Synthesis of dimeric analogs of adenophostin A that potently evoke Ca²⁺ release through IP₃ receptors

Abstract

Inositol 1,4,5-trisphosphate receptors (IP₃Rs) are tetrameric intracellular channels through which many extracellular stimuli initiate the Ca²⁺ signals that regulate diverse cellular responses. There is considerable interest in developing novel ligands of IP₃R. Adenophostin A (AdA) is a potent agonist of IP₃R and since some dimeric analogs of IP₃R ligands are more potent than the corresponding monomer; we considered whether dimeric AdA analogs might provide agonists with increased potency. We previously synthesized traizolophostin, in which a simple triazole replaced the adenine of AdA, and showed it to be equipotent to AdA. Here, we used click chemistry to synthesize four homodimeric analogs of triazolophostin, connected by oligoethylene glycol chains of different lengths. We evaluated the potency of these analogs to release Ca²⁺ through type 1 IP₃R and established that the newly synthesized dimers are equipotent to AdA and triazolophostin.

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Introduction

Inositol 1,4,5-trisphosphate (IP₃, 1, Fig. 1) is an important secondary messenger that evokes Ca²⁺ release from intracellular stores through its interaction with IP₃ receptors (IP₃R) in the endoplasmic reticulum.¹ IP₃R are large tetrameric proteins, within which IP₃ binding to each of the four subunits is required to initiate opening of the Ca²⁺-permeable channel.² High-resolution structures of the IP₃-binding core (IBC, residues 224–604) have defined the interactions of IP₃ with IP₃R.³ More recently, structures of the N-terminal region (residues 1–604)⁴ alongside a structure of the complete IP₃R derived from cryo-electron microscopy have begun to suggest how IP₃ binding might trigger the opening of the intrinsic pore of IP₃R.⁵

Fig. 1 The structures of IP₃ (1), adenophostin A (2) and triazolophostin (3).

There is continuing interest in the development of potent agonists and antagonists of IP₃R.⁶ The fungal metabolite, adenophostin A (AdA, 2, Fig. 1), binds to IP₃R with greater affinity than IP₃ and it is more potent than IP₃ in evoking Ca²⁺ release.⁷ AdA analogs with a nucleobase or base-surrogate are also more potent than IP₃.⁸ Molecular docking^8j,m,9 and mutation studies¹⁰ suggest that a cation–π interaction between the adenine moiety of AdA and Arg504 within the IBC contributes to the increased affinity of AdA. We recently reported synthesis of a library of active AdA analogs, triazolophostins, by using a click chemistry approach.¹¹ These potent analogs have substituted triazoles as adenine surrogates. The simplest analog, triazolophostin (3, Fig. 1) was equipotent with AdA.

Multimeric ligands often have greater affinity than monomeric ligands.¹² This can be due to simultaneous binding to more than one binding site or a statistical effect arising from the local increase in ligand concentration.¹³ The former is unlikely for IP₃R because the orientation of the IP₃-binding sites within the tetrameric IP₃R is unlikely to allow simultaneous binding of two ligands linked by a short tether.^4b,14

A few multimeric ligands of IP₃R have been reported. Before the location of the IP₃-binding sites within IP₃R was known, clustered bi- and tetra-dentate analogs of ribophostin (4, Fig. 2A) were synthesized, anticipating that if the spacing between the linked ligands was appropriate they might bind simultaneously to the four IP₃-binding sites.¹⁵ However, the potencies of the monomeric and polymeric ligands were rather similar. Several homodimeric¹⁶ and heterodimeric¹⁷ ligands of IP₃ (5–10, Fig. 2B), particularly those with short linkers, were shown to bind to IP₃R with increased affinity.^13d Very recently, dimers of 2-O-Bt-IP₄/IP₅ (11, Fig. 2C) were shown to be antagonists of IP₃Rs.¹⁸ These results demonstrate that dimeric IP₃R ligands can provide useful tools, some of which have greater affinity than the monomeric ligands. We therefore considered whether dimers of AdA might be more potent than AdA.

Fig. 2 The representative structures of (A) ribophostin dimer 4, (B) homo and hetero dimers of IP₃ (5–10) and (C) dimers of 2-Bt-IP₄/IP₅11.

Results and discussion

As the synthesis of AdA dimers is challenging, we decided to make oligoethylene glycol-tethered dimers of triazolophostin (Fig. 3). We envisaged that use of click reaction¹⁹ with a linker connected to alkyne at both termini would ensure both formation of triazole and link the two monomers in one step. Previous studies suggested that short linkers were most likely to improve the affinity of homodimers.^13d We therefore selected spacers smaller than hexaethylene glycol. The linkers 14a–d were synthesized by slightly modifying previously reported procedures.²⁰ The oligoethylene glycols were first co-evaporated with toluene and then treated with sodium hydride in the presence of excess propargyl bromide to get dipropargyl polyethylene glycols 14a–d in good to excellent yields. The azide 13 was synthesized from glucose and xylose by several protection–deprotection reactions followed by phosphorylation as reported earlier.¹¹ The azide 13 was then treated with dialkynyl polyethylene glycols 14a–d in the presence of Cu(I) catalyst to get fully protected triazolophostin dimers 15a–d in good yields. The debenzylation of protected triazolophostin dimers 15a–d was carried out using transfer hydrogenolysis in the presence of palladium and cyclohexene under reflux condition and the products were purified by ion-exchange chromatography to yield dimers 12a–d, in excellent yields (Scheme 1).

Fig. 3 The structure of dimeric analogs of triazolophostin 12.

Scheme 1 Synthesis of triazolophostin dimers. Reagents and conditions: (a) ref. 11; (b) Cu, CuSO₄, H₂O : ^tBuOH (1 : 1, v/v), rt, 24 h; (c) Pd(OH)₂/C, cyclohexene, MeOH : H₂O (10 : 1, v/v), 80 °C. 4 h; (a), n = 2; (b), n = 3; (c), n = 4; (d), n = 6.

The dimeric ligands 12a–d were screened for their abilities to evoke Ca²⁺ release through IP₃R (Table 1, Fig. 4). All four dimers were full agonists of IP₃R, more potent than IP₃, but similar in their potency to AdA and the monomer, triazolophostin. The similar potencies of 12a–d irrespective of their tether length suggest that these ligands might be interacting with IP₃R1 in monodentate fashion.

Table 1 Responses of IP₃R1 to IP₃ (1), monomer (3) and its dimeric analogs 12a–d^a

Ligand	pEC50	EC50 (nM)	EC50 w.r.t. 1^b	Max. response (%)	n H
a Maximal Ca^2+ release, the half-maximally effective ligand concentration (EC50), −log EC50 (pEC50) and Hill coefficient (nH) are shown as means ± SEM (n = 3). b The EC50 value of each ligand is also shown relative to that for IP3 (1) (EC^150/EC^analog50).
IP3 (1)	6.72 ± 0.12	190.5	1	69 ± 3	1.40 ± 0.16
Monomer (3)	7.86 ± 0.17	13.8	13.8	65 ± 1	1.66 ± 0.21
12a	7.83 ± 0.18	14.8	12.9	68 ± 2	1.33 ± 0.12
12b	7.85 ± 0.13	14.1	13.5	66 ± 1	1.89 ± 0.13
12c	7.62 ± 0.11	24.0	7.9	61 ± 3	1.60 ± 0.16
12d	7.84 ± 0.12	14.4	13.2	60 ± 1	1.94 ± 0.47

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Fig. 4 Summary of Ca²⁺ release from permeabilized DT40-IP₃R1 cells evoked by IP₃, monomer 3 and its dimeric analogs 12a–d.

Conclusions

In conclusion, based on several previous reports that dimeric IP₃R ligands can be more potent than the corresponding monomers, we anticipated that dimers of AdA might have increased potency. We used click chemistry to synthesize dimers of a potent analog of AdA (triazolophostin) linked by spacers of different length. In assays of Ca²⁺ release through IP₃R, the dimeric ligands were no more potent than the corresponding monomer (3). This suggests that whereas dimeric derivatives of IP₃ have reduced efficacy but improved affinity,^10,21 dimerization of AdA analogs does not improve their affinity.

Experimental section

General methods

The chemicals were purchased from commercial sources and used as received. The TLC plates were visualized under UV light and by dipping plates into either phosphomolybdic acid in MeOH or sulphuric acid in ethanol, followed by heating. All NMR experiments were carried out on a 500 MHz NMR spectrometer and at room temperature. Tetramethylsilane (TMS, δ 0.0 ppm) or the solvent reference (CDCl₃, δ 7.26 ppm; D₂O, δ 4.79 ppm) relative to TMS were used as the internal standard. The data are reported as follows: chemical shift in ppm (δ) (multiplicity [singlet (s), doublet (d), doublet of doublet (dd), triplet (t), quartet (q), and multiplet (m)], coupling constants [Hz], integration and peak identification). All NMR signals were assigned on the basis of ¹H NMR, ¹³C NMR, COSY and HMQC experiments. ¹³C NMR spectra were recorded with complete proton decoupling. Carbon chemical shifts are reported in ppm (δ) relative to TMS with the respective solvent resonance as the internal standard. The concentration of the compounds for ¹H NMR was 5 mg per 0.5 mL and for ¹³C NMR it was 20 mg per 0.5 mL for protected compounds and 5–7 mg per 0.5 mL for final compounds in case of ¹H and ¹³C NMR. Modified Brigg's phosphate assay²² was employed to quantify each triazolophostin 12a–d. Silica gel 230–400 mesh was used to perform flash column chromatography.

General procedure for syntheses of fully protected triazolophostin dimers

To a solution of azide 13 (0.144 mmol) and dialkynyl PEG 14a–d (0.072) in H₂O/^tBuOH (1/1, v/v, 2 mL) was added Cu (0.036 g, 0.57 mmol) and CuSO₄ (8 mg, 0.028 mmol) and stirred at room temperature for 24 h. The reaction was monitored by TLC. When the TLC showed complete disappearance of the azide 13, the mixture was filtered through a Celite bed and was partitioned between ethyl acetate and water. The organic layer was washed with brine. The organic layer was dried over anhyd. sodium sulphate, filtered and concentrated under reduced pressure. The residue thus obtained was purified by flash column chromatography using a mixture of acetone, diethyl ether and petroleum ether (4 : 2 : 15 v/v/v) as eluent to get pure 15a–d as a colourless gum.

Protected triazolophostin dimer 15a. Click reaction of azide 13 (0.2 g, 0.144 mmol) with diyne 14a (0.011 g, 0.072 mmol) gave the protected dimer 15a (0.18 g, 85%) as a colourless gum. ¹H NMR (500 MHz, CDCl₃) δ: 3.47–3.57 (m, 18H, H-2′′, H-4′′, H-6′′_A, and DEG-H), 3.73–3.75 (m, 2H, H-5′′), 4.20–4.23 (m, 2H, PhCH₂), 4.27–4.32 (m, 8H, H-5′_A and PhCH₂), 4.30–4.45 (m, 10H, H-3′, H-4′, H-5′_B, H-6′′_B and PhCH₂), 4.57–4.59 (m, 2H, PhCH₂), 4.63–4.66 (m, 4H, PhCH₂), 4.68–4.73 (m, 6H, PhCH₂), 4.80–4.93 (m, 16H, H-3′′, H-4′′ and PhCH₂), 5.11 (d, 2H, J = 3.2 Hz, H-1′′), 5.26–5.28 (m, 2H, H-2′) 6.24 (d, 2H, J = 5.0 Hz, H-1′), 7.00 (d, 4H, J = 7.0 Hz, Ar-H), 7.05–7.19 (m, 82H, Ar-H), 7.26 (d, 4H, J = 7.0 Hz, Ar-H), 7.60 (s, 2H, H-5); ¹³C NMR (125 MHz, CDCl₃) δ: 64.2, 68.3, 69.1, 69.2, 69.3, 69.5, 69.6, 69.7, 69.9, 70.1, 70.4, 71.9, 73.3, 73.5, 74.1, 78.0, 78.5, 82.8, 90.1, 95.7, 121.6, 127.6, 127.7, 127.9, 128.0, 128.1, 128.3, 128.4, 128.5, 135.2, 135.7, 135.8, 136.1, 137.3, 137.5, 138.0, 145.2; ³¹P NMR (202.4 MHz, CDCl₃) δ: −1.484, −1.928, −2.146; HRMS (ESI) mass calcd for C₁₅₈H₁₆₆N₆O₃₉P₆ [M]⁺ 2956.9616, found 2956.9620.

Protected triazolophostin dimer 15b. Click reaction of azide 13 (0.2 g, 0.144 mmol) with diyne 14b (0.016 g, 0.072 mmol) gave the protected dimer 15b (0.185 g, 86%) as a colourless gum. ¹H NMR (500 MHz, CDCl₃) δ: 3.44–3.57 (m, 22H, H-2′′, H-4′′, H-6′′_A, and TEG-H), 3.75 (bs, 2H, H-5′′), 4.21–4.30 (m, 10H, H-5′_A and PhCH₂), 4.42–4.43 (m, 10H, H-3′, H-4′, H-5′_B, H-6′′_B and PhCH₂), 4.57–4.59 (m, 2H, PhCH₂), 4.64–4.66 (m, 6H, PhCH₂), 4.68–4.73 (m, 4H, PhCH₂), 4.84–4.92 (m, 16H, H-3′′, H-4′′ and PhCH₂), 5.11 (bs, 2H, H-1′′), 5.27 (bs, 2H, H-2′) 6.24 (d, 2H, J = 5.0 Hz, H-1′), 7.00–7.25 (m, 90H, Ar-H), 7.61 (s, 2H, H-5); ¹³C NMR (125 MHz, CDCl₃) δ: 63.2, 67.3, 68.0, 68.3, 68.4, 68.6, 68.7, 69.1, 69.4, 70.9, 72.3, 75.7, 75.8, 75.9, 76.9, 77.5, 81.7, 89.0, 94.7, 120.6, 126.7, 127.0, 127.2, 127.4, 134.2, 134.6, 135.1, 136.3, 136.5, 137.0, 144.2; ³¹P NMR (202.4 MHz, CDCl₃) δ: −1.486, −1.935, −2.155; HRMS (ESI) mass calcd for C₁₆₀H₁₇₀N₆O₄₀P₆ [M]⁺ 3000.9879, found 3000.9877.

Protected triazolophostin dimer 15c. The reaction of azide 13 (0.2 g, 0.144 mmol) with diyne 14c (0.019 g, 0.072 mmol) gave the protected dimer 15c (0.175 g, 81%) as a colourless gum. ¹H NMR (500 MHz, CDCl₃) δ: 3.54–3.67 (m, 26H, H-2′′, H-4′′, H-6′′_A, and TetraEG-H), 3.84 (bs, 2H, H-5′′), 4.30–4.32 (m, 2H, PhCH₂), 4.37–4.39 (m, 8H, H-5′_A and PhCH₂), 4.48–4.53 (m, 10H, H-3′, H-4′, H-5′_B, H-6′′_B and PhCH₂), 4.66–4.68 (m, 2H, PhCH₂), 4.73–4.74 (m, 4H, PhCH₂), 4.78–4.82 (m, 6H, PhCH₂), 4.92–4.94 (m, 10H, H-3′′, H-4′′ and PhCH₂), 4.97–5.03 (m, 6H, PhCH₂), 5.20 (bs, 2H, H-1′′), 5.36 (bs, 2H, H-2′) 6.34 (d, 2H, J = 5.0 Hz, H-1′), 7.09–7.34 (m, 90H, Ar-H), 7.75 (s, 2H, H-5); ¹³C NMR (125 MHz, CDCl₃) δ: 64.2, 68.3, 69.1, 69.15, 69.2, 69.3, 69.39, 69.5, 69.5, 69.6, 69.8, 69.9, 70.4, 70.5, 70.55, 71.9, 73.3, 73.5, 76.7, 82.8, 95.7, 121.6, 127.6, 127.7, 127.78, 127.8, 127.9, 128.0, 128.1, 128.3, 128.37, 128.4, 128.5, 128.55, 128.6, 135.2, 136.1, 136.2, 137.3, 137.5, 138.0; ³¹P NMR (202.4 MHz, CDCl₃) δ: −1.468, −1.908, −2.138; HRMS (ESI) mass calcd for C₁₆₂H₁₇₄N₆O₄₁P₆ [M]⁺ 3045.0141, found 3045.0131.

Protected triazolophostin dimer 15d. The reaction of azide 13 (0.2 g, 0.144 mmol) with diyne 14d (0.026 g, 0.072 mmol) gave the protected dimer 15d (0.185 g, 82%) as a colourless gum. ¹H NMR (500 MHz, CDCl₃) δ: 3.53 (bs, 34H, H-2′′, H-4′′, H-6′′_A, and HEG-H), 3.74 (bs, 2H, H-5′′), 4.23–4.28 (m, 10H, H-5′_A and PhCH₂), 4.42 (bs, 10H, H-3′, H-4′, H-5′_B, H-6′′_B and PhCH₂), 4.56–4.58 (m, 2H, PhCH₂), 4.65–4.71 (m, 10H, PhCH₂), 4.83–4.91 (m, 16H, H-3′′, H-4′′ and PhCH₂), 5.11 (bs, 2H, H-1′′), 5.27 (bs, 2H, H-2′) 6.24 (d, 2H, J = 5.0 Hz, H-1′), 6.99–7.24 (m, 90H, Ar-H), 7.62 (s, 2H, H-5); ¹³C NMR (125 MHz, CDCl₃) δ: 64.2, 69.1, 69.16, 69.2, 69.3, 69.4, 69.5, 69.6, 69.7, 69.8, 69.9, 70.0, 70.4, 70.5, 71.9, 73.3, 73.5, 82.8, 95.7, 127.5, 127.8, 127.7, 127.75, 127.76, 127.8, 127.9, 128.0, 128.1, 128.2, 128.3, 128.4, 128.46, 128.49, 128.5, 128.6, 135.2, 136.1, 137.3, 137.5, 138.0; ³¹P NMR (202.4 MHz, CDCl₃) δ: −1.482, −1.919, −2.168; HRMS (ESI) mass calcd for C₁₆₆H₁₈₂N₆O₄₃P₆ [M]⁺ 3133.0665, found 3133.0669.

General procedure for syntheses of triazolophostin dimers 12a–d

The protected triazolophostin dimers 15a–d (0.15–0.175 g, 0.05–0.055 mmol) were treated with cyclohexene (3 mL) and Pd(OH)₂ (20% on carbon, 50 mg) in a mixture of methanol and water (9 : 1 v/v, 10 mL) at 80 °C for 4 h. The reaction mixture was then cooled, filtered through a membrane filter, washed successively with methanol and water. The combined filtrate was evaporated under reduced pressure. The crude product thus obtained was purified by ion-exchange column chromatography on Q-Sepharose matrix using 0–1.0 M TEAB as eluent to get pure triazolophostin dimers 12a–d.

Triazolophostin dimer 12a. The global debenzylation of 15a (0.15 g, 0.05 mmol) gave 46 mg (69%) of triazolophostin dimer 12a as a white hygroscopic solid: ¹H NMR (500 MHz, D₂O) δ: 3.63–3.65 (m, 8H, DEG-H), 3.70–3.83 (m, 12H, H-5′_A, H-2′′, H-6′′ and DEG-H), 4.09–4.10 (m, 2H, H-5′′), 4.41 (bs, 2H, H-4′), 4.48 (bs, 2H, H-5′_B), 4.62–4.65 (m, 6H, H-3′, H-3′′ and H-4′′), 5.16 (bs, 2H, H-2′), 5.24 (bs, 2H, H-1′′), 6.36 (bs, 2H, H-1′), 8.22 (s, 2H, H-5); ¹³C NMR (125 MHz, D₂O) δ: 60.1, 60.7, 62.8, 68.8, 69.4, 70.5, 71.5, 72.8, 73.7, 76.4, 77.9, 83.8, 90.9, 97.9, 124.3, 144.1; ³¹P NMR (202.4 MHz, D₂O) δ: 3.504, 3.583, 4.301; HRMS (ESI) mass calcd for C₃₂H₅₈N₆O₃₉P₆ [M]⁺, 1336.1165, found: 1336.1169.

Triazolophostin dimer 12b. The global debenzylation of 15b (0.155 g, 0.051 mmol) gave 51 mg (72%) of triazolophostin dimer 12b as a white hygroscopic solid: ¹H NMR (500 MHz, D₂O) δ: 3.56–3.60 (m, 12H, TEG-H), 3.69–3.74 (m, 12H, H-5′_A, H-2′′, H-6′′ and TEG-H), 4.06 (bs, 2H, H-5′′), 4.36 (bs, 2H, H-4′), 4.44 (bs, 2H, H-5′_B), 4.50–4.60 (m, 6H, H-3′, H-3′′ and H-4′′), 5.12 (bs, 2H, H-2′), 5.18 (bs, 2H, H-1′′), 6.31 (bs, 2H, H-1′), 8.18 (s, 2H, H-5); ¹³C NMR (125 MHz, D₂O) δ: 60.1, 60.7, 62.8, 68.8, 69.4, 69.48, 70.4, 71.5, 72.8, 73.7, 76.4, 77.8, 83.8, 90.8, 97.9, 124.3, 144.1; ³¹P NMR (202.4 MHz, D₂O) δ: 3.451 (2 × P), 4.224; HRMS (ESI) mass calcd for C₃₄H₆₂N₆O₄₀P₆ [M]⁺, 1380.1427, found: 1380.1420.

Triazolophostin dimer 12c. The global debenzylation of 15c (0.16 g, 0.052 mmol) gave 64 mg (85%) of triazolophostin dimer 12c as a white hygroscopic solid: ¹H NMR (500 MHz, D₂O) δ: 3.57–3.61 (m, 16H, TetraEG-H), 3.69–3.74 (m, 12H, H-5′_A, H-2′′, H-6′′ and TetraEG-H), 4.05 (bs, 2H, H-5′′), 4.37 (bs, 2H, H-4′), 4.44 (bs, 2H, H-5′_B), 4.58–4.61 (m, 6H, H-3′, H-3′′ and H-4′′), 5.12 (bs, 2H, H-2′), 5.19 (bs, 2H, H-1′′), 6.32 (bs, 2H, H-1′), 8.19 (s, 2H, H-5); ¹³C NMR (125 MHz, D₂O) δ: 60.1, 60.7, 62.9, 68.9, 69.4, 69.5, 70.5, 71.5, 72.8, 73.7, 76.4, 77.9, 83.8, 90.8, 98.0, 124.3, 144.0; ³¹P NMR (202.4 MHz, D₂O) δ: 3.478 (2 × P), 4.259; HRMS (ESI) mass calcd for C₃₆H₆₆N₆O₄₁P₆ [M]⁺, 1424.1690, found: 1424.1699.

Triazolophostin dimer 12d. The global debenzylation of 15d (0.175 g, 0.055 mmol) gave 65 mg (77%) of triazolophostin dimer 12d as a white hygroscopic solid: ¹H NMR (500 MHz, D₂O) δ: 3.58–3.72 (m, 24H, HEG-H), 3.77–3.81 (m, 12H, H-5′_A, H-2′′, H-6′′ and HEG-H), 4.01 (bs, 2H, H-5′′), 4.38–4.48 (m, 4H, H-4′ and H-5′_B), 4.58–4.63 (m, 6H, H-3′, H-3′′ and H-4′′), 5.12 (bs, 2H, H-2′), 5.20 (bs, 2H, H-1′′), 6.32 (bs, 2H, H-1′), 8.19 (s, 2H, H-5); ¹³C NMR (125 MHz, D₂O) δ: 60.2, 60.8, 62.9, 68.9, 69.4, 69.5, 70.8, 71.7, 72.6, 73.7, 76.3, 77.4, 83.8, 90.9, 97.9, 124.2, 144.2; ³¹P NMR (202.4 MHz, D₂O) δ: 3.482 (2 × P), 4.258; HRMS (ESI) mass calcd for C₄₀H₇₄N₆O₄₃P₆ [M]⁺, 1512.2214, found: 1512.2210.

Biological assay

Ca²⁺ release from the intracellular stores of saponin-permeabilized DT40 cells expressing only type 1 IP₃Rs was measured using a low-affinity Ca²⁺ indicator (Mag-fluo-4) trapped within the endoplasmic reticulum as described previously.¹¹ Briefly, Ca²⁺ uptake was initiated by addition of 1.5 mM MgATP in cytosol-like medium (140 mM KCl, 20 mM NaCl, 1 mM EGTA, 20 mM PIPES, pH 7.0, free [Ca²⁺] ∼220 nM after addition of ATP) containing p-trifluoromethoxyphenylhydrazone (FCCP) to inhibit mitochondria. After about 120 s, the triazolophostin analogs were added with cyclopiazonic acid (10 μM) to inhibit further Ca²⁺ uptake. Ca²⁺ release was assessed 10–20 s after addition of the analog, and expressed as a fraction of the ATP-dependent Ca²⁺ uptake.

Acknowledgements

AMV thanks the University Grants Commission (UGC) India for a Senior Research Fellowship (SRF) during this work. KMS thanks Department of Science and Technology (DST) India for Swarnajayanti Fellowship, Ramanujan Fellowship and for financial support. CWT and VK were supported by the Wellcome Trust.

Notes and references

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Footnote

† Electronic supplementary information (ESI) available: NMR spectral data for all the new compounds. See DOI: 10.1039/c6ra19413c

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