Agnieszka Glebocka

New 1α,25-Dihydroxy-19-norvitamin D3 Compounds Constrained in a Single A-Ring Conformation: Synthesis of the Analogues by Ring-Closing Metathesis Route and Their Biological Evaluation

Vitamin D compounds possessing A rings prohibited from flipping to the alternative chair form (i.e., analogues 2 and 26) were synthesized. The bicyclic fragment 22 consisting of the fused cyclohexane and dihydropyran rings was constructed via the ring-closing metathesis route. Also, a homologous synthon 23 with an attached dihydropyran ring was successfully synthesized using this strategy. The carbonyl deprotection in 22 yielded cyclohexanone 5 that was subjected to Julia coupling with the anion of the phenylthiazoline sulfone 25. In the resulting isomeric 19-norvitamins 2 and 26, their A rings can exist only in the alpha- and beta-conformation. The analogue 26 was 300 times more active in binding to the vitamin D receptor protein, 30 times more effective in causing HL-60 differentiation, and 10 times more active in transcription. These results confirm that the beta-chair form of the vitamin D ring A is necessary for the binding to the receptor.

Design, Synthesis, and Biological Evaluation of a 1α,25-Dihydroxy-19-norvitamin D3 Analogue with a Frozen A-Ring Conformation

To establish the conformation of vitamin D compounds responsible for biological activity, a 1alpha,25-dihydroxy-19-norvitamin D analogue 4 possessing a 1alpha-hydroxy group fixed in the axial orientation (beta-chair form) was synthesized. The starting compounds were bicyclic lactones 6, 7a, and 7b, derived from the quinic acid lactone, which were converted to the bicyclic ketone 13. Julia coupling of this compound with sulfone 15 produced the 19-norvitamin D analogue 4, possessing an additional ring connecting the 3beta-oxygen and C-2, and the isomeric 3beta-hydroxy compound 5. In vitro, both analogues 4 and 5 exhibit reduced activity compared to the natural hormone 1, but the binding, differentiation, and transcriptional activities of analogue 5 are markedly higher than that of 4 constrained in the alpha-chair conformation. Surprisingly, in vivo tests in mice showed that the analogue 4 significantly increases serum calcium at dose levels similar to 1alpha,25-(OH)2D3. These seemingly discordant results are discussed.

New derivative of 1α,25-dihydroxy-19-norvitamin D3 with 3′-alkoxypropylidene moiety at C-2: synthesis, biological activity and conformational analysis

Abstract In pursuit of novel biologically active Vitamin D compounds of potential therapeutic value, 1α,25-dihydroxy-2-[3′-(methoxymethoxy)propylidene]-19-norvitamin D 3 ( 7 ) was efficiently prepared in a convergent synthesis, starting with (−)-quinic acid and the protected 25-hydroxy Grundmann ketone 16 . The key synthetic step involved Lythgoe type Wittig–Horner coupling of 16 , with the phosphine oxide 15 . Molecular modeling was employed to establish the A-ring conformation of the synthesized Vitamin 7 . Also, preliminary modeling of its complex with the rVDR was performed and interactions between ligand and the binding domain analyzed. Analog 7 was found to be only six times less potent than 1α,25-(OH) 2 D 3 ( 1 ) in binding to the rat recombinant Vitamin D receptor (VDR). In comparison with hormone 1 , it also showed slightly lower cellular HL-60-differentiation activity. Preliminary in vivo tests indicated unusually high calcemic activity of 7 .

A-ring analogs of 1,25-dihydroxyvitamin D3

The growing interest in1α,25(OH)(2)D(3), the hormonally active form of vitamin D(3), has prompted numerous efforts to synthesize vitamin D analogs as potential therapeutic agents, and some of these are already on the market and in clinical development. Although most vitamin D preparations developed thus far have focused on side-chain modifications, providing many useful analogues with high potency and selectivity, in recent years, modifications of the A-ring has attracted much attention because it can afford useful analogues exhibiting unique activity profiles as well. In this review we will focus on the current understanding of the relationship between selected modifications in the A-ring of the 1α,25(OH)(2)D(3) molecule, such as epimerization and/or substitution at C-1 and C-3, substitution at C-2, and removal of the 10,19-exocyclic methylene group, and their effect on biological potency and selectivity. Finally, suggestions for the structure-based design of therapeutically valuable A-ring vitamin D analogs will conclude the review.

An analog of 1α,25-dihydroxy-19-norvitamin D3 with the 1α-hydroxy group fixed in the axial position lacks biological activity in vitro

Abstract The relationship between the A-ring chair conformation of vitamin D compounds and their ability to bind the vitamin D receptor (VDR) has long attracted the attention of many researchers. It was established that in the crystalline complexes of hVDRmt with the natural hormone, 1α,25-dihydroxyvitamin D 3 ( 1 ), and its side-chain analogs the vitamins exist in β-chair form with an equatorial orientation of 1α-OH. However, with all these ligands the interconversion between both A-ring forms would be possible in solution. In an attempt to verify the conformation of vitamin D compounds required for binding the VDR we prepared analog 4 , characterized by the presence of an axial 1α-hydroxy group. Since the additional ring connecting 3β-oxygen and C-2 prevents A-ring conformational flexibility, the synthesized vitamin 4 can exist exclusively in the α-chair form. The geometrical isomer 5 with a free 3β-OH group was also obtained. The analog 5 binds very poorly to VDR, whereas the vitamin 4 is practically devoid of binding ability. Both compounds also show very low HL-60-differentiating activity. When tested in vivo in mice the analogs 4 and 5 exhibit significant calcemic responses with analog 4 showing more activity than analog 5 .

New 1α,25-dihydroxy-19-norvitamin D3 analogs with a frozen A-ring conformation☆

We have recently described the synthesis of 1alpha,25-dihydroxy-19-norvitamin D3 analogs 2 and 3, possessing an additional ring connecting their 3beta-oxygen and C-2. Such structural constrains prevent the A-ring conformational flexibility and the analogs exist exclusively in the alpha-chair form with their 1alpha-hydroxy groups fixed in the axial position. The analogs bind very poorly to vitamin D receptor and are devoid of transcriptional activity. Rather unexpectedly, when tested in vivo in rats, they exhibited calcemic response significantly delayed compared to 1alpha,25-dihydroxyvitamin D3 (1). Such a response might be due to the metabolic conversion (ether cleavage?) of these compounds in the living organisms. It was therefore of interest to obtain and evaluate biologically the analogous compounds having an additional ring of purely hydrocarbon nature. Such analog 4 of 1alpha,25-dihydroxy-19-norvitamin D3, characterized by the presence of an equatorial 1alpha-hydroxy group (beta-chair form), has been synthesized by us and tested biologically. The geometrical isomer 5 having a fixed 3beta-hydroxy group was also obtained. These compounds were formed in the Julia coupling of the sulfone derived from the Grundmann ketone, and the A-ring fragment prepared in the multi-step synthesis from the (-)-quinic acid. Contrary to its counterpart 5, the analog 4 retained some affinity to vitamin D receptor.