Multiple myeloma (MM) is characterized by the accumulation of malignant plasma cells in the bone marrow and excessive production of a single monoclonal immunoglobulin. MM is the second leading cause of blood cancer worldwide, which accounts for approximately 10% of hematologic malignancies [1]. A recent increase in the incidence of MM has been described in Asian countries such as Japan, Korea, and Taiwan [2,3,4]. MM is more likely to occur in elderly men [4]. Generally, patients with early-stage MM are without symptoms (asymptomatic); however, symptoms develop following disease progression. The most typical clinical manifestations of MM include hypercalcemia, renal failure, anemia, and bone fractures (CRAB symptoms) [5]. Over the past several years, treatment strategies have expanded extensively for MM patients. The development of new drugs has led to significant improvements in prognosis and overall survival (OS) among patients. Key drugs for treatment are immunomodulatory imide drugs (IMiDs) (thalidomide, lenalidomide, and pomalidomide), proteasome inhibitors (bortezomib, carfilzomib, and ixazomib), monoclonal antibodies (daratumumab and elotuzumab), immune-based therapies (CAR-T cell and bispecific T-cell engager [BiTE]), and small-molecule inhibitors targeting FGFR3 and RAS/MAPK signaling pathways (erdafitinib, vemurafenib, and cobimetinib) [6, 7]. However, despite recent advances in therapeutic approaches in MM, it remains incurable. Although most patients generally respond to the standard first-line treatment, many of them would inevitably relapse or become refractory to the disease [6].
Conventional karyotyping and fluorescence in situ hybridization (FISH) are the gold standard techniques used for detecting cytogenetic abnormalities in MM. Conventional karyotypes are prepared from mitotic cells that have been arrested at the metaphase stage. About 30–50% of cytogenetic changes in MM can be detected using karyotyping; however, they are frequently acquired from advanced cases [8]. Conventional karyotyping tends to miss chromosomal aberration in early stages of the disease, due to its low mitotic index and number of malignant plasma cells. The low resolution of conventional karyotyping has also limited the identification of cryptic, subtle, or complex chromosomal changes. FISH, on the other hand, is more sensitive than classical cytogenetics. FISH uses interphase nuclei and thus does not require cell proliferation. It uses fluorescence-labeled probes to detect specific DNA sequences on the chromosome. Consequently, up to 80% of chromosomal aberration in MM can be identified by FISH [9]. The sensitivity of FISH testing can be enhanced with plasma cell enrichment of bone marrow specimens. Nevertheless, the main disadvantage of FISH is that it can only detect known chromosomal aberrations. Recently, with the progress in molecular techniques such as gene expression profiling (GEP) and next-generation sequencing (NGS), various driver gene mutations, oncogenic dependencies, structural variants (SVs), chromoplexy, and chromothripsis in relation to MM clonality and evolution have been discovered. Although GEP and NGS have paved a way toward a more effective management of MM patients, most of the developing countries in Asia, including Malaysia, are still relying on conventional methods (G-banding karyotype and FISH) to detect structural and numerical chromosomal aberrations in MM. The current challenge is whether new emerging molecular techniques should be adopted in the routine clinical practice of MM to improve the diagnosis and management of MM patients.
In this review, we describe the recurrent primary and secondary oncogenic events in MM and their clinical impacts on the prognosis of patients. We also summarize the most prominent findings concerning genomic changes (molecular signatures, new and rare somatic mutations) contributed by GEP and NGS studies of MM. We compiled the relevant articles for this review from electronic databases, including PubMed, Google Scholar, Scopus, and Web of Science. Keywords used for searches included MM, translocation, primary and secondary oncogenic events, chromosomal abnormalities, gene expression, NGS, mutation, clonal evolution, genomic landscape, and SVs. All relevant articles were carefully reviewed, and more articles were searched based on the citation in the articles.
MM is a heterogeneous and genetically complex disease. It is thought to occur via multiple primary and secondary oncogenic events (
Primary and secondary oncogenic events in the molecular pathogenesis of MM. MGUS, monoclonal gammopathy of undetermined significance; MM, multiple myeloma; SMM, smoldering multiple myeloma.
Common primary and secondary chromosomal abnormalities and their prognostic outcomes in MM
Hyperdiploidy | Trisomies of odd-numbered chromosomes | 50–60 | Standard risk | Favorable |
Trisomy 21-negative impact on OS |
[12,13,14,15,16,17] |
IgH translocations | 60 | Occur as early as the MGUS stage |
[12, 18, 19] | |||
t(4;14)(p16;q32) | 15 | High risk | Unfavorable/neutral |
Commonly developed from errors in CSR |
[10, 17, 20,21,22,23,24,25] | |
t(14;16)(q32;q23) | 5–7 | High risk | Unfavorable | [14, 19, 20, 26] | ||
t(14;20)(q32;q11) | 1–2 | High risk | Unfavorable | [14, 20, 27] | ||
t(11;14)(q13;q32) | 15–20 | Standard risk | Favorable/neutral | Commonly caused by SHM |
[10, 28,29,30] | |
Others: | ||||||
t(12;14)(p13;q32) | <1 | Standard risk | Favorable/neutral | [19] | ||
t(6;14)(p21;q32) | 2 | Standard risk | Favorable/neutral | |||
t(8;14)(q24.3;q32) | <1 | High risk | Unfavorable | |||
del(1p) | 30 | High risk | Unfavorable |
1p21 and 1p32 are the most frequently deleted regions | [31,32,33] | |
Gain (1q21) | 40 | High risk | Unfavorable |
Rarely found in MGUS |
[34,35,36,37] | |
Monosomy 13/ del(13q) | 50 | Neutral | 85% are monosomy; 15% are partial deletion |
[8, 32, 38,39,40] | ||
del(17p) | 5–10 NDMM |
High risk | Unfavorable |
Late event in pathogenesis |
[11, 41, 42] | |
MYC translocation | 15 early-stage MM |
Unfavorable/neutral |
[11, 12, 42,43,44,45,46,47,48,49] | |||
Others | ||||||
del(11q) | 7 | [12, 32, 50, 51] | ||||
del(16q) | 35 | Unfavorable | ||||
del(14q) | 38 | Unfavorable | ||||
del(12p) | Unfavorable | |||||
del(8p) | Unfavorable |
CSR, class-switch recombination; MGUS, monoclonal gammopathy of undetermined significance; MM, multiple myeloma; NDMM, newly diagnosed multiple myeloma; OS, overall survival; PFS, progression-free survival; RRMM, relapsed or refractory multiple myeloma; SHM, somatic hypermutation; SMM, smoldering multiple myeloma.
The co-occurrence of two or three high-risk genetic alterations is defined as double-/triple-hit MM. Based on the criteria stated in the IMWG, clinical outcomes of double-hit MM are similar regardless of whether they are classified as high-risk or low/standard-risk MM. Thus, defining the molecular features of double-hit MM is important to improve risk stratification and outcome prediction in MM. Double-hit MM is found in 1 of 33 newly diagnosed multiple myeloma (NDMM) [40]. According to Baysal et al. [52], the OS of MM patients with double-hit, single high-risk, and no high-risk MM were 6 months, 32 months, and 57 months, respectively. The same study found that the hazard ratio was worsened in triple-hit (7.30) than double-hit (5.55) and single high-risk (1.42) MM patients [52]. Another study revealed a close relationship between the co-occurrence of translocation t(4;14) and
Transcriptome changes play a critical role in myelomagenesis as early as in the monoclonal gammopathy of undetermined significance (MGUS) stage [55]. In the past decades, GEP or microarray has emerged as one of the most popular approaches in the identification of transcriptome changes, molecular mechanisms, and pathways underlying human cancers. GEP has contributed to the characterization of disease subtypes, risk stratification, prognosis, and outcomes in MM patients. One of the most prominent results from GEP was the discovery of a 70-gene signature model by the University of Arkansas for Medical Sciences (UAMS) in 2007 [56]. The 70-gene signature was found to be superior in risk and prognostic stratification of NDMM. They found that most of the crucial genes related to disease progression were localized in chromosome 1. From the same study, 17-gene subsets for high-risk MM were also revealed. Among the important genes found in the 17-gene subset prediction model are
Another significant study was reported by Broyl et al. [57]. They discovered seven unique clusters, which were associated with NDMM based on the gene expression profiles: i. translocation [MMSET/t(4;14), MAF/t(14;16)/t(14;20), cyclin D1/t(11;14), and cyclin D2/t(6;14)]; ii. hyperdiploidy (HY); iii. proliferation-associated genes (PR); iv. low percentage of bone disease (LB); v. overexpression of cancer testis antigens (CTAs); vi. NFκB pathway; and vii. overexpression of protein tyrosine phosphatases (PRLs) [57]. Among the seven clusters, three were newly identified (v–vii), while the rest were consistent with the UAMS classification [58].
The EMC-92 gene signature prediction model for high-risk MM was developed by another group of scientists from the Netherlands based on the findings from the GEP [59]. Survivin/
Although GEP has been successfully used to characterize MM into different clinicopathological subtypes and molecular signatures for diagnosis, staging, risk stratification, and prognostication of MM, data generated by GEP are inadequate to fully delineate the molecular biology of this highly genetically complex disease. In recent years, NGS has become a promising tool in the study of IgH translocations, V(D)J clonal rearrangements, IgH isotype, CNAs, and somatic mutations simultaneously in a more refined manner.
The very first mutational profiles generated by NGS data in the past 10 years revealed that there is no specific or unique mutation in MM. The observation remains valid in the latest findings [61, 62]. To date, approximately 250 mutated genes have been found in MM, and about a quarter of them are identified as driver genes [62].
Recurrent gene aberrations and their frequency in MM. MM, Multiple myeloma.
NGS findings not only confirm the heterogeneous complexity of MM but also show that most of the driver mutations are present in the subclonal population, and multiple mutations are detected in different genes within the same pathway, suggesting a diverse pattern of clonal evolution in MM, which evolves through space and time [10, 70]. The main clonal evolution in MM includes neutral evolution, branching evolution, and linear evolution [10, 71]. Neutral evolution occurs when all descendant subclones show a similar ability to survive under certain circumstances, and the evolutionary changes are not caused by selective pressure. Branching evolution is the early divergence of subclones with different mutations, which evolves further over time, possibly driven by the selective pressure of the bone marrow microenvironment and treatment, or inherent tumor characteristics, or both. In linear evolution, a single subclonal population is fully substituted by another highly adapted subclone [10, 71]. The main clonal evolution models in MM are visualized in
Main clonal evolution models in multiple myeloma. Each color represents a single subclone.
Despite the discovery of driver gene mutations and clonal evolution in MM, another major contribution by NGS was the uncovering of complex SVs in MM. Complex SVs are usually difficult to detect using conventional sequencing approaches. Common complex SVs in MM include chromothripsis, chromoplexy, and multiple templated insertions [73]. Generally, MM patients present with at least one complex SV (80%) [38]. Chromothripsis, a form of chromoanagenesis, was first detected in chronic lymphoblastic leukemia (CLL) [74]. Chromothripsis is characterized by ten to hundreds of chromosomes breaking and rejoining in confined genomic regions in one or a few chromosomes (
Complex chromosomal rearrangements in multiple myeloma: chromothripsis and chromoplexy.
NGS also contributes to the revelation of the activity-induced deaminase (
On the other hand,
The genomic spectrum of MGUS and SMM has been studied using NGS as well, although they are not as much as in MM. MGUS is a benign noncancerous condition, with a ~1% progression rate to MM per year. In contrast, SMM is a precancerous or early-stage MM, with a 10% annual progression risk to active MM in the first 5 years of diagnosis [86].
By comparing NGS data from paired SMM and MM from the same individual, Bolli et al. [53] postulated that most of the driver mutations such as hyperdiploidy,
In addition to comparing genomic data between SMM and MM, WGS has also been used to study the differences in the genomic landscape and temporal acquisition of myeloma-associated genomic events between clinically stable and progressive myeloma precursor conditions (MGUS and SMM) [87]. Compared with the clinically progressive myeloma precursor condition, the clinically stable myeloma precursor condition was associated with late initiation of the first clonal copy number alteration in patients’ life and absence or lower number of mutations in driver genes (genes involved in MAPK and NF-κB pathways,
In summary, advances in technology have expanded our knowledge of tumor heterogeneity, mutational landscape, clonal composition, and dynamic evolution of MM. Evidence has confirmed that myeloma is a highly heterogeneous disease, and one single treatment regimen does not fit all MM patients. Thus, precision medicine will become inevitably important in future myeloma therapy. NGS, which allows the analysis of the full spectrum of recurrent mutations and chromosomal abnormalities in MM, can become a powerful tool toward precision medicine in the treatment of MM. Importantly, NGS also enables rapid and accurate detection of clonal and subclonal mutations, which present at low variant allele frequencies in patients. Many of these subtle genetic changes have a significant influence on the drug efficacy and prognosis outcomes of the patients. NGS has been proven to provide significant clinical benefits and should be transitioned from research to clinical use. However, NGS is laborious, time-consuming, and remains expensive even today, thus preventing its routine use in clinical settings. Malaysia, one of the developing countries in Southeast Asia, also faces the same challenges. So far, we are relying on conventional karyotyping and FISH for routine diagnosis, risk assessment, and therapeutic selection of MM patients. As discussed earlier, karyotyping and FISH have limited resolution and therefore are inadequate to capture the complexity of the MM genome. As technological advances will continue to push NGS toward lower costs with more user-friendly methodologies and data analysis pipelines, we hope that NGS will be integrated into routine clinical practice at diagnosis/relapse to guide disease management and allow for precision medicine in future.