Targeting arginase-1 exerts antitumor effects in multiple myeloma and mitigates bortezomib-induced cardiotoxicity.

Multiple myeloma (MM) remains an incurable malignancy of plasma cells despite constantly evolving therapeutic approaches including various types of immunotherapy. Increased arginase activity has been associated with potent suppression of T-cell immune responses in different types of cancer. Here, we investigated the role of arginase 1 (ARG1) in Vκ*MYC model of MM in mice. ARG1 expression in myeloid cells correlated with tumor progression and was accompanied by a systemic drop in ʟ-arginine levels. In MM-bearing mice antigen-induced proliferation of adoptively transferred T-cells was strongly suppressed and T-cell proliferation was restored by pharmacological arginase inhibition. Progression of Vκ*MYC tumors was significantly delayed in mice with myeloid-specific ARG1 deletion. Arginase inhibition effectively inhibited tumor progression although it failed to augment anti-myeloma effects of bortezomib. However, arginase inhibitor completely prevented development of bortezomib-induced cardiotoxicity in mice. Altogether, these findings indicate that arginase inhibitors could be further tested as a complementary strategy in multiple myeloma to mitigate adverse cardiac events without compromising antitumor efficacy of proteasome inhibitors.

Qualitative analysis of enzymatic and chemical depolymerized low molecular weight heparins by UHPLC coupled with electrospray ionization quadrupole time-of-flight-mass spectrometry.

Complete heparin digestion with heparin lyase I and II results in a mixture of hexasaccharides and tetrasaccharides with 3-O-sulfo group-containing glucosamine residues at their reducing ends. Because these tetrasaccharides are derived from antithrombin III-binding sites of heparin, we examined whether this method could be applied to estimate the anticoagulant activity of heparin. Therefore, this paper presents a new low molecular weight heparin sample preparation method-chemical depolymerization. Qualitative analysis of the studied compounds and a comparison of their composition are an important contribution to the structural analysis of low molecular weight heparins, which has not been fully conducted so far. Qualitative on-line liquid chromatography-mass spectrometric analysis of these resistant oligosaccharides is also described in this paper.

Recent Developments in the Separation of Low Molecular Weight Heparin Anticoagulants.

The general function of anticoagulants is to prevent blood clotting and growing of the existing clots in blood vessels. In recent years, there has been a significant improvement in developing methods of prevention as well as pharmacologic and surgical treatment of thrombosis. For over the last two decades, low molecular weight heparins (LMWHs) have found their application in the antithrombotic diseases treatment. These types of drugs are widely used in clinical therapy. Despite the biological and medical importance of LMWHs, they have not been completely characterized in terms of their chemical structure. Due to both, the structural complexity of these anticoagulants and the presence of impurities, their structural characterization requires the employment of advanced analytical techniques. Since separation techniques play the key role in these endeavors, this review will focus on the presentation of recent developments in the separation of LMWH anticoagulants.

Bortezomib-Induced Muscle Toxicity in Multiple Myeloma.

Multiple myeloma (MM) accounts for ∼13% of all hematologic malignancies. Bortezomib treatment is effective in MM, but can be complicated with neurological side effects. We describe a patient with symptomatic MM who had a reversible metabolic myopathy associated with bortezomib administration and pathologically characterized by excessive storage of lipid droplets together with mitochondrial abnormalities. In a single-center prospective study, 14 out of 24 patients with symptomatic MM were treated with bortezomib and, among these, 7 developed muscular signs and/or symptoms. The myopathy was characterized by a proximal muscle weakness involving lower limbs and was an early complication. Complete resolution of muscle weakness occurred after treatment discontinuation. Conversely, none of the patients who received a treatment without bortezomib developed muscular symptoms. Experimental studies demonstrate that in primary human myoblasts bortezomib at low concentrations leads to excessive storage of lipid droplets together with structural mitochondrial abnormalities, recapitulating the pathologic findings observed in patient's muscle. Our data suggest that patients treated with bortezomib should be monitored for muscular signs and/or symptoms and muscle weakness should alert the clinician to the possibility of myopathy. Bortezomib-induced metabolic myopathy is a potentially reversible entity with important implications for management and treatment of patients with MM.

Selection of an optimal promoter for gene transfer in normal B cells.

Gene transfer into normal quiescent human B cells is a challenging procedure. The present study aimed to investigate whether it is possible to increase the levels of transgene expression by using various types of promoters to drive the expression of selected genes­of­interest. To produce lentiviral particles, the present study used the 2nd generation psPAX2 packaging vector and the vesicular stomatitis virus ­expressing envelope vector pMD2.G. Subsequently, lentiviral vectors were generated containing various promoters, including cytomegalovirus (CMV), elongation factor­1 alpha (EF1α) and spleen focus­forming virus (SFFV). The present study was unable to induce satisfactory transduction efficiency in quiescent normal B cells; however, infection of normal B cells with Epstein­Barr virus resulted in increased susceptibility to lentiviral transduction. In addition, the SFFV promoter resulted in a higher level of transgene expression compared with CMV or EF1α promoters. As a proof­of concept that this approach allows for stable gene expression in normal B cells, the present study used bicistronic lentiviral vectors with genes encoding fluorescent reporter proteins, as well as X­box binding protein­1 and binding immunoglobulin protein.

Characterization of Low-Molecular-Weight Heparins by Strong Anion-Exchange Chromatography.

Currently, detailed structural characterization of low-molecular-weight heparin (LMWH) products is an analytical subject of great interest. In this work, we carried out a comprehensive structural analysis of LMWHs and applied a modified pharmacopeial method, as well as methods developed by other researchers, to the analysis of novel biosimilar LMWH products; and, for the first time, compared the qualitative and quantitative composition of commercially available drugs (enoxaparin, nadroparin, and dalteparin). For this purpose, we used strong anion-exchange (SAX) chromatography with spectrophotometric detection because this method is more helpful, easier, and faster than other separation techniques for the detailed disaccharide analysis of new LMWH drugs. In addition, we subjected the obtained results to statistical analysis (factor analysis, t-test, and Newman-Keuls post hoc test).

Statins impair glucose uptake in human cells.

OBJECTIVE: Considering the increasing number of clinical observations indicating hyperglycemic effects of statins, this study was designed to measure the influence of statins on the uptake of glucose analogs by human cells derived from liver, adipose tissue, and skeletal muscle. DESIGN: Flow cytometry and scintillation counting were used to measure the uptake of fluorescently labeled or tritiated glucose analogs by differentiated visceral preadipocytes, skeletal muscle cells, skeletal muscle myoblasts, and contact-inhibited human hepatocellular carcinoma cells. A bioinformatics approach was used to predict the structure of human glucose transporter 1 (GLUT1) and to identify the presence of putative cholesterol-binding (cholesterol recognition/interaction amino acid consensus (CRAC)) motifs within this transporter. Mutagenesis of CRAC motifs in SLC2A1 gene and limited proteolysis of membrane GLUT1 were used to determine the molecular effects of statins. RESULTS: Statins significantly inhibit the uptake of glucose analogs in all cell types. Similar effects are induced by methyl-ß-cyclodextrin, which removes membrane cholesterol. Statin effects can be rescued by addition of mevalonic acid, or supplementation with exogenous cholesterol. Limited proteolysis of GLUT1 and mutagenesis of CRAC motifs revealed that statins induce conformational changes in GLUTs. CONCLUSIONS: Statins impair glucose uptake by cells involved in regulation of glucose homeostasis by inducing cholesterol-dependent conformational changes in GLUTs. This molecular mechanism might explain hyperglycemic effects of statins observed in clinical trials.

Isolation and determination of estrogens in water samples by solid-phase extraction using molecularly imprinted polymers and HPLC.

Advanced SPE with molecularly imprinted polymers (MIP) was used in this study. A noncovalent imprinting approach was applied to separate 17ß-estradiol, estriol, and estrone from water samples. Polymer material was prepared by bulk polymerization with methacrylic acid as a functional monomer, divinylbenzene and ethyleneglycol dimethacrylate as crosslinkers, and acetonitrile, acetonitrile/toluene (3:1, v/v) or isooctane/toluene (1:99, v/v) as a porogen. We also prepared an MIP film on a silica gel surface with methacrylic acid and ethyleneglycol dimethacrylate as monomers and acetonitrile as a solvent. Qualitative and quantitative hormone analyses were carried out by HPLC with various detection techniques, including UV/visible spectroscopic detection (diode array detection) and electrochemical detection (CoulArray). The results of the study indicate that MIP technology is an excellent method for the quality control of estrogens in environmental analyses with a low quantification limit for 17ß-estradiol of around 26 (diode array detection) and 0.25 ng/mL (electrochemical detection). The proposed method was found to be suitable for routine determinations of the analyzed compound in environmental laboratories.