Regulation Of Cellular Bioenergetics Research Articles

Regulation of CyR61 expression and release by 3-mercaptopyruvate sulfurtransferase in colon cancer cells

Cysteine-rich angiogenic inducer 61 (CYR61, also termed CCN family member 1 or CCN1), is a matricellular protein encoded by the CYR61 gene. This protein has been implicated in the regulation of various cancer-associated processes including tumor growth, angiogenesis, tumor cell adhesion, migration, and invasion as well as the regulation of anticancer drug resistance. Hydrogen sulfide (H2S) is a gaseous endogenous biological mediator, involved in the regulation of cellular bioenergetics, angiogenesis, invasion, and chemotherapeutic resistance in several types of cancer. H2S is produced by three enzymes: cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3-MST). The current studies were set up to investigate if CBS or 3-MST regulates CyR61 in colon cancer cells in the context of the regulation of proliferation, migration, and survival. The study mainly utilized HCT116 cells, in which two of the principal H2S-producing enzymes, CBS and 3-MST, are highly expressed. The H2S donor GYY4137 and the polysulfide donor Na2S3 activated the CyR61 promoter in a concentration-dependent fashion. Aminooxyacetic acid (AOAA), a pharmacological inhibitor of CBS as well as HMPSNE: 2-[(4-hydroxy-6- methylpyrimidin-2-yl)sulfanyl]-1-(naphthalen-1-yl)ethan-1-one, a pharmacological inhibitor of 3-MST inhibited CyR61 mRNA expression. This effect was more pronounced in response to HMPSNE than to AOAA and occurred through the modulation of S1PR via ATF1 and CREB. CyR61 was found to play an active, but relatively minor role in maintaining colon cell proliferation. HMPSNE markedly suppressed the secretion/release of CyR61 from the colon cancer cells. Moreover, HMPSNE promoted colon cancer cell apoptosis; endogenously produced CyR61 was found to counteract this effect, at least in part via RhoA activation. Taken together, we conclude that the upregulation of 3-MST in cancer cells exerts cytoprotective effects and confers the cancer cells a more aggressive phenotype – at least in part via the modulation of CyR61 expression and release.

Physiological roles of hydrogen sulfide in mammalian cells, tissues, and organs.

Over the last two decades, hydrogen sulfide (H2S) has emerged as an endogenous regulator of a broad range of physiological functions. H2S belongs to the class of molecules known as gasotransmitters, which typically include nitric oxide (NO) and carbon monoxide (CO). Three enzymes are recognized as endogenous sources of H2S in various cells and tissues: cystathionine γ-lyase (CSE), cystathionine β-synthase (CBS), and 3-mercaptopyruvate sulfurtransferase (3-MST). The present article reviews the regulation of these enzymes as well as the pathways of their enzymatic and nonenzymatic degradation and elimination. The multiple interactions of H2S with other labile endogenous molecules (e.g., NO) and reactive oxygen species are also outlined. Next, the various biological targets and signaling pathways are outlined, with special reference to H2S or oxidative posttranscriptional modification (persulfidation or sulfhydration) of proteins and the effect of H2S on various channels and intracellular second messenger pathways, the regulation of gene transcription and translation, and the regulation of cellular bioenergetics and metabolism. The pharmacological and molecular tools currently available to study H2S physiology are also reviewed, including their utility and limitations. In subsequent sections, the role of H2S in the regulation of various physiological and cellular functions is reviewed, including the regulation of membrane potential, endo- and exocytosis, regulation of various cell organelles (endoplasmic reticulum, Golgi, mitochondria), regulation of cell movement, cell cycle, cell differentiation, and physiological aspects of regulated cell death. Next, the physiological roles of H2S in various cell types and organ systems are overviewed, including the role of H2S in red blood cells, immune cells, the central and peripheral nervous systems (with focus on neuronal transmission, learning, and memory formation), and regulation of vascular function (including angiogenesis as well as its specialized roles in the cerebrovascular, renal, and pulmonary vascular beds) and the role of H2S in the regulation of special senses, vision, hearing, taste and smell, and pain-sensing. Finally, the roles of H2S in the regulation of various organ functions (lung, heart, liver, kidney, urogenital organs, reproductive system, bone and cartilage, skeletal muscle, and endocrine organs) are presented, with a focus on physiology (including physiological aging) but also extending to some common pathophysiological conditions. From these data, a wide array of significant roles of H2S in the physiological regulation of all organ functions emerges and the characteristic bell-shaped biphasic effects of H2S are highlighted. In addition, key pathophysiological aspects, debated areas, and future research and translational areas are identified.

Enzymatic Depletion of Mitochondrial Inorganic Polyphosphate (polyP) Increases the Generation of Reactive Oxygen Species (ROS) and the Activity of the Pentose Phosphate Pathway (PPP) in Mammalian Cells.

Inorganic polyphosphate (polyP) is an ancient biopolymer that is well preserved throughout evolution and present in all studied organisms. In mammals, it shows a high co-localization with mitochondria, and it has been demonstrated to be involved in the homeostasis of key processes within the organelle, including mitochondrial bioenergetics. However, the exact extent of the effects of polyP on the regulation of cellular bioenergetics, as well as the mechanisms explaining these effects, still remain poorly understood. Here, using HEK293 mammalian cells under Wild-type (Wt) and MitoPPX (cells enzymatically depleted of mitochondrial polyP) conditions, we show that depletion of polyP within mitochondria increased oxidative stress conditions. This is characterized by enhanced mitochondrial O2− and intracellular H2O2 levels, which may be a consequence of the dysregulation of oxidative phosphorylation (OXPHOS) that we have demonstrated in MitoPPX cells in our previous work. These findings were associated with an increase in basal peroxiredoxin-1 (Prx1), superoxide dismutase-2 (SOD2), and thioredoxin (Trx) antioxidant protein levels. Using 13C-NMR and immunoblotting, we assayed the status of glycolysis and the pentose phosphate pathway (PPP) in Wt and MitoPPX cells. Our results show that MitoPPX cells display a significant increase in the activity of the PPP and an increase in the protein levels of transaldolase (TAL), which is a crucial component of the non-oxidative phase of the PPP and is involved in the regulation of oxidative stress. In addition, we observed a trend towards increased glycolysis in MitoPPX cells, which corroborates our prior work. Here, for the first time, we show the crucial role played by mitochondrial polyP in the regulation of mammalian redox homeostasis. Moreover, we demonstrate a significant effect of mitochondrial polyP on the regulation of global cellular bioenergetics in these cells.

Towards understanding the mechanism of action of a polyherbal formulation using a multi-pronged strategy

Herein, we investigate the cognitive effects of a traditional polyherbal formulation, Brahmi Nei (BN) for its effect on cognitive health. Network pharmacological analysis of the bioactives reported in the phytoconstituents of BN was performed by retrieving information from various databases. The in-silico predictions were experimentally validated using in vitro and in vivo models through a combination of biochemical, behavioural and molecular studies. The network pharmacological analysis of the key molecules in BN revealed their ability to modulate molecular targets implicated in memory, cognition, neuronal survival, proliferation, regulation of cellular bioenergetics and oxidative stress. Behavioral studies performed on normal adult rats administered with BN showed a significant improvement in their cognitive performance. Microarray analysis of their brain tissues exhibited an up-regulation of genes involved in oxidative phosphorylation, learning, neuronal differentiation, extension, regeneration and survival while pro-inflammatory and pro-degenerative genes were down-regulated. The oxygen consumption rate in BN-treated hippocampal cells showed a significant improvement in the bioenergetic health index when compared to untreated cells due to the mitochondrial membrane fortifying effect and anti-inflammatory property of the BN constituents. The neuroregenerative potential of BN was manifested in increase in axonal length and neurite outgrowth. Western blots and 2D gel electrophoresis revealed a reduction in pro-apoptotic proteins while increasing Akt and cyclophilin proteins. Taken together, our data reveal that BN, although traditionally used to treat anxiolytic disorders can be explored as a nutraceutical to improve neuronal health as well as a therapeutic option to treat cognitive disorders.

ATP Synthase and Mitochondrial Bioenergetics Dysfunction in Alzheimer's Disease.

Alzheimer’s Disease (AD) is the most common neurodegenerative disorder in our society, as the population ages, its incidence is expected to increase in the coming decades. The etiopathology of this disease still remains largely unclear, probably because of the highly complex and multifactorial nature of AD. However, the presence of mitochondrial dysfunction has been broadly described in AD neurons and other cellular populations within the brain, in a wide variety of models and organisms, including post-mortem humans. Mitochondria are complex organelles that play a crucial role in a wide range of cellular processes, including bioenergetics. In fact, in mammals, including humans, the main source of cellular ATP is the oxidative phosphorylation (OXPHOS), a process that occurs in the mitochondrial electron transfer chain (ETC). The last enzyme of the ETC, and therefore the ulterior generator of ATP, is the ATP synthase. Interestingly, in mammalian cells, the ATP synthase can also degrade ATP under certain conditions (ATPase), which further illustrates the crucial role of this enzyme in the regulation of cellular bioenergetics and metabolism. In this collaborative review, we aim to summarize the knowledge of the presence of dysregulated ATP synthase, and of other components of mammalian mitochondrial bioenergetics, as an early event in AD. This dysregulation can act as a trigger of the dysfunction of the organelle, which is a clear component in the etiopathology of AD. Consequently, the pharmacological modulation of the ATP synthase could be a potential strategy to prevent mitochondrial dysfunction in AD.

Dysregulation of cystathionine γ‐lyase promotes prostate cancer progression and metastasis

Hydrogen sulfide (H2S), an endogenous signaling gaseous molecule, is involved in various physiological activities, including vessel relaxation, regulation of cellular bioenergetics, inflammation, and angiogenesis. By using xenograft orthotopic implantation of prostate cancer PC3 cells and subsequently comparing bone metastatic with primary tumor‐derived cancer cells, we find that H2S‐producing enzyme cystathionine γ‐lyase (CTH) is upregulated in bone‐metastatic PC3 cells. Clinical data further reveal that the expression of CTH is elevated in late‐stage prostate cancer patients, and higher CTH expression correlates with poor survival from The Cancer Genome Atlas (TCGA) prostate cancer RNA‐seq datasets. CTH promotes NF‐κB nuclear translocation through H2S‐mediated sulfhydration on cysteine‐38 of the NF‐κB p65 subunit, resulting in increased IL‐1β expression and H2S‐induced cell invasion. Knockdown of CTH in PC3 cells results in the suppression of tumor growth and distant metastasis, while overexpression of CTH in DU145 cells promotes primary tumor growth and lymph node metastasis in the orthotopic implanted xenograft mouse model. Together, our findings provide evidence that CTH generated H2S promotes prostate cancer progression and metastasis through IL‐1β/NF‐κB signaling pathways.

AMPK: An Epigenetic Landscape Modulator.

Activated by AMP-dependent and -independent mechanisms, AMP-activated protein kinase (AMPK) plays a central role in the regulation of cellular bioenergetics and cellular survival. AMPK regulates a diverse set of signaling networks that converge to epigenetically mediate transcriptional events. Reversible histone and DNA modifications, such as acetylation and methylation, result in structural chromatin alterations that influence transcriptional machinery access to genomic regulatory elements. The orchestration of these epigenetic events differentiates physiological from pathophysiological phenotypes. AMPK phosphorylation of histones, DNA methyltransferases and histone post-translational modifiers establish AMPK as a key player in epigenetic regulation. This review focuses on the role of AMPK as a mediator of cellular survival through its regulation of chromatin remodeling and the implications this has for health and disease.

Abstract 2715: Cell cycle regulated mitophagy: a key step at the G1/S metabolic checkpoint

Abstract How tumor cells coordinate their high demand for biosynthetic processes with nutrient availability, proliferation rate and oxidative stress tolerance is still not completely understood. We recently showed that highly proliferating cells, whether normal or tumorigenic, display different metabolic requirements throughout the cell cycle, in term of glucose and glutamine utilization. The role of autophagy in cancer cells is still debated, but recent evidence pointed to its involvement in the regulation of cellular bioenergetics and nutrients utilization. To investigate autophagy regulation during cancer cells proliferation and to characterize in detail the metabolic behaviour associated with tumor cell proliferation in the different phases of the cell cycle, we used synchronized HeLaS3, a human epithelial carcinoma cell line. Treatment of the cells with a double thymidine block (DTB) results in an arrest at the G1/S boundary and the possibility to study their progression through a complete division cycle. Here we show that in the highly proliferating HeLaS3 cancer cells basal autophagy is a cell cycle regulated phenomenon, responsible for mitochondria quality control and that it plays an essential role at the G1/S checkpoint. Both general autophagy genetic inhibition and mitophagy pharmacological inhibition with Mdivi-1 during cell cycle progression lead the cells to enter faster in S phase. However, probably due to the high levels of oxidative stress dictated by their proliferation rate, autophagy-incompetent cells start to accumulate insulted and less functional mitochondria. Autophagy is likely to be involved in the induction of resistance mechanisms to treatments in different types cancers. Because of this, there is an increased interest in the possibility to use inhibitors of the autophagic pathways as adjuvants for current anti-tumor therapies. Further investigations of the mechanisms here described will be of importance to elucidate whether drugs which inhibit autophagy may be a viable and effective approach in combination with chemotherapy for the treatment of highly proliferating cancers. Citation Format: Daniela Annibali. Cell cycle regulated mitophagy: a key step at the G1/S metabolic checkpoint. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2715.

Mitochondrial Oxidative Phosphorylation System (OXPHOS) Deficits in Schizophrenia: Possible Interactions with Cellular Processes.

Mitochondria are key players in the generation and regulation of cellular bioenergetics, producing the majority of adenosine triphosphate molecules by the oxidative phosphorylation system (OXPHOS). Linked to numerous signaling pathways and cellular functions, mitochondria, and OXPHOS in particular, are involved in neuronal development, connectivity, plasticity, and differentiation. Impairments in a variety of mitochondrial functions have been described in different general and psychiatric disorders, including schizophrenia (SCZ), a severe, chronic, debilitating illness that heavily affects the lives of patients and their families. This article reviews findings emphasizing the role of OXPHOS in the pathophysiology of SCZ. Evidence accumulated during the past few decades from imaging, transcriptomic, proteomic, and metabolomic studies points at OXPHOS deficit involvement in SCZ. Abnormalities have been reported in high-energy phosphates generated by the OXPHOS, in the activity of its complexes and gene expression, primarily of complex I (CoI). In addition, cellular signaling such as cAMP/protein kinase A (PKA) and Ca(+2), neuronal development, connectivity, and plasticity have been linked to OXPHOS function and are reported to be impaired in SCZ. Finally, CoI has been shown as a site of interaction for both dopamine (DA) and antipsychotic drugs, further substantiating its role in the pathology of SCZ. Understanding the role of mitochondria and the OXPHOS in particular may encourage new insights into the pathophysiology and etiology of this debilitating disorder.

Human Resistin Promotes Neutrophil Proinflammatory Activation and Neutrophil Extracellular Trap Formation and Increases Severity of Acute Lung Injury

Although resistin was recently found to modulate insulin resistance in preclinical models of type II diabetes and obesity, recent studies also suggested that resistin has proinflammatory properties. We examined whether the human-specific variant of resistin affects neutrophil activation and the severity of LPS-induced acute lung injury. Because human and mouse resistin have distinct patterns of tissue distribution, experiments were performed using humanized resistin mice that exclusively express human resistin (hRTN(+/-)(/-)) but are deficient in mouse resistin. Enhanced production of TNF-α or MIP-2 was found in LPS-treated hRtn(+/-/-) neutrophils compared with control Rtn(-/-/-) neutrophils. Expression of human resistin inhibited the activation of AMP-activated protein kinase, a major sensor and regulator of cellular bioenergetics that also is implicated in inhibiting inflammatory activity of neutrophils and macrophages. In addition to the ability of resistin to sensitize neutrophils to LPS stimulation, human resistin enhanced neutrophil extracellular trap formation. In LPS-induced acute lung injury, humanized resistin mice demonstrated enhanced production of proinflammatory cytokines, more severe pulmonary edema, increased neutrophil extracellular trap formation, and elevated concentration of the alarmins HMGB1 and histone 3 in the lungs. Our results suggest that human resistin may play an important contributory role in enhancing TLR4-induced inflammatory responses, and it may be a target for future therapies aimed at reducing the severity of acute lung injury and other inflammatory situations in which neutrophils play a major role.

Chapter Three - Heart Mitochondrial Nitric Oxide Synthase: A Strategic Enzyme in the Regulation of Cellular Bioenergetics

Heart mitochondria play a central role in cell energy provision and in signaling. Nitric oxide (NO) is a free radical which exerts an integral regulation of the cardiovascular system not only by adapting vascular smooth muscle tone but also by influencing ion channel function, myocyte contraction, energy metabolism, and hypertrophic myocardial remodeling. This chapter analyzes the available data about heart mitochondrial NOS (mtNOS) activity and identity. The regulation of heart mtNOS by the distinctive mitochondrial environment is described by showing the effects of Ca(2+), O2, L-arginine, NADPH, mitochondrial membrane potential (ΔΨ) and the metabolic states. Evidence about the regulation of heart mtNOS in chronic hypoxia and ischemia-reperfusion models is presented. Functional implications of heart mitochondrial NOS are delineated with emphasis on the chemical reactions through which NO interacts with mitochondrial targets and exerts some of its crucial roles.

PI3K/AKT signaling regulates bioenergetics in immortalized hepatocytes

Regulation of cellular bioenergetics by PI3K/AKT signaling was examined in isogenic hepatocyte cell lines lacking the major inhibitor of PI3K/AKT signaling, PTEN (phosphatase and tensin homolog deleted on chromosome 10). PI3K/AKT signaling was manipulated using the activator (IGF-1) and the inhibitor (LY 294002) of the PI3K/AKT pathway. Activation of PI3K/AKT signaling resulted in an enhanced anaerobic glycolysis and mitochondrial respiration. AKT, when phosphorylated and activated, translocated to mitochondria and localized within the membrane structure of mitochondria, where it phosphorylated a number of mitochondrial-resident proteins including the subunits α and β of ATP synthase. Inhibition of GSK3β by either phosphorylation by AKT or lithium chloride resulted in activation of pyruvate dehydrogenase, i.e., a decrease in its phosphorylated form. AKT-dependent phosphorylation of ATP synthase subunits α and β resulted in an increased complex activity. AKT translocation to mitochondria was associated with an increased expression and activity of complex I. These data suggest that the mitochondrial signaling pathway AKT/GSK3β/PDH, AKT-dependent phosphorylation of ATP synthase, and upregulation of mitochondrial complex I expression and activity are involved in the control of mitochondrial bioenergetics by increasing substrate availability and regulating the mitochondrial catalytic/energy-transducing capacity.

Hydrogen sulfide in cell signaling, signal transduction, cellular bioenergetics and physiology in C. elegans

Hydrogen sulfide (H2S), long viewed as a toxic gas and environmental hazard, is emerging as a biological mediator with remarkable physiological and pathophysiological relevance. H2S is now viewed as the third main gasotransmitter in the mammalian body. Its pharmacological characteristic possesses similarities to the other two gasotransmitters - nitric oxide (NO) and carbon monoxide (CO). Many of the biological effects of H2S follow a bell-shaped concentration-response; at low concentration or at lower release rates it has beneficial and cytoprotective effects, while at higher concentrations or fast release rates toxicity becomes apparent. Cellular bioenergetics is a prime example for this bell-shaped dose-response, where H2S, at lower concentrations/rates serves as an inorganic substrate and electron donor for mitochondrial ATP generation, while at high concentration it inhibits mitochondrial respiration by blocking the Complex IV in the mitochondrial electron transport chain. The current review is aimed to focus on the following aspects of H2S biology: 1) a general overview of the general pharmacological characteristics of H2S, 2) a summary of the key H2S-mediated signal transduction pathways, 3) an overview of role of H2S in regulation of cellular bioenergetics, 4) key aspects of H2S physiology in C. elegans (a model system) and, finally 5) the therapeutic potential of H2S donating molecules in various disease states.

The Role of Nitric Oxide as a Regulator of Cellular Bioenergetics in Lipopolysaccharide-Stimulated Macrophage

The Role of Nitric Oxide as a Regulator of Cellular Bioenergetics in Lipopolysaccharide-Stimulated Macrophage

LETM1 is an essential component of mitochondrial calcium uptake and regulation of cellular bioenergetics

Regulation of mitochondrial Ca2+‐dependent cellular bioenergetics has been implicated in a variety of pathophysiological settings including neurodegeneration, myocardial infarction and immunity. Recently, LETM1 was proposed as a mitochondrial Ca2+/H+ antiporter; however characterization of the functional role of LETM1‐mediated Ca2+ transfer has yet to be studied. Here we demonstrate that LETM1 knockdown significantly impairs mitochondrial Ca2+ uptake and results in bioenergetic collapse. We observed that mitochondrial Ca2+ uptake is impaired in shRNA‐mediated LETM1 knockdown and ΔEF hand LETM1 and D676AD688KLETM1 overexpression cells. Knockdown of LETM1 promotes LC3 positive multilamellar vesicle formation, indicative of autophagy induction. Furthermore, cellular bioenergetic parameters including ATP and oxygen consumption were reduced. In contrast, cellular NADH and mitochondrial membrane potential were unaltered in both control and shRNA LETM1 cells. Interestingly, knockdown of LETM1 by siRNA in primary neonatal cardiomyocytes resulted in a remarkable change in mitochondrial morphology and reduced Ca2+ uptake. We also observed that LETM1 knockdown results in increased ROS production. Our results represent the first time that LETM1 has been linked to cellular bioenergetics through regulation of mitochondrial Ca2+.Supported by NIH R01 HL086699, HL086699‐01A2S1, 1S10RR027327‐01

In Vitro and In Vivo Anti-Myeloma Activity of PRLX, an Orally-Bioavailable Agent Against Cells with Constitutive Activation of Ras or Its Downstream Pathway.

Ras mutations occur in 40–60% of multiple myeloma (MM) patients and are implicated in progression to advanced MM (including plasma cell leukemia/extramedullary lesions). The small molecule PRLX (Prolexys Pharmaceuticals) was identified in the context of “synthetic lethal” chemical screening for genotype-selective cytotoxicity against cells transformed with forced expression of mutant Ras (but not against isogenic normal cell counterparts) and was tested for anti-MM activity. In MTT survival assays, 34 of 46 human MM cell lines (74%) responded to 48hr treatment with sub-μM PRLX concentrations (achievable in preclinical pharmacokinetic studies). (24 MM lines had IC50 values <300nM). PRLX anti-MM activity compared favorably with its in vitro activity against cell lines from leukemias, lymphomas, and solid tumors. PRLX activity was not restricted to cells with known Ras mutations: it was also observed in cells with FGF-R3 overexpression and/or activating mutation, suggesting that PRLX may target cells with hyper-active signaling through either upstream receptors, Ras or its downstream effectors, rather than targeting Ras biochemical activity itself. PRLX was active against MM cells resistant to conventional (Dex, alkylators, anthracyclines) and/or novel (lenalidomide, CC-4047, bortezomib, multitargeted kinase inhibitors) anti-MM agents. No antagonism was observed when PRLX was combined in vitro with other anti-MM agents, including Dex, bortezomib or CC-4047. Co-culture with BMSCs did not protect MM cells against PRLX (at doses non-toxic to BMSCs). Cell death commitment assays revealed that a pharmacologically relevant 5hr pulse with 300nM PRLX is sufficient to commit OPM-2 MM cells to cell death. Gene expression profiles (with Affymetrix U133 2.0plus oligonucleotide microarrays) showed early (<2hr) PRLX-induced modulation of broad spectrum of genes involved in regulation of cellular bioenergetics. FACS analyses of PRLX-treated cells revealed decreased mitochondrial membrane potential followed by apoptotic and then necrotic features of cell death. Reducing agents, such as tocopherol, can protect MM cells against PRLX, suggesting that release of reactive oxidative species may be an ultimate downstream mediator of the mechanism of action of this agent. The in vivo anti-MM activity of PRLX was evaluated in 2 separate in vivo models of diffuse MM lesions in SCID-beige mice, generated by i.v.injections of OPM-2 and MM.1S cells, respectively. In both models, mice were sublethally irradiated, injected i.v. with 1x106 MM cells and then randomly assigned to receive by oral gavage either PRLX 100 mg/kg (n=14) or vehicle only (n=14) on cyclical schedule of 5 days-on/2 days-off treatment. In both models, >90% of PRLX-treated mice were alive at 140 and 64 days from treatment onset, in the OPM-2 and MM.1S model respectively. In contrast, median overall survival was 35 days (95% CI: 23–47) and 35 days (95%C.I.: 31–39) in the respective control groups (Kaplan-Meier analyses, p<0.0001 in both models, by log-rank test). PRLX represents a promising novel orally bioavailable agent that merits further evaluation for possible clinical trials in advanced MM.