Recent studies have shown that there is a considerable heterogeneity in the response of melanoma cell lines to MEK and BRAF inhibitors. In the current study, we address whether dysregulation of cyclin-dependent kinase 4 (CDK4) and/or cyclin D1 contribute to the BRAF inhibitor resistance of melanoma cells. Mutational screening identified a panel of melanoma cell lines that harbored both a BRAF V600E mutation and a CDK4 mutation: K22Q (1205Lu), R24C (WM39, WM46, and SK-Mel-28), and R24L (WM902B). Pharmacologic studies showed that the presence of a CDK4 mutation did not alter the sensitivity of these cell lines to the BRAF inhibitor. The only cell line with significant BRAF inhibitor resistance was found to harbor both a CDK4 mutation and a CCND1 amplification. Array comparative genomic hybridization analysis showed that CCND1 was amplified in 17% of BRAF V600E -mutated human metastatic melanoma samples, indicating the clinical relevance of this finding. As the levels of CCND1 amplification in cell lines are lower than those seen in clinical specimens, we overexpressed cyclin D1 alone and in the presence of CDK4 in a drug-sensitive melanoma line. Cyclin D1 overexpression alone increased resistance and this was enhanced when cyclin D1 and CDK4 were concurrently overexpressed. In conclusion, increased levels of cyclin D1, resulting from genomic amplification, may contribute to the BRAF inhibitor resistance of BRAF V600E -mutated melanomas, particularly when found in the context of a CDK4 mutation/ overexpression. [Mol Cancer Ther 2008;7(9):2876 -83]

Background: In gnathostomes, chemosensory receptors (CR) expressed in olfactory epithelia are encoded by evolutionarily dynamic gene families encoding odorant receptors (OR), trace amineassociated receptors (TAAR), V1Rs and V2Rs. A limited number of OR-like sequences have been found in invertebrate chordate genomes. Whether these gene families arose in basal or advanced vertebrates has not been resolved because these families have not been examined systematically in agnathan genomes.

BackgroundVertebrate pheromones are known to prime the endocrine system, especially the hypothalamic-pituitary-gonadal (HPG) axis. However, no known pheromone molecule has been shown to modulate directly the synthesis or release of gonadotropin releasing hormone (GnRH), the main regulator of the HPG axis. We selected sea lamprey (Petromyzon marinus) as a model system to determine whether a single pheromone component alters the output of GnRH.Sea lamprey male sex pheromones contain a main component, 7α, 12α, 24-trihydroxy-5α-cholan-3-one 24-sulfate (3 keto-petromyzonol sulfate or 3kPZS), which has been shown to modulate behaviors of mature females. Through a series of experiments, we tested the hypothesis that 3kPZS modulates both synthesis and release of GnRH, and subsequently, HPG output in immature sea lamprey.ResultsThe results showed that natural male pheromone mixtures induced differential steroid responses but facilitated sexual maturation in both sexes of immature animals (χ2 = 5.042, dF = 1, p < 0.05). Exposure to 3kPZS increased plasma 15α-hydroxyprogesterone (15α-P) concentrations (one-way ANOVA, p < 0.05) and brain gene expressions (genes examined: three lamprey (l) GnRH-I transcripts, lGnRH-III, Jun and Jun N-terminal kinase (JNK); one-way ANOVA, p < 0.05), but did not alter the number of GnRH neurons in the hypothalamus in immature animals. In addition, 3kPZS treatments increased lGnRH peptide concentrations in the forebrain and modulated their levels in plasma. Overall, 3kPZS modulation of HPG axis is more pronounced in immature males than in females.ConclusionsWe conclude that a single male pheromone component primes the HPG axis in immature sea lamprey in a sexually dimorphic manner.

Many environmental pollutants induce expression of the cytochrome P450 (CYP) 1A subfamily of genes. We integrated cellular and molecular biological techniques to examine the effects of β-naphthoflavone (BNF) exposure in lake trout brain CYP1A distribution and dynamics. Over a 32-day time-course, real time quantitative reverse transcription polymerase chain reaction (Q-RT-PCR) results showed that CYP1A mRNA induction in response to BNF exposure occurred rapidly and continued to rise in the BNF-treated lake trout after 4·h, with a peak at or after 2·days. Messenger RNA levels fell after 4·days, and this trend continued after 16·days of exposure. In situ hybridization indicated that CYP1A mRNA was universally elevated in the brain of BNF-exposed fish and was mainly expressed in the endothelia and occasionally in the glial cells. CYP1A immunoreactivity was induced in the olfactory bulb and valvula cerebelli of BNF-treated fish. Other brain areas showed constitutive CYP1A immunoreactivity in both control and BNF-treated fish. Some BNF-treated fish contained multifocal hemorrhages in the brain tissue, and these fish had overall depressed CYP1A immunoreactivity in the brain. The relationship between transcriptional and translational effects of BNF exposure in the brain of juvenile lake trout is discussed.