Current Advances in Non–Proteasome Inhibitor–Based Approaches to the Treatment of Relapsed/Refractory Multiple Myeloma

November 15, 2011

Multiple myeloma (MM) remains incurable despite the current approaches used in initial therapy, including more effective induction therapy, one or more autologous stem-cell transplants, and consolidation/maintenance strategies.

In the past decade, immunomodulatory drugs have been approved by the US Food and Drug Administration for the treatment of multiple myeloma (MM)-and a number of emerging agents that target the cellular pathways or proteins involved in the pathophysiology of MM are currently in development. Lenalidomide (Revlimid) and pomalidomide induce apoptosis and sensitize MM cells while demonstrating superior efficacy and better tolerability than thalidomide (Thalomid). Several novel classes of drugs, including the histone deacetylase (HDAC) inhibitors, heat shock protein (HSP) inhibitors, and monoclonal antibodies have been shown to have activity in myeloma in early-stage clinical trials. HDAC inhibitors, including vorinostat (Zolinza), panobinostat, and romidepsin (Istodax) are thought to affect multiple pathways involved in MM and correct the deregulation of genes involved in apoptosis and cell cycle arrest, thus potentially sensitizing MM cells to apoptosis. HSP inhibitors (eg, tanespimycin) decrease MM proliferation and suppress the long-term replicative potential of MM cells; they may also sensitize MM cells to other anticancer agents. The humanized monoclonal antibody elotuzumab induces antibody-dependent cell cytotoxicity–mediated apoptosis. It is likely that in the near future the treatment armamentarium for MM will undergo significant expansion as some of these additional target pathways become validated.

Introduction

Multiple myeloma (MM) remains incurable despite the current approaches used in initial therapy, including more effective induction therapy, one or more autologous stem-cell transplants, and consolidation/maintenance strategies. The improved survival of patients with myeloma and the incurable nature of the disease have led to an increased number of patients with relapsed disease who require treatment for continued control of their disease. Treatment of relapsed disease has improved considerably in the past decade and has resulted in improved outcomes in this group of patients.[1] Therapeutic advances during this decade have included thalidomide (Thalomid) and its immunomodulatory derivatives lenalidomide (Revlimid) and pomalidomide, as well as the proteasome inhibitor bortezomib (Velcade). Immunomodulatory drugs (IMiDs) represent a series of compounds that were developed based on thalidomide. Thalidomide was initially introduced as a sedative and used for morning sickness, but was withdrawn from the market in the early 1960s after it was found to be a teratogen. Given its antiangiogenic properties, it was evaluated in the treatment of several cancers, including myeloma. Based on promising initial results, thalidomide was evaluated in several phase II and phase III studies in the setting of relapsed and newly diagnosed disease. These studies demonstrated clear activity for the drug, and several combinations that include thalidomide have been developed since then. However, thalidomide was associated with significant toxicity, including peripheral neuropathy (PN), constipation, sedation, and deep venous thrombosis. Moreover, the teratogenic effect of the drug required that it be prescribed under strict regulations. Attempts to develop analogs with more acceptable toxicity and possibly improved activity led to the development of the IMiDs lenalidomide and pomalidomide.[2] More recently, several novel classes of drugs have been shown to have activity in myeloma in early-stage clinical trials. These include the histone deacetylase (HDAC) inhibitors, heat shock protein (HSP) inhibitors, mammalian target of rapamycin (mTOR) inhibitors, phosphoinositide 3-kinase (PI3k)/AKT inhibitors, and monoclonal antibodies. Here we review the recent developments and potential future options for management of relapsed/refractory MM.

Lenalidomide

Mechanisms of action

Lenalidomide is often referred to as an IMiD, but its mechanisms of action in myeloma have not been fully elucidated. It is likely that its antimyeloma activity is dependent on many effects, including direct cytotoxicity as well as indirect effects associated with modulation of the different cytokines, inhibition of angiogenesis, regulation of T-cell activity, and augmentation of natural killer (NK)-cell cytotoxicity.

Direct cytotoxic effects. Lenalidomide clearly has direct antiproliferative and apoptotic effects on MM cells, as has been shown with cell lines and primary myeloma cells.[3-5] Lenalidomide affects cyclin-dependent kinase (CDK) inhibitors through regulation of p21waf-1, a key cell-cycle regulator that modulates the activity of CDKs. Various mechanisms have been proposed for the apoptotic effects, including increased potentiation of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), inhibition of apoptosis protein-2, increased sensitivity to Fas-mediated cell death, upregulation of caspase-8 activation, down-regulation of caspase-8 inhibitors (FLIP, cIAP2), downregulation of nuclear factor kappa B (NFκB) activity, and inhibition of prosurvival effects of insulin-like growth factor (IGF)-1.[6] More recently, studies have shown that lenalidomide may act through CCAAT/enhancer binding protein (CEBP)-beta, a transcription factor that regulates the activity of interferon regulatory factor (IRF)-4, which appears to be a critical survival factor for myeloma cells.

Immunomodulatory effects. Lenalidomide inhibits the production of proinflammatory cytokines TNF-α, interleukin (IL)-1, IL-6, and IL-12 and increases the production of anti-inflammatory cytokine IL-10 from human peripheral blood mononuclear cells.[7,8] IL-6 is a critical cytokine in myeloma; it plays a role in cell proliferation and survival and is secreted both by myeloma cells and cells in the marrow microenvironment.[9] Lenalidomide down-regulates the production of IL-6 directly and also inhibits MM–bone marrow stromal cell (BMSC) interaction, leading to increased apoptosis of myeloma cells.[10,11] Lenalidomide is a much more potent inhibitor of TNF-α secretion-up to 50,000 times more potent than thalidomide-which may, to some extent, explain the differences in the clinical activity of these two agents.[2] Lenalidomide can lead to increased Th1-type cytokine response, resulting in increased expression of IL-2 and interferon (IFN)-γ, thereby enhancing T-cell and NK-cell-mediated lysis of myeloma cells.[7] CD28 is a co-stimulatory molecule that augments the T-cell response, and lenalidomide can induce tyrosine phosphorylation of CD28 on T cells, leading to activation of downstream targets such as PI3K, Grb2-SOS, and NFκB.[12,13] IMiDs have been shown to stimulate both cytotoxic CD8+ and helper CD4+ cells.[14] In vitro studies have shown an important role for NK-cell–mediated cytotoxicity against myeloma cells. Increased NK-cell numbers are seen following treatment with IMiDs and may be a result of increased IL-2 secretion.[15]

Antiangiogenic properties. Multiple in vitro and in vivo studies have demonstrated strong antiangiogenic properties of thalidomide and the IMiDs, but it remains unclear what the exact contribution of this effect is to antimyeloma activity.[16,17] Tumor-associated endothelial cells are more dependent on vascular endothelial growth factor (VEGF)-receptor signaling for growth and survival than are normal endothelial cells.[18,19] Lenalidomide significantly decreases the expression of angiogenic factors VEGF and IL-6 in MM, an effect that is still present when the myeloma cells are in the tumor microenvironment.[7,20] Apart from alteration in the levels of VEGF, lenalidomide partially inhibits AKT phosphorylation after VEGF stimulation in endothelial cells and also has inhibitory effects on phosphorylation of Gab1, a protein upstream of AKT1.[21,22] It is likely that multiple mechanisms underlie the antiangiogenic properties of lenalidomide and other IMiDs.

Effects on the microenvironment. In addition to the effect on myeloma cells and immune cells, lenalidomide can also have different effects on cells in the microenvironment. Interaction between osteoclasts and myeloma cells leads to increased production of IL-6 and other growth factors for MM cells as well as osteoclasts.[23] Lenalidomide affects the marrow microenvironment by decreasing the formation of osteoclasts.[10] It downregulates the important mediators of osteoclastogenesis, such as transcription factor PU.1 and pERK, and it reduces the levels of bone remodeling factor RANK. IMiDs also decrease cell surface adhesion molecules such as intercellular adhesion molecule (ICAM)-1, vascular cell adhesion molecule (VCAM)-1, and E-selectin, and they inhibit the adhesion of MM cells to bone marrow stromal cells.[24]

Adverse effects of lenalidomide

In general, lenalidomide is well tolerated and does not have many of the more bothersome adverse effects associated with thalidomide. There is no direct evidence of teratogenicity in humans, but lenalidomide’s structural similarity to thalidomide[25] calls for caution while using the drug in women with childbearing potential and in sexually active male patients.[26]

The most common grade 3 (or higher) adverse events reported in phase III trials were neutropenia, thromboembolic events, thrombocytopenia, anemia, and pneumonia.[27] Myelosuppression remains the most common problem associated with lenalidomide, with neutropenia much more common than thrombocytopenia and anemia. However, the frequency of febrile neutropenia is quite low.[28] Myelosuppression can usually be managed with growth factor support and/or lenalidomide dose reductions but may require discontinuation of treatment.[29]

The risk of venous thromboembolism (VTE) is low when lenalidomide is given as monotherapy but increases significantly when it is used in combination with dexamethasone, particularly high-dose dexamethasone, or when used with erythropoietic agents.[30-32] The risk is also higher when lenalidomide is used in combination with cytotoxic chemotherapy, particularly anthracyclines.[32] The incidence of VTE was 16% in patients treated with lenalidomide + dexamethasone without thromboprophylaxis in phase III trials.[27] Recent studies support the use of aspirin prophylaxis, which reduces the risk of VTE to less than 5%-with low-molecular-weight heparin reserved for those at high risk for thrombosis, especially immobilized patients and those with a history of VTE.[32-36]

Fatigue is another frequent side effect and is often the reason for treatment discontinuation.[37] This is usually manageable with dose reduction. Another bothersome complaint reported by many patients is muscle cramping. Nearly 20% of patients experience neurologic complications such as dizziness, headache, and/or insomnia. Unlike with thalidomide, new-onset neuropathy is rarely seen with lenalidomide alone, and worsening of preexisting neuropathy has not been widely reported.[38] Rashes have been described in 20% to 30% of patients with myeloma treated with lenalidomide with or without dexamethasone. Severe rashes requiring permanent discontinuation of lenalidomide therapy are rare.[39] Peripheral edema, dyspnea, constipation, diarrhea, and nausea are other common toxicities seen with this drug.

Clinical results with lenalidomide in the treatment of relapsed MM

Following the promising results seen in phase II trials in relapsed disease, two large, multicenter, randomized, placebo-controlled phase III trials were conducted comparing lenalidomide + dexamethasone with dexamethasone alone in patients with relapsed and/or refractory (R/R) MM: MM-009 (N = 353) in North America, and MM-010 (N = 351) in Europe, Australia, and Israel.[27,40,41] Patients were randomly assigned to receive either oral lenalidomide, 25 mg/d, or placebo for 3 weeks, along with 40 mg of oral dexamethasone for 4 days starting on days 1, 9, and 17 of each 28-day cycle for four cycles or until disease progression. After four cycles, dexamethasone (40 mg/d) was limited to days 1 through 4 only. The results of the two studies were largely the same, demonstrating similar and significant improvement in overall survival with lenalidomide + dexamethasone (with a median of 38.0 vs 31.6 months, P = .045).[27] In June 2006, based on the results of these trials, the US Food and Drug Administration (FDA) approved lenalidomide in combination with dexamethasone for the treatment of MM in patients who have received at least one earlier therapy. The European Medicines Agency (EMA) followed suit in June 2007. Subsequently, a large expanded-access study was performed in patients with relapsed MM; the results were in accord with those seen in the phase III studies.[29]

A large number of subsequent analyses have been done using data from these studies, addressing several important aspects of treatment with lenalidomide. Significant numbers of patients who initially had a partial response (PR) achieved a complete response (CR) or very good partial response (VGPR) with further treatment, especially among those who achieved a PR within the first four cycles.[42] Patients who achieved a CR/VGPR as their best response had significantly longer median response duration, time to progression, and overall survival compared with those with a PR.[42] Continuation of lenalidomide treatment until disease progression in patients who achieve a PR was associated with a significant survival advantage.[43]

Chromosomal aberrations such as deletion (17p) and translocations t(4;14), t(14;16), and t(14;20) are poor prognostic factors in MM and have been associated with poorer outcomes. The combination lenalidomide + dexamethasone can result in durable res-ponses in patients with relapsed t(4;14) disease but provides a limited duration of disease control in patients with del(17p).[44] Existing data suggest that lenalidomide may overcome the negative prognosis associated with del(13q).

Recently an expert panel published a consensus statement on the use of lenalidomide in R/R MM.[37] The panel stated that lenalidomide + dexamethasone is most effective when used at first relapse, a phenomenon unlikely to be specific for any one type of therapy. The optimal starting dosage of lenalidomide is 25 mg orally once daily on days 1 through 21 of each 28-day cycle, but should be modified in patients with renal dysfunction. The use of low-dose dexamethasone in combination with lenalidomide may result in better tolerability with no loss of efficacy compared with the standard regimen. The recommended dose of dexamethasone in combination with lenalidomide is 40 mg weekly, but this dose can be modified based on toxicity.[37]

While most studies used lenalidomide in combination with dexamethasone, Richardson et al studied the efficacy and safety of lenalidomide monotherapy in relapsed myeloma.[45] This phase II study enrolled more than 200 patients, among whom two-thirds had experienced three or more prior anti-MM treatment regimens, including prior autologous stem-cell transplants in 45% of the patients. Lenalidomide alone on days 1 through 21 of 28-day cycles induced a PR or better in more than 25% of patients. Myelosuppression was the most common adverse event and was manageable with dose reduction.[45]

TABLE 1


Clinical Studies of Lenalidomide in Relapsed Multiple Myeloma

Another multicenter, open-label, randomized phase II study evaluated two dosing regimens of lenalidomide (30 mg once daily or 15 mg twice daily) in relapsed MM.[46] Analysis showed a similar response rate (minor response [MR] or better in 25%) in the two groups, but grade 3/4 myelosuppression was noted more often in patients receiving 15 mg twice daily (80% vs 69%). However, this difference was not statistically significant. Although lenalidomide monotherapy was effective, the addition of dexamethasone in patients in whom lenalidomide either failed to achieve a response after two cycles or who subsequently progressed induced a response in 29% and stable disease in 21%.[46]

Lenalidomide has been combined with a variety of other drugs that were commonly used in myeloma, including anthracyclines and alkylating agents as well as newer drugs like bortezomib (Table 1). In particular, the combination of lenalidomide and bortezomib (VRD) has significant activity in relapsed disease and is well-tolerated. In addition to bortezomib, combinations with doxorubicin or cyclophosphamide have also been shown to be safe and effective with clinically relevant responses. Combinations of lenalidomide with novel agents such as panobinostat, bevacizumab (Avastin), SGN-40, perifosine, vorinostat (Zolinza), dasatinib (Sprycel), NPI-0002, everolimus (Afinitor), and carfilzomib are currently being investigated in phase I and phase II trials. Table 1 provides a summary of the important trials in this setting with lenalidomide.

Pomalidomide

Pomalidomide (CC-4047) is the most recent IMiD to be evaluated in clinical trials. It is derived from thalidomide and believed to have overlapping mechanisms of action with lenalidomide and thalidomide.[4,47] Preclinical studies showed that it significantly increases serum IL-2 receptor and IL-12 levels.[48] A decrease in CD8+/CD45RA+ cells and CD4+/CD45RA+ cells during the first month of study was accompanied by a corresponding increase in CD8+/CD45RO+ cells and CD4+/CD45RO+ cells, suggesting a switch from naive cells to activated effector T cells.[48] In vitro studies also have shown potent inhibitory effects on osteoclast differentiation.[49] Pomalidomide affects inflammation via transcriptional inhibition of cyclooxygenase-2 (COX-2) production, which is associated with increased prostaglandins in human lipopolysaccharide (LPS)-stimulated monocytes.[50]

TABLE 2


Clinical Studies of Pomalidomide in Relapsed Myeloma

Myelosuppression is the major and dose-limiting toxicity noted in all clinical trials. Grade 3/4 neutropenia has been seen in 30% to 60% of patients and is more common than thrombocytopenia or anemia. Thromboembolic complications occurred with a frequency similar to that reported with other IMiDs. New-onset neuropathy is infrequent, but worsening of preexisting neuropathy has been seen. Other common side effects include orthostatic hypotension, rash, and constipation. Like thalidomide, pomalidomide may have the potential for severe birth defects, so similar precautions should be taken.

Among patients with MM, pomalidomide has been studied extensively in the setting of relapsed disease (Table 2). Initial phase I trials established pomalidomide as well tolerated at a maximum tolerated dose (MTD) of 2 mg daily or 5 mg on alternate days.[48,51] Studies using pomalidomide predominantly as monotherapy have shown excellent activity with an overall response rate of 52%.[52]

The first phase II trial conducted by Lacy et al presented data on a cohort of 60 patients with relapsed myeloma, who had a history of two or three prior regimens. Patients were treated daily with pomalidomide, 2 mg orally, along with weekly dexamethasone, 40 mg orally.[53] Thirty-eight patients (63%) achieved confirmed response, including CR in 3 patients (5%), VGPR in 17 patients (28%), and PR in 18 patients (30%). Responses were seen in 40% of patients with lenalidomide-refractory disease, 37% of those with thalidomide-refractory disease, and 60% of those with bortezomib-refractory disease. In addition, 74% of patients with high-risk cytogenetic or molecular markers (such as hypodiploidy or karyotypic deletion of chromosome 13; fluorescence in situ hybridization [FISH] evidence of the presence of translocations t[4;14] or t[14;16] or deletion [del]17p; or plasma cell labeling index ≥ 3%) had a response. Pomalidomide was well tolerated, and the primary adverse effect was grade 3 or 4 hematologic toxicity, which was seen in one-third of the patients.[53] The median progression-free survival was 11.6 months and was not significantly different in patients with high-risk disease than in patients with standard-risk disease. To better define pomalidomide’s efficacy in lenalidomide-refractory disease, a subsequent phase II trial enrolled 34 patients whose disease was refractory to lenalidomide.[54] The best response was VGPR in 3 patients (9%), PR in 8 (23%), minimal response (MR) in 5 (15%), stable disease in 12 (35%) and progressive disease in 6 (18%), for an overall response rate (defined as MR or better) of 47%. Of the 14 patients who were considered to be at high risk, 8 (57%) had responses, including 4 who had PRs and 4 who had MRs. The median time to response was 2 months and response duration was 9.1 months. The median overall survival was 13.9 months in this group of patients with lenalidomide-refractory disease. In addition to its activity in patients with high-risk disease, pomalidomide is effective in the treatment of extramedullary disease, with a response rate of about 30%, including the extramedullary component.[55]

A third trial enrolled patients whose disease was refractory to both lenalidomide and bortezomib.[56] Patients whose disease is refractory to lenalidomide and bortezomib represent a group who have poor outcomes with current treatment strategies. Pomalidomide was given orally at either 2 mg daily
(n = 35 patients) or 4 mg daily (n = 35 patients), continuously in 28-day cycles along with dexamethasone (40 mg), which was given weekly. Patients enrolled at 2 mg could escalate their dose to 4 mg if there was lack of response or progression. Overall, the response rates were similar in the two groups, with 26% and 28% of patients in the 2-mg and 4-mg groups, respectively, achieving a PR or better. Adverse effects, especially hematologic toxicity, were higher in the 4-mg cohort. The optimal dose and schedule for pomalidomide remains a matter of debate. The original phase I studies showed the MTD to be 2 mg daily or 5 mg every other day.[48,51] The variations-continuous vs 3-out-of-4-weeks dosing, and the addition of dexamethasone in some trials-have led to considerable confusion about the correct strategy. In a recent phase I/II dose-escalation study, Richardson et al showed that 4 mg pomalidomide given daily for 3 of 4 weeks is the MTD for that dosing schedule.[57] Overall response rate in this study was 25%; the phase II study is currently ongoing. A study by Lacy et al has shown that starting with a higher pomalidomide dose (4 mg) has not demonstrated any superiority of response over starting with a 2-mg dose and is associated with a higher risk of myelosuppression.[56] The Intergroupe Francophone du Mylome performed a randomized phase II trial looking at two dosing schedules, 21 of 28 days or 28 of 28 days, with pomalidomide administered at 4 mg daily + weekly dexamethasone. The overall response rate and the duration of response were similar for the two strategies, as was the overall toxicity.[58]

Histone Deacetylase Inhibitors

Inhibition of histone deacetylase (HDAC) provides a novel approach to cancer treatment.[59] Histones are part of the core proteins of nucleosomes, and the acetylation and deacetylation of these proteins play an important role in the regulation of gene expression. Deacetylated histones bind tightly to DNA and limit access of transcription factors, thus inhibiting transcription. Acetylation neutralizes the charge of histones and generates a more open DNA conformation, allowing expression of the corresponding genes. The opposing activities of two groups of enzymes, histone acetyltransferase (HAT) and HDAC, control the amount of acetylation. In normal cells, a balance exists between HAT and HDAC activity. Several lines of evidence suggest that aberrant recruitment of HDAC and the resulting modification of chromatin structure may play a role in the changes in gene expression seen in transformed cells. For example, silencing of tumor suppressor genes at the chromatin level is common in human tumors[60-63], and HDAC complexes have been shown to be crucial to the activity of the acute myeloid leukemia (AML)-specific fusion proteins: promyelocytic leukemia zinc finger (PLZF)-retinoic acid receptor (RAR)-α, promyelocytic leukemia (PML)-RAR-α, and AML1/eight–twenty-one (ETO)[64]. HDAC inhibitors have been shown to induce differentiation, cell-cycle arrest, or apoptosis in cultured tumor cells and to inhibit the growth of tumors in animal models.[65-67] In addition, HDAC inhibitors have been shown to induce expression of p21, a key mediator of G1 arrest and differentiation.[68,69] HDAC inhibitors are thought to affect multiple pathways involved in MM and to correct the deregulation of genes involved in apoptosis and cell-cycle arrest, thus potentially sensitizing MM cells to apoptosis.[70,71]

Several HDAC inhibitors have been evaluated in the context of myeloma, including suberoylanilide hydroxamic acid (SAHA; vorinostat),[72] ITF2357,[73] LBH589 (panobinostat),[74] and romidepsin (Istodax).[75] Results so far suggest limited single-agent activity in patients with MM.[71]

A phase I trial of oral vorinostat (200 mg, 250 mg, or 300 mg twice daily for 5 days/week on a 4-week cycle or 200 mg, 300 mg, or 400 mg twice daily for 14 days on a 3-week cycle) was conducted in patients with R/R MM.[76] Thirteen patients were enrolled; MTD was not reached. Drug-related adverse effects included fatigue, anorexia, dehydration, diarrhea, and nausea, mostly grades 0 to 2. Of 10 evaluable patients, one had a minimal response and nine had stable disease.

Romidepsin (Istodax) is an HDAC inhibitor that has demonstrated cytotoxicity against MM cell lines in vitro. In a phase II trial, patients with MM whose disease was refractory to standard therapy were treated with romidepsin (13 mg/m2) given as a 4-hour intravenous infusion on days 1, 8, and 15 of a 28-day cycle.[75] No objective responses were seen among the 13 patients treated.

While this class of drugs does not seem to have significant single-agent activity, combinations of HDAC inhibitors with newer drugs (especially bortezomib) appear to be promising based on initial phase II trials. A phase I trial evaluated escalating doses of bortezomib (1 to 1.3 mg/m2 on days 1, 4, 8, and 11) and vorinostat (100 mg to 500 mg orally for 8 days of each 21-day cycle) in patients with R/R MM.[77] Twenty-three patients with a median of 7 prior regimens (range, 3 to 13), which included bortezomib in 19 patients, were enrolled. The most common toxicities were myelosuppression (n = 13), fatigue (n = 11), and diarrhea (n = 5). The overall response rate was 42%, including three PRs among nine bortezomib-refractory patients. In another phase I trial, patients with relapsed or refractory MM were randomly assigned to receive oral vorinostat (either 200 mg twice daily or 400 mg once daily for 14 days) in combination with bortezomib (0.7 mg/m2 or 0.9 mg/m2 on days 4, 8, 11, and 15; or 0.9 mg/m2, 1.1 mg/m2, or 1.3 mg/m2 on days 1, 4, 8, and 11 of a 21-day cycle).[78] The best responses observed in the 34 evaluable patients were PR 26%, MR 21%, and stable disease 53%, including PR in 38% of patients with previous bortezomib therapy. Vorinostat, 400 mg once daily, plus bortezomib, 1.3 mg/m2 on days 1, 4, 8, and 11, was considered the MTD.

The combination of panobinostat and bortezomib also has been explored in trials of early-stage disease. In a phase Ib trial, 29 patients were treated with escalating doses of panobinostat and bortezomib.[79] Overall, hematologic adverse events were frequent. Nonhematologic adverse effects included diarrhea, fever, nausea, fatigue, and asthenia. Encouraging clinical efficacy was observed, with 14 (50%) PRs (or better) among 28 evaluable patients, including 4 with a CR. The overall response rate was 64%, including minor responses, and activity was seen in patients with disease refractory to bortezomib alone. Similar results have also been noted with the combination of romidepsin and bortezomib.[80] These results have paved the way for two ongoing phase III trials that are evaluating the combinations of vorinostat or panobinostat with bortezomib compared with bortezomib alone. In addition, studies have also suggested that HDAC inhibitors can be combined with lenalidomide to show significant activity. This combination is going through clinical trials currently.

Heat Shock Protein 90 Inhibitors

Heat shock protein 90 (HSP90) is a molecular chaperone that is induced in response to cellular stress and that leads to stabilization of various client proteins involved in cell-cycle control and apoptotic signaling.[81] HSP90 overexpression may contribute to tumor cell survival by stabilizing aberrant signaling proteins, leading to increased proliferation and resistance to apoptosis. HSP90 inhibitors decrease MM proliferation, suppress the long-term replicative potential of MM cells, and may also sensitize MM cells to other anticancer agents.[82] Several HSP90 inhibitors have been evaluated in early-stage clinical trials.

Tanespimycin was one of the first HSP90 inhibitors to be tested in myeloma. In a phase I dose-escalation study, tanespimycin, 150 mg/m2 to 525 mg/m2, was given on days 1, 4, 8, and 11 of each 3-week cycle for up to 8 cycles to a group of heavily pretreated patients with R/R myeloma. Common adverse events included diarrhea, back pain, fatigue, nausea, anemia, and thrombocytopenia. One patient achieved MR, with a PFS of 3 months. Fifteen patients (52%) had stable disease with a median PFS of 2.1 months. Overall, tanespimycin monotherapy was well tolerated but had limited evidence of activity.[83]

Based on their mechanism of action (as with HDAC inhibitors), there is sufficient rationale to combine HSP90 inhibitors with bortezomib. In a multicenter phase I/II trial, tanespimycin (100 mg/m2 to 340 mg/m2) was combined with bortezomib (0.7 mg/m2 to 1.3 mg/m2) given on days 1, 4, 8, and 11 of each 21-day cycle.[84] The highest tested dose of tanespimycin (340 mg/m2) and bortezomib (1.3 mg/m2) was selected for the phase II portion of the study. Seventy-two patients with relapsed or relapsed and refractory MM were enrolled; 63 patients (89%) completed the study. The combination was well tolerated, and among 67 efficacy-evaluable patients there were two CRs (3%) and eight PRs (12%), for an overall response rate of 27%, including eight minimal responses (12%).

Inhibitors of the PI3k/AKT Pathway, Including mTOR Inhibitors

This pathway, which consists of a series of kinases, including PI3k/AKT, mammalian target of rapamycin (mTOR), and p70S6K, as well as several intervening signaling molecules, plays an important role in the regulation of cell growth, proliferation, and survival.[85-87] Once activated, PI3k converts phosphatidylinositol-4,5-bisphosphate (PIP2) to phosphatidylinositol-3,4,5-triphosphate (PIP3) at the inner surface of the plasma membrane. As a consequence of the interaction with PIP3, pyruvate dehydrogenase kinase (PDK)1 is recruited to the plasma membrane and activated. AKT is also recruited to the plasma membrane, where PDK1 catalyzes phosphorylation of AKT on Thr308, one of the two phosphorylations required for maximal AKT activation. After activation, AKT phosphorylates and alters the biologic activity of a variety of substrates.[88-90] One of the downstream molecules is the mTOR kinase that facilitates cell-cycle progression from G1 into S-phase by phosphorylating two important cell constituents, p70S6K and 4E-BP1,[91,92] resulting in activation of transcription factors S6 and eIF4E. This leads to increased transcriptional activity of genes encoding for cyclin D1; the transcription factors C-myc, hypoxia inducible factor (HIF)-1α, and signal transducer and activator of transcription (STAT)3; ornithine decarboxylase; the growth factors VEGF and fibroblast growth factor (FGF); and ribosomal proteins themselves.[85,90,93-96]

The PI3k/AKT pathway is critical for proliferation and survival of the myeloma cell and mediates some of the antiapoptotic and proliferative effects of IL-6[97], IGF-1[97,98], SDF-1α[99], and hepatocyte growth factor (HGF).[100] Increased phosphorylation of AKT is seen in myeloma cells compared with normal marrow cells and appears to correlate with advanced disease, especially plasma cell leukemia.[101] The pathway partly mediates the proliferative and antiapoptotic effects of IL-6 in myeloma cell lines.[102] IL-6 triggers activation of PI3k and its association with SHP2, deactivates caspase-9, and protects against dexamethasone-induced apoptosis. IL-6 triggers PI3k/AKT signaling, resulting in inactivation of forkhead transcriptional factor (FKHR), with related G1/S phase transition, whereas PI3k inhibitors, such as LY294002, block this signaling, resulting in upregulation of p27(KIP1) and G1 growth arrest.[102] IGF-1 stimulation of the myeloma cells leads to activation of this pathway with sustained activation of NFκB and AKT; phosphorylation of FKHRL-1; up-regulation of the antiapoptotic proteins FLIP, survivin, cIAP-2, A1/Bfl-1, and X-linked inhibitor of apoptosis (XIAP).[103] Activation of this pathway also mediates the migration of plasma cells induced by IGF-1.[104]

Perifosine is the best-studied AKT inhibitor in the setting of myeloma.[105] Initial trials focused on it as a single agent. Sixty-four patients with relapsed myeloma and a median of four lines of prior therapy were enrolled in a phase II trial of perifosine alone or with dexamethasone.[106] Among 48 patients evaluable for response, the best response to monotherapy after two cycles was MR in one patient (2%) and stable disease in 22 patients (46%). The addition of dexamethasone in 37 patients with disease progression led to a PR in 13% of the patients. The most common adverse events included nausea, vomiting, diarrhea, fatigue, increased serum creatinine level, and anemia. Subsequent trials examined the combination of perifosine with bortezomib or lenalidomide.[107,108]

The mTOR kinase, downstream in the PI3k/AKT pathway, is a serine/threonine kinase that receives multiple upstream signals regarding the nutrient and energy status of the cell.[85,96] On activation, mTOR facilitates cell-cycle progression from G1 into S-phase by phosphorylating two important cell constituents, p70S6K and 4E-BP1.[109,110] mTOR exists in mutually exclusive complexes with either Raptor (regulatory-associated protein of TOR) or Rictor (rapamycin-insensitive companion of TOR).[85,96,111] Note that the Raptor–mTOR complex is rapamycin-sensitive and is responsible for phosphorylation of p70S6K and 4E-BP1, whereas the Rictor–mTOR complex is rapamycin-insensitive and is one of the enzymes that can catalyze the activating phosphorylation of AKT at Ser473.[111] mTOR inhibitors include the macrolide rapamycin and its analogs temsirolimus (Torisel) and everolimus.[112] Preclinical studies confirm the antimyeloma activity of rapamycin and its analogs.[113,114] Both temsirolimus and everolimus have been studied in phase II trials in patients with relapsed disease and have shown very little clinically relevant antimyeloma activity. Better understanding of the reciprocal activity of TOR complex (TORC)1 and TORC2 has shed light on potential mechanisms of action and has led to the development of dual inhibitors. In addition, combined targeting of the PI3k/AKT/mTOR pathways may provide a way to enhance activity, and several dual inhibitors are currently going through early-phase trials.[71]

Monoclonal Antibodies

Monoclonal antibody therapy has been quite successful in lymphoid malignancies, as seen in the results with rituximab (Rituxan) in lymphoma and with alemtuzumab (Campath) in chronic lymphocytic leukemia. However, the search for monoclonal antibody–based therapy in myeloma has been beset by the heterogeneous expression of surface proteins in myeloma. More recently, early trials with the humanized monoclonal antibody elotuzumab (HuLuc63) have shown encouraging results. It induces antibody-dependent cell cytotoxicity–mediated apoptosis in vitro, and it significantly reduced tumor growth in preclinical myeloma models.[115] It has also shown significant activity in combination with bortezomib and lenalidomide in in vitro studies, which formed the basis for clinical evaluation of these combinations.[116] In a phase I study in patients who had received one to three prior therapies for myeloma, escalating doses of elotuzumab (2.5, 5, 10, and 20 mg/kg) were administered intravenously on days 1 and 11 in combination with bortezomib, 1.3 mg/m2, administered intravenously on days 1, 4, 8, and 11 of a 21-day cycle.[117] Dexamethasone, 20 mg orally on days 1, 2, 4, 5, 8, 9, 11, and 12 of subsequent cycles, was added for patients with disease progression. No disease-limiting toxicities (DLTs) were observed during cycle 1, and the MTD was not reached. The most frequent grade 3/4 adverse events were lymphopenia, fatigue, thrombocytopenia, neutropenia, hyperglycemia, PN, pneumonia, and anemia. A PR or better was observed in 13 of 27 evaluable patients (48%), including CR in 7% and PR in 41%. The results of the initial trials looking at the combination with lenalidomide were even more promising.[118] Escalating dose cohorts of elotuzumab (5, 10, and 20 mg/kg) were administered intravenously on days 1, 8, 15, and 22 of a 28-day cycle in the first two cycles, and then days 1 and 15 of each subsequent cycle, along with oral lenalidomide, 25 mg/d, on days 1 to 21, and oral dexamethasone, 40 mg weekly. No DLTs were observed at up to 20 mg/kg during the escalation phase and hence no MTD was established. The most frequent grade 3/4 toxicities were neutropenia and thrombocytopenia, and two patients experienced serious infusion-related reactions. A PR or better was seen in 82% (23 of 28) of treated patients and 96% (21 of 22) of lenalidomide-naive patients.

The inflammatory cytokine IL-6 is a survival factor for malignant plasma cells and is secreted by myeloma cells. Preclinical data suggest that CNTO328 (siltuximab), a novel human-mouse chimeric monoclonal antibody targeting IL-6, has an inhibitory effect on tumor burden and potentiates bortezomib-mediated apoptosis. Initial studies support the feasibility of combining the antibody with bortezomib in patients with relapsed myeloma.[119]

Conclusion

There are a number of classes of therapeutic agents in late-stage clinical trials for the treatment of myeloma. Among the IMiDs, both lenalidomide and pomalidomide represent significant additions to the armamentarium. While lenalidomide has been extensively studied in the setting of relapsed disease, current treatment patterns increasingly use it as part of initial therapy, especially in the United States. While studies suggest that retreatment with lenalidomide can have efficacy in selected groups of patients, pomalidomide is likely to fill that need once the drug is approved. Clearly, nonoverlapping mechanisms of action are in place given the activity of pomalidomide among patients whose disease is refractory to lenalidomide. The results of these trials need to be considered in the context of several promising drugs that are currently undergoing clinical trials. One potentially promising class of compounds are the HDAC inhibitors, including vorinostat and panobinostat. While their activity as single agents is limited, their sensitizing capacity and efficacy in combination are being vigorously investigated. HSP90 inhibitors such as tanespimycin fall into a similar category, with limited single-agent activity but signs of potential in combination settings. Monoclonal antibodies such as elotuzumab also offer significant promise both as monotherapies and in multidrug combinations. Encouraging preliminary data have also been seen with PI3K/AKT inhibitors and compounds targeting the mTOR pathway. It is likely that in the near future the treatment armamentarium for MM will undergo a significant expansion as a number of these additional target pathways become validated, offering additional hope for extending survival in patients with MM.

Acknowledgments:The authors would like to thank Brian Szente, PhD, of Fishawack Communications for his editorial assistance with the manuscript. Editorial support was funded by Onyx Pharmaceuticals.

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