Copyright © ARKAT USA, Inc The document attached has been archived with permission from the publisher.(2S,3S)-3-[N-Cbz-L-Leucinyl]amino-2-hydroxy-4-phenylbutanoic acid 5 dimerised on treatment with EDCI and HOBT to give the symmetrical [1,4]dioxane-2,5-dione-based peptidomimetic 6.Andrew D. Abell and Andrew J. Harve

Abell, A.

Harvey, A.

Adelaide Research &amp; Scholarship

Issue in Honor of Prof. Jim Coxon ARKIVOC 2006 (iii) 72-76 Synthesis of a [1,4]dioxane-2,5-dione based-peptidomimetic scaffold  Andrew D. Abell* and Andrew J. Harvey  Department of Chemistry, University of Canterbury, Private Bag 4800, Christchurch, New Zealand Email: andrew.abell@canterbury.ac.nz  Dedicated to Jim Coxon on the occasion of his 65th birthday   Abstract (2S,3S)-3-[N-Cbz-L-Leucinyl]amino-2-hydroxy-4-phenylbutanoic acid 5 dimerised on treatment with EDCI and HOBT to give the symmetrical [1,4]dioxane-2,5-dione-based peptidomimetic 6.  Keywords: Peptidomimetic scaffold, synthesis, dimer, cyclisation    Introduction  An endless array of organic compounds exist that exhibit an equally endless array of biological activities. These compounds function by simply presenting a well-defined topology to a complementary receptor or enzyme in spite of a diversity of molecular structure and biological mechanism of action. It therefore follows that new classes of bioactives would result from simply generating new molecular topologies. One way to achieve this objective is to tether known bioactive molecules onto an appropriate molecular scaffold to generate a new, overall molecular topology. The choice of scaffold needs to allow the ready chemical attachment of a range of simple molecules that possess a diversity of structure and function, perhaps in a combinatorial fashion. The result of such a union is a compound whose overall topology is controlled by the nature and combination of its constituents. This idea has been used to generate important bioactives that contain a core scaffold and a range of attached ligands that can number anywhere from a few (e.g. protease inhibitors that possess a core isosteric unit to which are attached enzyme binding domains)1 to many (e.g. a dendrimer).2 Such hybrid compounds provide leads for important biological probes and potential therapeutics with improved potency, stability and in many cases, vastly different biological properties and mechanisms of action relative to their separate components. There is, therefore, a clear need for new and general molecular scaffolds. In this paper we present a simple example of this idea that allows a peptide, or conceivably another suitable functionalized molecule, to be ISSN 1424-6376 Page 72 ©ARKAT Issue in Honor of Prof. Jim Coxon ARKIVOC 2006 (iii) 72-76 attached at two sites on a heterocyclic scaffold (see compound 6). The compound presented here is simple to prepare as a crystalline solid, and demonstrates this principle. Closely related compounds have been reported as inhibitors of Calpains,3 an over activity of which is associated with a number of important human diseases.4   Results and Discussion  The optically active peptidomimetic scaffold 6 was prepared as detailed in the Scheme below. In the key step, a solution of (2S,3S)-3-[N-Cbz-L-leucinyl]amino-2-hydroxy-4-phenylbutanoic acid 5 in DMF was reacted with a solution of 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide hydrochloride (EDCI) and hydroxybenztriazole (HOBT) in dichloromethane containing diisopropylethylamine (DIEA) to give 6 as a stable colorless solid in 74% yield. The starting material 5 was prepared in three steps as shown in the Scheme.  The known5 α-hydroxy β-amino acid 2 was N-deprotected with HBr in acetic acid to give a mixture of 3a and 3b (9:1), which was subsequently coupled with N-Cbz-L-leucine under standard EDCI conditions to give 4a and 4b (9:1). This mixture was finally hydrolysed to give the desired derivative 5 (see step iii) in 69% over two steps. It is interesting to note that the ratio of 3a to 3b obtained in step i was dependent upon the concentration of 2 used in the reaction. Product ratios of 4:1, 9:1, 12:1 and 19:1 were obtained for concentrations of 0.26, 0.97, 1.47 and 2.00 M, respectively. The dimeric nature of 6 was confirmed by a combination of 1H NMR, 13C NMR and mass spectrometry. The two-fold axis of symmetry in the dimer 6 was confirmed by the presence of only twenty peaks in the 13C NMR spectrum while high resolution mass spectral data was consistent with the dimeric structural formula [C48H57O10N4 (M+1)+].  In addition, the 1H NMR of 6 revealed a resonance at 5.49 ppm that was shown to correspond with H2 by a 2D NMR COSY experiment - a key correlation was observed in the COSY experiment between H3 (4.66 ppm) and H2 (5.49 ppm). Importantly, the chemical shift of H2 is consistent with O-acylation6 where a similar chemical shift was observed for H2 in 4b (5.53 ppm), whereas H2 in the corresponding alcohol 4a resonated further upfield at 4.29 ppm. In this paper we have presented the synthesis of an interesting and new class of peptidomimetic molecular scaffold whose architecture should be amenable to attaching a wide range of amino acid R groups and thus the presentation of a diverse range of topologies. ISSN 1424-6376 Page 73 ©ARKAT Issue in Honor of Prof. Jim Coxon ARKIVOC 2006 (iii) 72-76 OO HNNHONHHNOOPhPhOOOOOPhPh OHCO2HNHHNPhOOOPhOHCO2MeNHPhOOPhORCO2MeBr  H3NPhORCO2MeNHHNPhOOOPh2 3a R = H3b R = Ac4a R = H4b R = Ac56i iiiiiiv23  Sheme 1. Reagents and conditions: (i) 33% HBr in AcOH, 20 min, 9 (3a):1 (3b); (ii) N-Cbz-L-Leu, EDCI, HOBT, DIEA, DMF, DCM, 16 h, 9 (4a): 1 (4b); (iii) LiOH, MeOH, H2O, 4h; (iv) EDCO, HOBT, DIEA, DMF, DCM< 16 h, 74%.   Experimental Section  General Procedures. 1H NMR spectra were obtained on a Varian Inova spectrometer, operating at 500 MHz. Coupling constants (J) are quoted in Hz. 13C NMR spectra were obtained on a Varian Unity 300 spectrometer, operating at 75 MHz, with a delay (D1) of 1 s. Electron impact (EI) mass spectra were detected on a Kratos MS80 RFA mass spectrometer operating at 4000 V (accelerating potential) and 70 eV (ionisation energy) using a source temperature of 200-250°C.  (2S,3S)-3-[N-Cbz-L-Leucinyl]amino-2-hydroxybenzenebutanoic acid (5). Compound 25 (500 mg, 1.47 mmol) was dissolved in a solution of HBr in acetic acid (33%, 1.0 ml) and the resulting mixture was stirred at rt for 20 min. The addition of ether (2 mL) caused the amine salt to precipitate. The solvent was drawn off and the resulting residue retained. To the etheral solution was added petroleum ether (4 mL) to give a further precipitate. The solvent was drawn off and the two batches of solid were combined and dried to give a mixture (397 mg, 12:1 by 1H NMR) of the amine salts 3a (86%) and 3b(6%) as a brown tar. Selected data from the mixture: 1H NMR 3a (D2O) δ 7.19-7.31 (5H, m, ArH), 4.51 (1H, d, J = 2.5 Hz, CHCHOH), 4.03 (1H, m, CHCHOH), 3.38 (3H, s, CH3), 2.89 (2H, m, CH2Ph). 1H NMR 3b (D2O) δ 7.19-7.31 (5H, m, ArH), 5.41 (1H, d, J = 1.7 Hz, CHCHOAc), 4.25 (1H, m, CHCHOAc), 3.41 (3H, s, COOCH3), 3.00 (2H, m, CH2Ph), 2.18 (3H, s, OCOCH3). To a mixture of 3a and 3b (64 mg, 9:1 by 1H NMR, 0.22 mmol) were added EDCI (47 mg, 0.25 mmol) and HOBT (32 mg, 0.24 mmol). A solution (1.0 M) of N-Cbz-L-leucine in N-methylpyrrolidinone (240 µL, 0.24 mmol), DMF (250 µL) and DIEA (43 µL, 0.25 mmol) were then added and the reaction mixture was stirred overnight at rt under nitrogen. The reaction ISSN 1424-6376 Page 74 ©ARKAT Issue in Honor of Prof. Jim Coxon ARKIVOC 2006 (iii) 72-76 mixture was diluted with ethyl acetate (4 mL) and washed with 10% aqueous HCl (3 x 2 mL), saturated NaHCO3 (3 x 2 mL), brine (2 mL), dried over MgSO4 and concentrated in vacuo to give a mixture (9:1 by 1H NMR) of 4a and 4b (69 mg, 69%) as a white solid. Data from the mixture: [MS (FAB): 499 [(M+1)+ 4b], 457 [(M+1)+ 4a], 414, 307, 210, 154, 136]; mp 138-144 °C; IR (CHCl3) 3688, 3416, 2961, 1734, 1676, 1601, 1500, 1440 cm-1. 1H NMR 4a (CDCl3) δ 7.14-7.35 (10H, m, ArH), 6.51 (1H, d, J = 8.8 Hz, PheNH), 5.09 (3H, m, CbzCH2 & CbzNH), 4.51 (1H, ddd, J = 2.9, 7.3, 16.1 Hz, CHCHOH), 4.29 (1H, br s, CHCHOH), 4.11 (1H, m, NHCHCO), 3.59 (3H, s, OCH3), 2.79 (2H, m, PheCH2), 1.61 (3H, m, LeuCH & LeuCHA), 1.39 (1H, m, LeuCHB), 0.89 (6H, m, 2 x LeuCH3); 13C NMR 4a (CDCl3) δ 172.8 (COOMe), 172.0 (NHCHCO), 156.2 (CbzCO), 136.7 (PheArC), 136.0 (CbzArC), 129.3, 128.5, 128.3, 128.3, 128.1, 126.7 (ArCH), 72.0 (CHOH), 67.2 (CbzCH2), 53.7 (NHCHCO), 53.0 (CHCHOH), 52.5 (COOCH3), 41.0 (LeuCH2), 35.4 (PheCH2), 24.6 (LeuCH), 22.8 (Leu(CH3)A), 21.9 (Leu(CH3)B). 1H NMR 4b (CDCl3) δ 6.42 (1H, m, PheNH), 5.53 (1H, s, CHOAc), 3.63 (3H, s, COOCH3), 2.16 (3H, s, OCOCH3). To the mixture of 4a and 4b (60 mg, 0.13 mmol, 9:1 by 1H NMR) at 0 °C was added a solution of 0.25 M lithium hydroxide (1.5 parts) in 2:1 methanol/water. After the reaction mixture was stirred for 0.5-4 h, sufficient 1N aqueous HCl was added to acidify the solution (pH 5 by Universal Indicator) and the methanol was evaporated in vacuo. Water was added to the residue and the organics were extracted with ethyl acetate. The combined organic extracts were washed with water, brine, dried over MgSO4 and concentrated in vacuo to the acid 5 (58 mg, qu) as a white solid. 1H NMR (CD3OD) δ 7.82 (1H, d, J = 9.3 Hz, CbzNH), 7.33 (5H, m, ArH), 7.10-7.19 (5H, m, ArH), 5.07 (2H, m, CbzCH2), 4.49 (1H, m, CHCHOH), 4.17 (1H, d, J = 4.4 Hz, CHCHOH), 4.07 (1H, m, NHCHCO), 2.80 (2H, m, PheCH2), 1.58 (1H, m, LeuCH), 1.33 (2H, m, LeuCH2), 0.88 (6H, m, 2 x LeuCH3); 13C NMR (CD3OD) δ 175.8 & 175.2 (CHCONH & COOH), 158.6 (CbzCO), 139.6 (PheArC), 138.4 (CbzArC), 130.7, 129.8, 129.6, 129.3, 129.2, 127.6 (ArCH), 73.9 (CHOH), 68.0 (CbzCH2), 55.4 (NHCHCO), 54.8 (CHCHOH), 42.5 (LeuCH2), 36.4 (PheCH2), 26.1 (LeuCH), 23.6 (Leu(CH3)A), 22.2 (Leu(CH3)B); [MS (FAB): 465 (M+Na)+, 443, 399, 286, 176, 153, 136; HRMS (FAB) calcd for C24H30O6N2Na (M+Na)+ 465.2002, found 465.1996]; mp 131-134 °C; IR (CHCl3) 3433, 2963, 1720, 1660, 1510, 1456, 1340, 1217 cm-1. Dimer 6. To a solution of 5 (14 mg, 32 µmol) in DMF (1 mL) was added EDCI (8 mg, 41 µmol), HOBT (7 mg, 47 µmol), dichloromethane (1 mL) and DIEA (6 µL, 35 µmol). The reaction mixture was stirred overnight at rt before being diluted with dichloromethane (10 mL), washed with 10% aqueous HCl (10 mL), brine (10 mL), dried over MgSO4, and concentrated in vacuo to afford 6 (10 mg, 74%) as a white solid. 1H NMR (300Mz, DMSO) δ 8.28 (2H, d, J = 8.2 Hz, PheNH), 7.16-7.35 (22H, m, arom & CbzNH), 5.49 (2H, brs, H2), 5.01 (4H, s, CbzCH2), 4.66 (2H, m, H3), 4.05 (2H, m, Leu-αH), 2.81 (4H, m, PheCH2), 1.49 (2H, m, LeuCH), 1.36 - 1.27 (4H, m, LeuCH2), 0.81 (12H, m, LeuCH3); 13C NMR (DMSO) δ 172.1 (ring CO), 165.7 (LeuCO), 155.4 (CbzCO), 137.6, 136.7, 128.7, 128.1, 128.0, 127.6, 127.5, 126.1, 77.6 (C2), 65.2 (CbzCH2), 52.8 (Leu-αC), 49.9 (C3), 40.7 (LeuCH2), 33.8 (PheCH2), 23.9 (LeuCH), 22.6 ISSN 1424-6376 Page 75 ©ARKAT Issue in Honor of Prof. Jim Coxon ARKIVOC 2006 (iii) 72-76 ISSN 1424-6376 Page 76 ©ARKAT (LeuCH3), 21.3 (LeuCH3); [MS (ES): 871 (M+Na)+, 849 (M+1)+, 647, 556, 537, 464, 316, 289, 275, 249; HRMS (ES) calcd for C48H57O10N4 (M+1)+ 849.408, found 849.405]; RF 0.19 (1:4 ethyl acetate-dichloromethane); IR (CHCl3) 3628, 3354, 1767, 1722, 1683, 1228 cm-1; [α]D - 78 ° (c 0.082 acetonitrile).   Acknowledgements  The financial support of a Royal Society of New Zealand Marsden grant is gratefully acknowledged.   References  1. (a)Abdel-Rahman, H. M.; Al-karamany, G. S.; El-Koussi, N. A.; Youssef, A. F.; Kiso, Y.; Curr. Med. Chem. 2002, 9, 1905. (b)Lam, P. Y. S.; Jadhav, P. K.; Eyermann, C. J.; Hodge, C, N.; Ru, Y.; Bacheler, L. T.; Meek, J. L.; Otto, M. J.; Rayner, M. M.; Wong, Y. N.; Chang, C. H.; Weber, P. C.; Jackson, D. A.; Sharpe, T. R.; Ericksonviitanen, S. Science 1994, 263, 380. 2. (a)Boas, U.; Heegaard, P. M. H. Chem. Soc. Rev. 2004, 33, 43. (b) Dykes, G. M.; J. Chem. Technol. Biotechnol. 2001, 76, 903. 3. Alvarez, M. E.; Houck, D. R.; White, C. B.; Brownell, J. E.; Bobko, M. A.; Rodger, C. A.; Stawicki, M. B.; Sun, H. H.; Gillum, A. M.; Cooper, R. J. Antibiot. 1994, 47, 1195. 4. (a)Lin, G. D.; Chattopadhyay, D.; Maki, M.; Wang, K. K. W.; Carson, M.; Jin, L.; Yuen, P. W.; Takano, E.; Hatanaka, M.; DeLucas, L. J.; Narayana, S. V. L. Nat. Struct. Biol. 1997, 4, 539. (b) Chatterjee, S.; Gu, Z.-Q.; Dunn, D.; Tao, M.; Josef, K.; Tripathy, R.; Bihovsky, R.; Senadhi, S. E.; O'Kane, T. M.; McKenna, B. A.; Mallya, S.; Ator, M. A.; Bozyczko-Coyne, D.; Siman, R.; Mallamo, J. P. J. Med. Chem. 1998, 41, 2663. (c) Mathur, P.; Gupta, S. K.; Wegener, A. R.; Breipohl, W.; Ahrend, M. H.; Sharma, Y. D.; Gupta, Y. K.; Vajpayee, R. B. Curr. Eye Res. 2000, 21, 926. 5. Wasserman, H. H.; Ho, W.-B. J. Org. Chem. 1994, 59, 4364. 6. Kemp, W. Organic Spectroscopy; MacMillan: London, 1991; 3rd Edn., p. 171.  

Synthesis of a [1,4]dioxane-2,5-dione based-peptidomimetic scaffold

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Synthesis of a [1,4]dioxane-2,5-dione based-peptidomimetic scaffold

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Synthesis of a [1,4]dioxane-2,5-dione based-peptidomimetic scaffold

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