Review

RNA interference for multiple myeloma therapy: targeting signal transduction pathways 1.

Introduction

2.

RNA interference

3.

Signal transduction pathways in MM cells -- potential targets

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for RNAi therapy 4.

Conclusion

5.

Expert opinion

Jianfeng Guo, Sharon L McKenna, Michael E O’Dwyer, Mary R Cahill & Caitriona M O’Driscoll† †

University College Cork, School of Pharmacy, Pharmacodelivery Group, Cork, Ireland

Introduction: Multiple myeloma (MM) is a hematological malignancy characterized by infiltration of malignant plasma cells in the bone marrow (BM) and end-organ damage to the bone, BM, kidney and immune system. Although current treatments have improved the treatment of MM, it still remains an incurable disease. RNA interference (RNAi) effectors such as microRNAs and small interference RNAs have shown potential to selectively downregulate genes implicated in the pathology of a range of diseases. Signaling pathways that facilitate growth, survival and migration of MM cells, provide resistance to conventional therapies, and therefore, target these signaling pathways will prove promising for MM treatment. Areas covered: This review focuses on signaling pathways associated with the development of myeloma cells and how interaction of these cells with the tumor microenvironment impacts disease progression. Together these elements provide potential therapeutic targets for RNAi in the future. Expert opinion: Recent advances in oncogenomic studies have revealed the molecular pathogenesis of MM, thus providing new therapeutic targets for RNAi therapy. Pre-clinical evidence suggests that non-viral delivery technology offers the potential to translate this concept into the next generation of RNAi-based therapeutics for MM. Keywords: myeloma signaling pathways, non-viral delivery vectors, RNAi-based therapy, the bone marrow microenvironment Expert Opin. Ther. Targets [Early Online]

1.

Introduction

Multiple myeloma (MM) is a hematological malignancy characterized by multifocal proliferation of clonal, long-lived plasma cells (also known as post-germinal centre, terminally differentiated B cells) within the bone marrow (BM) (Figure 1). MM now accounts for 10% of all hematological malignancies and is the second most common after lymphoma [1]. Estimated numbers for new cases and deaths were 26,850 and 11,240 respectively in the USA in 2015 [2]. Malignant plasma cells secrete high levels of monoclonal antibody (immunoglobulin) or antibody light chains in the blood and/or urine. Clinical manifestations found in MM patients include fatigue due to anemia, recurrent infections due to immune suppression, bone pain due to lytic lesions and renal impairment due to glomerular damage from secreted proteins or high calcium levels [3]. Despite improvements in myeloma treatment achieved by novel drugs such as proteasome inhibitors and immunomodulatory drugs (ImiDs), in addition to conventional approaches and autologous transplant, long-term disease-free survival is rare, with most patients eventually relapsing and developing drug resistance [4]. In addition, MM is primarily diagnosed in the elderly, most of whom cannot tolerate aggressive chemotherapeutics and are unsuitable for transplant. 10.1517/14728222.2015.1071355 © 2015 Informa UK, Ltd. ISSN 1472-8222, e-ISSN 1744-7631 All rights reserved: reproduction in whole or in part not permitted

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Although advances in treatment from conventional options (chemotherapy and radiation therapy) including proteasome inhibitors and immunomodulatory drugs have improved the treatment of multiple myeloma (MM), it still remains an incurable disease. Consequently, novel targeted therapies are urgently needed. RNA interference (RNAi)-based therapies are currently being explored as a treatment option for a variety of diseases, including cancer, genetic disorders, autoimmune diseases and viral infections. When a pharmaceutical formulation that allows systemic administration of RNAi effectors is fully achieved in the future, it will be essential to have appropriate molecular targets. The interaction between MM cells and the BM microenvironment activates signaling pathways that develop the malignant clone and promote neoangiogenesis and osteoclastogenesis. Thus, MM signaling pathways are exciting therapeutic targets for RNAi. Signaling pathways that have been identified in the progression of myeloma cells and the interaction with the tumor microenvironment are discussed in this review presenting targets for RNAi therapy in the treatment of MM. Targeting signaling pathways using RNAi has mainly focused on functional studies rather than as a therapeutic strategy. Therefore, safe and efficacious RNAi delivery systems are urgently required to fulfill the therapeutic potential for MM.

This box summarizes key points contained in the article.

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Role of the BM microenvironment The BM microenvironment is composed of various extracellular matrix (ECM) proteins (i.e., collagen, fibronectin, osteopontin and laminin) and distinct cell populations (i.e., haematopoietic stem cells [HSCs], progenitor and precursor cells, immune cells, erythrocytes, BM stromal cells [BMSCs] and BM endothelial cells [BMECs]) [15]. An increasing number of studies indicate that the cross-talk between myeloma cells and components inside the BM niche is essential for progression of MM [16]. The signaling pathways that result from the mutual interactions can facilitate the migration of malignant cells, induce osteoclastogenesis and angiogenesis and promote resistance to traditional therapies [16]. For example, cell-adhesion-mediated drug resistance (CAM-DR) to conventional and high-dose therapeutics is caused by the interaction of MM cells with ECM proteins. It was reported that adhesion of MM cells to fibronectin results in upregulation of p27 (a cell cycle inhibitor protein that can stop or slow down the cell cycle) and induction of nuclear factor-kB (NF-kB; abnormal activation of NF-kB allows cancer cells to avoid apoptosis), leading to CAM-DR [4]. Recent advances in the pathogenesis of MM including mechanisms of BM homing, interactions of MM cells with BM accessory cells (i.e., BMSCs, osteoclasts, osteoblasts and BMECs) and impact of these interactions on MM, have been reviewed elsewhere [4,16,17] and are also summarized in Figure 2. 1.2

The myeloma stem cell The initiation, progression and relapse of cancers are driven by a rare population of cells termed cancer stem cells (CSCs) which are capable of regenerating tumors [5]. As CSCs often overexpress drug efflux transporters and spend most of their time in the non-dividing G0 cell cycle state, they are considered to be the major hurdle for conventional chemotherapeutics [6]. Although the existence of myeloma stem cells has been proposed, the exact nature of these cells and their relationship to normal plasma cell and B cell development is not fully understood. Several groups have attempted to identify myeloma stem cells based on the association between distinct phenotypes and specific functional growth [7-11]. For example, Matsui et al. showed that human myeloma patient plasma cells engrafted in irradiated non-obese diabetic/severe combined immunodeficiency (SCID) mice, following injection through the tail vein [8]. In this study, cells lacking CD138 were able to differentiate into mature CD138+ plasma cells, which were functionally able to produce circulating monoclonal proteins. In addition to CD138-, myeloma stem cells are reported to carry surface CD45+, CD20+, CD22+, CD27+, CD19+ and CD38-, similar to memory B cells [8-10]. 1.1

However, Yaccoby and Epstein reported that CD45-, CD19- and CD38+ MM patient cells achieved efficient engraftment when injected into human bone fragments of SCID-hu mice and caused several clinical features of MM including circulating M proteins, hypercalcemia and bone lysis [11]. These studies suggest that identification of a myeloma stem cell using immunophenotyping remains limited and therefore, functional definitions that confirm clonogenic growth potential are warranted. Future work in identification of myeloma stem cells needs careful experimental design using representative in vitro and in vivo models and appropriate primary specimens, which more closely mirror the clinical manifestations of MM [12]. Such studies will help reveal the key properties of myeloma stem cells that are attributed to various unexplained clinical features of the condition [5]. It is worth noting that although the expression of the aforementioned cell markers is a hallmark of myeloma stem cells, the mechanisms underlying cell marker upregulation/ downregulation in MM cells remain unclear. Recent studies have shown that myeloma cell marker expression may be regulated by hypoxia, a condition in which the body or a region of the body is deprived of adequate oxygen supply [13,14]. The data from these studies showed that hypoxia decreases the expression of CD138 and increases the expression of B-cell (CD20 and CD45) and stem cell (CD34) markers [14], indicating that a hypoxic microenvironment can affect the phenotype of MM cells, which may correlate with disease progression [13].

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RNAi for MM therapy

Dark zone HSC

Pro-B cell

Immature B cell

Clonal expansion

Somatic hypermutation

(A)

(CD138–CD45+ CD38–CD20+CD22+ CD19+CD27+) Memory B cell (CD138+CD45– CD38+CD20–CD22– CD19–CD27–)

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Mature native B cell

plasma cell Disadvantageous mutations

Short-lived plasma cell

Selection Class switching

lgM TH cell

(B)

lgG, lgA, lgE or lgD

Improved avidity

Follicular DC

Differentiation

(D)

Apoptosis

Apoptosis Light zone (C)

Mantle zone

Figure 1. Multiple myeloma (MM) is a malignancy of post-germinal centre, terminally differentiated B cells that home to the BM. (A) In the BM, HSCs are associated with specialized niches in close contact with MSCs, differentiating into lymphoid progenitors (the lymphoid lineage is primarily composed of T cells and B cells) [15]. In the BM, development of B cells is highly regulated to generate pre-B cells, pro-B cells and immature B cells [15]. (B) When mature native B cells leave the BM and encounter T cell-independent antigenic exposure in blood they differentiate into short-lived plasma cells, which produce lowaffinity IgM [84]. These IgM-secreting PCs do not have somatically mutated immunoglobulin genes and eventually undergo apoptosis in situ. (C) Mature native B cells migrate into secondary lymphoid tissues (i.e., spleen and lymph nodes) in which they establish germinal centers [84]. In the dark zone of a germinal center, mature native B cells are activated to proliferate and differentiate into centroblasts. The proliferating centroblasts initiate the rearrangement of their genes to create antibodies (this process is known as somatic hypermutation) [85]. These centroblasts subsequently migrate and become centrocytes inside the light zone of a germinal centre. The centrocytes improve their affinity when interacting with TH cells and FDCs and receive survival signals from them to avoid the apoptotic pathway; in contrast, centroblasts with unfavorable mutations go through apoptosis [85]. In addition, these centrocytes further undergo proliferation and selection. The immunoglobulin genes of many centrocytes are also remodeled by class switching, which replaces the originally expressed immunoglobulin heavy-chain constant region genes by those of another class (i.e., IgG, IgA, IgE or IgD) [85]. Finally, selected centrocytes differentiate into memory B cells or plasma cells and leave the germinal centre. (D) Post-germinal center plasma cells normally home to the BM, in which they receive survival signals such as IL-6 from stromal cells and become long lived. Recent studies have demonstrated that MM has phenotypic characteristics of long-lived plasma cells and is commonly evident in multiple sites in the BM. In addition, MM cells are somatically hypermutated and remain constant throughout the clinical course, implying that this malignancy mostly likely results from post-germinal center B cells. BM: Bone marrow; FDCs: Follicular dendritic cells; HSCs: Haematopoietic stem cells; IgM: Immunoglobulin M; MM: Multiple myeloma; MSCs: Mesenchymal stromal cells; TH: T helper.

Current and emerging therapeutics in MM A minority of patients with a plasma cell disorder are diagnosed as having solitary plasmacytomas, where localized tumor progression occurs without evidence of distant dissemination [18]. Radiation therapy is normally employed in this stage, and in the case of solitary soft tissue plasmacytomas it may be removed with surgery and irradiation [18]. In contrast, in MM patients, the malignant cells do not remain localized but are disseminated throughout the BM and at this stage, 1.3

drugs are chosen depending on the patient’s age and performance status and whether autologous hematopoietic cell transplantation is planned [18]. A variety of chemotherapeutics are currently available for the treatment of MM. Initial treatment in younger, transplant eligible patients usually involves a triple regimen incorporating a proteasome inhibitor (e.g., bortezomib/Velcade or carfilzomib/Kyprolis) and dexamethasone with either an ImiD (lenalidomide/Revlimid or pomalidomide/Pomalyst) [19]

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(A)

ECM

Collagen

Fibronectin

Syndecan 1

VLA4

(B) Osteoclast

Mesenchymal stem cell

IL6

(C) Blood vessels

IL6 MM cell

MIP1α OPG

IL6, IGF1, VEGF

IL3

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DKK1

Osteoblast

VEGF, HGF

Adhesion molecules IL6, IGF1, TNF-α, APRIL, HGF, etc.

HGF

NF-κB

OPG

VEGF HGF RANKL MIP1α

OPG Bone marrow stromal cell

Figure 2. Interaction of MM cells with the BM microenvironment. (A) The MM cells home to and localize into the BM where they selectively bind ECM proteins or BMSCs via adhesion molecules. For example, the VLA4 on MM cells binds to fibronectin on ECM and to VCAM1 on BMSCs, respectively [16]. In addition, syndecan 1 (a transmembrane heparin sulfate-bearing proteoglycan) on MM cells binds to collagen, inducing the expression of MMP1 that promotes bone resorption and tumor invasion [16]. Increased syndecan 1 is associated with increased tumor mass, decreased MMP9 activity and poor prognosis. (B) The binding of MM cells to BMSCs triggers adhesion- and cytokine-mediated MM cell growth, survival and migration. For example, MM cell binding to BMSCs activates p42/44 MAPK and NF-kB in BMSCs. Induction of IL-6 from BMSCs and MSCs and IGF1, APRIL (a proliferation-inducing ligand), TNF-a and HGF from BMSCs, activate the main signaling pathways in MM cells [17]. In addition, production of RANKL from BMSCs and MIP1a produced from BMSCs and MM cells stimulate osteoclastogenesis [17]. In contrast, osteoclastogenesis is inhibited by OPG produced from osteoblasts and BMSCs [17]. In addition, the secretion of IL3 and DKK1 from MM cells and HGF from BMSCs inhibits osteoblastogenesis [18]. (C) BMSCs secrete the SDF1a, which mediates the initial homing of MM cells to the BM stromal compartment through CXCR4 [17]. As a consequence of myeloma development, secretion of VEGF and HGF from MM cells and BMSCs stimulates BM angiogenesis (it is the formation of new blood vessels from pre-existing vasculature), which is able to promote MM cell growth by secreting IL6, IGF1 and VEGF, enhancing delivery of oxygen and nutrients and removing catabolites [17]. Recently, studies in MM cells and their BM microenvironment have identified new biomarkers and provided promising therapeutic strategies for MM treatment by means of inhibiting the adhesion of MM cells to ECM and BMSCs and production of cytokines and angiogenesis. BM: Bone marrow; BMSCs: Bone marrow stem cells; BMECs: Bone marrow endothelial cells; CXCR4: C-X-C chemokine receptor type 4; DKK1: Dickkopf 1; ECM: Extracellular matrix; HGF: Hepatocyte growth factor; MIP1a: Macrophage inflammatory protein-1a; MM: Multiple myeloma; MMP1: Matrix metalloproteinase 1; OPG: Osteoprotegerin; RANKL: Receptor activator of NF-kB ligand; SDF1a: Stromal-cell-derived factor 1a; VCAM1: Vascular cell adhesion molecule 1; VLA4: Very-late antigen 4.

or cyclophosphamide (a chemotherapeutic) [18]. Popular regimens include Revlimid-Velcade-Dexamethasone and cyclophosphamide-bortezomib-dexamethasone. In the nontransplant eligible population, commonly used regimens include doublets, such as lenalidomide with low-dose dexamethasone or triplets with bortezomib and prednisone combined with either melphalan or cyclophosphamide. Rarely used therapies include doxorubicin, liposomal doxorubicin and etoposide. Although the MM treatments have substantially improved, resistance to therapy remains a major problem. Therefore, novel biologically targeted agents including AKT inhibitors, Pim kinase inhibitors, heat-shock-protein (Hsp) inhibitors, histone deacetylase (HDAC) inhibitors 4

and monoclonal antibodies, are currently in development for the treatment of relapsed/refractory MM, achieving therapeutic effects on both MM cells and the BM tumor microenvironment (see reviews in [20,21]). However, the limitations of new approaches such as acquired drug resistance and toxicity (off-target effects), remain an obstacle to clinical application [22,23]. For example, although non-selective HDAC inhibitors are active in MM, combination therapy is limited by significant adverse effects including severe fatigue, gastrointestinal toxicity and myelosuppression [24]. In contrast, potential exists to use a combination of RNAi-based therapeutics with current therapies to achieve a synergistic clinic response with reduced toxicity (see below sections in this review).

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RNAi for MM therapy

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2.

RNA interference

RNA interference (RNAi) is achieved by small non-coding RNAs that mainly include micro RNA (miRNA) and small interference RNA (siRNA) (see reviews in [25-27]). miRNAs occur naturally in mammalian cells during differentiation and development, and are able to post-transcriptionally regulate gene expression. In the cytoplasm, miRNAs are recognized by the RNA-induced silencing complex and block translation by means of binding to the 3¢-untranslated regions of imperfectly complementary mRNA [26]. In a similar manner, siRNAs mediate gene knockdown via sequence-specific cleavage of perfectly matching mRNA strands [25]. In comparison with miRNAs, siRNAs have to be exogenously introduced into cells by either of two methods: 1) plasmid- or viral-based vectors enter the nucleus and result in the transcription of short hairpin RNAs (shRNAs) which are subsequently exported to the cytoplasm for RNAi pathway [27]; and 2) direct delivery of synthetic siRNAs into the cytoplasm using non-viral transfection systems [28,29]. Gene silencing via RNAi is widely used to identify new cellular pathways associated with disease development and progression (i.e., progression of MM stem cells, interaction of MM cells and the BM microenvironment, see Sections 1.1 and 1.2) [30]. In addition to playing a significant role in functional genomics and target validation, RNAi has presented the potential to selectively downregulate any gene including those normally defined as ‘undruggable’ [31]. Many nuclear transcriptions factors, which play a major role in haematological malignancies (e.g., MYC) are often referred to as ‘undruggable’. Current targeted treatments for MM are generally small-molecule inhibitors targeting specific types of ‘more druggable’ proteins such as kinases or other cytosolic enzymes, achieving pharmacologic inhibition. Therapeutic application of RNAi technology holds exciting potential due to the vastly increased range of potential targets (see Section 3). However, the application of this technology remains limited by barriers. For example, the negative charge (~ 40 anionic charges) and size (21 base-pair nucleic acids, ~ 13,300 g/mol) of siRNA suggest that this RNAi effector is unlikely to cross the cell membrane without assistance [25]. In addition, siRNA is susceptible to degradation from blood serum enzymes and is apt to trigger off-target effects causing nonspecific gene silencing and immunotoxicity (see review in [28]). Therefore, safe and effective RNAi delivery systems are urgently required to overcome the limitations of RNAi [32], including poor transfection, immunogenicity, in vivo instability, non-specific biodistribution, low endosomal escape capability, and disruption and saturation of endogenous RNA machinery [29]. Viral-based vectors encoding shRNA have demonstrated efficient in vitro and in vivo targeted gene silencing; however, they are prone to recognition by the immune system

(i.e., opsonin, complement, coagulation factors and virusspecific antibodies) in the systemic circulation, which can cause a variety of toxic effects limiting clinical application [33]. Recent advances in understanding non-viral delivery systems in terms of drug loading and encapsulation efficiency, delivery strategies, pharmacokinetic profiles and drug release profiles, have accelerated the development of multifunctional nanomedicines that are formulated from a combination of materials including lipids, proteins, carbohydrates, synthetic polymers and metals [34]. Indeed, a number of lipid nanoparticle (LNP) RNAi-based drugs have entered into clinical trials (see review in [29]). For example, an LNP formulation of siRNAs targeting vascular endothelial growth factor (VEGF) and kinesin spindle protein achieved the target downregulation and antitumor activity (i.e., complete regression of liver metastases in endometrial cancer) in patients with solid tumors [35]. To translate RNAi technology to a finished product suitable for the treatment of MM, however, major issues such as short plasma half-life, non-specific tissue distribution, poor intracellular uptake and trafficking need to be fully resolved [36]. By applying extensive knowledge of the aforementioned barriers of myeloma RNAi delivery to the design of novel delivery systems, researchers are moving towards the development of multifunctional non-viral vectors, which fulfill the criteria required for stability, efficiency and specificity [36]. In fact, a rational approach to formulation design by classification of the formulation components, referred to as an ‘ABCD’ system has been introduced by Kostarelos and Miller (see reviews in Figure 3) [37]. This ‘ABCD’ system has been substantially used to deliver functional effectors including ribozymes, DNA enzymes, antisense oligonucleotides and RNAi, for post-transcriptional gene silencing to treat many malignancies [38,39]. In addition, this concept has also presented the potential for transfecting engineered nucleases such as meganucleases, zinc finger nucleases, transcription activator-like effector nucleases and clustered regulatory interspaced short palindromic repeat/Cas systems, in order to achieve genome-editing approaches for cancer therapeutics [34,39]. Therefore, bio-responsive ‘smart’ nanoparticles (NPs) have potential to overcome the in vivo barriers to RNAi delivery in MM (Figure 3). Selective examples of RNAi delivery that have been used in targeting MM signaling pathways are summarized in Table 1 and discussed below.

Signal transduction pathways in MM cells -- potential targets for RNAi therapy

3.

MM is initiated as a result of acquired genetic defects involving plasma cells, such as translocations between immunoglobulin enhancers and oncogenes. For example, chromosomal abnormalities in MM include rearrangements of the switching regions of the immunoglobulin heave chain gene at 14q32 with a variety of partner genes, and these represent

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(1) NF-κB

(2)

BMSCs A

B

C

Wnt

Cytokines and growth factors

D

LRP5/6

Frizzled

Receptors

Receptors

Cytoplasm RAF

JAK

RAS PTEN

MEK

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STAT

MDC (i.e., GSK 3β, CK-1)

PI3K

Interaction

ERK/MAPK

Interaction

AKT

Interaction

Increased stability

Endosomal escape

β-catenin

mTORC2 mTORC1 RNAi release Nucleus

MM cell

Gene transcription

siRNA

miRNA

RISC Poly (A)

Cap

Protein synthesis for self-renewal, proliferation, survival and migration

Cleavage & translational arrest

Figure 3. Delivery of RNAi effectors using ABCD-based non-viral nanovectors against signaling pathways to inhibit the self-renewal, proliferation, survival and migration of MM cells. (1) When MM cells localize inside the BM the cytokine- and adhesion-mediated signaling cascades are activated. For example, the PI3K/AKT/mTOR pathway is associated with MM growth and survival. This cascade is negatively regulated through the action of the phosphatase and tensin homolog deleted on chromosome ten (PTEN), a tumor suppressor protein [20]. However, it is worth noting that the frequency of PTEN deletions and mutations is low in primary MM cells, implying that the pathogenetic role of this tumor suppressor requires further investigation [41]. In addition, RAF/MEK/ERK is activated by RAS mutations, which has been evident in 30 -- 40% of MM patients, and more so in advanced stages of MM [49]. The RAF protein family consists of A-RAF, B-RAF and RAF-1, which are involved in the regulation of proliferation, differentiation and apoptosis of MM. The downstream targets of the RAF/MEK/ERK cascade enter the nucleus in which they can regulate transcription factors, leading to activation of oncogenes. Furthermore, the biological functions of JAK/STAT include proliferation, differentiation, migration, apoptosis, and cell survival, depending on the signal, tissue, and cellular context. The IL6-induced STAT3 pathway plays a key role in transcriptional regulation mediating the expression of oncoproteins for survival and anti-apoptosis of MM cells. Moreover, WNT plays a central role in its pathway by inducing cell signaling at a receptor complex consisting of a Frizzled member and a co-receptor of the LDL receptor-related protein family, usually LRP5 or LRP6 (low-density lipoprotein receptor-related proteins 5 and 6) (this process is known as the canonical WNT signaling). The stabilized b-catenin forms a complex with accessory proteins, resulting in the transcription of important growth-promoting genes in MM cells. In addition, constitutive DNA-binding and transactivation activity of NF-kB has been reported in many cancers. Generally, there are two pathways of NF-kB activation: the classical or canonical pathway which involves dimers composed of either RelA or c-Rel and p50 and the alternative pathway which involves dimers containing RelB and p100 subunits [86]. In addition to activation of MM cells, NF-kB has been demonstrated to play a role in MM cells in the BM microenvironment, and activation of NF-kB in BMSCs and osteoclasts can enhance osteoclastogenesis [4], contributing to pathophysiology and pathogenesis of MM. (2) ABCD-based delivery systems are composed of RNAi effectors (miRNA and siRNA) (A), non-viral delivery materials by which RNAi effectors are either complexed or conjugated (B), biological stability enhancing moieties (e.g., PEG) (C) and distal targeting ligands (i.e., monoclonal antibodies targeting myeloma antigens) (D). Systemic delivery of ABCD-based nanovectors will avoid kidney filtration, nuclease degradation and the MPS, producing a prolonged blood half-life. Recently, myeloma target antigens, including CD38, CD138, CD56, CD74, CD40, IGF-1R, SLAMF7 and FcRL5, have been used for antibody-based therapies for MM [87]. The application of mAbs as a targeted delivery system for gene therapy may facilitate site-specific administration and avoid non-specific toxicity issues. Following cell-specific internalization (normally via ligand-receptor mediated pathway), RNAi effectors will be effectively released into cytoplasm from these bio-responsive ‘smart’ nanovectors that achieve successful endosomal/lysosomal escape. When RNAi effectors are recognized by the RISC the sense strand is removed whereas the antisense is incorporated to form antisense-RISC that will block translation and/or cause degradation of the corresponding mRNA, inhibiting the self-renewal, proliferation, survival and migration of MM cells. BM: Bone marrow; mAbs: Monoclonal antibodies; MM: Multiple myeloma; MPS: Mononuclear phagocyte system; RISC: RNA-induced silencing complex

RNAi for MM therapy

Table 1. Summary of in vitro and in vivo studies carried out using RNAi to target multiple myeloma signaling pathways. Signaling pathway

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PI3K/AKT/mTOR (in vitro studies)

RNAi technology (delivery vector and route where applicable) shRNA (Lentivirus)

Plasmid-based shRNA (Electroporation)

SmartPool siRNA (Lipofectamine 2000) siRNA (X-treme GENE)

PI3K/AKT/mTOR (in vivo studies)

shRNA (Lentivirus)

RAF/MEK/ERK (in vitro studies)

shRNA (Lentivirus)

shRNA (Lentivirus)

Plasmid-based shRNA (Electroporation)

Plasmid-based shRNA (Electroporation)

siRNA (Lipofectamine 2000)

RAF/MEK/ERK (in vivo studies)

shRNA (Retrovirus)

Comment

Ref.

Translation initiation factor eIF4F is one of the best-studied MYC effectors and a downstream target of the PI3K/mTOR pathway. shRNA-mediated knockdown of three subunits of the eIF4F cap-binding complex enhanced myeloma cell death in combination with dexamethasone, a corticosteroid used as frontline therapy in MM Inhibition of the PI3K/AKT/GSK3b pathway with electroporation-mediated shRNA decreased expression of the heat shock transcription factor 1 and downregulated constitutive and inducible heat shock protein 70 (Hsp70) expression, inducing apoptosis of myeloma cells Cell viability was significantly decreased with pan-class PI3K inhibition by down-regulation of all the isoforms a, b, g, and d in MM.1S cells using pooled siRNAs compared with a scrambled siRNA control Treatment of human myeloma IM-9 and 8226 cells with AKT siRNA resulted in significant inhibition of AKT protein expression, which induced the sensitization of cells to cisplatin for improved apoptosis SCID-Bg mice were injected with MM cells pre-treated with lentiviral shRNAs silencing different isoforms of PI3K (p110a, b, g and d). The data showed that tumor development was significantly reduced by shRNAmediated inhibition of P110b and d, suggesting that MM may be more dependent on PI3K p110b and p110d It was demonstrated that intracellular nicotinamide adenine nucleotide (NAD+) plays a major role in the regulation of several cellular processes, and a higher NAD+ turnover has been evident in malignant cells than normal cells. Downregulation of ERK1 and EKR2 using siRNA significantly enhanced MM cell death in combination with FK866 (a specific chemical inhibitor for Nampt that is a rate-limiting enzyme involved in NAD+ synthesis), suggesting that a combination of RNAi with chemical drugs may yield an efficient strategy for therapeutic intervention in drug-resistant MM malignancies that have mutations activating these pathways Knockdown of IRS-1 expression using shRNA diminished heparanasemediated ERK activation in tumor cells, revealing a novel mechanism whereby heparanase enhances activation of the insulin receptor signaling pathway leading to ERK activation and modulation of myeloma behavior Isoform-specific knockdown of RAS using shRNAs reduced the viability of MM cells harboring the respective oncogenic K- or N-RAS mutation. In addition, RAS mutation was not sensitive to AKT downregulation, and oncogenic RAS did not affect AKT activation, suggesting constitutively activated RAS and AKT contribute independently to MM cell survival The treatment of All-Trans Retinoic Acid (ATRA) caused the expression of MDR1 (the multidrug resistance 1), a known downstream target of Ape/ Ref-1. Transfection with siRNA specifically targeting MSK1 (the downstream target of ERK) reduced the constitutive expression of Ape/Ref1, potentially abolishing the induction of chemoresistance by ATRA Silencing of IL-16 using siRNA reduced the proliferation of end-stage myeloma cells and inhibited the colony-forming of myeloma precursors. In addition, this study suggests that IL16-mediated myeloma proliferation may be mediated by the involvement of transcription factors FOS and JUN in MAPK and PIK3 pathways The addition of exogenous DKK-1, MCP-1, and both, to the p38 MAPK shRNA-treated human MM ARP-1 cells, significantly restored the generation of osteoclasts, indicating that DKK-1 and MCP-1 are responsible for myeloma p38-mediated activation of osteoclast differentiation and activity in vitro and in vivo

[88]

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[89]

[46]

[90]

[91]

[92]

[93]

[94]

[95]

[96]

[97]

7

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Table 1. Summary of in vitro and in vivo studies carried out using RNAi to target multiple myeloma signaling pathways (continued). Signaling pathway

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JAK/STAT (in vitro studies)

RNAi technology (delivery vector and route where applicable) Plasmid-based shRNA (Electroporation)

siRNA (Electroporation)

JAK/STAT (in vivo studies)

siRNA (CpG-conjugation)

WNT/b-catenin (in vitro studies)

siRNA (Electroporation)

SmartPool siRNA (Electroporation)

siRNA (Lipofectamine 2000)

WNT/b-catenin (in vivo studies)

shRNA (Lentivirus)

siRNA (Atelocollagen)

NF-kB (in vitro studies)

shRNA (Retrovirus)

siRNA (Electroporation)

NF-kB (in vivo studies)

8

siRNA (PEI-based nanoparticle)

Comment

Ref.

The serine/threonine kinase serum/glucocorticoid-regulated kinase 1 (SGK1) is a prominent transcriptional target of cytokine-induced signaling in myeloma cells and is found in many human myeloma cell lines and primary myeloma cells. In addition, shRNA-mediated knock-down of STAT3 reduced SGK1 levels and resulted in decreased proliferation of myeloma cell lines and reduced cell numbers In this study, it was demonstrated that TNFa increased phosphorylation of STAT3 including c-Myc and cyclin D1. TNFa-stimulated IL-6 production was abolished by inhibition of JAK2 and IKKb or by small siRNA targeting TNF receptors (TNFR) but not by MEK, p38, and PI3K inhibitors, suggesting that blockage of JAK/STAT-mediated NF-kB activation was highly effective in controlling the growth of MM cells Chemically synthesized CpG-siRNA molecules achieved efficient downregulation of the STAT3 expression via the mediation of TLR9. They generated an enhanced antileukaemic effect in a subcutaneous mouse MM and model, without showing significant toxic side-effects The non-canonical Wnt agonist Wnt5a stimulates human osteoblastogenesis through its co-receptor Ror2. Wnt5a inhibition by siRNA reduced the human mesenchymal stromal cells (hMSCs) expression of osteogenic markers, suggesting that the Wnt5a/Ror2 pathway is involved in the pathophysiology of MM-induced bone disease siRNA-mediated gene silencing in MM cells demonstrated that accumulated b-catenin activates early endoplasmic reticulum stress signaling via eIF2a, C/EBP-homologous protein (CHOP), and p21, leading to immediate growth inhibition The WNT3-induced adhesion to the BM stromal cells caused cell adhesion--mediated drug resistance (CAMDR) of myeloma cells against doxorubicin. This CAMDR was significantly reduced by WNT3 siRNA, suggesting a role of RNAi for combined therapy in drug-resistant MM Overexpression of NEK2 activates both AKT and canonical Wnt signaling, causing increased cancer cell proliferation and drug resistance. Knockdown of NEK2 by shRNA inhibited myeloma cell growth and decreases drug resistance in vitro and in vivo Treatment of b-catenin siRNA formulated with atelocollagen significantly inhibited tumor growth in a subcutaneous MM mouse model, in comparison with treatments with scramble siRNA, PBS or atelocollagen alone NIK overexpression activates NF-kB signaling and blocks cell death in MM. Reduction of NIK using shRNA proved toxic to four cell lines with high NIK protein expression but not to four cell lines with low or absent NIK expression siRNA-mediated knockdown did not have any major short-term adverse effect on the viability of MM cells, indicating that careful selection in genetic background blockade of the NF-kB system is indispensable for RNAi-based therapy A polyethylenimine (PEI)-based nanoparticle containing anti-eIF5A siRNA efficiently reduced eIF5A mRNA and protein expression levels, which sensitized MM cells to apoptosis. In this study, inhibition of eIF5A using targeted siRNA, but not negative control siRNA, resulted in downregulation of ERK/MAPK and NF-kB, which significantly slowed down the myeloma growth in subcutaneous xenograft mouse models

[98]

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[99]

[57]

[100]

[101]

[67]

[102]

[68]

[103]

[104]

[53]

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the most common chromosomal abnormalities in MM (~ 70% of alterations) along with deletions of chromosome 13 [40]. In addition, abnormalities of chromosomes 17 and 18 involving the p53 and c-MYC genes are generally considered to be less frequent events (i.e., secondary events) but carry a poor prognosis [40]. These abnormalities are subsequently complemented by secondary events that lead to activation of growth and survival pathways and to disruption of apoptotic signaling [4]. These events can also enhance the myeloma--BM microenvironment interaction, which can either directly produce cytokines and growth factors for maintenance of MM, or indirectly promote tumor growth via stimulation of angiogenesis [41]. In addition, signals from MM cells can disturb the bone remodeling process by means of blocking osteoblast differentiation and inducing osteoclast activation, producing a pro-tumor microenvironment [18]. As these signaling pathways promote genetic instability, disease resistance and progression, targeting them may be an effective strategy for inhibiting MM progression and relapse. A comprehensive knowledge of the key signaling and survival pathways should provide better molecular targets and lead to the development of safe and efficient RNAi-based therapeutics for MM. In this section, signal transduction pathways involved in the development and drug-resistance of MM cells and their interaction with the tumor microenvironment will be discussed (Figure 3). PI3K/AKT/mTOR The PI3K/AKT/mTOR cascade is crucial to a variety of physiological processes including cell cycle progression, transcription, translation, differentiation, apoptosis, motility and metabolism [42]. The PI3K pathway mediates proliferative and anti-apoptotic signals in MM through both cytokinedependent and -independent mechanisms [20]. Eight classes of PI3K’s (Phosphatidylinositol-3-kinase’s) have been described in mammalian cells, but only the class I product (PIP3), which can function as second-messenger in intracellular signaling, has been implicated in oncogenesis [42]. AKT, downstream of PI3K, is the primary effector molecule of the PI3K signaling cascade (Figure 3). AKT is activated following PIP3 activation of PDK1. AKT can also be activated by mTORC2 (rapamycin-insensitive companion of mTOR complex), a complex that lies downstream of activated AKT itself [42]. Once activated, AKT is capable of translocating to the nucleus, within which it affects the activity of many transcriptional regulators (i.e., NF-kB and murine double minute 2 [MDM2]) [43]. In addition, AKT also activates the phosphorylation of several substrates, such as mTORC1 (mammalian target of rapamycin complex 1), FKHR (forkhead in rhabdomyosarcoma, also known as FOXO1, a transcription factor), GSK-3b (glycogen synthase kinase3 b) and BAD (member of B cell leukemia-2 gene product family, an antiapoptotic protein), which have a vital role in cell cycle regulation, proliferation and apoptosis, either 3.1

directly or indirectly via an intermediary, as reviewed elsewhere [44]. Recently, inhibition of PI3K/AKT/mTOR using RNAi has achieved a promising anti-myeloma effect. For example, it was reported that p110d, one of the eight distinct mammalian isoforms of class I PI3K, is expressed in many cancers, including colon and bladder carcinoma, glioblastoma, acute myeloid leukaemia and MM [45]. Knockdown of p110d using targeted siRNA, but not mock siRNA, significantly slowed down MM cell growth [45]. In addition, it was demonstrated that pooled siRNA successfully inhibited the expression of all PI3K isoforms resulting in increased MM cell death and reduced adhesion of MM cells with BMSCs [46]. This suggests that RNAi may disrupt the cross-talk between MM cells and their BM microenvironment achieving an anti-cancer effect independently of MM cell genetic signatures. RAS/RAF/MEK/ERK The RAF/MEK/ERK cascade transmits signals from multiple cell surface receptors to transcription factors in the nucleus, modulating the expression of key genes (Figure 3). This cascade interacts with a number of different signal transduction pathways including PI3K/AKT/mTOR and Janus kinase/ signal transducers and activators of the transcription (JAK/ STAT) (JAK STAT pathway) and mediates many fundamental biological processes, including cellular proliferation, survival, angiogenesis and migration [47]. Mutations in the RAS oncoproteins (NRAS, KRAS and BRAS) have frequently been found in MM patients with this cascade being one of the most commonly mutated signaling pathways in myeloma [48]. The prevalence of NRAS and KRAS mutations is ~ 15 -- 18% in newly diagnosed and relapsed MM patients, respectively. Evidence suggests that NRAS and KRAS mutations may have overlapping but non-identical effects [48]. In addition, BRAF mutations were reported in 4% of MM patients and a specific mutation in the BRAF gene (V600E) has been successfully treated with vemurafenib (a BRAF enzyme inhibitor) [48]. One of the primary downstream targets of RAF are MEK proteins (also known as MAPKK or MAPK kinase), the main physiological substrates of which are ERK (extracellular signal-regulated kinase) proteins (Figure 3). In addition, phosphorylation of ERK is activated via binding of IL6, IGF-1 (insulin-like growth factor 1), VEGF and APRIL (a proliferation-inducing ligand) to their respective receptors, as well as through the interaction of MM cells with BMSCs, which mediates ERK activation independently of cytokines/ growth factors [4]. ERK and its downstream targets enter the nucleus, where they can regulate transcription factors, leading to activation of oncogenes (such as FOS, JUN and MYC) [49]. Recently, Lu et al. delineated the role of proviral integration sites of Moloney 2 (Pim2) kinase in MM and described the mechanisms by which it mediates clonal plasma cell proliferation [50]. This up-regulation of the Pim2 gene is driven by IL6 and tumor necrosis factor (TNF) family cytokines (TNF-a, BAFF and APRIL) through signal transducer 3.2

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and activator of transcription 3 (STAT3) and nuclear factor kB (NK-kB) activation [51]. Treatment with Pim2 siRNA has successfully reduced MM cell proliferation [52]. In addition, it was reported that eukaryotic translation initiation factor 5A (eIF5A), a highly conserved protein is regulated by post-translational hypusine modification, which is involved in many cellular processes including proliferation, apoptosis, differentiation and inflammation [53]. High expression levels of hypusinated eIF5A have been associated with activation of oncogenes and with malignant transformation of hematopoietic cells. A polyethylenimine-based NP containing anti-eIF5A siRNA efficiently reduced eIF5A mRNA and protein expression levels, which sensitized MM cells to apoptosis [53]. In this study, inhibition of eIF5A using targeted siRNA, but not negative control siRNA, also resulted in downregulation of ERK/MAPK and NF-kB, which significantly slowed down the myeloma growth in subcutaneous xenograft mouse models. JAK/STAT Cellular responses to a number of cytokines and growth factors are mediated by the evolutionarily conserved JAK/ STAT signaling pathway [54]. In addition to STAT activation, the JAK family (JAK1, JAK2, TYK2, and JAK3) can activate other signaling pathways, such as MAPK and PI3K/AKT/ mTOR (Figure 3). Activation of STAT proteins, especially STAT5 and STAT3, is important in the pathogenesis of lymphoid and myeloid malignancies [55] and constitutive expression of STAT3 has been seen frequently in patients with MM [56]. The best characterized myeloma growth factor in the JAK/STAT pathway is IL-6, which leads to signaling through the IL-6 receptor (IL-6R) and triggers phosphorylation of STAT3 through JAK1 (Figure 3). The IL6-induced STAT3 pathway plays a key role in transcriptional regulation of oncoproteins (i.e., BCL-XL and MCL1), which are involved in survival and anti-apoptosis of MM cells [4]. It has recently been demonstrated that chemically synthesized CpG-siRNA molecules, produced by conjugation of a STAT3 siRNA to a CpG oligonucleotide, achieved efficient downregulation of STAT3 expression, via the mediation of TLR9 (toll-like receptor 9, is commonly expressed in myeloma) [57]. This TLR9-triggered CpG-siRNA displayed a primary immune response, which generated an enhanced anti-leukemic effect in human MM cells in vitro, as well as in a subcutaneous MM mouse model, without significant toxicity. 3.3

WNT/b-catenin The WNT/b-catenin cascade is involved in virtually every aspect of embryonic development and also controls homeostatic self-renewal in a number of adult tissues [58]. For example, the WNT/b-catenin cascade plays a critical role in self-renewal of HSCs and its dysregulation may be involved in a wide range of hematological malignancies including MM [59-61]. Self-renewal capacity, one key characteristic of 3.4

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myeloma stem cells, is normally associated with increased expression of WNT/b-Catenin signaling pathways [59-61]. The canonical WNT signaling can trigger the destabilization of the multi-protein destruction complex and the subsequent stabilization of b-catenin [61]. When cytoplasmic b-catenin is stabilized by WNT signaling, it forms a complex with accessory proteins, resulting in the transcription of important growth-promoting genes, such as c-MYC and Cyclin D1, both of which are frequently deregulated in MM [62,63]. MM depends on the BM microenvironment for growth and survival, and is characterized by extensive bone loss and osteolytic lesions located at sites of medullary plasmacytomas. Recent studies demonstrated that WNT directly affects both the osteoblast and osteoclast bone cell lineages and also indirectly affects these cells through cross-talk in the BM microenvironment, inducing an overall increase in bone formation and a decrease in bone resorption [64]. It is interesting to note that, despite a direct effect on increasing tumor growth at extraosseous sites, increasing WNT signaling in the BM microenvironment can prevent the development of myeloma bone disease and inhibit myeloma growth within bone in vivo [65]. However, production of WNT-signaling inhibitors DKK1, sFRP2 and sFRP3 by myeloma cells contributes to the development of osteolytic lesions through the direct suppression of osteoblast differentiation [66]. Kobune et al. reported that the overexpression of WNT3 is found in KMS-5 and ARH77 cells (human myeloma cell lines) which are tightly adhered to human BMSCs showing accumulation of b-catenin and GTP-bounded RhoA [67]. The WNT3-induced adhesion to BMSCs caused CAM-DR of myeloma cells against doxorubicin. This CAM-DR was significantly reduced by WNT3 siRNA, suggesting a role for RNAi in combination therapy in drug-resistant MM. In addition, Ashihara et al. demonstrated that treatment of b-catenin siRNA formulated with atelocollagen significantly inhibited tumor growth in a subcutaneous MM mouse model, in comparison with treatments with scrambled siRNA, PBS or atelocollagen alone [68]. NF-kB NF-kB (nuclear factor k-light-chain-enhancer of activated B cells, a protein complex that controls transcription) has been described as an important transcription factor for survival of healthy mature B cells, and therefore, mutations in the NF-kB pathway are highly associated with B-cell neoplasms including MM [69]. The NF-kB family of transcription factors are composed of five members, including RelA (p65), RelB, c-Rel, NF-kB1 (p50 and its precursor p105) and NF-kB2 (p52 and its precursor p100) [70]. The NF-kB transcription factor complex is assembled through dimerization of two subunits (they can be homodimeric or heterodimeric) and is subsequently sequestered in the cytoplasm due to their interaction with the regulatory proteins ‘inhibitors of NF-kB’ [71]. The activation of upstream signaling pathways results in the formation of the active NF--kB complex, which can 3.5

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then translocate to the nucleus and lead to the transcription of target genes (NF-kB targets in MM cells from the classic or alternative pathway, have been reviewed elsewhere [Figure 3] [69-71]). As NF-kB signaling is also critical for many immune cell functions, systemic blockade of this pathway may lead to detrimental side effects. Recently, Zhou et al. reported a safe and effective systemic delivery system for siRNA targeting the p65 subunit of NF-kB in rheumatoid arthritis (RA) [72]. In this study, a novel melittin-derived cationic amphiphilic peptide was used to complex siRNA forming NPs, which were able to specifically home to inflamed joints in a murine model of RA, resulting in reduced NF-kB mRNA and protein levels without off-target toxicities. This NP-based delivery system demonstrates the specificity of RNAi delivery into bone areas for NF-kB gene silencing, a critical feature that cannot be achieved with small-molecule inhibitors [72]. This novel NP may therefore have broad utility for the specific delivery of siRNA that can target and limit NF-kB-associated signaling for the treatment of a variety of diseases including MM. In addition, the rationale for inhibition of the proteasome as a therapeutic approach in MM was initially based on blocking activation of NF-kB signaling. However, NF-kB is important in other tumor types as well, yet proteasome inhibitors are not as active in these cancers. It is now reasoned that normal plasma cell biology requires stringent quality control for the large amounts of immunoglobulin proteins produced and that any disturbance in proteostasis (protein homeostasis) will compromise viability. The ubiquitin-proteasome system and autophagy (an intracellular degradation system that delivers cytoplasmic constituents to the lysosome and enables adaptation to stress) are integrated degradation networks that cooperate to maintain cellular proteostasis [73]. This importance of proteostasis in myeloma cells has led to targeting of endoplasmic reticulum stress response signaling (or the unfolded protein response, UPR) such as IRE1, XBP1 and heat shock proteins (see reviews in [74]). Recently, a loss-offunction RNAi screen using shRNA was applied to uncover essential pathways required for myeloma proliferation and survival, and in this study, human myeloma lines were shown to depend for their survival on interferon regulatory factor 4 (IRF4, a transcriptional factor), which, through a caspase 10-dependent mechanism, prevents excessive autophagy from executing non-apoptotic myeloma cell death [75]. Of note, inhibition of caspase-10 using shRNAs induced autophagy and was toxic to all myeloma cell lines tested, regardless of their various oncogenic aberrations, suggesting that targeting the proteostasis network using RNAi in MM may hold therapeutic potential for myeloma treatment. 4.

Conclusion

Recent advances in whole-genome sequencing and analysis of MM have revealed a complex signaling network that sustains survival of the malignant cell and mediates tumor progression

and drug resistance [76]. In addition to the aforementioned signaling pathways, Notch [77], Hsp [78] and p53/MDM2 [79] pathways also contribute to the maintenance of the myeloma signaling network. In addition, recent studies using next-generation sequencing have revealed significant subclonal evolution in myeloma [80]. These sub-clones have acquired novel mutations and phenotypes, which can facilitate subsequent relapse and resistance. Therefore the ultimate treatment will require multiple targeting to eradicate all clones, including sub-populations that have already acquired further survival signaling [80]. It is worth noting that inhibition of multiple cellular functions has been achieved by silencing several independent genes using a cocktail of siRNA [81]. Therefore, RNAi strategies that targeting the signaling pathways described above in relation to MM cells, tumor--BM interactions and/or the BM microenvironments, used alone or in combination, may offer enhanced therapeutic efficacy without the added toxicity seen in multiple drug combinations. Multiple targeting also offers the opportunity to overcome drug resistance that is often achieved by compensatory signaling pathways -- a common reason for failure of kinase inhibitors. Finally, development of a suitable vehicle or vector, to facilitate systemic (and potentially targeted) delivery, is required to exploit the full therapeutic potential of RNAi intervention. 5.

Expert opinion

Despite advances achieved by new strategies (i.e., small molecule inhibitors and immunomodulatory agents) used in combination with traditional chemotherapeutics, resistance inevitably develops and MM ultimately remains fatal, suggesting that it is necessary to develop other therapies for those who experience treatment failure and drug resistance. Promising results from several ongoing RNAi-based clinical trials demonstrate great therapeutic potential for a range of diseases, including cancer, genetic disorders, autoimmune diseases and viral infections (see review in [29]). Therefore, the application of RNAi technology is likely to present opportunities for the treatment of relapsed/refractory myeloma. As summarized in Table 1 the majority of research that has been conducted targeting signal transduction pathways with RNAi has focused on the use of this technology to determine the role of these molecules in MM progression rather than as a therapeutic strategy. The barrier in translating the rapidly expanding knowledge of RNAi into MM treatment is the lack of safe and effective delivery systems, which can penetrate into the BM. Therefore, when an ‘ideal’ formulation is developed in the future, RNAi-based therapeutics may advance therapeutic function with minimal side-effects for treatments of MM. Recently, it was reported that E-selectin expression is constitutive on the BM endothelium and limited to specific micro-domains that co-localize with homing sites of tumor cells, indicating E-selectin as an attractive biological target

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for the delivery of drug carriers to the BM endothelium [82]. A novel porous silicon microparticle developed by Mann and colleagues targeted the BM vasculature via specific binding of an E-selectin thioaptamer ligand to E-selectin [82]. In addition, Swami and colleagues developed a bone-homing polymeric NP system for spatiotemporal controlled delivery of therapeutics to bone for MM therapy, which diminishes off-target effects and increases local drug concentrations [83]. These NPs contain a bisphosphonate (alendronate) as a targeting ligand for bone mineral. The NPs containing bortezomib decreased tumor burden and enhanced survival in xenograft MM mice in comparison with empty-NPs (no drug) and unformulated drug [83], indicating a clinically relevant method of drug delivery that can inhibit MM progression. Moreover, a novel liposomal formulation named SMARTICLES was designed to deliver a mimic of the naturally occurring miRNA tumor suppressor, miR-34, which is lost or under expressed in tumors of patients with a wide variety of cancers (www.mirnarx.com). This product, trademarked as MRX34, is currently being studied in a multicentre, open-labeled Phase I clinical trial in patients with solid Bibliography Papers of special note have been highlighted as either of interest () or of considerable interest () to readers. 1.

Walker RE, Lawson MA, Buckle CH, et al. Myeloma bone disease: pathogenesis, current treatments and future targets. Br Med Bull 2014;111(1):117-38

2.

Siegel RL, Miller KD, Jemal A. Cancer statistics, 2015. CA Cancer J Clin 2015;65(1):5-29

3.

Anderson KC, Carrasco RD. Pathogenesis of myeloma. Annu Rev Pathol 2011;6:249-74 Discusses the molecular pathogenesis of multiple myeloma (MM) as new therapeutic targets.

.

4.

5.

6.

7.

12

Hideshima T, Mitsiades C, Tonon G, et al. Understanding multiple myeloma pathogenesis in the bone marrow to identify new therapeutic targets. Nat Rev Cancer 2007;7(8):585-98 Meacham CE, Morrison SJ. Tumour heterogeneity and cancer cell plasticity. Nature 2013;501(7467):328-37 Holohan C, Van Schaeybroeck S, Longley DB, Johnston PG. Cancer drug resistance: an evolving paradigm. Nat Rev Cancer 2013;13(10):714-26 Kirshner J, Thulien KJ, Martin LD, et al. A unique three-dimensional model for evaluating the impact of therapy on

tumors and blood malignancies including MM (ClinicalTrials.gov Identifier: NCT01829971). These encouraging advances suggest that multifunctional NP delivery systems will, in the future, address the critical barriers to the translation of RNAi therapeutics to the clinic, facilitating an advancement and improvement upon current MM treatment strategies.

Declaration of interests The authors acknowledge funding from: the Government of Ireland Postdoctoral Fellowship from Irish Research Council (GOIPD/2013/150) and the Irish Cancer Society via a project grant (PCI11ODR). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties. 14.

Muz B, de la Puente P, Azab F, et al. Hypoxia promotes stem cell-like phenotype in multiple myeloma cells. Blood Cancer J 2014;4:e262

15.

Morrison SJ, Scadden DT. The bone marrow niche for haematopoietic stem cells. Nature 2014;505(7483):327-34

16.

Manier S, Sacco A, Leleu X, et al. Bone marrow microenvironment in multiple myeloma progression. J Biomed Biotechnol 2012;2012:157496

Pilarski LM, Hipperson G, Seeberger K, et al. Myeloma progenitors in the blood of patients with aggressive or minimal disease: engraftment and self-renewal of primary human myeloma in the bone marrow of NOD SCID mice. Blood 2000;95(3):1056-65

17.

Oranger A, Carbone C, Izzo M, Grano M. Cellular mechanisms of multiple myeloma bone disease. Clin Dev Immunol 2013;2013:289458

18.

Podar K, Anderson KC. Multiple myeloma - a new era of treatment strategies. ISBN: 978-1-60805-609-5

11.

Yaccoby S, Epstein J. The proliferative potential of myeloma plasma cells manifest in the SCID-hu host. Blood 1999;94(10):3576-82

19.

12.

Hajek R, Okubote SA, Svachova H. Myeloma stem cell concepts, heterogeneity and plasticity of multiple myeloma. Br J Haematol 2013;163(5):551-64

Mitsiades CS, Chen-Kiang S. Immunomodulation as a therapeutic strategy in the treatment of multiple myeloma. Crit Rev Oncol Hematol 2013;88(Suppl 1):S5-13

20.

de la Puente P, Muz B, Azab F, et al. Molecularly targeted therapies in multiple myeloma. Leuk Res Treatment 2014;2014:976567

21.

Allegra A, Penna G, Alonci A, et al. Monoclonal antibodies: potential new therapeutic treatment against multiple myeloma. Eur J Haematol 2013;90(6):441-68

multiple myeloma. Blood 2008;112(7):2935-45 8.

9.

10.

13.

Matsui W, Huff CA, Wang Q, et al. Characterization of clonogenic multiple myeloma cells. Blood 2004;103(6):2332-6 Matsui W, Wang Q, Barber JP, et al. Clonogenic multiple myeloma progenitors, stem cell properties, and drug resistance. Cancer Res 2008;68(1):190-7

Kawano Y, Kikukawa Y, Fujiwara S, et al. Hypoxia reduces CD138 expression and induces an immature and stem celllike transcriptional program in myeloma cells. Int J Oncol 2013;43(6):1809-16

Expert Opin. Ther. Targets (2015) 19(12)

Expert Opin. Ther. Targets Downloaded from informahealthcare.com by Nyu Medical Center on 07/21/15 For personal use only.

RNAi for MM therapy

RNA interference therapeutic targeting VEGF and KSP in cancer patients with liver involvement. Cancer Discov 2013;3(4):406-17

47.

Chang-Yew Leow C, Gerondakis S, Spencer A. MEK inhibitors as a chemotherapeutic intervention in multiple myeloma. Blood Cancer J 2013;3:e105

48.

Kuehl WM, Bergsagel PL. Molecular pathogenesis of multiple myeloma and its premalignant precursor. J Clin Invest 2012;122(10):3456-63

49.

Guo J, Evans JC, O’Driscoll CM. Delivering RNAi therapeutics with nonviral technology: a promising strategy for prostate cancer? Trends Mol Med 2013;19(4):250-61

McCubrey JA, Steelman LS, Chappell WH, et al. Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR cascade inhibitors: how mutations can result in therapy resistance and how to overcome resistance. Oncotarget 2012;3(10):1068-111

50.

Godinho BM, Malhotra M, O’Driscoll CM, Cryan JF. Delivering a disease-modifying treatment for Huntington’s disease. Drug Discov Today 2015;20(1):50-64

Lu J, Zavorotinskaya T, Dai YM, et al. Pim2 is required for maintaining multiple myeloma cell growth through modulating TSC2 phosphorylation. Blood 2013;122(9):1610-20

51.

Neri P, Bahlis NJ. Pinning down myeloma with Pim2 inhibitors!. Blood 2013;122(9):1534-6

22.

Abdi J, Chen G, Chang H. Drug resistance in multiple myeloma: latest findings and new concepts on molecular mechanisms. Oncotarget 2013;4(12):2186-207

23.

Mahindra A, Laubach J, Raje N, et al. Latest advances and current challenges in the treatment of multiple myeloma. Nat Rev Clin Oncol 2012;9(3):135-43

24.

Andreu-Vieyra CV, Berenson JR. The potential of panobinostat as a treatment option in patients with relapsed and refractory multiple myeloma. Ther Adv Hematol 2014;5(6):197-210

37.

Kostarelos K, Miller AD. Synthetic, selfassembly ABCD nanoparticles; a structural paradigm for viable synthetic non-viral vectors. Chem Soc Rev 2005;34(11):970-94

25.

Guo J, Fisher KA, Darcy R, et al. Therapeutic targeting in the silent era: advances in non-viral siRNA delivery. Mol Biosyst 2010;6(7):1143-61

38.

26.

Wilson RC, Doudna JA. Molecular mechanisms of RNA interference. Annu Rev Biophys 2013;42:217-39

27.

Fellmann C, Lowe SW. Stable RNA interference rules for silencing. Nat Cell Biol 2014;16(1):10-18

28.

Guo JF, Bourre L, Soden DM, et al. Can non-viral technologies knockdown the barriers to siRNA delivery and achieve the next generation of cancer therapeutics? Biotechnol Adv 2011;29(4):402-17

29.

30.

31.

32.

33.

Kanasty R, Dorkin JR, Vegas A, Anderson D. Delivery materials for siRNA therapeutics. Nat Mater 2013;12(11):967-77 Mohr SE, Smith JA, Shamu CE, et al. RNAi screening comes of age: improved techniques and complementary approaches. Nat Rev Mol Cell Biol 2014;15(9):591-600 Wu SY, Lopez-Berestein G, Calin GA, Sood AK. RNAi therapies: drugging the undruggable. Sci Transl Med 2014;6(240):240ps247

36.

39.

Guo J, Cahill MR, McKenna SL, O’Driscoll CM. Biomimetic nanoparticles for siRNA delivery in the treatment of leukaemia. Biotechnol Adv 2014;32(8):1396-409

40.

Terpos E, Eleutherakis-Papaiakovou V, Dimopoulos MA. Clinical implications of chromosomal abnormalities in multiple myeloma. Leuk Lymphoma 2006;47(5):803-14

52.

Hiasa M, Teramachi J, Oda A, et al. Pim-2 kinase is an important target of treatment for tumor progression and bone loss in myeloma. Leukemia 2015;29(1):207-17

41.

Bommert K, Bargou RC, Stuhmer T. Signalling and survival pathways in multiple myeloma. Eur J Cancer 2006;42(11):1574-80 Discusses the therapeutic potential of signaling pathways in MM.

53.

Taylor CA, Liu Z, Tang TC, et al. Modulation of eIF5A expression using SNS01 nanoparticles inhibits NF-kappaB activity and tumor growth in murine models of multiple myeloma. Mol Ther 2012;20(7):1305-14 In vivo success of inhibiting NF-kB using siRNA for MM.

.

42.

43.

44.

Polivka J Jr, Janku F. Molecular targets for cancer therapy in the PI3K/AKT/ mTOR pathway. Pharmacol Ther 2014;142(2):164-75

54.

Morgan GJ, Walker BA, Davies FE. The genetic architecture of multiple myeloma. Nat Rev Cancer 2012;12(5):335-48

Carbone CJ, Fuchs SY. Eliminative signaling by Janus kinases: role in the downregulation of associated receptors. J Cell Biochem 2014;115(1):8-16

55.

Keane NA, Glavey SV, Krawczyk J, O’Dwyer M. AKT as a therapeutic target in multiple myeloma. Expert Opin Ther Targets 2014;18(8):897-915

Vainchenker W, Constantinescu SN. JAK/STAT signaling in hematological malignancies. Oncogene 2013;32(21):2601-13

56.

Munshi NC, Anderson KC, SpringerLink (Online service): Advances in Biology and Therapy of Multiple Myeloma Volume 1: Basic Science. ISBN: 978-1-4614-4666-8

57.

Zhang Q, Hossain DM, Nechaev S, et al. TLR9-mediated siRNA delivery for targeting of normal and malignant human hematopoietic cells in vivo. Blood 2013;121(8):1304-15 In vivo success of targeting STAT3 using siRNA for MM.

Williford JM, Wu J, Ren Y, et al. Recent advances in nanoparticle-mediated siRNA delivery. Annu Rev Biomed Eng 2014;16:347-70

45.

Yoo JW, Irvine DJ, Discher DE, Mitragotri S. Bio-inspired, bioengineered and biomimetic drug delivery carriers. Nat Rev Drug Discov 2011;10(7):521-35

Ikeda H, Hideshima T, Fulciniti M, et al. PI3K/p110{delta} is a novel therapeutic target in multiple myeloma. Blood 2010;116(9):1460-8

46.

Sahin I, Azab F, Mishima Y, et al. Targeting survival and cell trafficking in multiple myeloma and Waldenstrom macroglobulinemia using pan-class I PI3K inhibitor, buparlisib. Am J Hematol 2014;89(11):1030-6

34.

Yin H, Kanasty RL, Eltoukhy AA, et al. Non-viral vectors for gene-based therapy. Nat Rev Genet 2014;15(8):541-55

35.

Tabernero J, Shapiro GI, LoRusso PM, et al. First-in-humans trial of an

..

Expert Opin. Ther. Targets (2015) 19(12)

..

13

J. Guo et al.

58.

59.

60.

Expert Opin. Ther. Targets Downloaded from informahealthcare.com by Nyu Medical Center on 07/21/15 For personal use only.

61.

62.

63.

64.

65.

66.

67.

68.

..

69.

14

Clevers H, Nusse R. Wnt/beta-catenin signaling and disease. Cell 2012;149(6):1192-205

70.

Schmeel LC, Schmeel FC, Kim Y, et al. Targeting the Wnt/beta-catenin pathway in multiple myeloma. Anticancer Res 2013;33(11):4719-26

71.

Luis TC, Ichii M, Brugman MH, et al. Wnt signaling strength regulates normal hematopoiesis and its deregulation is involved in leukemia development. Leukemia 2012;26(3):414-21 Lento W, Congdon K, Voermans C, et al. Wnt signaling in normal and malignant hematopoiesis. Cold Spring Harb Perspect Biol 2013;5(2):pii: a008011

72.

.

Holien T, Vatsveen TK, Hella H, et al. Addiction to c-MYC in multiple myeloma. Blood 2012;120(12):2450-3

73.

Sewify EM, Afifi OA, Mosad E, et al. Amplification in Multiple Myeloma Is Associated With Multidrug Resistance Expression. Cl Lymph Myelom Leuk 2014;14(3):215-22

74.

Canalis E. Wnt signalling in osteoporosis: mechanisms and novel therapeutic approaches. Nat Rev Endocrinol 2013;9(10):575-83 Edwards CM, Edwards JR, Lwin ST, et al. Increasing Wnt signaling in the bone marrow microenvironment inhibits the development of myeloma bone disease and reduces tumor burden in bone in vivo. Blood 2008;111(5):2833-42 Baron R, Kneissel M. WNT signaling in bone homeostasis and disease: from human mutations to treatments. Nat Med 2013;19(2):179-92 Kobune M, Chiba H, Kato J, et al. Wnt3/RhoA/ROCK signaling pathway is involved in adhesion-mediated drug resistance of multiple myeloma in an autocrine mechanism. Mol Cancer Ther 2007;6(6):1774-84 Ashihara E, Kawata E, Nakagawa Y, et al. beta-catenin small interfering RNA successfully suppressed progression of multiple myeloma in a mouse model. Clin Cancer Res 2009;15(8):2731-8 In vivo success of suppressing b-catenin using siRNA for MM. Demchenko YN, Kuehl WM. A critical role for the NF-kappaB pathway in multiple myeloma. Oncotarget 2010;1(1):59-68

75.

76.

77.

Hoesel B, Schmid JA. The complexity of NF-kappaB signaling in inflammation and cancer. Mol Cancer 2013;12:86 Pateras I, Giaginis C, Tsigris C, et al. NF-kappaB signaling at the crossroads of inflammation and atherogenesis: searching for new therapeutic links. Expert Opin Ther Targets 2014;18(9):1089-101 Zhou HF, Yan H, Pan H, et al. Peptide-siRNA nanocomplexes targeting NF-kappaB subunit p65 suppress nascent experimental arthritis. J Clin Invest 2014;124(10):4363-74 siRNA nanocomplexes targeting NF-kB to suppress rheumatoid arthritis in vivo. Oliva L, Cenci S. Autophagy in plasma cell pathophysiology. Front Immunol 2014;5:103 Vincenz L, Jager R, O’Dwyer M, Samali A. Endoplasmic Reticulum Stress and the Unfolded Protein Response: Targeting the Achilles Heel of Multiple Myeloma. Mol Cancer Ther 2013;12(6):831-43 Lamy L, Ngo VN, Emre NC, et al. Control of autophagic cell death by caspase-10 in multiple myeloma. Cancer Cell 2013;23(4):435-49 Bolli N, Avet-Loiseau H, Wedge DC, et al. Heterogeneity of genomic evolution and mutational profiles in multiple myeloma. Nat Commun 2014;5:2997 Colombo M, Mirandola L, Platonova N, et al. Notch-directed microenvironment reprogramming in myeloma: a single path to multiple outcomes. Leukemia 2013;27(5):1009-18

78.

Zhang L, Fok JH, Davies FE. Heat shock proteins in multiple myeloma. Oncotarget 2014;5(5):1132-48

79.

Chesi M, Bergsagel PL. Epigenetics and microRNAs combine to modulate the MDM2/p53 axis in myeloma. Cancer Cell 2010;18(4):299-300

80.

Furukawa Y, Kikuchi J. Molecular pathogenesis of multiple myeloma. Int J Clin Oncol 2015;20(3):413-22

81.

Novobrantseva TI, Borodovsky A, Wong J, et al. Systemic RNAi-mediated gene silencing in nonhuman primate and rodent myeloid cells. Mol Ther Nucleic Acids 2012;1:e4

82.

Mann AP, Tanaka T, Somasunderam A, et al. E-Selectin-Targeted Porous Silicon Particle for Nanoparticle Delivery to the Expert Opin. Ther. Targets (2015) 19(12)

..

83.

..

Bone Marrow. Adv Mater 2011;23(36):H278-82 Demonstrates a systemic delivery system that can specifically target the BM vasculature. Swami A, Reagan MR, Basto P, et al. Engineered nanomedicine for myeloma and bone microenvironment targeting. Proc Natl Acad Sci USA 2014;111(28):10287-92 Develops a bone-homing nanoparticle system to deliver bortezomib and inhibit myeloma.

84.

McHeyzer-Williams M, Okitsu S, Wang N, McHeyzer-Williams L. Molecular programming of B cell memory. Nat Rev Immunol 2012;12(1):24-34

85.

Nutt SL, Tarlinton DM. Germinal center B and follicular helper T cells: siblings, cousins or just good friends? Nat Immunol 2011;12(6):472-7

86.

Walsh D, McCarthy J, O’Driscoll C, Melgar S. Pattern recognition receptors– molecular orchestrators of inflammation in inflammatory bowel disease. Cytokine Growth Factor Rev 2013;24(2):91-104

87.

Sherbenou DW, Behrens CR, Su Y, et al. The development of potential antibody-based therapies for myeloma. Blood Rev 2015;29(2):81-91

88.

Robert F, Roman W, Bramoulle A, et al. Translation initiation factor eIF4F modifies the dexamethasone response in multiple myeloma. Proc Natl Acad Sci USA 2014;111(37):13421-6

89.

Chatterjee M, Andrulis M, Stuhmer T, et al. The PI3K/Akt signaling pathway regulates the expression of Hsp70, which critically contributes to Hsp90-chaperone function and tumor cell survival in multiple myeloma. Haematologica 2013;98(7):1132-41

90.

Suzuki E, Daniels TR, Helguera G, et al. Inhibition of NF-kappaB and Akt pathways by an antibody-avidin fusion protein sensitizes malignant B-cells to cisplatin-induced apoptosis. Int J Oncol 2010;36(5):1299-307

91.

Sahin I, Moschetta M, Mishima Y, et al. Distinct roles of class I PI3K isoforms in multiple myeloma cell survival and dissemination. Blood Cancer J 2014;4:e204

92.

Cea M, Cagnetta A, Fulciniti M, et al. Targeting NAD+ salvage pathway

RNAi for MM therapy

induces autophagy in multiple myeloma cells via mTORC1 and extracellular signal-regulated kinase (ERK1/2) inhibition. Blood 2012;120(17):3519-29

Expert Opin. Ther. Targets Downloaded from informahealthcare.com by Nyu Medical Center on 07/21/15 For personal use only.

93.

99.

Purushothaman A, Babitz SK, Sanderson RD. Heparanase enhances the insulin receptor signaling pathway to activate extracellular signal-regulated kinase in multiple myeloma. J Biol Chem 2012;287(49):41288-96

94.

Steinbrunn T, Stuhmer T, Gattenlohner S, et al. Mutated RAS and constitutively activated Akt delineate distinct oncogenic pathways, which independently contribute to multiple myeloma cell survival. Blood 2011;117(6):1998-2004

95.

Liu Z, Li T, Jiang K, et al. Induction of chemoresistance by all-trans retinoic acid via a noncanonical signaling in multiple myeloma cells. PLoS One 2014;9(1):e85571

96.

Atanackovic D, Hildebrandt Y, Templin J, et al. Role of interleukin 16 in multiple myeloma. J Natl Cancer Inst 2012;104(13):1005-20

97.

He J, Liu Z, Zheng Y, et al. p38 MAPK in myeloma cells regulates osteoclast and osteoblast activity and induces bone destruction. Cancer Res 2012;72(24):6393-402

98.

multiple myeloma and supports the growth of myeloma cells. Oncogene 2011;30(28):3198-206

Fagerli UM, Ullrich K, Stuhmer T, et al. Serum/glucocorticoid-regulated kinase 1 (SGK1) is a prominent target gene of the transcriptional response to cytokines in

Lee C, Oh JI, Park J, et al. TNF alpha mediated IL-6 secretion is regulated by JAK/STAT pathway but not by MEK phosphorylation and AKT phosphorylation in U266 multiple myeloma cells. Biomed Res Int 2013;2013:580135

100.

Bolzoni M, Donofrio G, Storti P, et al. Myeloma cells inhibit non-canonical wnt co-receptor ror2 expression in human bone marrow osteoprogenitor cells: effect of wnt5a/ror2 pathway activation on the osteogenic differentiation impairment induced by myeloma cells. Leukemia 2013;27(2):451-63

101.

Raab MS, Breitkreutz I, Tonon G, et al. Targeting PKC: a novel role for beta-catenin in ER stress and apoptotic signaling. Blood 2009;113(7):1513-21

102.

Zhou W, Yang Y, Xia J, et al. NEK2 induces drug resistance mainly through activation of efflux drug pumps and is associated with poor prognosis in myeloma and other cancers. Cancer Cell 2013;23(1):48-62

103.

Annunziata CM, Davis RE, Demchenko Y, et al. Frequent engagement of the classical and alternative NF-kappaB pathways by diverse genetic abnormalities in multiple myeloma. Cancer Cell 2007;12(2):115-30

Expert Opin. Ther. Targets (2015) 19(12)

104. Rauert-Wunderlich H, Siegmund D, Maier E, et al. The IKK inhibitor Bay 11-7082 induces cell death independent from inhibition of activation of NFkappaB transcription factors. PLoS One 2013;8(3):e59292

Affiliation Jianfeng Guo1 PhD, Sharon L McKenna2 PhD, Michael E O’Dwyer3,4,5 MD, Mary R Cahill6 MD & Caitriona M O’Driscoll†7 PhD † Author for correspondence 1 Government of Ireland Postdoctoral Researcher, University College Cork, School of Pharmacy, Pharmacodelivery Group, Cork, Ireland 2 Principal Investigator, University College Cork, Cork Cancer Research Centre, Cork, Ireland 3 Consultant Haematologist and Professor of Haematology, Glycoscience Research Group, National University of Ireland, Galway, Ireland 4 Department of Hematology National University of Ireland, Galway, Ireland 5 Galway University Hospital, Galway, Ireland 6 Consultant Haematologist and Clinical Professor, Department of Haematology, Cork University Hospital, Cork, Ireland 7 Professor of Pharmaceutics, University College Cork, School of Pharmacy, Pharmacodelivery Group, Cork, Ireland Tel: +353 21 4901396; Fax: +353 21 4901656; E-mail: [email protected]

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RNA interference for multiple myeloma therapy: targeting signal transduction pathways.

Multiple myeloma (MM) is a hematological malignancy characterized by infiltration of malignant plasma cells in the bone marrow (BM) and end-organ dama...
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