SIMEPREVIR

Simeprevir

Inhibits HCV NS3/4A protease.

MEDIVIR ... originator

launched 2013

923604-59-5  CAS

C38H47N5O7S MF

749.93908  MW

IUPAC standard name
(1R, 4R, 6S, 15R, 17R)-N-(cyclopropanesulfonyl) -17 – ({7-methoxy-8-methyl-2-[4 - (propan-2-yl) -1,3-thiazol-2 -yl] quinolin-4-yl} oxy)-13-methyl-2 ,14-dioxo-3 ,13-diazatricyclo [13.3.0.0 4 , 6 ] octadec-7-ene-4-carboxamide
IUPAC traditional name
(1R, 4R, 6S, 15R, 17R)-N-(cyclopropanesulfonyl) -17 – {[2 - (4-isopropyl-1 ,3-thiazol-2-yl)-7-methoxy-8-methylquinolin-4- yl] oxy}-13-methyl-2 ,14-dioxo-3 ,13-diazatricyclo [13.3.0.0 4 , 6 ] octadec-7-ene-4-carboxamide

• Olysio
• Simeprevir
• TMC 435
• TMC 435350
• TMC-435
• TMC435
• TMC435350
• UNII-9WS5RD66HZ

November 22, 2013 -- The U.S. Food and Drug Administration  approved Olysio (simeprevir), a new therapy to treat chronic hepatitis C virus infection.

OLYSIO™ is the first once-daily protease inhibitor approved for the treatment of chronic hepatitis C in a combination antiviral regimen for adults with compensated liver disease

Hepatitis C is a viral disease that causes inflammation of the liver that can lead to diminished liver function or liver failure. Most people infected with the hepatitis C virus have no symptoms of the disease until liver damage becomes apparent, which may take several years. Most of these people then go on to develop chronic hepatitis C. Some will also develop scarring and poor liver function (cirrhosis) over many years, which can lead to complications such as bleeding, jaundice (yellowish eyes or skin), fluid accumulation in the abdomen, infections or liver cancer. According to the Centers for Disease Control and Prevention, about 3.2 million Americans are infected with the hepatitis C virus

Hepatitis C virus (HCV) infections affect approximately 3 percent of the worldwide population and often lead to cirrhosis and hepatocellular carcinoma. The standard therapy of pegylated- interferon and ribavirin induces serious side effects and provides viral eradication in less than 50% of patients. Combination therapy of HCV including ribavirin and interferonare currently is the approved therapy for HCV. Unfortunately, such combination therapy also produces side effects and is often poorly tolerated, resulting in major clinical challenges in a significant proportion of patients. Numerous direct acting agents (DAAs) have been or are being developed for treatment of HCV, such as telaprevir and boceprevir (both received MA approved in 2011 for use with interferon and ribavirin based therapy), however direct acting agents are linked to increased toxicity of treatment, the emergence of resistance, and to date do not provide a standard of care which is interferon free. The combination of direct acting agents can also result in drug-drug interactions. To date, no HCV therapy has been approved which is interferon free. There is therefore a need for new combination therapies which have reduced side effects, and interferon free, have a reduced emergence of resistance, reduced treatment periods and/or and enhanced cure rates.

Simeprevir (formerly TMC435) is an experimental drug candidate for the treatment of hepatitis C. It is being developed byMedivir and Johnson & Johnson‘s pharmaceutical division Janssen Pharmaceutica and is currently in Phase III clinical trials.^[1]

Simeprevir is a hepatitis C virus protease inhibitor.^[2]

Simeprevir is being tested in combination regimens with pegylated interferon alfa-2a and ribavirin,^[3] and in interferon-free regimens with other direct-acting antiviral agents including daclatasvir^[4] and sofosbuvir ^[5]

Simeprevir has been launched in 2013 in Japan by Janssen Pharmaceutical (JP) for use in combination with pegylated interferon (Peg-IFN) and ribavirin for the treatment of genotype 1 chronic hepatitis C virus (HCV) patients who are treatment naïve, prior non responders or relapsed following treatment with Peg-IFN with or without ribavirin. In 2013, the product has also been approved in the U.S. by Medivir and Janssen R&D Ireland for the oral treatment of chronic hepatitis C genotype 1 infection, in combination with peginterferon alfa and ribavirin in adults with compensated liver disease, including cirrhosis, who are treatment-naïve or who have failed previous interferon therapy (pegylated or non-pegylated) with ribavirin.

The drug candidate was originally developed at Medivir, which was acquired by Janssen R&D Ireland in 2012. In November 2004, Medivir entered into a license and research collaboration agreement with Tibotec, a Johnson & Johnson subsidiary, for the discovery and development of orally active protease inhibitors of the NS3/4A protease of HCV. In 2011, a codevelopment agreement between Pharmasset (now Gilead Sciences) and Tibotec was signed for the treatment of chronic hepatitis C (HCV) in combination with PSI-7977. Also in 2011, fast track designation was received in the U.S. for the treatment of chronic hepatitis C (CHC) genotype-1 infection.

In 2011, Tibotec Therapeutics, Division of Centocor Ortho Biotech Products, L.P. announced that it had changed its name to Janssen Therapeutics, Division of Janssen Products, LP.

“Hepatitis C is a complex disease and Janssen is committed to working with the HCV community, caregivers, and health care systems to address this global epidemic,” said Gaston Picchio, Hepatitis Disease Area Leader, Janssen Research & Development. “We are pleased that the FDA has granted simeprevir Priority Review, as it is a significant step forward in making this therapy available to physicians and their hepatitis C patients.”

Hepatitis C virus (HCV) is the leading cause of chronic liver disease worldwide.

Following initial acute infection, a majority of infected individuals develop chronic hepatitis because HCV replicates preferentially in hepatocytes but is not directly cytopathic. Chronic hepatitis can progress to liver fibrosis leading to cirrhosis, end- stage liver disease, and HCC (hepatocellular carcinoma), making it the leading cause of liver transplantations. This and the number of patients involved, has made HCV the focus of considerable medical research. Replication of the genome of HCV is mediated by a number of enzymes, amongst which is HCV NS3 serine protease and its associated cofactor, NS4A. NS3 serine protease is considered to be essential for viral replication and has become an attractive target for drug discovery.

Current anti-HCV therapy is based on (pegylated) interferon-alpha (IFN-α) in combination with ribavirin. Not only does this therapy result in a limited efficacy in that only part of the patients are treated successfully, but it also faces significant side effects and is poorly tolerated in many patients. Hence there is a need for further HCV inhibitors that overcome the disadvantages of current HCV therapy such as side effects, limited efficacy, poor tolerance, the emergence of resistance, as well as compliance failures.

Various agents have been described that inhibit HCV NS3 serine protease. WO05/073195 discloses linear and macrocyclic NS3 serine protease inhibitors with a central substituted proline moiety and WO 05/073216 with a central cyclopentyl moiety. Amongst these, the macrocyclic derivatives are attractive by overcoming one or more of the disadvantages of current anti-HCV therapy

(I)  simeprevir

The compound of formula (I) is an inhibitor of the Hepatitis C virus (HCV) serine protease and is described in WO 2007/014926, published on 8 February 2007. This compound overcomes several of the disadvantages of current anti-HCV therapy and in particular shows pronounced activity against HCV, has an attractive pharmacokinetic profile, and is well-tolerated. Following the synthesis procedure described in Example 5 of WO 2007/014926, an amorphous solid form is obtained.

It now has been found that the compound of formula (I) can be converted into crystalline forms, which can advantageously be used as active ingredients in anti-HCV therapy. To that purpose, these crystalline forms are converted into pharmaceutical formulations.

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SIMEPREVIR

..............................

simeprevir

OLYSIO (simeprevir) is an inhibitor of the HCV NS3/4A protease.

The chemical name for simeprevir is (2R,3aR,10Z,11aS,12aR,14aR)-N-(cyclopropylsulfonyl)-2[[2-(4-isopropyl-1,3-thiazol-2-yl)-7-methoxy-8-methyl-4-quinolinyl]oxy]-5-methyl-4,14-dioxo2,3,3a,4,5,6,7,8,9,11a,12,13,14,14atetradecahydrocyclopenta[c]cyclopropa[g][1,6]diazacyclotetradecine-12a(1H)-carboxamide. Its molecular formula is C₃₈H₄₇N₅O₇S₂ and its molecular weight is 749.94. Simeprevir has the following structural formula:

Simeprevir drug substance is a white to almost white powder. Simeprevir is practically insoluble in water over a wide pH range. It is practically insoluble in propylene glycol, very slightly soluble in ethanol, and slightly soluble inacetone. It is soluble in dichloromethane and freely soluble in some organic solvents (e.g., tetrahydrofuran and N,N-dimethylformamide).

OLYSIO (simeprevir) for oral administration is available as 150 mg strength hard gelatin capsules. Each capsule contains 154.4 mg of simeprevir sodium salt, which is equivalent to 150 mg of simeprevir. OLYSIO (simeprevir) capsules contain the following inactive ingredients: colloidal anhydrous silica, croscarmellose sodium, lactose monohydrate, magnesium stearate and sodium lauryl sulphate. The white capsule contains gelatin and titanium dioxide (E171) and is printed with ink containing iron oxide black (E172) and shellac (E904).

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Synthesis

WO2008092954A2

Example 1 : preparation of 17-[2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methyl- quinolin-4-yloxy]- 13-methyl-2, 14-dioxo-3, 13-diazatricyclo[ 13.3.0.0⁴'⁶]octadec-7-ene- 4-carboxylic acid (16)

Synthesis of 4-hydroxy-2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methylquinoline (6) Step 1 : synthesis of Λ/-(tert-butyloxycarbonyl)-3-methoxy-2-methylaniline (2)

¹                                                                                               2

Triethylamine (42.4 mL, 302 mmol) was added to a suspension of 3-methoxy-2- methylbenzoic acid (45.6 g, 274 mmol) in dry toluene (800 mL). A clear solution was obtained. Then, dppa (65.4 mL, 302 mmol) in toluene (100 mL) was slowly added. After 1 h at room temperature, the reaction mixture was successively heated at 50⁰C for 0.5 h, at 70⁰C for 0.5 h then at 100⁰C for 1 h. To this solution, t-BuOH (30.5 g, 411 mmol) in toluene (40 mL) was added at 100⁰C and the resulting mixture was refluxed for 7h. The solution was cooled to room temperature then successively washed with water, 0.5 N HCl, 0.5 N NaOH and brine, dried (Na₂SO₄), and evaporated to give 67 g of the target product: m/z = 237 (M)⁺.

_2: synthesis of 3-methoxy-2-methylaniline (3)

TFA (40.7 mL, 548 mmol) was added to a solution of jV-(teτt-butyloxycarbonyl)- 3-methoxy-2-methylaniline, in dichloro methane (500 mL). After 2 h at room temperature, TFA (40.7 mL, 548 mmol) was added and the resulting mixture was stirred at room temperature overnight. Then, volatiles were evaporated. The residue was triturated with toluene (100 mL) and diisopropylether (250 mL), filtered off and washed with diisopropyl ether (100 mL) to give 56.3 g of the title product as a TFA salt: m/z = 138 (M+H)⁺. The TFA salt was transformed to the free aniline by treatment with NaHCO₃.

Step 3: synthesis of (2-amino-4-methoxy-3-methylphenyl)(methyl)ketone (4)

A solution Of BCl₃ (1.0 M, 200 mL, 200 mmol) in CH₂Cl₂ was slowly added under nitrogen to a solution of 3-methoxy-2-methylaniline (26.0 g, 190 mmol) in xylene (400 mL). The temperature was monitored during the addition and was kept below 10⁰C. The reaction mixture was stirred at 5°C for 0.5 h. Then, dry acetonitrile (13 mL, 246 mmol) was added at 5°C. After 0.5 h at 5°C, the solution was transferred into a dropping funnel and slowly added at 5°C to a suspension OfAlCl₃ (26.7 g, 200 mmol) in CH₂Cl₂ (150 mL). After 45 min at 5°C, the reaction mixture was heated at 70⁰C under a nitrogen stream. After evaporation Of CH₂Cl₂, the temperature of the reaction mixture reached 65°C. After 12 h at 65°C, the reaction mixture was cooled at 0⁰C, poured onto ice (300 g), and slowly heated to reflux for 7h. After 2 days at room temperature, 6 N NaOH (50 mL) was added. The pH of the resulting solution was 2-3. The xylene layer was decanted. The organic layer was extracted with CH₂Cl₂. The xylene and CH₂Cl₂ layers were combined, successively washed with water, IN NaOH, and brine, dried (Na₂SO₄) and evaporated. The residue was triturated in diisopropyl ether at O⁰C, filtered off and washed with diisopropylether to give 13.6 g (40 %) of the title product as a yellowish solid: m/z = 180 (M+H)⁺.

Step 4: synthesis of 2'-[[(4-isopropylthiazole-2-yl)(oxo)methyl]amino]-4'-methoxy-3 '- methylacetophenone (5)

A solution of the compound 4 (18.6 g, 104 mmol) in dioxane (50 rnL) was added under nitrogen to a suspension of 4-isopropylthiazole-2-carbonyl chloride in dioxane (250 rnL). After 2 h at room temperature, the reaction mixture was concentrated to dryness. Then, the residue was partitioned between an aqueous solution of NaHCOs and AcOEt, organic layer was washed with brine, dried (Na₂SO₄), and evaporated. The residue was triturated in diisopropyl ether, filtered off and washed with diisopropyl ether to give 30.8 g (90 %) of the title product 5.

Step 5: synthesis of 4-hydroxy-2-(4-isopropylthiazole-2-yl)-7-methoxy-8- methylquinoline (6)

Potassium tert-butoxide (21.8 g, 195 mmol) was added to a suspension of the compound 5 (30.8 g, 92.7 mmol) in tert-butanol. The resulting reaction mixtures was heated at 100⁰C overnight. Then, the reaction mixture was cooled at room temperature and diluted with ether (100 mL). The precipitate was filtered off and washed with Et₂O to give a powder (fraction A). The mother liquor was concentrated in vacuo, triturated in ether, filtered off, and washed with ether to give a powder (fraction 2). Fractions 1 and 2 were mixed and poured into water (250 mL). The pH of the resulting solution was adjusted to 6-7 (control with pH paper) with HCl IN. The precipitate was filtered off, washed with water and dried. Then, the solid was triturated in diisopropyl ether, fϊltered off and dried to give 26 g (88%) of the compound 6 as a brownish solid: m/z = 315 (M+H)⁺.

Synthesis of (hex-5-enyl)(methyl)amine (8)

O CF,

FX N Br' N O NH

H 7

(a) Sodium hydride (1.05 eq) was slowly added at 0⁰C to a solution of JV-methyl- trifluoro-acetamide (25 g) in DMF (140 mL). The mixture was stirred for Ih at room temperature under nitrogen. Then, a solution of bromohexene (32,1 g) in DMF

(25 mL) was added dropwise and the mixture was heated to 70⁰C for 12 hours. The reaction mixture was poured on water (200 mL) and extracted with ether (4 x 50 mL), dried (MgSO₄), filtered and evaporated to give 35 g of the target product 7 as a yellowish oil which was used without further purification in the next step.

(b) A solution of KOH (187.7 g) in water (130 mL) was added dropwise to a solution of 7 (35 g) in methanol (200 mL). The mixture was stirred at room temperature for

12 hours. Then, the reaction mixture was poured on water (100 mL) and extracted with ether (4 x 50 mL), dried (MgSO₄), filtered and the ether was distilled under atmospheric pressure. The resulting oil was purified by distillation under vacuum (13 mm Hg pressure, 50⁰C) to give 7,4 g (34 %) of the title product 8 as a colourless oil: ¹H-NMR (CDCl₃): δ 5.8 (m, IH), 5 (ddd, J = Yl 2 Hz, 3.5 Hz, 1.8 Hz, IH), 4.95 (m, IH), 2.5 (t, J = 7.0 Hz, 2H), 2.43 (s, 3H), 2.08 (q, J= 7.0 Hz, 2H), 1.4 (m, 4H), 1.3 (br s, IH).

Preparation of 17-[2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methylquinolin-4-yloxyl- 13-methyl-2, 14-dioxo-3, 13-diazatricyclo[ 13.3.0.0⁴'⁶loctadec-7-ene-4-carboxylic acid (16)

3-Oxo-2-oxa-bicyclo[2.2.1]heptane-5-carboxylic acid 9 (500 mg, 3.2 mmol) in 4 mL DMF was added at 0⁰C to HATU (1.34 g, 3.52 mmol) and JV-methylhex-5-enylamine (435 mg, 3.84 mmol) in DMF (3 mL), followed by DIPEA. After stirring for 40 min at 0⁰C, the mixture was stirred at room temperature for 5 h. Then, the solvent was evaporated, the residue dissolved in EtOAc (70 rnL) and washed with saturated NaHCOs (IO mL). The aqueous layer was extracted with EtOAc (2 x 25 mL). The organic phases were combined, washed with saturated NaCl (20 mL), dried (Na₂SO₄), and evaporated. Purification by flash chromatography (EtO Ac/petroleum ether, 2:1) afforded 550 mg (68%) of the target product 10 as a colorless oil: m/z = 252 (M+H)⁺.

A solution of LiOH (105 mg in 4 mlof water) was added at 0⁰C to the lactone amide 10. After Ih, the conversion was completed (HPLC). The mixture was acidified to pH 2 - 3 with IN HCl, extracted with AcOEt, dried (MgSO₄), evaporated, co-evaporated with toluene several times, and dried under high vacuum overnight to give 520 mg (88%) of the target product 11: m/z = 270 (M+H)⁺.

The l-(amino)-2-(vinyl)cyclopropanecarboxylic acid ethyl ester hydrochloride 12

(4.92 g, 31.7 mmol) and HATU (12.6 g, 33.2 mmol) were added to 11 (8.14 g,

30.2 mmol). The mixture was cooled in an ice bath under argon, and then DMF (100 mL) and DIPEA (12.5 mL, 11.5 mmol) were successively added. After 30 min at 0⁰C, the solution was stirred at room temperature for an additional 3 h. Then, the reaction mixture was partitioned between EtOAc and water, washed successively with 0.5 N HCl (20 mL) and saturated NaCl (2 x 20 mL), and dried (Na₂SO₄). Purification by flash chromatography (AcOEt/CH₂Cl₂/Petroleum ether, 1 :1 :1) afforded 7.41 g (60%) of the target product 13 as a colorless oil: m/z = 407 (M+H)⁺.

DIAD (1.02 niL, 5.17 mmol) was added at -15°C under nitrogen atmosphere to a solution of 13 (1.5 g, 3.69 mmol), quinoline 6 (1.39 g, 4.43 mmol) and triphenyl- phosphine (1.26 g, 4.80 mmol) in dry THF (40 mL). After 4.5 h, at -15°C, the reaction mixture was partitioned between ice-cold water and AcOEt, dried (Na₂SO₄) and evaporated. The crude material was purified by flash column chromatography (gradient of petroleum AcOEt/CH₂Cl₂, 1 :9 to 2:8) to give 1.45 g (56 %) of the target product 14: m/z = 703 (M+H)⁺.

A solution of 14 (1.07 g, 1.524 mmol) and Hoveyda-Grubbs 1^st generation catalyst (33 mg, 0.03 eq) in dried and degassed 1 ,2-dichloroethane (900 mL) was heated at 75°C under nitrogen for 12 h. Then, the solvent was evaporated and the residue purified by silica gel chromatography (25% EtOAc in CH₂Cl₂). 620 mg (60%) of pure macrocycle 15 were obtained, m/z = 674 (M+H)⁺. ¹H NMR (CDCl₃): 1.18-1.39 (m, 12H), 1.59 (m, IH), 1.70-2.08 (m, 5H), 2.28 (m, IH), 2.38 (m, IH), 2.62 (m, 2H), 2.68 (s, 3H), 2.83 (m, IH), 3.06 (s, 3H), 3.19 (sept, J= 6.7 Hz, IH), 3.36 (m, IH), 3.83 (m, IH), 3.97 (s, 3H), 4.09 (m, 2H), 4.65 (td, J= 4 Hz, 14 Hz, IH), 5.19 (dd, J= 4 Hz,

10 Hz, IH), 5.31 (m, IH), 5.65 (td, J= 4 Hz, 8 Hz, IH), 7.00 (s, IH), 7.18 (s, IH), 7.46

(d, J= 9 Hz, IH), 7.48 (s, IH), 8.03 (d, J= 9 Hz, IH).

A solution of lithium hydroxide (1.65 g, 38.53 mmol) in water (15 rnL) was added to a stirred solution of ester 15 (620 mg, 0.920 mmol) in THF (30 mL) and MeOH (20 mL). After 16 h at room temperature, the reaction mixture was quenched with NH₄Cl sat., concentrated under reduced pressure, acidified to pH 3 with HCl IN and extracted with CH₂Cl₂, dried (MgSO₄) and evaporated to give 560 mg (88%) of carboxylic acid 16. m/z = 647 (M+H)⁺. ¹H NMR (CDCl₃): 1.11-1.40 (m, 8H), 1.42-1.57 (m, 2H), 1.74 (m, 2H), 1.88-2.00 (m, 2H), 2.13 (m, IH), 2.28 (m, IH), 2.40 (m, IH), 2.59 (m, 2H), 2.67 (s, 3H), 2.81 (m, IH), 2.97 (s, 3H), 3.19 (m, IH), 3.31 (m, IH), 3.71 (m, IH), 3.96 (s, 3H), 4.56 (dt, J= 4 Hz, 12 Hz, IH), 5.23 (m, 2H), 5.66 (m, IH), 7.01 (s, IH), 7.10 (s, IH), 7.22 (d, J= IO Hz, IH), 7.45 (s, IH), 8.00 (d, J= 10 Hz, IH).

Example 2: Preparation of Λ/-[17-[2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methyl- quinolin-4-yloxy]- 13-methyl-2, 14-dioxo-3, 13-diazatricyclo[ 13.3.0.0⁴'⁶]octadec-7-ene- 4-carbonyll(cvclopropyl)sulfonamide (17) SIMEPREVIR

A solution of the compound 16 (560mg, 0.867 mmol) prepared according to Example 4, and carbonyldiimidazole (308 mg, 1.90 mmol) in dry THF (10 mL) was stirred at reflux under nitrogen for 2h. The reaction mixture was cooled to room temperature and cyclopropylsulfonamide (400 mg, 3.301 mmol) and DBU (286 mg, 1.881 mmol) were added. This solution was heated at 50⁰C for 15 h. Then, the reaction mixture was cooled down at room temperature and concentrated under reduced pressure. The residue was partitioned between CH₂Cl₂ and HCl 1 N, the organic layer was washed with brine, dried (MgSO₄) and evaporated. Purification by flash chromatography (gradient of EtOAc (0 to 25%) in CH₂Cl₂) afforded 314 mg of an off-white solid which was further washed with water, then isopropylether, and dried in the vacuum oven to deliver 282 mg (40%) of the pure title product 17, which is the compound of formula (I)  SIMEPREVIR , as a white powder: m/z = 750 (M+H)⁺.

¹H NMR (CDCl₃): 0.99-1.52 (m, 14H), 1.64-2.05 (m, 4H), 2.77 (m, IH), 2.41 (m, 2H), 2.59 (m, 2H), 2.69 (s, 3H), 2.92 (m, 2H), 3.04 (s, 3H), 3.19 (m, IH), 3.40 (m, 2H), 3.98 (s, 3H), 4.60 (t, J= 13 Hz, IH), 5.04 (t, J= 11 Hz, IH), 5.37 (m, IH), 5.66 (m, IH), 6.21 (s, IH), 7.02 (s, IH), 7.22 (d, J= IO Hz, IH), 7.45 (s, IH), 7.99 (d, J= 10 Hz, IH), 10.82 (broad s, IH).

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SYNTHESIS

WO2007014926A1

Example 4: preparation of 17-[2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methyl- quinolin-4-yloxy] - 13 -methyl-2, 14-dioxo-3 , 13 -diazatricyclo[ 13.3.0.0⁴'⁶]octadec-7-ene- 4-carboxylic acid (46) FREE ACID

Synthesis of 4-hvdroxy-2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methylquinoline (36) Step 1: synthesis of iV-(tert-butyloxycarbonyl)-3-methoxy-2-methylaniline (32)

31 32

Triethylamine (42.4 mL, 302 mmol) was added to a suspension of 3-methoxy-2- methylbenzoic acid (45.6 g, 274 mmol) in dry toluene (800 mL). A clear solution was obtained. Then, dppa (65.4 mL, 302 mmol) in toluene (100 mL) was slowly added. After 1 h at room temperature, the reaction mixture was successively heated at 50°C for 0.5 h, at 70°C for 0.5 h then at 100°C for 1 h. To this solution, t-BuOH (30.5 g, 411 mmol) in toluene (40 mL) was added at 100°C and the resulting mixture was refluxed for 7h. The solution was cooled to room temperature then successively washed with water, 0.5 N HCl, 0.5 N NaOH and brine, dried (Na₂SO₄), and evaporated to give 67 g of the target product: m/z = 237 (M)⁺.

Step 2: synthesis of 3-methoxy-2-methylaniline (33)

TFA (40.7 mL, 548 mmol) was added to a solution of iV-(tert-butyloxycarbonyl)-3- methoxy-2-methylaniline, in dichloromethane (500 mL). After 2 h at room temperature, TFA (40.7 mL, 548 mmol) was added and the resulting mixture was stirred at room temperature overnight. Then, volatiles were evaporated. The residue was triturated with toluene (100 mL) and diisopropylether (250 mL), filtered off and washed with diisopropyl ether (100 mL) to give 56.3 g of the title product as a TFA salt: m/z = 138 (M+H)⁺. The TFA salt was transformed to the free aniline by treatment with NaHCO₃.

Step 3: synthesis of (2-amino-4-methoxy-3-methylphenyl)(methyl)ketone (34)

A solution OfBCl₃ (1.0 M, 200 mL, 200 mmol) in CH₂Cl₂ was slowly added under nitrogen to a solution of 3-methoxy-2-methylaniline (26.0 g, 190 mmol) in xylene (400 mL). The temperature was monitored during the addition and was kept below 10°C. The reaction mixture was stirred at 5°C for 0.5 h. Then, dry acetonitrile (13 mL, 246 mmol) was added at 5°C. After 0.5 h at 5°C, the solution was transferred into a dropping funnel and slowly added at 5°C to a suspension OfAlCl₃ (26.7 g, 200 mmol) in CH₂Cl₂ (150 mL). After 45 min at 5°C, the reaction mixture was heated at 70°C under a nitrogen stream. After evaporation Of CH₂Cl₂, the temperature of the reaction mixture reached 65°C. After 12 h at 65°C, the reaction mixture was cooled at 0°C, poured onto ice (300 g), and slowly heated to reflux for 7h. After 2 days at room temperature, 6 N NaOH (50 mL) was added. The pH of the resulting solution was 2-3. The xylene layer was decanted. The organic layer was extracted with CH₂Cl₂. The xylene and CH₂Cl₂ layers were combined, successively washed with water, IN NaOH, and brine, dried (Na₂SO₄) and evaporated. The residue was triturated in diisopropyl ether at O⁰C, filtered off and washed with diisopropylether to give 13.6 g (40 %) of the title product as a yellowish solid: m/z = 180 (M+H)⁺.

Step 4: synthesis of 2'-[[(4-isopropylthiazole-2-yl)(oxo)methyl]amino]-4'-methoxy-3 '- methylacetophenone (35)

A solution of (2-amino-4-methoxy-3-methylphenyl)(methyl)ketone (18.6 g, 104 mmol) in dioxane (50 mL) was added under nitrogen to a suspension of 4-isopropylthiazole-2- carbonyl chloride in dioxane (250 mL). After 2 h at room temperature, the reaction mixture was concentrated to dryness. Then, the residue was partitioned between an aqueous solution OfNaHCO₃and AcOEt, organic layer was washed with brine, dried (Na₂SO₄), and evaporated. The residue was triturated in diisopropyl ether, filtered off and washed with diisopropyl ether to give 30.8 g (90 %) of the title product 35.

Step 5: synthesis of 4-hydroxy-2-(4-isopropylthiazole-2-yl)-7-methoxy-8- methylquinoline (36)

Potassium tert-butoxide (21.8 g, 195 mmol) was added to a suspension of 2'-[[(4-iso- propylthiazole-2-yl)(oxo)methyl]amino]-4'-methoxy-3'-methylacetophenone (35, 30.8 g, 92.7 mmol) in tert-butanol. The resulting reaction mixtures was heated at 100°C overnight. Then, the reaction mixture was cooled at room temperature and diluted with ether (100 mL). The precipitate was filtered off and washed with Et₂O to give a powder (fraction A). The mother liquor was concentrated in vacuo, triturated in ether, filtered off, and washed with ether to give a powder (fraction 2). Fractions 1 and 2 were mixed and poured into water (250 mL). The pH of the resulting solution was adjusted to 6-7 (control with pH paper) with HCl IN. The precipitate was filtered off, washed with water and dried. Then, the solid was triturated in diisopropyl ether, filtered off and dried to give 26 g (88%) of the title product 36 as a brownish solid: m/z = 315 (M+H)⁺.

Synthesis of (hex-5-enyl)(methyl)amine (38)

Sodium hydride (1.05 eq) was slowly added at 0°C to a solution of iV-methyltrifluoro- acetamide (25 g) in DMF (140 mL). The mixture was stirred for Ih at room temperature under nitrogen. Then, a solution of bromohexene (32,1 g) in DMF (25 mL) was added dropwise and the mixture was heated to 70°C for 12 hours. The reaction mixture was poured on water (200 mL) and extracted with ether (4 x 50 mL), dried (MgSO₄), filtered and evaporated to give 35 g of the target product 37 as a yellowish oil which was used without further purification in the next step.

Step B:

A solution of potassium hydroxide (187.7 g) in water (130 mL) was added dropwise to a solution of 37 (35 g) in methanol (200 mL). The mixture was stirred at room temperature for 12 hours. Then, the reaction mixture was poured on water (100 mL) and extracted with ether (4 x 50 mL), dried (MgSO₄), filtered and the ether was distilled under atmospheric pressure. The resulting oil was purified by distillation under vacuum (13 mm Hg pressure, 50°C) to give 7,4 g (34 %) of the title product 38 as a colourless oil: ¹H-NMR (CDCl₃): δ 5.8 (m, IH), 5 (ddd, J= 17.2 Hz, 3.5 Hz, 1.8 Hz, IH), 4.95 (m, IH), 2.5 (t, J= 7.0 Hz, 2H), 2.43 (s, 3H), 2.08 (q, J= 7.0 Hz, 2H), 1.4 (m, 4H), 1.3 (br s, IH).

Preparation of 17-r2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methylquinolin-4-yloxyl-

13-methyl-2,14-dioxo-3,13-diazatricvclori3.3.0.0⁴'⁶loctadec-7-ene-4-carboxylic acid

£46}

3-Oxo-2-oxa-bicyclo[2.2.1]heptane-5-carboxylic acid 39 (500 mg, 3.2 mmol) in 4 mlDMF was added at 0°C to HATU (1.34 g, 3.52 mmol) and iV-methylhex-5- enylamine (435 mg, 3.84 mmol) in DMF (3 mL), followed by DIPEA. After stirring for 40 min at 0°C, the mixture was stirred at room temperature for 5 h. Then, the solvent was evaporated, the residue dissolved in EtOAc (70 mL) and washed with saturated NaHCO₃ (10 mL). The aqueous layer was extracted with EtOAc (2 x 25 mL). The organic phases were combined, washed with saturated NaCl (20 mL), dried (Na₂SO₄), and evaporated. Purification by flash chromatography (EtOAc/petroleum ether, 2:1) afforded 550 mg (68%) of the target product 40 as a colorless oil: m/z = 252 (M+H)⁺.

A solution of LiOH (105 mg in 4 mlof water) was added at 0°C to the lactone amide 40. After Ih, the conversion was completed (HPLC). The mixture was acidified to pH 2 - 3 with IN HCl, extracted with AcOEt, dried (MgSO₄), evaporated, co-evaporated with toluene several times, and dried under high vacuum overnight to give 520 mg (88%) of the target product 41: m/z = 270 (M+H)⁺.

The l-(amino)-2-(vinyl)cyclopropanecarboxylic acid ethyl ester hydrochloride 42 (4.92 g, 31.7 mmol) and HATU (12.6 g, 33.2 mmol) were added to 41 (8.14 g, 30.2 mmol). The mixture was cooled in an ice bath under argon, and then DMF (100 mL) and DIPEA (12.5 mL, 11.5 mmol) were successively added. After 30 min at 0°C, the solution was stirred at room temperature for an additional 3 h. Then, the reaction mixture was partitioned between EtOAc and water, washed successively with 0.5 N HCl (20 mL) and saturated NaCl (2 x 20 mL), and dried (Na₂SO₄). Purification by flash chromatography (AcOEt/CH₂Cl₂/Petroleum ether, 1:1:1) afforded 7.41 g (60%) of the target product 43 as a colorless oil: m/z = 407 (M+H)⁺.

DIAD (1.02 mL, 5.17 mmol) was added at -15°C under nitrogen atmosphere to a solution of 43 (1.5 g, 3.69 mmol), quinoline 36 (1.39 g, 4.43 mmol) and triphenyl- phosphine (1.26 g, 4.80 mmol) in dry THF (40 mL). After 4.5 h, at -15°C, the reaction mixture was partitioned between ice-cold water and AcOEt, dried (Na₂SO₄) and evaporated. The crude material was purified by flash column chromatography (gradient of petroleum AcOEt/CH₂Cl₂, 1 :9 to 2:8) to give 1.45 g (56 %) of the target product 44: m/z = 703 (M+H)⁺.

A solution of 44 (1.07 g, 1.524 mmol) and Hoveyda-Grubbs 1^st generation catalyst (33 mg, 0.03 eq) in dried and degassed 1,2-dichloroethane (900 mL) was heated at 75°C under nitrogen for 12 h. Then, the solvent was evaporated and the residue purified by silica gel chromatography (25% EtOAc in CH₂Cl₂). 620 mg (60%) of pure macrocycle 45 were obtained, m/z = 674 (M+H)⁺. ¹H NMR (CDCl₃): 1.18-1.39 (m, 12H), 1.59 (m, IH), 1.70-2.08 (m, 5H), 2.28 (m, IH), 2.38 (m, IH), 2.62 (m, 2H), 2.68 (s, 3H), 2.83 (m, IH), 3.06 (s, 3H), 3.19 (sept, J= 6.7 Hz, IH), 3.36 (m, IH), 3.83 (m, IH), 3.97 (s, 3H), 4.09 (m, 2H), 4.65 (td, J= 4 Hz, 14 Hz, IH), 5.19 (dd, J= 4 Hz, 10 Hz, IH), 5.31 (m, IH), 5.65 (td, J= 4 Hz, 8 Hz, IH), 7.00 (s, IH), 7.18 (s, IH), 7.46 (d, J= 9 Hz, IH), 7.48 (s, IH), 8.03 (d, J= 9 Hz, IH).

Step F

A solution of lithium hydroxide (1.65 g, 38.53 mmol) in water (15 mL) was added to a stirred solution of ester 45 (620 mg, 0.920 mmol) in THF (30 mL) and MeOH (20 mL). After 16 h at room temperature, the reaction mixture was quenched with NH₄Cl sat., concentrated under reduced pressure, acidified to pH 3 with HCl IN and extracted with CH₂Cl₂, dried (MgSO₄) and evaporated to give 560 mg (88%) of carboxylic acid 46. m/z = 647 (M+H)⁺. ¹H NMR (CDCl₃): 1.11-1.40 (m, 8H), 1.42-1.57 (m, 2H), 1.74 (m, 2H), 1.88-2.00 (m, 2H), 2.13 (m, IH), 2.28 (m, IH), 2.40 (m, IH), 2.59 (m, 2H), 2.67 (s, 3H), 2.81 (m, IH), 2.97 (s, 3H), 3.19 (m, IH), 3.31 (m, IH), 3.71 (m, IH), 3.96 (s, 3H), 4.56 (dt, J= 4 Hz, 12 Hz, IH), 5.23 (m, 2H), 5.66 (m, IH), 7.01 (s, IH), 7.10 (s, IH), 7.22 (d, J= 10 Hz, IH), 7.45 (s, IH), 8.00 (d, J= 10 Hz, IH).

Example 5: Preparation of JV-ri7-r2-(4-isopropylthiazole-2-yl)-7-methoxy-8- methylquinolin-4- yloxyl - 13 -methyl-2, 14-dioxo-3 , 13 -diazatricyclol^" 13.3.0.0⁴'⁶loctadec- 7-ene-4-carbonyll (cvclopropyPsulfonamide (47) SIMEPREVIR

A solution of 17-[2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methylquinolin-4-yloxy]- 13-methyl-2, 14-dioxo-3, 13-diazatricyclo[l 3.3.0.04,6]octadec-7-ene-4-carboxylic acid 46 (560mg, 0.867 mmol) prepared according to Example 4, and carbonyldiimidazole (308 mg, 1.90 mmol) in dry THF (10 mL) was stirred at reflux under nitrogen for 2h. The reaction mixture was cooled to room temperature and cyclopropylsulfonamide (400 mg, 3.301 mmol) and DBU (286 mg, 1.881 mmol) were added. This solution was heated at 50°C for 15 h. Then, the reaction mixture was cooled down at room temperature and concentrated under reduced pressure. The residue was partitioned between CH2CI2 and HCl 1 N, the organic layer was washed with brine, dried (MgSO₄) and evaporated. Purification by flash chromatography (gradient of EtOAc (0 to 25%) in CH2CI2) afforded 314 mg of an off-white solid which was further washed with water, then isopropylether, and dried in the vacuum oven to deliver 282 mg (40%) of the pure title product 47  SIMEPREVIR as a white powder: m/z = 750 (M+H)⁺.

¹H NMR (CDCl₃): 0.99-1.52 (m, 14H), 1.64-2.05 (m, 4H), 2.77 (m, IH), 2.41 (m, 2H), 2.59 (m, 2H), 2.69 (s, 3H), 2.92 (m, 2H), 3.04 (s, 3H), 3.19 (m, IH), 3.40 (m, 2H), 3.98 (s, 3H), 4.60 (t, J= 13 Hz, IH), 5.04 (t, J= 11 Hz, IH), 5.37 (m, IH), 5.66 (m, IH), 6.21 (s, IH), 7.02 (s, IH), 7.22 (d, J= 10 Hz, IH), 7.45 (s, IH), 7.99 (d, J= 10 Hz, IH), 10.82 (broad s, IH).

.......................

REFERENCES

1.  “Medivir Announces That Simeprevir (TMC435) Data Will Be Presented at the Upcoming AASLD Meeting”. Yahoo News. October 1, 2012. Retrieved November 6, 2012.
2.  Lin, TI; Lenz, O; Fanning, G; Verbinnen, T; Delouvroy, F; Scholliers, A; Vermeiren, K; Rosenquist, A et al. (2009). “In vitro activity and preclinical profile of TMC435350, a potent hepatitis C virus protease inhibitor”. Antimicrobial agents and chemotherapy 53 (4): 1377–85. doi:10.1128/AAC.01058-08. PMC 2663092. PMID 19171797. |displayauthors= suggested (help)
3.  “Phase 3 Studies Show Simeprevir plus Interferon/Ribavirin Cures Most Patients in 24 Weeks”. hivandhepatitis.com. December 27, 2012.
4.  Medivir announces TMC435 in an expanded clinical collaboration. Medivir. 18 April 2012.
5.  Results from a phase IIa study evaluating Simeprevir and Sofosbuvir in prior null responder Hepatitis C patients have been presented at CROI. 6 March 2013.
6. TMC-435350
   Drugs Fut 2009, 34(7): 545
7. Structure-activity relationship study on a novel series of cyclopentane-containing macrocyclic inhibitors of the hepatitis C virus NS3/4A protease leading to the discovery of TMC435350
   Bioorg Med Chem Lett 2008, 18(17): 4853
8. Synthesis of enantiomerically pure trans-3,4-substituted cyclopentanols by enzymatic resolution
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7. WO 2011113859
8. WO 2013041655

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WO2005073195A2 *	28 Jan 2005	11 Aug 2005	Per-Ola Johansson	Hcv ns-3 serine protease inhibitors
WO2007014926A1 *	28 Jul 2006	8 Feb 2007	Tibotec Pharm Ltd	Macrocyclic inhibitors of hepatitis c virus

The compound ritonavir, and pharmaceutically acceptable salts thereof, and methods for its preparation are described in WO94/14436. For preferred dosage forms of ritonavir, see US6,037, 157, and the documents cited therein: US5,484, 801, US08/402,690, and WO95/07696 and WO95/09614. Ritonavir has the following formula:

Glenmark conferred with Best Biotech New Molecular Entity Patent award « New Drug Approvals

Glenmark conferred with Best Biotech New Molecular Entity Patent award « New Drug Approvals:

'via Blog this'

Afinitor, Everolimus

Afinitor (everolimus)

40-O-(2-hydroxyethyl)-rapamycin

Company: Novartis
Approval Status: Approved July 2012
Treatment Area: hormone receptor-positive, HER2-negative breast cancer

Everolimus is a derivative of Rapamycin (sirolimus), and works similarly to Rapamycin as an mTOR (mammalian target of rapamycin) inhibitor. It is currently used as an immunosuppressant to prevent rejection of organ transplants. In a similar fashion to other mTOR inhibitors Everolimus' effect is solely on the mTORC1 protein and not on the mTORC2 protein.

Also known as: Afinitor, Certican, Zortress, SDZ-RAD, RAD001, Everolimus [USAN], 42-O-(2-Hydroxyethyl)rapamycin, RAD 001

Molecular Formula: C₅₃H₈₃NO₁₄   Molecular Weight: 958.22442

159351-69-6  CAS NO

BRANDS

Afinitor	Novartis
Certican	Novartis
VOTUBIA	Novartis
Zortress	Novartis

Afinitor (everolimus), an inhibitor of mTOR (mammalian target of rapamycin), is an antineoplastic agent.

Afinitor is specifically approved for the treatment of postmenopausal women with advanced hormone receptor-positive, HER2-negative breast cancer (advanced HR+ BC) in combination with exemestane, after failure of treatment with letrozole or anastrozole.

Afinitor is supplied as a tablet for oral administration. The recommended dose of Afinitor for breast cancer is 10 mg, to be taken once daily, at the same time every day, either consistently with food or consistently without food.

FDA Approval
The FDA approval of Afinitor for the treatment of advanced hormone receptor-positive, HER2-negative breast cancer was based on a randomized, double-blind, multicenter study in 724 postmenopausal women with estrogen receptor-positive, HER 2/neu-negative advanced breast cancer with recurrence or progression following prior therapy with letrozole or anastrozole.

Everolimus is indicated for the treatment of postmenopausal women with advanced hormone receptor-positive, HER2-negative breast cancer (advanced HR+ BC) in combination with exemestane, after failure of treatment with letrozole or anastrozole. Indicated for the treatment of adult patients with progressive neuroendocrine tumors of pancreatic origin (PNET) with unresectable, locally advanced or metastatic disease. Indicated for the treatment of adult patients with advanced renal cell carcinoma (RCC) after failure of treatment with sunitinib or sorafenib. Indicated for the treatment of adult patients with renal angiomyolipoma and tuberous sclerosis complex (TSC), not requiring immediate surgery. Indicated in pediatric and adult patients with tuberous sclerosis complex (TSC) for the treatment of subependymal giant cell astrocytoma (SEGA) that requires therapeutic intervention but cannot be curatively resected.

Everolimus (RAD-001) is the 40-O-(2-hydroxyethyl) derivative of sirolimus and works similarly to sirolimus as an inhibitor of mammalian target of rapamycin (mTOR).

It is currently used as an immunosuppressant to prevent rejection of organ transplants and treatment of renal cell cancer and other tumours. Much research has also been conducted on everolimus and other mTOR inhibitors for use in a number of cancers.

It is marketed by Novartis under the tradenames Zortress (USA) and Certican (Europe and other countries) in transplantation medicine, and Afinitor in oncology.

EVEROLIMUS

AFINITOR (everolimus), an inhibitor of mTOR, is an antineoplastic agent.

The chemical name of everolimus is (1R,9S,12S,15R,16E,18R,19R,21R,23S,24E,26E,28E,30S,32S,35R)-1,18- dihydroxy-12-{(1R)-2-[(1S,3R,4R)-4-(2-hydroxyethoxy)-3-methoxycyclohexyl]-1-methylethyl}-19,30-dimethoxy15,17,21,23,29,35-hexamethyl-11,36-dioxa-4-aza-tricyclo[30.3.1.0^4,9]hexatriaconta-16,24,26,28-tetraene-2,3,10,14,20pentaone.

The molecular formula is C₅₃H₈₃NO₁₄ and the molecular weight is 958.2. The structural formula is:

AFINITOR Tablets are supplied for oral administration and contain 2.5 mg, 5 mg, 7.5 mg, or 10 mg of everolimus. The tablets also contain anhydrous lactose, butylated hydroxytoluene, crospovidone, hypromellose, lactose monohydrate, and magnesium stearate as inactive ingredients.

AFINITOR DISPERZ (everolimus tablets for oral suspension) is supplied for oral administration and contains 2 mg, 3 mg, or 5 mg of everolimus. The tablets for oral suspension also contain butylated hydroxytoluene, colloidal silicon dioxide, crospovidone, hypromellose, lactose monohydrate, magnesium stearate, mannitol, and microcrystalline cellulose as inactive ingredients.

1. R.N Formica Jra, K.M Lorberb, A.L Friedmanb, M.J Biaa, F Lakkisa, J.D Smitha, M.I Lorber (March 2004). “The evolving experience using everolimus in clinical transplantation”. Elsevier 36 (2): S495–S499.
2.  “Afinitor approved in US as first treatment for patients with advanced kidney cancer after failure of either sunitinib or sorafenib” (Press release). Novartis. 2009-03-30. Retrieved April 6, 2009.
3. “Novartis receives US FDA approval for Zortress (everolimus) to prevent organ rejection in adult kidney transplant recipients” (Press release). Novartis. 2010-04-22. Retrieved April 26, 2010.
4. “Novartis’ Afinitor Cleared by FDA for Treating SEGA Tumors in Tuberous Sclerosis”. 1 Nov 2010.
5. http://www.fda.gov/NewsEvents/Newsroom/PressAnnouncements/ucm254350.htm
6. “US FDA approves Novartis drug Afinitor for breast cancer”. 20 Jul 2012.

PATENTS

Country	Patent Number	Approved	Expires (estimated)
United States	6440990	1993-09-24	2013-09-24
Canada	2145383	2004-11-16	2013-09-24
Canada	2225960	2004-05-11	2016-07-12
United States	7297703	1999-12-06	2019-12-06

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WO1994009010A1	Sep 24, 1993	Apr 28, 1994	Sandoz Ag	O-alkylated rapamycin derivatives and their use, particularly as immunosuppressants
WO2007135397A1 *	May 18, 2007	Nov 29, 2007	Christoph Beckmann	36 -des (3 -methoxy-4 -hydroxycyclohexyl) 36 - (3 -hydroxycycloheptyl) derivatives of rapamycin for the treatment of cancer and other disorders
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US5665772	Sep 24, 1993	Sep 9, 1997	Sandoz Ltd.	O-alkylated rapamycin derivatives and their use, particularly as immunosuppressants
US20030125800	Apr 24, 2002	Jul 3, 2003	Shulze John E.	Drug-delivery endovascular stent and method for treating restenosis

...........................................

Rapamycin is a known macrolide antibiotic produced by Streptomvces hvgroscopicus. having the structure depicted in Formula A:

See, e.g., McAlpine, J.B., et al., J. Antibiotics (1991) 44: 688; Schreiber, S.L., et al., J. Am. Chem. Soc. (1991) J_13: ⁷⁴³³'- US Patent No. 3 929 992. Rapamycin is an extremely potent immunosuppressant and has also been shown to have antitumor and antifungal activity. Its utility as a pharmaceutical, however, is restricted by its very low and variable bioavailabiiity as well as its high toxicity. Moreover, rapamycin is highly insoluble, making it difficult to formulate stable galenic compositions.

Everolimus, 40-O-(2-hydroxyethyl)-rapamycin of formula (1) is a synthetic derivative of rapamycin (sirolimus) of formula (2), which is produced by a certain bacteria strain and is also pharmaceutically active.

(1)                                                                                                               (2)

Everolimus is marketed under the brand name Certican for the prevention of rejection episodes following heart and kidney transplantation, and under the brand name Afinitor for treatment of advanced kidney cancer.

Due to its complicated macrolide chemical structure, everolimus is, similarly as the parent rapamycin, an extremely unstable compound. It is sensitive, in particular, towards oxidation, including aerial oxidation. It is also unstable at temperatures higher than 25°C and at alkaline pH.

Everolimus and a process of making it have been disclosed in WO 94/09010

Synthesis

Alkylation of rapamycin (I) with 2-(tert-butyldimethylsilyloxy)ethyl triflate (II) by means of 2,6-lutidine in hot toluene gives the silylated target compound (III), which is deprotected by means of 1N HCl in methanol (1). (Scheme 21042401a) Manufacturer Novartis AG (CH). References 1. Cottens, S., Sedrani, R. (Sandoz-Refindungen VmbH; Sandoz-Patent GmbH; Sandoz Ltd.). O-Alkylated rapamycin derivatives and their use, particularly as immunosuppressants. EP 663916, EP 867438, JP 96502266, US 5665772, WO 9409010.EP 0663916; EP 0867438; JP 1996502266; JP 1999240884; US 5665772; WO 9409010

..............

SYNTHESIS

https://www.google.com/patents/WO2012103960A1

(US 5,665,772, EP 663916). The process principle is shown in the scheme below, wherein the abbreviation RAP-OH has been used as an abbreviation for the rapamycin structure of formula (2) above, L is a leaving group and P is a trisubstituted silyl group serving as a OH- protective group.

RAP-OH + L-CH₂-CH₂-0-P — --> RAP-O-CH2-CH2-O-P — - > RAP-O-CH2-CH2-OH

(2)                                                 (4)                                                                 (1)

Specifically, the L- group is a trifluoromethanesulfonate (triflate) group and the protective group P- is typically a tert-butyldimethylsilyloxy- group. Accordingly, the known useful reagent within the above general formula (3) for making everolimus from rapamycin is 2-(tert-butyldimethylsilyloxy)ethyl triflate of formula (3 A):

According to a known synthetic procedure disclosed in Example 8 of WO 94/09010 and in Example 1 of US application 2003/0125800, rapamycin (2) reacts in hot toluene and in the presence of 2,6-lutidine with a molar excess of the compound (3 A), which is charged in several portions, to form the t-butyldimethylsilyl-protected everolimus (4A). This compound is isolated and deprotected by means of IN aqueous HC1 in methanol. Crude everolimus is then purified by column chromatography. Yields were not reported.

(2)                                       (3A)                              (4A)                                (1)

In an article of Moenius et al. (J. Labelled Cpd. Radiopharm. 43, 113-120 (2000)), which used the above process for making C14-labelled and tritiated everolimus, a diphenyl- tert.butylsilyloxy -protective group was used as the alkylation agent of formula (3B).

Only 8% yield of the corresponding compound (4B)

and 21% yield of the compound (1) have been reported.

Little is known about the compounds of the general formula (3) and methods of their preparation. The synthesis of the compound (3 A) was disclosed in Example 1 of US application 2003/0125800. It should be noted that specification of the reaction solvent in the key step B of this synthesis was omitted in the disclosure; however, the data about isolation of the product allow for estimation that such solvent is dichloromethane. Similarly also a second article of Moenius et al. (J. Labelled Cpd. Radiopharm.42, 29-41 (1999)) teaches that dichloromethane is the solvent in the reaction.

It appears that the compounds of formula (3) are very reactive, and thus also very unstable compounds. This is reflected by the fact that the yields of the reaction with rapamycine are very low and the compound (3) is charged in high molar extent. Methods how to monitor the reactivity and/or improve the stability of compounds of general formula (3), however, do not exist.

Thus, it would be useful to improve both processes of making compounds of formula (3) and, as well, processes of their application in chemical synthesis.

xample 6: 40-O-[2-((2,3-dimethylbut-2-yl)dimethylsilyloxy)ethyl]rapamycin

In a 100 mL flask, Rapamycin (6 g, 6.56 mmol) was dissolved in dimethoxyethane (4.2 ml) and toluene (24 ml) to give a white suspension and the temperature was raised to 70°C. After 20 min, N,N-diisopropylethylamine (4.56 ml, 27.6 mmol) and 2-((2,3-dimethylbutan-2- yl)dimethylsilyloxy)ethyl trifluoromethanesulfonate (8.83 g, 26.3 mmol) were added in 2 portions with a 2 hr interval at 70°C. The mixture was stirred overnight at room temperature, then diluted with EtOAc (40 ml) and washed with sat. NaHC0₃ (30 ml) and brine (30 ml). The organic layer was dried with Na₂S0₄, filtered and concentrated. The cmde product was chromatographed on a silica gel column (EtOAc/heptane 1/1 ; yield 4.47 g).

Example 7: 40-O-(2-hydroxyethyl)-rapamycin [everolimus]

In a 100 mL flask, 40-O-[2-((2,3-dimethylbut-2-yl)dimethylsilyloxy)ethyl]rapamycin (4.47 g, 4.06 mmol) was dissolved in methanol (20 ml) to give a colorless solution. At 0°C, IN aqueous hydrochloric acid (2.0 ml, 2.0 mmol) was added and the mixture was stirred for 90 min. The reaction was followed by TLC (ethyl acetate/n-heptane 3 :2) and HPLC. Then 20 ml of saturated aqueous NaHC03 were added, followed by 20 ml of brine and 80 ml of ethyl acetate. The phases were separated and the organic layer was washed with saturated aqueous NaCl until pH 6/7. The organic layer was dried by Na₂S0₄, filtered and concentrated to yield 3.3 g of the product.

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SYNTHESIS

https://www.google.co.in/patents/WO1994009010A1

Example 8: 40-O-(2-Hydroxy)ethyl-rapamycin

a) 40-O-[2-(t-Butyldimethylsilyl)oxy]ethyl-rapamycin

A solution of 9.14 g (10 mmol) of rapamycin and 4.70 mL (40 mmol) of 2,6-lutidine in 30 mL of toluene is warmed to 60°C and a solution of 6.17 g (20 mmol) of 2-(t-butyldimethylsilyl)oxyethyl triflate and 2.35 mL (20 mmol) of 2,6-lutidine in 20 mL of toluene is added. This mixture is stirred for 1.5h. Then two batches of a solution of 3.08 g (10 mmol) of triflate and 1.2 mL (10 mmol) of 2,6-lutidine in 10 mL of toluene are added in a 1.5h interval. After addition of the last batch, stirring is continued at 60°C for 2h and the resulting brown suspension is filtered. The filtrate is diluted with ethyl acetate and washed with aq. sodium bicarbonate and brine. The organic solution is dried over anhydrous sodium sulfate, filtered and concentrated. The residue is purified by column chromatography on silica gel (40:60 hexane-ethyl acetate) to afford 40-O-[2-(t-butyldimethylsilyl)oxy]ethyl-rapamycin as a white solid: ¹H NMR (CDCl₃) δ 0.06 (6H, s), 0.72 (1H, dd), 0.90 (9H, s), 1.65 (3H, s), 1.75 (3H, s), 3.02 (1H, m), 3.63 (3H, m), 3.72 (3H, m); MS (FAB) m/z 1094 ([M+Na]⁺), 1022 ([M-(OCH₃+H₂O)]⁺).

b) 40-O-(2-Hydroxy)ethyl-rapamycin

To a stirred, cooled (0°C) solution of 4.5 g (4.2 mmol) of 40-O-[2-(t-butyldimethylsilyl)oxy]ethyl-rapamycin in 20 mL of methanol is added 2 mL of IN HCl. This solution is stirred for 2h and neutralized with aq. sodium bicarbonate. The mixture is extracted with three portions of ethyl acetate. The organic solution is washed with aq.

sodium bicarbonate and brine, dried over anhydrous sodium sulfate, filtered and

concentrated. Purification by column chromatography on silica gel (ethyl acetate) gave the title compound as a white solid:¹H NMR (CDCl₃) δ 0.72 (1H, dd), 1.65 (3H, s), 1.75 (3H, s), 3.13 (5H, s and m), 3.52-3.91 (8H, m); MS (FAB) m/z 980 ([M+Na]⁺), 926 ([M-OCH₃]⁺), 908 ([M-(OCH₃+H₂O)]⁺), 890 ([M-(OCH₃+2H₂O)]⁺), 876 ([M-(2CH₃OH+OH)]⁺), 858 ([M-(OCH₃+CH₃OH+2H₂O)]⁺).

MBA (rel. IC50) 2.2

IL-6 dep. prol. (rel. IC50) 2.8

MLR (rel. IC50) 3.4

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synthesis

Everolimus (Everolimus) was synthesized by the Sirolimus (sirolimus, also known as rapamycin Rapamycin) ether from. Sirolimus is from the soil bacterium Streptomyces hygroscopicus isolated metabolites. Activation end sirolimus (triflate, Tf) the other end of the protection (t-butyldimethylsilyl, TBS) of ethylene glycol 1 reaction of 2 , because the hydroxyl group 42 hydroxyl site over the 31-bit resistance is small, so the reaction only occurs in 42. Compound 2under acidic conditions TBS protection is removed everolimus.