A high-content biosensor-based screen identifies cell-permeable activators and inhibitors of EGFR function: implications in drug discovery.

Early success of kinase inhibitors has validated their use as drugs. However, discovery efforts have also suffered from high attrition rates due to lack of cellular activity. We reasoned that screening for such candidates in live cells would identify novel cell-permeable modulators for development. For this purpose, we have used our recently optimized epidermal growth factor receptor (EGFR) biosensor assay to screen for modulators of EGFR activity. Here, we report on its validation under high-throughput screening (HTS) conditions displaying a signal-to-noise ratio of 21 and a Z' value of 0.56-attributes of a robust cell-based assay. We performed a pilot screen against a library of 6912 compounds demonstrating good reproducibility and identifying 82 inhibitors and 66 activators with initial hit rates of 1.2% and 0.95%, respectively. Follow-up dose-response studies revealed that 12 of the 13 known EGFR inhibitors in the library were confirmed as hits. ZM-306416, a vascular endothelial growth factor receptor (VEGFR) antagonist, was identified as a potent inhibitor of EGFR function. Flurandrenolide, beclomethasone, and ebastine were confirmed as activators of EGFR function. Taken together, our results validate this novel approach and demonstrate its utility in the discovery of novel kinase modulators with potential use in the clinic.

[Anti-inflammatory activity of a topical glucocorticoid, fludroxycortide tape in experimental animals (author's transl)].

Fludroxycortide tape is a thin plastic tape which contains a synthetic glucocorticoid, fludroxycortide of 4 microgram/cm2. fludroxycortide tape with a topical application of 1 cm2 inhibited significantly contact hypersensitivities to oxazolone and picryl chloride in the ear skin of mice and PCA caused by IgE-like antibodies in the depilated abdominal skin of rats. Topical application of 1 to 4 cm2 of fludroxycortide tape produced a significant inhibition against histamine-induced vascular permeability, skin edema induced by intradermal injection of carrageenin into the depilated back and ear edema induced by topical application of croton oil in rats. Topical to oxazolone in dose-dependent manner, and had no thymolytic action, while in a dose of 100 microgram/ear, thymus atrophy occurred. Ear edema induced by croton oil was markedly inhibited by topical application of fludroxycortide (0.4 microgram/ear). Thus, fludorxycortide tape has topical anti-inflammatory activity against both allergic and non-allergic inflammation in mice and rats, and the topical anti-inflammatory activity of fludroxycortide appears to be favourably dissociated from its thymolytic action.

Effects of occlusive tape systems on the mitotic activity of epidermis. With and without corticosteroids.

This study was designed to elaborate on the effects of occlusion and corticosteroids on human epidermal mitotic activity. The experimental variables included tape delivery systems that afforded total, partial (50%), or no occlusion to normal human skin. In some experiments, hydrocortisone, flurandrenolide, betamethasone, betamethasone valerate, and fluorometholone were added to these tape systems. Using demecolcine cream, biopsies were taken, and mitotic figures were determined. Results suggest that potent antimitotic effects occur with semiocclusive and nonocclusive tapes that contain corticosteroids, and that total occlusion is not necessary for this physiological effect. This suggests the possibility that the presumably better tolerated nonocclussive and partially occlusive systems might be used as dermatological drug delivery systems.

Measurement of dermal atrophy induced by topical steroids using a radiographic technique.

A modified radiographic technique was used to assess dermal atrophy induced by topical steriods. The method proved reliable, and significant atrophy occurred with fluocinolone acetonide, clobetasone butyrate, hydrocortisone 17-butyrate and flurandrenolone, but not with hydrocortisone.

A separation of clinical from epidermal thinning effect in the topical glucocorticoid clobetasone butyrate.

Clobetasone butyrate ointment has been shown to be more effective in treating psoriasis and eczema than flurandrenolone ointment yet to cause less epidermal thinning in a human experimental model. This is an indication that the clinical activity of topical glucocorticoids may not necessarily be inseparable from their propensity to cause atrophy of the skin.

The effect of clobetasone butyrate and other topical steroids on skin thickness of the domestic pig.

A new glucocorticoid, clobetasone butyrate, has been shown in patients to have good topical anti-inflammatory activity and a minimal effect on hypothalamic-pituitary-adrenal function. Among a group of topically active corticosteroids, compared in a controlled study in the domestic pig, clobetasone butyrate is shown to cause less epidermal thinning than any of the others except only 1% hydrocortisone. This evidence of a lesser atrophogenic effect may indicate further dissociation of unwanted from wanted properties in clobetasone butyrate.

Experimentally induced steroid atrophy in the domestic pig and man.

A histological method of ranking topical steroid preparations according to the degree of epidermal thinning they cause in pig skin has been shown to be applicable to human skin. A 0-05% ointment of the new compound, clobetasone butyrate (Molivate), has been shown to cause less thinning than either fluocinolone acetonide 0-025% (Synalar) or flurandrenolone 0-0125% (Haelan) when applied without the use of occlusive dressings for 3 weeks to the human forearm.

Steroid-induced atrophy in an animal and human model.

A ratchet-controlled micrometer screw gauge has been used to measure the changes in skin thickness produced by topical corticosteroids. Measurable thinning of the mouse ear followed repeated daily application of corticosteroids. This response in the animal was used to assess the atrophic potential in a variety of standard formulations. The same compounds were applied to human skin and the changes in thickness measured. Patterns of response and the rank order of thinning obtained from the animal model were found to parallel closely the human results.

Flurandrenolone tape. A new preparation for occlusive therapy.

A new variation of plastic film occlusion is described, using a plastic tape into the adhesive side of which flurandrenolone has been incorporated. In a series of 10 patients with psoriasis, the efficacy of the tape is compared in a controlled manner with that of plastic film wrapping. The results with the tape are at least as good as, and probably better than, the results with the plastic film wrapping. The noteworthy finding is that all 10 of the patients treated preferred the tape to the plastic wrap because of the tape's greater convenience.