Transgenic mice expressing a mutant form of loricrin reveal the molecular basis of the skin diseases, Vohwinkel syndrome and progressive symmetric erythrokeratoderma

Mutations in the cornified cell envelope protein loricrin have been reported recently in some patients with Vohwinkel syndrome (VS) and progressive symmetric erythrokeratoderma (PSEK). To establish a causative relationship between loricrin mutations and these diseases, we have generated transgenic mice expressing a COOH-terminal truncated form of loricrin that is similar to the protein expressed in VS and PSEK patients. At birth, transgenic mice (ML.VS) exhibited erythrokeratoderma with an epidermal barrier dysfunction. 4 d after birth, high-expressing transgenic animals showed a generalized scaling of the skin, as well as a constricting band encircling the tail and, by day 7, a thickening of the footpads. Histologically, ML. VS transgenic mice also showed retention of nuclei in the stratum corneum, a characteristic feature of VS and PSEK. Immunofluorescence and immunoelectron microscopy showed the mutant loricrin protein in the nucleus and cytoplasm of epidermal keratinocytes, but did not detect the protein in the cornified cell envelope. Transfection experiments indicated that the COOH-terminal domain of the mutant loricrin contains a nuclear localization signal. To determine whether the ML.VS phenotype resulted from dominant-negative interference of the transgene with endogenous loricrin, we mated the ML.VS transgenics with loricrin knockout mice. A severe phenotype was observed in mice that lacked expression of wild-type loricrin. Since loricrin knockout mice are largely asymptomatic (Koch, P.K., P. A. de Viragh, E. Scharer, D. Bundman, M.A. Longley, J. Bickenbach, Y. Kawachi, Y. Suga, Z. Zhou, M. Huber, et al., J. Cell Biol. 151:389-400, this issue), this phenotype may be attributed to expression of the mutant form of loricrin. Thus, deposition of the mutant protein in the nucleus appears to interfere with late stages of epidermal differentiation, resulting in a VS-like phenotype.     

Predicting microscale shifts in the distribution of the butterfly Plebejus argus at the northern edge of its range

Species are often observed to occur in restricted patches of particularly warm microclimate at their high latitude/altitude geographic range margin. In these areas, global warming is expected to cause small‐scale expansion of the occupied area, but most previous studies of range expansion have used very coarse scale data. Using high resolution microclimate models together with detailed field surveys, we tested whether the butterfly Plebejus argus, occurring on limestone grassland in north Wales, was responding as might be expected due to climate change in the last 30–40 yr. The abundance of adult Plebejus argus at 100 m resolution in 2011 was strongly affected by elevation and near‐ground temperatures in May. A statistical model including microclimate, fitted to 2011 data, was successful (67% correct) at hindcasting the occurrence of Plebejus argus in 1983 when the average May air temperature was 1.4°C cooler. However, the model was less accurate at hindcasting occurrences in 1972 (50% correct). Given the distribution of micro‐sites in this landscape, we predict that further warming of approximately 1°C would make the majority of sites highly microclimatically suitable for this species. There are a growing number of long‐term studies of range change, and investigations into the mechanisms driving them, but still surprisingly few that explicitly make and test predictions with independent data. Our tests are a valuable example of how accurate predictions of distribution change can be, but also of the inevitable uncertainties. Improved understanding of how well models predict will be very important to plan robust climate change adaptation measures.     

Releasable PEGylation of mesothelin targeted immunotoxin SS1P achieves single dosage complete regression of a human carcinoma in mice

Recombinant immunotoxins exhibit targeting and cytotoxic functions needed for cell-specific destruction. However, antitumor efficacy, safety, and pharmacokinetics of these therapeutics might be improved by further macromolecular engineering. SS1P is a recombinant anti-mesothelin immunotoxin in clinical trials in patients with mesothelin-expressing tumors. We have modified this immunotoxin using several PEGylation strategies employing releasable linkages between the protein and the PEG polymers, and observed superior performance of these bioconjugates when compared to similar PEG derivatives bearing permanent linkages to the polymers. PEGylated derivatives displayed markedly diminished cytotoxicity on cultured mesothelin-overexpressing A431-K5 cells; however, the releasable PEGylated immunotoxins exhibited increased antitumor activity in A431-K5 xenografts in mice, with a diminished animal toxicity. Most significantly, complete tumor regressions were achievable with single dose administration of the bioconjugates but not the native immunotoxin. Pharmacokinetic analysis of the releasable PEGylated derivatives in mice demonstrated an over 80-fold expansion of the area under the curve exposure of bioactive protein when compared to native immunotoxin. A correlation in degree of derivatization, release kinetics, and polymer size with potency was observed in vivo, whereas in vitro cytotoxicity was not predictive of efficacy in animal models. The potent antitumor efficacy of the releasable PEGylated mesothelin-targeted immunotoxins was not exhibited by similar untargeted PEG immunotoxins in this model. Since the bioconjugates can also exhibit the attributes of passive targeting via enhanced permeability and retention, this is the first demonstration of a pivotal role of active targeting for immunotoxin bioconjugate efficacy.     

Potent achiral agonists of the ghrelin (growth hormone secretagogue) receptor. Part I: Lead identification

High throughput screening combined with efficient datamining and parallel synthesis led to the discovery of a novel series of indolines showing potent in vitro ghrelin receptor agonist activity and acceleration of gastric emptying in rats.