Predictive value of ERCC1 single-nucleotide polymorphism in patients receiving platinum-based chemotherapy for locally-advanced and advanced non-small cell lung cancer--a pilot study

Platinum-based chemotherapy is the main type of I-line treatment of advanced and non-operative NSCLC patients without EGFR gene mutation. The excision repair cross-complementation group 1 (ERCC1) is an enzyme that executes the incision of the damaged DNA strand and removes platinum-induced DNA adducts. We investigated whether ERCC1 gene polymorphism has an effect on the response to chemotherapy and survival in 43 patients with NSCLC treated with platinum-based chemotherapy. ERCC1 19007 T>C SNPs were assessed using a PCR-RFLP methods in DNA isolated from peripheral blood lymphocytes. Disease control occurred significantly (p = 0.045) more frequently in patients with CC or CT genotype compared to patients with TT genotype. Median PFS and OS for CC homozygous were 4 and 10.5 months, 4 and 12.5 months for CT heterozygous, but only 0.3 and 1.5 months for TT homozygous patients, respectively. The probability of PFS was significantly higher (HR = 0.438, 95% CI: 0.084-0.881, p = 0.03) and probability of OS was insignificantlyhigher (HR = 0.503, 95% CI: 0.129-1.137, p = 0.084) in patients with CC or CT genotype than in patients with TT genotype. Uncommon TT genotype of ERCC1 19007 T>C polymorphism could predict poor response and shortening of progression free survival in NSCLC patients treated with platinum-based I-line chemotherapy. The analysis of this polymorphism may serve as a promising tool in the qualification of advanced NSCLC patients for appropriate chemotherapy.     

Dinuclear Rh(II) complexes with one polypyridyl ligand, structure, properties and antitumor activity

Rhodium(II) complexes [Rh₂(μ-OAc)₂(OAc)(bpy)(H₂O)₂]PF₆ (1), [Rh₂(μ-OAc)₂(OAc)(phen)(H₂O)₂](PF₆)·H₂O (2), [Rh₂(μ-OOCCH₃)₃(OOCCH₃)(phen)] (3) and [Rh₂(μ-O₂CCH₃)₃(O₂CCH₃)(Ph₂phen)] (4) (Ph₂phen = 4,7-diphenyl-1,10-phenanthroline) have been synthesized and characterized by means of NMR, IR and UV-Vis spectroscopic methods. X-ray structure of complex 4·1.5(CH₃COCH₃) has been determined and its geometry and electronic structure has been elucidated using OPBE and B3LYP DFT methods. The compounds are active cytostatic agents against tumor cells.     

MDM2 promotes cell motility and invasiveness through a RING-finger independent mechanism

Recent studies connect MDM2 with increased cell motility, invasion and/or metastasis proposing an MDM2-mediated ubiquitylation-dependent mechanism. Interestingly, in renal cell carcinoma (RCC) p53/MDM2 co-expression is associated with reduced survival which is independently linked with metastasis. We therefore investigated whether expression of p53 and/or MDM2 promotes aggressive cell phenotypes. Our data demonstrate that MDM2 promotes increased motility and invasiveness in RCC cells (N.B. similar results are obtained in non-RCC cells). This study shows for the first time both that endogenous MDM2 significantly contributes to cell motility and that this does not depend upon the MDM2 RING-finger, i.e. is independent of ubiquitylation (and NEDDylation). Our data suggest that protein-protein interactions provide a likely mechanistic basis for MDM2-promoted motility which may constitute future therapeutic targets.     

Fluid perfusion as a method of cerebrospinal fluid formation rate--critical appraisal

The aim of the study was to evaluate whether or not cerebrospinal fluid formation rate (Vf) calculated according to the equation of Heisey et al., truly show the produced cerebrospinal fluid. For this reason Vf was simulated (40.6 microL/min) by an infusion pump in a plastic cylinder and the evaluation was done by comparing the results obtained between the calculated Vf and the simulated one. In both cases the result should be the same (40.6 micro/min). Other types of experiments were carried out by ventriculocisternal perfusion (92.4 microL/min) on anaesthetized and sacrificed cats. If the equation is correct, the calculated Vf for sacrificed animals should be zero, because there is no Vf in dead animals. The fact that the calculated Vf (46.5 microL/min) in the plastic cylinder was different (p < 0.0001) from the simulated one (40.6 microL/min) and that Vf was calculated even for dead animals (3-5 microL/min) clearly shows the that perfusion method may not be an accurate method for determination of Vf.     

Thermostable Pyrococcus woesei beta-D-galactosidase--high level expression, purification and biochemical properties

The gene encoding beta-D-galactosidase from Pyrococcus woesei was PCR amplified, cloned, expressed in Escherichia coli under the control of an inducible T7 promoter, purified and characterized. The expression system was developed by the construction of recombinant plasmid, based on the high copy number pUET1 vector, giving four times more efficient expression of P. woesei beta-D-galactosidase (20 mg of enzyme from 1 liter of culture) than that obtained from a previously constructed one. The recombinant enzymes were purified in a two-step procedure: double heat-denaturation of E. coli cell proteins and affinity chromatography on p-aminobenzyl 1-thio-beta-D-galactopyranoside-agarose. To achieve efficient purification of P. woesei beta-D-galactosidase by immobilized metal-ion affinity chromatography (IMAC), a His-tag was placed either at the N- or the C-terminal of the coding sequence. The obtained fusion proteins revealed the same specific activity of approximately 5400 U/mg, which was 10 times lower than the wild-type beta-D-galactosidase (51100 U/mg). The activity of P. woesei beta-D-galactosidase was enhanced by thiol compounds, Mg(2+) ions and D-galactose, and was inhibited by heavy metal ions and D-glucose, while Ca(2+) ions had no effect.     

Insights into a mummy: a paleoradiological analysis

The aim of the study was to analyze possible human skeletal remains within the wrappings of a mummy from the Archaeological Museum, Zagreb, Croatia through the use of the multidetector CT (MDCT) technology. Plain X-ray films and MDCT images of the mummy were taken in both frontal and lateral views. In a single volumetric acquisition of the whole body by MDCT 0.75 mm axial slices were obtained and combined with sagittal and coronal reformatting and three-dimensional (3D) reconstruction. Sex and age was assessed visually using standard anthropological methods. The results suggest that the mummy was of an adult female, most likely over 40 years of age at death. Pathologies observed included degenerative changes on the vertebral column and healed fractures of the lower right arm. Damage of the ethmoid bone at the roof of the nasal cavity was most likely caused by mortuary brain removal practice. Remnants of a resin and an unusual object were found inside the cranial cavity. An elongated metal object and additional three metal "belts" can be seen on the lower portion of the body. All internal organs were removed and thoracic and abdominal cavities were filled with various substances, most likely mud and pieces of linen cloth. Our results show that the MDCT is a very useful technique for assessing the human remains in archeological samples, especially in comparison to the use of plain film (X-ray), where important details are obscured and 3D imaging impossible.     

Genetic screening for glucocorticoid-remediable aldosteronism (GRA): experience of three clinical centres in Poland

Glucocorticoid-remediable aldosteronism (GRA), also known as familial hyperaldosteronism type I (FH-I, OMIM 103900), is a monogenic form of inherited hypertension caused by the presence of a chimaeric gene originating from an unequal cross-over between the CYP11B1 (11beta-hydroxylase) and CYP11B2 (aldosterone synthase) genes. The hybrid gene has the CYP11B1 sequence at the 5' end, including the promoter, and the CYP11B2 sequence at the 3' end. The aim of our study was to evaluate the prevalence of GRA in a Polish population of 129 patients with primary hyperaldosteronism (PHA) and 132 patients with essential hypertension (EH), through the use of a PCR-based test revealing the chimaeric gene. None of our PHA or EH patients was positive for the CYP11B1/CYP11B2 chimaeric gene. These data suggest that GRA is unlikely to be a common cause of hypertension in Polish subjects. However, the real prevalence of GRA in Poland, both in the high-risk group of individuals with primary hyperaldosteronism and in the general population, remains to be established.     

Kinetic intermediates of unfolding of dimeric prostatic phosphatase

Kinetics of guanidine hydrochloride (GdnHCl)-induced unfolding of human prostatic acid phosphatase (hPAP), a homodimer of 50 kDa subunit molecular mass was investigated with enzyme activity measurements, capacity for binding an external hydrophobic probe, 1-anilinonaphtalene-8-sulfonate (ANS), accessibility of thiols to reaction with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) and 2-(4'-maleimidylanilino)naphthalene-6-sulfonate (MIANS) and ability to bind Congo red dye. Kinetic analysis was performed to describe a possible mechanism of hPAP unfolding and dissociation that leads to generation of an inactive monomeric intermediate that resembles, in solution of 1.25 M GdnHCl pH 7.5, at 20 degrees C, in equilibrium, a molten globule state. The reaction of hPAP inactivation in 1.25 M GdnHCl followed first order kinetics with the reaction rate constant 0.0715 +/- 0.0024 min(-1) . The rate constants of similar range were found for the pseudo-first-order reactions of ANS and Congo red binding: 0.0366 +/- 0.0018 min(-1) and 0.0409 +/- 0.0052 min(-1), respectively. Free thiol groups, inaccessible in the native protein, were gradually becoming, with the progress of unfolding, exposed for the reactions with DTNB and MIANS, with the pseudo-first-order reaction rate constants 0.327 +/- 0.014 min(-1) and 0.216 +/- 0.010 min(-1), respectively. The data indicated that in the course of hPAP denaturation exposure of thiol groups to reagents took place faster than the enzyme inactivation and exposure of the protein hydrophobic surface. This suggested the existence of a catalytically active, partially unfolded, but probably dimeric kinetic intermediate in the process of hPAP unfolding. On the other hand, the protein inactivation was accompanied by exposure of a hydrophobic, ANS-binding surface, and with an increased capacity to bind Congo red. Together with previous studies these results suggest that the stability of the catalytically active conformation of the enzyme depends mainly on the dimeric structure of the native hPAP.     

Saproxylic moths reveal complex within-group and group-environment patterns

We investigated saproxylic moths of the family Tineidae, a neglected group inhabiting wood-decaying fungi and dead wood, within the Bia?owie?a Primeval Forest in Poland. Study data were obtained from the collection of 280 fruiting bodies of wood-decaying fungi and the subsequent rearing of adults. Spatial and statistical distribution of saproxylic moths, relationship among species and the influence of environment reflected by interaction with tree and fungal species together with tree and stand level characteristics were studied. Fifteen species and 533 individuals of saproxylic moths were reared. The fungal species, number of fruiting bodies and standing position of a tree influenced species richness. Moth abundance was influenced by fungal species, coniferous trees, increasing tree diameter, number of fruiting bodies and brown rot. Moth abundance was also enhanced by standing dead trees located in managed forests with higher canopy closure. Analyses indicate that several moth species favor a particular rot type and that some fungi host a richer fauna than others. Furthermore, our results indicate mutually independent fungal colonization events by saproxylic moth species, and thus a possible mechanism exists for competition avoidance with other saproxylic moths. Saproxylic moths revealed complex within-group patterns that responded differently to environmental variables. Thus, potential conservation of these organisms requires various approaches including ecosystem management, especially in the context of addressing their diverse habitat requirements.