Chutes and ladders in hepatitis C nucleoside drug development

Abstract

Chutes and Ladders is an exciting up-and-down-again game in which players race to be the first to the top of the board. Along the way, they will find ladders to help them advance, and chutes that will cause them to move backwards. The development of nucleoside analogs for clinical treatment of hepatitis C presents a similar scenario in which taking shortcuts may help quickly advance a program, but there is always a tremendous risk of being sent backwards as one competes for the finish line. In recent years the treatment options for chronic hepatitis C virus (HCV) infection have expand due to the development of a replicon based in vitro evaluation system, allowing for the identification of multiple drugable viral targets along with a concerted and substantial drug discovery effort. Three major drug targets have reached clinical study for chronic HCV infection: the NS3/4A serine protease, the large phosphoprotein NS5A, and the NS5B RNA-dependent RNA polymerase. Recently, two oral HCV protease inhibitors were approved by the FDA and were the first direct acting anti-HCV agents to result from the substantial research in this area. There are currently many new chemical entities from several different target classes that are being evaluated worldwide in clinical trials for their effectiveness at achieving a sustained virologic response (SVR) (Pham et al., 2004; Radkowski et al., 2005). Clearly the goal is to develop therapies leading to a cure that are safe, widely accessible and available, and effective against all HCV genotypes (GT), and all stages of the disease. Nucleoside analogs that target the HCV NS5B polymerase that have reached human clinical trials is the focus of this review as they have demonstrated significant advantages in the clinic with broader activity against the various HCV GT and a higher barrier to the development of resistant viruses when compared to all other classes of HCV inhibitors.

Keywords: Antiviral; Clinical trials; HCV; Nucleoside analog; Prodrug.

Fig. 1

Structures for ribavirin (RBV), boceprevir (Victrelis) and telaprevir (Incivek).

Fig. 2

Nucleosides transform via three successive cellular kinases to the biologically active nucleoside analog triphosphates. Natural ribo- and 2′-deoxyribonucleoside bases.

Fig. 3

Synthesis of acylsalicylic acid from salicylic acid.

Fig. 4

Chemical structures of NM107 and NM283.

Fig. 5

4′-Azidocytidine analogs used in the human clinical treatment of HCV.

Fig. 6

Metabolism and metabolites of RG7348: (a) esterases; (b) CYP3A4; (c) kinases.

Fig. 7

Metabolism of PSI-6130 leads to both the active triphosphate and the inactive nucleoside PSI-6206.

Fig. 8

Structures of setrobuvir (ANA598, non-nucleoside polymerase inhibitor) and danoprevir (RG7227, protease inhibitor).

Fig. 9

Chemical structure of HCV protease inhibitor IDX320.

Fig. 10

Structure of 2′-F-2′-C-methyluridine, PSI-6206, and its triphosphate PSI-6206-TP.

Fig. 11

Structures, anti-HCV replication activity and cytotoxicity of phosphoramidate prodrug PSI-7851 and its diastereoisomers PSI-7976 and PSI-7977.

Fig. 12

Structures of daclatasvir, ledipasvir and simeprevir.

Fig. 13

Structure of prodrug TMC649128 and related nucleosides.

Fig. 14

Structure of GSK2336805.

Fig. 15

Structure of BMS-986094.

Scheme 1

Synthesis of valopicitabine (NM283). Reagents and conditions: (a) i. CaO, water; ii. CO₂, oxalic acid, 18%; (b) BzCl, TEA, DMAP, DME, 83%; (c) Red-Al/EtOH, toluene, 99%; (d) BzCl, TEA, DMAP, DME, 52%; (e) bis(trimethylsilyl)acetamide, cytosine, SnCl₄, CAN; (f) MeONa, MeOH, r.t., 90% over two steps; (g) N-Boc-L-valine, CDI, DMAP, TEA, DMF/THF, 91%; (h) HCl, EtOH, 97%.

Scheme 2

Synthesis of R1479 and its prodrug balapiravir (R1626). Reagents and conditions: (a) I₂, PPh₃, imidazole, THF, 83%; (b) NaOMe, MeOH, 75%; (c) BnN(Et)₃Cl, NaN₃, 4-NMM, I₂, CH₃CN; (d) BzCl, Pyr., 74% over two-steps; (e) K₂HPO₄, m-CPBA, CH₂Cl₂, 71%; (f) 1,2,4-triazole, POCl₃, Et₃N, MeCN, 88%; (g) NH₄OH, MeOH, H₂SO₄-isopropanol, 95%; (h) isobutyric anhydride, KOH, DMAP, THF, 80%.

Scheme 3

Synthesis of PSI-6130. Reagents and conditions: (a) Ph₃PC(Me)CO₂Et, DCM, −40°C; (b) KMnO₄, acetone, 0 °C, 53% over two steps; (c) i. SOCl₂, TEA, DCM, 0 °C; ii. aq. NaOCl, TEMPO, NaHCO₃, CH₃CN, 0 °C; (d) i. TEAF, dioxane, 100 °C; ii. (MeO)₂C(Me)₂, conc. HCl, dioxane, r.t.; (e) EtOH, conc. HCl, r.t.; (f) BzCl, pyridine, r.t., 47% from (14); (g) i. Li(O-tBu)₃AlH, THF, −20 °C; ii. Ac₂O, DMAP, −20 °C; (h) i. silylated N⁴-benzoylcytosine, SnCl₄, PhCl, 65 °C; ii. NH₃, MeOH, r.t.. 29% from (18).

Scheme 4

Synthesis of RG7128. Reagents and conditions: (a) DMAP, THF/H₂O, TEA, isobutyryl chloride, pH 6.4, 0 °C.

Scheme 5

Synthesis of IDX184. Reagents and conditions: (a) TrCl, Et₃N, DMAP, CH₂Cl_2, reflux, overnight; (b) NaOH, dioxane, reflux, 16 h, 92% over two steps; (c) i. CDI, toluene/DMF (2/1, v/v), r.t., 30 min; ii. 2-mercaptoethanol, toluene/DMF (2/1, v/v), −10 °C, 3 h; (d) i. H₃PO₃, PivCl, pyridine, 0 °C to r.t., 3 h; ii. TEAB, 90% over two steps; (e) (23), PivCl, pyridine, −15 °C, 2 h, 32%; (f) Benzylamine, CCl₄, r.t., 1 h, quantitative yield; (g) TFA, CH₂Cl₂, r.t., 30 min, 39%.

Scheme 6

Synthesis of PSI-7976 and PSI-7977 (sofosbuvir, GS-7977) via column purification. Reaction conditions: (a) 70% aqueous acetic acid, 100 °C; (b) 25% methanolic ammonia, 0–15 °C, 78% overall yield; (c) (28a) (R_p:S_p = 1:1) (6.5 equiv), NMI (8 equiv), CH₂Cl₂, r.t., overnight, 6%, 1:1 mix of PSI-7976 and PSI-7977; (d) (28b) (R_p:S_p ≈ 1:1) (2 equiv), t-BuMgCl (2.1 equiv), THF, r.t., 48 h, 47%, 3:1 mix PSI-7977:PSI-7976.

Scheme 7

Synthesis of single isomer PSI-7977 (sofosbuvir, GS-7977). Reaction conditions: (a) THF, t-BuMgCl (2.1 equiv) added over 30 min at r.t., stirred 30 min then (28c), r.t., 60 h, 40 %; (b) THF, t-BuMgCl (2.1 equiv) added over 30 min at −5 °C, stirred 30 min, warmed up to 20 °C and stirred for 30 min and then cooled to 5 °C then (28d) (1.2 equiv) was added over 30 min, stirred for 18 h at 5 °C, 68%.

Scheme 8

Synthesis of PSI-938 intermediate lactone 29. Reagents and conditions: (a) p-Cl-BzCl, pyr., r.t., 70%.

Scheme 9

Synthesis of PSI-938. Reagents and conditions: (a) LiAl(t-BuO)₃H, THF, −20 °C, 61%; (b) CBr₄, PPh₃, CH₂Cl₂, −20 °C, 1 h, 79%; (c) 2-amino-6-chloropurine, t-BuOK, t-BuOH, CH₃CN, 50 °C, 16 h, 65%; (d) NaOEt, EtOH, reflux, 1 h, 92%; (e) isopropyldichlorophosphate, NMI, Et₃N, CH₂Cl₂, 52%.

Scheme 10

Synthesis of 4′-azido-2′-deoxy-2′-methyl cytidine (TMC649128). Reagents and conditions: (a) bis(trimethylsilyl)uracil, SnCl₄, CH₃CN, r.t., 20 h, 96%; (b) NH₃, CH₃OH, 0–4 °C, 2 days, >99%; (c) TIPDSCl₂, imidazole, DMF, r.t., 2 h, 65%; (d) ClCOCO₂Me, DMAP, CH₃CN, r.t., 1 h; (e) n-Bu₃SnH, AIBN, toluene, reflux, 2 h, 97% (β/α 93:7, de 86%) over two steps; (f) TBAF/THF, r.t., 1 h, >99% (β/α 93:7); (g) I₂, PPh₃, imidazole, THF, 76%; (h) NaOMe, MeOH, 75%; (i) i. I₂, NMO, Bn(Et)₃NN₃; ii. BzCl, NMP, DMAP, THF, 81% over 2 steps; (j) m-CPBA, (NH₄)₂SO₄, DCM, 68%; (k) i. pyridine, 2-chlorophenyldichlorophosphate, 1-H-tetrazole; ii. dioxane, NH₃; iii. NH₃, MeOH, 61% over 3 steps; (l) i-PrCOCl, TEA, DCM, r.t., >99%.

Scheme 11

Synthesis of GS-6620. Reagents and conditions: (a) (44), 1,1,4,4-tetramethyl-1,4-dichlorodisilylethylene (1.0 equiv) and n-BuLi (3.3 equiv), THF, −78 °C followed by addition of (43), 1 h, 65–80%; (b) TMSCN (6 equiv), TMSOTf (4 equiv), CH₂Cl₂, −15 to 0 °C, 2 h, 85–95%; (c) BCl₃ (3 equiv), CH₂Cl₂, 0 °C, 1 h, 90%; (d) N-methylimidazole, trimethylphosphate and THF, 0 °C to r.t., 2 h, overall yield 59%; (e) i. N,N-dimethylformamide dimethyl acetal, CH₃CN, r.t., 1 h; ii. isobutyric acid, DCC, CH₃CN, r.t., 48 h; iii. water-TFA, 0 °C to r.t., 64 h, overall yield 73%.

Scheme 12

Synthesis of BMS-986094. Reagents and conditions: (a) 2-amino-6-chloropurine, DBU, MeCN, TMSOTf, 65 °C, 4 h, 79%; (b) NaOMe, MeOH, r.t., 24 h, 76%; (c) POCl₃, Et₃N, Et₂O, −78 °C, 1 h then r.t., overnight, 95%; (d) tBuCH₂-L-valine ester, Et₃N, DCM, −78 °C, 1 h then r.t., 2 h; (e) NMI, THF, r.t., overnight, 18%.