Anti-inflammatory and anti-invasive effects of α-melanocyte-stimulating hormone in human melanoma cells

Alpha-melanocyte stimulating hormone (alpha-MSH) is known to have pleiotrophic functions including pigmentary, anti-inflammatory, antipyretic and immunoregulatory roles in the mammalian body. It is also reported to influence melanoma invasion with levels of alpha-, beta- and gamma-MSH correlated clinically with malignant melanoma development, but other studies suggest alpha-MSH acts to retard invasion. In the present study, we investigated the action of alpha-MSH on three human melanoma cell lines (HBL, A375-SM and C8161) differing in metastatic potential. alpha-melanocyte-simulating hormone reduced invasion through fibronectin and also through a human reconstructed skin composite model for the HBL line, and inhibited proinflammatory cytokine-stimulated activation of the NF-kappaB transcription factor. However, A375-SM and C8161 cells did not respond to alpha-MSH. Immunofluorescent microscopy and Western blotting identified melanocortin-1 receptor (MC-1R) expression for all three lines and MC-2R on HBL and A375-SM lines. Receptor binding identified a similar affinity for alpha-MSH for all three lines with the highest number of binding sites on HBL cells. Only the HBL melanoma line demonstrated a detectable cyclic adenosine monophosphate (cAMP) response to alpha-MSH, although all three lines responded to acute alpha-MSH addition (+(-)-N(6)-(2-phenylisopropyl)-adenosine (PIA)) with an elevation in intracellular calcium. The nonresponsive lines displayed MC-1R polymorphisms (C8161, Arg (wt) 151/Cys 151; A375-SM, homozygous Cys 151), whereas the HBL line was wild type. Stable transfection of the C8161 line with wild-type MC-1R produced cells whose invasion was significantly inhibited by alpha-MSH. From this data, we conclude that alpha-MSH can reduce melanoma cell invasion and protect cells against proinflammatory cytokine attack in cells with the wild-type receptor (HBL).Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

Mutations in or near the Transmembrane Domain Alter PMEL Amyloid Formation from Functional to Pathogenic

PMEL is a pigment cell-specific protein that forms physiological amyloid fibrils upon which melanins ultimately deposit in the lumen of the pigment organelle, the melanosome. Whereas hypomorphic PMEL mutations in several species result in a mild pigment dilution that is inherited in a recessive manner, PMEL alleles found in the Dominant white (DW) chicken and Silver horse (HoSi)—which bear mutations that alter the PMEL transmembrane domain (TMD) and that are thus outside the amyloid core—are associated with a striking loss of pigmentation that is inherited in a dominant fashion. Here we show that the DW and HoSi mutations alter PMEL TMD oligomerization and/or association with membranes, with consequent formation of aberrantly packed fibrils. The aberrant fibrils are associated with a loss of pigmentation in cultured melanocytes, suggesting that they inhibit melanin production and/or melanosome integrity. A secondary mutation in the Smoky chicken, which reverts the dominant DW phenotype, prevents the accumulation of PMEL in fibrillogenic compartments and thus averts DW–associated pigment loss; a secondary mutation found in the Dun chicken likely dampens a HoSi–like dominant mutation in a similar manner. We propose that the DW and HoSi mutations alter the normally benign amyloid to a pathogenic form that antagonizes melanosome function, and that the secondary mutations found in the Smoky and Dun chickens revert or dampen pathogenicity by functioning as null alleles, thus preventing the formation of aberrant fibrils. We speculate that PMEL mutations can model the conversion between physiological and pathological amyloid

Ultrasonographic measures of synovitis in an early phase clinical trial: a double-blind, randomised, placebo and comparator controlled phase IIa trial of GW274150 (a selective inducible nitric oxide synthase inhibitor) in rheumatoid arthritis.

OBJECTIVES: To test the sensitivity to change of ultrasonographic endpoints in early phase clinical trials in subjects with active rheumatoid arthritis (RA). METHODS: A double-blind, placebo and comparator controlled, randomised, two-centre study investigated the effect on synovial thickness and vascularity of 28 days repeat daily oral dosing of 60 mg of the inducible nitric oxide synthase inhibitor GW274150 or 7.5 mg prednisolone in RA. Fifty patients with DAS28 scores ≥4.0 were assigned to 3 treatment arms of 17, 19 and 14 (on placebo, GW274150 and prednisolone respectively). Synovial thickness and vascularity of all 10 metacarpophalangeal joints were assessed by ultrasonography using a semi-quantitative scale at baseline (Day 1), Day 15 and Day 28. Vascularity was also measured quantitatively by power Doppler area. RESULTS: At Day 28, the GW274150 group showed a trend towards reduction in synovial thickness compared with placebo, with an adjusted mean decrease of 33% (p=0.072); the prednisolone group decreased significantly by 44% (p=0.011). Similarly, there was a trend to reduced synovial vascularity with GW274150 by 42% compared with placebo (p=0.075); prednisolone resulted in a statistically significant decrease of 55% (p=0.012). There was a 55% decrease in power Doppler area for GW274150, compared with placebo although the result was not statistically significant (p=0.375). Prednisolone 7.5 mg resulted in a highly statistically significant decrease of 95% (p=0.003). CONCLUSIONS: This study advocates the use of ultrasonographic measures of metacarpophalangeal joint synovitis as an endpoint for clinical studies assessing therapeutic potential of new compounds in small patient cohorts over 28 days

Specific Targeting of Melanotic Cells with Peptide Ligated Photosensitizers for Photodynamic Therapy

Abstract A strategy combining covalent conjugation of photosensitizers to a peptide ligand directed to the melanocortin 1 (MC1) receptor with the application of sequential LED light dosage at near-IR wavelengths was developed to achieve specific cytotoxicity to melanocytes and melanoma (MEL) with minimal collateral damage to surrounding cells such as keratinocytes (KER). The specific killing of melanotic cells by targeted photodynamic therapy (PDT) described in this study holds promise as a potentially effective adjuvant therapeutic method to control benign skin hyperpigmentation or superficial melanotic malignancy such as Lentigo Maligna Melanoma (LMM)