Comprehensive small RNA-sequencing of primary myeloma cells identifies miR-105-5p as a predictor of patient survival.

BACKGROUND: Small RNAs (sRNAs), a heterogenous group of non-coding RNAs, are emerging as promising molecules for cancer patient risk stratification and as players in tumour pathogenesis. Here, we have studied microRNAs (miRNAs) and other sRNAs in relation to survival and disease severity in multiple myeloma. METHODS: We comprehensively characterised sRNA expression in multiple myeloma patients by performing sRNA-sequencing on myeloma cells isolated from bone marrow aspirates of 86 myeloma patients. The sRNA expression profiles were correlated with the patients' clinical data to investigate associations with survival and disease subgroups, by using cox proportional hazards (coxph) -models and limma-voom, respectively. A publicly available sRNA dataset was used as external validation (n = 151). RESULTS: We show that multiple miRNAs are differentially expressed between ISS Stage I and III. Interestingly, we observed the downregulation of seven different U2 spliceosomal RNAs, a type of small nuclear RNAs in severe disease stages. Further, by a discovery-based approach, we identified miRNA miR-105-5p as a predictor of poor overall survival (OS) in multiple myeloma. Multivariate analysis showed that miR-105-5p predict OS independently of established disease markers. CONCLUSIONS: Overexpression of miR-105-5p in myeloma cells correlates with reduced OS, potentially improving prognostic risk stratification in multiple myeloma.

Monoclonal immunoglobulins promote bone loss in multiple myeloma.

Most patients with multiple myeloma develop a severe osteolytic bone disease. The myeloma cells secrete immunoglobulins, and the presence of monoclonal immunoglobulins in the patient's sera is an important diagnostic criterion. Here, we show that immunoglobulins isolated from myeloma patients with bone disease promote osteoclast differentiation when added to human preosteoclasts in vitro, whereas immunoglobulins from patients without bone disease do not. This effect was primarily mediated by immune complexes or aggregates. The function and aggregation behavior of immunoglobulins are partly determined by differential glycosylation of the immunoglobulin-Fc part. Glycosylation analyses revealed that patients with bone disease had significantly less galactose on immunoglobulin G (IgG) compared with patients without bone disease and also less sialic acid on IgG compared with healthy persons. Importantly, we also observed a significant reduction of IgG sialylation in serum of patients upon onset of bone disease. In the 5TGM1 mouse myeloma model, we found decreased numbers of lesions and decreased CTX-1 levels, a marker for osteoclast activity, in mice treated with a sialic acid precursor, N-acetylmannosamine (ManNAc). ManNAc treatment increased IgG-Fc sialylation in the mice. Our data support that deglycosylated immunoglobulins promote bone loss in multiple myeloma and that altering IgG glycosylation may be a therapeutic strategy to reduce bone loss.

Revealing the influence of electron beam melted Ti-6Al-4V scaffolds on osteogenesis of human bone marrow-derived mesenchymal stromal cells.

Porous Titanium-6Aluminum-4Vanadium scaffolds made by electron beam-based additive manufacturing (AM) have emerged as state-of-the-art implant devices. However, there is still limited knowledge on how they influence the osteogenic differentiation of bone marrow-derived mesenchymal stromal cells (BMSCs). In this study, BMSCs are cultured on such porous scaffolds to determine how the scaffolds influence the osteogenic differentiation of the cells. The scaffolds are biocompatible, as revealed by the increasing cell viability. Cells are evenly distributed on the scaffolds after 3 days of culturing followed by an increase in bone matrix development after 21 days of culturing. qPCR analysis provides insight into the cells' osteogenic differentiation, where RUNX2 expression indicate the onset of differentiation towards osteoblasts. The COL1A1 expression suggests that the differentiated osteoblasts can produce the osteoid. Alkaline phosphatase staining indicates an onset of mineralization at day 7 in OM. The even deposits of calcium at day 21 further supports a successful bone mineralization. This work shines light on the interplay between AM Ti64 scaffolds and bone growth, which may ultimately lead to a new way of creating long lasting bone implants with fast recovery times.

Monitoring multiple myeloma by quantification of recurrent mutations in serum.

Circulating tumor DNA is a promising biomarker to monitor tumor load and genome alterations. We explored the presence of circulating tumor DNA in multiple myeloma patients and its relation to disease activity during long-term follow-up. We used digital droplet polymerase chain reaction analysis to monitor recurrent mutations, mainly in mitogen activated protein kinase pathway genes NRAS, KRAS and BRAF Mutations were identified by next-generation sequencing or polymerase chain reaction analysis of bone marrow plasma cells, and their presence analyzed in 251 archived serum samples obtained from 20 patients during a period of up to 7 years. In 17 of 18 patients, mutations identified in bone marrow during active disease were also found in a time-matched serum sample. The concentration of mutated alleles in serum correlated with the fraction in bone marrow plasma cells (r=0.507, n=34, P<0.002). There was a striking covariation between circulating mutation levels and M protein in ten out of 11 patients with sequential samples. When relapse evaluation by circulating tumor DNA and M protein could be directly compared, the circulating tumor DNA showed relapse earlier in two patients (3 and 9 months), later in one patient (4 months) and in three patients there was no difference. In three patients with transformation to aggressive disease, the concentrations of mutations in serum increased up to 400 times, an increase that was not seen for the M protein. In conclusion, circulating tumor DNA in myeloma is a multi-faceted biomarker reflecting mutated cells, total tumor mass and transformation to a more aggressive disease. Its properties are both similar and complementary to M protein.

Toll-like receptor signaling in multiple myeloma cells promotes the expression of pro-survival genes B-cell lymphoma 2 and MYC and modulates the expression of B-cell maturation antigen.

Infections are common in plasma cell cancer multiple myeloma (MM) due to disease-related immune deficiencies and cancer treatment. Myeloma cells express Toll-like receptors (TLRs), and TLR activation has been shown to induce proliferative and pro-survival signals in cancer cells. MM is a complex and heterogeneous disease, and expression levels of TLRs as well as downstream signaling components are likely to differ between patients. Here, we show that in a large cohort of patients, TLR1, TLR4, TLR6, TLR9, and TLR10 are the most highly expressed in primary CD138+ cells. Using an MM cell line expressing TLR4 and TLR9 as a model, we demonstrate that TLR4 and TLR9 activation promoted the expression of well-established pro-survival and oncogenes in MM such as MYC, IRF4, NFKB, and BCL2. TLR4 and TLR9 activation inhibited the efficacy of proteasome inhibitors bortezomib and carfilzomib, drugs used in the treatment of MM. Inhibiting the autophagosome-lysosome protein degradation pathway by hydroxychloroquine (HCQ) diminished the protective effect of TLR activation on proteasome inhibitor-induced cytotoxicity. We also found that TLR signaling downregulated the expression of TNFRSF17, the gene encoding for B-cell maturation antigen (BCMA). MYC, BCL2, and BCL2L1 were upregulated in approximately 50% of primary cells, while the response to TLR signaling in terms of TNFRSF17 expression was dichotomous, as an equal fraction of patients showed upregulation and downregulation of the gene. While proteasome inhibitors are part of first-line MM treatment, several of the new anti-MM immune therapeutic drugs target BCMA. Thus, TLR activation may render MM cells less responsive to commonly used anti-myeloma drugs.

Interpreting single-cell messages in normal and aberrant hematopoiesis with the Cell Marker Accordion.

Single-cell technologies offer a unique opportunity to explore cellular heterogeneity in hematopoiesis, reveal malignant hematopoietic cells with clinically significant features and measure gene signatures linked to pathological pathways. However, reliable identification of cell types is a crucial bottleneck in single-cell analysis. Available databases contain dissimilar nomenclature and non-concurrent marker sets, leading to inconsistent annotations and poor interpretability. Furthermore, current tools focus mostly on physiological cell types, lacking extensive applicability in disease. We developed the Cell Marker Accordion, a user-friendly platform for the automatic annotation and biological interpretation of single-cell populations based on consistency weighted markers. We validated our approach on peripheral blood and bone marrow single-cell datasets, using surface markers and expert-based annotation as the ground truth. In all cases, we significantly improved the accuracy in identifying cell types with respect to any single source database. Moreover, the Cell Marker Accordion can identify disease-critical cells and pathological processes, extracting potential biomarkers in a wide variety of contexts in human and murine single-cell datasets. It characterizes leukemia stem cell subtypes, including therapy-resistant cells in acute myeloid leukemia patients; it identifies malignant plasma cells in multiple myeloma samples; it dissects cell type alterations in splicing factor-mutant cells from myelodysplastic syndrome patients; it discovers activation of innate immunity pathways in bone marrow from mice treated with METTL3 inhibitors. The breadth of these applications elevates the Cell Marker Accordion as a flexible, faithful and standardized tool to annotate and interpret hematopoietic populations in single-cell datasets focused on the study of hematopoietic development and disease.

Diet-induced obesity reduces bone marrow T and B cells and promotes tumor progression in a transplantable Vk*MYC model of multiple myeloma.

Obesity is associated with an increased risk of developing multiple myeloma (MM). The molecular mechanisms causing this association is complex and incompletely understood. Whether obesity affects bone marrow immune cell composition in multiple myeloma is not characterized. Here, we examined the effect of diet-induced obesity on bone marrow immune cell composition and tumor growth in a Vk*MYC (Vk12653) transplant model of multiple myeloma. We find that diet-induced obesity promoted tumor growth in the bone marrow and spleen and reduced the relative number of T and B cells in the bone marrow. Our results suggest that obesity may reduce MM immune surveillance and thus may contribute to increased risk of developing MM.

Anti-c-MET Nanobody - a new potential drug in multiple myeloma treatment.

BACKGROUND: c-MET is the tyrosine kinase receptor of the hepatocyte growth factor (HGF). HGF-c-MET signaling is involved in many human malignancies, including multiple myeloma (MM). Recently, multiple agents have been developed directed to interfere at different levels in HGF-c-MET signaling pathway. Nanobodies are therapeutic proteins based on the smallest functional fragments of heavy-chain-only antibodies. In this study, we wanted to determine the anticancer effect of a novel anti-c-MET Nanobody in MM. METHODS: We examined the effects of an anti-c-MET Nanobody on thymidine incorporation, migration, adhesion of MM cells, and osteoblastogenesis in vitro. Furthermore, we investigated the effects of the Nanobody on HGF-dependent c-MET signaling by Western blotting. RESULTS: We show that the anti-c-MET Nanobody effectively inhibited thymidine incorporation of ANBL-6 MM cells via inhibition of an HGF autocrine growth loop and thymidine incorporation in INA-6 MM cells induced by exogenous HGF. HGF-induced migration and adhesion of INA-6 were completely and specifically blocked by the Nanobody. Furthermore, the Nanobody abolished the inhibiting effect of HGF on bone morphogenetic protein-2-induced alkaline phosphatase activity and the mineralization of human mesenchymal stem cells. Finally, we show that the Nanobody reduced phosphorylation of tyrosine residues in c-MET, MAPK, and Akt. We also compared the Nanobody with anti-c-MET monoclonal antibodies and revealed the similar or better effect. CONCLUSIONS: The anti-c-MET Nanobody inhibited MM cell migration, thymidine incorporation, and adhesion, and blocked the HGF-mediated inhibition of osteoblastogenesis. The anti-c-MET Nanobody might represent a novel therapeutic agent in the treatment of MM and other cancers driven by HGF-c-MET signaling.

The pro-tumorigenic cytokine IL-32 has a high turnover in multiple myeloma cells due to proteolysis regulated by oxygen-sensing cysteine dioxygenase and deubiquitinating enzymes.

IL-32 is a pro-inflammatory cytokine expressed by several types of cancer cells and immune cells. Currently, no treatment targeting IL-32 is available, and its intracellular and exosomal localization make IL-32 less accessible to drugs. We previously showed that hypoxia promotes IL-32 expression through HIF1α in multiple myeloma cells. Here, we demonstrate that high-speed translation and ubiquitin-dependent proteasomal degradation lead to a rapid IL-32 protein turnover. We find that IL-32 protein half-life is regulated by the oxygen-sensing cysteine-dioxygenase ADO and that deubiquitinases actively remove ubiquitin from IL-32 and promote protein stability. Deubiquitinase inhibitors promoted the degradation of IL-32 and may represent a strategy for reducing IL-32 levels in multiple myeloma. The fast turnover and enzymatic deubiquitination of IL-32 are conserved in primary human T cells; thus, deubiquitinase inhibitors may also affect T-cell responses in various diseases.

The effects of surface treatments on electron beam melted Ti-6Al-4V disks on osteogenesis of human mesenchymal stromal cells.

Additive manufactured (AM) Titanium-6Aluminum-4Vanadium (Ti64) scaffolds display unique mechanical and biological properties for implant devices. The elastic modulus can be tailored by adjusting the porosity, further facilitating bone ingrowth. Although Ti64 implants are biocompatible, the effects of AM surfaces without porous structures, and how the topography and surface chemistry of the respective surfaces affect the osteogenesis of bone marrow-derived mesenchymal stromal cells (BMSCs) has not yet been revealed. In this paper, we cultured BMSCs on solid electron beam melted Ti64 disks subjected to three surface treatments: chemical etching (HF), atomic-layer deposition of TiO2 (TiO2), and polished (POL), or left untreated (AB). The biocompatibility and osteogenic properties of these surfaces were investigated, and the results were compared to cells cultured in regular tissue-culture polystyrene culturing wells (TCPS). The surfaces were hydrophobic, except for the polished surface which was hydrophilic. All surface treatments are biocompatible and allow for osteogenic differentiation, as revealed by viability assays and gene expression analysis. Scanning electron microscopy shows that cells adhere differently depending on the surface properties, with more filopodia on the rougher surfaces, AB and TiO2 disks, and more lamellipodia on the smoother surfaces, HF and POL disks. All groups stimulated with beta glycerophosphate, ascorbic acid, and dexamethasone, have elevated expression of genes related to matrix formation, where the cells cultured on the disks treated with TiO2, HF and POL have the overall highest expression. The AB group appears to be less favorable in regards to matrix formation. Considering the matrix mineralization, the rougher surfaces, AB and TiO2, are able to induce matrix mineralization, with an elevated gene expression of vitamin D receptors and calcium deposition of unstimulated cells. Finally, imaging at day 21 revealed an even amount of cells and matrix, covering most of the partially melted particles. Our results suggests that surface topography is more important to osteogenesis than the wettability of the surface. Overall, the present study contributes to the understanding of using surface modifications to AM Ti64 implant materials and reveals how they affect bone growth.

IL-32 is induced by activation of toll-like receptors in multiple myeloma cells.

Multiple myeloma (MM) is a hematological cancer characterized by accumulation of malignant plasma cells in the bone marrow. The patients are immune suppressed and suffer from recurrent and chronic infections. Interleukin-32 is a non-conventional, pro-inflammatory cytokine expressed in a subgroup of MM patients with a poor prognosis. IL-32 has also been shown to promote proliferation and survival of the cancer cells. Here we show that activation of toll-like receptors (TLRs) promotes expression of IL-32 in MM cells through NFκB activation. In patient-derived primary MM cells, IL-32 expression is positively associated with expression of TLRs. Furthermore, we found that several TLR genes are upregulated from diagnosis to relapse in individual patients, predominantly TLRs sensing bacterial components. Interestingly, upregulation of these TLRs coincides with an increase in IL-32. Taken together, these results support a role for IL-32 in microbial sensing in MM cells and suggest that infections can induce expression of this pro-tumorigenic cytokine in MM patients.

Secondary hyperparathyroidism but stable bone-mineral density in patients with chronic myeloid leukemia treated with imatinib.

Imatinib is currently the standard treatment for chronic myeloid leukemia(CML). Previous studies have shown that imatinib affects bone metabolism in CML patients. However, these effects are not well-studied prospectively. The authors studied bone-mineral density (BMD) and bone metabolism in 17 CML patients and matched controls in 2007 and now repeated the analyses prospectively in 2011. All CML patients were in complete cytogenetic remission during this 4-year period and treated with 400 mg imatinib q.d. (n 5 15) or 600 mg imatinib q.d. (n 5 2). Mean treatment duration was 102 months (range 69­129) in 2011. The authors found that serum levels of parathyroid hormone increased significantly in the patients between 2007 and 2011, and seven out of 17 patients had secondary hyperparathyroidism in 2011. However, the mean areal and volumetric BMDs were stable in the CML patients over the 4-year-observation period. Moreover, the CML patients had significantly higher volumetric BMD in the cortical compartment when compared with controls in 2011 and 2007. Thus, despite a high incidence of secondary hyperparathyroidism,there were no signs of osteoporosis or osteomalacia in imatinib-treated CML patients as suggested earlier.

Imatinib inhibits proliferation of human mesenchymal stem cells and promotes early but not late osteoblast differentiation in vitro.

Altered bone metabolism has been reported in patients with chronic myeloid leukemia treated with the tyrosine kinase inhibitor imatinib. Several studies have shown that imatinib inhibits the differentiation and activity of osteoclasts in vitro, whereas the effects of imatinib on osteoblast differentiation are less clear. In this study osteoblast differentiation was induced in human mesenchymal stem cells (hMSCs) by treatment with bone morphogenetic protein 2 in vitro. Imatinib inhibited proliferation of hMSCs in a dose-dependent manner. Even though imatinib promoted early osteoblast differentiation assessed by alkaline phosphate activity, mineralization measured by Alizarin Red staining (ARS) was reduced by imatinib. Moreover, the inhibitory effect of imatinib on mineralization was most prominent at low concentrations of imatinib. When we measured the relative mRNA expression levels of Runx2, we found that Runx2 expression was higher in imatinib-treated (5 µM) cultures at early time points during differentiation. On the other hand, the expression of Osterix late during differentiation was lower in imatinib-treated (5 µM) cultures, corresponding to the ARS results. Thus, the effect of imatinib on osteoblast differentiation is not only dependent on the drug concentration, but indeed also on the maturation stage of the cells. This finding might partly explain why previous studies on the effects of imatinib osteoblast differentiation have shown different results.

Association between high plasma levels of hepatocyte growth factor and progression of radiographic damage in the joints of patients with rheumatoid arthritis.

OBJECTIVE: Hepatocyte growth factor (HGF) and dickkopf-1 (Dkk-1) inhibit osteoblast differentiation. The present study was thus undertaken to investigate whether plasma levels of HGF and Dkk-1 are related to bone damage in patients with rheumatoid arthritis (RA). METHODS: RA patients with a disease duration of <4 years were followed up prospectively with collection of demographic, clinical, and radiographic data at study enrollment (1992) and after 1, 2, 5, and 10 years. Hand radiographs were scored according to the modified Sharp/van der Heijde score (SHS). Levels of HGF and Dkk-1 in stored plasma samples obtained from 136 patients at the time of enrollment were measured by enzyme-linked immunosorbent assay. Associations between cytokine levels and radiographic progression were examined by linear regression analysis. RESULTS: Patients with above-median levels of HGF at enrollment had a significantly greater change in the SHS after 1, 2, 5, and 10 years than did patients with below-median levels of HGF (P < 0.001, P = 0.006, P = 0.01, and P = 0.01, respectively). In an unadjusted analysis, baseline HGF levels predicted the severity of joint damage at all time points studied. After adjustment for other known predictors of radiographic progression, baseline HGF levels remained significantly associated with radiographic progression. Plasma levels of Dkk-1 at enrollment were not significantly associated with radiographic progression at any time point studied. CONCLUSION: Plasma levels of HGF predict joint damage in RA, and this finding suggests that HGF plays a role in RA joint destruction. The role of HGF as a potential prognostic biomarker or target for treatment warrants further exploration.

CpG-oligodeoxynucleotide inhibits Smad-dependent bone morphogenetic protein signaling: effects on myeloma cell apoptosis and in vitro osteoblastogenesis.

The TLR9 agonist CpG-oligodeoxynucleotide (CpG-ODN) with a phosphorothioate backbone (PTO-CpG-ODN) is evaluated in clinical trials as a vaccine adjuvant or as treatment of cancers. Bone morphogenetic proteins (BMPs) regulate growth and differentiation of several cell types, and also induce apoptosis of cancer cells. Cross-talk between BMP- and TLR-signaling has been reported, and we aimed to investigate whether CpG-ODN influenced BMP-induced osteoblast differentiation or BMP-induced apoptosis of malignant plasma cells. We found that PTO-CpG-ODN inhibited BMP-2-induced osteoblast differentiation from human mesenchymal stem cells. Further, PTO-CpG-ODN counteracted BMP-2- and BMP-6-induced apoptosis of the human myeloma cell lines IH-1 and INA-6, respectively. In contrast, PTO-CpG-ODN did not antagonize the antiproliferative effect of BMP-2 on hMSCs or IH-1 cells. Inhibition of Smad-signaling and p38 MAPK-signaling indicated that apoptosis of IH-1 cells is dependent on Smad-signaling downstream of BMP, whereas the antiproliferative effect of BMP-2 on IH-1 cells also involves p38 MAPK-signaling. Together, the data suggested a specific inhibition by PTO-CpG-ODN on BMP-Smad-signaling. Supporting this we found that PTO-CpG-ODN inhibited BMP-induced phosphorylation of receptor-Smads in human mesenchymal stem cells and myeloma cell lines. This effect appeared to be independent of TLR9 because GpC-ODN and other ODNs with the ability to form multimeric structures inhibited Smad-signaling as efficiently as PTO-CpG-ODNs, and because knockdown of TLR9 by small interfering RNA in INA-6 cells did not blunt the effect of PTO-CpG-ODN. In conclusion, our results demonstrate that PTO-CpG-ODN inhibits BMP-signaling, and thus might provoke unwanted TLR9-independent side effects in patients.

Automated Quantification of Human Osteoclasts Using Object Detection.

A balanced skeletal remodeling process is paramount to staying healthy. The remodeling process can be studied by analyzing osteoclasts differentiated in vitro from mononuclear cells isolated from peripheral blood or from buffy coats. Osteoclasts are highly specialized, multinucleated cells that break down bone tissue. Identifying and correctly quantifying osteoclasts in culture are usually done by trained personnel using light microscopy, which is time-consuming and susceptible to operator biases. Using machine learning with 307 different well images from seven human PBMC donors containing a total of 94,974 marked osteoclasts, we present an efficient and reliable method to quantify human osteoclasts from microscopic images. An open-source, deep learning-based object detection framework called Darknet (YOLOv4) was used to train and test several models to analyze the applicability and generalizability of the proposed method. The trained model achieved a mean average precision of 85.26% with a correlation coefficient of 0.99 with human annotators on an independent test set and counted on average 2.1% more osteoclasts per culture than the humans. Additionally, the trained models agreed more than two independent human annotators, supporting a more reliable and less biased approach to quantifying osteoclasts while saving time and resources. We invite interested researchers to test their datasets on our models to further strengthen and validate the results.

Paired miRNA- and messenger RNA-sequencing identifies novel miRNA-mRNA interactions in multiple myeloma.

Multiple myeloma (MM) is an incurable cancer of terminally differentiated plasma cells that proliferate in the bone marrow. miRNAs are promising biomarkers for risk stratification in MM and several miRNAs are shown to have a function in disease pathogenesis. However, to date, surprisingly few miRNA-mRNA interactions have been described for and functionally validated in MM. In this study, we performed miRNA-seq and mRNA-seq on CD138 + cells isolated from bone marrow aspirates of 86 MM patients to identify novel interactions between sRNAs and mRNAs. We detected 9.8% significantly correlated miRNA-mRNA pairs of which 5.17% were positively correlated and 4.65% were negatively correlated. We found that miRNA-mRNA pairs that were predicted by in silico target-prediction algorithms were more negatively correlated than non-target pairs, indicating functional miRNA targeting and that correlation between miRNAs and mRNAs from patients can be used to identify miRNA-targets. mRNAs for negatively correlated miRNA-mRNA target pairs were associated with gene ontology terms such as autophagy, protein degradation and endoplasmic stress response, reflecting important processes in MM. Targets for two specific miRNAs, miR-125b-5p and miR-365b-3p, were functionally validated in MM cell line transfection experiments followed by RNA-sequencing and qPCR. In summary, we identified functional miRNA-mRNA target pairs by correlating miRNA and mRNA data from primary MM cells. We identified several target pairs that are of potential interest for further studies. The data presented here may serve as a hypothesis-generating knowledge base for other researchers in the miRNA/MM field. We also provide an interactive web application that can be used to exploit the miRNA-target interactions as well as clinical parameters associated to these target-pairs.

Intracellular IL-32 regulates mitochondrial metabolism, proliferation, and differentiation of malignant plasma cells.

Interleukin-32 (IL-32) is a nonclassical cytokine expressed in cancers, inflammatory diseases, and infections. Its expression is regulated by two different oxygen sensing systems; HIF1α and cysteamine dioxygenase (ADO), indicating that IL-32 may be involved in the response to hypoxia. We here demonstrate that endogenously expressed, intracellular IL-32 interacts with components of the mitochondrial respiratory chain and promotes oxidative phosphorylation. Knocking out IL-32 in three myeloma cell lines reduced cell survival and proliferation in vitro and in vivo. High-throughput transcriptomic and MS-metabolomic profiling of IL-32 KO cells revealed that cells depleted of IL-32 had perturbations in metabolic pathways, with accumulation of lipids, pyruvate precursors, and citrate. IL-32 was expressed in a subgroup of myeloma patients with inferior survival, and primary myeloma cells expressing IL-32 had a gene signature associated with immaturity, proliferation, and oxidative phosphorylation. In conclusion, we demonstrate a previously unrecognized role of IL-32 in the regulation of plasma cell metabolism.

Molecular interactions and functions of IL-32.

IL-32 is a multifaceted cytokine associated with several diseases and inflammatory conditions. Its expression is induced in response to cellular stress such as hypoxia, infections, and pro-inflammatory cytokines. IL-32 can be secreted from cells and can induce the production of pro-inflammatory cytokines from several cell types but are also described to have anti-inflammatory functions. The intracellular form of IL-32 is shown to play an important role in various cellular processes, including the defense against intracellular bacteria and viruses and in modulation of cell metabolism. In this review, we discuss current literature on molecular interactions of IL-32 with other proteins. We also review data on the role of intracellular IL-32 as a metabolic regulator and its role in antimicrobial host defense.

Bystander Memory T Cells and IMiD/Checkpoint Therapy in Multiple Myeloma: A Dangerous Tango?

In this review article we discuss the role of the memory T cells in multiple myeloma (MM) and how they may influence immune responses in patients that received immunomodulating drugs and check point therapy.