The serine protease matriptase inhibits migration and proliferation in multiple myeloma cells.

BACKGROUND: Multiple myeloma (MM) is an incurable malignancy of plasma cells. The serine protease matriptase is frequently dysregulated in human carcinomas, which facilitates tumor progression and metastatic dissemination. The importance of matriptase in hematological malignancies is yet to be clarified. In this study, we aimed to characterize the role of matriptase in MM. MATERIALS AND METHODS: mRNA expression of matriptase and its inhibitors hepatocyte growth factor activator inhibitor (HAI)-1 and HAI-2 was studied in primary MM cells from patient samples and human myeloma cell lines (HMCLs). We further investigated the effect of matriptase on migration and proliferation of myeloma cells in vitro. By use of the CoMMpass database, we assessed the clinical relevance of matriptase in MM patients. RESULTS: Matriptase was expressed in 96% of patient samples and all HMCLs tested. Overexpression of matriptase in vitro reduced proliferation, and significantly decreased cytokine-induced migration. Conversely, matriptase knockdown significantly enhanced migration. Mechanistically, overexpression of matriptase inhibited activation of Src kinase. CONCLUSIONS: Our findings may suggest a novel role of matriptase as a tumor suppressor in MM pathogenesis.

Highly expressed genes in multiple myeloma cells - what can they tell us about the disease?

Cancer cells can convert proto-oncoproteins into oncoproteins by increasing the expression of genes that are oncogenic when expressed at high levels. Such genes can promote oncogenesis without being mutated. To find overexpressed genes in cancer cells from patients with multiple myeloma, we retrieved mRNA expression data from the CoMMpass database and ranked genes by their expression levels. We grouped the most highly expressed genes based on a set of criteria and we discuss the role a selection of them can play in the disease pathophysiology. The list was highly concordant with a similar list based on mRNA expression data from the PADIMAC study. Many well-known "myeloma genes" such as MCL1, CXCR4, TNFRSF17, SDC1, SLAMF7, PTP4A3, and XBP1 were identified as highly expressed, and we believe that hitherto unrecognized key players in myeloma pathogenesis are also enriched on the list. Highly expressed genes in malignant plasma cells that were absent or expressed at only a low level in healthy plasma cells included IFI6, IFITM1, PTP4A3, SIK1, ALDOA, ATP5MF, ATP5ME, and PSMB4. The ambition of this article is not to validate the role of each gene but to serve as a guide for studies aiming at identifying promising treatment targets.

Inhibition of Cytosolic Phospholipase A2α Induces Apoptosis in Multiple Myeloma Cells.

Cytosolic phospholipase A2α (cPLA2α) is the rate-limiting enzyme in releasing arachidonic acid and biosynthesis of its derivative eicosanoids. Thus, the catalytic activity of cPLA2α plays an important role in cellular metabolism in healthy as well as cancer cells. There is mounting evidence suggesting that cPLA2α is an interesting target for cancer treatment; however, it is unclear which cancers are most relevant for further investigation. Here we report the relative expression of cPLA2α in a variety of cancers and cancer cell lines using publicly available datasets. The profiling of a panel of cancer cell lines representing different tissue origins suggests that hematological malignancies are particularly sensitive to the growth inhibitory effect of cPLA2α inhibition. Several hematological cancers and cancer cell lines overexpressed cPLA2α, including multiple myeloma. Multiple myeloma is an incurable hematological cancer of plasma cells in the bone marrow with an emerging requirement of therapeutic approaches. We show here that two cPLA2α inhibitors AVX420 and AVX002, significantly and dose-dependently reduced the viability of multiple myeloma cells and induced apoptosis in vitro. Our findings implicate cPLA2α activity in the survival of multiple myeloma cells and support further studies into cPLA2α as a potential target for treating hematological cancers, including multiple myeloma.

PRL-3 induces a positive signaling circuit between glycolysis and activation of STAT1/2.

Multiple myeloma (MM) is an incurable hematologic malignancy resulting from the clonal expansion of plasma cells. MM cells are interacting with components of the bone marrow microenvironment such as cytokines to survive and proliferate. Phosphatase of regenerating liver (PRL)-3, a cytokine-induced oncogenic phosphatase, is highly expressed in myeloma patients and is a mediator of metabolic reprogramming of cancer cells. To find novel pathways and genes regulated by PRL-3, we characterized the global transcriptional response to PRL-3 overexpression in two MM cell lines. We used pathway enrichment analysis to identify pathways regulated by PRL-3. We further confirmed the hits from the enrichment analysis with in vitro experiments and investigated their function. We found that PRL-3 induced expression of genes belonging to the type 1 interferon (IFN-I) signaling pathway due to activation of signal transducer and activator of transcription (STAT) 1 and STAT2. This activation was independent of autocrine IFN-I secretion. The increase in STAT1 and STAT2 did not result in any of the common consequences of increased IFN-I or STAT1 signaling in cancer. Knockdown of STAT1/2 did not affect the viability of the cells, but decreased PRL-3-induced glycolysis. Interestingly, glucose metabolism contributed to the activation of STAT1 and STAT2 and expression of IFN-I-stimulated genes in PRL-3-overexpressing cells. In summary, we describe a novel signaling circuit where the key IFN-I-activated transcription factors STAT1 and STAT2 are important drivers of the increase in glycolysis induced by PRL-3. Subsequently, increased glycolysis regulates the IFN-I-stimulated genes by augmenting the activation of STAT1/2.

Phosphatase of regenerating liver-3 regulates cancer cell metabolism in multiple myeloma.

Cancer cells often depend on microenvironment signals from molecules such as cytokines for proliferation and metabolic adaptations. PRL-3, a cytokine-induced oncogenic phosphatase, is highly expressed in multiple myeloma cells and associated with poor outcome in this cancer. We studied whether PRL-3 influences metabolism. Cells transduced to express PRL-3 had higher aerobic glycolytic rate, oxidative phosphorylation, and ATP production than the control cells. PRL-3 promoted glucose uptake and lactate excretion, enhanced the levels of proteins regulating glycolysis and enzymes in the serine/glycine synthesis pathway, a side branch of glycolysis. Moreover, mRNAs for these proteins correlated with PRL-3 expression in primary patient myeloma cells. Glycine decarboxylase (GLDC) was the most significantly induced metabolism gene. Forced GLDC downregulation partly counteracted PRL-3-induced aerobic glycolysis, indicating GLDC involvement in a PRL-3-driven Warburg effect. AMPK, HIF-1α, and c-Myc, important metabolic regulators in cancer cells, were not mediators of PRL-3's metabolic effects. A phosphatase-dead PRL-3 mutant, C104S, promoted many of the metabolic changes induced by wild-type PRL-3, arguing that important metabolic effects of PRL-3 are independent of its phosphatase activity. Through this study, PRL-3 emerges as one of the key mediators of metabolic adaptations in multiple myeloma.

Targeting phosphoglycerate dehydrogenase in multiple myeloma.

BACKGROUND: Multiple myeloma (MM) is a hematological malignancy characterized by the clonal expansion of plasma cells in the bone marrow. To date, this disease is still incurable and novel therapeutic approaches are required. Phosphoglycerate dehydrogenase (PHGDH) is the first and rate-limiting enzyme in the de novo serine synthesis pathway, and it has been attributed to bortezomib-resistance in MM. METHODS: Two different PHGDH inhibitors, CBR5884 and NCT-503, were tested against human myeloma cell lines, primary MM cells from patients, and peripheral blood mononuclear cells isolated from healthy donors. The PHGDH inhibitors were then tested in combination with proteasome inhibitors in different MM cell lines, including proteasome-resistant cell lines. Furthermore, we confirmed the effects of PHGDH inhibition through knocking down PHGDH and the effect of NCT-503 in vivo in the 5T33MM mouse model. RESULTS: All the tested myeloma cell lines expressed PHGDH and were sensitive to doses of NCT-503 that were tolerated by peripheral blood mononuclear cells isolated from healthy donors. Upon testing bortezomib in combination with NCT-503, we noticed a clear synergy in several HMCLs. The sensitivity to bortezomib also increased after PHGDH knockdown, mimicking the effect of NCT-503 treatment. Interestingly, targeting PHGDH reduced the intracellular redox capacity of the cells. Furthermore, combination treatment with NCT-503 and bortezomib exhibited a therapeutic advantage in vivo. CONCLUSIONS: Our study shows the therapeutic potential of targeting PHGDH in MM, and suggest it as a way to overcome the resistance to proteasome inhibitors.

Conversion of ATP to adenosine by CD39 and CD73 in multiple myeloma can be successfully targeted together with adenosine receptor A2A blockade.

BACKGROUND: PD1/PDL1-directed therapies have been unsuccessful for multiple myeloma (MM), an incurable cancer of plasma cells in the bone marrow (BM). Therefore, other immune checkpoints such as extracellular adenosine and its immunosuppressive receptor should be considered. CD39 and CD73 convert extracellular ATP to adenosine, which inhibits T-cell effector functions via the adenosine receptor A2A (A2AR). We set out to investigate whether blocking the adenosine pathway could be a therapy for MM. METHODS: Expression of CD39 and CD73 on BM cells from patients and T-cell proliferation were determined by flow cytometry and adenosine production by Liquid chromatograpy-mass spectrometry (HPCL/MS). ENTPD1 (CD39) mRNA expression was determined on myeloma cells from patients enrolled in the publicly available CoMMpass study. Transplantable 5T33MM myeloma cells were used to determine the effect of inhibiting CD39, CD73 and A2AR in mice in vivo. RESULTS: Elevated level of adenosine was found in BM plasma of MM patients. Myeloma cells from patients expressed CD39, and high gene expression indicated reduced survival. CD73 was found on leukocytes and stromal cells in the BM. A CD39 inhibitor, POM-1, and an anti-CD73 antibody inhibited adenosine production and reduced T-cell suppression in vitro in coculture of myeloma and stromal cells. Blocking the adenosine pathway in vivo with a combination of Sodium polyoxotungstate (POM-1), anti-CD73, and the A2AR antagonist AZD4635 activated immune cells, increased interferon gamma production, and reduced the tumor load in a murine model of MM. CONCLUSIONS: Our data suggest that the adenosine pathway can be successfully targeted in MM and blocking this pathway could be an alternative to PD1/PDL1 inhibition for MM and other hematological cancers. Inhibitors of the adenosine pathway are available. Some are in clinical trials and they could thus reach MM patients fairly rapidly.

Activins as Dual Specificity TGF-ß Family Molecules: SMAD-Activation via Activin- and BMP-Type 1 Receptors.

Activins belong to the transforming growth factor (TGF)-ß family of multifunctional cytokines and signal via the activin receptors ALK4 or ALK7 to activate the SMAD2/3 pathway. In some cases, activins also signal via the bone morphogenetic protein (BMP) receptor ALK2, causing activation of the SMAD1/5/8 pathway. In this study, we aimed to dissect how activin A and activin B homodimers, and activin AB and AC heterodimers activate the two main SMAD branches. We compared the activin-induced signaling dynamics of ALK4/7-SMAD2/3 and ALK2-SMAD1/5 in a multiple myeloma cell line. Signaling via the ALK2-SMAD1/5 pathway exhibited greater differences between ligands than signaling via ALK4/ALK7-SMAD2/3. Interestingly, activin B and activin AB very potently activated SMAD1/5, resembling the activation commonly seen with BMPs. As SMAD1/5 was also activated by activins in other cell types, we propose that dual specificity is a general mechanism for activin ligands. In addition, we found that the antagonist follistatin inhibited signaling by all the tested activins, whereas the antagonist cerberus specifically inhibited activin B. Taken together, we propose that activins may be considered dual specificity TGF-ß family members, critically affecting how activins may be considered and targeted clinically.

BMPR2 inhibits activin and BMP signaling via wild-type ALK2.

TGF-ß/BMP superfamily ligands require heteromeric complexes of type 1 and 2 receptors for ligand-dependent downstream signaling. Activin A, a TGF-ß superfamily member, inhibits growth of multiple myeloma cells, but the mechanism for this is unknown. We therefore aimed to clarify how activins affect myeloma cell survival. Activin A activates the transcription factors SMAD2/3 through the ALK4 type 1 receptor, but may also activate SMAD1/5/8 through mutated variants of the type 1 receptor ALK2 (also known as ACVR1). We demonstrate that activin A and B activate SMAD1/5/8 in myeloma cells through endogenous wild-type ALK2. Knockdown of the type 2 receptor BMPR2 strongly potentiated activin A- and activin B-induced activation of SMAD1/5/8 and subsequent cell death. Furthermore, activity of BMP6, BMP7 or BMP9, which may also signal via ALK2, was potentiated by knockdown of BMPR2. Similar results were seen in HepG2 liver carcinoma cells. We propose that BMPR2 inhibits ALK2-mediated signaling by preventing ALK2 from oligomerizing with the type 2 receptors ACVR2A and ACVR2B, which are necessary for activation of ALK2 by activins and several BMPs. In conclusion, BMPR2 could be explored as a possible target for therapy in patients with multiple myeloma.This article has an associated First Person interview with the first author of the paper.