Maximilian Grassberger

Discovery of topical calcineurin inhibitors and pharmacological profile of pimecrolimus

Using a newly developed model of allergic contact dermatitis in pigs, calcineurin inhibitors of the tacrolimus and ascomycin type were shown to have a highly anti-inflammatory action after topical application. These findings provided the first pharmacological evidence of the efficacy of this novel class of topical agents in the treatment of inflammatory skin diseases, and, thus, their potential to become the first alternative to corticosteroids in more than 40 years. As a result of a large research program into ascomycins, pimecrolimus (Elidel®, SDZ ASM 981) was selected for development due to its favorable pharmacology and safety profile, alongside tacrolimus (Protopic®, FK 506). In vitro, pimecrolimus inhibits the transcription and release of pro-inflammatory cytokines in T cells. Similar to the corticosteroids, betamethasone-17-valerate and dexamethasone, pimecrolimus is effective at nanomolar concentrations. Targeting mainly T cells, pimecrolimus has, however, a more specific mode of action. Moreover, in contrast to corticosteroids, pimecrolimus has no effect on Langerhans’ cells, the professional antigen- presenting dendritic cells of the skin that are crucial for local immunosurveillance. When applied topically, pimecrolimus exerts a high and selective anti-inflammatory activity in the skin, shows minimal percutaneous absorption, and has a low potential to affect systemic immunoreactions. Pimecrolimus cream 1% has proven to be well tolerated, safe, and highly effective in clinical studies in patients with atopic dermatitis.

Synthesis and oxidative cleavage of the major equilibrium products of ascomycin and FK 506

Abstract Ascomycin and FK 506 are converted into the 9-ketal isomers in high yields by simple treatment with Lewis acids. The structure of the 9-ketal isomer was confirmed by oxidative cleavage.

Efficacy and safety of pimecrolimus cream 1% in adult patients with vitiligo: results of a randomized, double-blind, vehicle-controlled study.

Background: Vitiligo is an acquired, pigmentary skin disorder which is disfiguring and difficult to treat. In an earlier open label study in adult patients with vitiligo, pimecrolimus cream 1% was reported to have similar efficacy as clobetasol propionate 0.05%. We performed a double‐blind, intrapatient comparison of pimecrolimus cream 1% with placebo cream.

Highly selective reactions of FK506 with diazomethane

Abstract Three modes of diazomethane reactivity, namely oxirane formation, O-methylation and cyclopropanation, can be accomplished with excellent selectivity on the multifunctional molecule FK506.

Isolation of an isomer of FK-506 from fermentation of Streptomyces tsukubaensis and its chemical synthesis from FK-506

Abstract From the fermentation broth of Streptomyces tsukubaensis 9993 an isomer 2a of the immunosuppressant FK-506 1a was isolated as a minor metabolite. From NMR- and FAB-MS data follows that in 2a the size of the macrocycle is reduced by two carbon atoms due to a shift of the lactone-acyl from 0–26 to 0–24. The compound was synthesized from FK-506 and proved to be identical with the natural product 2a.

PIMECROLIMUS: ABSORPTION, DISTRIBUTION, METABOLISM, AND EXCRETION IN HEALTHY VOLUNTEERS AFTER A SINGLE ORAL DOSE AND SUPPLEMENTARY INVESTIGATIONS IN VITRO

The absorption and disposition of pimecrolimus, a calcineurin inhibitor developed for the treatment of inflammatory skin diseases, was investigated in four healthy volunteers after a single oral dose of 15 mg of [3H]pimecrolimus. Supplementary information was obtained from in vitro experiments. Pimecrolimus was rapidly absorbed. After tmax (1–3 h), its blood concentrations fell quickly to 3% of Cmax at 24 h, followed by a slow terminal elimination phase (average t1/2 62 h). Radioactivity in blood decreased more slowly (8% of Cmax at 24 h). The tissue and blood cell distribution of pimecrolimus was high. The metabolism of pimecrolimus in vivo, which could be well reproduced in vitro (human liver microsomes), was highly complex and involved multiple oxidative O-demethylations and hydroxylations. In blood, pimecrolimus was the major radiolabeled component up to 24 h (49% of radioactivity area under the concentration-time curve0–24 h), accompanied by a large number of minor metabolites. The average fecal excretion of radioactivity between 0 and 240 h amounted to 78% of dose and represented predominantly a complex mixture of metabolites. In urine, 0 to 240 h, only about 2.5% of the dose and no parent drug was excreted. Hence, pimecrolimus was eliminated almost exclusively by oxidative metabolism. The biotransformation of pimecrolimus was largely catalyzed by CYP3A4/5. Metabolite pools generated in vitro showed low activity in a calcineurin-dependent T-cell activation assay. Hence, metabolites do not seem to contribute significantly to the pharmacological activity of pimecrolimus.

SYNTHETIC MODIFICATIONS OF ASCOMYCIN. I: A CHEMOSELECTIVE REMOVAL OF THE CYCLOHEXYL RESIDUE OF ASCOMYCIN

Abstract An efficient semisynthetic preparation of des-28-(cyclohexyl)methylene-28-oxo-ascomycin derivatives starting from 24,33- O -bis( tert -butyldimethylsilyl)-ascomycin ( 1 ) is described. The strategy for preparing 28-oxo-ascomycin derivatives involves the reduction of C-22 carbonyl group, followed by 5-endo-cyclization of the resulting C-22 alcohol with the C-19/C-20 double bond using an oxymercuration reaction; ozonolysis of the C-28/C-29 double bond and regeneration of the C-19/C-20 double bond. Further, the 20-mercury-substituted ascomycin derivatives could be reduced to the corresponding metal free cyclic ethers using n -Bu 3 SnH.

C9-Imino and C10-amino derivatives of ascomycin (21-ethyl-FK 506)

Abstract Treatment of Ascomycin (21-ethyl-FK 506) with ammonia in alcoholic solution yeilds C9-imine 5 and C10-amine 6 , whereas methylamine gives only C9-methylimine 10 .

CARBONYL TO METHYLENE CONVERSIONS AT THE TRICARBONYL-PORTION OF ASCOMYCIN DERIVATIVES

Abstract Treatment of ascomycin or its O-TBDMS-derivatives with hydrogen sulfide and pyridine in dimethylformamide solution results in deoxygenation reactions at the tricarbonyl sequence of the binding domain. 9-deoxo-ascomycins ( 5a-c ) are obtained in high yields (75–85%) together with small amounts (2–14% yield) of 10-deoxo-ascomycins ( 6a-c ). The novel derivative 10-deoxo-ascomycin ( 6a ) is accessible in excellent yield (85%) from the 10-amino-analog of ascomycin upon reaction with hydrogen sulfide in the absence of base.

Regioselective Carbon–Carbon Bond Formation in the Effector Domain of Ascomycin

Abstract Starting from 33- O -silyl-protected ascomycin or its 23,24-dehydration product, regio- and stereo-selective formal aldol reactions under carbon–carbon bond formation at C-22, C-23, or C-24 are demonstrated. Additionally, with the 22-silyl enol ether ascomycin derivative a rhodium(I)-catalysed shift of the 19,20 double bond into the 19,38 exocyclic position is observed at elevated temperature.