Room-Temperature Synthesis of Re(I) and Tc(I) Metallocarboranes

Article pubs.acs.org/Organometallics

Room-Temperature Synthesis of Re(I) and Tc(I) Metallocarboranes Mohamed E. El-Zaria,† Nancy Janzen, and John F. Valliant* Department of Chemistry and Chemical Biology, McMaster University, 1280 Main Street W., Hamilton, Ontario, L8S 4M1, Canada S Supporting Information *

ABSTRACT: A series of carborane derivatives bearing guanidine substituents were prepared and characterized, and their reactivity toward Re(I) and Tc(I) in aqueous media was evaluated. Guanidinylation was achieved by treating 1-aminomethyl-1,2closo-dodecaborane with N1,N2-di-Boc-1H-pyrazole-1-carboxamidine, and the associated N-ethyl derivative, which produced the desired products in good (circa 50%) yield. These were deprotected and converted to the corresponding nido-carboranes, which, when combined with [M(CO)3(H2O)3]+ (M = Re and 99mTc) at room temperature for 3 h or 35 °C for 1 h, afforded the corresponding η5-metallocarborane complexes. Corresponding reactions involving carboranes without basic substituents generally require microwave heating at temperatures greater than 150 °C. The rate, yields, and the temperature of the reaction appear to be dependent on the basicity of the guanidines tested. The biodistribution of two of the 99mTc complexes, which are stable indefinitely in solution, were evaluated in vivo in CD1 mice and showed that the 99mTc−carboranyl guanidine complexes clear key nontarget organs and tissues within one half-life (6 h) and have properties that are desirable for developing targeted molecular imaging probes.

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INTRODUCTION

synthesis of Re and Tc metallocarboranes still requires microwave heating at high temperatures (150−200 °C).7 The high labeling temperature notwithstanding, carboranes are attractive synthons for preparing radiopharmaceuticals8 because the ligands can be prepared bearing a wide range of different functional groups at one or more of the carbon and/or boron vertices using well-established synthetic procedures. This confers flexibility when developing bioconjugation targeting strategies.9 There is also a wealth of carborane−biomolecule derivatives that have been prepared and screened for boron neutron capture therapy (BNCT), which represent a pool of promising and advanced candidates to design new radiopharmaceuticals for cancer imaging applications.10 Unfortunately, the high temperature required to form Tc/Re metallocarboranes in aqueous solutions is a barrier that must be overcome before the cluster’s attractive properties can be exploited. To form Tc(I) metallocarborane complexes, the bridging hydrogen atom on the nido-carborane must be removed (deprotonation).11 During the preparation of metal complexes of pyridine−carborane derivatives, we noted a significant reduction in the labeling temperature compared with that of the aryl and cyclohexyl analogues.7c These results suggested

Organometallic chemistry plays a key role in diagnostic nuclear medicine where a 99mTc(I)−hexakisisonitrile complex has been used for decades for myocardial perfusion and breast cancer imaging.1 More recently, the focus has shifted away from perfusion-type tracers to developing targeted molecular imaging probes derived from Tc(I).2 The latter is made possible by the discovery of a convenient method to produce [Tc(CO)3(OH2)3]+ in water3 from 99mTcO4−, which can be obtained from commercial 99Mo/99mTc generators. The majority of targeted compounds derived from [Tc(CO)3(OH2)3]+ are prepared using bifunctional tridentate chelates linked to some type of delivery vector.4 As an alternative to coordination complexes, the availability of the [Tc(CO)3]+ core affords the opportunity to prepare η5-type organometallic compounds as compact synthons for creating targeted nuclear imaging agents. Work to date has utilized cyclopentadiene (Cp)5 and carborane ligands, which form robust complexes with Tc(I). One of the limitations for both classes of ligands, however, is that reaction temperatures that are required to achieve good radiochemical yields are not compatible with most targeting vectors. In the case of Cp, direct labeling at reduced temperatures (99% in each case. Furthermore, the purification procedure removed the excess ligand used during the labeling, making it possible to produce the complexes in high effective specific activity.

The stabilities of the metallocarboranes were assessed by dissolving the 99mTc complexes in 5% EtOH−saline and analyzing the samples by HPLC every hour for 8 h. Under multiple elution conditions in which the pH and composition of the HPLC eluent were varied, there was no evidence for degradation over the entire duration of the study for either complex. This is consistent with other reports, which showed that Tc-metallocarboranes are robust.7 Biodistribution Studies. The introduction of guanidine ancillary groups makes it possible to form metallocarborane complexes under mild conditions. The strategies that could be employed to make targeted derivatives would be to link a vector to the other cluster carbon atom or the guanidine residue itself. Irrespective of the approach, a key question is 5944

dx.doi.org/10.1021/om300521j | Organometallics 2012, 31, 5940−5949

Organometallics

Article

Figure 6. In vivo biodistribution data for 15 in female CD1 mice. Mice were injected with 0.40 MBq of 15 sacrificed at the indicated time points. Data are expressed as %ID/g.

Figure 7. In vivo biodistribution data for 16 in female CD1 mice. Mice were injected with 0.26 MBq of 16 and sacrificed at the indicated time points. Data are expressed as %ID/g.

uptake. The major sites for initial localization were the gall bladder, liver, kidneys, stomach, intestines, thyroid, and bladder. The uptake of 15 and 16 in the liver was similar at 1 h, where levels of 15 decreased significantly, reaching 3.24 ± 0.48%ID/g at 24 h, which is 3-fold lower than that of 16. While 15 showed the highest concentrations in the gall bladder (68.01 ± 15.03% ID/g at 1 h; 42.11 ± 16.99%ID/g at 4 h; 1.07 ± 0.20%ID/g at 24 h), indicating clearance via the hepatobiliary system, there was markedly reduced uptake in the gall bladder for 16 (13.06 ± 5.74%ID/g at 1 h; 11.56 ± 5.57%ID/g at 4 h; 0.67 ± 0.24% ID/g at 24 h) and significantly more activity in the urine/ bladder. In general, the reduction in background (nonspecific)

with regards to the impact of the guanidine group on the in vivo distribution of the cluster. Hydrophilic substituents are attractive in that they decrease the hydrophobic nature of carboranes, which tend to promote unwanted accumulation of metallocarboranes in the liver. To assess their in vivo properties, the 99mTc−carboranyl guanidine complexes were administered separately to healthy female CD1 mice (260−400 kBq per mouse). Tissues were collected at 1, 4, and 24 h postinjection (pi) and counted (Figures 6 and 7), and the data were reported as the percent injected dose per gram (%ID/g). Compounds 15 and 16 cleared the blood rapidly and showed low heart and lung 5945

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Organometallics

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saturated sodium bicarbonate, and brine. After drying with sodium sulfate and filtering, the solvent was removed under reduced pressure. The desired product was isolated by silica gel chromatography using CH2Cl2−hexane (2:1) as a mobile phase. Compound 3 was obtained as a white crystalline solid (0.10 g, 25%). Rf = 0.47, mp = 136−138 °C. 1 H NMR (600 MHz, CDCl3): δ 11.30 (s, 1H, NHBoc), 8.80 (t, 1H, J = 6.0 Hz, NHCH2), 4.24 (s, 1H, CH−carborane), 4.08 (d, 2H, J = 6.0 Hz, CH2N), 1.60−2.80 (bm, 10H, BH), 1.52, 1.48 (s, 18H, CH3). 13C NMR (150 MHz, CDCl3): δ 162.7, 156.4 (CO), 152.9 (CN), 84.2, 79.9 (C(CH3)3), 74.6 (C−carborane), 60.9 (CH−carborane), 45.4 (CH2), 28.2, 28.0 (CH3). 11B NMR (192 MHz, CDCl3): δ −1.8, −4.8, −9.6, −11.5, −12.8. IR (KBr, cm−1): ν 3308, 3270, 2596, 1733, 1540. HRMS (ESI, positive) m/z for C14H33B10N3O4: Calcd 416.3560. Found 416.3551 [M + H]+. Method B: A stirring solution of 1 (0.20 g, 0.95 mmol) and triethylamine (0.19 g, 266 μL, 1.90 mmol) in dry CH2Cl2 (5 mL) was treated with N1,N2-di-Boc-1H-pyrazole-1-carboxamidine (2) (0.30 g, 0.95 mmol), and the mixture was allowed to react at room temperature for 24 h. The mixture was diluted with CH2Cl2 (10 mL) and extracted with 2 M sodium bisulfate, saturated sodium bicarbonate, and brine. The solution was subsequently filtered through Celite, which was washed twice with CH2Cl2, and the filtrates were combined and evaporated. The desired product was isolated by silica gel chromatography (CH2Cl2−hexane mixtures 1:2) as a white colorless crystalline solid (0.22 g, 55%). Synthesis of [(HCB10H10C)CH2NHC(NH2)NH2][CF3CO2] (4). Trifluoroacetic acid (5 mL) was added to a solution of 3 (0.10 g, 0.24 mmol) in CH2Cl2 (5 mL) at 0 °C, and the mixture was stirred at room temperature for 45 min. The reaction mixture was evaporated under vacuum to dryness, and the resulting white residue was redissolved in a minimum amount of CH2Cl2 (2 mL) and treated with hexane (1 mL). After 30 min at room temperature, 4, as a colorless crystalline solid (0.08 g, 97%), was isolated by filtration. mp = 152−54 °C. 1H NMR (600 MHz, d6-DMSO): δ 7.94 (t, 1H, J = 6.0 Hz, NHCH2), 7.38 (bs, NH), 5.09 (s, 1H, CH−carborane), 4.06 (d, 2H, J = 6.0 Hz, CH2N), 2.55−1.5 (bm, 10H, BH). 13C NMR (150 MHz, d6DMSO): δ 159.4 (m, CO), 156.9 (CN), 117.9 (m, CF3), 75.5 (C−carborane), 62.5 (CH−carborane), 44.2 (CH2). 11B NMR (192 MHz, d6-DMSO): δ −3.2, −5.6, −9.1, −11.4, −13.0, −13.5. IR (KBr, cm−1): 3472, 3372, 2589. HRMS (ESI, positive) m/z for C4H18B10N3: Calcd 217.2471. Found 217.2431 [M + H]+. Synthesis of (HCB9H10C)CH2NHC(NH2)NH2 (5). Compound 4 (0.05 g, 0.15 mmol) and sodium fluoride (0.03 g, 0.75 mmol) were combined in ethanol−water (3:2, 3 mL), and the mixture was heated to 85 °C for 3 h. Afterward, the solution was concentrated to dryness and water (5 mL) was added. The suspension was filtered, and the white solid was washed with water (2 mL) and dried on the filter to give 5 as a colorless solid (0.03 g, 95%). Rf = 0.36 in CH2Cl2/MeOH (9:1), mp = 263−265 °C. 1H NMR (600 MHz, d6-DMSO): δ 7.12 (t, 1H, J = 6.0 Hz, HNCH2), 6.65 (bs, NH), 3.21 (m, 1H, CH2N), 3.16 (d, 1H, CH−carborane), 3.06 (m, 1H, CH2N), 2.4 to −0.55 (bm, 10H, BH), −2.71 (bm, 1H, μ-H). 13C NMR (150 MHz, d6-DMSO): δ 156.2 (CN), 56.8 (C−carborane), 49.2 (CH2), 44.4 (CH− carborane). 11B NMR (192 MHz, d6-DMSO): δ −10.9, −15.02, −16.5, −18.2, −19.2, −22.0, −32.8, −37.1. IR (KBr, cm−1): 3474, 3372, 2534. HRMS (ESI, positive) m/z for C4H18B9N3Na: Calcd 228.2310. Found 228.2344 [M + Na]+. Synthesis of [(HCB10H10C)CH2NHC(NH2)NH2][NO3] (6). Nitric acid (2.50 mL, 6 M) was added to a solution of 3 (0.10 g, 0.24 mmol) in MeOH (5 mL), and the mixture was heated to 60 °C. After approximately 10 min, a white crystalline solid formed, which dissolved after an additional 15 min; heating was continued for 2 h. The reaction mixture was cooled to room temperature, giving a white crystalline solid that was collected by filtration and washed twice with cold water (2 mL) to yield 6 as small needle-shaped crystals (0.06 g, 97%). mp > 300 °C. 1H NMR (600 MHz, d6-DMSO): δ 7.92, 7.37 (bs, NH), 5.09 (s, 1H, CH−carborane), 4.08 (d, 2H, J = 6.0 Hz, CH2N), 3.81(bs, 1H, NH), 2.65−1.55 (bm, 10H, BH). 13C NMR (150 MHz, d6-DMSO): δ 156.6 (CN), 75.3 (C−carborane), 62.5 (CH− carborane), 44.2 (CH2). 11B NMR (192 MHz, d6-DMSO): δ −3.1,

binding in critical organs and tissues after 4 h is sufficient to warrant the use of the ligands to prepare targeted agents. In addition, it is apparent that changing the nature of the guanidine not only impacts the labeling temperature and yield but also can be used to fine-tune pharmacokinetics and route of clearance in vivo.

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CONCLUSIONS A series of carborane guanidine derivatives were prepared in good to excellent yields, and X-ray structures of several closo and nido clusters were obtained. Labeling with Re and 99mTc was achieved at room temperature for the first time, where isomerization from the 3,1,2 to the 2,1,8 clusters was not observed. The yields of the reaction appear to be related to the basicity of the guanidine with the most basic substituent producing the metal complexes in the highest yields. These results now make it possible to radiolabel targeted carboranes at room temperature in aqueous solution, opening the door to the preparation of a new generation of organometallic molecular imaging probes.

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EXPERIMENTAL SECTION

General Methods. Chemicals and reagents were purchased from Sigma-Aldrich, with the exception of decaborane, which was purchased from Katchem (Czech Rep.). Solvents were purchased from Caledon and dried using a PureSolv drying apparatus (Innovative Technology). [99mTc(CO)3(OH2)3]+,3 Re(CO)3(OH2)3]Br,25 N1,N2-di-Boc-N3-trifluoromethanesulfonyl guanidine,20a and 1-aminomethyl-1,2-dicarbacloso-dodecaborane hydrochloride salt21 were prepared according to literature methods. 1H, 13C, and 11B NMR spectra were measured on a Bruker Avance AV-600 (1H = 600.13 MHz, 13C = 150.90 MHz, 11B = 192.55 MHz) spectrometer. 1H NMR and 13C NMR chemical shifts are expressed in parts per million (ppm, δ units), and coupling constants are expressed in hertz (Hz). The chemical shifts of 11B NMR were reported relative to an external standard of BF3·Et2O in CDCl3. IR spectra were recorded on a Nicolet 6700 FT-IR spectrometer. Lowresolution mass spectra were obtained on an Agilent 630 ion trap electron spray ionization (ESI) instrument, using a 1200 series LC system H2O/MeOH (1:1). High-resolution mass spectra (HRMS) were obtained using a Waters Micromass Global Ultima Q-TOF in ESI mode. A 99Mo/99mTc generator was provided by Lantheus Medical Imaging (Hamilton). A CAPINTEC CR-25R dose calibrator was used for measuring the amount of radioactivity employed during the radiosynthesis protocols. For 99mTc, high-performance liquid chromatography (HPLC) was performed with a Waters 1525 Binary HPLC system fitted with a 2998 photodiode array detector (254 nm) and a Bioscan γ detector. For analysis of compounds, a Phenomenex Gemini column (5 μm, 4.6 × 250 mm, C18) at a flow rate of 1.0 mL/ min and monitoring at 254 nm was used. For semipreparative HPLC, a Phenomenex Luna column (5 μm, 10.0 × 250 mm, C18) at a flow rate of 4.0 mL/min and monitoring at 254 nm was employed. HPLC protocols used were as follows: Analytical and semipreparative HPLC: solvent A = water containing 0.1% TFA; solvent B = acetonitrile containing 0.1% TFA; gradient elution, 40% B (0−20 min), 100% B (20−22 min), 40% B (22−25 min). When producing 99mTc-labeled compounds for biodistribution studies, the same gradient was used; however, TFA was omitted from the eluents. Analytical thin-layer chromatography (TLC) was performed on glass plates of silica gel 60 GF254 (Merck). Column chromatography was conducted on silica gel (Merck Kieselgel 70−230 mesh). Development of plates was performed using a 0.1 M PdCl2 in 3 M HCl spray. Synthesis of (HCB10H10C)CH2NHC(NBoc)NHBoc (3). Method A: Compound 1 (0.20 g, 0.95 mmol) was added to a solution of N1,N2-di-Boc-N3-trifluoromethanesulfonyl guanidine (0.34 g, 0.95 mmol) and triethylamine (0.19 g, 266 μL, 1.90 mmol) in dry CH2Cl2 (5 mL) at room temperature. After 2 days, the mixture was diluted with CH2Cl2 (10 mL) and extracted with 2 M sodium bisulfate, 5946

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Article

−5.3, −9.5, −11.8, −13.2. IR (KBr, cm−1): 3345, 3197, 2580. HRMS (ESI, positive) m/z for C4H18N3B10: Calcd 217.2471. Found 217.2444 [M]+. Synthesis of [(HCB10H10C)CH2NHC(NH2)NNO2] (7). Concentrated sulfuric acid (1 mL) in a 10 mL flask was cooled to −5 °C, and compound 6 (0.20 g, 0.76 mmol) was added slowly in small portions such that the reaction was kept below 0 °C. After 2 h at 0 °C, the reaction was quenched with ice water, whereupon a precipitate formed when the mixture was allowed to stand with cooling at 0 °C for another 1 h. Following filtration, the residue was washed with water and then vigorously with hot water (5 mL, ×2) prior to drying. The desired product 7 was obtained as a white solid (0.14 g, 70%). mp = 203−205 °C. 1H NMR (600 MHz, d6-DMSO): δ 8.15 (bs, NH), 7.88 (m, 1H, NHCH2), 7.34 (bs, CNH2), 5.04 (s, 1H, CH−carborane), 4.01 (m, 2H, CH2N), 2.65−1.55 (bm, 10H, BH). 13C NMR (150 MHz, d6-DMSO): δ 158.8 (CN), 75.3 (C−carborane), 61.9 (CH− carborane), 44.2 (CH2). 11B NMR (192 MHz, d6-DMSO): δ −3.0, −5.5, −9.9, −11.7,−13.0. IR (KBr, cm−1): 3415, 3321, 2593, 1528. HRMS (ESI, positive) m/z for C4H16B10N4O2Na: Calcd 282.2212. Found: 282.2243 [M + + Na]. HRMS (ESI, negative) m/z C4H15B10N4O2: Calcd 259.2201. Found 259.2219 [M − H]−. Synthesis of (HCB9H10C)CH2NHC(NH2)NHNO2 (8). Compound 7 (0.05 g, 0.19 mmol) and sodium fluoride (0.04 g, 0.95 mmol) were combined in 3:2 v/v aqueous ethanol (3 mL), and the suspension was heated to 85 °C for 5 h. After concentrating the solution to dryness, the desired product 8 was isolated by silica gel chromatography (9:1 CH2Cl2/MeOH as a colorless oil (0.03 g, 75%). Rf = 0.23. 1H NMR (600 MHz, d6-DMSO): δ 8.91, 7.60, 6.75 (bs, NH), 3.16 (d, 2H, J = 6.0 Hz, CH2N), 1.78 (s, 1H, CH−carborane), 2.17 to −0.33 (bm, 10H, BH), −2.78 (bs, 1H, μ-H). 13C NMR (150 MHz, d6-DMSO): δ 158.5 (CN), 57.2 (C−carborane), 48.9 (CH−carborane), 44.4 (CH2). 11B NMR (192 MHz, d6-DMSO): δ −10.8, −15.01, −16.4, −18.5, −19.0, −22.0, −33.0, −37.1. IR (KBr, cm−1): 3419, 3304, 2523, 1535. HRMS (ESI, negative) m/z for C4H16B9N4O2: Calcd 250.2151. Found 250.2119 [M]−. N1,N2-Bis(tert-butyloxycarbonyl)-N2-ethyl-1H-pyrazole-1carboxamidine (9). To a suspension of sodium hydride (0.02 g, 1.00 mmol) in dry DMF (5 mL) was added 2 (0.31 g, 1.00 mmol). The suspension turned homogeneous after 5 min at room temperature, whereupon bromoethane (0.11 g, 1.00 mmol) in DMF (2 mL) was added dropwise. Once TLC indicated the complete consumption of N1,N2-di-Boc-1H-pyrazole-1-carboxamidine (typically 3 days), the reaction mixture was partitioned between water and ethyl acetate, and the desired product 9 was isolated by silica gel column chromatography (1:4 ethyl acetate/hexane) as a colorless oil (0.31 g, 90%). Rf = 0.37. 1H NMR (600 MHz, CDCl3): δ 7.9 (s, 1H, ArH), 7.62 (s, 1H, ArH), 6.36 (s, 1H, ArH), 3.67 (s, 2H, CH2NBoc), 1.44 (s, 9H, NBoc), 1.22 (m, 12H, CH3). 13C NMR (150 MHz, CDCl3): δ 157.5 (CO), 152.2 (CN), 143.0 (CAr), 129.8 (CAr), 109.0 (CAr), 82.3 (O−C), 43.9 (NCH2), 27.9 (CH3), 21.7 (CH3), 13.4 (CH3). IR (KBr, cm−1): 3347, 3258, 1790, 1726. HRMS (ESI, positive) m/z for C16H26N4O4: Calcd 339.2032. Found 339.2015 [M + H]+. Synthesis of (HCB10H10C)CH2NHC(NBoc)N(Et)Boc (10). Compound 1 (0.21 g, 1.00 mmol) was added to a solution of 9 (0.34 g, 1.00 mmol) and triethylamine (0.20 g, 280 μL, 2.0 mmol) in dry CH2Cl2 (5 mL) under argon. The mixture was stirred at room temperature for 24 h and subsequently diluted with CH2Cl2 (10 mL) and extracted with 2 M sodium bisulfate, saturated sodium bicarbonate, and brine. After drying with sodium sulfate and filtering, the solvent was removed under reduced pressure. The desired product 10 was isolated by silica gel chromatography (3:1 CH2Cl2−hexane) as a white crystalline solid (0.20 g, 45%). Rf = 0.29, mp = 138−140 °C. 1H NMR (600 MHz, CDCl3): δ 8.47 (t, 1H, J = 6.0 Hz, NHCH2), 4.17 (s, 1H, CH− carborane), 3.90 (m, 2H, CH2N), 3.66 (bs, 2H, CH2NBoc), 1.80−2.53 (bm, 10H, BH), 1.50 (s, 18H, CH3), 1.17 (t, 3H, J = 6.0 Hz, CH3). 13 C NMR (150 MHz, CDCl3): δ 153.9 (CO), 152.9 (CN), 83.9, 83.0 (C(CH3)3), 82.1 (C−carborane), 59.9 (CH−carborane), 45.0 (HNCH2), 42.7 (NCH2), 28.2, 28.1 (BocCH3), 13.9 (CH3). 11B NMR (192 MHz, CDCl3): δ −2.8, −5.3, −9.2, −11.4, −12.9. IR (KBr,

cm −1 ): 3270, 2589, 1748. HRMS (ESI, positive) m/z for C16H37B10N3O4: Calcd 444.3874. Found 444.3901 [M + H]+. Synthesis of [(HCB10H10C)CH2NHC(NH2)NHEt][CF3CO2] (11). Compound 11 was prepared from 10 (0.10 g, 0.22 mmol) and trifluoroacetic acid (10 mL) following the procedure described for 4, to give 11 (0.08 g, 100%) as a colorless crystalline solid. mp = 131− 133 °C. 1H NMR (600 MHz, d6-DMSO): δ 8.14 (t, 1H, J = 6.0 Hz, HNEt), 7.99 (t, 1H, J = 6.0 Hz, NHCH2), 7.75 (bs, CNH2), 5.12 (s, 1H, CH−carborane), 4.11 (d, 2H, J = 12.0 Hz, CH2N), 3.18 (q, 2H, J = 12.0 Hz, CH2), 2.55−1.55 (bm, 10H, BH), 1.10 (t, 3H, J = 6.0 Hz, CH3). 13C NMR (150 MHz, d6-DMSO): δ 158.5 (m, CO), 155.3 (CN), 118.1 (m, CF3), 75.6 (C−carborane), 62.4 (CH−carborane), 44.1 (CH2NH), 36.1 (CH2), 13.9 (CH3). 11B NMR (192 MHz, d6DMSO): δ −3.1, −5.3, −9.5, −11.8, −13.0. IR (KBr, cm−1): 3306, 3193, 2578. HRMS (ESI, positive) m/z for C6H22B10N3: Calcd 244.2820. Found 244.2815 [M]+. Synthesis of (HCB9H10C)CH2NHC(NH2)NHEt (12). Compound 12 was prepared from 11 (0.10 g, 0.28 mmol) and NaF (0.06 g, 1.42 mmol) following the procedure described for 5, to give 12 (0.06 g, 100%) as a colorless crystalline solid. mp = 285−287 °C. 1H NMR (600 MHz, d6-DMSO): δ 7.24, 6.91 (bs, NH), 3.24 (m, 1H, CH− carborane), 3.11 (m, 4H, CH2, NCH2), 2.48 to −0.75 (bm, 10H, BH), 1.08 (t, 3H, J = 6.0 Hz, CH3), −2.78 (bm, 1H, μ-H). 13C NMR (150 MHz, d6-DMSO): δ 155.0 (CN), 56.9 (C−carborane), 49.1 (CH2NH), 44.4 (CH−carborane), 35.9 (CH2), 14.1 (CH3). 11B NMR (192 MHz, d6-DMSO): δ −10.9, −15.0, −16.6, −18.3, −19.1, −19.5, −22.0, −32.9, −37.2. IR (KBr, cm−1): 3447, 3237, 2522. HRMS (ESI, negative) m/z for C6H21B9N3: Calcd 233.2614. Found 233.2600 [M]−. Synthesis of (η5-Re(CO)3HCB9H9C)CH2NHC(NH2)NH2 (13). Compound 5 (0.05 g, 0.21 mmol) was combined with Re[(CO)3(H2O)3]Br (0.09 g, 0.21 mmol) in 1:1 v/v aqueous ethanol (3 mL). The pH of the resulting reaction mixture was adjusted to 10 using 1 M NaOH. The reaction was stirred at room temperature for 3 h or 35 °C for 1 h, and the solvent was removed by rotary evaporation. Water (1 mL) was added to the residue, and the solution was filtered and lyophilized, giving a white solid prior to purification by silica gel chromatography (1:9 MeOH/CH2Cl2). The compound was further purified by semipreparative HPLC to yield 13 as a white solid (0.03 g, 30%). Rf = 0.49, mp = 225−227 °C. 1H NMR (600 MHz, d6-DMSO): δ 7.36 (bs, CNH2), 7.26 (t, 1H, J = 6.0 Hz, NHCH2), 6.96 (bs, CNH2), 4.01 (s, 1H, CH−carborane), 3.74 (m, 1H, CH2N), 3.65 (m, 1H, CH2N), 2.53−1.19 (bm, 10H, BH). 13C NMR (150 MHz, d6DMSO): δ 197.9 (Re(CO)3), 157.7 (m, CF3CO), 156.2 (CN), 118.6 (m, CF3), 60.8 (C−carborane), 51.7 (CH−carborane), 46.2 (CH2). 11B NMR (192 MHz, d6-DMSO): δ −6.0, −9.9, −10.4, −14.6, −20.6, −22.1. IR (KBr, cm−1): 3464, 3178, 2527, 2006, 1904. HRMS (ESI, negative) m/z for ReC7H16B9N3O3: Calcd 476.1600. Found 476.1557 [M]−. Synthesis of (η5-Re(CO)3HCB9H9C)CH2NHC(NH2)NHEt (14). Compound 14 was prepared from 12 (0.05 g, 0.19 mmol) and Re[(CO)3(H2O)3]Br (0.08 g, 0.19 mmol) following the procedure described for 13, to give 14 (0.05 g, 47%) as a colorless crystalline solid. Rf = 0.55 (MeOH/CH2Cl2 1:9), mp = 195−197 °C. 1H NMR (600 MHz, d6-DMSO): δ 7.38, 7.36, 6.99 (bs, NH,), 3.96 (s, 1H, CH− carborane), 3.74 (m, 1H, CH2N), 3.65 (m, 1H, CH2N), 3.14 (m, 2H, CH2), 2.55−1.05 (bm, 10H, BH), 1.08 (t, 3H, J = 6.0 Hz, CH3). 13C NMR (150 MHz, d6-DMSO): δ 197.9 (Re(CO)3), 157.8 (m, CF3CO), 154.8 (CN), 118.4 (m, CF3), 51.6 (CH−carborane), 46.1 (carboraneCH2), 40.0 (C−carborane), 35.9 (CH2), 14.1 (CH3). 11 B NMR (192 MHz, d6-DMSO): δ −6.1, −9.7, −10.1, −14.7, −20.8, −22.5. IR (KBr, cm−1): 3497, 3396, 2537, 1999, 1893. HRMS (ESI, negative) m/z for ReC9H20B9N3O3: Calcd 502.1955. Found: 502.1967 [M]−. Synthesis of (η5-Tc(CO)3HCB9H9C)CH2NHC(NH2)NH2 (15). A 500 μL portion of [99mTc(CO)3(H2O)3]+ (218 MBq) and 500 μL of a 1 mg/mL (EtOH/H2O 1:1) solution of 5 were combined in a reaction vial (0.5−2 mL) purged with argon. The vial was heated to 35 °C for 1 h. The reaction was monitored by analytical HPLC by first diluting a small sample (∼1 μL) with 500 μL of ethanol. The product was 5947

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Organometallics subsequently purified by semipreparative HPLC, where the solvent was removed using a Biotage V10 evaporator system to yield 15 in 47% radiochemical yield. tr = 11.43 min. Synthesis of (η5-Tc(CO)3HCB9H9C)CH2NHC(NH2)NHEt (16). Compound 16 was prepared from 12 and [99mTc(CO)3(H2O)3]+ (500 μL, 206 MBq) following the procedure described for 15, to give 16 in 69% radiochemical yield. tr = 12.75 min. Stability Study. Following isolation, the 99mTc−carboranyl guanidine complexes 15 and 16 were dissolved in 5% EtOH in 0.9% saline (3 mL). The stability of the complexes was assessed by HPLC using different elution conditions (mobile phase and pH). Biodistribution Study. Biodistribution of compounds 15 and 16 was performed using female CD1 mice (Charles River Laboratories, Senneville, QC, Canada). Three mice were used per time point (t = 1, 4, and 24 h. The mice were administered 0.40 MBq or 0.26 MBq of 15 and 16, respectively (100 μL in 5% EtOH/saline), via tail vein injection. Animals were anaesthetized with 3% isoflurane and euthanized by cervical dislocation. Blood, heart, lungs, liver, gall bladder, spleen, kidneys, adrenals, stomach (with contents), small intestines (with contents), large intestine and cecum (with contents), adipose, brain, thyroid/trachea, bone, skeletal muscle, bladder and urine, and tail were collected, weighed, and counted in a PerkinElmer Wizard 1470 Automated Gamma Counter. Decay correction was used to normalize organ activity measurements to time of dose preparation for data calculations with respect to injected dose (i.e., %ID/g). X-ray Structure Determination of 4, 5, 7, 11, and 12. Colorless single crystals of 4 and 11 suitable for X-ray analyses were obtained from a mixture of CH2Cl2−hexane at room temperature, whereas single crystals of 5, 7, and 12 were obtained by slow evaporation of methanol−water solution at room temperature. Crystals suitable for analysis were chosen and mounted in oil on a MiTeGen head and placed in the cold stream (100 K) of the diffractometer. Data were collected on a Bruker APEX2 diffractometer equipped with a SMART6000 area detector, using Cu Kα radiation (λ = 1.54178 Å) or Mo Kα radiation (λ = 0.71073 Å), and ω and φ scans. Unit cell parameters were determined using at least 50 frames from three different orientations. Data were processed using SAINT26 and corrected for absorption using SADABS, then solved using direct methods with the SHELXTL27 program suite. All non-hydrogen atoms were refined anisotropically, and hydrogen atoms were located and refined isotropically. Hydrogen atoms at the ethyl terminus were placed in idealized positions, riding on their constituent atoms, and updated with each cycle of refinement. Hydrogen atoms on N1 and N2 were located and then fixed, and updated with each cycle of refinement. Hydrogen atoms on the carborane cage were located and allowed to refine isotropically without fixing to their parent atoms.

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ACKNOWLEDGMENTS

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REFERENCES

The authors gratefully acknowledge the financial support of the Natural Science and Engineering Research Council (NSERC) of Canada and McMaster University for this work.

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ASSOCIATED CONTENT

S Supporting Information *

X-ray crystallographic files for 4, 5, 7, 11, and 12 in CIF format; tables of complete data for tissue distribution of 15 and 16 in CD1 mice; HRMS spectra; 11B, 11B{1H}, 1H, 13C, and 13C NMR spectra of compounds (3−15); and HPLC chromatograms of compounds 5, 12, 13, and 14. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Present Address †

Department of Chemistry, Faculty of Science, University of Tanta, 31527-Tanta, Egypt. Notes

The authors declare no competing financial interest. 5948

dx.doi.org/10.1021/om300521j | Organometallics 2012, 31, 5940−5949

Organometallics

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dx.doi.org/10.1021/om300521j | Organometallics 2012, 31, 5940−5949