No abstract

Loss of p53 suppressor function, through mutations or inactivation of the p53 pathway, occurs in most human cancers. SGT-53 is a liposomal nanocomplex designed for systemic, tumor-targeting delivery of the wt p53 gene. In this nanodelivery system, an anti-transferrin receptor single-chain antibody fragment serves as the targeting moiety. In an initial phase 1 trial in patients with advanced solid tumors, SGT-53 demonstrated tumor-specific targeting, was shown to be well tolerated, and was associated with an antitumor effect in several patients. Our preclinical studies have also demonstrated enhanced antitumor activity with the combination of SGT-53 and docetaxel. Thus, this dose-escalation trial was undertaken to assess the combination of SGT-53 and docetaxel for safety and potential efficacy in 14 advanced cancer patients. Results reveal that the combination of SGT-53 (maximum dose, 3.6 mg DNA/infusion) and docetaxel (75 mg/m(2)/infusion) was well tolerated. Moreover, clinical activity involving 12 evaluable patients was observed. Three of these patients achieved RECIST-verified partial responses with tumor reductions of -47%, -51%, and -79%. Two others had stable disease with significant shrinkage (-25% and -16%). These results support phase 2 testing of SGT-53 in combination with docetaxel.

Estradiol (E(2)) acts as a potent feedback molecule between the ovary and hypothalamic GnRH neurons, and exerts both positive and negative regulatory actions on GnRH synthesis and secretion. However, the extent to which these actions are mediated by estrogen receptors (ERs) expressed in GnRH neurons has been controversial. In this study, Single-cell RT-PCR revealed the expression of both ERalpha and ERbeta isoforms in cultured fetal and adult rat hypothalamic GnRH neurons. Both ERalpha and ERbeta or individual ERs were expressed in 94% of cultured fetal GnRH neurons. In adult female rats at diestrus, 68% of GnRH neurons expressed ERs, followed by 54% in estrus and 19% in proestrus. Expression of individual ERs was found in 24% of adult male GnRH neurons. ERalpha exerted marked G(i)-mediated inhibitory effects on spontaneous action potential (AP) firing, cAMP production, and pulsatile GnRH secretion, indicating its capacity for negative regulation of GnRH neuronal function. In contrast, increased E(2) concentration and ERbeta agonists increase the rate of AP firing, GnRH secretion, and cAMP production, consistent with ERbeta-dependent positive regulation of GnRH secretion. Consonant with the coupling of ERalpha to pertussis toxin-sensitive G(i/o) proteins, E(2) also activates G protein-activated inwardly rectifying potassium channels, decreasing membrane excitability and slowing the firing of spontaneous APs in hypothalamic GnRH neurons. These findings demonstrate that the dual actions of E(2) on GnRH neuronal membrane excitability, cAMP production, and GnRH secretion are mediated by the dose-dependent activation of ERalpha and ERbeta expressed in hypothalamic GnRH neurons.

The G protein-coupled receptor 54 (GPR54) and its endogenous ligand, kisspeptin, are essential for activation and regulation of the hypothalamic-pituitary-gonadal axis. Analysis of RNA extracts from individually identified hypothalamic GnRH neurons with primers for GnRH, kisspeptin-1, and GPR54 revealed expression of all three gene products. Also, constitutive and GnRH agonist-induced bioluminescence resonance energy transfer between Renilla luciferase-tagged GnRH receptor and GPR54 tagged with green fluorescent protein, expressed in human embryonic kidney 293 cells, revealed heterooligomerization of the two receptors. Whole cell patch-clamp recordings from identified GnRH neurons showed initial depolarizing effects of kisspeptin on membrane potential, followed by increased action potential firing. In perifusion studies, treatment of GT1-7 neuronal cells with kisspeptin-10 increased GnRH peak amplitude and duration. The production and secretion of kisspeptin in cultured hypothalamic neurons and GT1-7 cells were detected by a specific RIA and was significantly reduced by treatment with GnRH. The expression of kisspeptin and GPR54 mRNAs in identified hypothalamic GnRH neurons, as well as kisspeptin secretion, indicate that kisspeptins may act as paracrine and/or autocrine regulators of the GnRH neuron. Stimulation of GnRH secretion by kisspeptin and the opposing effects of GnRH on kisspeptin secretion indicate that GnRH receptor/GnRH and GPR54/kisspeptin autoregulatory systems are integrated by negative feedback to regulate GnRH and kisspeptin secretion from GnRH neurons.