Synthesis of (+)-ribostamycin by catalytic, enantioselective hydroamination of benzene

Aminoglycosides (AGs) represent a large group of pseudoglycoside natural products, in which several different sugar moieties are harnessed to an aminocyclitol core. AGs constitute a major class of antibiotics that target the prokaryotic ribosome of many problematic pathogens. Hundreds of AGs have been isolated to date, with 1,3-diaminocyclohexanetriol, known as 2-deoxystreptamine (2-DOS), being the most abundant aglycon core. However, owning to their diverse and complex architecture, all AG-based drugs are either natural substances or analogues prepared by late-stage modifications. Synthetic approaches to AGs are rare and lengthy; most studies involve semi-synthetic reunion of modified fragments. Here we report a bottom-up chemical synthesis of the 2-DOS-based AG antibiotic ribostamycin, which proceeds in ten linear operations from benzene. A key enabling transformation involves a Cu-catalyzed, enantioselective, dearomative hydroamination, which set the stage for the rapid and selective introduction of the remaining 2-DOS heteroatom functionality. This work demonstrates how the combination of a tailored, dearomative logic and strategic use of subsequent olefin functionalizations can provide practical and concise access to the AG class of compounds.


General Procedures
All chemicals were purchased from commercial suppliers and used as received, unless otherwise noted.
MTAD was synthesized according to literature protocol. 1 C18-derivatized SiO2 was prepared according to the literature procedure. 2 Diethyl ether (ACS grade), dichloromethane (ACS grade), tetrahydrofuran (HPLC grade), and toluene (ACS grade) were dried for reactions using the MB-SPS solvent purification system containing activated alumina manufactured by MBRAUN. Reaction temperatures correspond to the external temperature of the reaction vessel and reactions were run under an atmosphere of dry nitrogen and in predried reaction vessels unless otherwise noted.
Analytical thin-layer chromatography (TLC) was performed on Merck silica gel 60 F254 aluminum sheets.
Visualization was accomplished with UV light and/or potassium permanganate (KMnO4). Retention factor (Rf) values reported were measured using a 5 × 2 cm TLC plate in a developing chamber containing the solvent system described.
Electrospray ionization (ES+) spectra were performed using a time-of-flight (TOF) mass analyzer. Data are reported in the form of m/z. Infrared (IR) spectra were measured neat on a Perkin-Elmer Spectrum Two FT-IR ATR spectrometer. Peaks are reported in cm -1 with indicated relative intensities: s (strong, 0 -33% T); m (medium, 34 -66% T), w (weak, 67 -100% T), and br (broad). Melting points were measured on a Buchi B-540 melting point apparatus and are uncorrected. Optical rotations were recorded on a Jasco P-2000 polarimeter at 589 nm and are S4 reported in units of 10 -1 (deg cm 2 g -1 ). Single crystal diffraction experiments were carried out in George L. Clark X-Ray laboratory of the School of Chemical Sciences, UIUC. Single crystals were mounted on Cryo-loops using Paratone-N or Krytox oils. Data collection was performed on either Bruker Venture D8 / Photon II or Bruker APEX-II CCD diffractometer. Data was integrated using APEX III program suite and absorption correction was then applied using SADABS. Space group was determined with XPREP software and then solved with either Direct Methods (XS) or Intrinsic Phasing (XT), then refined using full-matrix least squares against F2 as implemented in XL.  To an oven-dried test tube was added sublimed MTAD (18, 114.0 mg, 1.0 mmol, 1.0 eq.). The tube was evacuated and backfilled with nitrogen three times, then CH2Cl2 (8 mL) and benzene (10.0 mmol, 0.90 mL, 10.0 eq.)

Abbreviations
were sequqentially added. The tube was cooled to -78 °C and irradiated with white LEDs at that temperature until the pink color disappeared overnight. In a separate vial, copper source (0.05 mmol, 5.0 mol%), ligand (0.055 mmol, 5.5 mol%), and KOtBu (123.4 mg, 1.1 mmol, 1.1 eq.) were dissolved in toluene (2 mL) and stirred 30 minutes. The catalyst solution was then added to the reaction mixture dropwise and followed by the dropwise addition of diethoxymethylsilane (DEMS, 400 µL, 2.5 mmol, 2.5 eq.). Then, the LEDs and the chiller were turned off, and the reaction was allowed to -5 °C (approximately 5 hours until reaction Dewar reaches this temperature). The reaction was then placed in a water bath and allowed to reach room temperature before the addition of (CH3)2SO4 (946 µL, 10.0 mmol, 10 eq.) and K2CO3 (691.0 mg, 5.0 mmol, 5.0 eq.). Thereafter, the reaction was stirred at 35 °C for 1.5 hours. Then the mixture was quenched with NH4OH (5%, aq. sol., 2 mL) and water (2 mL), and the aqueous phase was extracted with CH2Cl2 (3 × 5 mL). The combined organic phase was washed with sodium chloride (sat. aq. sol., 20 mL), dried over anhydrous MgSO4 and concentrated under reduced pressure. NMR yields of the corresponding product 13-Me were taken using nitromethane as a standard. The crude residue was purified by flash column chromatography (Al2O3, hexane:EtOAc = 4:1), and the e.r. was determined with HPLC analysis using Diacel Chiracel OD-3 column, 10% i-PrOH in n-hexane, 0.8 mL/min.

Synthesis of Ribostamycin (2)
Diene 13. MTAD (18, 2.83 g, 25 mmol, 1.0 eq.) was dissolved in anhydrous dichloromethane (167 mL) and degassed benzene (14, 11.2 mL, 125 mmol, 5.0 eq.) was added. It should be noted that multiple attempts of enantioenrichment by crystallization of diene 13 or other downstream intermediates (i.e., 28-31) were unsuccessful. With regard to material taken through the remainder of the synthesis, the diene 13, produced with 96:4 e.r., was taken forward and used for characterization. Moreover, the material from the larger scale (100 mmol, produced with 79:21 e.r.) was also carried through the identical sequence and optical enrichment was achieved after the first glycosylation with the enantiopure sugar. The corresponding pseudodisaccharide 33 could be readily separated from its minor diastereoisomer using flash chromatography conditions described in the protocol. S10 Allylic Epoxide 28. To a cool (0 °C) solution of diene 13 (10.0 g, 52.0 mmol, 1.0 eq.) and 4-dimethylaminopyridine (0.95 g, 7.8 mmol, 0.15 eq.) in CH2Cl2 (610 mL) was added Boc2O (18 mL, 78 mmol, 1.5 eq.). The reaction was allowed to warm to room temperature and stirred for 30 minutes. Thereafter, the mixture was recooled to 0 °C and treated with solid sodium bicarbonate (6.1 g, 72 mmol, 1.4 eq.) followed by portion-wise addition of mCPBA (75 wt%, 15.5 g, 67.3 mmol, 1.3 eq.). The contents were allowed to warm to room temperature and stirred 12 hours. The reaction was quenched by addition of sodium bicarbonate (sat. aq. sol., 50 mL) and sodium thiosulfate (10% aq. sol., 50 mL). The organic layer was collected and the resulting aqueous layer was extracted with CH2Cl2 (3 × 250 mL) and

Compounds and Data Supporting Mechanistic Studies
For the synthesis of deutero substrate 29-d6, the same procedure as 13 to 29 was utilized. See

Computational Methods
All density functional theory (DFT) calculations were carried out using the Gaussian 16 program 9 on Pitt CRC and XSEDE supercomputers. Geometries of intermediates and transition states were optimized in the gas phase using B3LYP functional 10 with a mixed basis set of SDD 11 (i.e. the Stuttgart/Dresden effective core potentials and the associated valence electron basis set) for Fe and Cu and 6-31G(d) for other atoms.
Vibrational frequency calculations were performed to confirm if each structure is a local minimum or a transition state. Gibbs free energies were calculated at 223.15K using quasi-harmonic approximation with Grimme's approach 12 using the GoodVibes package. 13 Single-point energy calculations were carried out using the M06 functional 14 with the SDD basis set for Fe and Cu and 6-311+G(d,p) for other atoms.
Solvation energy corrections were calculated in single point energy calculations using the SMD solvation model 15 in dichloromethane. CYLview 16 was used to generate all 3D structures.

Dimeric Copper(I) Hydride Supported by (S,Sp)-TaniaPhos Ligand
The DFT calculations indicated that the dimeric (S,Sp)-TaniaPhos-supported CuH catalyst 23 is 5.9 kcal/mol more stable than the monomeric CuH species 24. The Cylview 3D structure of the most stable ground state of 23 is provided in Supplementary Figure 3.
Supplementary Figure 3. Reaction Gibbs free energy of dissociation of dimeric copper(I) hydride 23 to form monomeric copper(I) hydride 24.

Stereoisomers of the Hydrocupration Transition State and the Alkyl Copper(I) Intermediate
Eight possible stereoisomers of the hydrocupration transition state (Supplementary Figure 4)

Allylic Substitution Pathway
The competing allylic substitution pathway was studied using DFT calculations. The Cu(III) intermediate

Calculated Total Energies and Thermochemical Corrections
Cartesian coordinates of all optimized geometries are provided in the Supplementary Information in a separate file in the .xyz format.  Figure S11. HPLC trace of racemic diene 13-Me-d6.