CITED2 and NCOR2 in anti-oestrogen resistance and progression of breast cancer.

BACKGROUND: Endocrine therapies of breast cancer are effective but ultimately fail because of the development of treatment resistance. We have previously revealed several genes leading to tamoxifen resistance in vitro by retroviral insertion mutagenesis. To understand the manner in which these genes yield tamoxifen resistance, their effects on global gene expression were studied and those genes resulting in a distinct gene expression profile were further investigated for their clinical relevance. METHODS: Gene expression profiles of 69 human breast cancer cell lines that were made tamoxifen resistant through retroviral insertion mutagenesis were obtained using oligonucleotide arrays and analysed with bioinformatic tools. mRNA levels of NCOR2 and CITED2 in oestrogen receptor-positive breast tumours were determined by quantitative RT-PCR. mRNA levels were evaluated for association with metastasis-free survival (MFS) in 620 patients with lymph node-negative primary breast cancer who did not receive systemic adjuvant therapy, and with clinical benefit in 296 patients receiving tamoxifen therapy for recurrent breast cancer. RESULTS: mRNA expression profiles of most tamoxifen-resistant cell lines were strikingly similar, except for the subgroups of cell lines in which NCOR2 or CITED2 were targeted by the retrovirus. Both NCOR2 and CITED2 mRNA levels were associated with MFS, that is, tumour aggressiveness, independently of traditional prognostic factors. In addition, high CITED2 mRNA levels were predictive for a clinical benefit from first-line tamoxifen treatment in patients with advanced disease. CONCLUSIONS: Most retrovirally targeted genes yielding tamoxifen resistance in our cell lines do not impose a distinctive expression profile, suggesting that their causative role in cell growth may be accomplished by post-transcriptional processes. The associations of NCOR2 and CITED2 with outcome in oestrogen receptor-positive breast cancer patients underscore the clinical relevance of functional genetic screens to better understand disease progression, which may ultimately lead to the development of improved treatment options.

Impact of discrepancies between the Abbott realtime and cobas TaqMan assays for quantification of human immunodeficiency virus type 1 group M non-B subtypes.

Viral loads in 249 clinical samples from individual patients infected with human immunodeficiency virus type 1 non-B subtypes were determined with both the Abbott RealTime and Cobas TaqMan assays. The differences exceeded 0.5 log for about 20% of samples and 1 log for 3%, with higher values always from the Abbott assay in the latter cases.

Mithramycin A activates Fas death pathway in leukemic cell lines.

Mithramycin A (MMA, trade name Plicamycin) can facilitate TNFalpha- (Tumor Necrosis Factor) and Fas ligand-induced apoptosis. Besides, several drugs play their anticancer effect through Fas apoptotic pathway. So we investigated the effect of MMA on Fas signaling. In this study we show that MMA induces apoptosis in Fas sensitive Jurkat cells and Fas resistant KG1a cells. This effect involves Fas apoptotic pathway: cell exposure to MMA leads to Fas clustering at the cell surface, DISC (Death Inducing Signaling Complex) formation and caspase cleavage. This phenomenon is independent of Fas ligand/Fas interaction and blockade of Fas death pathway partially inhibits MMA-induced apoptosis. Moreover the activation of Fas apoptotic pathway by MMA is correlated to the modulation of c-Flip(L) expression. Finally, pre-treatment with sub-lethal doses of MMA sensitizes KG1a cells to chemotherapeutic agents. Thus all these results may have important implications to improve clinical treatments.

Cerebral venous sinus thrombosis in children: risk factors, presentation, diagnosis and outcome.

Neuroimaging and management advances require review of indications for excluding cerebral venous sinus (sinovenous) thrombosis (CSVT) in children. Our goals were to examine (i) clinical presentations of CSVT, (ii) prothrombotic risk factors and other predisposing events, (iii) clinical and radiological features of brain lesions in CSVT compared with arterial stroke, and (iv) predictors of outcome. We studied 42 children with CSVT from five European paediatric neurology stroke registries. Patients aged from 3 weeks to 13 (median 5.75) years (27 boys; 64%) presented with lethargy, anorexia, headache, vomiting, seizures, focal signs or coma and with CSVT on neuroimaging. Seventeen had prior chronic conditions; of the 25 previously well patients, 23 had recent infections, eight became dehydrated and six had both. Two children had a history compatible with prior CSVT. Anaemia and/or microcytosis (21 probable iron deficiency, five haemolytic, including two with sickle cell disease and one with beta-thalassaemia) was as common (62%) as prothrombotic disorder (13/21 screened). High factor VIII and homozygosity for the thermolabile methylene tetrahydrofolate reductase polymorphism were the commonest prothrombotic disorders. The superficial venous system was involved in 32 patients, the deep in six, and both in four. Data on the 13 children with bland infarction and the 12 with haemorrhage in the context of CSVT were compared with those from 88 children with ischaemic (AIS) and 24 with haemorrhagic (AHS) arterial stroke. In multiple logistic regression, iron deficiency, parietal infarction and lack of caudate involvement independently predicted CSVT rather than arterial disease. Five patients died, three acutely, one after recurrence and one after 6 months being quadriparetic and blind. Follow-up ranged from 0.5 to 10 (median 1) years. Twenty-six patients (62%) had sequelae: pseudotumour cerebri in 12 and cognitive and/or behavioural disabilities in 14, associated with epilepsy in three, hemiparesis in two and visual problems in two. Eighteen patients, including six with haemorrhage, were anticoagulated. Older age [odds ratio (OR) 1.54, 95% confidence limits (CI) 1.12, 2.13, P = 0.008], lack of parenchymal abnormality (OR 0.17, 95% CI 0.02, 1.56, P = 0.1), anticoagulation (OR 24.2, 95% CI 1.96, 299) and lateral and/or sigmoid sinus involvement (OR 16.2, 95% CI 1.62, 161, P = 0.02) were independent predictors of good cognitive outcome, although the last predicted pseudotumour cerebri. Death was associated with coma at presentation. Of 19 patients with follow-up magnetic resonance (MR) venography, three had persistent occlusion, associated with anaemia and longer prodrome. A low threshold for CT or MR venography in children with acute neurological symptoms is essential. Nutritional deficiencies may be modifiable risk factors. A paediatric anticoagulation trial may be required, after the natural history has been further established from registries of cases with and without treatment.

Differentiating the mechanisms of antiresorptive action of nitrogen containing bisphosphonates.

Bisphosphonates (BPS) inhibit bone resorption and are divided into two classes according to their chemical structure and mechanism of action: nonnitrogen containing BPS such as etidronate and clodronate that are of low potency and inhibit osteoclast function via metabolism into toxic ATP-metabolites and nitrogen-containing BPS (NBPS), such as alendronate and risedronate that inhibit the enzyme of the mevalonate biosynthetic pathway farnesyl pyrophosphate synthase (FPPS), resulting in inhibition of the prenylation of small GTP-binding proteins in osteoclasts and disruption of their cytoskeleton. Previously, studies in various cell types suggested, however, that pamidronate functions by mechanism(s) additional or independent of the mevalonate pathway. To examine if such mechanism(s) are also involved in the action of NBPS on osteoclastic bone resorption, we examined the action of alkyl and heterocyclic NBPS with close structural homology on FPPS/isopentenyl pyrophosphate isomerase (IPPI) activity, on osteoclastic resorption, and on reversibility of this effect with GGOH. As expected, both pamidronate and alendronate suppressed bone resorption and FPPS/IPPI activity, the latter with greater potency than the first. Surprisingly, however, unlike alendronate, the antiresorptive effect of pamidronate was only partially reversible with GGOH, indicating the involvement of mechanism(s) of action additional to that of suppression of FPPS. Comparable results were obtained with the heterocyclic NBP NE-21650, a structural analog of risedronate. Thus, despite an effect on FPPS, the actions on bone resorption of some NBPS may involve mechanisms additional to suppression of FPPS. These findings may lead to identification of additional pathways that are important for bone resorption and may help to differentiate among members of the NBP class which are currently distinguished only according to their potency to inhibit bone resorption.

Neuroanatomy of cerebellum and olfactory bulb in a substrain of C57BL/6J inbred mice carrying a spontaneous mutation.

Mice of the inbred C57BL/6JNmg substrain carry a mutation decreasing the size of the zinc-rich hippocampal intra- and infrapyramidal mossy fibre (IIPMF) terminal fields. In the present experiment, it was investigated whether this neurological mutation has also effects on other characteristics of the brain. No morphological differences were found in two other laminated neural structures, the olfactory bulb, where the accessory granular layer is also rich in zinc terminals, and the cerebellum. However, the mutants had a somewhat inferior performance on a motor function task known to test cerebellar involvement. The present findings confirm that previously found effects of this mutation on different types of behaviour are most probably due to the IIPMF. These substrains provide a powerful tool to localise the gene involved and subsequently investigate the plausible pathways leading from gene to behaviour.

Mitochondria-targeting drugs arsenic trioxide and lonidamine bypass the resistance of TPA-differentiated leukemic cells to apoptosis.

Exposure of U937 human leukemic cells to the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) induces their differentiation into monocyte/macrophage-like cells. This terminal differentiation is associated with a resistant phenotype to apoptosis induced by the topoisomerase II inhibitor etoposide. The inhibition occurs upstream of the mitochondrial release of cytochrome c and the activation of procaspase-2, -3, -6, -7, -8, and -9. By using cell-free systems, it was demonstrated that the mitochondrial pathway to cell death that involves mitochondrial membrane depolarization, cytochrome c release and cytosolic activation of procaspases by cytochrome c/dATP remains functional in TPA-differentiated U937 cells. Accordingly, 2 drugs recently shown to target the mitochondria, namely lonidamine and arsenic trioxide, bypass the resistance of TPA-differentiated U937 cells to classical anticancer drugs. Cell death induced by the 2 compounds is associated with mitochondrial membrane depolarization, release of cytochrome c and Smac/Diablo from the mitochondria, activation of caspases, poly(ADP-ribose) polymerase cleavage and internucleosomal DNA fragmentation. Moreover, the decreased glutathione content associated with the differentiation process amplifies the ability of arsenic trioxide to activate the mitochondrial pathway to cell death. Similar results were obtained by comparing undifferentiated and TPA-differentiated human HL60 leukemic cells. These data demonstrate that mitochondria-targeting agents bypass the resistance to classical anticancer drugs induced by TPA-mediated leukemic cell differentiation. (Blood. 2001;97:3931-3940)

The endometrial approach in contraception.

PIP: Contraceptive techniques that selectively impair endometrium receptivity have the potential to reduce the risk of cardiovascular complications associated with hormonal methods. However, further research is needed to provide better treatment when oral estrogen is contraindicated, to improve tolerance of progestin-only contraceptives, and identify alternative techniques that will not interfere with the endocrine events of the cycle. This article presents an overview of the endometrial approach to contraception. Discussed are endometrium physiology and the mechanisms of hormonal contraception, endometrial modifications following hormonal contraceptive use, emergency contraception, and intrauterine steroid administration. Outstanding research issues include the mechanisms of endometrial bleeding, definition of molecular and cellular targets for an endometrial approach to contraception, progesterone action, integrins, placental protein 14, insulin growth factor binding protein-1, and plasminogen activators. Short-term developments in this area are expected to include the intrauterine administration of steroids and continuous administration of antiprogestins at doses that do not interfere with secretion. Long-term development will focus on defining new targets for a pharmacological approach in a tissue-specific manner.^ieng

[In vitro activity of a new fluoroquinolone, marbofloxacin (RO 09-1168) against strictly anaerobic bacteria and some bacteria from human fecal flora]. / Activité in vitro d'une nouvelle fluoroquinolone, la marbofloxacine (RO 09-1168) sur les anaérobies stricts et quelques bactéries de la flore fécale humaine.

As recommended by the EEC, the in vitro activity of a veterinary quinolone has to be evaluated on human strains isolated from the gut and especially against anaerobes. Thus, the MICs for Marbofloxacin were determined by the reference agar method (Norma NCCLS M11 A3) using Wilkins Chalgren medium for the 124 anaerobic strains and Mueller Hinton agar for E. coli, Proteus, Enterococcus and Lactobacillus. On the whole aerobes, the activity of Marbofloxacin was equal or slightly greater to that of ofloxacin. The modal MIC was 0.03 mg/l for Enterobacteriaceae, 2 mg/l and 16 mg/l for Enterococcus and Lactobacillus strains, respectively. Considering the anti-anaerobic activity of this new drug, in comparison with that of ofloxacin, MICs for ofloxacin were generally one log2 higher for Gram negative bacilli and two log2 higher for Gram positive bacilli. Modal MICs for marbofloxacin were 0.12 mg/l for Clostridium perfringens, 0.5 mg/l for Bacteroides fragilis, Fusobacterium, Prevotella, and Peptostreptococcus strains, 1 mg/l for Bifidobacteria, 2 mg/l for non-fragilis members of the B. fragilis group, and 4 mg/l for C. difficile strains, respectively. It appears in vitro that Marbofloxacin was a more potent agent than ofloxacin on the whole anaerobes.

[Perspectives of contraception]. / Perspectives de la contraception.

According to demographic provision, the world population will soon reach 6 billions. It is therefore urgent to increase women's access to well tolerated, efficient, low cost contraceptives. The primary goal is to inform women on their reproductive health and on the use of contraceptives. Secondly, new molecules should be shortly available: low dose estroprogestins containing new non androgenic progestins and 17 beta-estradiol, RU 486 as a post-coïtal pill, levonorgestrel or desogestrel containing IUDs. The daily intake of low dose antiprogestins to interfere with endometrial maturation is also currently studied. In the future, anti-FSH molecules or tissue-specific steroids could be designed.

A single injection of a gonadotropin-releasing hormone (GnRH) antagonist (Cetrorelix) postpones the luteinizing hormone (LH) surge: further evidence for the role of GnRH during the LH surge.

OBJECTIVES: To assess the ability of a new third-generation GnRH antagonist, Cetrorelix (Asta Medica AG, Frankfurt am Main, Germany), to postpone the LH surge after a single injection during the late follicular phase. DESIGN: A single 5-mg (group 1, n = 7) or 3-mg (group 2, n = 3) dose SC of Cetrorelix was administered during the late follicular phase, on the day of the cycle when plasma E2 exceeded 150 pg/mL (550 pmol/L). Estradiol, LH, FSH, and P levels were measured daily from day 5 of the cycle until day 10 after antagonist administration. Transvaginal ultrasonographies were performed on the day of injection and after antagonist treatment. SUBJECTS: Ten normal women with regular ovulatory menstrual cycles. RESULTS: In group 1, Cetrorelix was administered on day 14.6 +/- 5 (mean +/- SD) of the cycle, when the mean plasma E2 level was 181 +/- 32 pg/mL (664 +/- 117 pmol/L) (mean +/- SD). Plasma LH and FSH decreased by 56% +/- 19% and 29.5% +/- 16% (mean +/- SD), respectively, reaching the nadir 24 hours after Cetrorelix administration. Estradiol decreased by 85% +/- 17%, reaching the nadir 48 hours after antagonist injection. In group 2, Cetrorelix was administered on day 14.3 +/- 1.2 of the cycle when the mean plasma E2 level was 169 +/- 21 pg/mL (618 +/- 77 pmol/L). Plasma LH and FSH decreased by 66% +/- 18% and 32% +/- 6%, respectively, reaching a nadir 24 hours after Cetrorelix administration. Estradiol decreased by 81% +/- 9%, reaching the nadir 24 to 48 hours after antagonist administration. The LH surge was interrupted in every case. In six of seven subjects from group 1, the LH surge was delayed, occurring 6 to 17 days after the antagonist injection. In the remaining woman, Cetrorelix was administered at the beginning of the LH surge (LH = 13 IU/L): the LH level fell immediately by 54%, and the surge was postponed by 3 days. In group 2, in three of three subjects, the LH surge was delayed, occurring 6 to 9 days after the antagonist injection. No adverse effects were observed, except for very slight and transient erythema and pruritus at the injection site. CONCLUSION: Cetrorelix is a very potent new GnRH antagonist. A single injection during the late follicular phase delays the LH surge, even if the latter has already begun. In addition, this new-generation GnRH antagonist is very well tolerated and simple to use. Our data reinforce the role of GnRH during the LH surge and point to a role for new GnRH antagonists in controlled ovarian hyperstimulation to avoid premature LH surges and subsequent luteinization.