Bioorganic & Medicinal Chemistry Letters 25 (2015) 904–907
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Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl
Two new protein tyrosine phosphatase 1B inhibitors, hyattellactones A and B, from the Indonesian marine sponge Hyattella sp. Delfly B. Abdjul a, Hiroyuki Yamazaki a,⇑, Ohgi Takahashi a, Ryota Kirikoshi a, Remy E. P. Mangindaan b, Michio Namikoshi a a b
Faculty of Pharmaceutical Sciences, Tohoku Pharmaceutical University, Sendai 981-8558, Japan Faculty of Fisheries and Marine Science, Sam Ratulangi University, Kampus Bahu, Manado 95115, Indonesia
a r t i c l e
i n f o
Article history: Received 25 November 2014 Revised 12 December 2014 Accepted 16 December 2014 Available online 30 December 2014 Keywords: Hyattellactone Scalarane sesterterpene Marine sponge Hyattella sp. PTP1B inhibitor Phyllofolactone
a b s t r a c t Two unique sesterterpenes, hyattellactones A (1) and B (2), together with two known sesterterpenes, phyllofolactones F (3) and G (4), were isolated from the Indonesian marine sponge Hyattella sp. The structures of the two new compounds, 1 and 2 were assigned based on their spectroscopic data. Hyattellactone A (1) was a scalarane sesterterpene with an a,b-unsaturated-c-lactone ring and C-ethyl group, while B (2) was an epimer of 1 at the C-24 position. Compounds 1 and 3 inhibited PTP1B activity with IC50 values of 7.45 and 7.47 lM, respectively. On the other hand, compounds 2 and 4 (24S-isomers of 1 and 3, respectively) showed much reduced activity than the 24R-isomers. Ó 2014 Elsevier Ltd. All rights reserved.
Protein tyrosine phosphatase 1B (PTP1B) is an effective and attractive therapeutic target for the treatment of type-2 diabetes and plays a key role as a negative regulator in insulin and leptin signaling. Therefore, the development of PTP1B inhibitors has been expected to provide new medicines for type-2 diabetes mellitus and obesity.1 To date, approximately 300 PTP1B inhibitors have been reported from various natural resources.2 However, the activities and selectivities of these natural products against PTP1B were not satisfactory. Accordingly, the search for more potent and selective PTP1B inhibitors from natural resources is one of the most important subjects in natural product chemistry. In the course of our research on PTP1B inhibitors from marine organisms, we have reported polybromodiphenyl ethers and dehydroeuryspongin from marine sponges.3 Further investigations on extracts from marine invertebrates revealed that an extract from the Indonesian marine sponge Hyattella sp. exhibited prominent inhibitory activity against PTP1B. The bioassay-guided separation of this extract led to the isolation of two new sesterterpenes, hyattellactones A (1) and B (2), together with two known sesterterpenes, phyllofolactones F (3)4 and G (4)4 (Fig. 1). Compounds 1–4 were unique pentacyclic scalarane sesterterpenes that possessed an a,b-unsaturated-c-lactone ring and C-ethyl ⇑ Corresponding author. Tel./fax: +81 22 727 0218. E-mail address: [email protected] (H. Yamazaki). http://dx.doi.org/10.1016/j.bmcl.2014.12.058 0960-894X/Ó 2014 Elsevier Ltd. All rights reserved.
group. Although a number of compounds in this class have been reported from several marine sponges,5 compounds 1 and 2 are the first examples to possess an ethyl group at C-10. We herein described the isolation, structure elucidation including stereochemistry, and biological activities of compounds 1–4. The marine sponge (487 g, wet weight)6 was extracted with ethanol, and the extract (11.3 g) was partitioned between hexane and 90% CH3OH. The hexane extract (0.51 g) was separated by a Silica gel column (50 g) to give five fractions. The bioactive fraction was purified by repeated HPLC using an ODS column to yield compounds 1 (4.3 mg), 2 (1.8 mg), 3 (1.1 mg), and 4 (1.7 mg) as colorless oils.7 Compounds 3 and 4 were identified as the known sesterterpenes, phyllofolactones F and G, by comparing the spectral data obtained with reported values.4c Compounds 3 and 4 were the epimers at C-24 and were previously reported in several marine sponges of the genus Phyllospongia.4 The physicochemical properties of 1 and 2 were very similar to those of 3 and 4, which suggested that these compounds shared the same skeleton.8–11 The molecular formula of hyattellactone A (1)10 was assigned as C27H42O3 from HREIMS (m/z 414.3151 [M]+, D +1.7 mmu) and NMR data for 1 (Table 1). The 1H and 13C NMR signals of 1 (in pyridine-d5) were classified into six methyls, nine methylenes, three sp3 methines, two sp3 oxygenated methines, four sp3 quaternary carbons, two sp2 quaternary carbons, and
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OH
O
O
18
12
OH
O
O
OH
O
O
OH
O
O
24
14
1
H 4
8
H
H
H
H
H
H
H
6
H
H
Hyattellactone A (1)
H
Hyattellactone B (2)
H
Phyllofolactone F (3)
Phyllofolactone G (4)
Figure 1. Structures of compounds 1–4 isolated from the marine sponge Hyattella sp. collected in North Sulawesi, Indonesia.
Table 1 H and 13C NMR data for hyattellactones A (1) and B (2) in C5D5N
1
Position
1 dC
dH (J in Hz)
1
35.6
2
18.8
3
42.3
0.58 2.09 1.15 1.37 1.11 1.39
ddd d m m m m
4 5 6
33.2 58.3 18.0
7
42.3
0.77 1.40 1.73 0.72 1.71
m m m m m
8 9 10 11
37.6 59.1 40.5 29.5
12 13 14 15
77.0 42.6 55.8 16.7
16
24.9
17 18 19 20 21 22 23 24 25 26 27 12-OH
167.6 135.6 34.7 22.2 17.2 21.2 16.8 80.0 175.7 18.1 10.7
2
(12.8,12.8,3.6) (12.8)
0.91 1.51 1.78 1.54 2.25
0.89 0.83 0.95 1.62 1.25 4.95
1.36 d 0.85 brd 6.49 brs
35.7
0.61 2.11 1.15 1.32 1.11 1.32
ddd m m m m m
0.78 1.40 1.72 0.72 1.71
m m m m d
42.2 33.2 58.5 18.1 42.6
(13.4, 2.4)
37.6 59.3 40.5 29.5 76.7 42.5 55.5 16.9
m m m m m
s s s q s m
dH (J in Hz)
18.8
0.80 m 1.82 dd 2.16 m 3.65 m
dC
24.6
(7.6)
(6.8) (7.6)
one carbonyl carbon, and then assigned by an analysis of 2D NMR spectra (Table 1). The presence of an OH group was elucidated from 1H NMR data (d 6.49) and IR absorption at 3386 cm 1. The 1 H–1H COSY spectrum of 1 revealed the partial structures I–VI, as shown by the blue bold lines in Figure 2. The connectivities of partial structures I–VI were elucidated from the HMBC spectrum of 1 (Fig. 1) and confirmed by a comparison of NMR data for 1 with those for phyllofolactones F (3) and G (4). The position of an ethyl group (partial structure III) was assigned at C-10 based on HMBC correlations from H2-22 (d 1.62) to C-1 (d 35.6), C-9 (d 59.1), and C-10 (d 40.5) and from H3-27 (d 0.85) to C-10 (Fig. 2). The relative stereochemistry of 1 was determined by an analysis of the NOESY spectrum in pyridine-d5 (Fig. 3) and comparison of NMR data with those for 3 and 4. The ring junctions at A/B, B/C, and C/D were revealed as the trans-configuration from NOE correlations between H3-20 (d 0.83)/H2-22 (d 1.62), H3-21
167.7 135.6 34.7 22.3 17.2 21.3 17.0 79.7 175.7 18.3 10.7
(12.9,12.9,3.4)
(12.4)
0.80 m 1.82 dd 2.20 m 3.65 m 0.91 1.50 1.75 1.54 2.33
m m m m dd
0.89 0.84 0.95 1.62 1.23 4.99
s s s q s m
1.28 d 0.86 brd 6.50 brs
(13.4, 2.4)
(19.3, 5.6)
(7.6)
(6.8) (7.6)
Figure 2. 1H–1H COSY and key HMBC data for hyattellactones A (1) and B (2).
(d 0.95)/H2-22, H3-21/H3-23 (d 1.25), Ha-1 (d 0.58)/H-5 (d 0.77), H-5/H-9 (d 0.80), and H-12 (d 3.65)/H-14 (d 0.91) (Fig. 3). An
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Figure 3. Stereostructure and key NOESY correlations of hyattellactone A (1).
1. Consequently, the absolute structures of 1–4 were assigned as shown in Figure 1. More than 60 scalarane sesterterpenes reported from various marine sponges have a C-ethyl group in their structures.5 The ethyl group of these compounds is attached at the C-4 position, except for one example that had the ethyl group at C-13.16 Therefore, hyattellactones A (1) and B (2) are the first examples to possess the ethyl group at the C-10 position. Scalarane sesterterpenes have been reported to exhibit a wide range of biological activities such as cytotoxicity against several cancer cell lines as well as antifeedant, ichthyotoxicity, antiinflammatory, antimicrobial including antitubercular and antiMRSA, antithrombocyte, and vasodilatory activities.5 However, the biological activities of phyllofolactones F (3) and G (4) have not yet been reported.4 The inhibitory activities of compounds 1–4 against PTP1B were evaluated according to a previously described method.3 Two (24R)isomers, 1 and 3 inhibited PTP1B activity with IC50 values of 7.45 and 7.47 lM, respectively. Oleanolic acid, a positive control,17 had an IC50 value of 1.03 lM in the same bioassay. The (24S)-isomer of 1, compound 2, showed 42% inhibition at 24.2 lM, and the (24S)-isomer of 3, compound 4, was inactive at 24.2 lM. Therefore, stereochemistry at the C-24 position was very important for the inhibitory activities of these scalarane sesterterpenes. Moreover, the cytotoxic activities of compounds 1–4 were examined against human T-cell lymphoma Jurkat cells,18 and 1–4 did not inhibit cell proliferation at 24.2 lM. Acknowledgments
Figure 4. Experimental CD spectrum of hyattellactone A (1) (green line) and calculated ECD spectra of 1 (black line) and its enantiomer (dashed line).
NOE correlation between 12-OH (d 6.49) and H3-23 indicated the b-orientation of the OH group. The stereochemistry of C-24 (C-26) was deduced to be the same as that of 3 (a-orientation) from an NOE correlation between H-14 (d 0.91)/Ha-16 (d 2.25) and Ha-16/H3-26 (d 1.36), and confirmed by a comparison of chemical shifts at the 24 position of 1 (dH 4.84 and dC 79.5 in CDCl3) with those of 3 (dH 4.84 and dC 79.5 in CDCl3).8 Based on the NOE data for 1, a Monte Carlo conformational analysis was performed with an MMFF94 force field utilizing Spartan’14,12 as shown in Figure 3. Hyattellactone B (2)11 had the same molecular formula (C27H42O3) as 1, and its 1H and 13C NMR spectra and physicochemical properties were also very similar to those of 1. 1H–1H COSY and HMBC data for 2 revealed that 1 and 2 had the same skeletal structure, as shown in Figure 2. A marked difference in 1H signals between 1 and 2 was detected for H-24 and H3-26. Therefore, compound 2 was assigned the structure as an epimer of 1 at C-24, which was also supported by the comparison of chemical shifts at the 24 position of 2 (dH 4.86 and dC 79.1 in CDCl3) and 4 (dH 4.86 and dC 79.1 in CDCl3).9 Thus, the structure of compound 2 was assigned as 24-epi-hyttellactone A. The absolute configurations of compounds 1–4 were deduced from a comparison of the experimental Circular Dichroism (CD) spectra of 1–4 with the calculated Electronic Circular Dichroism (ECD) spectra.13 The experimental CD spectrum of 1 (green line) was very similar to the ECD spectrum calculated for the (5S,8R,9S,10S,12R,13S,14S,24R)-isomer (black solid line) (Fig. 4). The CD and ECD spectra of 2–4 were superimposable to those of
This work was supported in part by the Foundation for Japanese Chemical Research to H.Y. We are grateful to the Center for Biomedical Research, Institute of Development, Aging and Cancer, Tohoku University for providing the human cancer cell lines, Dr. K. Ogawa of Z. Nakai Laboratory for identification of the marine sponge, and to Mr. T. Matsuki and S. Sato of Tohoku Pharmaceutical University for measurements of mass and NMR spectra. References and notes 1. (a) Barr, A. J. Future Med. Chem. 2010, 2, 1563; (b) Zhang, S.; Zhang, Z. Y. Drug Discovery Today 2007, 12, 373; (c) Lee, S.; Wang, Q. Med. Res. Rev. 2007, 27, 553. 2. Jiang, C. S.; Liang, L. F.; Guo, Y. W. Acta Pharmacol. Sin. 2012, 33, 1217. 3. (a) Yamazaki, H.; Sumilat, D. A.; Kanno, S.; Ukai, K.; Rotinsulu, H.; Wewengkang, D. S.; Ishikawa, M.; Mangindaan, R. E.; Namikoshi, M. J. Nat. Med. 2013, 67, 730; (b) Yamazaki, H.; Nakazawa, T.; Sumilat, D. A.; Takahashi, O.; Ukai, K.; Takahashi, S.; Namikoshi, M. Bioorg. Med. Chem. Lett. 2013, 23, 2151. 4. (a) Wan, Y.; Li, Q.; Zeng, L.; Su, J. Acta Scientiarum Naturalium 1998, 37, 81; (b) Reddy, M. V. R.; Venkateswarlu, Y.; Rao, J. V. Indian J. Chem. B 1993, 32, 1196; (c) Ponomarenko, L. P.; Kalinovsky, A. I.; Stonik, V. A. J. Nat. Prod. 2004, 67, 1507. 5. Gonzalez, M. A. Curr. Bioact. Compd. 2010, 6, 178. 6. The marine sponge was collected through scuba diving in the coral reef at the Lembeh Strait, North Sulawesi, Indonesia, in September 2013 and was subsequently identified as Hyattella sp. A voucher specimen was deposited at the Faculty of Fisheries and Marine Science, Sam Ratulangi University and the Faculty of Pharmaceutical Sciences, Tohoku Pharmaceutical University as 13L01. 7. The sponge (487.0 g, wet weight) was cut into small pieces and soaked in ethanol (1.5 L) on a boat immediately after its collection. The ethanol extract (11.3 g), after the evaporation of ethanol, was suspended in 200 mL of 90% CH3OH and extracted with hexane. The hexane extract (0.51 g) was separated into five fractions (Fr. 1–Fr. 5) by a silica gel column (50 g) with 200 mL of hexane, a mixture (v/v) of hexane–ethyl acetate (4:1, 3:2 and 2:3), and then ethyl acetate. The active Fr. 2 (138.6 mg) was subjected to preparative HPLC [column, PEGASIL ODS (Senshu Sci. Co. Ltd, i.d. 10 mm 250 mm); solvent, 85% CH3OH containing 0.05% TFA; flow rate, 2.0 mL/min; detection, UV 210 nm] to give hyattellactone A (1) (4.3 mg, tR = 63.8 min), phyllofolactone F (3) (1.1 mg, tR = 73.1 min), and five fractions (Fr. 2-1–Fr. 2-5). The active Fr. 2-3 was further separated by preparative HPLC [column, PEGASIL ODS (i.d. 10 mm 250 mm); solvent, 80% CH3OH containing 0.05% TFA; flow rate, 2.0 mL/min; detection, UV 210 nm] to afford hyattellactone B (2) (1.8 mg, tR = 75.3 min) and phyllofolactone G (4) (1.7 mg, tR = 86.1 min).
D.B. Abdjul et al. / Bioorg. Med. Chem. Lett. 25 (2015) 904–907 8. Phyllofolactone F (3): a colorless oil; [a]23 D +9.7 (c 0.04, CHCl3); IR (KBr) mmax 3387, 2928, 1710, 1652, 1064 cm 1; UV (CH3OH) kmax 220 (e 9442); CD (CH3CN) kextermum (De) 202 (2.6), 244 ( 3.1); EIMS m/z 414 [M]+; HREIMS m/z 414.3131 ([M]+; calcd for C27H42O3, 414.3134); 1H NMR (CDCl3) d 5.98 (1H, br s), 4.84 (1H, q, J = 6.7 Hz), 3.65 (1H, m), 2.33 (1H, m), 2.20 (1H, m), 1.93 (1H, m), 1.91 (1H, m), 1.86 (1H, m), 1.82 (1H, m), 1.74 (1H, m), 1.66 (1H, m), 1.62 (1H, m), 1.56 (1H, m), 1.48 (1H, m), 1.38 (3H, d, J = 6.7 Hz), 1.16 (2H, m), 1.12 (3H, s), 1.02 (1H, m), 0.90 (2H, m), 0.87 (3H, s), 0.86 (3H, s), 0.84 (2H, m), 0.78 (4H, br s), 0.74 (3H, t, J = 7.5 Hz); 13C NMR (CDCl3) d 174.9, 166.0, 135.5, 79.5, 75.6, 58.8, 58.4, 55.2, 42.1, 42.0, 40.2, 37.5, 37.3, 36.7, 36.1, 28.6, 25.8, 24.8, 24.5, 18.3, 18.17, 18.17, 17.2, 16.78, 16.76, 16.4, 8.6. 9. Phyllofolactone G (4): a colorless oil; [a]23 D +4.7 (c 0.10, CHCl3); IR (KBr) mmax 3385, 2927, 1711, 1652, 1065 cm 1; UV (CH3OH) kmax 220 (e 14173); CD (CH3CN) kextermum (De) 208 (2.7), 244 ( 2.3); EIMS m/z 414 [M]+; HREIMS m/z 414.3151 ([M]+; calcd for C27H42O3, 414.3134); 1H NMR (CDCl3) d 5.98 (1H, br s), 4.86 (1H, q, J = 6.7 Hz), 3.67 (1H, m), 2.37 (1H, m), 2.18 (1H, m), 1,95 (1H, m), 1.92 (1H, m), 1.87 (1H, m), 1.82 (1H, m), 1.74 (1H, m), 1.68 (1H, m), 1.62 (1H, m), 1.56 (1H, m), 1.48 (1H, m), 1.40 (3H, d, J = 6.7 Hz), 1.19 (2H, m), 1.11 (3H, s), 1.02 (1H, m), 0.89 (1H, m), 0.88 (4H, br s), 0.86 (3H, s), 0.85 (2H, m), 0.79 (4H, br s), 0.74 (3H, t, J = 7.5 Hz); 13C NMR (CDCl3) d 175.0, 165.9, 135.5, 79.1, 75.5, 58.8, 58.5, 55.0, 42.03, 42.00, 40.2, 37.5, 37.3, 36.7, 36.1, 28.5, 25.8, 24.5, 24.4, 18.4, 18.3, 18.2, 17.2, 16.9, 16.8, 16.6, 8.6. 10. Hyattellactone A (1): a colorless oil; [a]23 D +11.3 (c 0.10, CHCl3); IR (KBr) mmax 3386, 2930, 1712, 1652, 1063 cm 1; UV (CH3OH) kmax 220 (e 7837); CD (CH3CN) kextermum (De) 206 (3.0), 245 ( 2.7); EIMS m/z 414 [M]+; HREIMS m/z 414.3151 ([M]+; calcd for C27H42O3, 414.3134); 1H NMR (CDCl3) d 6.00 (1H, br s), 4.84 (1H, q, J = 6.7 Hz), 3.51 (1H, m), 2.33 (2H, m), 2.21 (1H, m), 2.18 (1H, m), 2.03 (1H, m), 2.00 (1H, m), 1.95 (1H, m), 1.85 (1H, m), 1.68 (1H, m), 1.66 (2H, m), 1.49 (4H, m), 1.41 (1H, m), 1.38 (3H, d, J = 6.7 Hz), 1.13 (4H, br s), 1.01 (4H, br s), 0.95 (2H, m), 0.90 (3H, t, J = 7.6 Hz), 0.84 (7H, br s); 13C NMR (CDCl3) d 175.0, 166.1, 135.4, 79.5, 76.5, 59.3, 58.6, 55.9, 42.7, 42.3, 42.1, 40.4, 37.6, 35.6, 34.6, 33.2, 28.7, 24.8, 22.1, 20.9, 18.6, 18.2, 17.8, 17.2, 16.6, 16.5, 10.6; 1H and 13 C NMR (Pyridine-d5), see Table 1.
907
11. Hyattellactone B (2): a colorless oil; [a]23 D +9.4 (c 0.10, CHCl3); IR (KBr) mmax 3378, 2924, 1708, 1136 cm 1; UV (CH3OH) kmax 220 (e 23290); CD (CH3CN) kextermum (De) 208 (2.0), 245 ( 1.8); EIMS m/z 414 [M]+; HREIMS m/z 414.3126 ([M]+; calcd for C27H42O3, 414.3134); 1H NMR (CDCl3) d 6.02 (1H, br s), 4.86 (1H, q, J = 6.8 Hz), 3.52 (1H, m), 2.40 (2H, m), 2.21 (1H, m), 2.18 (1H, m), 2.03 (1H, m), 2.02 (1H, m), 1.94 (1H, m), 1.87 (1H, m), 1.68 (1H, m), 1.66 (2H, m), 1.49 (4H, m), 1.41 (1H, m), 1.40 (3H, d, J = 6.8 Hz), 1.13 (4H, br s), 1.01 (4H, br s), 0.93 (2H, m), 0.90 (3H, t, J = 7.5 Hz), 0.85 (7H, br s); 13C NMR (CDCl3) d 175.1, 166.0, 135.4, 79.1, 76.3, 59.4, 58.6, 55.7, 42.7, 42.2, 42.0, 40.4, 37.6, 35.6, 34.6, 33.2, 28.8, 24.6, 22.2, 20.9, 18.5, 18.4, 17.8, 17.1, 16.70, 16.68, 10.6; 1H and 13C NMR (Pyridine-d5), see Table 1. 12. Spartan’14, Wavefunction, Irvine, CA. 13. The most stable conformer of hyattellactone A was predicted using Spartan’1412 through a preliminary conformational analysis with the MMFF94 force field, followed by geometry optimization using the density functional theory (DFT) with the B3LYP functional and 6-31G(d) basis set. The ECD spectrum in acetonitrile was calculated for the predicted most stable conformer using Gaussian 0314 by time-dependent DFT (TDDFT) with the B3LYP functional and 6-31+G(d,p) basis set. The solvent effect was introduced by the polarizable continuum model (PCM). Ten low-lying excited states were calculated (corresponding to the wavelength region down to approximately 196 nm). The calculated spectrum was displayed using GaussView 5.0.915 with the peak half-width at half height being 0.333 eV. The calculated spectrum was shifted by 15 nm to match the experimental spectrum. 14. Frisch, M. J.; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.; Robb, M. A.; Cheeseman, J. R.; Montgomery, J. A., Jr.; Vreven, T.; Kudin, K. N.; Burant, J. C., et al. Gaussian 03, revision E.1; Gaussian: Wallingford, CT, USA, 2004. 15. Dennington, R.; Keith, T.; Millam, J. GaussView, version 5; Semichem: Shawnee Mission, KS, USA, 2009. 16. De Rosa, S.; Crispino, A.; De Giulio, A.; Iodice, C.; Tommonaro, G. Tetrahedron 1998, 54, 6185. 17. Liu, J. J. Ethnopharmacol. 1995, 49, 57. 18. Mosmann, T. J. Immunol. Methods 1983, 65, 55.
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl
Two new protein tyrosine phosphatase 1B inhibitors, hyattellactones A and B, from the Indonesian marine sponge Hyattella sp. Delfly B. Abdjul a, Hiroyuki Yamazaki a,⇑, Ohgi Takahashi a, Ryota Kirikoshi a, Remy E. P. Mangindaan b, Michio Namikoshi a a b
Faculty of Pharmaceutical Sciences, Tohoku Pharmaceutical University, Sendai 981-8558, Japan Faculty of Fisheries and Marine Science, Sam Ratulangi University, Kampus Bahu, Manado 95115, Indonesia
a r t i c l e
i n f o
Article history: Received 25 November 2014 Revised 12 December 2014 Accepted 16 December 2014 Available online 30 December 2014 Keywords: Hyattellactone Scalarane sesterterpene Marine sponge Hyattella sp. PTP1B inhibitor Phyllofolactone
a b s t r a c t Two unique sesterterpenes, hyattellactones A (1) and B (2), together with two known sesterterpenes, phyllofolactones F (3) and G (4), were isolated from the Indonesian marine sponge Hyattella sp. The structures of the two new compounds, 1 and 2 were assigned based on their spectroscopic data. Hyattellactone A (1) was a scalarane sesterterpene with an a,b-unsaturated-c-lactone ring and C-ethyl group, while B (2) was an epimer of 1 at the C-24 position. Compounds 1 and 3 inhibited PTP1B activity with IC50 values of 7.45 and 7.47 lM, respectively. On the other hand, compounds 2 and 4 (24S-isomers of 1 and 3, respectively) showed much reduced activity than the 24R-isomers. Ó 2014 Elsevier Ltd. All rights reserved.
Protein tyrosine phosphatase 1B (PTP1B) is an effective and attractive therapeutic target for the treatment of type-2 diabetes and plays a key role as a negative regulator in insulin and leptin signaling. Therefore, the development of PTP1B inhibitors has been expected to provide new medicines for type-2 diabetes mellitus and obesity.1 To date, approximately 300 PTP1B inhibitors have been reported from various natural resources.2 However, the activities and selectivities of these natural products against PTP1B were not satisfactory. Accordingly, the search for more potent and selective PTP1B inhibitors from natural resources is one of the most important subjects in natural product chemistry. In the course of our research on PTP1B inhibitors from marine organisms, we have reported polybromodiphenyl ethers and dehydroeuryspongin from marine sponges.3 Further investigations on extracts from marine invertebrates revealed that an extract from the Indonesian marine sponge Hyattella sp. exhibited prominent inhibitory activity against PTP1B. The bioassay-guided separation of this extract led to the isolation of two new sesterterpenes, hyattellactones A (1) and B (2), together with two known sesterterpenes, phyllofolactones F (3)4 and G (4)4 (Fig. 1). Compounds 1–4 were unique pentacyclic scalarane sesterterpenes that possessed an a,b-unsaturated-c-lactone ring and C-ethyl ⇑ Corresponding author. Tel./fax: +81 22 727 0218. E-mail address: [email protected] (H. Yamazaki). http://dx.doi.org/10.1016/j.bmcl.2014.12.058 0960-894X/Ó 2014 Elsevier Ltd. All rights reserved.
group. Although a number of compounds in this class have been reported from several marine sponges,5 compounds 1 and 2 are the first examples to possess an ethyl group at C-10. We herein described the isolation, structure elucidation including stereochemistry, and biological activities of compounds 1–4. The marine sponge (487 g, wet weight)6 was extracted with ethanol, and the extract (11.3 g) was partitioned between hexane and 90% CH3OH. The hexane extract (0.51 g) was separated by a Silica gel column (50 g) to give five fractions. The bioactive fraction was purified by repeated HPLC using an ODS column to yield compounds 1 (4.3 mg), 2 (1.8 mg), 3 (1.1 mg), and 4 (1.7 mg) as colorless oils.7 Compounds 3 and 4 were identified as the known sesterterpenes, phyllofolactones F and G, by comparing the spectral data obtained with reported values.4c Compounds 3 and 4 were the epimers at C-24 and were previously reported in several marine sponges of the genus Phyllospongia.4 The physicochemical properties of 1 and 2 were very similar to those of 3 and 4, which suggested that these compounds shared the same skeleton.8–11 The molecular formula of hyattellactone A (1)10 was assigned as C27H42O3 from HREIMS (m/z 414.3151 [M]+, D +1.7 mmu) and NMR data for 1 (Table 1). The 1H and 13C NMR signals of 1 (in pyridine-d5) were classified into six methyls, nine methylenes, three sp3 methines, two sp3 oxygenated methines, four sp3 quaternary carbons, two sp2 quaternary carbons, and
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OH
O
O
18
12
OH
O
O
OH
O
O
OH
O
O
24
14
1
H 4
8
H
H
H
H
H
H
H
6
H
H
Hyattellactone A (1)
H
Hyattellactone B (2)
H
Phyllofolactone F (3)
Phyllofolactone G (4)
Figure 1. Structures of compounds 1–4 isolated from the marine sponge Hyattella sp. collected in North Sulawesi, Indonesia.
Table 1 H and 13C NMR data for hyattellactones A (1) and B (2) in C5D5N
1
Position
1 dC
dH (J in Hz)
1
35.6
2
18.8
3
42.3
0.58 2.09 1.15 1.37 1.11 1.39
ddd d m m m m
4 5 6
33.2 58.3 18.0
7
42.3
0.77 1.40 1.73 0.72 1.71
m m m m m
8 9 10 11
37.6 59.1 40.5 29.5
12 13 14 15
77.0 42.6 55.8 16.7
16
24.9
17 18 19 20 21 22 23 24 25 26 27 12-OH
167.6 135.6 34.7 22.2 17.2 21.2 16.8 80.0 175.7 18.1 10.7
2
(12.8,12.8,3.6) (12.8)
0.91 1.51 1.78 1.54 2.25
0.89 0.83 0.95 1.62 1.25 4.95
1.36 d 0.85 brd 6.49 brs
35.7
0.61 2.11 1.15 1.32 1.11 1.32
ddd m m m m m
0.78 1.40 1.72 0.72 1.71
m m m m d
42.2 33.2 58.5 18.1 42.6
(13.4, 2.4)
37.6 59.3 40.5 29.5 76.7 42.5 55.5 16.9
m m m m m
s s s q s m
dH (J in Hz)
18.8
0.80 m 1.82 dd 2.16 m 3.65 m
dC
24.6
(7.6)
(6.8) (7.6)
one carbonyl carbon, and then assigned by an analysis of 2D NMR spectra (Table 1). The presence of an OH group was elucidated from 1H NMR data (d 6.49) and IR absorption at 3386 cm 1. The 1 H–1H COSY spectrum of 1 revealed the partial structures I–VI, as shown by the blue bold lines in Figure 2. The connectivities of partial structures I–VI were elucidated from the HMBC spectrum of 1 (Fig. 1) and confirmed by a comparison of NMR data for 1 with those for phyllofolactones F (3) and G (4). The position of an ethyl group (partial structure III) was assigned at C-10 based on HMBC correlations from H2-22 (d 1.62) to C-1 (d 35.6), C-9 (d 59.1), and C-10 (d 40.5) and from H3-27 (d 0.85) to C-10 (Fig. 2). The relative stereochemistry of 1 was determined by an analysis of the NOESY spectrum in pyridine-d5 (Fig. 3) and comparison of NMR data with those for 3 and 4. The ring junctions at A/B, B/C, and C/D were revealed as the trans-configuration from NOE correlations between H3-20 (d 0.83)/H2-22 (d 1.62), H3-21
167.7 135.6 34.7 22.3 17.2 21.3 17.0 79.7 175.7 18.3 10.7
(12.9,12.9,3.4)
(12.4)
0.80 m 1.82 dd 2.20 m 3.65 m 0.91 1.50 1.75 1.54 2.33
m m m m dd
0.89 0.84 0.95 1.62 1.23 4.99
s s s q s m
1.28 d 0.86 brd 6.50 brs
(13.4, 2.4)
(19.3, 5.6)
(7.6)
(6.8) (7.6)
Figure 2. 1H–1H COSY and key HMBC data for hyattellactones A (1) and B (2).
(d 0.95)/H2-22, H3-21/H3-23 (d 1.25), Ha-1 (d 0.58)/H-5 (d 0.77), H-5/H-9 (d 0.80), and H-12 (d 3.65)/H-14 (d 0.91) (Fig. 3). An
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Figure 3. Stereostructure and key NOESY correlations of hyattellactone A (1).
1. Consequently, the absolute structures of 1–4 were assigned as shown in Figure 1. More than 60 scalarane sesterterpenes reported from various marine sponges have a C-ethyl group in their structures.5 The ethyl group of these compounds is attached at the C-4 position, except for one example that had the ethyl group at C-13.16 Therefore, hyattellactones A (1) and B (2) are the first examples to possess the ethyl group at the C-10 position. Scalarane sesterterpenes have been reported to exhibit a wide range of biological activities such as cytotoxicity against several cancer cell lines as well as antifeedant, ichthyotoxicity, antiinflammatory, antimicrobial including antitubercular and antiMRSA, antithrombocyte, and vasodilatory activities.5 However, the biological activities of phyllofolactones F (3) and G (4) have not yet been reported.4 The inhibitory activities of compounds 1–4 against PTP1B were evaluated according to a previously described method.3 Two (24R)isomers, 1 and 3 inhibited PTP1B activity with IC50 values of 7.45 and 7.47 lM, respectively. Oleanolic acid, a positive control,17 had an IC50 value of 1.03 lM in the same bioassay. The (24S)-isomer of 1, compound 2, showed 42% inhibition at 24.2 lM, and the (24S)-isomer of 3, compound 4, was inactive at 24.2 lM. Therefore, stereochemistry at the C-24 position was very important for the inhibitory activities of these scalarane sesterterpenes. Moreover, the cytotoxic activities of compounds 1–4 were examined against human T-cell lymphoma Jurkat cells,18 and 1–4 did not inhibit cell proliferation at 24.2 lM. Acknowledgments
Figure 4. Experimental CD spectrum of hyattellactone A (1) (green line) and calculated ECD spectra of 1 (black line) and its enantiomer (dashed line).
NOE correlation between 12-OH (d 6.49) and H3-23 indicated the b-orientation of the OH group. The stereochemistry of C-24 (C-26) was deduced to be the same as that of 3 (a-orientation) from an NOE correlation between H-14 (d 0.91)/Ha-16 (d 2.25) and Ha-16/H3-26 (d 1.36), and confirmed by a comparison of chemical shifts at the 24 position of 1 (dH 4.84 and dC 79.5 in CDCl3) with those of 3 (dH 4.84 and dC 79.5 in CDCl3).8 Based on the NOE data for 1, a Monte Carlo conformational analysis was performed with an MMFF94 force field utilizing Spartan’14,12 as shown in Figure 3. Hyattellactone B (2)11 had the same molecular formula (C27H42O3) as 1, and its 1H and 13C NMR spectra and physicochemical properties were also very similar to those of 1. 1H–1H COSY and HMBC data for 2 revealed that 1 and 2 had the same skeletal structure, as shown in Figure 2. A marked difference in 1H signals between 1 and 2 was detected for H-24 and H3-26. Therefore, compound 2 was assigned the structure as an epimer of 1 at C-24, which was also supported by the comparison of chemical shifts at the 24 position of 2 (dH 4.86 and dC 79.1 in CDCl3) and 4 (dH 4.86 and dC 79.1 in CDCl3).9 Thus, the structure of compound 2 was assigned as 24-epi-hyttellactone A. The absolute configurations of compounds 1–4 were deduced from a comparison of the experimental Circular Dichroism (CD) spectra of 1–4 with the calculated Electronic Circular Dichroism (ECD) spectra.13 The experimental CD spectrum of 1 (green line) was very similar to the ECD spectrum calculated for the (5S,8R,9S,10S,12R,13S,14S,24R)-isomer (black solid line) (Fig. 4). The CD and ECD spectra of 2–4 were superimposable to those of
This work was supported in part by the Foundation for Japanese Chemical Research to H.Y. We are grateful to the Center for Biomedical Research, Institute of Development, Aging and Cancer, Tohoku University for providing the human cancer cell lines, Dr. K. Ogawa of Z. Nakai Laboratory for identification of the marine sponge, and to Mr. T. Matsuki and S. Sato of Tohoku Pharmaceutical University for measurements of mass and NMR spectra. References and notes 1. (a) Barr, A. J. Future Med. Chem. 2010, 2, 1563; (b) Zhang, S.; Zhang, Z. Y. Drug Discovery Today 2007, 12, 373; (c) Lee, S.; Wang, Q. Med. Res. Rev. 2007, 27, 553. 2. Jiang, C. S.; Liang, L. F.; Guo, Y. W. Acta Pharmacol. Sin. 2012, 33, 1217. 3. (a) Yamazaki, H.; Sumilat, D. A.; Kanno, S.; Ukai, K.; Rotinsulu, H.; Wewengkang, D. S.; Ishikawa, M.; Mangindaan, R. E.; Namikoshi, M. J. Nat. Med. 2013, 67, 730; (b) Yamazaki, H.; Nakazawa, T.; Sumilat, D. A.; Takahashi, O.; Ukai, K.; Takahashi, S.; Namikoshi, M. Bioorg. Med. Chem. Lett. 2013, 23, 2151. 4. (a) Wan, Y.; Li, Q.; Zeng, L.; Su, J. Acta Scientiarum Naturalium 1998, 37, 81; (b) Reddy, M. V. R.; Venkateswarlu, Y.; Rao, J. V. Indian J. Chem. B 1993, 32, 1196; (c) Ponomarenko, L. P.; Kalinovsky, A. I.; Stonik, V. A. J. Nat. Prod. 2004, 67, 1507. 5. Gonzalez, M. A. Curr. Bioact. Compd. 2010, 6, 178. 6. The marine sponge was collected through scuba diving in the coral reef at the Lembeh Strait, North Sulawesi, Indonesia, in September 2013 and was subsequently identified as Hyattella sp. A voucher specimen was deposited at the Faculty of Fisheries and Marine Science, Sam Ratulangi University and the Faculty of Pharmaceutical Sciences, Tohoku Pharmaceutical University as 13L01. 7. The sponge (487.0 g, wet weight) was cut into small pieces and soaked in ethanol (1.5 L) on a boat immediately after its collection. The ethanol extract (11.3 g), after the evaporation of ethanol, was suspended in 200 mL of 90% CH3OH and extracted with hexane. The hexane extract (0.51 g) was separated into five fractions (Fr. 1–Fr. 5) by a silica gel column (50 g) with 200 mL of hexane, a mixture (v/v) of hexane–ethyl acetate (4:1, 3:2 and 2:3), and then ethyl acetate. The active Fr. 2 (138.6 mg) was subjected to preparative HPLC [column, PEGASIL ODS (Senshu Sci. Co. Ltd, i.d. 10 mm 250 mm); solvent, 85% CH3OH containing 0.05% TFA; flow rate, 2.0 mL/min; detection, UV 210 nm] to give hyattellactone A (1) (4.3 mg, tR = 63.8 min), phyllofolactone F (3) (1.1 mg, tR = 73.1 min), and five fractions (Fr. 2-1–Fr. 2-5). The active Fr. 2-3 was further separated by preparative HPLC [column, PEGASIL ODS (i.d. 10 mm 250 mm); solvent, 80% CH3OH containing 0.05% TFA; flow rate, 2.0 mL/min; detection, UV 210 nm] to afford hyattellactone B (2) (1.8 mg, tR = 75.3 min) and phyllofolactone G (4) (1.7 mg, tR = 86.1 min).
D.B. Abdjul et al. / Bioorg. Med. Chem. Lett. 25 (2015) 904–907 8. Phyllofolactone F (3): a colorless oil; [a]23 D +9.7 (c 0.04, CHCl3); IR (KBr) mmax 3387, 2928, 1710, 1652, 1064 cm 1; UV (CH3OH) kmax 220 (e 9442); CD (CH3CN) kextermum (De) 202 (2.6), 244 ( 3.1); EIMS m/z 414 [M]+; HREIMS m/z 414.3131 ([M]+; calcd for C27H42O3, 414.3134); 1H NMR (CDCl3) d 5.98 (1H, br s), 4.84 (1H, q, J = 6.7 Hz), 3.65 (1H, m), 2.33 (1H, m), 2.20 (1H, m), 1.93 (1H, m), 1.91 (1H, m), 1.86 (1H, m), 1.82 (1H, m), 1.74 (1H, m), 1.66 (1H, m), 1.62 (1H, m), 1.56 (1H, m), 1.48 (1H, m), 1.38 (3H, d, J = 6.7 Hz), 1.16 (2H, m), 1.12 (3H, s), 1.02 (1H, m), 0.90 (2H, m), 0.87 (3H, s), 0.86 (3H, s), 0.84 (2H, m), 0.78 (4H, br s), 0.74 (3H, t, J = 7.5 Hz); 13C NMR (CDCl3) d 174.9, 166.0, 135.5, 79.5, 75.6, 58.8, 58.4, 55.2, 42.1, 42.0, 40.2, 37.5, 37.3, 36.7, 36.1, 28.6, 25.8, 24.8, 24.5, 18.3, 18.17, 18.17, 17.2, 16.78, 16.76, 16.4, 8.6. 9. Phyllofolactone G (4): a colorless oil; [a]23 D +4.7 (c 0.10, CHCl3); IR (KBr) mmax 3385, 2927, 1711, 1652, 1065 cm 1; UV (CH3OH) kmax 220 (e 14173); CD (CH3CN) kextermum (De) 208 (2.7), 244 ( 2.3); EIMS m/z 414 [M]+; HREIMS m/z 414.3151 ([M]+; calcd for C27H42O3, 414.3134); 1H NMR (CDCl3) d 5.98 (1H, br s), 4.86 (1H, q, J = 6.7 Hz), 3.67 (1H, m), 2.37 (1H, m), 2.18 (1H, m), 1,95 (1H, m), 1.92 (1H, m), 1.87 (1H, m), 1.82 (1H, m), 1.74 (1H, m), 1.68 (1H, m), 1.62 (1H, m), 1.56 (1H, m), 1.48 (1H, m), 1.40 (3H, d, J = 6.7 Hz), 1.19 (2H, m), 1.11 (3H, s), 1.02 (1H, m), 0.89 (1H, m), 0.88 (4H, br s), 0.86 (3H, s), 0.85 (2H, m), 0.79 (4H, br s), 0.74 (3H, t, J = 7.5 Hz); 13C NMR (CDCl3) d 175.0, 165.9, 135.5, 79.1, 75.5, 58.8, 58.5, 55.0, 42.03, 42.00, 40.2, 37.5, 37.3, 36.7, 36.1, 28.5, 25.8, 24.5, 24.4, 18.4, 18.3, 18.2, 17.2, 16.9, 16.8, 16.6, 8.6. 10. Hyattellactone A (1): a colorless oil; [a]23 D +11.3 (c 0.10, CHCl3); IR (KBr) mmax 3386, 2930, 1712, 1652, 1063 cm 1; UV (CH3OH) kmax 220 (e 7837); CD (CH3CN) kextermum (De) 206 (3.0), 245 ( 2.7); EIMS m/z 414 [M]+; HREIMS m/z 414.3151 ([M]+; calcd for C27H42O3, 414.3134); 1H NMR (CDCl3) d 6.00 (1H, br s), 4.84 (1H, q, J = 6.7 Hz), 3.51 (1H, m), 2.33 (2H, m), 2.21 (1H, m), 2.18 (1H, m), 2.03 (1H, m), 2.00 (1H, m), 1.95 (1H, m), 1.85 (1H, m), 1.68 (1H, m), 1.66 (2H, m), 1.49 (4H, m), 1.41 (1H, m), 1.38 (3H, d, J = 6.7 Hz), 1.13 (4H, br s), 1.01 (4H, br s), 0.95 (2H, m), 0.90 (3H, t, J = 7.6 Hz), 0.84 (7H, br s); 13C NMR (CDCl3) d 175.0, 166.1, 135.4, 79.5, 76.5, 59.3, 58.6, 55.9, 42.7, 42.3, 42.1, 40.4, 37.6, 35.6, 34.6, 33.2, 28.7, 24.8, 22.1, 20.9, 18.6, 18.2, 17.8, 17.2, 16.6, 16.5, 10.6; 1H and 13 C NMR (Pyridine-d5), see Table 1.
907
11. Hyattellactone B (2): a colorless oil; [a]23 D +9.4 (c 0.10, CHCl3); IR (KBr) mmax 3378, 2924, 1708, 1136 cm 1; UV (CH3OH) kmax 220 (e 23290); CD (CH3CN) kextermum (De) 208 (2.0), 245 ( 1.8); EIMS m/z 414 [M]+; HREIMS m/z 414.3126 ([M]+; calcd for C27H42O3, 414.3134); 1H NMR (CDCl3) d 6.02 (1H, br s), 4.86 (1H, q, J = 6.8 Hz), 3.52 (1H, m), 2.40 (2H, m), 2.21 (1H, m), 2.18 (1H, m), 2.03 (1H, m), 2.02 (1H, m), 1.94 (1H, m), 1.87 (1H, m), 1.68 (1H, m), 1.66 (2H, m), 1.49 (4H, m), 1.41 (1H, m), 1.40 (3H, d, J = 6.8 Hz), 1.13 (4H, br s), 1.01 (4H, br s), 0.93 (2H, m), 0.90 (3H, t, J = 7.5 Hz), 0.85 (7H, br s); 13C NMR (CDCl3) d 175.1, 166.0, 135.4, 79.1, 76.3, 59.4, 58.6, 55.7, 42.7, 42.2, 42.0, 40.4, 37.6, 35.6, 34.6, 33.2, 28.8, 24.6, 22.2, 20.9, 18.5, 18.4, 17.8, 17.1, 16.70, 16.68, 10.6; 1H and 13C NMR (Pyridine-d5), see Table 1. 12. Spartan’14, Wavefunction, Irvine, CA. 13. The most stable conformer of hyattellactone A was predicted using Spartan’1412 through a preliminary conformational analysis with the MMFF94 force field, followed by geometry optimization using the density functional theory (DFT) with the B3LYP functional and 6-31G(d) basis set. The ECD spectrum in acetonitrile was calculated for the predicted most stable conformer using Gaussian 0314 by time-dependent DFT (TDDFT) with the B3LYP functional and 6-31+G(d,p) basis set. The solvent effect was introduced by the polarizable continuum model (PCM). Ten low-lying excited states were calculated (corresponding to the wavelength region down to approximately 196 nm). The calculated spectrum was displayed using GaussView 5.0.915 with the peak half-width at half height being 0.333 eV. The calculated spectrum was shifted by 15 nm to match the experimental spectrum. 14. Frisch, M. J.; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.; Robb, M. A.; Cheeseman, J. R.; Montgomery, J. A., Jr.; Vreven, T.; Kudin, K. N.; Burant, J. C., et al. Gaussian 03, revision E.1; Gaussian: Wallingford, CT, USA, 2004. 15. Dennington, R.; Keith, T.; Millam, J. GaussView, version 5; Semichem: Shawnee Mission, KS, USA, 2009. 16. De Rosa, S.; Crispino, A.; De Giulio, A.; Iodice, C.; Tommonaro, G. Tetrahedron 1998, 54, 6185. 17. Liu, J. J. Ethnopharmacol. 1995, 49, 57. 18. Mosmann, T. J. Immunol. Methods 1983, 65, 55.
