normal B cells to produce antibodies [10]. Lenalidomide can also normalize functional T-cell subsets [15, 16]. Lenalidomide’s immunostimulatory actions may also be responsible for specific toxicities observed with lenalidomide treatment in CLL, such as tumor flare reaction (TFR) [9, 17].

Toxicities of Lenalidomide in CLL Upon initiation of the first clinical trials of lenalidomide in CLL, lenalidomide had already been in routine use for

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patients with myeloma and MDS. Toxicities such as myelosuppression, fatigue, and rash were well recognized, and did not forewarn of the distinctive toxicities in CLL that were to emerge. In the first published report of lenalidomide in CLL, Chanan-Khan reported two cases of tumor lysis syndrome (TLS) [18]. An unusual predisposition to TLS was corroborated in subsequent studies, even when doses of lenalidomide lower than routinely used in other diseases were administered [19, 20]. TLS in this setting typically occurs within the first 2 weeks of therapy, and may be associated with pre-existing bulky nodal disease, renal dysfunction, and increased uric acid levels [21]. Interventions that have been successful in mitigating the risk of TLS include prophylaxis with allopurinol and hydration, careful laboratory monitoring, and most importantly, use of lower starting doses of lenalidomide (2.5–5 mg daily dose), followed by slow dose escalation. TFR is also observed frequently with lenalidomide therapy in CLL. TFR is characterized clinically by rapidly enlarging, painful lymph nodes, sometimes associated with fever, localized erythema, or rash. Onset may occur within hours of the first dose, and although it is observed most frequently with the first cycle of therapy, it can be recurrent for months when using lenalidomide intermittently with regular off-drug periods [19]. TFR is not unique to lenalidomide nor to CLL, but TFR rates as high as 58 % when lenalidomide is used for relapsed/refractory CLL and 88 % when used for first-line therapy are notable [18–20, 22•, 23•]. Fortunately, TFR can be managed symptomatically with a short course of nonsteroidal anti-inflammatory or steroid therapy, and does not appear to impact negatively on lenalidomide efficacy. In fact, TFR has been postulated by some to predict response to lenalidomide [9], though this correlation has not been uniformly observed [17, 19, 20]. The mechanisms by which lenalidomide causes TFR symptoms, appear to be related to immune activation. Lenalidomide-induced B cell activation (upregulation of CD40, CD86, CD80) [17, 24], T cell activation (upregulation of CD69), and inflammatory cytokine release (TNFa, IL-6) [13, 20] have all been proposed as TFR mechanisms. Myelosuppression due to lenalidomide is common to all diseases, including CLL. Even with low doses of lenalidomide, the incidence of severe neutropenia in CLL is high (up to 83 % of patients) and growth factor support may be required [18–20, 22•, 23•]. Grade 3–4 thrombocytopenia is less common (up to 45 % of patients) [18–20, 22•, 23•]. Given that many CLL patients will have compromised hematopoiesis at baseline, drug-related myelosuppression may be severe and rapid in onset. This again supports the use of low starting doses of lenalidomide that may be adequate to debulk the tumor burden in the bone marrow and facilitate recovery of normal hematopoeisis. This approach may allow for later dose escalation as needed to optimize response.

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Other commonly observed lenalidomide-associated toxicities in CLL include fatigue, gastrointestinal symptoms (diarrhea and/or constipation), and skin rash. In CLL, skin rashes are common (13–64 % of patients) [18–20, 22•, 23•] and remarkably heterogeneous in presentation (urticaria, nodules, diffuse erythema, eczematous scaling). Rash can manifest as part of TFR at the start of therapy, but may likewise present idiosyncratically later in the treatment course. Up to 5.8 % of patients treated on single agent lenalidomide studies have develop venous thromboembolism (VTE) [18, 20, 23•]. Lenalidomide appears to induce upregulation of soluble vascular endothelial adhesion molecule 1 (sVCAM1), thrombomodulin, and acute phase reactants such as TNFα, C-reactive protein, and FVIII, with particularly high levels of sVCAM1 and TNFα reported in a subset of CLL patients who developed VTE [25]. This suggests that inflammation and endothelial dysfunction may contribute to an increased thrombotic risk in CLL.

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lenalidomide (25 mg versus 10 mg daily for 21 days of a 28 daycycle) in relapsed or refractory disease was initiated [23•]. Of the first 18 patients enrolled in this study, four developed severe TLS, and therefore the study was amended to a Phase 1 trial using a low starting dose of 2.5 mg daily, increasing to 5 mg after 28 days, with subsequent 5 mg increments to one of three dose levels: 10 mg, 15 mg, or 20 mg. Of the 52 patients enrolled, no further TLS was reported, but other toxicities, particularly infections, were common. Even with the conservative starting doses used in this study, one-third of patients could not escalate beyond 2.5 mg, although those who reached the 20 mg dose did not suffer any dose-limiting toxicities. The low response rates achieved in this study (best response PR in six patients; 11.5 %) could be attributed in part to the low doses of lenalidomide used and the heavily pretreated, high-risk characteristics of this cohort (48 % del(17p) and/or del (11q), 52 % fludarabine-refractory). Previously Untreated CLL

Clinical Trials of Single Agent Lenalidomide Relapsed/Refractory CLL Chanan-Khan et al. reported the first clinical trial of single agent lenalidomide in CLL [18]. In this Phase 2 trial of 45 patients with relapsed or refractory CLL, lenalidomide was initiated at 25 mg daily for 21 days of a 28-daycycle. After the occurrence of two cases of TLS in the first 29 patients enrolled, the starting daily dose was reduced to 5 mg, with subsequent 5 mg dose escalations every 1–2 weeks to a maximum of 25 mg. No further TLS was noted. Responses were observed in 21 patients (47 %), with four patients (9 %) achieving a complete response (CR) and 17 (38 %) a partial response (PR). Three patients with progressive disease (PD) were able to achieve PR after the addition of rituximab. Given that 51 % of patients on this study were fludarabine-refractory, these response data were highly encouraging. However, with the two cases of TLS, and rates of TFR and grade 3–4 neutropenia of 58 % and 70 %, respectively, this study demonstrated that administration of lenalidomide in CLL would not be a simple generalization from treatment in other hematologic malignancies. Ferrajoli et al. subsequently studied single agent lenalidomide in 44 patients with relapsed and refractory CLL [20]. Using a more cautious dosing schedule (starting at 10 mg daily continuously, increasing by 5 mg every 28 days to a maximum of 25 mg daily), a more modest overall response rate of 32 % (7 % CR) was achieved. Although grade 3–4 neutropenia was still common using these lower doses (41 %), TFR was significantly reduced (all grades, 12 %) and no cases of TLS were reported. In order to evaluate optimal dosing of lenalidomide in CLL, a sponsor-initiated trial comparing two doses of

Two studies evaluating single-agent lenalidomide as firstline therapy have been published [19, 22•]. In our Phase 2 study, lenalidomide was initiated at 10 mg daily for 21 days of a 28 daycycle, with plans for weekly dose escalations to a maximum daily dose of 25 mg [19]. Once again, TLS and severe myelosuppression were encountered with the first two patients, and the protocol was amended to implement a starting dose of 2.5 mg, with slower monthly dose escalations to a target dose of only 10 mg daily (dose escalation to 25 mg was allowed if needed for response). Even with these low doses, an encouraging overall response rate of 56 % (no CR, 40 % stable disease (SD)) was achieved at a median follow-up of 20.7 months. With a prolonged follow-up to 47 months, this response rate improved to 72 %, including three patients who upgraded to CR status (Chen et al., unpublished data). Hence, high quality, durable responses can be achieved with low doses of lenalidomide, though they may be slow to achieve. Unfortunately in our study, long-term use of lenalidomide was associated with a high rate of severe, often recurrent, neutropenia (76 %). Badoux et al. studied the effects of lenalidomide as firstline therapy for patients older than age 65, using a 5 mg starting dose and monthly dose escalations [22•]. They too reported an encouraging response rate of 65 % (CR 10 %), though grade 3–4 neutropenia was very frequent in this elderly cohort (83 %). Both first-line studies reported somewhat higher rates of TFR (52–88 %) and rash (50–64 %) than are seen in the pretreated population, perhaps suggesting a greater tendency for immunostimulation in these treatment-naïve patients. A large, multicenter, randomized trial comparing single-agent lenalidomide to oral chlorambucil in elderly patients (age 65 or above) is ongoing

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(Clinicaltrials.gov NCT00910910). In this study, the starting dose of lenalidomide is 5 mg to a maximum of 15 mg daily continuously. Predictors of Response Given the small number of CLL studies evaluating singleagent lenalidomide to date, identification of response predictors is limited. A wide number of pre-treatment, baseline variables have been analyzed. In our Phase 2 study, we could correlate the presence of higher pre-study neutrophil counts with response, hypothesizing that higher neutrophils may allow patients to tolerate the myelosuppressive effects of lenalidomide and stay on therapy longer [19]. Similarly, Badoux et al. reported that higher pre-treatment hemoglobin levels correlated with higher tolerated median daily doses of lenalidomide, which in turn correlated with higher likelihood of response [22•]. In the same study, patients with 17p deletion by FISH were less likely to respond, though numbers were small (six of 60 patients). In a subgroup analysis of the Phase 2 trial of lenalidomide in relapsed/refractory CLL by Chanan-Khan, 16 patients were identified as high risk with deletion 11q (ten patients) and deletion 17p (six patients) [26]. Only six of 16 (38 %) patients responded to lenalidomide therapy, with only three CR in the del(11q) patients. Median PFS for the 16 patients was 12.1 months. Summarizing from the limited data available, patients with del(17p) may respond to single-agent lenalidomide, but achievement of CR remains elusive, and it is unclear whether prolongation of PFS or overall survival is feasible. The development of TFR has been reported as a clinical predictor of response [27], but this correlation has not been consistently observed across studies. Although CRBN expression is mandatory for lenalidomide’s anti-tumor activity in multiple myeloma, it is not yet clear whether the same applies to CLL. We have studied CRBN expression by gene expression profiling and Western blot in 19 of the 25 patients treated on our single agent lenalidomide Phase 2 trial [19]. CRBN expression was uniformly identified in all 19 patients at baseline, regardless of subsequent response (Chen et al., unpublished data). Whether serial testing will reveal loss of CRBN expression and correlation with response is currently under investigation at our center. Dosing and Schedule of Lenalidomide As is apparent from published clinical reports of single agent lenalidomide therapy to date, toxicities can be disease-specific and therefore administration details cannot be generalized from other hematologic diseases to CLL. In CLL, it is mandatory to initiate therapy with a low starting dose (2.5–5 mg daily) and TLS precautions. Low doses, however, do not appear optimal for efficacy [22•], and

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therefore dose escalations can be undertaken safely with 5 mg increments every 28 days. More rapid increments may be safe, but have not yet been thoroughly evaluated. Using this approach, only a minority of patients reach the maximum 25 mg dose with median daily doses of 4.5-15 mg achieved [18–20, 22•, 23•]. Both continuous daily dosing and intermittent dosing (21 days on, 7 days off, each 28-day cycle), as is used in myeloma, have been utilized in CLL. In our experience, the continuous daily dosing eliminates the fluctuating peripheral lymphocyte count seen during the week off therapy, and appears as well tolerated as the intermittent schedule. Longer breaks off therapy when using an intermittent schedule does not appear to significantly reduce toxicities [28]. Duration of therapy in all published studies to date have not predefined a set number of cycles, rather continuing until progression and/or CR. Given that initial responses can occur as late as 19 months [19], prolonged and sustained therapy is a reasonable target. Shorter treatment durations may, however, be effective with combination therapy.

Clinical Trials of Lenalidomide in Combination Therapy With the encouraging efficacy of single-agent lenalidomide in CLL, numerous studies evaluating combination therapy have been initiated. None of these studies have been published in full, and therefore only a brief review of promising combinations will be discussed. The combination of lenalidomide and rituximab has been evaluated as both first-line and salvage therapy. Interestingly, there are conflicting in vitro data in primary CLL cells treated with this combination. Lapalombella et al. [29] have reported that lenalidomide downregulates the CD20 antigen and antagonizes the apoptotic and ADCC activity of rituximab, whereas Wu et al. reported that lenalidomide enhances NK cell-mediated ADCC by rituximab [30]. Regardless, this combination appears active, with Ferrajoli et al. reporting overall responses of 66 % (CR 10 %) and median time to treatment failure of 24 months in relapsed CLL [31]. In this study, rituximab 375 mg/m2 was given weekly for 4 weeks, followed by one dose every 4 weeks and lenalidomide 10 mg was administered continuously daily on a 4-weekcycle for 12 cycles. In comparison to single agent lenalidomide, this combination did not appear to increase toxicity, and in fact, TFR appeared less frequently (27 %) than expected. Furthermore, a promising overall response rate of 54 % (13 % CR) was achieved in 15 patients with del(17p). This combination was also evaluated in the frontline setting, leading to overall responses of greater than 90 %, CR 20 % in 69 patients, but mature follow-up is pending [32]. These results are undergoing confirmation in two large, multicenter, sponsor-initiated trials in untreated and relapsed patients

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(Clinical Trials.gov NCT00628238 and NCT01199575). Given the encouraging data from the combination rituximab and lenalidomide studies, Ferrajoli et al. are testing a combination of ofatumumab with lenalidomide in relapsed CLL [33]. Ofatumumab is given weekly for 4 weeks, followed monthly for 5 months, then every other month thereon, and lenalidomide 10 mg continuously, both for total of 24 months. Of 34 evaluable patients thus far, responses were noted in 65 %, CR 15 %. Not unexpectedly, neutropenia was the most common toxicity (44 % grade 3–4) and the median tolerated daily dose of lenalidomide was only 5 mg. Lenalidomide has been incorporated into fludarabinebased combinations, with varying results thus far. In a Phase I trial reported by Brown et al., lenalidomide was concurrently combined with fludarabine and rituximab [34]. Both lenalidomide and fludarabine were dose-escalated in a 3+3 design, starting with lenalidomide at 2.5 mg daily for days 1–21 of a 28-daycycle and fludarabine 25 mg/m2 intravenously for 3 days, with a fixed dose of rituximab. Even at the lowest dose level, dose-limiting toxicities occurred in two of the first four patients enrolled (tumor flare with prolonged neutropenia, rash and rhabdomyolysis). After further dose reduction, recurrence of toxicities (rash, TFR, cytopenias) ultimately led to study discontinuation. Similarly, Flinn et al. used fixed standard doses of fludarabine and rituximab, with two doses of lenalidomide, 2.5 mg and 5 mg, days 1–21 of each 28-day cycle. Toxicities in the first four patients led to a modification to delay the start of lenalidomide to days 8–28, with significantly improved tolerance [35]. Using a similar sequential dosing of lenalidomide in a Phase I trial of the same combination, Egle et al. were able to dose escalate to 25 mg in one-third of the patients, although myelosuppression was common and ultimately limited dosing in 42 % of patients [36]. A remarkable CR rate of 49 % was attained with this approach (overall responses 87 %). From the experience thus far of combination studies, it is clear that combining agents with lenalidomide requires careful consideration of dosing, scheduling, sequencing, and cumulative toxicities in order to maximize anti-tumor activity. Continued vigilance in monitoring for unanticipated toxicities is mandatory.

Future Directions in the Clinical Use of Lenalidomide in CLL Given lenalidomide’s activity in CLL, convenience for administration as an oral agent, and positive experience as a maintenance agent in multiple myeloma, lenalidomide is currently under evaluation for either maintenance or consolidation in CLL. To date, there are limited published data. In one Phase 2 trial using 6 months of lenalidomide as consolidation

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following PCR, six of 23 patients were able to improve the quality of response [37]. A large randomized, placebocontrolled, study evaluating the role of lenalidomide maintenance is ongoing, but preliminary data are not yet available (CONTINUUM study; Clinicaltrials.gov NCT00774345). Future directions in lenalidomide combinations should focus on enhancing lenalidomide’s multiple mechanisms of action based on current understanding. Preclinically, there is promising synergy between lenalidomide and CAL101 (a PI3K-δ inhibitor) [38], SGN-40 (a humanized IgG1 monoclonal antibody) [39], and AT-101 (a BH3 mimetic BCL2 antagonist) [40]. Combination therapies using agents that lack crosstoxicity with lenalidomide, in particular myelosuppression, would be of particular interest.

Conclusion Lenalidomide has emerged as an important therapeutic agent in CLL. Although much has been learned from current experience, lenalidomide remains a complicated agent to understand, both from a mechanistic and a clinical treatment perspective. Lessons learned from studies thus far and in carefully designed studies from hereon, should inform further on the intricacies of dosing, scheduling, ideal combinations, and prevention/management of toxicities of this highly active agent. Disclosure Dr. Chen has received research funding and honoraria from Celgene.

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