Glut-1 Expression Correlates with Basal-like Breast Cancer

INTRODUCTION: Glucose transporter 1 (Glut-1) is a facilitative glucose transporter expressed in many cancers including breast cancer. Basal-like breast cancer (BLBC) is a high-risk disease associated with poor prognosis and lacks the benefit of targeted therapy. The aim of this study was to characterize the immunohistochemical (IHC) expression of Glut-1 in patients with BLBC compared with non-BLBC. MATERIALS AND METHODS: We identified 523 cases of invasive breast carcinoma from our database. The clinicopathologic findings and the biologic markers including estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (Her2) status were reviewed. IHC stains for cytokeratin 5/6 (CK5/6), epidermal growth factor receptor (EGFR), p53, and Glut-1 were performed on tissue microarray using standard procedures. BLBC was defined as ER-,PR-, Her2-, and CK5/6+ and/or EGFR+. RESULTS: Of informative cases, 14.7% were categorized as BLBC versus 85.3% as non-BLBC. Glut-1 was expressed in 42 (76.4%) of 55 BLBCs, whereas only 55 (23.8%) of 231 non-BLBCs showed immunostaining for Glut-1 (P < .001). Overall, Glut-1 expression was significantly associated with high histologic grade, ER negativity, PR negativity, CK5/6 positivity, EGFR expression, and high p53 expression (P < .001). However, there was no correlation between Glut-1 immunostaining and patient's outcome. CONCLUSIONS: Our results show that Glut-1 is significantly associated with BLBC and might be a potential therapeutic target for this aggressive subgroup of breast cancer, and this warrants further investigations.     

8-Modified-2'-deoxyadenosine analogues induce delayed polymerization arrest during HIV-1 reverse transcription

The occurrence of resistant viruses to any of the anti-HIV-1 compounds used in the current therapies against AIDS underlies the urge for the development of new drug targets and/or new drugs acting through novel mechanisms. While all anti-HIV-1 nucleoside analogues in clinical use and in clinical trials rely on ribose modifications for activity, we designed nucleosides with a natural deoxyribose moiety and modifications of position 8 of the adenine base. Such modifications might induce a steric clash with helix αH in the thumb domain of the p66 subunit of HIV-1 RT at a distance from the catalytic site, causing delayed chain termination. Eleven new 2'-deoxyadenosine analogues modified on position 8 of the purine base were synthesized and tested in vitro and in cell-based assays. In this paper we demonstrate for the first time that chemical modifications on position 8 of 2'-deoxyadenosine induce delayed chain termination in vitro, and also inhibit DNA synthesis when incorporated in a DNA template strand. Furthermore, one of them had moderate anti-HIV-1 activity in cell-culture. Our results constitute a proof of concept indicating that modification on the base moiety of nucleosides can induce delayed polymerization arrest and inhibit HIV-1 replication.     

Role of BMI-associated loci identified in GWAS meta-analyses in the context of common childhood obesity in European Americans

Obesity is a serious health concern for children and adolescents, particularly in Western societies, where its incidence is now considered to have reached epidemic proportions. A number of genetic determinants of adult BMI have already been established through genome wide association studies (GWAS), most recently from the GIANT meta-analysis of such datasets combined. In this current study of European Americans, we examined the 32 loci detected in that GIANT study in the context of common childhood obesity within a cohort of 1,097 cases (defined as BMI ≥95th percentile), together with 2,760 lean controls (defined as BMI <50th percentile), aged between 2 and 18 years old. Nine of these single-nucleotide polymorphims (SNPs) yielded at least nominal evidence for association with common childhood obesity, namely at the FTO, TMEM18, NRXN3, MC4R, SEC16B, GNPDA2, TNNI3K, QPCTL, and BDNF loci. However, overall 28 of the 32 loci showed directionally consistent effects to that of the adult BMI meta-analysis. We conclude that among the 32 loci that have been reported to associate with adult BMI in the largest meta-analysis of BMI to date, at least nine also contribute to the determination of common obesity in childhood in European Americans, as demonstrated by their associations in our pediatric cohort.     

FIRST EVIDENCE OF PALYTOXIN ANALOGUES FROM AN OSTREOPSIS MASCARENENSIS (DINOPHYCEAE) BENTHIC BLOOM IN SOUTHWESTERN INDIAN OCEAN1

Benthic dinoflagellates of the genus Ostreopsis Schmidt are common in tropical and subtropical water, and some species produce toxins potentially involved in human intoxication events. A benthic bloom of Ostreopsis mascarenensis Quod was observed near Rodrigues Island during a survey of benthic dinoflagellates in the southwestern Indian Ocean. The morphology of O. mascarenensis was studied by LM and SEM. Preliminary screening of a crude extract of an O. mascarenensis bloom revealed neurotoxicity in mice similar to that induced by palytoxin. After partition of the crude extract, the highest toxicity was retained in the butanol‐soluble fraction, which retained hemolytic activity suggestive of palytoxin analogues. Two new toxins, mascarenotoxin‐A and ‐B, were resolved from this fraction by HPLC coupled to a diode array detector. The closed mass spectrum profile and fragmentation pattern obtained by advanced nano–electrospray ionization quadrupole time‐of‐flight mass spectrometry between purified toxins and a reference palytoxin confirmed the mascarenotoxins as palytoxin analogues. These results were confirmed by tandem mass spectrometry with the identification of specific fragment ion m/z 327. An on‐line liquid chromatography protocol coupled to tandem mass spectrometry was developed for detection of these palytoxin analogues. The present study describes the first purification, chemical, and toxicological characterization of new palytoxin analogues isolated from a benthic bloom of O. mascarenensis. These results suggest that O. mascarenensis, which is largely distributed in the southwestern Indian Ocean, could be a source of palytoxin poisoning in this tropical area.     

Chromatographic analysis of carbamazepine binding to human serum albumin

In this study, high-performance affinity chromatography was used to characterize the binding of carbamazepine to an immobilized human serum albumin (HSA) column. Frontal analysis was first used to determine the association equilibrium constant and binding capacity for carbamazepine on this column at various temperatures. The non-specific binding of carbamazepine within the column was also considered. The results indicated that carbamazepine had a single binding site on HSA with an association equilibrium constant of 5.3 x 10(3)M(-1) at pH 7.4 and 37 degrees C. This was confirmed through zonal elution self-competition studies. The value of DeltaG for this reaction was -5.35 kcal/mol at 37 degrees C, with an associated change in enthalpy (DeltaH) of -6.45 kcal/mol and a change in entropy (DeltaS) of -3.56 cal/molK. The location of this binding region was examined by competitive zonal elution experiments using probe compounds with known sites on HSA. It was found that carbamazepine had direct competition with l-tryptophan, a probe for the indole-benzodiazepine site of HSA, but allosteric interactions with probes for the warfarin, tamoxifen and digitoxin sites. Changes in the pH, ionic strength, and organic modifier content of the mobile phase were used to identify the predominant forces in the carbamazepine-HSA interaction.     

Transferrin's mechanism of interaction with receptor 1

The kinetics and thermodynamics of the interactions of transferrin receptor 1 with holotransferrin and apotransferrin in neutral and mildly acidic media are investigated at 37 degrees C in the presence of CHAPS micelles. Receptor 1 interacts with CHAPS in a very fast kinetic step (<1 micros). This is followed in neutral media by the interaction with holotransferrin which occurs in two steps after receptor deprotonation, with a proton dissociation constant (K(1a)) of 10.0 +/- 1.5 nM. The first step is detected by the T-jump technique and is associated with a molecular interaction between the receptor and holotransferrin. It occurs with a first-order rate constant (k(-1)) of (1.6 +/- 0.2) x 10(4) s(-1), a second-order rate constant (k(1)) of (3.20 +/- 0.2) x 10(10) M(-1) s(-1), and a dissociation constant (K(1)) of 0.50 +/- 0.07 microM. This step is followed by a slow change in the conformation with a relaxation time (tau(2)) of 3400 +/- 400 s and an equilibrium constant (K(2)) of (4.6 +/- 1.0) x 10(-3) with an overall affinity of the receptor for holotransferrin [(K'1)(-1)] of (4.35 +/- 0.60) x 10(8) M(-1). Apotransferrin does not interact with receptor 1 in neutral media, between pH 4.9 and 6, it interacts with the receptor in two steps after a receptor deprotonation (K(2a) = 2.30 +/- 0.3 microM). The first step occurs in the range of 1000-3000 s. It is ascribed to a slow change in the conformation which rate-controls a fast interaction between apotransferrin and receptor 1 with an overall affinity constant [(K(3))(-1)] of (2.80 +/- 0.30) x 10(7) M(-1). These results imply that receptor 1 probably exists in at least two forms, the neutral species which interacts with holotransferrin and not with apotransferrin and the acidic species which interacts with apotransferrin. At first, the interaction of the neutral receptor with holotransferrin is extremely fast. It is followed by the slow change in conformation, which leads to an important stabilization of the thermodynamic structure. In the acidic media of the endosome, the interaction of apotransferrin with the acidic receptor is sufficiently strong and rate-controlled by a very slow change in conformation which allows recycling back to the plasma membrane.     

Chromatographic analysis of allosteric effects between ibuprofen and benzodiazepines on human serum albumin

The effects of (R)- and (S)-ibuprofen on the binding of benzodiazepines to human serum albumin (HSA) were examined by biointeraction chromatography. The displacement of benzodiazepines from HSA by (R)- and (S)-ibuprofen was found to involve negative allosteric interactions (or possible direct competition) for most (R)-benzodiazepines. However, (S)-benzodiazepines gave positive or negative allosteric effects and direct competition when displaced by (R)- or (S)-ibuprofen. Association equilibrium constants and coupling constants measured for these effects indicated that they involved two classes of ibuprofen binding regions (i.e., low- and high-affinity sites). Based on these results, a model was proposed to explain the binding of benzodiazepines to HSA and their interactions with ibuprofen. This model gave good agreement with previous reports examining the binding of benzodiazepines to HSA.     

Functional impairment of CD4 T cells despite normalization of T cell number in HIV

Using an in vitro model, we demonstrate that when CD4 T cells from HIV infected subjects are enriched from total blood lymphocytes the immune response to antigen is augmented. However, augmentation of this response is confined to HIV infected subjects with relatively preserved CD4 T cell counts. Enriching for CD4 T cells had no effect on antigen responses in patients with low CD4 lymphocyte counts. These findings support the concept that CD4 T cells in late stage HIV have inherent qualitative defects.