Comptes rendus des seances de la Societe de biologie et de ses filiales最新文献

We use the knockout mice strategy to investigate the contribution of the 5-HT1B receptor in mediating the effects of selective serotonin reuptake inhibitors (SSRI). Using microdialysis in awake 129/Sv mice, we show that the absence of the 5-HT1B receptor in mutant mice (KO 1B -/-) potentiated the effect of paroxetine on extracellular 5-HT levels in the ventral hippocampus, but not in the frontal cortex compared to wild-type mice (WT). Furthermore, using the forced swimming test, we demonstrate that SSRIs decreased immobility of WT mice, and this effect is absent in KO 1B -/- mice showing therefore that activation of 5-HT1B receptors mediate the antidepressant-like effects of SSRIs. Taken together these findings suggest that 5-HT1B autoreceptors limit the effects of SSRI particularly in the hippocampus while postsynaptic 5-HT1B receptors are required for the antidepressant activity of SSRIs.

This short review presents the current stage of knowledge of our laboratory on the mechanism of action of cathepsin D and estrogens on tumor progression, mostly based on studies of human breast and ovarian cancer cell lines. Cathepsin D (cath-D) overexpression in breast cancer cells is associated with increased risk of metastasis in patients as confirmed by a recent meta-analysis of clinical studies on node negative breast cancer patients. Transfection of a human cDNA cath-D expression vector increases the metastatic potential of a rat tumor cells line when intravenously injected into nude mice. The mechanism of cath-D induced metastasis seems to require maturation of the pro-enzyme, mostly in large acidic compartments identified as phagosomes. Cath-D is mitogenic in different cell types, and different substrates (growth inhibitors, precursors of growth factor etc.) are proposed to mediate this activity. A mitogenic effect of the pro-enzyme on transmembrane receptor is not totally excluded. The mitogenic activity of estrogens in several estrogen receptor positive breast and ovarian cancer cell lines is well established in our and other laboratories. By contrast the role of estrogens during early steps of metastasis, involving cell invasion through the basement membrane and cell motility is more controversial. The motility of several estrogen receptor (ER) positive breast (MCF7, T47D) and ovarian (BG-1, SKOV3, PEO4) cancer cell lines were studied in our laboratory using a modified Boyden chamber assay. We observed, in all cases, estradiol-induced inhibition of cancer cell invasion and motility. A similar inhibitory effect of estradiol was found when the wild-type ER was stably transfected in the ER-negative MDA-MB231 cells and 3Y1-Ad12 cancer cells. The mechanism of this inhibitory effect is unknown. In ovarian cancer, however it may involve intermediary proteins such as fibulin-1, an extracellular matrix protein that strongly interacts with fibronectin and which is induced by estrogen and secreted by ovarian cancer cells. In breast cancer cells other estrogen regulated proteins may be involved. We conclude that estrogens in ER-positive breast and ovarian cancers have a dual effect, since they stimulate tumor growth but inhibit invasion and motility. This may be consistent with the good initial prognostic value of ER-positive breast cancers compared to ER negative breast cancers noted in several clinical studies, and with the better prognosis of breast cancer occurring after a prolonged treatment of menopause by estrogen as described by the collaborative group on hormonal factors in breast cancer.

Cyclin A is a positive regulatory component of kinases required for the progression through S phase and for the transition between the G2 and M phases of the cell division cycle. Previous studies conducted in established cell lines and in primary human T lymphocytes, have demonstrated that the promoter of its gene is under negative transcriptional control in quiescent cells. The DNA sequences mediating this repression have been delineated through in vitro mutagenesis as well as in vivo genomic footprinting experiments. Indirect observations suggest the involvement of proteins related to the retinoblastoma tumor suppressor protein (pRb). Using primary fibroblasts from either pRb(-/-), p107(-/-), p130(-/-) or p107(-/-)/p130(-/-) mice, we show in this work that mutation of the pRb gene has the more profound effect on cyclin A transcription. Finally, normal fibroblasts cultured in suspension fail to express cyclin A and can no longer enter S phase and proliferate, revealing thus a dependence of cyclin A expression on cell anchorage. Our work suggests the existence of at least two sets of regulators controlling cell cycle progression. On the one hand, proteins like cyclin D1, whose expression is a direct consequence of the activation of the ras signalling pathway and on the other hand, proteins like cyclin A which are secondary response effectors. As a result, growth factor stimulation leads to a transcriptional activation of the former set, while the transcription of the latter set is under the control of a repressor whose effect is alleviated after triggering the ras cascade. The status of pRb thus dictates whether cells continue their progression through the cell cycle when ras is mutated, probably by allowing the uncontrolled expression of critical genes like cyclin A.

The D3 receptor is recognized with high affinity by all antipsychotics and selectively expressed in limbic brain areas participating in the central of emotions, motivation and reward. In transfected cultured cells, stimulation of the D3 receptor inhibits cAMP formation and increases mitogenesis, which, in turn, is potentiated by activation of the cAMP cascade. This suggests that both opposite and synergistic interactions occur between the D3 receptor and the cydic AMP pathway, possibly underlying D1/D3 receptor interactions. In fact, D1 and D3 receptors colocalize in the islands of Calleja, in which they interact in opposition on c-fos mRNA expression, and in the shell of nucleus accumbens, in which they interact in synergy on substance P mRNA expression. The expression of the D3 receptor is highly dependent of the dopamine innervation: lesion of ascending dopamine neurons reduces D3 receptor mRNA and binding in the shell of nudeus accumbens, by deprivation of an unknown factor of dopamine neurons, distinct form dopamine and its cotransmitters. In agreement, expression of the D3 receptor in neurons during rat brain development starts after the settlement of dopamine innervation during the first postnatal week. However, in adult rats with a unilateral lesion of dopamine neurons, repeated treatment with levodopa rescues D3 receptor expression in the shell of nudeus accumbens and induces this expression in the dorsal striatum, a region controlling movements in which the D3 receptor is normally absent. This induction seems responsible for the behavioral sensitisation, i.e. increased responsiveness to levodopa. These observations suggest a role of the D3 receptor in the progressive increase in the therapeutic efficacy of levodopa in the initial treatment of Parkinson's disease, and/or its adversive motor and psychopathological effects during long-term treatment. Finally, various pharmacological and genetic data suggest a role of the D3 receptor in drug addiction and schizophrenia, the treatment of which could benefit from selective D3R agents.

Mice lacking the the plasma membrane dopamine transporter (DAT), following gene inactivation or knock out, show an increase in their spontaneous locomotor activity that is of the same magnitude than in normal mice treated with amphetamine or cocaine, known to increase levels of dopamine in the basal ganglia. Many adaptive responses have occurred in these animals than could not compensate for the hyper activity of the dopamine system. Surprisingly, while intracellular dopamine levels were of only 5%, extracellular dopamine levels were increased by 300%. We investigated the regulation of tyrosine hydroxylase (TH), the rate limiting enzyme of dopamine synthesis, and found that this enzyme is regulated at the levels of mRNA, protein, trafficking as well as in its regional, cellular and subcellular organization. Our results establish not only the central importance of the transporter as the key element controlling dopamine levels in the brain, but also its role in the behavioral and biochemical action of amphetamine, cocaine and morphine. In addition, these mice have provided key elements leading to possible clinical and social implications for illnesses such as Parkinson disease, attention deficit disorder and drug addiction.

Advances in medical oncology have been obtained with compounds having new structure/mechanism of action. Pharmaceutical research is largely focused on "prospective" targets identified by basic science such as the oncogenic signal transduction pathways. Ras proteins stand as converging targets that could be blocked by different approaches including inhibition of the isoprenylation of the proteins. Many academic institutions and pharmaceutical companies have embarked on the research of farnesyl transferase inhibitors. Two approaches have been developed: 1) random screening of compounds from natural/synthetic origin; 2) synthesis of compounds able to mimic the C-terminal tetrapeptidic "CAAX box" and to inhibit consequently the critical step of farnesylation. This peptidomimetic approach has been successfull since active and specific inhibitors of Ras proteins farnesylation have been synthesized in the nM range. However, several major drawbacks have been identified: in particular, most of the time, the preclinical evaluation has been done with biochemical and biological materials implicating the activated Ha-ras oncogene (very unfrequently activated in human tumors) instead of the activated Ki-ras oncogene which is the relevant target in human carcinomas. This has resulted in the selection of compounds with preferential activity on Ha-ras tumors. Nevertheless, evidence has been now generated that inhibition of farnesyl transferase of ras proteins can lead to significant experimental antitumor effects. The most convincing data are those obtained with the Merck inhibitor L739,749 which is able to cause tumor regressions of carcinomas in transgenic mice.