Emerging therapies for the treatment of relapsed or refractory multiple myeloma

European Journal of Haematology REVIEW ARTICLE Emerging therapies for the treatment of relapsed or refractory multiple myeloma Meletios A. Dimopoulos 1, Jesus F. San-Miguel 2, Kenneth C. Anderson 3 1 Department of Clinical Therapeutics, University of Athens School of Medicine, Athens, Greece; 2 University Hospital of Salamanca, Salamanca, Spain; 3 Dana-Farber Cancer Institute, Boston, MA, USA Abstract Encouraging progress has been made in the treatment of patients with relapsed refractory multiple myeloma (MM). The rapidly evolving understanding of key pathways responsible for tumor growth and survival has led to the development of novel agents (including immunomodulatory drugs, proteasome inhibitors, histone deacetylase inhibitors, and other targeted agents) with the potential to provide significant improvements in response and survival, and influence treatment guidelines. This review summarizes recent advances in understanding of the biology of relapsed refractory MM and clinical trials with novel targeted agents that are currently under investigation for patients with this disease. Key words multiple myeloma; immunomodulatory drug; proteasome inhibitor; Akt inhibitor; histone deacetylase inhibitor Correspondence Meletios A. Dimopoulos, MD, Department of Clinical Therapeutics, University of Athens School of Medicine, Alexandra Hospital, 80 Vas. Sofias, Athens 11528, Greece. Tel: +(30210) 3381541; Fax: + (30210) 3381511; e-mail: mdimop@med.uoa.gr Accepted for publication 10 October 2010 doi:10.1111/j.1600-0609.2010.01542.x Multiple myeloma (MM) is the second most common hematologic malignancy and was responsible for an estimated 21 000 deaths in the European Union in 2008 and more than 10 000 deaths in the United States in 2009 (1, 2). Newly diagnosed MM is responsive to treatment with combinations of melphalan, prednisone, dexamethasone, doxorubicin, immunomodulatory drugs (IMiDs; such as thalidomide and lenalidomide), and proteasome inhibitors (PIs, such as bortezomib) (3) or autologous stem cell transplant following high-dose chemotherapy in appropriate patients (4, 5). However, most patients eventually relapse or become refractory to treatment, owing in part to the changing biology of the tumor and development of aggressive, drug-resistant phenotypes within the tumor. Although some agents used as initial therapy (including thalidomide, lenalidomide, and bortezomib) have also shown activity and improved outcomes in patients with relapsed or refractory MM (6 11), these responses are often of limited duration (12 14). Thus, there is an urgent unmet need to develop targeted agents that provide durable disease control and symptomatic relief in patients with MM that has relapsed or is refractory to currently approved agents. There are three distinct patient populations within the relapsed refractory MM setting: patients who are relapsed but not refractory to treatment, patients with primary refractory disease, and patients who are relapsed and refractory (15). Historically, the definition of relapsed vs. refractory disease was based on sensitivity to the vincristine, doxorubicin, and dexamethasone regimen, but the introduction of bortezomib, thalidomide, and lenalidomide has outdated this distinction. A more relevant definition of relapse is the presence of clinically active disease in patients who have received one or more prior therapies. Similarly, it has been suggested that refractory MM be defined as either progressive disease (PD) or stable disease (SD) while on prior therapy or PD within 3 months of the last dose of prior therapy. Patients with relapsed and refractory disease would be those who had achieved at least a minor response (MR) before disease progression within 60 d of the last treatment (15). This review is focused on the current management of patients with relapsed or refractory MM who have experienced disease progression within 60 d of the most recent treatment, although some of the studies reviewed ª 2010 John Wiley & Sons A/S 1

Emerging therapies in multiple myeloma Dimopoulos et al. may have used the older definitions (15). The aims of this review are to discuss the biology of the advanced stages of the disease, examine the current therapeutic options for patients, and review clinical data on currently approved and emerging treatment options for patients who have relapsed or become refractory to treatment. Biology of relapsed/refractory multiple myeloma Multiple myeloma is characterized by the accumulation of clonally identical plasma cells in the bone marrow that appear to develop from post-germinal center B cells (16). Current criteria for the diagnosis of MM have been summarized elsewhere (17). The growth, survival, adhesion, migration, and apoptotic resistance of MM cells are mediated by a large number of cytokines and adhesion molecules found in the bone marrow and tumor microenvironment (18). Very late antigen 4 and intercellular adhesion molecule 1 are important for the adhesion of MM cells to extracellular matrix bone marrow stromal cells (BMSCs) (19, 20). Interleukin-6 (IL-6), IL-21, tumor necrosis factor-a (TNF-a), insulin-like growth factor 1, vascular endothelial growth factor (VEGF), and stromal cell-derived factor 1-a have been shown to mediate MM cell survival, growth, and or resistance to apoptosis (18). In addition to clonal expansion of myeloma cells, the complex interplay of soluble factors in the bone marrow microenvironment and various receptor-mediated signaling pathways on BMSCs, osteoblasts osteoclasts, and myeloma cells [e.g., the receptor activator of nuclear factor-kappa B (NF-jB) osteoprotegerin system and NF-jB pathway] mediates the development of destructive bone disease and potentially life-threatening hypercalcemia (21, 22). Surrogate cytokine markers of time-to-event outcomes have been investigated in relapsed refractory MM and may have prognostic potential. For example, low baseline levels of VEGF were shown to be an independent prognostic factor for reduced response and shorter progression-free survival (PFS) in patients treated with thalidomide (23). Different somatic genetic abnormalities reflect the complex biology and pathogenesis of MM and have prognostic value in patients with newly diagnosed MM. Chromosomal translocations such as t(4;14) and the 17p13 deletion (del17p13; associated with low expression of TP53 gene) are associated with early relapse in newly diagnosed patients treated with high-dose therapy (24). The presence of t(4;14) and deletion of chromosome 13 (del13) have been associated with a significantly lower likelihood of response (defined as a >90% reduction in M-protein concentration) to up-front thalidomide plus dexamethasone in newly diagnosed MM patients (25). Similarly, the combination of lenalidomide and dexamethasone was not able to overcome the adverse prognostic effects of del(13) or t(4;14) in patients with relapsed refractory MM (26). The influence of cytogenetics has also been examined in patients with relapsed refractory MM, with varying results. As an example, bortezomib in combination with doxorubicin and dexamethasone showed comparable activity in relapsed refractory patients with or without del13q (27). In contrast, relapsed refractory MM patients treated with lenalidomide plus dexamethasone exhibited comparable time to progression (TTP) and overall survival (OS) regardless of del13q or t(4;14) status, whereas patients with del17p13 experienced worse time-to-event outcomes (28). Similarly, the presence of a non-hyperdiploid karyotype, other poor-risk cytogenetic abnormalities [i.e., presence of del13q, del17p, add1q21, t(4;14), or t(14;16)], and thalidomide-refractory disease were associated with reduced responses [less than a partial response (PR)] to treatment with lenalidomide and dexamethasone alone or in combination with bortezomib (29). These findings suggest that the presence of high-risk karyotypic abnormalities may define subsets of patients more likely to benefit from targeted therapies. Furthermore, new agents that improve PFS or OS in high-risk patients are of particular interest in the treatment of relapsed refractory MM. Current treatment options for relapsed/ refractory multiple myeloma Several factors should be considered in the selection of appropriate treatment options for patients with relapsed refractory MM, including response to prior therapies, type of relapse (e.g., aggressive), and individual patient characteristics (e.g., comorbidities, life expectancy, and quality of life) (30). Currently, multiple targeted agents, such as IMiDs and bortezomib, are approved for the treatment of relapsed refractory MM. Immunomodulatory drugs Before the advent of immunomodulatory drugs, few effective treatment options were available for patients with relapsed refractory MM. As an example, vincristine in combination with doxorubicin and dexamethasone (VAD) was associated with overall response rates (ORR) ranging from 25% to 61% in patients with relapsed refractory MM.(31 34) Although the VAD regimen was an improvement over high-dose melphalan (35, 36), the clinical benefits were limited, with response duration and OS 12 months (32 34). Thus, the development of novel targeted agents with tumor-specific mechanisms of action is an important advance in the treatment of 2 ª 2010 John Wiley & Sons A/S

Dimopoulos et al. Emerging therapies in multiple myeloma relapsed refractory MM, as it has led to a significant improvement in 5-yr survival (increasing from approximately 29% in 1990 1992 to 35% in 2002 2004, P < 0.001) and 10-yr survival rates (increasing from approximately 11 17% during the same time periods, P < 0.001) (37). Thalidomide and lenalidomide Immunomodulatory drugs target similar pathways including inhibition of cytokine expression (e.g., IL-6, TNF-a) by BMSCs and inhibition of angiogenesis that contribute to decreased growth of MM cells (38). For example, thalidomide stimulates T lymphocytes with IL- 2 and interferon-c release and subsequent NK cell activation, leading to the destruction of myeloma cells (38). A review of the literature shows that treatment of relapsed refractory MM with thalidomide (alone or in combination with dexamethasone) is associated with ORR ranging from 25% to 65% (39). When combined with bortezomib, melphalan, and prednisone or dexamethasone, thalidomide produced ORR ranging from 55% to 67% (40 42). However, peripheral neuropathy occurred in 6 16% of relapsed refractory MM patients who received thalidomide alone or in combination with high-dose chemotherapy and was the primary cause of thalidomide dose reduction (39). Moreover, an increased incidence of thromboembolic events was observed, particularly when thalidomide was given in combination with dexamethasone and doxorubicin (39). Consequently, the thalidomide analog lenalidomide was developed in an effort to reduce the toxicity associated with thalidomide, while maintaining or improving its efficacy. Studies have shown that combined treatment of relapsed refractory MM with lenalidomide and dexamethasone was associated with an ORR [complete response (CR) PR] of approximately 60%, significantly improved TTP, and significantly longer OS, regardless of previous exposure to thalidomide (8, 9). Although low rates of grade 3 peripheral neuropathy were observed (<2% of patients), venous thromboembolic events were reported in approximately 10 15% of patients receiving lenalidomide plus dexamethasone (8, 9). More recently, lenalidomide has shown clinical activity in combination with bortezomib and dexamethasone in patients with relapsed refractory MM. In a phase II study in 64 patients (77% had previously received thalidomide and 55% had received bortezomib therapy), the combination of lenalidomide (15 mg on days 1 14), bortezomib (1.0 mg m 2, days 1, 4, 8, and 11), and dexamethasone (40 mg 20 mg, cycles 1 4 5 8, days of after bortezomib dosing) produced an objective response (21% CR near CR, 68% at least PR, 84% at least MR) in 62 evaluable patients, irrespective of high-risk disease features and prior therapies (43). The median duration of response was 24 wk. Adverse events (AEs) were generally manageable (primarily grade 1 2 myelosuppression); two patients developed deep vein thrombosis while receiving aspirin prophylaxis, two patients had grade 3 atrial fibrillation, and one patient experienced grade 3 peripheral neuropathy. Other recent clinical studies in patients with relapsed refractory MM suggest that the ORR to lenalidomide in combination with targeted agents (e.g., bevacizumab, dacetuzumab, and dasatinib) may vary widely depending on the patient population and molecular drug target involved (44 47). Proteasome inhibitors In MM, it is thought that the ubiquitin proteasome system may affect tumor growth and progression via proteolysis of key proteins, including NF-jB signaling pathways; proapoptotic caspases; and various cytokines involved in the regulation of tumor cell growth, apoptosis resistance, and angiogenesis (48). Bortezomib The reversible PI bortezomib affects expression of a number of proteins involved in cell cycle arrest and apoptosis (e.g., NF-jB, caspase-9) (49). In patients with relapsed refractory MM, bortezomib alone significantly improved TTP (6.2 vs. 3.5 month; hazard ratio, 0.55; P < 0.001) and the response rate (CR + PR, 38% vs. 18%; P < 0.001) compared with high-dose dexamethasone (50). Alternating combination regimens have been explored in an effort to improve the response to bortezomib-based salvage therapy. In a study in 20 patients with relapsed refractory MM (30% had previously received bortezomib; 5% had received previous IMiD therapy), of bortezomib (1.3 mg m 2, days 1, 4, 8, and 11) in combination with melphalan (9 mg m 2, days 1 4), prednisone (60 mg m 2, days 1 4), and doxorubicin (conventional, 40 mg m 2 on day 1; liposomal, 30 mg m 2 on day 1) were alternated with of thalidomide (200 mg daily, days 1 28) in combination with cyclophosphamide (50 mg daily, days 1 28) and dexamethasone (40 mg daily, days 1 4). This approach resulted in an ORR of 95% (immunofixation-negative CR, 42%; near CR, 16%; PR, 47%) in nine evaluable patients, including CR in three of seven patients (42%) with highrisk cytogenetic abnormalities [e.g., t(4;14) or delrb] (51). The use of alternating combination therapy regimens was associated with manageable toxicities, including grade 3 thrombocytopenia (30%), neutropenia (30%), and infection (16%); grade 1 2 peripheral neuropathy occurred in three patients (15%) (51). The synergistic effects of combined treatment with bortezomib and pegylated liposomal doxorubicin (PLD; ª 2010 John Wiley & Sons A/S 3

Emerging therapies in multiple myeloma Dimopoulos et al. a novel formulation of doxorubicin with improved cardiac safety) (52) have been evaluated in patients with relapsed refractory MM. In a phase III study in bortezomib-naive patients (none had progressed on anthracycline-based therapy), 21-d cycles of bortezomib (1.3 mg m 2, days 1, 4, 8, and 11) in combination with PLD (30 mg m 2 on day 4) produced a significantly greater quality of response [i.e., CR + very good partial response (VGPR) rate] (27% vs. 19%; P = 0.0157), TTP (9.3 vs. 6.5 month; hazard ratio, 1.82; P = 0.000004), duration of response (10.2 vs. 7.0 month; P = 0.0008), and 15-month survival (76% vs. 65%; P = 0.03) compared with bortezomib alone (11). Several subgroup analyses have also been conducted in this study population to determine whether patientrelated factors influence the response to treatment. These analyses have shown that the significantly longer TTP seen with the bortezomib-pld regimen (relative to bortezomib alone) is consistent, even in relapsed refractory MM patients with prior IMiD exposure, prior stem cell transplant, and poor prognostic factors (e.g., serum beta-2 microglobulin 5.5 mg ml, refractory disease) (53 55). Moreover, the bortezomib-pld regimen is associated with significant improvements in TTP in both elderly (276 vs. 205 d; hazard ratio, 1.82; P = 0.0056) and younger patients (295 vs. 190 d; hazard ratio, 1.75; P = 0.0008) (56). Importantly, combined treatment with bortezomib and PLD did not increase the incidence of grade 3 cardiac events, thromboembolic events, or peripheral neuropathy compared with bortezomib alone (11, 54). Further investigations should establish the clinical benefits of bortezomib-based combinations with liposomal doxorubicin in elderly and high-risk MM patients. Investigational options for relapsed/refractory multiple myeloma Although currently available agents can provide clinical benefit in relapsed MM, not all patients will respond, and even those who do respond will ultimately relapse or become refractory to salvage therapy. Consequently, several new agents from a range of therapeutic classes are being examined in the relapsed refractory setting. Specific agents in development for the treatment of bortezomib- or lenalidomide-resistant MM include new IMiDs (e.g., pomalidomide), second-generation PIs (e.g., carfilzomib, NPI-0052), the signal transduction modulator perifosine, monoclonal antibody therapy (e.g., elotuzumab), and histone deacetylase (HDAC) inhibitors (e.g., panobinostat, romidepsin, and vorinostat). Although the goal with all of these newer agents is to improve patient outcomes, the rationale for use in MM varies with the drug class. Immunomodulatory drugs Pomalidomide Pomalidomide, an IMiD derived from thalidomide, has demonstrated greater activity in vitro (e.g., inhibition of osteoclast formation, cell cycle arrest) than thalidomide (57, 58). In a phase I II study, pomalidomide (2 5 mg daily on days 1 21 of each 28-d cycle) demonstrated a 38% ORR and up to 46% SD when administered alone or in combination with dexamethasone in 32 patients with relapsed refractory MM (59). In a phase II study in 60 relapsed MM patients (62% had received prior thalidomide or lenalidomide treatment), the combination of pomalidomide (2 mg daily during each 28-d cycle) and dexamethasone (40 mg daily, days 1, 8, 15, and 22) resulted in an ORR of 63% (5% CR, 28% VGPR, and 30% PR), including confirmed responses in 74% of patients classified as high risk (60). The primary toxicity was grade 3 myelosuppression; grade 3 neuropathy and a thromboembolic event were each reported in a single patient. Taken together, these studies suggest that pomalidomide may overcome resistance to the IMiDs currently used as initial or second-line therapy, with a lower incidence of neurotoxic and thromboembolic events. Preliminary results from recent clinical studies of pomalidomide are summarized in Table 1. Proteasome inhibitors The rationale for developing new PIs is similar to the rationale for developing new IMiDs: potential improvements in efficacy and or tolerability and potentially incomplete cross-resistance within the drug class. Carfilzomib In patients who have become resistant to bortezomib, the use of a new PI with a different chemical backbone could overcome this resistance. Carfilzomib is a secondgeneration PI that is structurally similar to epoxomicin. Unlike bortezomib, which has a reversible effect, carfilzomib irreversibly targets the same proteasomal subunit (20S chymotrypsin-like b5 subunit) and has shown activity (e.g., caspase activation, inhibition of proliferation) against bortezomib-resistant MM cell lines, as well as cells from MM patients with clinical evidence of bortezomib resistance (61). In a phase I study, 19 patients who had relapsed following or became refractory to previous bortezomib and IMiD therapy received carfilzomib 15 27 mg m 2 on days 1, 2, 8, 9, 15, and 16 of each 28-d cycle. Treatment with carfilzomib resulted in an ORR of approximately 17%, with 33% of patients achieving an MR or better. No treatment-related or newly emergent peripheral neuropathy was reported in response to carfilzomib (62). Two 4 ª 2010 John Wiley & Sons A/S

Dimopoulos et al. Emerging therapies in multiple myeloma Table 1 Targeted agents in clinical investigation for the treatment of relapsed refractory multiple myeloma Agent Author N n Dosing regimen Confirmed responses, % Time-to-event outcome Most common toxicities, % Pomalidomide Lacy (116) 34 Richardson (71) Carfilzomib Niesvizky (117) 32 20 Siegel (118) 35 33 Wang (119) 57 51 NPI-0052 Richardson (66) 27 Perifosine Richardson (59) 84 73 Tanespimycin Richardson (87) 72 Badros (86) 22 Panobinostat Berenson (99) 15 12 San Miguel (101) 29 28 POM 2 mg daily, days 1 28 DEX 40 mg, days 1, 8, 15, 22 POM 2 5 mg, days 1 21 POM 2 5 mg, days 1 21 DEX 40 mg, weekly (after 4 cycles for lack of response or PD) CFZ 15 27 mg m 2, days 1, 2, 8, 9, 15, 16 LEN 10 27 mg, days 1 21 DEX 40 mg, days 1, 8, 15, 22 (monthly after cycle 5) CFZ 20 mg m 2, days 1, 2, 8, 9, 15, 16 CFZ 20 mg m 2, days 1, 2, 8, 9, 15, 16 NPI 0.025 0.7 mg m 2, days 1, 8, 15 21-d cycles PER 50 mg daily BTZ 1.3 mg m 2, days 1, 4, 8, 11 DEX 20 mg, day of and day after BTZ for PD 21-d cycles TSP 100 340 mg m 2, days 1, 4, 8, 11 BTZ 0.7 1.3 mg m 2, days 1, 4, 8, 11 21-d cycles TSP 50 340 mg m 2, days 1, 4, 8, 11 BTZ 1.3 mg m 2, days 1, 4, 8, 11 PAN 20 mg, days 1, 3, 5, 8, 10 MLP 0.05 mg kg, days 1, 3, 5 21-d cycles PAN 10 30 mg, 3 times weekly BTZ 1.3 mg m 2, days 1,4, 8, 11 ORR, 26 (all PR) NR Grade 3 4 Neutropenia, 21; anemia, 12; thrombocytopenia, 9; fatigue, 9; non-infectious pneumonitis, 3; hyperglycemia, 3; edema, 3; skin rash, 3; no TEEs observed POM alone: ORR MR, 38 POM + DEX: ORR MR, 38 DOR, 11 wk TTP, 8.3 wk DOR, 14.2 wk TTP, 20 wk Grade 3 4 Neutropenia, thrombocytopenia CR VGPR PR, 55 NR Grade 3 4 Thrombocytopenia 15; anemia 15; neutropenia 8 CR PR, 18 NR Grade 3 4 Anemia, 14; neutropenia, 11; peripheral neuropathy, 3 CR VGPR PR, 45 NR Grade 3 4 Thrombocytopenia, 9; fatigue, 9; neutropenia, 7; lymphopenia, 7; anemia, 5; pneumonia, 5; hyperglycemia, 5 NR 1 unconfirmed PR (71% flm-protein after 3 cycles) 8SD ORR PR, 38 CR PR, 20 ORR MR: BTZ-naive (n = 21), 48 BTZ-pretreated (n = 23), 22 BTZ-refractory (n = 23), 13 NR DLTs observed (grade 3 fatigue, mental status change and loss of balance, 1 patient each at highest dose) TTP, 6.4 mo OS, 22.5 mo Grade 3 4 Thrombocytopenia; neutropenia; anemia; hyponatremia; diarrhea ( 5% each) DOR, 12 mo Grade 3 4 thrombocytopenia, 25; neutropenia, 3 VGPR PR MR, 14 NR Grade 3 4 Thrombocytopenia, 27; neutropenia, 18; peripheral neuropathy 5 CR PR, 33 NR Grade 3 4 Neutropenia; thrombocytopenia CR PR, 50 NR Grade 3 4 Thrombocytopenia; neutropenia; anemia; pneumonia; fatigue; significant QT c did not occur ª 2010 John Wiley & Sons A/S 5

Emerging therapies in multiple myeloma Dimopoulos et al. Table 1 (Continued ) Agent Author N n Dosing regimen Confirmed responses, % Time-to-event outcome Most common toxicities, % Romidepsin Harrison (105) 25 18 Vorinostat Jagannath (110) 34 9 Siegel (112) 28 25 Voorhees (113) 9 7 RMD 8 14 mg m 2, days 1, 8, 15 BTZ 1.3 mg m 2, days 1,4, 8, 11 DEX 20 mg, days 1, 2, 4, 5, 8, 11, 12 VOR 200 mg BID; or 400 mg, days 1 14 BTZ 0.7 or 0.9 mg m 2, days 4, 8, 11, 15; or 0.9 1.3 mg m 2, days 1, 4, 8, 11 DEX 20 mg, days 1 4, 9 12 for PD VOR 300 400 mg, daily days 1 7, 15 21 LEN 10 25 mg, days 1 21 DEX 40 mg, days 1, 8, 15, 22 VOR 200 400 mg daily, days 4 11 BTZ 1.3 mg m 2, days 1,4, 8, 11 PLD 30 mg m 2, day 4 CR ncr VGPR PR, 67 NR Dose limiting No DLTs at MTD (romidepsin 10 mg m 2 ); no reports of QT c prolongation PR MR, 78 TTP, 9.8 mo Grade 3 4: Neutropenia; drug-related toxicities included diarrhea, nausea, and fatigue (all grades) CR ncr VGPR PR, 64 NR Dose limiting Grade 3 diarrhea in 1 patient at highest dose (VOR 400 mg) CR VGPR PR, 86 NR Grade 3 4 Sensory neuropathy; neutropenia; lymphopenia; thrombocytopenia BID, twice daily; BTZ, bortezomib; CFZ, carfilzomib; CR, complete response; DEX, dexamethasone; DLT, dose-limiting toxicity; DOR, duration of response; LEN, lenalidomide; MLP, melphalan; MR, minimal response; MTD, maximum tolerated dose; ncr, near complete response; NPI, NPI-0052; NR, not reported; ORR, overall response rate; OS, overall survival; PAN, panobinostat; PER, perifosine; PD, progressive disease; PLD, pegylated liposomal doxorubicin; POM, pomalidomide; PR, partial response; QT c, corrected QT interval; RMD, romidepsin; SD, stable disease; TEE, thromboembolic events; TSP, tanespimycin; TTP, time to progression; VGPR, very good partial response; VOR, vorinostat. ongoing phase II trials are investigating the efficacy, safety, and tolerability of carfilzomib as monotherapy in patients with relapsed refractory MM and prior treatment with bortezomib and thalidomide or lenalidomide (63, 64). Preliminary data from ongoing clinical studies are summarized in Table 1. NPI-0052 Like carfilzomib, the non-peptide-based inhibitor NPI-0052 also targets all three proteasome units (i.e., the caspase-, chymotrypsin-, and trypsin-like subunits) and irreversibly inhibits the 20S proteasome (65). In a phase I study, NPI-0052 (0.025 0.075 mg m 2, days 1, 8, and 15 of a 28-d cycle) exhibited more potent proteasome inhibitory activity than bortezomib, with no reports of peripheral neuropathy or myelosuppression in patients (N = 27) with relapsed refractory MM (Table 1) (66). Inhibitors of signal transduction and cell adhesion Although conventional and targeted agents have dramatically improved response rates, MM remains incurable. As noted earlier, the interactions of MM cells within the bone marrow microenvironment are complex and depend on a number of cell ligand and cell cell interactions that activate signal transduction processes controlling cell migration, growth, and survival. Signal transduction modulators affect a variety of cellular processes, including cell growth, differentiation, and death, making them rational targets for new therapies. Perifosine Perifosine is thought to target cell membranes and indirectly affect the phosphatidylinositol 3-kinase Akt pathway, which is a critical regulator of cell survival and cell growth and may underlie the pathogenesis of resistance to conventional agents (e.g., dexamethasone, doxorubicin) in MM (67, 68). In a phase I dose-escalation study in 32 heavily pretreated patients (94% received prior dexamethasone, 83% prior thalidomide, and 47% prior bortezomib therapy), treatment with perifosine (50 or 100 mg daily during a 28-d cycle) in combination with lenalidomide (15 or 25 mg, days 1 21) and dexamethasone (20 mg, days 1 4, 9 12, and 17 20 for 4 cycles; days 1 4 thereafter) resulted in a 50% ORR (PR or better) in evaluable patients (n = 30) (69). Patients who achieved a PR or better exhibited a longer median TTP (31 vs. 23 wk in all evaluable patients). The most common grade 3 5 AEs were neutropenia, hypophosphatemia, thrombocytopenia, anemia, and fatigue (69). 6 ª 2010 John Wiley & Sons A/S

Dimopoulos et al. Emerging therapies in multiple myeloma In a phase II study in 64 patients with relapsed or relapsed refractory MM (95% received prior dexamethasone, 89% prior thalidomide, 73% prior bortezomib, and 30% prior lenalidomide), perifosine alone (150 mg daily for a 21-d cycle) showed modest clinical activity, producing best responses [according to European Blood and Marrow Transplant (EBMT) criteria] of MR (n = 1) and SD (n = 22).(70) However, when administered in combination with dexamethasone 20 mg twice weekly to patients with PD, perifosine showed greater clinical activity (38% PR + MR) in 12 of 31 evaluable patients; an additional 15 patients (47%) achieved SD (70). The most common grade 3 5 AEs were nausea, vomiting, fatigue, anemia, increased creatinine, and reversible neutropenia. Peripheral neuropathy and deep vein thrombosis were not reported (70). Perifosine has also been evaluated in combination with bortezomib and dexamethasone in 84 relapsed refractory MM patients previously treated with bortezomib (71). As shown in Table 1, this regimen was associated with an ORR of 38% (CR PR, 20%) and an OS of 22.5 months (median not yet reached); myelosuppression, hyponatremia, and diarrhea were the most common grade 3 5 events. Elotuzumab Further improvements in the management of relapsed and refractory MM may be achieved using monoclonal antibody (MAb) therapy. Elotuzumab (HuLuc63) is a humanized MAb that targets CS1, a cell surface glycoprotein involved in cell adhesion that is selectively expressed on MM cells and colocalizes with CD138 in these cells (72, 73). High rates of tumor cell lysis were observed when CD138+ cells isolated from patients with refractory (and newly diagnosed) MM were treated with elotuzumab in the presence of autologous peripheral blood mononuclear cells, including natural killer cells (72). Importantly, elotuzumab-induced tumor cell lysis was enhanced in MM cells that had been pretreated with subtherapeutic doses of diverse types of targeted agents (i.e., bortezomib, lenalidomide, perifosine) (72, 74). These promising preclinical findings have been validated in early clinical trials in patients with relapsed refractory MM. In a phase I study in 28 patients with relapsed refractory MM (31% had received prior bortezomib), 21-d cycles of elotuzumab (2.5 20 mg kg, days 1 and 11) in combination with bortezomib (1.3 mg m 2, days 1, 4, 8, and 11) produced a best response ( MR) of 60% (40% PR) in 20 evaluable patients who had completed at least 2 treatment cycles (75). Elotuzumab has also been evaluated in combination with lenalidomide in a phase I II study in 29 patients (69% had received prior bortezomib, 59% received thalidomide, and 21% received lenalidomide). Treatment with elotuzumab (5 20 mg kg weekly for the first 2 28-d cycles, then every other week) combined with lenalidomide (25 mg, days 1 21) produced an ORR of 82% (18% VGPR; 64% PR) in 28 evaluable patients (76). Interestingly, an ORR of 95% (23% VGPR; 73% PR) was seen in the 22 lenalidomide-naive patients enrolled in the study. Further investigation is needed to determine the optimal role of elotuzumab in the treatment of MM. High-dose chemotherapy and targeted agents Bendamustine, a bifunctional alkylating agent that crosslinks DNA and induces apoptosis and mitotic catastrophe (77), may be another option for salvage therapy in patients with relapsed refractory MM. This agent has shown some clinical activity as monotherapy (ORR, 36 55%) (78, 79), prompting the evaluation of combination regimens in the treatment of MM. In a phase I study in 28 evaluable patients with relapsed refractory MM (14% had received prior bortezomib and 7% received thalidomide), of bendamustine (60 mg m 2, days 1, 8, and 15) in combination with prednisolone (100 mg, days 1, 8, 15, and 22) and thalidomide (50, 100, or 200 mg, days 1 28) produced an ORR of 86% (CR + PR), including those patients who had relapsed on prior conventional chemotherapy or high-dose chemotherapy and autologous stem cell transplant (SCT) (80). Overall, the median duration of response was 11 months, and median OS was 19 months; however, OS was longer in patients who had relapsed on prior chemotherapy (32+ vs. 16 month; P = 0.03) compared with SCT (80). The most common grade 3 AEs were hematologic in nature, and thromboembolic events were not observed (80). The feasibility of adding bendamustine to bortezomib and dexamethasone therapy has been explored in patients with <MR to 1 cycle of bortezomib plus dexamethasone. In this study, a total of seven patients with relapsed refractory MM who failed to respond adequately to bortezomib plus dexamethasone received 21-d cycles of bortezomib (1.3 mg m 2, days 1, 4, 8, and 11) combined with dexamethasone (40 mg, days 1, 4, 8, and 11) and bendamustine (50 100 mg m 2, days 1 and 8). In this non-responding patient population, the combination resulted in an ORR of 86% (57% PR; 29% MR) (81). Further clinical trials are needed to establish the role of bendamustine alone and in combination with other targeted agents in the treatment of relapsed refractory MM. Targeted inhibition of heat shock protein Novel treatment approaches targeting diverse pathways complementary to those targeted by conventional and ª 2010 John Wiley & Sons A/S 7

Emerging therapies in multiple myeloma Dimopoulos et al. newer approved agents show promise for inducing myeloma cell cytotoxicity and downregulating signaling pathways that induce myeloma growth, survival, and therapeutic resistance. Heat shock proteins (e.g., HSP27, HSP90) are potential therapeutic targets because expression of these Bcl-2-like proteins interferes with the mitochondrial stress response and activation of proapoptotic signaling (e.g., activation of Bax, caspase-3) that can result in the development of drug resistance (82, 83). For example, overexpression of HSP27 correlated with resistance to dexamethasone in myeloma cells, whereas blockade of HSP27 restored sensitivity to bortezomib (84, 85). Tanespimycin Tanespimycin, an inhibitor of HSP90, has shown activity in combination with bortezomib in MM patients (Table 1) (86, 87). In a phase I II study in 72 pretreated MM patients (74% had received prior bortezomib therapy and 69% had received prior lenalidomide), tanespimycin [340 mg m 2 intravenously (IV), days 1, 4, 8, and 11 of each 21-d cycle] in combination with bortezomib (0.7 1.3 mg m 2 IV, days 1, 4, 8, and 11) inhibited HSP90 and proteasome activity and showed antitumor activity based on modified EBMT criteria. The ORR (defined as MR or better) was 48% in bortezomib-naive patients, 22% in bortezomib-pretreated patients, and 13% in bortezomib-refractory patients, with a median response duration of 12 months. There were no reports of grade 3 5 peripheral neuropathy (87). In another study in 22 heavily pretreated MM patients (96% had received prior thalidomide therapy), the combination of tanespimycin (50, 175, or 340 mg m 2, days 1, 4, 8, and 11 of each 21-d cycle) with bortezomib (1.3 mg m 2, days 1, 4, 8, and 11) demonstrated clinical activity, with 14% of patients achieving MR or better. The most common grade 3 5 AEs were hematologic in nature, and one patient experienced grade 3 peripheral neuropathy (86). Histone deacetylase inhibitors Histone deacetylase inhibitors are another new class of molecules that show promise as a complementary approach for the treatment of relapsed refractory MM, and a number of phase I and II studies have recently been conducted with these agents. HDAC inhibition promotes acetylation of histone and non-histone proteins (Fig. 1). Histone acetylation affects higher-order DNA chromatin structure, and HDAC inhibition leads to increased transcription of genes that have been downregulated by histone acetylation (88). Therefore, inhibition of HDAC affects epigenetic mechanisms that help restore or increase expression of genes that may play a critical role in the control of tumor growth and survival. As is the case with PIs, HDAC inhibitors may help restore or increase the expression of proapoptotic proteins in tumors. Non-histone proteins are also regulated by acetylation, with evidence for non-histone-mediated effects on tumor cell growth. Transcription factor acetylation disrupts control of cell cycle transit and apoptosis in cancer. Direct acetylation of p53 affects its growth-regulatory and proapoptotic functions. Treatment of a variety of Condensed chromatin HAT Ac Decondensed chromatin Ac Ac Ac Gene transcription activation/repression HDAC HDACi Deacetylated protein HDAC HAT Ac Ac Acetylated protein Ac Ac Transcription factors E2F p53 NF-κB STAT-1 Non-histone proteins VEGF hsp90 hif1α α-tubulin Apoptosis Cell cycle arrest Immune modulation Angiogenesis inhibition Figure 1 Effects of histone deacetylase (HDAC) inhibitors on histone protein acetylation and chromatin structure, acetylation of transcription factors resulting in changes in gene expression, and acetylation of other non-histone proteins leading to diverse biologic effects underlying the pathogenesis and treatment of multiple myeloma. Reprinted with permission. Paik PK, Krug LM. HDAC inhibitors in malignant pleural mesothelioma: preclinical rationale and clinical trials. J Thorac Oncol 2010; 5: 275 279. 8 ª 2010 John Wiley & Sons A/S

Dimopoulos et al. Emerging therapies in multiple myeloma tumor cells with HDAC inhibitors resulted in hyperacetylation of p53 and induction of p21 Waf Cip1 mediated cell cycle arrest, increased expression of proapoptotic proteins (e.g., cytochrome c, BAX, Bid, activated caspase), and downregulated expression of antiapoptotic proteins (e.g., Bcl-2) (89 91). In addition, increased acetylation of HSP90 can disrupt its chaperone function, resulting in decreased intracellular levels of progrowth and antiapoptotic proteins (e.g., Akt), possibly through enhanced proteasomal degradation of proteins (92, 93). HDAC6 links acetylation of HSP90 with aggresome formation and the accumulation of ubiquinated proteins (94, 95). Panobinostat Panobinostat (LBH589) is an oral HDAC inhibitor (96) that is currently being investigated alone (97) and in combination with lenalidomide and dexamethasone (98), melphalan (99), or bortezomib for the treatment of patients with relapsed or relapsed refractory MM (100, 101). In a phase II study, single-agent panobinostat showed clinical activity with a durable VGPR and MR (based on EBMT criteria) in 2 of 38 patients with heavily pretreated (including bortezomib, lenalidomide, and thalidomide) refractory MM, and there were no reports of significant thromboembolic events (97). Panobinostat is also being investigated as a component of combination regimens for the treatment of relapsed or refractory MM (summarized in Table 1). In a phase I study, panobinostat in combination with melphalan demonstrated clinical activity with an ORR of 33% (one immunofixation-positive CR, three PR) in 12 patients with relapsed or refractory MM previously treated with melphalan (99). The most common grade 3 5 AEs were reversible neutropenia (n = 6) and thrombocytopenia (n = 6) (99). Combination therapy with panobinostat and bortezomib is also being investigated in patients with advanced MM. In a phase IB study in 29 heavily pretreated patients (55% had received prior bortezomib therapy), treatment with panobinostat combined with bortezomib resulted in at least a PR in 14 of 28 evaluable patients (50%), including four patients with immunofixationnegative CR (101). Importantly, an objective response (PR + MR) was observed in 6 of 10 (60%) evaluable patients who were refractory to previous bortezomib therapy (101). The most common grade 3 5 AEs were thrombocytopenia (n = 25), neutropenia (n = 18), and anemia (n = 6) (101). Romidepsin Romidepsin, an HDAC inhibitor administered as a 4-h infusion, is approved for the treatment of patients with relapsed or refractory cutaneous T-cell lymphoma who have received at least 1 prior systemic therapy (102). In patients with relapsed or refractory MM, romidepsin is currently being evaluated in combination with bortezomib (103, 104) or with bortezomib and dexamethasone (105). In a phase I study in relapsed or refractory MM patients (N = 25; 37% had received prior vincristine, 50% received prior thalidomide, and 25% received prior bortezomib therapy), treatment with romidepsin at the maximum tolerated dose (MTD) of 10 mg m 2 in combination with bortezomib resulted in an ORR of 71% (based on EBMT criteria), including one CR, three PR, and one MR among seven evaluable patients (103). No grade 3 or 4 non-hematologic AEs or dose-limiting toxicities were reported (103). In a phase I II study, romidepsin was shown to have clinical activity in combination with bortezomib and dexamethasone in patients with relapsed or refractory MM (Table 1) (105). No dose-limiting toxicities were reported among seven evaluable patients who completed at least two cycles (range, 1 8) of treatment with romidepsin 8 or 10 mg m 2 once weekly; however, grade 3 fatigue (n = 2), peripheral neuropathy (n = 1), neutropenia (n = 1), and sepsis (n = 2) were reported in this small cohort of patients (105). Importantly, 12 of 18 evaluable patients (67%) experienced at least a PR (four CR near CR, four VGPR, four PR); five additional patients (28%) achieved an MR (105). Of the seven patients receiving long-term maintenance therapy with romidepsin (10 mg m 2 on days 1 and 8 of every 28-d cycle), four experienced disease progression, including three who had progressed on a previous bortezomib maintenance regimen. Vorinostat Vorinostat is an oral HDAC inhibitor that was approved in the United States in 2006 for the treatment of patients with cutaneous T-cell lymphoma who have progressive, persistent, or recurrent disease on or following 2 systemic therapies (106 108). In patients with relapsed refractory MM, vorinostat is currently being investigated in combination with bortezomib (109 111), in combination with lenalidomide and dexamethasone (112), or in combination with PLD and bortezomib (113). In a phase I study in 23 heavily pretreated patients (100% had received prior thalidomide; 83% had received prior bortezomib), vorinostat demonstrated clinical activity at the MTD of 400 mg daily on days 4 11 in combination with bortezomib 1.3 mg m 2 on days 1, 4, 8, and 11 of each 21-d cycle, with 55% of patients achieving PR or better (109). The most frequent grade 3 AEs were reversible myelosuppression and fatigue; 1 patient had grade 3 peripheral neuropathy. ª 2010 John Wiley & Sons A/S 9

Emerging therapies in multiple myeloma Dimopoulos et al. In another phase I study, the addition of vorinostat 200 400 mg daily on days 4 to 11 to PLD 30 mg m 2 on day 4 and bortezomib 1.3 mg m 2 on days 1, 4, 8, and 11 in 21-d cycles showed clinical activity in six of seven evaluable patients (1 CR, 1 VGPR, and 4 PR) based on International Myeloma Working Group criteria (114) and was generally well tolerated. No dose-limiting toxicities, serious AEs, or deaths were reported, although some neurologic (grade 3 sensory neuropathy in two of nine patients) and hematologic toxicities (grade 3 neutropenia, lymphopenia, and thrombocytopenia in 2, 3, and two patients, respectively) were identified in this small cohort of patients (113). Another study (summarized in Table 1) showed that extended treatment ( 12 cycles) with vorinostat 200 mg twice daily or 400 mg once daily in combination with bortezomib was well tolerated (one patient had grade 4 neutropenia, and five patients had grade 3 treatment-related AEs). Long-term clinical activity was observed, with five PR, two MR, and two SD among nine evaluable patients. The duration of PR ranged from 147 to 609 d (110). The safety and tolerability of vorinostat has been well documented in patients with hematologic malignancies and those with solid tumors. In an analysis of 341 patients who had received vorinostat as monotherapy, the most common grade 3 4 drug-related adverse events were fatigue (12% of patients) and thrombocytopenia (11%). In an analysis of 157 patients who had received vorinostat in combination, the most common grade 3 4 drug-related adverse event was fatigue (13% of patients). Furthermore, an analysis in more than 1845 vorinostattreated patients revealed that the rate of thromboembolic events related to treatment was <2.6% (115). Ongoing clinical studies include a pivotal phase IIB study and a randomized, blinded, phase III study to determine the ORR, TTP, PFS, OS, and tolerability of vorinostat in combination with IV bortezomib. Conclusions The development and application of targeted therapies, such as bortezomib and lenalidomide, have improved treatment outcomes in patients with relapsed refractory MM. Emerging studies suggest that new combination therapies targeting complementary signaling pathways may further improve prognosis in treating this advanced form of MM. Completion of the numerous ongoing clinical investigations should determine which if any of these newly emerging therapies are viable treatment options for patients with relapsed refractory MM. Regardless of outcome, the clinical study results will further improve our understanding of the biology of MM and the role for targeted therapies in providing durable disease control and symptomatic relief in MM patients. Disclosures Dr. Dimopoulos has received honoraria from Merck, Sharp, and Dohme, a subsidiary of Merck & Co., Inc., Celgene Corporation, and Centocor Ortho Biotech. Dr. Anderson has served as an advisor for Celgene Corporation, Novartis Oncology, Millenium Pharmaceuticals, Inc., Onyx Pharmaceuticals, and Nereus Pharmaceuticals. Dr. San-Miguel has served as an advisor for Celgene Corporation, Novartis Oncology, Millenium Pharmaceuticals, Inc., and Janssen-Cilag. Acknowledgements The authors wish to thank Craig Albright, PhD, for writing and editorial assistance during the development of this manuscript. 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