Since the discovery of 1α,25-dihydroxyvitamin D3 [1α,25-(OH)2D3, calcitriol, 1; Fig. 1] as an active form of vitamin D3 resulting from its two sequential hydroxylations in liver and kidney [1], there has been a rapid development of studies on this compound and its derivatives [2]. Intensive research efforts carried out at academic institutions and industries resulted in the synthesis of more than three thousand of its structural analogs with modified biological potencies [3]. It is well-established that the biological actions of calcitriol, commonly referred to as the vitamin D hormone [4], are primarily regulation of calcium and phosphorous homeostasis. It was proved that the vast majority of the biological actions of 1 are realized through vitamin D receptor (VDR)-mediated regulation of the gene expression [5]. Thus, the important genes regulated in such manner are also possibly responsible for control of immunomodulation [6], inflammation [7] and oxidation as well as cancer cells growth arresting, differentiating and apoptosis [8]. The established presence of the receptor and its ligand in most of the human tissues [9] clearly indicates that the role of this hormone in the living organisms must be significantly broader and its further exploration is desirable. The fact that vitamin D compounds can have important applications in new areas has been confirmed by numerous findings during the COVID-19 epidemic. Clinical studies strongly suggested that supplementation with calcitriol [10] and its analogs [11] played an important role in counteracting this disease and was associated with the improvement of survival in patients with COVID-19.

VDR, belonging to the nuclear receptor superfamily [12], binds calcitriol with high affinity and forms a heterodimer with the retinoid X receptor (RXR). After binding to a specific DNA motif, the DNA-bound VDR/RXR heterodimer recruits coregulators [13]. When calcitriol or its agonists are present in the complexed VDR, its conformational change allows (or prevents) interaction with coregulators and further transcriptional processes [14]. Taking into account high pharmaceutical potency of the calcitriol analogs, the knowledge of their interactions with the amino acids lining the ligand-binding domain (LBD) of the VDR seemed to be of significant importance; it could facilitate the discovery of the vitamin D compounds showing high cell differentiation potency and similar affinity for VDR as calcitriol but with decreased (undesirable in this case) calcemic action.

Important information regarding the action of the vitamin D analogs could be obtained via X-ray diffraction analysis of their crystal complexes with VDR. The human VDR LBD complexed with the natural hormone 1 [15], has already been known for two decades and its solved crystal structure can be considered as one of the breakthroughs in the area of vitamin D studies. The crystal structure of hVDR-1 complex (PDB: 1DB1) shows the important hydrogen bonds anchoring the three hydroxyl groups of the ligand to the LBD: 1α-OH interacts with Ser-237 and Arg-274, 3β-OH with Tyr-143 and Ser-278, whereas the side chain 25-OH with His-305 and His-397. As it can be expected from the structure of the vitamin D ligand, it is surrounded in the LBD mainly by hydrophobic amino acids. Not surprisingly, the ring D also interacts (4 Å cutoff) with hydrophobic residues, such as: Leu233, Val234, Ile268, Ile271, Met272, Trp286 and Leu313.

Since then, VDR has also been crystallized with a significant number of other synthetic calcitriol analogs and to-date more than 180 such complexes have been described [16]. It was found that the position of helix 12 of the LBD is of critical importance for the agonistic, antagonistic and nonagonistic conformation of the VDR. The structural details of the binding domain allow researchers to design new vitamin D analogs with improved pharmacological properties.

To design vitamin D analogs with novel modification of the carbon skeleton of 1α,25-(OH)2D3, we have turned our attention to the enlargement of the ring D. Analysis of the literature reveals that this structural change has already attracted some attention of researchers half a century ago [17] but it has been limited to vitamin D compounds with a six-membered ring D. Four C/D-ring modified trans-decalin 1α,25-dihydroxyvitamin D3 analogs 4–7 were synthesized by De Clercq et al., in 2003 [18]. Except compound 6, all analogs were characterized by high binding ability to the VDR, comparable to calcitriol. (20R)-Analogs 4 and 5 proved to be calcemic and exhibited significantly higher cell differentiation activity in comparison with their (20S)-counterparts 6 and 7. Thus, the presence of a trans-decalin C/D-ring system represents one of the rare examples where the 20-epi-vitamin D compounds show a reduced biological activity when compared to their epimers with the natural side-chain configuration. Moreover, the synthesis of the vitamin D compounds 8–11 possessing a cyclohexane ring D but lacking the ring C was reported [19]. The same research group also prepared other des-C,D-homo analogs possessing double bonds in the ring D [20], or different alkyl substituents [21]. It was found that such skeletal modifications generally lead to compounds of decreased calcemic potency.

The studies on D-homo vitamin D analogs described in the literature also include compounds with an aromatic ring D. The first report on such class of compounds was published by Mourino et al., in 2010 [22]; an analog 12 had a strong tendency to undergo isomerization to its previtamin form. Eight years later, highly active and noncalcemic analogs 13–17, possessing an aromatic m-phenylene ring D and an alkyl substituent at C-8, were synthesized by the same group [23].

Taking into account all the data presented above, we decided to synthesize calcitriol analogs with a seven-membered ring D to check the effect of such D-ring enlargement on the biological activity of vitamin D compounds and their interaction with the VDR.