Structural comparison of protiated, H/D-exchanged and deuterated human carbonic anhydrase IX.

Human carbonic anhydrase IX (CA IX) expression is upregulated in hypoxic solid tumours, promoting cell survival and metastasis. This observation has made CA IX a target for the development of CA isoform-selective inhibitors. To enable structural studies of CA IX-inhibitor complexes using X-ray and neutron crystallography, a CA IX surface variant (CA IXSV; the catalytic domain with six surface amino-acid substitutions) has been developed that can be routinely crystallized. Here, the preparation of protiated (H/H), H/D-exchanged (H/D) and deuterated (D/D) CA IXSV for crystallographic studies and their structural comparison are described. Four CA IXSV X-ray crystal structures are compared: two H/H crystal forms, an H/D crystal form and a D/D crystal form. The overall active-site organization in each version is essentially the same, with only minor positional changes in active-site solvent, which may be owing to deuteration and/or resolution differences. Analysis of the crystal contacts and packing reveals different arrangements of CA IXSV compared with previous reports. To our knowledge, this is the first report comparing three different deuterium-labelled crystal structures of the same protein, marking an important step in validating the active-site structure of CA IXSV for neutron protein crystallography.

Do surface-active lipids in food increase the intestinal permeability to toxic substances and allergenic agents?

The incidence of many common diseases has increased during the last decades. High fat intake is a risk factor for many diseases. We propose that some of the negative effects of fat are caused by lipid-induced damage of the gastrointestinal epithelium, thus compromising the epithelial function as a barrier for passage of toxic substances and allergenic agents to the circulatory system. Monoglycerides (MGs), phospholipids and fatty acids (FAs) are surface-active molecules that in pharmaceutical studies act as permeability enhancers for hydrophilic drugs with low absorption. Three possible mechanisms were proposed: (a) lipid-induced alterations in intracellular events may cause destabilization of tight junctions between the GI epithelial cells, (b) lipids may destabilize cell membranes, (c) lipids cause intestinal cell damage, which increase the permeability of the GI epithelium. These "side effects" of lipids may partly explain the association between fat intake and disease observed in epidemiological studies.

Treatment of severe psoriasis with etretin (RO 10-1670).

Eighty patients with severe psoriasis were treated in a double-blind fashion with either an initial dose of 10 mg, 25 mg or 50 mg of etretin daily or with placebo. Follow-up examinations were carried out monthly and the efficacy of treatment was evaluated by using the PASI score. Adverse effects of the treatment were recorded monthly; liver enzymes, cholesterol and triglycerides were measured. After 2 months of treatment the maintenance dose was reduced in some of the patients either because of complete remission or adverse effects. After 2 months treatment, groups receiving 25 mg/day and 50 mg/day showed significantly lower PASI scores than the placebo group. The 10 mg/day group showed a response intermediate between the 25 mg and 50 mg groups and the placebo group. Thus, the optimal initial dose seems to be approximately 25 mg/day and the maintenance dose somewhat lower. Six months after the start of treatment there were no significant differences between the four groups; the last follow-up examination took place during the summer and some of the patients probably experienced spontaneous improvement. Although clinical adverse effects were frequent in all groups, severe side effects, namely hair loss and paronychia, occurred frequently only among patients treated with an initial dose of 50 mg of etretin daily. The effect of treatment on liver enzymes, cholesterol and triglycerides was minimal.