Concurrent lipidomics and proteomics on malignant plasma cells from multiple myeloma patients: Probing the lipid metabolome.
Mohamed Ahmed,Collins Joel,Jiang Hui,Molendijk Jeffrey,Stoll Thomas,Torta Federico,Wenk Markus R,Bird Robert J,Marlton Paula,Mollee Peter,Markey Kate A,Hill Michelle M
PloS one
BACKGROUND:Multiple myeloma (MM) is a hematological malignancy characterized by the clonal expansion of malignant plasma cells. Though durable remissions are possible, MM is considered incurable, with relapse occurring in almost all patients. There has been limited data reported on the lipid metabolism changes in plasma cells during MM progression. Here, we evaluated the feasibility of concurrent lipidomics and proteomics analyses from patient plasma cells, and report these data on a limited number of patient samples, demonstrating the feasibility of the method, and establishing hypotheses to be evaluated in the future. METHODS:Plasma cells were purified from fresh bone marrow aspirates using CD138 microbeads. Proteins and lipids were extracted using a bi-phasic solvent system with methanol, methyl tert-butyl ether, and water. Untargeted proteomics, untargeted and targeted lipidomics were performed on 7 patient samples using liquid chromatography-mass spectrometry. Two comparisons were conducted: high versus low risk; relapse versus newly diagnosed. Proteins and pathways enriched in the relapsed group was compared to a public transcriptomic dataset from Multiple Myeloma Research Consortium reference collection (n = 222) at gene and pathways level. RESULTS:From one million purified plasma cells, we were able to extract material and complete untargeted (~6000 and ~3600 features in positive and negative mode respectively) and targeted lipidomics (313 lipids), as well as untargeted proteomics analysis (~4100 reviewed proteins). Comparative analyses revealed limited differences between high and low risk groups (according to the standard clinical criteria), hence we focused on drawing comparisons between the relapsed and newly diagnosed patients. Untargeted and targeted lipidomics indicated significant down-regulation of phosphatidylcholines (PCs) in relapsed MM. Although there was limited overlap of the differential proteins/transcripts, 76 significantly enriched pathways in relapsed MM were common between proteomics and transcriptomics data. Further evaluation of transcriptomics data for lipid metabolism network revealed enriched correlation of PC, ceramide, cardiolipin, arachidonic acid and cholesterol metabolism pathways to be exclusively correlated among relapsed but not in newly-diagnosed patients. CONCLUSIONS:This study establishes the feasibility and workflow to conduct integrated lipidomics and proteomics analyses on patient-derived plasma cells. Potential lipid metabolism changes associated with MM relapse warrant further investigation.
10.1371/journal.pone.0227455
Serum lipidomics for exploring biomarkers of bortezomib therapy in patients with multiple myeloma.
Maekawa Keiko,Ri Masaki,Nakajima Miki,Sekine Akihiro,Ueda Ryuzo,Tohkin Masahiro,Miyata Naoki,Saito Yoshiro,Iida Shinsuke
Cancer science
Although the proteasome inhibitor bortezomib (BTZ) shows excellent efficacy in multiple myeloma (MM), a fraction of patients has a suboptimal or no response to this agent. In addition, BTZ-induced peripheral neuropathy (BiPN), a frequent side-effect of this therapy, limits its use in some patients. This study aimed to explore serum lipid biomarker candidates to predict the response to BTZ and the severity of BiPN. Fifty-nine serum samples were collected from patients with MM prior to receiving BTZ plus low-dose dexamethasone therapy. Serum levels of phospholipids, sphingolipids, neutral lipids, and polyunsaturated fatty acids and their oxidation products were measured by a comprehensive lipidomic study. Overall, 385 lipid metabolites were identified in patients' sera; lower levels of several glycerophospholipids, sphingolipids, and cholesteryl esters were associated with a poor treatment response. Metabolites related to platelet-activating factor biosynthesis and cholesterol metabolism appeared particularly relevant. Furthermore, several lysophosphatidylcholines, phosphatidylcholines, ceramides, neutral lipids, and oxidative fatty acids were significantly increased or decreased in patients with BiPN grades ranging from G0 to G3. Among these compounds, mediators reportedly inducing myelin breakdown and stimulating inflammatory responses were prominent. Although further study is necessary to validate these biomarker candidates, our results contribute to the development of predictive biomarkers for response to BTZ treatment, or ensuing severe BiPN, in patients with MM.
10.1111/cas.14178
Cholesterol levels in patients with multiple myeloma.
Yavasoglu Irfan,Tombuloglu Murat,Kadikoylu Gurhan,Donmez Ayhan,Cagirgan Seckin,Bolaman Zahit
Annals of hematology
Hypocholesterolemia is seen in solid tumors and some hematological malignancies. We assessed cholesterol levels and the relationship between these levels and types and stages of multiple myeloma (MM) in the patients with MM. One-hundred two patients (60 male and 42 female) of mean age 59 +/- 11 years with MM were enrolled to this study. While 71.6% of the patients were Ig G type, 80.4% of the patients were at stage III. In the control group, there were 71 healthy persons (42 male and 29 female) of mean age 58 +/- 8 years. The levels of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) in the patients with MM were significantly lower than the controls (p < 0.001). There was no difference for the levels of very-low-density lipoprotein cholesterol and triglyceride between the two groups (p > 0.05). Lipid parameters were not different between Ig types (p > 0.05). The levels of TC and LDL-C in the patients with stage I were higher than those of stages II and III (p < 0.001 and p < 0.005, respectively). The levels of TC and LDL-C in the controls were not higher than the patients with stage I (p > 0.05). HDL-C levels in the patients with stage III were lower than controls (p < 0.001). Hypocholesterolemia are seen in the patients with MM. Hypocholesterolemia may be due to increased LDL clearance and utilization of cholesterol by myeloma cells.
10.1007/s00277-007-0375-6
[The clinic and pathologic significance of plasma cell myeloma with CCND1].
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi
OBJECTIVE:To study the clinical and pathologic features of multiple myeloma(MM) with CCND1. METHODS:Retrospectively analyzed the clinical and pathologic profiles of 158 patients with MM from 2010 to 2013. The clinical and morphologic features of bone marrow aspiration, biopsy and immunophenotypic analysis which was carried out by flow cytometry and immunohistochemistry were analyzed in all patients with MM respectively. CCND1 translocation was studied by FISH method in all cases. Classical cytogenetic studies of bone marrow were performed in 24 cases whose CCND1 was positive. RESULTS:In the 158 patients with MM, CCND1 was detected in 31 patients (19.6%）. In 31 patients, type IgA, IgD, IgG, IgM, light-chain only and nonsecretory MM were 4 cases,4 cases,11 cases,1 case, 6 cases and 5 cases respectively. A high incidence of CCND1 was observed in IgD and nonsecretory MM comparied with IgA and IgG respectively (P<0.05). but no statistical significance was reached between κ and λ type patients (P=0.627). The morphology of plasma cell in bone marrow biopsies were small Lymphocyte- Like 24 cases,mature plasma cell 6 cases and immature plasma cell 1 case. Immunophenotype of all 31 cases was CD38⁺CD138⁺CD19⁻CD45⁻, (CD56⁺ in 11 cases, CD20⁺ in 9 cases, CD117⁺ in 3 cases. MM with CCND1 showed a strong association with CD20 expression, the lack of CD56 expression. Immunohistochemistry showed positive for cyclinD1 in 22 cases. CONCLUSION:A high incidence of CCND1 was detected in the IgD and nonsecretory MM, and correlated with Small Lymphocyte- Like, higher positive rate of CD20, cyclinD1 and the lack of CD56 expression. MM with CCND1 must be distinguished from LPL and other mature B cell lymphomas which have plasmacytoid differentiation.
10.3760/cma.j.issn.0253-2727.2015.09.012
Global real-time quantitative reverse transcription-polymerase chain reaction detecting proto-oncogenes associated with 14q32 chromosomal translocation as a valuable marker for predicting survival in multiple myeloma.
Inagaki Atsushi,Tajima Emi,Uranishi Miyuki,Totani Haruhito,Asao Yu,Ogura Hiroka,Masaki Ayako,Yoshida Tatsuya,Mori Fumiko,Ito Asahi,Yano Hiroki,Ri Masaki,Kayukawa Satoshi,Kataoka Takae,Kusumoto Shigeru,Ishida Takashi,Hayami Yoshihito,Hanamura Ichiro,Komatsu Hirokazu,Inagaki Hiroshi,Matsuda Yasufumi,Ueda Ryuzo,Iida Shinsuke
Leukemia research
CCND1, FGFR3 and c-MAF mRNA expression of tumor samples from 123 multiple myeloma patients were analyzed by global RQ/RT-PCR. CCND1, FGFR3 and c-MAF were positive in 44 (36%), 28 (23%) and 16 (13%) of patients, respectively. In 7 patients, both FGFR3 and c-MAF were positive. The expression of c-MAF was independent unfavorable prognostic factors for overall survival (OS). Autologous stem cell transplantation improved progression-free survival of CCND1-positive patients. Bortezomib, thalidomide or lenalidomide extended OS of FGFR3 and/or c-MAF-positive patients. Thus, CCND1, FGFR3 and c-MAF mRNA expression can predict survival and is useful for planning stratified treatment strategies for myeloma patients.
10.1016/j.leukres.2013.09.026
Future of Personalized Therapy Targeting Aberrant Signaling Pathways in Multiple Myeloma.
Anwer Faiz,Gee Kevin Mathew,Iftikhar Ahmad,Baig Mirza,Russ Atlantis Dawn,Saeed Sabina,Zar Muhammad Abu,Razzaq Faryal,Carew Jennifer,Nawrocki Steffan,Al-Kateb Hussam,Cavalcante Parr Nadia Nunes,McBride Ali,Valent Jason,Samaras Christy
Clinical lymphoma, myeloma & leukemia
Multiple myeloma (MM) is a genetically complex disease. Identification of mutations and aberrant signaling pathways that contribute to the progression of MM and drug resistance has potential to lead to specific targets and personalized treatment. Aberrant signal pathways include RAS pathway activation due to RAS or BRAF mutations (targeted by vemurafenib alone or combined with cobimetinib), BCL-2 overexpression in t(11:14) (targeted by venetoclax), JAK2 pathway activation (targeted by ruxolitinib), NF-κB pathway activation (treated with DANFIN combined with bortezomib), MDM2 overexpression, and PI3K/mTOR pathway activation (targeted by BEZ235). Cyclin D1 (CCND1) and MYC are also emerging as key potential targets. In addition, histone deacetylase inhibitors are already in use for the treatment of MM in combination therapy, and targeted inhibition of FGFR3 (AZD4547) is effective in myeloma cells with t(4;14) translocation. Bromodomain and extra terminal (BET) protein antagonists decrease the expression of MYC and have displayed promising antimyeloma activity. A better understanding of the alterations in signaling pathways that promote MM progression will further inform the development of precision therapy for patients.
10.1016/j.clml.2019.03.017
Immunohistochemistry for identification of CCND1, NSD2, and MAF gene rearrangements in plasma cell myeloma.
Murase Takayuki,Ri Masaki,Narita Tomoko,Fujii Keiichiro,Masaki Ayako,Iida Shinsuke,Inagaki Hiroshi
Cancer science
The t(11;14)/CCND1-IGH, t(4;14)/NSD2(MMSET)-IGH, and t(14;16)/IGH-MAF gene rearrangements detected by fluorescence in situ hybridization (FISH) are used for risk stratification in patients with multiple myeloma (MM). Compared with conventional FISH techniques using fresh cells, immunohistochemistry (IHC) is much more cost- and time-efficient, and can be readily applied to routinely prepared formalin-fixed, paraffin-embedded (FFPE) materials. In this study, we performed tissue FISH and IHC employing FFPE specimens, and examined the usefulness of IHC as a tool for detecting CCND1, NSD2, and MAF gene rearrangements. CD138 signals were used to identify plasma cells in tissue FISH and IHC analyses. With cohort 1 (n = 70), we performed tissue FISH and subsequently IHC, and determined IHC cut-off points. In this cohort, the sensitivity and specificity for the 3 molecules were ≥.90 and ≥.96, respectively. With cohort 2, using MM cases with an unknown gene status (n = 120), we performed IHC, and the gene status was estimated using the cut-off points determined with cohort 1. The subsequent FISH analysis showed that the sensitivity and specificity for the 3 molecules were ≥.92 and ≥.98, respectively. CCND1, NSD2, and MAF gene rearrangements were estimated accurately by IHC, suggesting that conventional FISH assays can be replaced by IHC.
10.1111/cas.14109
Genomic analysis of multiple myeloma using targeted capture sequencing in the Japanese cohort.
Kanamori Takashi,Sanada Masashi,Ri Masaki,Ueno Hiroo,Nishijima Dai,Yasuda Takahiko,Tachita Takuto,Narita Tomoko,Kusumoto Shigeru,Inagaki Atsushi,Ishihara Rei,Murakami Yuki,Kobayashi Nobuhiko,Shiozawa Yusuke,Yoshida Kenichi,Nakagawa Masahiro M,Nannya Yasuhito,Shiraishi Yuichi,Chiba Kenichi,Tanaka Hiroko,Miyano Satoru,Horibe Keizo,Handa Hiroshi,Ogawa Seishi,Iida Shinsuke
British journal of haematology
Previous genomic studies have revealed the genomic landscape of myeloma cells. Although some of the genomic abnormalities shown are believed to be correlated to the molecular pathogenesis of multiple myeloma and/or clinical outcome, these correlations are not fully understood. The aim of this study is to elucidate the correlation between genomic abnormalities and clinical characteristics by targeted capture sequencing in the Japanese multiple myeloma cohort. We analysed 154 patients with newly diagnosed multiple myeloma. The analysis revealed that the study cohort consisted of a less frequent hyperdiploid subtype (37·0%) with relatively high frequencies of KRAS mutation (36·4%) and IGH-CCND1 translocation (26·6%) compared with previous reports. Moreover, our targeted capture sequencing strategy was able to detect rare IGH-associated chromosomal translocations, such as IGH-CCND2 and IGH-MAFA. Interestingly, all 10 patients harboured MAX mutations accompanied by 14q23 deletion. The patients with del(17p) exhibited an unfavourable clinical outcome, and the presence of KRAS mutation was associated with shorter survival in patients with multiple myeloma, harbouring IGH-CCND1. Thus, our study provides a detailed landscape of genomic abnormalities, which may have potential clinical application for patients with multiple myeloma.
10.1111/bjh.16720
Post-transcriptional Modifications Contribute to the Upregulation of Cyclin D2 in Multiple Myeloma.
Misiewicz-Krzeminska Irena,Sarasquete María E,Vicente-Dueñas Carolina,Krzeminski Patryk,Wiktorska Katarzyna,Corchete Luis Antonio,Quwaider Dalia,Rojas Elizabeta A,Corral Rocío,Martín Ana A,Escalante Fernando,Bárez Abelardo,García Juan Luis,Sánchez-García Isidro,García-Sanz Ramón,San Miguel Jesús F,Gutiérrez Norma C
Clinical cancer research : an official journal of the American Association for Cancer Research
PURPOSE:Dysregulation of one of the three D-cyclin genes has been observed in virtually all multiple myeloma tumors. The mechanisms by which CCND2 is upregulated in a set of multiple myeloma are not completely deciphered. We investigated the role of post-transcriptional regulation through the interaction between miRNAs and their binding sites at 3'UTR in CCND2 overexpression in multiple myeloma. EXPERIMENTAL DESIGN:Eleven myeloma cell lines and 45 primary myeloma samples were included in the study. Interactions between miRNAs deregulated in multiple myeloma and mRNA targets were analyzed by 3'UTR-luciferase plasmid assay. The presence of CCND2 mRNA isoforms different in length was explored using qRT-PCR, Northern blot, mRNA FISH, and 3' rapid amplification of cDNA ends (RACE)-PCR. RESULTS:We detected the presence of short CCND2 mRNA, both in the multiple myeloma cell lines and primary cells. The results obtained by 3'RACE experiments revealed that changes in CCND2 3'UTR length are explained by alternative polyadenylation. The luciferase assays using plasmids harboring the truncated CCND2 mRNA strongly confirmed the loss of miRNA sites in the shorter CCND2 mRNA isoform. Those multiple myelomas with greater abundance of the shorter 3'UTR isoform were associated with significant higher level of total CCND2 mRNA expression. Furthermore, functional analysis showed significant CCND2 mRNA shortening after CCND1 silencing and an increased relative expression of longer isoform after CCND1 and CCND3 overexpression, suggesting that cyclin D1 and D3 could regulate CCND2 levels through modifications in polyadenylation-cleavage reaction. CONCLUSIONS:Overall, these results highlight the impact of CCND2 3'UTR shortening on miRNA-dependent regulation of CCND2 in multiple myeloma.
10.1158/1078-0432.CCR-14-2796