Osteopontin overexpression in breast cancer: knowledge gained and possible implications for clinical management

Osteopontin (OPN) is a secreted protein that is overexpressed in a number of human cancers, and has been associated with increased metastatic burden and poor prognosis in breast cancer patients. The OPN protein contains several conserved structural elements including heparin- and calcium-binding domains, a thrombin-cleavage site, a CD44 binding site, and two integrin-binding sites. Experimental studies have shown that the ability of OPN to interact with a diverse range of factors, including cell surface receptors (integrins, CD44), secreted proteases (matrix metalloproteinases, urokinase plasminogen activator), and growth factor/receptor pathways (TGFalpha/EGFR, HGF/Met) is central to its role in malignancy. These complex signaling interactions can result in changes in gene expression, which ultimately lead to alterations in cell properties involved in malignancy such as adhesion, migration, invasion, enhanced tumor cell survival, tumor angiogenesis, and metastasis. Therefore, OPN is not merely associated with cancer, but rather it plays a multi-faceted functional role via complex molecular cross-talk with other factors. This review will focus on the role of OPN in breast cancer, in particular on the malignancy-promoting aspects of OPN that may reveal opportunities for new approaches to the clinical management of breast cancer.     

A randomized controlled clinical trial to determine the optimum duration of G-CSF priming prior to BM stem cell harvesting

BACKGROUND: Harvesting of hemopoietic stem cells (HSC) from G-CSF-primed BM for autologous transplantation is an alternative to collection of unprimed BM or G-CSF-primed peripheral blood (PB). However, the optimum number of days of G-CSF administration for this purpose is unknown. We set out to determine whether cell yields could be optimized by varying the number of days of G-CSF administration prior to BM stem cell harvesting. METHODS: We conducted a randomized controlled single-center trial of 6 days (the standard) vs. 4 days of G-CSF administration and compared yields of total nucleated cells (TNC), CD34(+) HSC and CFU-GM cells per kilogram patient body weight. Statistical analysis was by Student's t-test. RESULTS: Twenty-four patients were enrolled; 13 received 6 days and 11 received 4 days of G-CSF administration. Analysis of the first harvest aspirate showed higher proportions of CD34(+) HSC (P=0.02) and CFU-GM (P=0.03) in the 4-day group. For the 6-day and 4-day groups, respectively, the median yield of TNC/kg was 6.5 x 10(8) and 5.4 x 10(8) (P=0.28), of CD34(+) cells/kg 0.56 x 10(6) and 0.98 x 10(6) (P=0.04) and of CFU-GM cells/kg 1.66 x 10(5) and 1.55 x 10(5) (P=0.75). DISCUSSION: These results suggest that by 6 days the HSC-stimulating effect of G-CSF has passed its peak and that 4 days should be adopted as the standard for G-CSF priming prior to BM stem cell harvesting for autologous transplantation.     

Laboratory evolution of cytochrome p450 BM-3 monooxygenase for organic cosolvents

Cytochrome p450 BM-3 (EC 1.14.14.1) catalyzes the hydroxylation and/or epoxidation of a broad range of substrates, including alkanes, alkenes, alcohols, fatty acids, amides, polyaromatic hydrocarbons, and heterocycles. For many of these notoriously water-insoluble compounds, p450 BM-3's K(m) values are in the millimolar range. Polar organic cosolvents are therefore added to increase substrate solubility and achieve high catalytic efficiency. Using p450 BM-3 as a catalyst for these important transformations requires that we improve its ability to tolerate the cosolvents. By directed evolution, we improved the activity of p450 BM-3 in the presence of dimethylsulfoxide (DMSO) and tetrahydrofuran (THF), achieving increases in specific activity up to 10-fold in 2% (v/v) THF and 6-fold in 25% (v/v) DMSO. The engineered p450 BM-3's are also significantly more resistant to acetone, acetonitrile, dimethylformamide, and ethanol as cosolvents in the reaction.     

The X-Ray Crystal Structure of Escherichia coli Succinic Semialdehyde Dehydrogenase; Structural Insights into NADP+/Enzyme Interactions

Background

In mammals succinic semialdehyde dehydrogenase (SSADH) plays an essential role in the metabolism of the inhibitory neurotransmitter γ-aminobutyric acid (GABA) to succinic acid (SA). Deficiency of SSADH in humans results in elevated levels of GABA and γ-Hydroxybutyric acid (GHB), which leads to psychomotor retardation, muscular hypotonia, non-progressive ataxia and seizures. In Escherichia coli, two genetically distinct forms of SSADHs had been described that are essential for preventing accumulation of toxic levels of succinic semialdehyde (SSA) in cells.

Methodology/Principal Findings

Here we structurally characterise SSADH encoded by the E coli gabD gene by X-ray crystallographic studies and compare these data with the structure of human SSADH. In the E. coli SSADH structure, electron density for the complete NADP⁺ cofactor in the binding sites is clearly evident; these data in particular revealing how the nicotinamide ring of the cofactor is positioned in each active site.

Conclusions/Significance

Our structural data suggest that a deletion of three amino acids in E. coli SSADH permits this enzyme to use NADP⁺, whereas in contrast the human enzyme utilises NAD⁺. Furthermore, the structure of E. coli SSADH gives additional insight into human mutations that result in disease.     

Contraceptives,counselling, and pregnancy in women with sickle cell disease.

Sickle cell disease is listed in the manufacturers'' data sheets in the United Kingdom as a contraindication to the use of most combined contraceptive pills; the result is confused advice on family planning to a group of women who are at substantial risk from both planned and unplanned pregnancy. A study in north London on the use of contraceptives by women with sickle cell disease indicates that the use of combined oral contraceptives is common. Although medical staff usually advised against pregnancy, such advice was almost always ignored. Over half of the women surveyed had some knowledge about antenatal diagnosis. Family planning advice should be an integral part of the care of women with sickle cell disease. In the absence of specific data to the contrary all methods of contraception may be considered, although with appropriate caution.     

Synthesis and evaluation of aminobenzothiazoles as blockers of N- and T-type calcium channels

Both N- and T-type calcium ion channels have been implicated in pain transmission and the N-type channel is a well-validated target for the treatment of neuropathic pain. An SAR investigation of a series of substituted aminobenzothiazoles identified a subset of five compounds with comparable activity to the positive control Z160 in a FLIPR-based intracellular calcium response assay measuring potency at both Ca_V2.2 and Ca_V3.2 channels. These compounds may form the basis for the development of drug leads and tool compounds for assessing in vivo effects of variable modulation of Ca_V2.2 and Ca_V3.2 channels.