A High‐Yielding Synthesis of EIDD‐2801 from Uridine**

A simple reordering of the reaction sequence allowed the improved synthesis of EIDD‐2801, an antiviral drug with promising activity against the SARS‐CoV‐2 virus, starting from uridine. Compared to the original route, the yield was enhanced from 17 % to 61 %, and fewer isolation/purification steps were needed. In addition, a continuous flow procedure for the final acetonide deprotection was developed, which proved to be favorable toward selectivity and reproducibility.


General Methods
All solvents and chemicals were obtained from standard commercial vendors (TCI, Sigma-Aldrich/Merck or VWR) and were used without any further purification, unless otherwise noted. 1 H NMR spectra were recorded on a Bruker 300 MHz instrument. 13 C NMR spectra were recorded on the same instrument at 75 MHz. Che i al shifts δ are e pressed i pp do field fro TMS as i ter al sta dard. The letters s, d, t, q, sept, dd and m are used to indicate singlet, doublet, triplet, quadruplet, septet, doublet of doublets and multiplet. Analytical HPLC analysis was carried out on a Shimadzu instrument using a C18 reversedphase RP a al ti al olu × . , parti le size μ at °C usi g o ile phases A (H 2 O/MeCN (90:10 v/v) + 0.1% TFA) and B (MeCN + 0.1% TFA) at a flow rate of 1.5 mL/min. The following gradient was applied: start at 3 % solvent B, increase to 5 % solvent B until 3 min, increase to 30 % solvent B until 7 min and finally increase to 100 % solvent B until 10 min. LC-MS analysis was carried out on a Shimadzu instrument using a C18 reversed-phase (RP) analytical column (150 mm × 4.6 mm, particle size μ using mobile phases A (H 2   [a] Equivalents given in parenthesis.

Optimization Studies: One-pot Acetonide Protection and Esterification
[b] Area% determined by HPLC at 260 nm.
The hydrolysis product 7 was identified by LC-MS analysis (see Figure S1). The generation of 7 was additionally verified, since upon reaction with isobutyric anhydride, the corresponding ester 8 was obtained. The identity of 8 was confirmed by NMR analysis (see Figures S2 and S3).  Scheme S1. One-pot acetonide protection and esterification.
In the protection step, the reaction mixture was stirred first for 30 min at rt, then the stepwise azeotropic distillation was performed. We chose this reaction regime out of precautionary reasons, because the bp of DMP (85 °C) is close to the bp of the azeotrope MeOH/MeCN (63 °C). Therefore, in order not to potentially distill DMP during the azeotropic distillation, initially stirring at rt was preferred. A 73% conversion to 2 was achieved after 30 min, and full conversion was accomplished during the azeotropic distillation ( Figure S4). Reaction monitoring of the esterification step could be done visually, as with full conversion of 23 the suspension became a clear solution. The reaction time was dependent on the scale: On a 600 mg scale the reaction went to completion within 40 min ( Figure S4), while on a 5 g scale the reaction time needed to be prolonged to 1 h (Scheme S1). In general, the reaction sequence proved to be remarkably clean, as no side products were detected. After extractive work-up, 3 was isolated in quantitative yield and a purit of ≥ %.

Optimization Studies: Acetonide Deprotection in Batch
General procedure for the acid screening: A 1 mL HPLC vial was charged with 4 (50 mg, 0.135 mmol) and 1 mL of acid. The vial was crimped and stirred at rt or heated in an aluminum heating block at 60 °C. [a] Acids were 1 M in the respective solvent.
[b] Area% determined by HPLC at 260 nm. Except for entry 9, further unidentified impurities were detected.
General procedure for the one-pot hydroxyamination and acetonide deprotection: A microwave vial was charged with 3 (50 mg, 0.123 mmol). i-PrOH (617 µL) and hydroxylamine (50w% in water, 11.1 µL, 1.5 equiv.) were added and the reaction mixture was stirred at room temperature for 20 minutes to ensure full conversion to 4. Next, conc H 2 SO 4 was added dropwise under stirring, the microwave vial crimped and subjected to microwave heating.
As can be seen in Table 3 (entries 1 and 2), the preformation of hydroxylamine 4 is required in order to drive the reaction toward EIDD-2801 while concomitantly reduce the formation of side products 5 and 6. Nevertheless, we experienced reproducibility issues in this optimization study, most likely because of the exotherm upon addition of H 2 SO 4 , which proved to be difficult to control in batch. Unfortunately, the reproducibility could not be improved by diluting the conc H 2 SO 4 with i-PrOH.

Optimization Studies: Acetonide Deprotection in Continuous Flow
The flow set-up consisted of a . L rea tio oil PFA, / " OD, . ID i ersed i a heated oil bath, a Syrris Asia syringe pump module (P1 and P2) equipped with two injection valves, two sample loops SL a d SL , L a d . L, respe ti el ; PFA, / " OD, . ID, each) and a Zaiput back pressure regulator which kept a constant pressure of 5 bar.
General procedure: For each experiment, a 2 mL solution containing 0.18 M of compound 3 (146 mg, 0.36 mmol) and 1.5 equiv. of NH 2 OH . μL, t% in H 2 O) in the corresponding solvent was stirred for 15 min at room temperature to ensure full conversion to 4. This reaction mixture was then directly transferred to SL2. SL1 was filled with neat HCOOH or with a solution of H 2 SO 4 , CF 3 COOH or TfOH prepared in the same solvent as the substrate solution. The liquid feeds were combined in a Y-mixer, and the resulting stream was directed through the heated reaction coil. In each run, approx. 0.5 mL sample of product mixture was collected which was next analyzed directly by HPLC at 260 nm.  [a] Neat HCOOH ≥ % as employed which corresponds to approx. 26 M.
[b] Area% determined by HPLC at 260 nm.
[c] Further unidentified impurities.  [c] CF 3 COOH was employed as 6.5 M solution in MeOH.

Acetonide Deprotection: Analysis of Side Products
In the acetonide deprotection reactions, ester hydrolysis and the exchange of the hydroxyl amine moiety occurred as most prominent side reactions yielding compounds 5 and 6 as side products as detailed above. The identity of these substances was corroborated by means of LC-MS.

Synthesis of Triazolated Uridine 1
An oven dried 500 mL 2-necked round bottom flask was flushed with argon and charged with uridine (5.0 g, 20.5 mmol) and 164 mL of anhydrous MeCN (AcroSeal TM , max. 0.001% H 2 O). N-Methylpyrrolidine (31.9 mL, 307 mmol, 15 equiv) and TMSCl (13 mL, 102 mmol, 5 equiv) were added, and the reaction mixture was allowed to stir for 1 h at room temperature. The solution was cooled in an ice bath to 0 °C. POCl 3 (3.74 mL, 41.0 mmol, 2 equiv) was added. After stirring for 10 min, 1,2,4-triazole (14.1 g, 205 mmol, 10 equiv) was added, and stirring was continued at 0 °C for 1 h and at room temperature for another 2 h. The yellow solution was then poured onto 700 mL triethylammonium phosphate buffer (0.5 M, pH 7) and extracted with DCM (3 × 100 mL). The combined organic phases were dried over Na 2 SO 4 and the solvent evaporated under reduced pressure. A mixture of MeOH:AcOH (4:1 v/v) was added to the residue and the reaction mixture was stirred at room temperature overnight. The precipitated product was collected by filtration, washed with diethyl ether and dried under reduced pressure. 1 was obtained as a white solid in 88% yield (5.34 g) and ≥99% purity.

Telescoped Synthesis of EIDD-2801 in Continuous Flow
The flow set-up used was identical with the one described in Section 5 ( Figure S6, see also Figure 1 in the main text). Sample loops of 5 mL (SL1) and 10 mL (SL2) were used (PFA, / " OD, . ID, ea h . Scale-out was performed under optimum flow conditions (see Table S5, entry 9) as follows: A 10.5 mL solution containing 0.18 M of compound 3 (766 mg, 1.89 mmol) and 1.5 equiv. of NH 2 OH (170 μL, t% in H 2 O) in MeOH was prepared and was stirred for 15 min at room temperature. This reaction mixture was then directly transferred to SL2. SL1was filled with 1 M H 2 SO 4 solution in MeOH. P1 was set to μL/ i a d P to μL/min. The liquid feeds were combined in a Y-mixer, and the resulting stream was directed through a 3.5-mL reaction coil at 100 °C. The product mixture leaving the coil was collected for 16 min under steady state conditions. The collected mixture was neutralized (pH 7) with a 4 M aq. NaOH solution and was next purified by column chromatography using a 6-16% gradient of MeOH in CH 2 Cl 2 as eluent. EIDD-2801 was isolated in 69% yield (307 mg) and ≥99% purity as a white solid.