Review
Expert Review of Molecular Diagnostics Downloaded from informahealthcare.com by Nyu Medical Center on 10/12/14 For personal use only.
The potential of miRNAs as biomarkers for multiple myeloma Expert Rev. Mol. Diagn. Early online, 1–13 (2014)
Alberto Rocci1, Craig C Hofmeister2 and Flavia Pichiorri*2 1 Department of Haematology, Manchester Royal Infirmary Hospital, Manchester, UK 2 Division of Hematology, Department of Internal Medicine, Comprehensive Cancer Center, The Ohio State University, Columbus, OH, USA *Author for correspondence: [email protected]
Since the discovery of the link between miRNA and cancer, miRNAs have been investigated in virtually all tumors. Their ability to add a novel level of gene regulation and to target genes apparently not linked to each other has greatly intrigued researchers and physicians alike. In this review, the role of miRNAs in multiple myeloma (MM) is summarized, with particular attention to their potential as biomarkers. The promising role of circulating miRNAs in diagnosis and risk stratification is also discussed, as well as preliminary results of miRNA-based therapeutic approaches. Finally, the critical issues in miRNA analysis in MM and ongoing strategies to solve them are discussed. The ability to standardize miRNA analysis procedures will permit the inclusion of miRNA evaluation alongside available stratification tools, paving the way for personalized medicine in MM. KEYWORDS: biomarkers • circulating miRNAs • exosomes • miRNAs • multiple myeloma • staging system
Multiple Myeloma (MM) is the second most frequent hematological cancer in developed countries, and is estimated to account for 24,000 new diagnoses and 11,000 deaths in 2014 in the USA [1]. MM is characterized by the expansion of clonal plasma cells (PCs) in the bone marrow (BM) leading to bone disease, anemia, kidney failure and eventually death. In all cases, MM is preceded by a premalignant condition known as monoclonal gammopathy of undetermined significance (MGUS) that can directly evolve to MM or can be characterized by a slower progression through a smoldering MM (sMM) phase [2–5]. The biologic transition from normal PCs to MGUS and then to MM consists of many oncogenic events, including dysregulation of a cyclin D genes [1–5], activating mutations of NRAS and KRAS and constitutive activation of NF-kB (FIGURE 1). Other molecular alterations that characterize this progression include chromosomal translocations of several oncogenes to the heavy chain Ig (IgH) locus causing their overexpression, deletion of chromosomal arms causing tumor suppressor downregulation and mutations in several critical genes. Distinct clones can become predominant in different stages during the natural history of MM, resulting in drug resistance and ultimately refractoriness to treatment [6,7]. informahealthcare.com
10.1586/14737159.2014.946906
MM cells are also strongly dependent on the BM microenvironment (e.g., fibroblasts/bone marrow stromal cells [BMSCs], endothelial cells and osteoclasts). Such cells have a pivotal role in the regulation of MM cell growth, survival and drug resistance [6–9] via the production of supportive growth factors, for example, IL-6, IL-8, IGF-1, VEGF and integrin contacts [10–13]. In the last 10 years, the treatment of MM has radically changed with the introduction of novel drugs into routine practice and, in fit patients, of autologous stem cell transplant. The inclusion of immunomodulatory agents (e.g., lenalidomide, pomalidomide) and proteasome inhibitors (e.g., bortezomib, carfilzomib) together with alkylating agents (melphalan, cyclophosphamide) and steroids has resulted in a concrete improvement in overall response rate, progression-free survival (PFS) and overall survival (OS) for MM patients [14,15], with a few able to achieve longterm remission. However, the clinical behavior of the disease is extremely variable; some patients become quickly refractory to any treatment with an OS of less than 24 months, while others can have an OS greater than 10 years. Our ability to predict patient outcome and stratify them to distinct risk classes is currently based on the International Staging System
2014 Informa UK Ltd
ISSN 1473-7159
1
Review
Rocci, Hofmeister & Pichiorri
MGUS
MM
PCL
Secondary translocations MYC translocation
Expert Review of Molecular Diagnostics Downloaded from informahealthcare.com by Nyu Medical Center on 10/12/14 For personal use only.
Cyclin D1, D2, D3 overexpression Primary chromosomal abnormalities: hyperdiploid vs nonhyperdiploid
NRAS, NFκB, FGFR3 activation KRAS, BRAF activation TP53 and RB inactivation miR-21, miR-25, miR-181s, miR-106a miR-15a, miR-16, miR-19s, miR-32
Bone resorption Angiogenesis
Figure 1. Schematic representation of multiple myeloma pathogenesis. Together with chromosomal abnormalities and gene expression alterations, aberrant miRNA expression has been found to be responsible for MM pathogenesis. Modulation of specific miRNAs characterizes the PCs of MM patients, determining abnormal expression of particular genes involved in cell growth and apoptosis. miRNAs also play a role in the interaction of PCs with surrounding cells in creating a microenvironment that stimulates tumor cell survival and growth. MGUS: Monoclonal gammopathy of undetermined significance; MM: Multiple myeloma.
(ISS) and on the presence or absence of distinct cytogenetic abnormalities or mutations [2–16]. The ability to predict response to therapy, clinical outcome and even to identify patients prone to develop a idiosyncratic drug-specific toxicity would be enormously helpful in deciding which treatment is most suitable for a particular patient. Due to the significant heterogeneity, both of patients and of MM itself, it is difficult to create an algorithm to help physicians in the decision-making process. With increased therapeutic options, there is an even greater need for strong biomarkers to inform the decision process and to identify patients who could achieve benefit from a specific drug. There is widespread evidence that epigenetic control mechanisms, including miRNAs, become deregulated in cancers and are involved in tumor initiation and progression. This is also true for MM, highlighting the importance of a new level of investigation in this field [17–20]. miRNAs are short non-coding RNAs that bind to complementary sequences on target mRNAs transcripts and in turn induce mRNA translational repression or degradation, leading to protein downregulation [21]. Aberrant miRNA expression has been shown to have a prognostic impact in patients with chronic lymphocytic leukemia (CLL) [22,23], acute myeloid leukemia [24,25] and other cancers [26]. The interest in a possible biologic and prognostic role of miRNAs is not limited to those found intracellularly, but recently has also been extended to those found in different body fluids [27,28]. doi: 10.1586/14737159.2014.946906
Extracellular miRNAs may be encapsulated in extracellular vesicles (EV) or associated with the Argonaute-2 (AGO-2) complex outside EV, and are extremely stable in either context [29–32]. Emerging studies have shown that circulating miRNAs can be detected in patients with a variety of malignancies, including MM [33,34], and that they could be potentially used as non-invasive diagnostic markers [34,35]. Previous studies suggested a relevant biologic role for cellular and circulating miRNAs in MM, but whether this translates into a clinical impact remains unknown [8,36,37]. In this review, we will summarize how miRNAs are abnormally expressed in MM, their biological significance in MM pathogenesis and their possible use as biomarkers. We will also analyze factors that might limit their routine clinical use, with an overview on their future potential in MM prognosis, diagnosis and treatment. Prognostic markers in MM
Several parameters have been investigated as prognostic markers in MM, but so far only two approaches have been largely employed: the combination of albumin and b2-microglobulin as part of the ISS, and the presence of chromosomal abnormalities detected via FISH. However, these two systems fail to reliably predict clinical outcome, and novel tools to improve patient stratification are required. The ISS, proposed in 2005, was developed using data from 10,750 newly diagnosed MM patients [38]. It is based on the measurement of b2-microglobulin and albumin, two factors that together reflect disease severity and patient condition. Patients with a b2-microglobulin
Expert Review of Molecular Diagnostics Downloaded from informahealthcare.com by Nyu Medical Center on 10/12/14 For personal use only.
The potential of miRNAs as biomarkers for multiple myeloma Expert Rev. Mol. Diagn. Early online, 1–13 (2014)
Alberto Rocci1, Craig C Hofmeister2 and Flavia Pichiorri*2 1 Department of Haematology, Manchester Royal Infirmary Hospital, Manchester, UK 2 Division of Hematology, Department of Internal Medicine, Comprehensive Cancer Center, The Ohio State University, Columbus, OH, USA *Author for correspondence: [email protected]
Since the discovery of the link between miRNA and cancer, miRNAs have been investigated in virtually all tumors. Their ability to add a novel level of gene regulation and to target genes apparently not linked to each other has greatly intrigued researchers and physicians alike. In this review, the role of miRNAs in multiple myeloma (MM) is summarized, with particular attention to their potential as biomarkers. The promising role of circulating miRNAs in diagnosis and risk stratification is also discussed, as well as preliminary results of miRNA-based therapeutic approaches. Finally, the critical issues in miRNA analysis in MM and ongoing strategies to solve them are discussed. The ability to standardize miRNA analysis procedures will permit the inclusion of miRNA evaluation alongside available stratification tools, paving the way for personalized medicine in MM. KEYWORDS: biomarkers • circulating miRNAs • exosomes • miRNAs • multiple myeloma • staging system
Multiple Myeloma (MM) is the second most frequent hematological cancer in developed countries, and is estimated to account for 24,000 new diagnoses and 11,000 deaths in 2014 in the USA [1]. MM is characterized by the expansion of clonal plasma cells (PCs) in the bone marrow (BM) leading to bone disease, anemia, kidney failure and eventually death. In all cases, MM is preceded by a premalignant condition known as monoclonal gammopathy of undetermined significance (MGUS) that can directly evolve to MM or can be characterized by a slower progression through a smoldering MM (sMM) phase [2–5]. The biologic transition from normal PCs to MGUS and then to MM consists of many oncogenic events, including dysregulation of a cyclin D genes [1–5], activating mutations of NRAS and KRAS and constitutive activation of NF-kB (FIGURE 1). Other molecular alterations that characterize this progression include chromosomal translocations of several oncogenes to the heavy chain Ig (IgH) locus causing their overexpression, deletion of chromosomal arms causing tumor suppressor downregulation and mutations in several critical genes. Distinct clones can become predominant in different stages during the natural history of MM, resulting in drug resistance and ultimately refractoriness to treatment [6,7]. informahealthcare.com
10.1586/14737159.2014.946906
MM cells are also strongly dependent on the BM microenvironment (e.g., fibroblasts/bone marrow stromal cells [BMSCs], endothelial cells and osteoclasts). Such cells have a pivotal role in the regulation of MM cell growth, survival and drug resistance [6–9] via the production of supportive growth factors, for example, IL-6, IL-8, IGF-1, VEGF and integrin contacts [10–13]. In the last 10 years, the treatment of MM has radically changed with the introduction of novel drugs into routine practice and, in fit patients, of autologous stem cell transplant. The inclusion of immunomodulatory agents (e.g., lenalidomide, pomalidomide) and proteasome inhibitors (e.g., bortezomib, carfilzomib) together with alkylating agents (melphalan, cyclophosphamide) and steroids has resulted in a concrete improvement in overall response rate, progression-free survival (PFS) and overall survival (OS) for MM patients [14,15], with a few able to achieve longterm remission. However, the clinical behavior of the disease is extremely variable; some patients become quickly refractory to any treatment with an OS of less than 24 months, while others can have an OS greater than 10 years. Our ability to predict patient outcome and stratify them to distinct risk classes is currently based on the International Staging System
2014 Informa UK Ltd
ISSN 1473-7159
1
Review
Rocci, Hofmeister & Pichiorri
MGUS
MM
PCL
Secondary translocations MYC translocation
Expert Review of Molecular Diagnostics Downloaded from informahealthcare.com by Nyu Medical Center on 10/12/14 For personal use only.
Cyclin D1, D2, D3 overexpression Primary chromosomal abnormalities: hyperdiploid vs nonhyperdiploid
NRAS, NFκB, FGFR3 activation KRAS, BRAF activation TP53 and RB inactivation miR-21, miR-25, miR-181s, miR-106a miR-15a, miR-16, miR-19s, miR-32
Bone resorption Angiogenesis
Figure 1. Schematic representation of multiple myeloma pathogenesis. Together with chromosomal abnormalities and gene expression alterations, aberrant miRNA expression has been found to be responsible for MM pathogenesis. Modulation of specific miRNAs characterizes the PCs of MM patients, determining abnormal expression of particular genes involved in cell growth and apoptosis. miRNAs also play a role in the interaction of PCs with surrounding cells in creating a microenvironment that stimulates tumor cell survival and growth. MGUS: Monoclonal gammopathy of undetermined significance; MM: Multiple myeloma.
(ISS) and on the presence or absence of distinct cytogenetic abnormalities or mutations [2–16]. The ability to predict response to therapy, clinical outcome and even to identify patients prone to develop a idiosyncratic drug-specific toxicity would be enormously helpful in deciding which treatment is most suitable for a particular patient. Due to the significant heterogeneity, both of patients and of MM itself, it is difficult to create an algorithm to help physicians in the decision-making process. With increased therapeutic options, there is an even greater need for strong biomarkers to inform the decision process and to identify patients who could achieve benefit from a specific drug. There is widespread evidence that epigenetic control mechanisms, including miRNAs, become deregulated in cancers and are involved in tumor initiation and progression. This is also true for MM, highlighting the importance of a new level of investigation in this field [17–20]. miRNAs are short non-coding RNAs that bind to complementary sequences on target mRNAs transcripts and in turn induce mRNA translational repression or degradation, leading to protein downregulation [21]. Aberrant miRNA expression has been shown to have a prognostic impact in patients with chronic lymphocytic leukemia (CLL) [22,23], acute myeloid leukemia [24,25] and other cancers [26]. The interest in a possible biologic and prognostic role of miRNAs is not limited to those found intracellularly, but recently has also been extended to those found in different body fluids [27,28]. doi: 10.1586/14737159.2014.946906
Extracellular miRNAs may be encapsulated in extracellular vesicles (EV) or associated with the Argonaute-2 (AGO-2) complex outside EV, and are extremely stable in either context [29–32]. Emerging studies have shown that circulating miRNAs can be detected in patients with a variety of malignancies, including MM [33,34], and that they could be potentially used as non-invasive diagnostic markers [34,35]. Previous studies suggested a relevant biologic role for cellular and circulating miRNAs in MM, but whether this translates into a clinical impact remains unknown [8,36,37]. In this review, we will summarize how miRNAs are abnormally expressed in MM, their biological significance in MM pathogenesis and their possible use as biomarkers. We will also analyze factors that might limit their routine clinical use, with an overview on their future potential in MM prognosis, diagnosis and treatment. Prognostic markers in MM
Several parameters have been investigated as prognostic markers in MM, but so far only two approaches have been largely employed: the combination of albumin and b2-microglobulin as part of the ISS, and the presence of chromosomal abnormalities detected via FISH. However, these two systems fail to reliably predict clinical outcome, and novel tools to improve patient stratification are required. The ISS, proposed in 2005, was developed using data from 10,750 newly diagnosed MM patients [38]. It is based on the measurement of b2-microglobulin and albumin, two factors that together reflect disease severity and patient condition. Patients with a b2-microglobulin
