Antitubercular Bis-Substituted Cyclam Derivatives - ACS Publications

Mar 20, 2018 - Tuberculosis Research Program, Centenary Institute, Royal Prince Alfred Hospital, Missenden Road, Camperdown, Sydney, NSW. 2050 ...
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Article Cite This: J. Med. Chem. 2018, 61, 3595−3608

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Antitubercular Bis-Substituted Cyclam Derivatives: Structure− Activity Relationships and in Vivo Studies Malcolm Spain,† Joseph K.-H. Wong,† Gayathri Nagalingam,‡ James M. Batten,† Elinor Hortle,§ Stefan H. Oehlers,∥ Xiao Fan Jiang,† Hasini E. Murage,† Jack T. Orford,† Patrick Crisologo,† James A. Triccas,‡ Peter J. Rutledge,*,† and Matthew H. Todd*,† †

School of Chemistry, The University of Sydney, Sydney, NSW 2006, Australia Microbial Immunity and Pathogenesis Group, Department of Infectious Diseases and Immunology, Sydney Medical School, The University of Sydney, Sydney, NSW 2006, Australia § Tuberculosis Research Program, Centenary Institute, Royal Prince Alfred Hospital, Missenden Road, Camperdown, Sydney, NSW 2050, Australia ∥ Central Clinical School, Sydney Medical School, The University of Sydney, Sydney, NSW 2006, Australia ‡

S Supporting Information *

ABSTRACT: We recently reported the discovery of nontoxic cyclam-derived compounds that are active against drug-resistant Mycobacterium tuberculosis. In this paper we report exploration of the structure−activity relationship for this class of compounds, identifying several simpler compounds with comparable activity. The most promising compound identified, possessing significantly improved water solubility, displayed high levels of bacterial clearance in an in vivo zebrafish embryo model, suggesting this compound series has promise for in vivo treatment of tuberculosis.

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INTRODUCTION Antimicrobial resistance represents a current worldwide threat to human health and an increasing burden upon healthcare.1−4 An estimated 1.7 billion people were infected with latent tuberculosis (TB) in 2014, leading to 10.4 million incident cases and 1.8 million deaths in 2015.5,6 In the treatment of TB, multidrug therapy of first-line drugs is used over long periods of time (typically 6 months) to address the challenge of resistance.7 However, poor adherence to the long treatment regimens has been implicated in the rise of drug resistant TB strains.7,8 In 2015, 480 000 cases of multidrug-resistant TB (MDR-TB) were reported, requiring longer and more expensive treatment programs.6,7 Furthermore, mismanagement of MDR-TB has led to extensively drug-resistant TB (XDR-TB) being reported in 117 countries.7 The rise in drugresistant forms of TB is a global public health crisis and requires the urgent development of new small molecule drugs.7,9,10 During M. tuberculosis infection in humans, host phagocytes take up bacteria, which can replicate within these cells; as the infection progresses, aggregation of further immune cells leads to granulomas where bacteria can remain for long periods of time.11 Commonly, TB results from reactivation of established granulomas in individuals infected with latent TB, with a 10% chance of activation over a lifetime.11 As such, granulomas represent an important and characteristic feature of TB infections. Recently, zebrafish (Danio rerio) infected with © 2018 American Chemical Society

Figure 1. Most important hit compound 1 against M. tuberculosis (for our previous study see ref 13), a bis-substituted cyclam core with pendant naphthalimide groups linked through 1,2,3-triazoles.

Mycobacterium marinum, a close relative of M. tuberculosis, have been used to study mycobacterial granulomas. The translucency of the zebrafish allows the infection, and hence the impact of treatment, to be observed over time. The zebrafish-M. marinum infection platform is thus amenable for investigating the efficacy of novel antibiotic compounds within an in vivo infection context.12 We have shown the promising activity of cyclam-based ligands and their copper and zinc complexes against virulent and drug-resistant M. tuberculosis strains.13 These compounds are nontoxic and active against intracellular M. tuberculosis. Herein we present our progress in determining the structure− Received: January 16, 2018 Published: March 20, 2018 3595

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

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Scheme 1. Synthesis of Compounds To Investigate the Pendant Groups

Figure 2. Pendant group SAR. Potencies are MIC vs H37Rv using concentration of compound as determined by the resazurin viability assay (see the Experimental Section for details).

Scheme 2. Synthesis of compounds To Investigate the Linker Group

Figure 3. Linker group SAR. Potencies are MIC vs H37Rv.

To investigate the pendant groups, compounds 5a−h were synthesized through copper-catalyzed azide−alkyne cycloaddition (CuAAC) between bis-alkyne 2 and the corresponding azides 3a−h to yield 4a−h followed by Boc deprotection (Scheme 1). Naphthalimides represent key structural features in a range bioactive compounds14 including antitumor, antiviral, and antibacterial compounds, while their fluorescent properties make them widely used probes.13,15 However, inclusion of the naphthalimide group can lead to compounds of low water

activity relationship (SAR) of the cyclam compound 1 (Figure 1), which possesses a cyclic cyclam core linked to two identical naphthalimide pendants via triazole linkers.

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RESULTS AND DISCUSSION

The SAR was determined through the symmetric modification of the two pendant groups of 1, the linker groups, and investigations into the impact of metal complexation upon biological activity. 3596

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

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Table 1. Activity of Metal Complexes 14a−g and 15a,b

compd d

1 14ad 14bd 14c 14d 14e 14f 14g 5a 15a 15b

complexationa

ionic radius (Å)b

MIC vs H37Rv (μM)c

n/a 1 + Zn(ClO4)2 1 + Cu(ClO4)2 1 + Fe(ClO4)2 1 + Fe(ClO4)3 1 + Sm(ClO4)3 1 + Co(ClO4)2 1 + Mn(ClO4)2 n/a 5a + Zn(ClO4)2 5a + Cu(ClO4)2

n/a 0.74 0.73 0.61 0.55 0.96 0.65 0.67 n/a 0.74 0.73

3.13−6.25 3.13−6.25 6.25 3.13 3.13 1.56−3.13 3.13−6.25 3.13 25−50 3.13 3.13

a

All compounds were complexed as detailed in the Experimental Section. bIonic radii based on the hexacoordinate metals.19 cSee Experimental Section for details. dSee ref 13 for details.

Table 2. Activity of Zn2+ and Cu2+ Metal Complexes (16a− 20b) of Compounds 9a−e

solubility. In an attempt to lower molecular weight and increase water solubility relative to parent compound 1, a range of pendant groups was tested, maintaining the cyclam core and triazole linkers (Figure 2). Pleasingly replacing the naphthalimide in 1 (MIC = 3.13 μM) with simple naphthalene-based pendant groups (5a,b) maintained some of the superior activity of the naphthalimide.16 However, single aromatic ring pendants 5c−h15c including a diversity of aromatic electron density, with or without a methylene spacer, exhibited low potency. Thus, while the naphthalimide can be replaced by simpler 1-naphthyl groups, single aromatic rings were not found to be suitable replacements. Due to the ease of synthesis and promising activity of simple naphthyl groups, the 1-naphthyl pendant group was maintained while other linking groups were investigated. Compound 9a was synthesized through CuAAC between bis-azide 6 and the corresponding alkyne 7a to yield 8a followed by Boc deprotection (Scheme 2). To investigate the simple alkyl linker groups, compound 13 was synthesized through alkylation of 10 with bromide 11 followed by hydrolysis and hydrogenation of intermediate 12 to yield 13 (Scheme 2). Interestingly, the alkane linker in 13 maintained similar bioactivity (25−50 μM) versus the triazole-based compound 5a

compd

complexationa

MIC vs H37Rv (μM)b

9a 16a 16b 9bc 17ac 17bc 9c 18a 18b 9dc 19ac 19bc 9e 20a 20b

n/a 9a + Zn(ClO4)2 9a + Cu(ClO4)2 n/a 9b + Zn(ClO4)2 9b + Cu(ClO4)2 n/a 9c + Zn(ClO4)2 9c + Cu(ClO4)2 n/a 9d + Zn(ClO4)2 9d + Cu(ClO4)2 n/a 9e + Zn(ClO4)2 9e + Cu(ClO4)2

6.25 6.25 3.13 25 25 6.25 6.25 3.13 3.13 50 50 6.25 >100 >100 6.25

a

Free amines were complexed with the relevant metal perchlorate salts detailed in the Experimental Section. bSee Experimental Section for details. cSee ref 15c for synthetic details.

(Figure 3), suggesting that flexible and noncoordinating linker groups can maintain bioactivity. Importantly, introducing an extra methylene group and changing the 1,2,3-triazole connectivity to be C-linked to the pendant group in 9a resulted in a more potent compound (6.25 μM), suggesting that positioning of the pendants, as opposed to the flexibility of the linker, is a key determinant of potency. Previous medicinal chemistry studies on the treatment of HIV with cyclam-derived compounds noted that the coordinating metal ion impacted on biological activity.17 Thus, we investigated a variety of complexes of 1 (Table 1) using metal

Figure 4. Pendant group SAR with the alternative linker topology. 3597

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

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Table 3. Effect of N-Functionalization on Water Solubility and Biological Potency

compd

complexationa

thermodynamic solubility at pH 7.0 (mM)b

MIC vs H37Rv (μM)c

14ad 21a 21b

1 + Zn(ClO4)2 5i + Zn(ClO4)2 5i + ZnCl2

97% from Sigma-Aldrich, contains approximately 17% of 2(chloromethyl)naphthalene by 1H NMR) and NaN3 (0.745 g, 11.5 mmol, 1.8 equiv) in MeCN (10 mL, 0.6 M) were refluxed for 17 h. The reaction was carefully concentrated, diluted with H2O (10 mL) and extracted with diethyl ether (3 × 10 mL), and the organic layers were dried over MgSO4 and carefully concentrated to yield 3b as a colorless oil (0.770 g, 4.2 mmol, 64%, contains 2-(azidomethyl)naphthalene impurities that could not be separated by HPLC). 1H NMR (300 MHz, CDCl3): δ 4.71 (s, 2 H), 7.41−7.57 (m, 4 H), 7.81− 7.87 (m, 2 H), 8.00 (d, J 8.0, 1 H). LRMS (ESI+): m/z Not found. 1H NMR matched those previously described.23 1-Azido-4-nitrobenzene (3d). Similar to the procedure for 3a, 4nitroaniline (443 mg, 3.21 mmol, 1.0 equiv), H2O (2 mL), conc HCl (1 mL), NaNO2 (289 mg, 4.19 mmol, 1.3 equiv) in H2O (1 mL), NaN3 (251 mg, 3.87 mmol, 1.2 equiv) in H2O (1 mL) for 1 h extracted, dried over Na2SO4, and concentrated to yield 3d as a yellow solid (525 mg, 3.20 mmol, 99%). 1H NMR (300 MHz, CDCl3): δ 7.10−7.20 (m, 2 H), 8.24−8.30 (m, 2 H). 13C NMR (50 MHz, CDCl3): δ 119.4, 125.6, 144.7, 146.9. FTIR (ATR) νmax/cm−1: 2120, 2100, 1611. Spectroscopic properties matched those previously described.21 1-Azido-4-methoxybenzene (3e). Similar to the procedure for 3a, p-anisidine (396 mg, 3.21 mmol, 1.0 equiv), H2O (2 mL), conc HCl (1 mL), NaNO2 (289 mg, 4.19 mmol, 1.3 equiv) in H2O (1 mL), NaN3 (251 mg, 3.87 mmol, 1.2 equiv) in H2O (1 mL) for 1 h

CONCLUSIONS We have shown through SAR that simple naphthyl groups can be used as replacements for naphthalimides in this compound series and that the activity is relatively insensitive to the linker groups tested. Importantly, the metal coordinating ability of the triazole is not essential. This links to the observation that in the primary series of compounds 14a−g, activity is apparently insensitive to the coordinated metal ion or its identity. However, Cu2+ coordination does increase the activity of several compounds of the “reversed” triazole geometry 16a− 20b, and thus Cu2+ may be involved in the bioactivity of these compounds. Encouragingly, progression of the hit compound 1 to our current lead compound 21b with good water solubility has shown that these compounds reduce bacterial load in vivo. Combining the in vivo activity of this unusual class of antitubercular drugs with our previous demonstration of their effectiveness against drug-resistant M. tuberculosis leads us to prioritize further optimization of potency alongside studies toward understanding the mechanism of action.

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EXPERIMENTAL SECTION

General Procedures. Acetonitrile, methanol, and THF were collected from a PureSolv MD 7 solvent purification system with anhydrous alumina columns. All commercially available reagents and solvents were purchased from Sigma-Aldrich, Alfa Aesar, Matrix Scientific, Merck, or Ajax Finechem and used without purification. Flash column chromatography was performed on Davisil Grace Davison 40−63 μm (230−400 mesh) silica gel. Automated flash column chromatography was performed on a Biotage Isolera Spektra One using Biotage SNAP KP-Sil cartridges (filled with Grace 40−63 μm (230−400 mesh) silica gel) at their default flow rates. Reversedphase automated chromatography was carried out on a Biotage Isolera Spektra One using Biotage SNAP KP-C18-HS (30 g) column using a flow rate of 25 mL/min; mobile phases of 0.1% TFA in Milli-Q water and 0.1% TFA in MeCN, 0−100% over 10 CV were used unless stated otherwise. Preparative reversed-phase high performance liquid chromatography (HPLC) was carried out on a Waters 600 controller with a Waters 600 pump and a 2998 photodiode array (PDA) detector. A Waters SunFire C18 OBD preparative 217 column (5 μm, 19 mm × 150 mm) was used at a flow rate of 7 mL/min; mobile phases of 0.1% TFA in Milli-Q water and 0.1% TFA in MeCN in different ratios were used. Analytical HPLC analysis was conducted on a Waters Alliance 2695 instrument using a SunFire C18 column (5 μm, 2.1 mm × 150 mm) with a Waters 2996 PDA detector and a flow rate of 0.2 mL/min; mobile phases of 0.1% TFA in Milli-Q water and 0.1% TFA in MeCN, 0−100% over 90 min, were used. Melting points were recorded on a Stanford Research Systems Optimelt automated melting system and are uncorrected. 1H and 13C NMR spectra were recorded on a Bruker Avance DPX 200, 300, 400, or 500 MHz spectrometer. Chemical shifts are reported in ppm relative to tetramethylsilane or residual solvent resonance as internal standard. Coupling constants J are reported in Hz to the nearest 0.1 Hz. Mass spectra (LRMS) were acquired on a Thermo Classic LCQ mass spectrometer or a Bruker amaZon SL mass spectrometer. Accurate mass measurements (HRMS) were performed on a Bruker Apex Qe 7T Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometer. Infrared spectra were recorded on a Bruker ALPHA FT-IR spectrophotometer (ZnSe or diamond ATR). Elemental analyses were carried out by the Campbell Microanalytical Laboratory (University of Otago, New Zealand) on a Carlo Erba EA 1108 elemental analyzer or by the Chemical Analysis Facility (Macquarie University, Australia) on a PerkinElmer PE2400 CHN elemental analyzer. Synthesis and characterization of compounds 1 and 14a,b have been reported previously in ref 13 and their purities determined through elemental analysis; additionally compounds 5c, 9b, 9d, 17a,b, 19a,b were previously reported in ref 15c and their purities determined through elemental analysis. Elemental analysis or analytical HPLC was 3599

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

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mmol, 2.4 equiv) and 2 (591 mg, 1.25 mmol, 1 equiv), then THF (13 mL) and H2O (13 mL) (0.16 M of cyclam in the 1:1 mixture of THF/ H2O), and the reaction mixture was backfilled with N2 (×5, fast), CuSO4·5H2O (63 mg, 20 mol %), and sodium ascorbate (110 mg, 50 mol %) was added and backfilled with N2 (× 3, fast), and the reaction mixture was stirred at rt for 20 h. The reaction mixture was concentrated to remove THF and diluted with aqueous saturated NH4Cl (20 mL) and extracted with DCM (3 × 50 mL). The combined organic layers were dried over Na2SO4 and concentrated by rotary evaporation to yield the crude product which was purified using automated flash chromatography 0−100% EtOAc−petroleum benzene over 10 CV, to yield 4a as a brown foamy solid (584 mg, 58%). Mp 85−96 °C. 1H NMR (200 MHz, CDCl3): δ 1.34−1.40 (br s, 18 H), 1.70−2.00 (m, 4 H), 2.60 (t, J 6.0, 4 H), 2.72 (t, J 6.0, 4 H), 3.24−3.61 (m, 8 H), 3.91 (s, 4 H), 7.39−7.67 (m, 12 H), 7.85−8.05 (m, 4 H). 13 C NMR (75 MHz, CDCl3): δ 26.5, 28.4, 46.9, 50.1, 51.9, 53.6, 61.9, 79.3, 122.3, 124.7, 124.9, 126.9, 127.8, 128.2, 128.4, 130.1, 133.7, 134.0, 140.4, 155.7 (1× quaternary aromatic signal not seen or overlapping). LRMS (ESI+): m/z 815.49 ([M + H]+, 100%). Di-tert-butyl 4,11-Bis((1-(4-nitrophenyl)-1H-1,2,3-triazol-4yl)methyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (4d). Similar to the procedure for 4a. 3d (0.84 mmol, 137.2 mg, 2.09 equiv) and 2 (238 mg, 0.4 mmol, 1 equiv) in THF (1.7 mL) and H2O (1.7 mL), CuSO4·5H2O (20 mg, 20 mol %) and sodium ascorbate (35 mg, 50 mol %) and stirred at rt for 1 h. The reaction mixture was concentrated and purified by flash chromatography (0− 100% DCM−MeOH, over 5 CV to remove azide), then 100% MeOH (5 CV)) to yield 4d as an orange foamy solid (171 mg, 0.21 mmol, 53%). 1H NMR (500 MHz, CDCl3): δ 1.19−1.53 (m, 18 H), 1.76− 1.86 (br m, 4 H), 2.53−2.59 (br m, 4 H), 2.60−2.71 (br m, 4 H), 3.26−3.60 (m, 8 H), 3.86 (br s, 4 H), 7.86−8.15 (m, 6 H), 8.26−8.47 (m, 4 H). 13C NMR (126 MHz, CDCl3): δ 27.0, 28.4, 46.9, 47.5, 50.2, 52.2, 54.5, 79.5, 120.3, 120.7, 125.4, 141.1, 147.0, 147.2, 155.9. LRMS (ESI+): m/z 805.46 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C38H53N12O8+ 805.4109, found 805.4109. Di-tert-butyl 4,11-Bis((1-(4-methoxyphenyl)-1H-1,2,3-triazol-4-yl)methyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (4e). Similar to the procedure to 4a. 3e (0.84 mmol, 125 mg, 2.09 equiv) and 2 (238 mg, 0.4 mmol, 1 equiv) in THF (1.7 mL) and H2O (1.7 mL), CuSO4·5H2O (20 mg, 20 mol %) and sodium ascorbate (35 mg, 50 mol %) and stirred at rt for 1 h. The reaction mixture was concentrated and purified by flash chromatography (0− 100% DCM−MeOH, over 5 CV to remove excess 3e) and 100% MeOH (5 CV) to yield 4e as a yellow-green solid (140 mg, 0.18 mmol, 45%). 1H NMR (500 MHz, CDCl3): δ 1.22−1.50 (br s, 18 H), 1.82−1.86 (br m, 4 H), 2.51−2.65 (br m, 4 H), 2.63−2.71 (br m, 4 H), 3.26−3.58 (m, 8 H), 3.81−3.86 (br m, 10 H), 6.90−6.95 (br m, 4 H), 7.56−7.62 (br m, 4 H), 7.74−7.80 (br s, 2 H). 13C NMR (126 MHz, CDCl3): δ 26.9, 28.4, 46.8, 47.4, 50.0, 52.0, 55.5, 79.6, 114.6, 115.1, 121.9, 122.1, 130.1, 155.9, 159.6. LRMS (ESI+): m/z 775.51 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C40H59N10O6+ 775.4619 found 775.4620. Di-tert-butyl 4,11-Bis((1-benzyl-1H-1,2,3-triazol-4-yl)methyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (4f). Similar to the procedure for 4a, 3f (0.84 mmol, 160 mg, 2.09 equiv) and 2 (238 mg, 0.4 mmol, 1 equiv) in THF (1.7 mL) and H2O (1.7 mL) were added CuSO4·5H2O (20 mg, 20 mol %) and sodium ascorbate (35 mg, 50 mol %) and stirred at rt for 1 h. The reaction mixture was concentrated and purified by flash chromatography (0− 15% DCM−MeOH, over 10 CV) to yield 4f as a white foamy solid (95 mg, 0.13 mmol, 32%). 1H NMR (500 MHz, CDCl3): δ 1.39 (br s, 18 H), 1.62−1.76 (m, 4 H), 2.35−2.48 (m, 4 H), 2.49−2.64 (m, 4 H), 3.14−3.41 (m, 8 H), 3.62−3.81 (m, 4 H), 5.49 (d, J 14.5, 4 H), 7.24 (br s, 4 H), 7.31−7.38 (br m, 8 H). 13C NMR (126 MHz, CDCl3): δ 26.5, 28.4, 46.4, 47.0, 50.4, 52.0, 53.6, 54.0, 79.2, 122.3, 127.9, 128.6, 129.0, 134.9, 145.0, 155.7. LRMS (ESI+): m/z 743.20 ([M + H]+, 100%). Di-tert-butyl 4,11-Bis((1-(4-nitrobenzyl)-1H-1,2,3-triazol-4yl)methyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (4g). Similar to the procedure for 4a, 3g (0.84 mmol, 150 mg, 2.09 equiv) and 2 (238 mg, 0.4 mmol, 1 equiv) in THF (1.7 mL) and

extracted, dried over Na2SO4, and concentrated to yield 3e as a dark brown solid (319 mg, 2.15 mmol, 67%). 1H NMR (300 MHz, CDCl3): δ 3.82 (s, 3 H), 6.90−6.94 (m, 2 H), 6.96−7.00 (m, 2 H). 13 C NMR (50 MHz, CDCl3): δ 55.6, 115.2, 120.0, 132.4, 157.1. FTIR (ATR) νmax/cm−1: 2108, 1602. Spectroscopic properties matched those previously described.21 Azidomethylbenzene (3f). Based on the literature procedure.24 To a solution of benzyl bromide (1 mL, 8.4 mmol, 1 equiv) in acetone (1 mL) and H2O (1 mL) was added NaN3 (1.9 g, 30.2 mmol, 3.6 equiv). The reaction mixture was stirred at rt for 2 h. The reaction was diluted with H2O (10 mL) and extracted with Et2O (3 × 10 mL) and dried over Na2SO4 and concentrated to yield 3f as a colorless oil (770 mg, 5.8 mmol, 69%). 1H NMR (300 MHz, CDCl3): δ 4.37 (s, 2 H), 7.32−7.45 (m, 5 H). 13C NMR (50 MHz, CDCl3): δ 54.8, 128.3, 128.4, 128.9, 135.5. FTIR (ATR) νmax/cm−1: 2089, 1605. Spectroscopic properties matched those previously described.25 1-(Azidomethyl)-4-nitrobenzene (3g). Similar to the procedure for 3f, 4-nitrobenzyl bromide (416 mg, 2 mmol, 1 equiv), NaN3 (484 mg, 7.4 mmol, 3.7 equiv), acetone:H2O (2 mL, 3:1) for 2 h. The reaction was diluted with H2O (10 mL), extracted with Et2O (3 × 20 mL), and dried over Na2SO4 and concentrated to yield 3g as a colorless oil (332 mg, 1.86 mmol, 93%). 1H NMR (300 MHz, CDCl3): δ 7.52 (d, J 9.0, 2 H), 8.21 (d, J 9.0, 2 H). 13C NMR (50 MHz, CDCl3): δ 53.6, 123.8, 128.4, 142.6, 147.6. FTIR (ATR) νmax/ cm−1: 2107, 1607. Spectroscopic properties matched those previously described.26 1-(Azidomethyl)-4-methoxybenzene (3h). Following a literature procedure,27 to a vial charged with 4-methoxybenzyl chloride (313 mg, 2.0 mmol, 1 equiv) and NaN3 (484 mg, 7.4 mmol, 3.7 equiv) was added THF (2 mL) and stirred at rt for 1 h. The reaction was diluted with H2O (10 mL), extracted with Et2O (3 × 20 mL), dried over Na2SO4, and concentrated to yield 3h as a colorless oil (280 mg,1.88 mmol, 94%). 1H NMR (300 MHz, CDCl3): δ 3.78 (s, 3 H), 4.23 (s, 2 H), 6.89 (d, J 8.5, 2 H), 7.22 (d, J 8.5, 2 H). 13C NMR (50 MHz, CDCl3): δ 54.4, 55.3, 114.2, 127.4, 129.6, 156.6. FTIR (ATR) νmax/cm−1: 2096. Spectroscopic properties matched those previously described.27 6-Azido-2-(3-hydroxypropyl)-1H-benzo[de]isoquinoline1,3(2H)-dione (3i). Following a literature 2-step procedure,15a to a suspension of 4-bromo-1,8-naphthalic anhydride (7.5 g, 27.1 mmol, 1 equiv) in ethanol (0.54 M, 50 mL) was added 3-aminopropan-1-ol (2.1 mL, 27.1 mmol, 1.0 equiv) and heated to reflux for overnight. The reaction was cooled to rt and filtered, collecting to yield the intermediate 6-bromo-2-(3-hydroxypropyl)-1H-benzo[de]isoquinoline-1,3(2H)-dione as a light brown powder (8.6 g, 25.7 mmol, 95%). 1H NMR (300 MHz, CDCl3): δ 2.00 (app quin, J 6.0, 2 H), 2.96 (t, J 6.0, 1 H), 3.61 (app q, J 6.0, 2 H), 4.36 (t, J 6.0, 2 H), 7.88 (dd, J 8.5, 7.0, 1 H), 8.07 (d, J 8.0, 1 H), 8.45 (d, J 8.0, 1 H), 8.61 (dd, J 8.5, 1.0, 1 H), 8.69 (dd, J 7.0, 1.0, 1 H). 13C NMR (75 MHz, CDCl3): δ 31.0, 37.0, 58.9, 121.7, 122.6, 128.1, 128.8, 130.5, 130.6, 131.1, 131.4, 132.2, 133.4, 164.1, 164.3. FTIR (ATR) νmax/cm−1: 1701, 1655. Spectroscopic properties matched those previously described.15a The intermediate was then converted to the azide product using a reaction carried out at a maximum of 3.0 g scale for safety reasons. To 6-bromo-2-(3-hydroxypropyl)-1H-benzo[de]isoquinoline-1,3(2H)dione (3.00 g, 9.0 mmol, 1.0 equiv) in NMP (10 mL, 0.9 M) was added NaN3 (3.27 g, 50.2 mmol, 5.6 equiv), and the mixture was heated to 110 °C for 45 min and cooled to rt. The reaction mixture was diluted with water (60 mL) and filtered collecting the precipitate which was dried under high vacuum to yield 3i as a brown solid (2.58 g, 0.87 mmol, 97%). 1H NMR (300 MHz, CDCl3): δ 1.85−1.97 (m, 2 H), 3.52 (t, J 5.5, 2 H), 4.26 (t, J 6.0, 2 H), 7.39 (d, J 8.0, 1 H), 7.67 (m, 1 H), 8.37 (d, J 8.5, 1 H), 8.50 (d, J 8.0, 1 H), 8.56 (d, J 7.0, 1 H). 13 C NMR (75 MHz, CDCl3): δ 30.9, 36.8, 58.9, 114.7, 118.4, 122.2, 124.2, 124.3, 126.9, 129.0, 132.0, 132.5, 143.3, 162.3, 164.5. FTIR (ATR) νmax/cm−1: 2125, 1697, 1654, 1589. Spectroscopic properties matched those previously described.15a Di-tert-butyl 4,11-bis((1-(naphthalen-1-yl)-1H-1,2,3-triazol4-yl)methyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (4a). To a flask charged with N2 were added 3a (511 mg, 3.02 3600

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

Journal of Medicinal Chemistry

Article

H2O (1.7 mL) and CuSO4·5H2O (20 mg, 20 mol %) and sodium ascorbate (35 mg, 50 mol %) for 1 h at rt. The reaction mixture was concentrated and purified by flash chromatography (0−15% DCM− MeOH, over 10 CV) to yield 4g as a yellow foamy solid (120 mg, 0.14 mmol, 36%). 1H NMR (500 MHz, CDCl3): δ 1.40 (br s, 18 H), 1.60− 1.84 (br m, 4 H), 2.36−2.50 (m, 4 H), 2.52−2.63 (m, 4 H), 3.18−3.48 (br m, 8 H), 3.72 (br s, 4 H), 5.64 (br s, 4 H), 7.40 (d, J 8.5, 4 H), 7.45−7.65 (br s, 2 H), 8.22 (d, J 8.5, 4 H). 13C NMR (126 MHz, CDCl3): δ 26.7, 28.4, 46.6, 49.8, 50.5, 52.1, 53.0, 54.3, 79.4, 122.7, 124.3, 128.5, 142.0, 148.0, 155.7, 158.1. LRMS (ESI+): m/z 833.44 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C40H57N12O8+ 833.4422, found 833.4429. Di-tert-butyl 4,11-Bis((1-(4-methoxybenzyl)-1H-1,2,3-triazol4-yl)methyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (4h). Similar to the procedure for 4a, 3h (0.84 mmol, 137 mg, 2.09 equiv) and 2 (238 mg, 0.4 mmol, 1 equiv) in THF (1.7 mL) and H2O (1.7 mL) and CuSO4·5H2O (20 mg, 20 mol %) and sodium ascorbate (35 mg, 50 mol %) for 1 h at rt. The reaction mixture was concentrated and purified by flash chromatography (0−15% DCM− MeOH, over 10 CV) to yield 4h as a yellow foamy solid (155 mg, 0.19 mmol, 48%). 1H NMR (500 MHz, CDCl3): δ 1.34−1.43 (br m, 18 H), 1.63−1.74 (br m, 4 H), 2.38−2.46 (br m, 4 H), 2.51−2.60 (br m, 8 H), 3.15−3.40 (m, 8 H), 3.64−3.74 (br m, 4 H), 3.78 (s, 6 H), 6.87 (d, J 8.5, 4 H), 7.14−7.25 (br m, 4 H), 7.31 (br s, 2 H). 13C NMR (126 MHz, CDCl3): δ 27.1, 28.4, 46.5, 49.7, 50.4, 51.9, 53.6, 53.6 (HSQC), 55.3, 79.3, 114.4, 114.8, 122.1, 126.9, 129.6, 155.8, 159.8. HRMS (ESI+): m/z calcd for C42H63N10O6+ 803.4932, found 803.4934. Di-tert-butyl 4,11-Bis((1-(2-(3-hydroxypropyl)-1,3-dioxo-2,3dihydro-1H-benzo[de]isoquinolin-6-yl)-1H-1,2,3-triazol-4-yl)methyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (4i). To a flask charged with 3i (1.86 g, 6.29 mmol, 2.09 equiv) and 2 (1.5 g, 3.15 mmol, 1 equiv) were added THF (14 mL) and H2O (14 mL) (0.11 M of cyclam in the 1:1 mixture of THF/H2O) backfilled with N2 at this point (×3). CuSO4·5H2O (157 mg, 20 mol % 0.63 mmol) and sodium ascorbate (0.281 g, 50 mol %, 1.6 mmol) were added, and the reaction was backfilled with N2 (×3). The reaction was stirred at rt (reached using a water bath at 25 °C) for 1 h. The reaction mixture was concentrated by rotary evaporation and purified by column chromatography (5% MeOH−DCM, to remove the 3i increasing to 40% MeOH−DCM). Fractions were combined which were >98% purity by HPLC (analytical run, 0−100% over 60 min; MeCN−H2O (with 0.1% TFA) over 90 min) to yield 4i as a yellowbrown solid (1.80 g, 1.68 mmol, 52%). 1H NMR (300 MHz, CDCl3): δ 1.34−1.43 (br m, 22 H), 1.64−2.03 (br m, 4 H), 2.55−2.59 (br m, 4 H), 2.70−2.75 (br m, 4 H), 3.20−3.42 (br m, 8 H), 3.61−3.65 (br m, 4 H), 3.91−4.02 (br m, 4 H), 4.35−4.39 (br m, 4 H), 7.85−7.92 (br m, 4 H), 8.26−8.39 (br m, 4 H), 8.72−8.70 (m, 4 H). 13C NMR (75 MHz, CDCl3): δ 28.4, 30.9, 37.2, 45.6, 47.5, 59.1, 79.7, 122.6, 123.4, 123.6, 125.1, 126.4, 128.7, 129.1, 130.0, 131.1, 132.4, 138.5, 155.7, 155.8, 163.2. FTIR (ATR) νmax/cm−1: 3055, 1662, 1647. Spectroscopic properties matched those previously described.15a 1,8-Bis((1-(naphthalen-1-yl)-1H-1,2,3-triazol-4-yl)methyl)1,4,8,11-tetraazacyclotetradecane (5a). To a solution of 4a (489 mg, 0.60 mmol) in 1,4-dioxane (3.0 mL, 0.2 M) was added HCl in dioxane (4.0 M, 4.8 mmol, 1.2 mL) and stirred at rt for 1 h. 1,4Dioxane was removed by rotary evaporation to yield the salt of 5a·0.8 C4H8O2·3.8 H2O·3.4HCl as pale brown powder (447 mg, 98%). Mp 250−262 °C. Anal. Calcd for C36H42N10·0.8C4H8O2·3.8H2O·3.4HCl: C 53.64 H 6.82 N 15.96. Found: C 53.64 H 6.77 N 15.96. Conversion of the salt (100 mg) to the amine was performed by reverse phase automated chromatography (0−100% MeCN−H2O (with 0.1% TFA) over 10 CV) and concentrated and extracted from 2.5 M aqueous NaOH (10 mL) with DCM (3 × 30 mL). The combined organic layers were dried over Na2SO4 and concentrated to yield 5a as a brown solid (35 mg, 0.057 mmol, 9.5% (43% based on the % purified), >95% purity by analytical HPLC). 1H NMR (300 MHz, CDCl3): δ 1.81− 1.92 (br s, 4 H), 2.55−2.62 (m, 4 H), 2.64−2.70 (br m, 4 H), 2.71− 2.81 (m, 8 H), 3.86 (s, 4 H), 7.45−7.60 (m, 10 H), 7.71 (s, 2 H), 7.83−7.91 (m, 2 H), 7.91−7.99 (m, 2 H). 13C NMR (75 MHz,

CDCl3): δ 25.9, 47.3, 47.4, 49.9, 51.4, 54.2, 122.3, 123.5, 124.9, 125.2, 126.9, 127.7, 128.1, 128.5, 130.1, 133.7, 134.0, 143.2. LRMS (ESI+): m/z 615.38 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C36H43N10 615.3672 found 615.3666. Anal. Calcd for C36H42N10·0.8 C4H8O2·3.8H2O·3.4HCl: C 53.64 H 6.82 N 15.96. Found: C 53.64 H 6.77 N 15.96. 1,8-Bis((1-(naphthalen-1-ylmethyl)-1H-1,2,3-triazol-4-yl)methyl)-1,4,8,11-tetraazacyclotetradecane (5b). Similar to the procedure for 4a, 3b (112 mg, 0.61 mmol, 2.4 equiv, contained 2(azidomethyl)naphthalene as a minor impurity), 2 (123 mg, 0.26 mmol, 1 equiv), CuSO4·5H2O (7 mg, 10 mol %), and sodium ascobate (9 mg, 20 mol %) were reacted for 6 h at 50 °C and concentrated by rotary evaporation. Saturated NH4Cl (50 mL) was added and extracted with DCM (3 × 50 mL) and dried over MgSO4 and concentrated to yield 4b as a crude mixture (crude yield 148 mg, 0.24 mmol, 92%). 1H NMR (300 MHz, CDCl3): δ 1.32−1.43 (br s, 18 H), 1.46−1.60 (br m, 4 H), 1.61−1.77 (br m, 4 H), 2.24−2.38 (br m, 4 H), 2.38−2.51 (br m, 4 H), 2.95−3.27 (br m, 8 H), 3.49−3.69 (br s, 4 H), 5.75−6.03 (br m, 2 H) 7.34−7.54 (br m, 10 H), 7.84 (d, J 8.0, 4 H), 7.94 (br s, 2 H). LRMS (ESI+): m/z 843.50 [M + H]+, m/z 865.44 [M + Na]+. The crude products of 4b (148 mg) were dissolved in 1,4-dioxane (4 mL) and HCl in dioxane (16 equiv, 4.0 M) was added and stirred at rt for 16 h. The reaction mixture was concentrated and 16 mg of this mixture was purified by preparative RP-HPLC (gradient 20% to 40% MeCN in H2O (with 0.1% TFA), over 90 min) to yield 5b·3.6TFA·0.5H2O as a white solid (4 mg, 0.004 mmol, 3%). 1 H NMR (300 MHz, D2O): δ 1.92 (br m, 4 H), 2.66 (br m, 4 H), 2.94 (br m, 8 H), 3.23 (br m, 4 H), 3.52−5.64 (br s, 4 H), 5.90−6.02 (br s, 4 H), 7.35−7.52 (br s, 2 H), 7.53−7.65 (br m, 2 H), 7.65−7.81 (br m, 6 H), 8.05−8.19 (br m, 6 H). 5b·3.6TFA·0.5H2O (5 mg) was converted to the amine (5b) for 13C NMR analysis (by extracting the compound with DCM (3 × 3 mL) from 2.5 M aqueous NaOH (5 mL). 1H NMR (500 MHz, CDCl3): δ 1.64−1.76 (m, 4 H), 2.38 (apparent t, J 5.5, 4 H), 2.48−2.53 (m, 4 H), 2.53−2.59 (m, 8 H), 3.61 (s, 4 H), 5.61 (s, 4 H), 7.29 (dd, J 8.5, 2.0, 2 H), 7.36 (br s, 2 H), 7.44−7.50 (m, 4 H), 7.69 (s, 2 H), 7.74−7.81 (m, 6 H). 13C NMR (126 MHz, CDCl3): δ 25.5, 47.1, 48.1, 49.3, 51.7, 53.2, 54.1, 122.6, 125.2, 126.6, 126.7, 127.3, 127.7, 127.9, 129.1, 132.2, 133.1, 133.2, 144.4. LRMS (ESI+): m/z 643.37 ([M (free amine) + H]+, 100%). HRMS (ESI+): m/z calculated for C38H47N10 643.3956, found 643.3985 [M + H]+. Anal. Calcd for C38H46N10·3.6CF3CO2H· 0.5H2O: C 51.10 H 4.80 N 13.18. Found: C 51.29 H 4.88 N 13.10. 1,8-Bis((1-(4-nitrophenyl)-1H-1,2,3-triazol-4-yl)methyl)1,4,8,11-tetraazacyclotetradecane (5d). To 4d (80 mg, 0.1 mmol, 1 equiv) in 1,4-dioxane (1 mL) was added HCl in dioxane (0.25 mL, 4.0 M, 8 equiv) and the reaction mixture stirred at rt for 30 min (typically precipitation occurs after 5 min, the reactions can be followed by TLC, an aliquot containing solid was added to a vial containing NaHCO3 (sat., 0.5 mL) and EtOAc (0.5 mL), the amine products have an RF of 0.00 in EtOAc, 10% MeOH−DCM and 1% NH4OH (aq) in 10% MeOH−DCM). The reaction mixture was concentrated by rotary evaporation and purified by reverse-phase automated chromatography (0−100% MeCN−H2O (with 0.1% TFA) over 10 CV). The fractions were concentrated by rotary evaporation (to remove MeCN), and aqueous NaOH (5 mL, 2.5 M) was added and extracted with DCM (1 × 5 mL). The extracts were dried over Na2SO4 and concentrated to yield 5d as an orange solid (38 mg, 0.63 mmol, 63%, >98% purity by analytical HPLC). 1H NMR (500 MHz, CDCl3): δ 1.87−1.95 (m, 4 H), 2.58−2.63 (m, 4 H), 2.78−2.84 (m, 4 H) 2.84−2.96 (m, 8 H), 3.80 (s, 4 H), 8.09 (d, J 9.0, 4 H), 8.36 (d, J 9.0, 4 H), 8.64 (br s, 2 H). 13C NMR (126 MHz, CDCl3): δ 24.8, 47.1, 48.5, 50.5, 51.8, 53.4, 120.3, 121.6, 125.4, 141.1, 145.8, 147.0. LRMS (ESI+): m/z 605.29 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C28H37N12O4+ 605.3061, found 605.3060. FTIR (ATR) νmax/cm−1: 2963, 2823, 1598, 1523. 1,8-Bis((1-(4-methoxyphenyl)-1H-1,2,3-triazol-4-yl)methyl)1,4,8,11-tetraazacyclotetradecane (5e). Similar to the procedure for 5d, 4e (78 mg, 0.1 mmol, 1 equiv) in 1,4-dioxane (1 mL) and HCl in dioxane (4.0 M, 10 equiv, 0.25 mL) for 30 min at rt. The reaction mixture was concentrated and purified by reverse phase by automated 3601

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

Journal of Medicinal Chemistry

Article

6,6′-(((1,4,8,11-Tetraazacyclotetradecane-1,8-diyl)bis(methylene))bis(1H-1,2,3-triazole-4,1-diyl))bis(2-(3-hydroxypropyl)-1H-benzo[de]isoquinoline-1,3(2H)-dione) (5i). 4i (500 mg, 0.47 mmol, 1 equiv) was dissolved in 1,4-dioxane (6 mL, 0.08M), and HCl in dioxane (4.0 M, 0.94 mL, 3.74 mmol, 8 equiv) was added (instantaneous change giving a yellow-white precipitate upon addition of HCl). The reaction mixture was left to stir at rt for 2 h and concentrated by rotary evaporation. The residue was dissolved in a mixture of EtOH (200 mL) and CHCl3 (200 mL), and Ambersep 900 resin (hydroxide form, 200 mg, washed with MeOH (3 × 10 mL)) was added, and the reaction stirred for 1 h at rt. The resin was removed by filtration (without washing the resin) and the filtrate concentrated by rotary evaporation to yield 5i as a light yellow powder (348 mg, 0.40 mmol, 85%). 1H NMR (300 MHz, D2O): δ 1.78−1.97 (m, 4 H), 2.08−2.22 (m, 4 H), 2.82−3.00 (m, 4 H), 3.03−3.17 (m, 4 H), 3.40− 3.51 (m, 4 H), 3.53−3.63 (very br m, 4 H), 3.64−3.72 (m, 4 H), 3.94−4.03 (m, 4 H), 4.06−4.15 (m, 4 H), 7.20−7.33 (m, 2 H), 7.56− 7.71 (m, 4 H), 8.06−8.13 (m, 2 H), 8.21−8.27 (m, 2 H), 8.29−8.33 (m, 2 H. 1H NMR (300 MHz, CDCl3 (10% MeOD-d4)): δ 1.54−1.77 (m, 8 H), 2.31−2.63 (m, 16 H), 3.05 (br s, 2 H), 3.39 (t, J 6.0, 4 H), 3.61−3.78 (br m, 4 H), 4.00 (t, J 7.0, 4 H), 7.49−7.65 (m, 4 H), 7.93 (d, J 8.5, 2 H), 7.99 (s, 2 H), 8.35 (d, J 7.5, 4 H). 13C NMR (75 MHz, CDCl3 (10% MeOD-d4)): δ 24.7, 30.3, 37.3, 46.5, 49.8, 52.3, 53.3, 54.6, 59.1, 122.1, 123.1, 123.3, 125.6, 125.9, 128.1, 128.5, 129.2, 130.3, 131.7, 137.8, 143.2, 163.0, 163.6. HRMS (ESI+): m/z calcd for C46H53N12O6+ [M + H]+ 869.4211, found 869.4188. 1-Ethynylnaphthalene (7a). To a solution of 1-bromonaphthalene (285 mg, 1.38 mmol) in diisopropylamine (10 mL) were added Pd(PPh3)2Cl2 (48 mg, 68 μmol, 5 mol %) and CuI (13 mg, 68 μmol, 5 mol %) under N2. Trimethylsilylacetylene (292 μL, 2.07 mmol) was added dropwise, and the reaction mixture was stirred for 16 h at 80 °C. The reaction mixture was filtered and concentrated by rotary evaporation. The residue was dissolved in CHCl3 (25 mL), washed with H2O (2 × 25 mL), dried over Na2SO4, and concentrated by rotary evaporation. The residue was purified by flash column chromatography (petroleum benzine) to yield the TMS-protected intermediate as a yellow oil (259 mg, 84%). 1H NMR (400 MHz, CDCl3): δ 0.33 (s, 9 H), 7.40 (dd, J 8.0, 7.0, 1 H), 7.51 (ddd, J 8.0, 7.0, 1.5, 1 H), 7.57 (ddd, J 8.5, 7.0, 1.5, 1 H), 7.70 (dd, J 7.0, 1.0, 1 H), 7.79−7.86 (m, 2 H), 8.31−8.36 (m, 1 H). LRMS (EI): m/z 209.2 ([M − CH3]+, 100%), 224.1 (M+, 58%). Spectroscopic properties matched those previously described.28 To a solution of the TMS-protected intermediate (255 mg, 1.14 mmol) in MeOH (10 mL) was added a solution of KOH (77 mg, 1.4 mmol) in H2O (2 mL). The reaction mixture was stirred for 16 h at rt. The volatiles were removed and H2O (25 mL) was added. The product was extracted with Et2O (2 × 25 mL), and the extracts were combined, washed with H2O (25 mL), dried over Na2SO4, and the solvent was removed by rotary evaporation. The residue was purified by flash column chromatography (petroleum benzine) to yield 7a as an orange oil (143 mg, 83%). 1 H NMR (400 MHz, CDCl3): δ 3.47 (s, 1 H), 7.42 (dd, J 8.0, 7.0, 1 H), 7.49−7.55 (m, 1 H), 7.55−7.61 (m, 1 H), 7.74 (d, J 7.0, 1 H), 7.85 (d, J 8.0, 2 H), 7.36 (d, J 8.5, 1 H). LRMS (EI): m/z 152.2 (M+, 100%). Spectroscopic properties matched those previously described.29 2-Ethynylnaphthalene (7c). To a solution of 2-bromonaphthalene (500 mg, 2.42 mmol) in diisopropylamine (10 mL) were added Pd(PPh3)2Cl2 (102 mg, 6 mol %) and CuI (28 mg, 6 mol %) under N2. Trimethylsilylacetylene (512 μL, 3.62 mmol) was added dropwise, and the reaction mixture was stirred for 16 h at 80 °C. The reaction mixture was filtered and the solvent was removed by rotary evaporation. The residue was dissolved in DCM (50 mL), washed with H2O (2 × 25 mL), dried over Na2SO4, and concentrated by rotary evaporation. The residue was purified by automated flash column chromatography (100% petroleum benzine for 12 CV) to yield the TMS-protected intermediate as a brown oil (460 mg, 85%). 1H NMR (400 MHz, CDCl3): δ 0.28 (s, 9 H), 7.43−7.53 (m, 3 H), 7.72−7.83 (m, 3 H), 8.00 (s, 1 H). LRMS (EI): m/z 209.1 ([M − CH3]+, 100%), 224.0 (M+, 38%). Spectroscopic properties matched those previously described.30 To a solution of the TMS-protected intermediate (443

reverse phase column chromatography ((0−100%) MeCN−H2O (with 0.1% TFA) over 10 CV). After concentration of the fractions and freeze-drying the products were made into the free-amine by addition of 2.5 M NaOH (5 mL) and extracted with DCM (5 mL). The organic layer was dried over Na2SO4 and concentrated to yield 5e as a yellow solid (30 mg, 0.052 mmol, 52%, >98% purity by analytical HPLC). 1H NMR (500 MHz, CDCl3): δ 1.81−1.92 (m, 4 H), 2.59 (t, J 5.5, 4 H), 2.75−2.84 (m, 8 H), 2.85−2.95 (m, 4 H), 3.74 (s, 4 H), 3.82 (s, 6 H), 6.91−7.04 (m, 4 H), 7.59−7.69 (m, 4 H), 8.05 (s, 2 H). 13 C NMR (126 MHz, CDCl3): δ 24.8, 47.0, 48.4, 48.5, 50.9, 51.7, 55.5, 114.7, 121.2, 121.8, 130.4, 144.6, 159.6. LRMS (ESI+): m/z 575.35 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C30H43N10O2+ 575.3571, found 575.3568. FTIR (ATR) νmax/cm−1: 2948, 2823, 1685, 1513. 1,8-Bis((1-benzyl-1H-1,2,3-triazol-4-yl)methyl)-1,4,8,11tetraazacyclotetradecane (5f). Similar to the procedure for 5d, 4f (74 mg, 0.1 mmol, 1 equiv) in 1,4-dioxane (1 mL) was added HCl in dioxane (4.0 M, 10 equiv, 0.25 mL) for 30 min at rt. The reaction mixture was concentrated and purified by automated reverse phase column chromatography ((0−100%) MeCN−H2O (with 0.1% TFA) over 10 CV). After concentration of the fractions and freeze-drying the products were made into the free amine by addition of 2.5 M NaOH (5 mL) and extracted with DCM (5 mL). The organic layer was dried over Na2SO4 and concentrated to yield 5f as a yellow solid (31 mg, 0.041 mmol, 41%, >98% purity by analytical HPLC). 1H NMR (500 MHz, CDCl3): δ 1.75−1.84 (br m, 4 H), 2.44−2.54 (br m, 4 H), 2.62−2.78 (br m, 8 H), 2.75−2.84 (br m, 4 H), 3.63 (s, 4 H), 5.45 (s, 4 H), 7.19−7.27 (m, 4 H), 7.27−7.34 (m, 6 H), 7.62 (s, 2 H). 13C NMR (126 MHz, CDCl3): δ 24.6, 29.7, 46.8, 48.7, 51.4, 51.8, 54.0, 123.0, 128.1, 128.7, 129.0, 134.9, 144.6. LRMS (ESI+): m/z 543.35 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C30H43N10+ 543.3672, found 543.3669. FTIR (ATR) νmax/cm−1: 2935, 2815, 1459. 1,8-Bis((1-(4-nitrobenzyl)-1H-1,2,3-triazol-4-yl)methyl)1,4,8,11-tetraazacyclotetradecane (5g). Similar to the procedure for 5d, 4g (83 mg, 0.1 mmol, 1 equiv) in 1,4-dioxane (1 mL) and HCl in dioxane (4.0 M, 10 equiv, 0.25 mL) for 30 min at rt. The reaction mixture was concentrated and purified by reverse phase by automated reverse phase column chromatography ((0−100%) MeCN−H2O (with 0.1% TFA) over 10 CV). After concentration of the fractions and freeze-drying the products were made into the free amine by addition of 2.5 M NaOH (5 mL) and extracted with DCM (5 mL). The organic layer was dried over Na2SO4 and concentrated to yield 5g as a yellow solid (37 mg, 0.058 mmol, 58%, >98% purity by analytical HPLC). 1H NMR (500 MHz, CDCl3): δ 1.76−1.85 (m, 4 H), 2.46− 2.55 (m, 4 H), 2.65−2.76 (m, 8 H), 2.78−2.85 (m, 4 H), 3.68 (s, 4 H), 5.65 (s, 4 H), 7.46 (d, J 8.5, 4 H), 7.83 (s, 2 H), 8.16 (d, J 8.5, 4 H). 13 C NMR (126 MHz, CDCl3): δ 24.7, 47.0, 48.4, 48.8, 51.2, 51.7, 52.9, 123.5, 124.1, 128.8, 141.9, 144.6, 147.9. LRMS (ESI+): m/z 633.31 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C30H41N12O4+ 633.3374, found 633.3372. FTIR (ATR) νmax/cm−1: 2959, 2829, 1678, 1519. 1,8-Bis((1-(4-methoxybenzyl)-1H-1,2,3-triazol-4-yl)methyl)1,4,8,11-tetraazacyclotetradecane (5h). Similar to the procedure for 5d, 4h (80 mg, 0.1 mmol, 1 equiv) in 1,4-dioxane (1 mL) and HCl in dioxane (4.0 M, 10 equiv, 0.25 mL) for 30 min at rt. The reaction mixture was concentrated and purified by reverse phase by automated reverse phase column chromatography ((0−100%) MeCN−H2O (with 0.1% TFA) over 10 CV). After concentration of the fractions and freeze-drying the products were made into the free amine by addition of 2.5 M NaOH (5 mL) and extracted with DCM (5 mL). The organic layer was dried over Na2SO4 and concentrated to yield 5h as a yellow solid (37 mg, 0.061 mmol, 61%, >98% purity by analytical HPLC). 1H NMR (500 MHz, CDCl3): δ 1.70−1.79 (br m, 4 H), 2.41−2.50 (m, 4 H), 2.56−2.69 (m, 8 H), 2.71−2.78 (m, 4 H), 3.62 (s, 4 H), 3.76 (s, 6 H), 5.38 (s, 4 H), 6.83 (d, J 8.5, 4 H), 7.18 (d, J 8.5, 4 H), 7.47 (s, 2 H). 13C NMR (126 MHz, CDCl3): δ 24.8, 46.8, 48.4, 48.7, 51.5, 51.9, 53.5, 55.2, 114.3, 122.5, 126.7, 129.5, 144.3, 159.8. LRMS (ESI+): m/z 603.36 ([M + H]+, 100%). HRMS (ESI+): m/z calcd for C32H47N10O2+ 603.3884, found 603.3881. FTIR (ATR) νmax/ cm−1: 2948, 2839, 1688, 1610. 3602

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

Journal of Medicinal Chemistry

Article

by rotary evaporation and the residue was purified by flash column chromatography (EtOAc/petroleum benzine = 1:1 to EtOAc to MeOH/EtOAc 1:9) to yield 8e as an off-white gum (147 mg, 60%). 1 H NMR (300 MHz, CDCl3): δ 1.44 (s, 18 H), 1.45−1.57 (m, 4 H), 2.38 (t, J 6.5, 4 H), 2.53 (t, J 5.5, 4 H), 2.85 (t, J 6.5, 4 H), 3.01−3.19 (m, 8 H), 4.06 (s, 4 H), 4.26 (t, J 6.5, 4 H), 7.15−7.34 (m, 12 H). 13C NMR (75 MHz, CDCl3): δ 28.5, 32.2, 46.6, 46.8, 48.3, 52.5, 53.5, 55.3, 73.9, 79.5, 122.0, 126.4, 128.5, 128.7, 139.1, 147.3, 155.6. LRMS (ESI +): m/z 771.2 ([M + H]+, 100%), 793.1 ([M + Na]+, 45%). HRMS (ESI+): m/z calcd for C42H63N10O4+ [M + H]+ 771.5028, found 771.5035. FTIR (ATR) νmax/cm−1: 2974, 1678, 1494, 1454, 1413, 1391, 1365, 1248, 1154, 1074, 1048, 911, 774, 726, 698, 646, 463. 1,8-Bis(2-(4-(naphthalen-1-yl)-1H-1,2,3-triazol-1-yl)ethyl)1,4,8,11-tetraazacyclotetradecane (9a). To 8a (44 mg, 0.052 mmol) in 1,4-dioxane (2 mL) was added HCl in dioxane (4.0 M, 0.10 mL, 8 equiv) and stirred for 16 h at rt. The residue was concentrated under reduced pressure and washed with EtOAc to yield 9a·3(HCl)· 3H2O as a beige solid (37 mg, 94%). Mp 240 °C (dec). 1H NMR (400 MHz, DMSO-d6): δ 1.81−1.93 (m, 4 H), 2.62−2.73 (m, 4 H), 2.80− 2.92 (m, 4 H), 3.01−3.10 (m, 4 H), 3.13 (t, J 6.5, 4 H), 3.15−3.22 (m, 4 H), 4.65 (t, J 6.5, 4 H), 7.48−7.61 (m, 6 H), 7.74 (dd, J 7.0, 1.0, 2 H), 7.92−8.03 (m, 4 H), 8.44−8.51 (m, 2 H), 8.71 (s, 2 H), 8.74 (br s, 2 H, NH). 13C NMR (100 MHz, DMSO-d6): δ 22.7, 43.3, 44.6, 45.4, 48.6, 50.0, 50.4, 124.5, 125.2, 125.5, 126.0, 126.6, 126.7, 127.8, 128.5, 128.5, 130.1, 133.5, 145.6. LRMS (ESI+): m/z 643.3 (free base [M + H]+, 100%). HRMS (ESI+): m/z calcd for C38H47N10+ 643.3980, found 643.3984. FTIR (ATR) νmax/cm−1: 3396, 2956, 2615, 2418, 1653, 1586, 1453, 1393, 1354, 1259, 1220, 1049, 1018, 946, 800, 776, 743. Anal. Calcd for C38H46N10·3HCl·3H2O: C 56.61, H 6.88, N 17.37. Found: C 56.44, H 6.76, N 17.20. 1,8-Bis(2-(4-(naphthalen-2-yl)-1H-1,2,3-triazol-1-yl)ethyl)1,4,8,11-tetraazacyclotetradecane (9c). 8c (59 mg, 0.070 mmol) was dissolved in a mixture of TFA (4 mL), H2O (0.5 mL) and CH2Cl2 (0.5 mL) and stirred for 4 h at rt. The reaction mixture was concentrated by rotary evaporation, and the residue was triturated with EtOAc and purified by HPLC (0% to 100% MeCN in H2O (with 0.1% TFA) over 20 min) to yield 9c·2.5(TFA) as a white solid (42 mg, 68%). Mp 213 °C (dec). 1H NMR (400 MHz, DMSO-d6): δ 1.67− 1.78 (m, 4 H), 2.57−2.66 (m, 4 H), 2.67−2.75 (m, 4 H), 2.82−2.92 (m, 4 H), 3.02 (t, J 6.5, 4 H), 3.07−3.16 (m, 4 H), 4.59 (t, J 6.5, 4 H), 7.48−7.57 (m, 4 H), 7.88−8.02 (m, 8 H), 8.15 (br s, 4 H, NH), 8.37 (s, 2 H), 8.78 (s, 2 H). 13C NMR (100 MHz, DMSO-d6): δ 23.4, 43.6, 44.2, 45.9, 48.6, 50, 50.6, 117.3 (q, JC−F 298.0, TFA) 122.8, 123.8, 124, 126.7, 127.2, 128.2, 128.4, 128.5, 129.1, 133, 133.6, 147, 158.8 (q, JC−F 31.5, TFA). LRMS (ESI+): m/z 643.1 (free base [M + H]+, 100%). HRMS (ESI+): m/z calcd for C38H47N10+ free base [M + H]+ 643.3980, found 643.3981. FTIR (ATR) νmax/cm−1: 3055, 2961, 2848, 1674, 1461, 1431, 1199, 1179, 1128, 1073, 828, 799, 755, 720, 477. Anal. Calcd for C38H46N10·2.5CF3CO2H: C 55.66, H 5.27, N 15.10. Found: C 55.80, H 5.12, N 14.88. 1,8-Bis(2-(4-benzyl-1H-1,2,3-triazol-1-yl)ethyl)-1,4,8,11tetraazacyclotetradecane (9e). Similar to the procedure for 9a, 8e (141 mg, 0.183 mmol) in 1,4-dioxane (2 mL) was added HCl in dioxane (4.0 M, 0.10 mL, 8 equiv) and stirred for 16 h at rt. The residue was concentrated under reduced pressure and washed with EtOAc to yield 9e·4(HCl)·2.5H2O as a white solid (122 mg, 98%). Mp 189 °C (dec). 1H NMR (400 MHz, D2O): δ 1.58−1.68 (m, 4 H), 2.26 (t, J 7.0, 4 H), 2.47−2.58 (m, 8 H), 2.78−2.87 (m, 4 H), 2.97 (t, J 5.5, 4 H), 4.08 (s, 4 H), 4.52 (t, J 5.5, 4 H), 7.27−7.46 (m, 10 H), 8.01 (s, 2 H). 13C NMR (100 MHz, D2O): δ 23.7, 31.4, 43.3, 44.1, 48.2, 48.3, 50.4, 51.1, 124.9, 127.6, 129.4, 129.7, 139.6, 148.5. LRMS (ESI +): m/z 571.3 (free base [M + H]+, 100%). HRMS (ESI+): m/z calcd for C32H47N10+ free base [M + H]+ 571.3980, found 571.3979. FTIR (ATR) νmax/cm−1: 3355, 2998, 2365, 1494, 1452, 1225, 1076, 1053, 1028, 968, 729, 699, 583, 463. Anal. Calcd for C32H46N10·4HCl· 2.5H2O: C 50.46, H 7.28, N 18.29. Found: C 50.53, H 7.43, N 17.79. 1,4,8,11-Tetraazatricyclo[9.3.1.14,8]hexadecane (10). To a solution of cyclam (5.00 g, 25.0 mmol, 1 equiv) in deionized H2O (300 mL, 0.08 M) (requires heating and/or sonication to dissolve) was added formaldehyde (∼37% in H2O, 4.50 mL, 55.0 mmol, 2.2 equiv)

mg, 1.97 mmol) in MeOH (10 mL) was added a solution of KOH (133 mg, 2.37 mmol) in H2O (2 mL). The reaction mixture was stirred at for 16 h at rt. The volatiles were removed, and H2O (25 mL) was added. The product was extracted with Et2O (2 × 25 mL), and the extracts were combined, dried over Na2SO4, and the solvent was removed by rotary evaporation. The residue was purified by flash column chromatography (petroleum benzine) to yield 7c as a colorless oil (204 mg, 68%). 1H NMR (400 MHz, CDCl3): δ 3.14 (s, 1 H), 7.46−7.56 (m, 3 H), 7.75−7.86 (m, 3 H), 8.03 (br s, 1 H). LRMS (EI): m/z 152.1 ([M]+, 100%). Spectroscopic properties matched those previously described.30 Di-tert-butyl 4,11-Bis(2-(4-(naphthalen-1-yl)-1H-1,2,3-triazol-1-yl)ethyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (8a). To 6 (102 mg, 0.189 mmol, 1 equiv) and 7a (69 mg, 0.45 mmol, 2.4 equiv) in THF (7 mL) was added an orange cloudy mixture of CuSO4·5H2O (5 mg, 10 mol %) and sodium ascorbate (6 mg, 20 mol %) in H2O (3 mL). (NOTE: Ideally this should not be cloudy but result in an oxygen sensitive clear brown solution if mixed under an inert atmosphere; our preferred procedure is to add the CuSO4·5H2O and sodium ascorbate as solids to degassed solvents or to the backfilled reaction flask containing solvents.) The reaction mixture was stirred under N2 for 16 h at 50 °C. The reaction was quenched with saturated aqueous NH4Cl (25 mL), THF removed by rotary evaporation and extracted with EtOAc (2 × 25 mL). The combined organic extracts were dried over Na2SO4 and concentrated by rotary evaporation. The residue was purified by flash column chromatography (EtOAc/petroleum benzine = 1:1 to 3:2 to EtOAc) to yield 8a as a yellowish oil (112 mg, 70%). 1H NMR (500 MHz, CDCl3): δ 1.44 (s, 18 H), 1.49−1.58 (m, 4 H), 2.37 (t, J 6.5, 4 H), 2.55 (t, J 5.5, 4 H), 2.86 (t, J 6.5, 4 H), 3.14 (t, J 5.0, 4 H), 3.21 (t, J 7.0, 4 H), 4.30 (t, J 5.0, 4 H), 7.41−7.52 (m, 6 H), 7.66−7.75 (m, 4 H), 7.77−7.86 (m, 4 H), 8.43 (d, J 8.0, 4 H). 13C NMR (125 MHz, CDCl3): δ 26.8, 28.4, 46.6, 46.8, 48.3, 52.4, 53.8, 55.1, 79.6, 123.3, 125.2, 125.5, 125.9, 126.5, 126.9, 128.0, 128.3, 128.7, 130.9, 133.8, 146.5, 155.7. LRMS (ESI+): m/z 843.3 ([M + H]+, 100%), 865.4 ([M + Na]+, 22%). HRMS (ESI+): m/z calcd for C48H63N10O4+ [M + H]+ 843.5028, found 843.5038. FTIR (ATR) νmax/cm−1: 2975, 2933, 1673, 1455, 1416, 1393, 1366, 1250, 1159, 1049, 910, 801, 777, 730, 646. Di-tert-butyl 4,11-Bis(2-(4-(naphthalen-2-yl)-1H-1,2,3-triazol-1-yl)ethyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (8c). Similar to the procedure for 8a, 7c (180 mg, 0.33 mmol, 1 equiv) and 6 (122 mg, 0.80 mmol, 2.4 equiv) in THF (7 mL), CuSO4·5H2O (9 mg, 10 mol %) and sodium ascorbate (11 mg, 20 mol %) in H2O (3 mL) for 16 h at 50 °C. The reaction was quenched with saturated aqueous NH4Cl (25 mL), THF removed by rotary evaporation and extracted with EtOAc (2 × 25 mL). The combined organic extracts were dried over Na2SO4 and concentrated by rotary evaporation and the residue purified by automated column chromatography (15% EtOAc in petroleum benzine over 1 CV, 15% to 100% over 10 CV, 100% over 1 CV) to yield 8c as an off-white gum (191 mg, 68%). 1H NMR (400 MHz, CDCl3): δ 1.37−1.51 (m, 4 H), 1.47 (s, 18 H), 2.29 (t, J 6.0, 4 H), 2.45−2.58 (m, 4 H), 2.69−2.81 (m, 4 H), 2.98−3.10 (m, 4 H), 3.10−3.24 (m, 4 H), 4.00−4.17 (m, 4 H), 7.40−7.51 (m, 4 H), 7.71 (s, 2 H), 7.77−7.84 (m, 2 H), 7.84−7.93 (m, 4 H), 7.93−8.01 (m, 2 H), 8.40 (s, 2 H). 13C NMR (100 MHz, CDCl3): δ 26.7, 28.6, 46.4, 47.2, 48.5, 52.6, 54.2, 55.4, 79.7, 121.4, 123.8, 124.2, 126.1, 126.5, 127.8, 128.1, 128.2, 128.6, 133.1, 133.6, 147, 155.9. LRMS (ESI+): m/z 843.2 ([M + H]+, 29%), 865.1 ([M + Na]+, 100%). HRMS (ESI+): m/z calcd for C48H63N10O4+ [M + H]+ 843.5028, found 843.5021. FTIR (ATR) νmax/cm−1: 2973, 1682, 1457, 1415, 1366, 1246, 1159, 1050, 908, 861, 820, 731, 476. Di-tert-butyl 4,11-Bis(2-(4-benzyl-1H-1,2,3-triazol-1-yl)ethyl)-1,4,8,11-tetraazacyclotetradecane-1,8-dicarboxylate (8e). Similar to the procedure for 8a, 6 (172 mg, 0.32 mmol, 1 equiv) and 3-phenyl-1-propyne (7e) (95 μL, 0.76 mmol, 2.4 equiv) in THF (7 mL), CuSO4·5H2O (8 mg, 10 mol %) and sodium ascorbate (11 mg, 20 mol %) in H2O (3 mL) for 16 h at 50 °C. The reaction was quenched with saturated aqueous NH4Cl (25 mL), THF removed by rotary evaporation and extracted with EtOAc (2 × 25 mL). The combined organic extracts were dried over Na2SO4 and concentrated 3603

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

Journal of Medicinal Chemistry

Article

needle overnight. The reaction mixture was filtered through Celite (eluting with MeOH) and concentrated and purified by automated reverse phase chromatography (0−100% H2O−MeCN (with 0.1% TFA), over 10 CV). MeCN was removed by rotary evaporation and the aqueous phase taken to pH 14 using 2.5 M aqueous NaOH solution and extracted with DCM (3 × 30 mL). The combined organic layers were dried over Na2SO4 and concentrated to yield 13 as a yellow waxy solid (28 mg, 0.522 mmol, 25%, >95% by HPLC). 1H NMR (300 MHz, CDCl3): δ 1.64−1.77 (m, 4 H), 1.77−1.89 (m, 4 H), 2.43−2.56 (m, 12 H), 2.59−2.67 (br m., 4 H), 2.66−2.78 (m, 8 H), 7.19 (d, J 7.0, 2 H), 7.31 (t, J 7.5, 2 H), 7.39−7.52 (m, 4 H), 7.63 (d, J 8.0, 2 H), 7.76−7.82 (m, 2 H), 7.90 (d, J 8.0, 2 H). 13C NMR (126 MHz, CDCl3): δ 22.6, 25.7, 26.3, 29.7, 30.7, 50.8, 53.1, 54.3, 123.8, 125.4, 125.5, 125.7, 126.5, 127.9, 128.7, 131.8, 133.8, 138.3. LRMS (ESI+): m/z 559.38 ([M + Na]+, 100%). HRMS (ESI+): m/z calcd for C36H49N4+ 537.3957, found 537.3952. FTIR (ATR) νmax/ cm−1: 2914, 2872, 1683, 1610. Fe(ClO 4 ) 2 Complex of 6,6′-(((1,4,8,11-Tetraazacyclotetradecane-1,8-diyl)bis(methylene))bis(1H-1,2,3-triazole-4,1diyl))bis(2-ethyl-1H-benzo[de]isoquinoline-1,3(2H)-dione) (14c). To a solution of 1 (200 mg, 248 μmol, 1.00 equiv) in EtOH (0.1 M, 2.5 mL) was added Fe(ClO4)2·H2O (67.6 mg, 248 μmol, 1.00 equiv) in EtOH (0.1 M, 2.5 mL) at rt, and the reaction mixture was heated at reflux for 24 h. The reaction mixture was cooled in an ice bath and the precipitate collected by vacuum filtration. The precipitate was washed with ice-cold ethanol (3 × 2 mL) and diethyl ether (3 × 2 mL) and dried under high vacuum to yield 14c·H2O as a purple solid (146 mg, 55%). Mp 203 °C. UV−vis (CH3CN) λmax/nm 344, ε 44249. FS (CH3CN) λex/nm 344, λem/nm 345, 419. 1H NMR (300 MHz, DMSO-d6): δ 1.02−1.39 (m, 6 H), 1.73−2.15 (m, 4 H), 2.64−3.05 (m, 8 H), 3.75−4.24 (m, 8 H) 7.78−8.29 (br m, 6 H), 8.30−9.14 (br m, 6 H) (8 × H missing or overlapping). 13C NMR (126 MHz, DMSO-d6): δ 13.1, 30.7, 35.1, 46.6, 123.1, 123.9, 124.6, 126.1, 127.4, 128.8, 129.3, 129.6, 130.9, 132.0, 138.0, 162.9, 163.4 (5 × C missing or overlapping). LRMS (ESI+) m/z 432.16 [M − 2ClO4]2+, 809.45 ([C44H48N12O4 + H]+, 909.50 ([C44H48ClN12O8] − [2H]+). Anal. Calcd for C44H48Cl2FeN12O12·H2O: C 48.86 H 4.66 N 15.54. Found: C 49.22 H 4.60 N 15.22. FTIR (ATR) νmax/cm−1 1702, 1660, 1590, 1342, 1245, 1085, 785, 755, 623. Fe(ClO4)3 Complex of 6,6′-(((1,4,8,11-Tetraazacyclotetradecane-1,8-diyl)bis(methylene))bis(1H-1,2,3-triazole-4,1-diyl))bis(2-ethyl-1H-benzo[de]isoquinoline-1,3(2H)-dione) (14d). Similar to the procedure for 14c, 1 (200 mg, 248 μmol, 1.00 equiv) in EtOH (0.1 M, 2.5 mL) and Fe(ClO4)3·H2O (92.0 mg, 248 μmol, 1.00 equiv) in EtOH (0.1 M, 2.5 mL) were refluxed for 24 h and then collected and washed to yield 14d·H2O as a golden yellow solid (189 mg, 66%). Mp 213 °C. UV−vis (CH3CN) λmax/nm 344, ε 30428. FS (CH3CN) λex/nm 344, λem/nm 345, 419. 1H NMR (300 MHz, DMSO-d6): δ 1.25 (t, J 7.0, 6 H), 1.95−2.11 (m, 4 H), 2.69−2.89 (m, 8 H), 4.02−4.17 (m, 8 H), 7.94 (t, J 8.0, 2 H), 8.05 (d, J 8.0, 2 H), 8.17 (d, J 8.5, 2 H), 8.33−8.70 (br m, 4 H), 8.89 (s, 2 H) (8 × H missing or overlapping). 13C NMR (126 MHz, DMSO-d6): δ 13.5, 22.6, 35.6, 44.2, 44.9, 46.9, 49.6, 52.1, 123.1, 123.9, 124.7, 126.2, 127.6, 128.8, 129.2, 129.6, 130.9, 132.0, 138.0, 142.4, 162.9, 163.4. LRMS ( A P C I ) m / z 8 09 . 3 6 ( [ C 4 4 H 4 8 N 1 2 O 4 ] + H ) + , 9 0 9 .4 2 ([C44H48ClN12O8] − [2H]+). Anal. Calce for C44H48Cl3FeN12O16· H2O: C 48.06 H 4.77 N 15.28. Found: C 48.40 H 4.80 N 15.26. FTIR (ATR) νmax/cm−1 1670, 1657, 1591, 1341, 1245, 1067, 791, 756, 623. Sm(ClO 4 ) 3 Complex of 6,6′-(((1,4,8,11-Tetraazacyclotetradecane-1,8-diyl)bis(methylene))bis(1H-1,2,3-triazole-4,1diyl))bis(2-ethyl-1H-benzo[de]isoquinoline-1,3(2H)-dione) (14e). Similar to the procedure for 14c, 1 (200 mg, 248 μmol, 1.00 equiv) in EtOH (0.1 M, 2.5 mL) and Sm(ClO4)3 (278 mg, 248 μmol, 1.00 equiv) in EtOH (0.1 M, 2.5 mL) were refluxed for 24 h and then collected and washed to yield 14e·H2O as a light yellow solid (243 mg, 78%). Mp 290−293 °C. UV−vis (CH3CN) λmax/nm 344, ε 45175. FS (CH3CN) λex/nm 344, λem/nm 345, 420. 1H NMR (300 MHz, DMSO-d6): δ 1.22 (t, J 7.0, 6 H), 1.73−1.85 (m, 4 H), 2.53−2.60 (m, 4 H), 2.65−2.72 (m, 4 H), 2.78−2.85 (m, 4 H), 2.89−2.96 (m, 4 H), 3.76 (s, 4 H), 4.09 (q, J 7.0, 4 H), 7.89 (t, J 8.0, 2 H), 7.97 (d, J 8.0, 2 H), 8.07 (d, J 8.5, 2 H), 8.53−8.59 (m, 4 H), 8.69 (s, 2 H). 13C NMR

dropwise at 0 °C. The reaction mixture was allowed to warm to rt, and was stirred for 5 h resulting in a white suspension. The precipitate was collected by vacuum filtration, washed with H2O (3 × 50 mL), and dried under high vacuum to yield 10 as a white solid (4.98 g, 89%, typically yields vary from 70 to 90% over 5−18 h). Mp 107−110 °C (lit.15,31 106−108 °C). 1H NMR (200 MHz, CDCl3): δ 1.18 (d, J 13.0, 2 H), 2.13−2.34 (m, 2 H), 2.34−2.50 (m, 4 H), 2.63 (td, J 12.0, 3.0, 4 H), 2.78−2.84 (m, 4 H), 2.91 (d, J 11.0, 2 H), 3.04−3.25 (m, 4 H), 5.44 (d, J 11.0, 2 H). LRMS (ESI+) m/z 225.16 ([M + H]+, 100%). Spectroscopic properties matched those previously described.15,31 1-(3-Bromoprop-1-yn-1-yl)naphthalene (11). Using a 2-step procedure 3-(naphthalen-1-yl)prop-2-yn-1-ol was prepared and converted to the bromide based on literature procedure.32 Under an argon atmosphere, to a flask charged with 1-iodonaphthalene (5.08 g, 20 mmol, 1 equiv) in THF (40 mL) were added Pd(PPh3)2Cl2 (0.24 g, 5 mol %), CuI (0.12 g, 10 mol %), and DIPEA (6.0 mL, 39.0 mmol, 3.72 equiv) followed by the addition of propargyl alcohol (4 mL, 65.6 mmol, 3.3 equiv). The reaction mixture stirred at rt for 2.5 h. The reaction mixture was diluted with saturated ammonium chloride solution (20 mL) and extracted with EtOAc (3 × 100 mL). The combined organic layers were dried over Na2SO4 and loaded onto silica for purification. The compound was purified by automated column chromatography (0−20% MeOH/DCM, 10 CV) to yield 3(naphthalen-1-yl)prop-2-yn-1-ol as a yellow oil (2.41 g, 13.2 mmol, 66%). 1H NMR (300 MHz, CDCl3): δ 4.67 (s, 2 H), 7.43 (t, J 8.0, 1 H), 7.49−7.63 (m, 2 H), 7.69 (d, J 7.0, 1 H), 7.80−7.87 (m, 2 H), 8.35 (d, J 8.0, 1 H). 13C NMR (75 MHz, CDCl3): δ 51.8, 83.8, 92.1, 120.2, 125.1, 126.0, 126.4, 126.8, 128.2, 129.0, 130.6, 133.1, 133.2. Spectroscopic properties matched those previously described.32 Upon the basis of a general Appel procedure,33 3-(naphthalen-1yl)prop-2-yn-1-ol (1000 mg, 5.49 mmol, 1 equiv) was dissolved in DCM (16 mL, 0.33M) in an ice bath, and triphenylphosphine (1.1 equiv, 6.04 mmol, 1.58 g) followed by CBr4 (1.1 equiv, 6.04 mmol, 2.00 g) was added and stirred for 20 min before concentrating the reaction mixture. The crude reaction mixture was purified by automated column chromatography (100% petroleum ether) to yield 11 as a yellow oil (2.55 g, 49% purity by 1H NMR using duroquinone as an internal standard, 5.34 mmol, 97% yield by 1H NMR, mass impurities may be CBr4). 1H NMR (300 MHz, CDCl3): δ 4.33 (s, 2 H), 7.43 (t, J 8.0, 1 H), 7.50−7.64 (m, 2 H), 7.69 (d, J 7.0, 1 H), 7.86 (d, J 8.0, 2 H), 8.31 (d, J 8.0, 1 H). 13C NMR (75 MHz, CDCl3): δ 15.5, 85.1, 89.1, 119.8, 125.2, 126.0, 126.6, 127.1, 128.4, 129.4, 131.0, 133.1, 133.4. Spectroscopic properties matched those previously described.34,35 1,8-Bis(3-(naphthalen-1-yl)prop-2-yn-1-yl)-1,4,8,11tetraazacyclotetradecane (12). To 10 (250 mg, 1.1 mmol, 1 equiv) in MeCN (12 mL, 0.1 M) were added 11 (1.34 g, 49% purity, 2.68 mmol, 2.4 equiv) and MeCN (12 mL, 0.1 M) and stirred overnight at rt to yield a white suspension. The reaction mixture was concentrated and diluted with 2.5 M aqueous NaOH (25 mL), DCM (50 mL) and shaken vigorously; after a biphasic solution formed the organic layer was dried over Na2SO4 and concentrated by rotary evaporation to give a crude brown oil (389 mg). A portion of the crude material (259 mg) was purified by automated reverse phase chromatography (0−100% H2O−MeCN (with 0.1% TFA), over 10 CV). MeCN was removed by rotary evaporation and the aqueous phase taken to pH 14 with aqueous NaOH (2.5 M) solution and extracted with DCM (3 × 30 mL). The combined organic layers were dried over Na2SO4 and concentrated to yield 12 as a brown oil (168 mg, 0.32 mmol, 29%, 43% yield based on 2/3 purified). 1H NMR (300 MHz, CDCl3): δ 1.80−1.93 (m, 4 H), 2.68−2.98 (m, 16 H), 3.82−3.81−3.94 (br m, 4 H), 7.35−7.45 (m, 2 H), 7.45−7.59 (m, 4 H), 7.63−7.69 (m, 2 H), 7.77−7.88 (m, 4 H), 8.35 (d, J 8.0, 2 H). 13C NMR (300 MHz, CDCl3): sample decomposed. LRMS (ESI+): m/z not found. FTIR (ATR) νmax/cm−1 2938, 2841, 2199, 1654. 1,8-Bis(3-(naphthalen-1-yl)propyl)-1,4,8,11-tetraazacyclotetradecane (13). To 12 (111 mg, 0.21 mmol, 1 equiv) in MeOH (10 mL, 0.02M) was backfilled with N2 (×3) and Pd(C) (1g, 10 wt %, 100 mol %) was added and the reaction mixture backfilled with N2 (×3) followed by bubbling of H2 through the solution using a long 3604

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

Journal of Medicinal Chemistry

Article

(126 MHz, DMSO-d6): δ 13.5, 24.6, 35.5, 46.7, 47.4, 48.0, 48.8, 50.9, 122.3, 123.6, 124.4, 126.0, 127.0, 128.6, 129.1, 129.5, 130.8, 131.8, 138.0, 144.1, 162.8, 163.3. LRMS (APCI) No molecular ion or fragments observed. Anal. Calce for C44H48Cl3SmN12O16·H2O: C 41.43 H 3.95 N 13.18. Found: C 41.36 H 4.33 N 12.87. FTIR (ATR) νmax/cm−1 1702, 1658, 1591, 1341, 1246, 1066, 790, 756, 623. Co(ClO4)2 Complex of 6,6′-(((1,4,8,11-Tetraazacyclotetradecane-1,8-diyl)bis(methylene))bis(1H-1,2,3-triazole-4,1-diyl))bis(2-ethyl-1H-benzo[de]isoquinoline-1,3(2H)-dione) (14f). Similar to the procedure for 14c, 1 (200 mg, 248 μmol, 1.00 equiv) in EtOH (0.1 M, 2.5 mL) and Co(ClO4)2.6H2O (90.4 mg, 248 μmol, 1.00 equiv) in EtOH (0.1 M, 2.5 mL) were refluxed for 24 h and then collected and washed to yield 14f·H2O as a beige solid (189.9 mg, 72%). Mp 250−255 °C. UV−vis (CH3CN) λmax/nm 335, ε 21044. 1H NMR (300 MHz, DMSO-d6): δ 1.26 (m, 8 H), 2.13−2.30 (m, 4 H), 2.70−3.06 (m, 16 H), 4.15 (m, 4 H), 7.98−8.17 (m, 2 H), 8.23−8.48 (m, 4 H), 8.68−8.88 (m, 4 H), 9.38 (s, 2 H), (4 × H peaks missing or overlapping). 13C NMR (126 MHz, DMSO-d6): δ 13.5, 35.7, 47.4, 123.5, 125.7, 126.4, 127.6, 128.8, 129.9, 130.8, 132.5, 136.8, 162.9, 163.3 (8 × C peaks missing or overlapping). LRMS (APCI) m/z 809.43 ([C44H48N12O12] + H)+, 433.15 (M − 2ClO4)2+, 405.20 ([C44H48N12O12] + 2 H)2+, 289.09 (M + H − 2ClO4)3+. Anal. Calcd for C44H48Cl2CoN12O12·H2O: C 44.98 H 5.15 N 14.31. Found: C 44.69 H 4.66 N 14.34. FTIR (ATR) νmax/cm−1 1702, 1659, 1590, 1341, 1245, 1083, 784, 622. Mn(II) Complex of 6,6′-(((1,4,8,11-Tetraazacyclotetradecane-1,8-diyl)bis(methylene))bis(1H-1,2,3-triazole-4,1diyl))bis(2-ethyl-1H-benzo[de]isoquinoline-1,3(2H)-dione) (14g). Similar to the procedure for 14c, 1 (200 mg, 248 μmol, 1.00 equiv) in EtOH (0.01 M, 2.5 mL) and Mn(ClO4)2·6H2O (89.4 mg, 248 μmol, 1.00 equiv) in EtOH (0.01 M, 2.5 mL) were refluxed for 24 h then collected and washed to yield 14g·5H2O as a brown solid (102.0 mg, 39%). Mp 289−295 °C. UV−vis (CH3CN) λmax/nm 343, ε 28270. 1H NMR (300 MHz, DMSO-d6): δ 1.22 (t, J 6.5, 6 H), 1.72− 1.81 (m, 4 H), 2.25−2.30 (m, 4 H), 2.62−2.69 (m, 4 H), 2.71−2.80 (m, 4 H), 3.76 (s, 4 H), 4.03−4.15 (m, 4 H), 7.86−7.93 (m, 2 H), 7.97 (d, J 7.5, 2 H), 8.08 (d, J 7.5, 2 H), 8.53−8.59 (m, 4 H), 8.67 (s, 2 H), (2 × H peaks missing or overlapping). 13C NMR (126 MHz, DMSOd6): δ 13.5, 24.6, 35.5, 46.7, 47.4, 48.0, 48.8, 50.9, 123.0, 123.7, 124.4, 126.1, 127.0, 128.6, 129.2, 129.4, 130.7, 131.8, 138.0, 144.0, 162.8, 163.3. LRMS (APCI) m/z 431.67 (M − 2ClO4)2+. Anal. Calcd for C44H48Cl2MnN12O12·3H2O: C 47.32 H 4.87 N 15.05. Found: C 47.33 H 4.47 N 15.12. FTIR (ATR) νmax/cm−1 1702, 1659, 1590, 1341, 1245, 1090, 782, 755, 622. Zn(ClO4)2 Complex of 1,8-Bis((1-(naphthalen-1-yl)-1H-1,2,3triazol-4-yl)methyl)-1,4,8,11-tetraazacyclotetradecane (15a). Similar to the procedure for 14c, 5d (82 mg, 0.133 mmol, 1.0 equiv) and Zn(ClO4)2·6H2O (49 mg, 0.133 mmol, 1.0 equiv) in EtOH (3 mM, 43 mL) were refluxed for 2 h and then collected and washed and azeotroped with toluene to yield 15a·0.1Zn(ClO4)2·0.75C7H8 as a beige solid (34 mg, 28%). Mp 241−290 °C (dec). 1H NMR (200 MHz, DMSO-d6): δ 1.99−2.12 (m, 4 H), 2.61−2.97 (br m, 8 H), 2.97−3.15 (br m, 4 H), 3.64−3.82 (br m, 4 H), 6.90−7.10 (br m, 2 H), 7.48−7.85 (m, 10 H), 8.07−8.36 (m, 4 H), 8.55−8.65 (br m, 0.6 H), 8.70 (s, 0.3 H), 8.87 (s, 0.6 H), 8.90 (s, 0.2 H), 8.92 (s, 0.1 H), 9.02 (s, 0.1 H), 9.09 (s, 0.1 H). LRMS (ESI+): m/z 340.17 [5d + Zn]2+, 615.38 [5d + H]+. Anal. Calcd for C36H42Cl2N10O8Zn·0.10 Zn(ClO4)2·0.75C7H8: C 50.88, H 4.88, N 14.38. Found: C 51.22 H 4.54 N 14.07. Cu(ClO4)2 Complex of 1,8-Bis((1-(naphthalen-1-yl)-1H-1,2,3triazol-4-yl)methyl)-1,4,8,11-tetraazacyclotetradecane (15b). Similar to the procedure for 14c, 5d (81 mg, 0.132 mmol, 1.0 equiv) and Cu(ClO4)2·6H2O (49 mg, 0.133 mmol, 1.0 equiv) in EtOH (3 mM, 43 mL) were refluxed for 2 h and then collected and washed to yield 15b·0.11Cu(ClO4)2 as a blue powder (62 mg, 52%). Mp 253−280 °C (dec). LRMS (ESI+) m/z 338.67 [5d + Cu]2+, 776.24 [5d + Cu + ClO4]+. Anal. Calcd for C36H42Cl2N10O8Cu· 0.11Cu(ClO4)2: C 47.72, H 4.67, N 15.46. Found: C 47.79 H 4.49 N 15.33. Zn(ClO4)2 Complex of 1,8-Bis(2-(4-(naphthalen-1-yl)-1H1,2,3-triazol-1-yl)ethyl)-1,4,8,11-tetraazacyclotetradecane

(16a). To a solution of 9a·3(HCl) (37 mg, 0.050 mmol) in EtOH (2 mL) was added an excess of Ambersep 900 resin (hydroxide form, preswelled with H2O for 30 min and EtOH for 30 min) in EtOH (10 mL). The mixture was stirred for 15 min and filtered. To the filtrate was added a solution of Zn(ClO4)2·6H2O (20 mg, 0.054 mmol) in EtOH (1 mL). The reaction mixture was stirred at temperature rt for 16 h. The reaction mixture was centrifuged and the precipitate was washed with ice-cold EtOH and Et2O, dried under high vacuum and lyophilized to yield 16a·H2O as an off-white solid (32 mg, 72%). Mp 188 °C (dec). LRMS (ESI+): m/z 353.1 ([M − 2ClO4]2+, 100%), 805.1 ([M − ClO4]+, 37%). HRMS (ESI+): m/z calcd for C38H46ClN10O4Zn+ [M − ClO4]+ 805.2678, found 805.2681. FTIR (ATR) νmax/cm−1: 1455, 1339, 1259, 1214, 1090, 948, 930, 885, 804, 779, 736, 622, 435. Anal. Calcd for C38H46Cl2N10O8Zn·H2O: C 49.33, H 5.23, N 15.14. Found: C 49.48, H 5.09, N 15.08. Cu(ClO4)2 Complex of 1,8-Bis(2-(4-(naphthalen-1-yl)-1H1,2,3-triazol-1-yl)ethyl)-1,4,8,11-tetraazacyclotetradecane (16b). Similar to the procedure to 16a, 9a·3(HCl) (24 mg, 0.032 mmol), Cu(ClO4)2·6H2O (13 mg, 0.035 mmol) and stirred at rt for 16 h to yield 16b as a black solid (23 mg, 80%). Mp 182 °C (dec). LRMS (ESI+): m/z 352.5 ([M − 2ClO4]2+, 100%), 803.9 ([M − ClO4]+, 23%). HRMS (ESI+): m/z calcd for C38H46ClCuN10O4+ [M − 2ClO4]2+ 352.6596, found 352.6598. FTIR (ATR) νmax/cm−1: 1462, 1427, 1352, 1085, 946, 880, 803, 778, 740, 671, 622, 565, 433. Anal. Calcd for C38H46Cl2CuN10O8: C 50.42, H 5.12, N 15.47. Found: C 50.40, H 5.40, N 15.30. Cu(ClO4)2 Complex of 6,6′-(1,1′-((1,4,8,11-Tetraazacyclotetradecane-1,8-diyl)bis(ethane-2,1-diyl))bis(1H-1,2,3-triazole4,1-diyl))bis(2-ethyl-1H-benzo[de]isoquinoline-1,3(2H)-dione) (17b). Compound 9b as a TFA salt and its Zn complex (17a) have been reported previously.15c Similar to the procedure for 17a, 9b· 3TFA (41 mg, 0.036 mmol) was complexed with Cu(ClO4)2·6H2O (13 mg, 0.035 mmol) at reflux for 16 h to yield 17b·2H2O as a brown solid (29 mg, 74%). Mp 263 °C (dec). LRMS (ESI+): m/z 449.5 ([M − 2ClO4]2+, 100%), 997.9 ([M + ClO4]+, 16%). HRMS (ESI+): m/z calcd for C46H52ClCuN12O8+ [M − ClO4]+ 998.3010, found 998.3020. FTIR (ATR) νmax/cm−1: 1694, 1650, 1613, 1587, 1447, 1385, 1371, 1341, 1244, 1062, 952, 930, 894, 872, 841, 784, 758, 622. Anal. Calcd C46H52Cl2CuN12O12·2H2O C 48.66, H 4.97, N 14.80. Found: C 48.68, H 4.93, N 14.62. Zn(ClO4)2 Complex of 1,8-Bis(2-(4-(naphthalen-2-yl)-1H1,2,3-triazol-1-yl)ethyl)-1,4,8,11-tetraazacyclotetradecane (18a). Similar to the procedure for 16a, 9c·2.5TFA (33 mg, 0.037 mmol) was complexed with Zn(ClO4)2·6H2O (14 mg, 0.038 mmol) at rt for 16 h to yield 18a as an off-white solid (27 mg, 87%). Mp 252 °C (dec). LRMS (ESI+): m/z 353.1 ([M − 2ClO4]2+, 100%), 804.9 ([M − ClO4]+, 8%). HRMS (ESI+): m/z calcd for C38H46ClN10O4Zn+ [M − ClO4]+ 805.2678, found 805.2684. FTIR (ATR) νmax/cm−1: 1454, 1335, 1238, 1075, 1057, 1013, 994, 972, 938, 930, 907, 886, 873, 842, 819, 764, 738, 623, 482. Anal. Calcd C38H46Cl2N10O8Zn: C 50.31, H 5.11, N 15.44. Found: C 50.5, H 5.11, N 15.15. Cu(ClO4)2 Complex of 1,8-Bis(2-(4-(naphthalen-2-yl)-1H1,2,3-triazol-1-yl)ethyl)-1,4,8,11-tetraazacyclotetradecane (18b). Similar to the procedure for 16a, 9c·2.5TFA (33 mg, 0.037 mmol) was complexed with Cu(ClO4)2·6H2O (14 mg, 0.037 mmol) at rt for 16 h to yield 18b·H2O as a purple solid (28 mg, 90%). Mp 191 °C (dec). LRMS (ESI+): m/z 352.5 ([M − 2ClO4]2+, 100%), 804.0 ([M + ClO4]+, 4%). HRMS (ESI+): m/z calcd for C38H46ClCuN10O4+ [M − ClO4]+ 804.2683, found 804.2680. FTIR (ATR) νmax/cm−1: 1458, 1433, 1231, 1088, 995, 938, 905, 866, 844, 819, 753, 622, 477. Anal. Calcd C38H46Cl2CuN10O8·H2O: C 49.43, H 5.24, N 15.17. Found: C 49.67, H 5.18, N 15.20. Cu(ClO4)2 Complex of 1,8-Bis(2-(4-phenyl-1H-1,2,3-triazol-1yl)ethyl)-1,4,8,11-tetraazacyclotetradecane (19b). Compound 9d as a TFA salt and Zn complex (19a) has been reported previously.15c Similar to the procedure for 16a, 9d·3TFA (25 mg, 0.028 mmol) was complexed with Cu(ClO4)2·6H2O (9.4 mg, 0.025 mmol) at reflux for 16 h to yield 19b·H2O as a purple solid (32.2 mg, 71%). Mp 186−189 °C. HRMS (ESI+): m/z calcd for C30H42ClCuN10O4+ [M − ClO4]+ 704.2370, found 704.2365. FTIR (ATR) νmax/cm−1: 1467, 1072, 976, 929, 828, 769, 738, 697, 622, 513. 3605

DOI: 10.1021/acs.jmedchem.7b01569 J. Med. Chem. 2018, 61, 3595−3608

Journal of Medicinal Chemistry

Article

Anal. Calcd for C30H42Cl2CuN10O8·H2O: C 43.77, H 5.39, N 17.02. Found: C 43.58, H 5.24, N 16.89. Zn(ClO4)2 Complex of 1,8-Bis(2-(4-benzyl-1H-1,2,3-triazol-1yl)ethyl)-1,4,8,11-tetraazacyclotetradecane (20a). Similar to the procedure for 16a, 9e·3(HCl) (40 mg, 0.059 mmol) was complexed with Zn(ClO4)2·6H2O (24 mg, 0.064 mmol) at rt for 16 h to yield 20a as a white solid (42 mg, 86%). Mp 129−132 °C. LRMS (ESI+): m/z 317.1 ([M − 2ClO4]2+, 100%), 733.0 ([M − ClO4]+, 23%). HRMS (ESI+): m/z calcd for C32H46ClN10O4Zn+ [M − ClO4]+ 733.2678, found 733.2677. FTIR (ATR) νmax/cm−1: 3261, 2873, 1495, 1454, 1331, 1228, 1071, 1030, 974, 930, 906, 886, 838, 809, 729, 700, 621, 464. Anal. Calcd for C32H46Cl2N10O8Zn: C 46.03, H 5.55, N 16.77. Found: C 45.91, H 5.43, N 16.63. Cu(ClO4)2 Complex of 1,8-Bis(2-(4-benzyl-1H-1,2,3-triazol-1yl)ethyl)-1,4,8,11-tetraazacyclotetradecane (20b). Similar to the procedure for 16a, 9e·3(HCl) (40 mg, 0.059 mmol) was complexed with Cu(ClO4)2·6H2O (24 mg, 0.065 mmol) at rt for 16 h to yield 20b as a vibrant purple solid (27 mg, 55%). Mp 146−148 °C LRMS (ESI +): m/z 316.5 ([M − 2ClO4]2+, 100%), 732.1 ([M − ClO4]+, 16%). HRMS (ESI+): m/z calcd for C32H46ClCuN10O4+ [M − ClO4]+ 732.2683, found 732.2680. FTIR (ATR) νmax/cm−1: 1495, 1454, 1428, 1070, 1029, 929, 834, 730, 699, 621, 463. Anal. Calcd for C32H46Cl2CuN10O8: C 45.15, H 5.68, N 16.45. Found: C 45.15, H 5.61, N 16.38. Zn(ClO 4 ) 2 Complex of 6,6′-(((1,4,8,11-Tetraazacyclotetradecane-1,8-diyl)bis(methylene))bis(1H-1,2,3-triazole-4,1diyl))bis(2-(3-hydroxypropyl)-1H-benzo[de]isoquinoline1,3(2H)-dione) (21a). 5i (38.5 mg, 0.045 mmol, 1.0 equiv) was dissolved in EtOH (1.4 mL), was heated to reflux, and Zn(ClO4)2 (17.8 mg, 0.048 mmol, 1.0 equiv) in Et2O (0.46 mL) was added and heated at reflux for 24 h. The solution was allowed to cool to rt and the precipitate collected by vacuum filtration and washed with EtOH (3 × 3 mL) and dissolved in MeCN/H2O (5 mL) and filtered, then freeze-dried to yield 21a·0.45H2O as a yellow fluffy solid (35 mg, 0.031 mmol, 69%). 1H NMR (300 MHz, D2O) δ 1.82−1.94 (m, 4 H), 2.09− 2.22 (m, 4 H), 2.85−3.00 (m, 4 H), 3.05−3.16 (m, 4 H), 3.43−3.52 (m, 4 H), 3.64−3.73 (m, 4 H), 3.96−4.04 (m, 4 H), 4.05−4.16 (m, 4 H), 7.22−7.32 (m, 2 H), 7.59−7.68 (m, 4 H), 8.05−8.14 (m, 2 H), 8.21−8.28 (m, 2 H), 8.29−8.33 (m, 2 H) (8 × H missing or overlapping). Anal. Calcd for C46Cl2H52N12O14Zn·0.45H2O: C 48.64, H 4.64, N 14.80. Found: C 49.00, H 5.02, N 14.60. ZnCl2 Complex of 6,6′-(((1,4,8,11-Tetraazacyclotetradecane1,8-diyl)bis(methylene))bis(1H-1,2,3-triazole-4,1-diyl))bis(2-(3hydroxypropyl)-1H-benzo[de]isoquinoline-1,3(2H)-dione) (21b). 5i (1000 mg, 1.15 mmol, 1.0 equiv) in H2O (Milli-Q, 70 mL) was heated to reflux, and ZnCl2 (1.0 M in diethyl ether stock solution, 1.15 mmol, 1.0 equiv) was added and refluxed for 2 h. The solution was cooled to rt and concentrated under reduced pressure (to