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Rotamers are stereoisomers produced by rotation (twisting) about σ bonds and are often rapidly interconverting at room temperature. Xylitol—massively produced sweetener—(2R,3r,4S)-pentane-1,2,3,4,5-pentol) forms rotamers from the linear conformer by rotation of a xylitol fragment around the C2–C3 bond (rotamer 1) or the C3–C4 bond (rotamer 2). The rotamers form two distinguishable structures. Small differences in geometry of rotamers of the main carbon chain were confirmed by theoretical calculations; however, they were beyond the capabilities of the X-ray powder diffraction technique due to the almost identical unit cell parameters. In the case of rotamers of similar compounds, the rotations occurred mostly within hydroxyl groups likewise rotations in L-arabitol and D-arabitol, which are discussed in this work. Our results, supported by theoretical calculations, showed that energetic differences are slightly higher for rotamers with rotations within hydroxyl groups instead of a carbon chain.

Two new square planar ONNO nickel(II) complexes C2_core and C3_core have been synthesized and characterized by single crystal X-ray diffraction, NMR spectroscopy, thermogravimetry, and DFT calculations. The experimental results revealed the effect of the length of diamine bridge in the ligand on the behavior of the studied complexes in the reaction with N-heterocyclic aromatic amines, while DFT calculations provided a basis for the rationalization of this observation. The complex with propylenediamine bridge (C3_core) readily reacts with pyridine and its derivatives to form high-spin (paramagnetic) complexes with octahedral geometry as characterized by X-ray diffraction; electron-donating substituents on the pyridine ring facilitate the coordination of axial ligands. In contrast, the complex with ethylenediamine bridge (C2_core) does not undergo such a reaction because of the high deformation energy of the core required for the formation of C2_Py complex.

A new series of highly biologically active (20S,22R)-1α,25-dihydroxy-22-methyl- 2-methylene-vitamin D3 analogs, possessing different side chains, have been efficiently prepared as potential agents for medical therapy. Design of these synthetic targets was based on the analysis of the literature data and molecular docking experiments. The synthetic strategy involved Sonogashira coupling of the known A-ring dienyne with the C,D-ring enol triflates, obtained from the corresponding Grundmann ketones. All synthesized vitamin D compounds were characterized by high in vitro potency and, moreover, they proved to be very calcemic in vivo exerting high activity on bone with particularly elevated intestinal calcium transport.

New and more efficient routes of chemical synthesis of vitamin D(3) (D(3)) hydroxy (OH) metabolites, including 20S(OH)D(3), 20S,23S(OH)(2)D(3) and 20S,25(OH)(2)D(3), that are endogenously produced in the human body by CYP11A1, and of 20S,23R(OH)(2)D(3) were established. The biological evaluation showed that these compounds exhibited similar properties to each other regarding inhibition of cell proliferation and induction of cell differentiation but with subtle and quantitative differences. They showed both overlapping and differential effects on T-cell immune activity. They also showed similar interactions with nuclear receptors with all secosteroids activating vitamin D and aryl hydrocarbon receptors in functional assays and also as indicated by molecular modeling. They functioned as substrates for CYP27B1 with enzymatic activity being the highest towards 20S,25(OH)(2)D(3) and the lowest towards 20S(OH)D(3). In conclusion, defining new routes for large scale synthesis of endogenously produced D(3)-hydroxy derivatives by pathways initiated by CYP11A1 opens an exciting era to analyze their common and differential activities in vivo, particularly on the immune system and inflammatory diseases.

Continuing our search for vitamin D analogues, we explored the modification of the steroidal side chain and inserted a methylene moiety in position C-22 together with either lengthening the side chain or introducing a ring at the terminal end. Our conformational studies confirmed that the presence of a methylene group attached to C-22 restricts the conformational flexibility of the side chain, which can result in changes in biological characteristics of a molecule. All synthesized 1α,25-dihydroxy-2,22-dimethylene-19-norvitamin D3 analogues proved equal to calcitriol in their ability to bind to the vitamin D receptor, and most of them exert significantly higher differentiation and transcriptional activity than calcitriol. The most active compounds were characterized by the presence of an elongated side chain or 26,27-dimethylene bridge. The synthetic strategy was based on the Wittig-Horner coupling of the known A-ring phosphine oxide with the corresponding Grundmann ketones prepared from a 20-epi-Inhoffen-Lythgoe diol derived from vitamin D2.

Continuing our studies aimed at A-ring modified vitamin D compounds, we designed novel 19-norcalcitriol derivatives bearing at C-2 pegylated chains of different lengths. The terminal fragments of these substituents contain hydroxyls or moieties possessing nitrogen and/or sulfur atoms capable of transition metal ions complexation. Also, two conjugate-type platinum(II) complexes of 19-norcalcitriol were obtained in which l-methionine served as chelating moiety. The convergent synthesis of the target 19-norcalcitriol analogs involved several steps with the crucial one being condensation of A-ring phosphine oxide and the known Grundmann ketone by Wittig-Horner reaction. Further elaboration of the 2-alkylidene substituent provided all final compounds which were then tested to determine their affinity for the vitamin D receptor and cytotoxic activity.

Herein, a homologous series of cholesterol-based liquid crystalline bent-shaped dimers with different lengths of a terminal alkoxy chain were synthesized and characterized by polarizing optical microscopy, differential scanning calorimetry, and powder and single-crystal X-ray diffraction. The photosensitivity of the mesogenic molecules provides an opportunity to manipulate the structure of the liquid crystalline phase using light. The compound with the shortest alkoxy tail exhibited the heliconical twist-bend nematic phase. Elongation of the terminal alkoxy chain prevented the formation of the Ntb phase; for compounds with longer tails, instead of the Ntb phase, SmA was observed, which was rather unique for bent-shaped dimeric mesogens. The crystal structures presented herein shed new light on the possible molecular conformations and arrangements of molecules in the liquid crystalline phase. We have shown that in addition to the model of intercalated molecules, special attention should be paid to the model of strongly bent molecules. A new model of the molecular packing of bent-shaped dimers with flexible linkers in the SmA phase is proposed.

Hormonally active vitamin D3 metabolite, calcitriol, plays an important role in calcium-phosphate homeostasis, immune system actions and cell differentiation. Although anticancer activity of calcitriol is well documented and thousands of its analogs have been synthesized, none has been approved as a potential drug against cancer. Therefore, we attempted to introduce the cytotoxic effect to the calcitriol molecule by its linking to cisplatin. Herein, we present the synthesis of vitamin D compounds, designed on the basis of molecular modeling and docking experiments to the vitamin D receptor, and characterized by the presence of significantly different two side chains attached to C-20. In this study, a new synthetic approach to Gemini analogs was developed. Preparation of the target 19-norcalcitriol compounds involved separate syntheses of several building blocks (the A-ring, C/D-rings and side-chain fragments). The convergent synthetic strategy was used to combine these components by the different coupling processes, the crucial one being Wittig-Horner reaction of the Grundmann ketone analog with the known 2-methylene A-ring phosphine oxide. Due to the nature of the constructed steroidal side chains (bidentate ligands), which allowed coordination of metal ions, the first conjugate-type platinum(II) complexes of the vitamin D analogs were also successfully prepared and characterized. The target vitamin D compounds, displaying significant affinity for a vitamin D receptor, were assessed in vitro for their anti-proliferative activities towards several cell lines.

Prompted by results of molecular modeling performed on the seco-d-ring-vitamins D, we turned our attention to such analogs, having reversed configurations at C-13 and C-14, as the next goals of our studies on the structure-activity relationship for vitamin D compounds. First, we developed an efficient total synthesis of the "upper" C/seco-d-ring fragment with a 7-carbon side chain. Then, we coupled it with A-ring fragments using Sonogashira or Wittig-Horner protocol, providing the targeted D-seco analogs of 1α,25-dihydroxyvitamin D3 and 1α,25-dihydroxy-19-norvitamin D3 possessing a vinyl substituent at C-14 and a double bond between C-17 and C-20. The affinities of the synthesized vitamin D analogs to the full-length recombinant rat VDR were examined, as well as their differentiating and transcriptional activities. In these in vitro tests, they were significantly less active compared to 1α,25-(OH)2D3. Moreover, it was established that the analogs tested in vivo in rats showed no calcemic potency.

As a continuation of our efforts directed to the structure-activity relationship studies of vitamin D compounds, we present in this paper the synthesis of new analogues of 1α,25-(OH)2D3 characterized by numerous structural modifications, especially a cleaved D ring. Total synthesis of the CD fragment required for the construction of the target vitamins was based on the Stork approach. The structure of the key intermediate - bicyclic hydroxy lactone - was established by crystallographic and electronic circular dichroism (ECD) spectral analysis. Following the attachment of the hydroxyalkyl side chain, the formed D-seco Grundmann ketone was subjected to Wittig-Horner coupling with the corresponding A-ring phosphine oxides providing two desired D-seco analogues of 19-nor-1α,25-(OH)2D3, one without a substituent at C-2 and the other possessing a 2-exomethylene group. Both compounds were biologically tested and the latter was found to be more active in in vitro tests. Despite so many structural changes introduced in its structure, the biological activity of the 2-methylene analogue approached that of the natural hormone. The synthesized D-seco vitamins, however, proved to be inactive on bone and intestine in vivo.

A series of bile acid derived 1,2- and 1,3-diamines as well as their platinum(II) complexes were designed and synthesized in hope to get a highly cytotoxic compound by the combination of two bioactive moieties. All complexes obtained were subjected to cytotoxicity assays in vitro and some hybrid molecules showed an expected activity.

The application of 3D NMR experiments and DFT calculations enabled the structure investigation of C-17 epimer of 3-(25-hydroxycholest-5-enyl) acetate is presented. The H-17 and H-20 protons features the same values of 1H chemical shift, what causes that the structure elucidation require additional resolution enabled by 3D NMR experiments. The NMR experiments and theoretical calculations allowed for: the resonance assignment (3D COSY-HMBC and 3D TOCSY-HSQC techniques), the prediction of spatial structure (3D NOESY-HSQC and 3D ROESY-HSQC experiments), and the precise measurement of heteronuclear coupling constants (3D HSQC-TOCSY spectra with E.COSY-type multiplets).

The results presented in this paper constitute an extension of our synthetic efforts focused on 19-norvitamin D compounds possessing elongated 2-alkylidene substituents. Based on our previous results, molecular modeling studies, and docking experiments, we selected a novel 19-norcalcitriol analog with long chain at C-2 containing several ether moieties and terminated by 2-(pyridin-2′-yl)ethylamino fragment. It was expected that such structural modification might allow binding of transition metal by the ligand, increase solubility of the formed complexes as well as improve their affinity to the VDR. For comparison, a 19-norcalcitriol analog was also obtained with the terminal hydroxyl group at its pegylated 2-alkylidene substituent. The synthesis of the target vitamin D compounds described in this work was performed using the Wittig-Horner approach. The respective A-ring phosphine oxide was obtained starting from the D-(-)-quinic acid and then coupled with the known Grundmann ketone.

Continuing the structure-activity relationship studies in the vitamin D area, we designed and synthesized novel C-9 substituted calcitriol analogues, possessing different nonpolar groups at this position. 9α-Methyl-1α,25-(OH)2D3, both epimers of 9-methylene-10,19-dihydro-1α,25-(OH)2D3 as well as the parent vitamin with the "reversed" triene system, 9-methylene-19-nor-1α,25-(OH)2D3, were obtained from the previtamin D precursors, constructed by either Suzuki-Miyaura, Sonogashira, or Stille couplings of the corresponding A- and C,D-ring fragments. An alternative synthetic path, leading to the latter vitamin and its homologue with 9-ethylidene group, involved formation of dienynes as precursors of the respective 19-norprevitamin D compounds. 9β-Methyl-19-nor-1α,25-(OH)2D3 was prepared by homogeneous hydrogenation with Wilkinson catalyst, and this analogue was found to be the most active in vitro. Moreover, 9α-methyl-1α,25-(OH)2D3 and 9-methylene-19-nor-1α,25-(OH)2D3 showed some in vitro activity, however, the in vivo assays indicated only weak calcemic potency of these compounds in the intestinal calcium transport.

Continuing our structure-activity studies on the vitamin D analogs with the altered intercyclic seco-B-ring fragment, we designed compounds possessing dienyne system conjugated with the benzene D ring. Analysis of the literature data and the docking experiments seemed to indicate that the target compounds could mimic the ligands with a good affinity to the vitamin D receptor (VDR). Multi-step synthesis of the C/D-ring building block of the tetralone structure was achieved and its enol triflate was coupled with the known A-ring fragments, possessing conjugated enyne moiety, using Sonogashira protocol. The structures of the final products were confirmed by NMR, UV and mass spectroscopy. Their binding affinities for the full-length human VDR were determined and it was established that compound substituted at C-2 with exomethylene group showed significant binding to the receptor. This analog was also able to induce monocytic differentiation of HL-60 cells.

On the basis of the literature data, our previous research work and docking experiments, we designed novel 19-norvitamin D compounds having elongated 2-alkylidene substituents. These 19-norcalcitriol derivatives have attached 2-(3'-aminopropylidene) substituent in which the nitrogen atom bears acyl residue derived from succinic acid and l-methionine. Both compounds were obtained by the same synthetic strategy involving Julia coupling of the A-ring ketone with the known C/D-ring sulfone. In the obtained 1α,25-dihydroxy-19-norvitamin D3 derivative, the alkylidene substituent at C-2 was further elaborated to the desired structures.

The 20R- and 20S-isomers of 25-hydroxy-2-methylene-vitamin D3 and 3-desoxy-1α,25-dihydroxy-2-methylene-vitamin D3 have been synthesized. Two alternative synthetic routes were devised for preparation of the required A-ring synthons, starting from the chiral compound derived from the (-)-quinic acid and, alternatively, from the commercially available achiral precursor, monoprotected 1,4-cyclohexanedione. The A-ring dienynes were coupled by the Sonogashira process with the respective C,D-ring fragments, the enol triflates derived from the protected (20R)- or (20S)-25-hydroxy Grundmann ketones. All four compounds possessed significant in vivo activity on bone calcium mobilization and intestinal calcium transport. The presence of a 2-methylene group increased intestinal calcium transport activity of all four analogues above that of the native hormone, 1α,25-dihydroxyvitamin D3. In contrast, bone calcium mobilization was equal to that produced by 1α,25-dihydroxyvitamin D3 in compounds having a (20S)-configuration or diminished to one-tenth that of 1α,25-dihydroxyvitamin D3 in compounds with a (20R)-configuration.

9-Alkylidene analogs of 19-nor-1α,25-(OH) 2 D 3 were synthesized, possessing a ‘reversed’ triene system compared to the natural hormone. The conjugated triene moiety of the novel analogs was constructed by coupling an enyne anion, representing an A-ring synthon, with a 9α-substituted Grundmann ketone derivative. Regioselective dehydration followed by semihydrogenation under Lindlar conditions, provided the desired 9-alkylated 19-norprevitamins which were thermally isomerized to the corresponding 9-methylene and 9-ethylidene analogs of 19-norcalcitriol. It was established that only the former compound had significant binding affinity to the full-length recombinant rat vitamin D receptor. The remaining in vitro studies show very low activity of both analogs. This article is part of a Special Issue entitled ‘Vitamin D Workshop’.

We have recently obtained 1-desoxy and 3-desoxy analogs of (20S)-1α,25-dihydroxy-2-methylene-19-norvitamin D3 (2MD), a compound exerting significantly enhanced calcemic activity and currently being evaluated as a potential drug for osteoporosis. In order to further explore this class of pharmacologically important vitamin D compounds we have decided to synthesize analogs characterized by the presence of two A-ring exocyclic methylene groups attached to C-2 and C-10. The Sonogashira coupling of a triflate enol of the protected (20R)- or (20S)-25-hydroxy Grundmann ketone and the corresponding dienyne A-ring fragment provided the target compounds. A new synthetic path was elaborated, leading to the desired A-ring synthon, that started from commercially available 1,4-cyclohexanedione monoethylene acetal. Biological in vitro and in vivo activities of the synthesized 25-hydroxy-2-methylene-vitamin D3 compounds, belonging to 20R- and 20S-series, were evaluated and discussed. This article is part of a Special Issue entitled 'Vitamin D Workshop'.

The steroid hormone 1α,25-dihydroxyvitamin D 3 [1α,25-(OH) 2 D 3 ] is the most active metabolite of vitamin D 3 which exerts its control over a multitude of biological processes related to calcium and phosphorus homeostasis, cell proliferation and differentiation, and immune regulation. Unfortunately, the therapeutic application of 1α,25-(OH) 2 D 3 is limited by induction of hypercalcemia. The need for vitamin D compounds with selective biological profiles has stimulated the synthesis of more than three thousand analogs of 1α,25-(OH) 2 D 3 . Most of these compounds have structural modifications in the side chain and A-ring; there is also an increasing number of modifications in the CD-rings and limited number in the triene system (seco-B ring). Herein, we report the synthesis and biological evaluation of seco-A-19-nor analogs of 1α,25-dihydroxyvitamin D 3 , developed to study the role of ring A in the biological activity of 1α,25-(OH) 2 D 3 . Interestingly, compounds 2 and 4 show substantial ability to bind the vitamin D receptor and the former is also characterized by selective intestinal calcium transport activity. This article is part of a Special Issue entitled ‘Vitamin D Workshop’.

As a continuation of our studies on structure–activity relationship of vitamin D compounds we synthesized new calcitriol analogs characterized by the presence of an exomethylene substituent at C-2. The A-ring dienyne synthon was prepared from commercially available quinic acid by two different synthetic routes, and it was then coupled with the triflate enol derived from the corresponding (20 R )- and (20 S )-Grundmann ketone by palladium catalyzed Sonogashira reaction. The obtained 1α,25-dihydroxy-2-methylene-vitamin D 3 analogs, epimeric at C-20, were biologically evaluated by in vitro and in vivo studies. Both isomers exhibited unique activity profiles and greater biological potency than 1α,25-(OH) 2 D 3 . It was established that the biological profiles of the newly obtained vitamin D compounds depend on the configuration at C-20. Thus, introduction of 2-methylene substituent to the calcitriol molecule together with alteration of stereochemistry of its side chain induces remarkable changes in a VDR-mediated signaling response and enhances biological activity. This article is part of a Special Issue entitled ‘Vitamin D Workshop’.

As a continuation of our efforts directed to the structure-activity relationship studies of vitamin D compounds, we present in this paper the synthesis of new analogues of 1α,25-(OH)

As a continuation of our studies focused on the vitamin D compounds lacking the C,D-hydrindane system, 13,13-dimethyl-des-C,D analogues of (20S)-1α,25-dihydroxy-2-methylene-19-norvitamin D3 (2, 2MD) were prepared by total synthesis. The known cyclohexanone 30, a precursor of the desired A-ring phosphine oxide 11, was synthesized starting with the keto acetal 13, whereas the aldehyde 12, constituting an acyclic ‘upper’ building block, was obtained from the isomeric esters 34, prepared previously in our laboratory. The commercial 1,4-cyclohexanedione monoethylene ketal (13) was enantioselectively α-hydroxylated utilizing the α-aminoxylation process catalyzed by l-proline, and the introduced hydroxy group was protected as a TBS, TPDPS, and SEM ether. Then the keto group in the obtained compounds 15–17 was methylenated and the allylic hydroxylation was performed with selenium dioxide and pyridine N-oxide. After separation of the isomers, the newly introduced hydroxy group was protected and the ketal group hydrolyzed to yield the corresponding protected (3R,5R)-3,5-dihydroxycyclohexanones 30–32. The esters 34, starting compounds for the C,D-fragment 12, were first α-methylated, then reduced and the resulted primary alcohols 36 were deoxygenated using the Barton-McCombie protocol. Primary hydroxy group in the obtained diether 38 was deprotected and oxidized to furnish the aldehyde 12. The Wittig–Horner coupling of the latter with the anion of the phosphine oxide 11, followed by hydroxyl deprotection furnished two isomeric 13,13-dimethyl-des-C,D analogues of 2MD (compounds 10 and 42) differing in configuration of their 7,8-double bond. Pure vitamin D analogues were isolated by HPLC and their biological activity was examined. The in vitro tests indicated that, compared to the analogue 7, unsubstituted at C-13, the synthesized vitamin D analogue 10 showed markedly improved VDR binding ability, significantly enhanced HL-60 differentiation activity as well as increased transcriptional potency. Docking simulations provided a rational explanation for the observed binding affinity of these ligands to the VDR. Biological in vivo tests proved that des-C,D compound 10 retained some intestinal activity. Its geometrical isomer 42 was devoid of any biological activity.

As a continuation of our efforts directed to vitamin D compounds of promising biological properties, 19-norvitamins 9-13, possessing a 3'-hydroxypropylidene fragment attached to C-2 and shortened 17β-alkyl chains, were synthesized. A new synthetic pathway providing the CD-ring ketones 20-24 is described starting from the epimeric aldehydes 25 and 26. The hydrindanones 20-24 were subjected to the Wittig-Horner reaction with the phosphine oxide 14, and the vitamin D compounds 9-13 were obtained after hydroxyl deprotection. In comparison to 1α,25-(OH)(2)D(3) (1), the prepared analogues, except for the 20R-compound 12, were only ca. 3 times less potent in binding to VDR. Compounds 9-11 and 13 exhibited HL-60 cellular activity 5-20 times lower and transcriptional activity ca. 10 times decreased related to those for the hormone 1. When tested in vivo, all the analogues showed no ability to mobilize calcium from bone, and intestinal calcium transport activity was observed only at high doses of the vitamins 10, 12, and 13.

Among more than 3000 analogs of 1α,25-dihydroxyvitamin D3 synthesized to date, only a few were characterized by structural modifications in the seco-B-ring. The compounds alkylated at C-6 seemed to be interesting targets for synthetic efforts. Such vitamin D analogs easily undergo thermal conversion to their previtamin forms. The results of molecular modeling indicate that significant deviation from planarity must be present in their molecules associated with the interaction of the 6-alkyl substituent and hydrogens from the C-ring. The synthesis of the analog of 1α,25-(OH)2D3, being characterized by the presence of the 6-methyl group, is reported here, together with the results of preliminary testing of its biological potency. This 6-alkylated compound was efficiently prepared using a novel stereoconvergent strategy in which the ring A and the triene unit of the vitamin D skeleton are constructed by a one-pot Pd-catalyzed tandem cyclization-Negishi coupling process. © 2010 Elsevier Ltd.

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.

The 18-nor (7), 21-nor (8) and 18,21-dinor (9) analogs of (20S)-1alpha,25-dihydroxy-2-methylene-19-norvitamin D(3) (6, 2MD) were prepared by convergent syntheses. The known phosphine oxide 10 was coupled by the Wittig-Horner process with the corresponding C,D-fragments (13-15), obtained by a multi-step procedure from commercial vitamin D(2). The goal of our studies was to examine the influence of removal of the methyl groups located at carbons 13 and 20 on the biological potency of 2MD in the hope of finding analogs with improved therapeutic profiles. Replacement of the 20-methyl with hydrogen in 2-methylene-19-nor-(20S)-1alpha,25-dihydroxyvitamin D(3) (2MD) did not affect binding to the rat vitamin D receptor and had little effect on transcription activity and on HL-60 differentiation. However, the mobilization of calcium from bone was largely eliminated while intestinal calcium transport remained strong. Curiously, removal of both the C-13-methyl and 20-methyl restored slightly the bone calcium mobilizing activity. Thus, the 21-nor analog of 2MD may provide a potent analog with a greater margin of safety than 2MD.

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.

Analogs of (20S)-1alpha,25-dihydroxy-2-methylene-19-norvitamin D(3) (2, 2MD), substituted at C-13 but lacking both C and D rings, were prepared in convergent syntheses, starting with the chiral ester 14 and the phosphine oxide 29. Two of the synthesized vitamins (11 and 32) were analogs in which the 13-methyl group constituted a substituent of an unsaturated fragment, that is, C(13)-C(17) double bond, whereas in the two other cases (12 and 13), the methyl group belonged to a ternary carbon stereogenic center. The aim of these studies was to further explore extensive modifications in the 'upper' part of the vitamin D skeleton in the hope of finding biologically active analogs of potential therapeutic value. The commercial (R)-(-)-methyl-3-hydroxy-2-methylpropionate (14) was converted in six steps to alcohol 18, the vitamin D side chain fragment. Its subsequent three-step transformation led to aldehyde 20 which was subjected to the Still-Gennari HWE reaction with anion derived from ester 21. The obtained alpha,beta-unsaturated esters 22 and 23 served as convenient starting compounds to the syntheses of the corresponding chiral acyclic aldehydes, beta,gamma-unsaturated (28) and saturated (39 and 40), required for the final Wittig-Horner coupling with the anion of the phosphine oxide 29. After hydroxyl deprotection, the synthesized vitamin D analogs 11-13 and 32 were purified and biologically tested. Only the (13R,20S)-analog 12 retained substantial, although 30 times lower than 1alpha,25-(OH)(2)D(3), binding ability to the full-length rat recombinant vitamin D receptor (VDR). This analog was also very effective in differentiation of HL-60 cells, and it exerted significant transcriptional activity (2 times and 15 times less potent, respectively, as compared to the native hormone). The in vivo tests showed that all synthesized vitamin D analogs were devoid of calcemic activity.

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.

Vitamin D analogs are being developed that retain therapeutic effects but are less calcemic and phosphatemic, a concern in CKD patients who are prone to vascular calcification. We tested a new analog of vitamin D, 2MbisP, and found that it suppresses PTH at doses that do not affect serum Ca or P. Introduction: Calcitriol is used for the treatment of secondary hyperparathyroidism. However, its use is often limited by the development of hypercalcemia and hyperphosphatemia, an important consideration in patients with chronic kidney disease (CKD) because they are prone to vascular calcification. To minimize this toxicity, structural modifications in the vitamin D molecule have led to the development of calcitriol analogs with selective actions. Materials and Methods: In this study, we compared the effects of 1,25(OH)2D3 and a new analog, 2-methylene-19-nor-(20S)-1α-hydroxy-bishomopregnacalciferol (2MbisP), on the development of secondary hyperparathyroidism and established secondary hyperparathyroidism in uremic rats and on mobilization of calcium and phosphorus from bone in parathyroidectomized rats. The clearance from circulation, half-life, and binding affinities to the vitamin D–binding protein and vitamin D receptor of this compound were also evaluated. Results: Uremia produced a marked rise in plasma PTH, but treatment every other day for 2 wk with either 1,25(OH)2D3 (4 ng) or 2MbisP (250, 750, 1500, or 3000 ng) suppressed this increase by >50%. The suppression by 1,25(OH)2D3, however, was accompanied by increases in ionized calcium, phosphorus, and the calcium × phosphorus product, whereas these three parameters were unchanged by 2MbisP. The binding affinity of 2MbisP was 10–20 times less for the vitamin D receptor and 1000 times less for the serum vitamin D–binding protein compared with 1,25(OH)2D3. Also, 2MbisP was cleared more rapidly from the circulation (t½ = 10 min) than 1,25-(OH)2D3 (t½=7–9 h). In parathyroidectomized rats fed calcium-or phosphorus-deficient diets, daily injections of 2MbisP (1500 or 3000 ng), unlike 1,25(OH)2D3 (50 ng), had no effect on calcium or phosphorus mobilization from bone. Conclusions: In uremic rats, 2MbisP can suppress PTH at doses that do not affect plasma calcium, phosphorus, and calcium × phosphorus product. This new vitamin D analog may represent an important tool in the treatment of secondary hyperparathyroidism in patients with CKD.

The UV-induced photochemical reaction of 1α,25-dihydroxy-9-methylene-19-norvitamin D3 has been investigated. The pentacyclic structure of the isolated product has been unequivocally established by X-ray crystallographic analysis. The possible reaction paths of the examined photochemical transformation are discussed. Biological in vivo and in vitro tests proved that the photoproduct is devoid of calcemic activity.

In pursuit of novel biologically active Vitamin D compounds of potential therapeutic value, 1alpha,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 1alpha,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.

Since the discovery of the active metabolite of vitamin D, i.e., 1α,25-dihydroxyvitamin D 3 , there has been a continuous effort to synthesize analogs able to carry out many of the functions of the native hormone without raising serum calcium concentration. The present report provides a series of previously undescribed analogs wherein this goal is realized. We have prepared 2-methylene-19-nor-1α-hydroxyvitamin D analogs of 1,25-(OH) 2 D 3 that possess only two to four carbons of the side chain without a hydroxyl thereon. Compared to 1,25-(OH) 2 D 3 , these analogs are slightly less active in binding to the vitamin D receptor, in causing HL-60 differentiation, and are slightly less active in in vitro transcription assays using the 24-hydroxylase promoter attached to a luciferase reporter gene. Of considerable importance is that these analogs, given to rats at daily doses of up to 70 μg/kg of body weight per day, are either unable or only slightly able to raise serum calcium concentration but are nevertheless able to suppress parathyroid hormone levels in plasma up to 100% and induce 24-hydroxylase mRNA in skin. Because of their ability to act in vivo without raising serum calcium levels, they may be of considerable interest for the systemic treatment of diseases such as psoriasis, cancer, and secondary hyperparathyroidism of renal failure, where a rise in serum calcium is undesirable.

B3LYP calculations in conjunction with natural bond orbital population analysis have been performed for a previtamin D model and corresponding transition structures for the [1,7]-hydrogen migration. In addition the 19,19-difluoro, 19-methoxy and 19-fluoro substituted analogs were investigated. The calculated activation barriers decrease in the following order: CHF2>CH3>CH2OCH3 (24.8, 23.5 and 20.1 kcal/mol). This is in qualitative agreement with experiments. It has been suggested that a decrease of the barrier by a 19-methoxy substituent and its increase by a 19,19-difluoro substituent are phenomena of different origin. In the case of 19-methoxy substitution, the effect is due to the charge redistribution in the triene system and the decrease of the C(19)–H bond energy. The effect of two fluorine substituents at C-19 on the activation barrier is suggested to originate from the combination and balance of several factors: electrostatic repulsion between the negative fluorine atom and the π-electron cloud over the conjugated system, an increase of the HOMO–LUMO gap, and geminal difluoro substitution affecting C–F and C–C bond energies.

The aryl hydrocarbon receptor (AHR) is a ligand-inducible transcription factor that is best known because it mediates the actions of polycyclic and halogenated aromatic hydrocarbon environmental toxicants such as 3-methylcholanthrene and 2,3,7,8-tetrachlorodibenzo- p -dioxin. We report here the successful identification of an endogenous ligand for this receptor; ≈20 μg was isolated in pure form from 35 kg of porcine lung. Its structure was deduced as 2-(1′H-indole-3′-carbonyl)-thiazole-4-carboxylic acid methyl ester from extensive physical measurements and quantum mechanical calculations. In a reporter gene assay, this ligand activates the AHR with a potency five times greater than that of β-naphthoflavone, a prototypical synthetic AHR ligand. 2-(1′H-indole-3′-carbonyl)-thiazole-4-carboxylic acid methyl ester competes with 2,3,7,8-[ 3 H]tetrachlorodibenzo- p -dioxin for binding to human, murine, and fish AHRs, thus showing that AHR activation is caused by direct receptor binding, and that recognition of this endogenous ligand is conserved from early vertebrates (fish) to humans.

Novel 19-nor analogues of 1alpha,25-dihydroxyvitamin D(3) were prepared and substituted at C-2 with an ethylidene group. The synthetic pathway was via Wittig-Horner coupling of the corresponding A-ring phosphine oxides with the protected 25-hydroxy Grundmann's ketones. Selective catalytic hydrogenation of 2-ethylidene analogues provided the 2alpha- and 2beta-ethyl compounds. The 2-ethylidene-19-nor compounds with a methyl group from the ethylidene moiety in a trans relationship to the C(6)-C(7) bond (E-isomers) were more potent than the corresponding Z-isomers and the natural hormone in binding to the vitamin D receptor. Both geometrical isomers (E and Z) of (20S)-2-ethylidene-19-norvitamin D(3) and both 2alpha-ethyl-19-norvitamins (in the 20R- and 20S-series) have much higher HL-60 differentiation activity than does 1alpha,25-(OH)(2)D(3). Both E-isomers (20R and 20S) of 2-ethylidene vitamins are characterized by very high calcemic activity in rats. The three-dimensional structure model of the rat vitamin D receptor and the computational docking of four synthesized (20R)-19-norvitamin D(3) analogues into its binding pocket are also reported.

Two novel vitamin D analogues of the hormone 1α,25-(OH)2D3 modified at C-7, namely, 7-methyl-1α,25-(OH)2D3 (12) and 7-methyl-1α,25-(OH)2-19-nor-D3 (26), were synthesized and biologically evaluated to gain further insights into the structure-function relationships of vitamin D. Key steps in the synthesis of 12 include the functionalization at C-7 by an efficient regioselective hydrostannylation of an allene precursor, and the construction of the triene framework by a palladium-catalyzed intramolecular cyclization-Suzuki-Miyaura coupling cascade. Since the calcitriol analogue 12 was prone to conversion into its previtamin D form by thermal equilibration, the corresponding 19-nor-compound 26 was also synthesized. The diene moiety of compound 26 was constructed by a modified Julia coupling. UV data as well as X-ray analysis indicate that introduction of the methyl group at C-7 results in a significant deviation from planarity of the 5,7-diene moiety. The new vitamin D analogues 12 and 26 retained good VDR binding ability.

7α- and 7β-Methyl derivatives of cholest-5-en-3β-ol (cholesterol), cholest-4-en-3-one and cholest-4-en-3,6-dione were prepared. Methylation of 3α,5-cyclo-5α-cholestan-6-one at C-7 was followed by stereoselective reduction with LiAlH 4 . The key transformation was cycloreversion of the mixture of 7α-methyl-3α,5-cyclo-5α-cholestan-6α-ol and 7β-methyl-3α,5-cyclo-5α-cholestan-6β-ol. The reaction of the latter was much faster under the standard conditions enabling easy separation of the C-7 isomers.

The synthesis of provitamin D analogue 19-(phenylsulfonyl)androsta-5,7-diene-3β,17β-diyl 3-acetate 17-pivalate (20) has been accomplished from 19-hydroxyandrost-5-ene-3β,17β-diyl 3-acetate 17-pivalate. 19-(Phenylsulfonyl)androst-5-ene-3β,17β-diyl 3-acetate 17-pivalate (10), a precursor of 20, was first obtained in low yield in the nucleophilic displacement reactions of 19-halogenated-5-ene steroids with sodium benzenesulfinate. Then a more efficient method has been used, which involves protection of the double bond as an epoxide. Introduction of the C(7)-C(8) double bond into olefin 10 has been also achieved in two ways. The first involved bromination-dehydrobromination and the other consisted of an allylic oxidation of olefin 10 leading to enone and the Bamford-Stevens reaction of its tosylhydrazone. UV irradiation of 5,7-diene 20 resulted in formation of a complex mixture of products. The structures of five isolated compounds were established on the basis of their 1H NMR spectra and mechanistic rationale.

A simple method for the synthesis of yet unknown 5E-vitamin D3 analogs with an additional six-membered ring connecting C-6 and C-19 was developed. Ring-closing metathesis (RCM) was used for efficient formation thereof from the corresponding 5E-isomers of 6-alkenyl vitamin D3 compounds which in turn were prepared from the 6-oxo-3,5-cyclovitamin D3. Reinvestigation of the Grignard reactions of this latter compound as well as the following acid-catalyzed cycloreversions showed discrepancies with the literature data describing the course of such processes.

This article describes efficient preparation of isomeric allyl phosphine oxides possessing a protected cyclohexanediol fragment. Their base-catalyzed interconversions are examined and reactions with the Grundmann ketone provide an adduct containing the rearranged vinyl phosphine oxide moiety, instead of 19-norvitamin D3 analogs, the expected products of the Horner–Wittig process.

3-Desoxy-20-oxo-19-nor-pregnacaliferol (6a) and its 2-oxo analog 6c were prepared from 22-acetoxy-Grundmann's ketone 1 and the corresponding (cyclohexylidene)ethyl phosphine oxides 2a, b and their binding to the progesterone receptor examined.

Triol diester 1 was converted into the B-ring 5,7-diene 6 representing the first example of the provitamin D analog where the 10β angular methyl group is connected to ring C by a C11/C19 ether linkage. Ultraviolet light irradiation of 6 resulted in the formation of stereoisomeric 9β, 10α-compound 8a. The structure of the photoproduct was established by analysis of vicinal 1H–1H coupling constants and by molecular mechanics. Keywords: 11β, 19-oxido steroids, lumisterol analog, 5,7-androstadienes, electrocyclic reactions, photochemistry.

Two A-ring stereoisomeric 1-hydroxy-6,7-diaza-19-norvitamin D 3 compounds 7a , 8a and their 25-hydroxylated analogs, 7c , 8c have been prepared starting with enantiomeric trans-3,5-dihydroxycyclohexanones 3c,d and the CD-ring 8-ketones 4a and 4c . 1,25-Dihydroxy azavitamins 7c and 8c are essentially devoid of biological activity, demonstrating the importance of the 5,7-diene structure of vitamin D.

The reaction of Grundmann's ketone 2a with ruthenium tetroxide, generated in situ from ruthenium trichloride and sodium metaperiodate, has been investigated. In addition to the main 25-hydroxylated compound 2b the minor oxidation products have been isolated and identified. The examined catalytic process results in the oxidation of methylene groups and hydroxylation at teriary positions occurring with retention of configuration.

We have recently described the synthesis and biological evaluation of 1α,25-dihydroxy-6-methylvitamin D 3 which binds the vitamin D receptor (VDR) very effectively. Unfortunately, this compound has a strong tendency to rearrange to its previtamin form. Taking this into consideration, we decided to synthesize a series of 6-substituted analogs of 1α,25-dihydroxy-19-norvitamin D 3 which lack the exomethylene moiety at C-10 and thus are unable to undergo a conversion to their respective previtamin forms. The synthesis of analogs bearing different substituents at C-6 was accomplished by Suzuki–Miyaura cross-coupling reaction of a bicyclic organoboron derivative (CD-ring fragment) with the respective alkenyl halides (A-ring synthons). The biological in vitro studies of the affinity of synthesized analogs to the full-length recombinant rat VDR indicate that presence of a bulky, polar substituent at C-6 results in decrease in binding ability. Moreover, introduction of a 6-methyl substituent into the 1α,25-dihydroxy-19-norvitamin D 3 molecule results in the 9 times lower affinity of the homolog to the VDR while the same modification of calcitriol has not influenced binding activity. This article is part of a Special Issue entitled ‘Vitamin D Workshop’.