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ABCD
The Process Development of New Hepatitis C Protease Inhibitors
Vittorio Farina,
Dept. of Chemical Development,
Boehringer Ingelheim, Ridgefield CT
New Methods in Process Chemistry, Univ. of Zurich, 14 Oct. 2004
ABCDTherapeutic Need
• Over 170 million people are infected with hepatitis C virus (HCV) worldwide (WHO 1997).
• > 70% of infected individuals will develop chronic hepatitis C. Chronic hepatitis C is a progressive condition leading to end-stage liver disease (cirrhosis and hepatocellular carcinoma).
• Available interferon-containing therapies have limited efficacy and have significant side effects.
ABCDHepatitis C - Standard Therapy
• Interferon or (subcutaneous administration).
• Rebetron: Intron A + oral ribavirin, launched in 1998 (USA).
• New standard therapy: 50% sustained response rate(genotype I virus).
• Treatment cost: ~ $17,000/year ($1,440/month) per patient.
ABCDHepatitis C Virus NS3 Protease
•Cleaves at NS3/4A; NS4A/4B; NS4B/5A and NS5A/5B. Good medicinal target.
•In in vitro assays, it was found that protease is inhibited by a hexapeptide (N-terminal) produced by cleavage of substrate from NS5A/5B [DDIVPC].
•This can be the basis for rational drug design.
•Rapid peptide screening shows: DDIVPC IC50 = 71MDdIVPC IC50 = 4 M (replaced L-Asp with D-Asp)
M. Llinàs-Brunet et al. Bioorg. Med. Chem. Lett. 1998, 8, 1713
ABCD
H2NNH
HN
NH
N
HO2C
O
O
O
O
CO2H
O NH
CO2H
SH
PEPTIDE LEAD / DDIVPC / IC50= 71 M
AcHNNH
HN
NH
N
HO2C
O
O
O
OO N
HCO2H
CO2H
O
MODIFIED PEPTIDE IC50 = 0.013 M
Amino Cap
Modified P4
Huge LipophilicPocket at P2
Modified P1
L-Aspto D-Glu
Initial SAR Work
M. Llinàs-Brunet et al. Bioorg. Med. Chem. Lett. 2000, 10, 2267
ABCDFrom Peptides to Peptidomimetics
NHN
OH
O
OO
NH
O
O
O
N
MeO
S
N HN
S
S
S
R
R
BILN 2061Clinical Candidate
Reversible inhibitor of HCV genotypeIa and Ib, Ki = 0.3-0.66 nM.Good PK, metabolic stability.
Structure-Activity Relationships:Angew. Chem. Int. Ed. Engl. 2003, 42, 1356J. Med. Chem. 2004, 47, 123J. Med. Chem. 2004, 47, 1605J. Med. Chem. 2004, 47, 2511Nature 2003, 426, 186
Discovery Synthesis:Org. Lett. 2004, 4, 2901.
ABCDBILN 2061: Clinical Proof of Principle
•Administered in solution (Water, PEG 400, EtOH) at 200 mg bid, for 2 days.
•Viral titer dropped from an average of 106 copies/mL to <1,500 (LOD) in most patients.
•Viral titer rebounded in 6-13 days after treatment was discontinued (as expected).
H. Hinrichsen et al. Gastroenterology, in press.
ABCDBILN 2061: Retrosynthetic Analysis
NHN
OH
O
OO
NH
O
O
O
N
MeO
S
N HN
H2NOH
OO
NHO
O
HO
N
MeO
S
N HN
OH
N
O
OH
OH
S
S
S
R
R
BILN 2061
H
INRF3 (P1) INRF15 (P3) INRF13INRF1 (P2)
C40H50N6O8SMW 774.955 Asymmetric Centers(Z) -Alkene15-Membered Ring
Amenable to convergentassembly
ABCDProgram Goals
• Develop “Expedient Route” in order to supply early batches for toxicologyand phase Ia studies (0.5 – 1.0 Kg).
• Develop cost-effective “Practical Route” and develop it into a process that can form the basis for commercial manufacturing:
- Highly Reproducible.- No chromatographies.- Control of all organic impurities > 0.1 %.- Control of all residual organic volatiles (ICH guidelines).- Control of inorganic impurities (Ru?).- Control of solid-state properties of Active Pharmaceutical Ingredients (API) (polymorph, crystal habit, particle size).
N.G. Anderson “Practical Process Research and Development”, Academic Press,San Diego, 2000.
ABCDThe BILN 2061 Chemistry Team
BIPI Process ChemistryDr. Nathan YeeDr. Yannis HoupisDr. Vittorio FarinaDr. Nizar HaddadDr. Rogelio FrutosDr. Fabrice GallouDr. Xiao-jun WangDr. Xudong WeiDr. Robert SimpsonDr. XuWu FengVictor FuchsYibo XuJonathan TanLi ZhangJinghua XuLana SmithJana VitousEarl Spinelli
BIKG Biberach, ProcessDr. Volker EhrigDr. Rolf HerterDr. Juergen SchnaubeltDr. Rainer Soyka
BI Laval Discovery ChemistryDr. Pierre BeaulieuDr. Anne-Marie FaucherDr. Youla TsantritzosDr. Montse Llinàs-BrunetDr. Murray BaileyDr. Stephen KawaiDr. Bruno SimoneauDr. Jean-Marie FerlandChristian BrochuJean-Simon DuceppeElise GhiroVida GorysTed HalmosMartin PoirierJames GillardBruno HachéColette Boucher
BIKG Ingelheim, ProcessDr. Wolfgang DerschDr. Wendelin SamstagDr. Thomas NicolaDr. Kai Donsbach
ABCDSynthesis of Racemic P1 Unit
O
OEtNPh
CO2EtNPh
PhPh
+CO2EtN
Ph Ph
BrBr
tBuOK, THF
-78 oC
NPh
Ph
[3,3]
CO2Et
HN
PhPh
CO2Et
organic soluble
2:1HCl
CO2EtH2N
E, isolated from aqueous(ca. 30%)
E
Z
Very rapid entry into racemicINRF3 Et ester.• Diastereoselectivity is modest.• Yield must be increased.• Cryogenic conditions must be removed.• Needs resolution.
Discovery Chemistry, Laval
ABCDRoute to P1: Optimization
Imine: Benzaldehyde best (selected).
Counterion effect (tBuO base) on diastereoselectivity: Li > Na > K (d.e.=20:1 with Li, 10:1 with Na, 2:1 with K).
Solvents: Non-polar solvents (PhMe) give best yields.
Esters: Et best, Me 5-8% lower yield.
Concentration: No difference in yield over practicable range (0.28-0.7 M).
Temperature: No difference in range 20-45 0C . No improvements at lower temperature, just prolonged reaction times.
ABCDP1:Scalable Process
Process Chemistry, Ridgefield and BIKG Biberach
CO2MeN
ClH3N CO2Me
PhCHO
+
1 eq HC(OMe)3
PhMe, NMP (8:1)NEt3, 15-25oC 5h
N CO2MePh
aqueous wash, concentrate vol. 60%
BrBr
tBuOLi
MTBE, 15-20 oC
3h
Ph
washed with aqueous
aq. HCl / pH=2
2.5 h , rtCO2MeH2N
NaOH, Boc2O
acetone
CO2MeBocHN
Solvent change to acetone
1.7 M conc.
distil acetone
work-up MTBE
waterRacemate
ABCDP1: Scalable Process
Process Chemistry, Ridgefield and Biberach
CO2MeBocHN
Alcalase, pH 8.15
Na2HPO4
10:1 water / acetone, 40 oC
70 h
0.12 - 0.13M
Racemate
CO2MeBocHN
31-32%in situ yield overall
>98% ee
work-up
evap. acetone,extractionwater/MTBE
1 equiv pTsOH
MIBK
40 oC, 2h.the xtal at 2-3 oC
CO2MeTsOH . H2N
>99.9 % ee
28.7% overall
Features:• No isolations till the end.• Use of NMP in first step to increase solubility and throughput (>10 fold).• Scalable to >100 Kg•Tosylate aids isolation.
ABCDP3 Unit: Scalable Synthesis
ClBr
MgBr ClNHAc
CO2Et
CO2Et
NHAc
CO2H
NH2
CO2H
O
ClOHN
CO2H / HNCy2
O
O
Li2CuCl4ether/THF
1. KOHH2O/EtOH
Acylase 1 (0.5% w/w)pH = 7.6 - 7.75
2. citric acid
+
90% assay
NHAc
EtO2C CO2Et
DMF, K2CO3, Cs2CO3
80 oC
85% assay12
3 (isolated ,cryst. IPA/H2O)
4 (>99% ee)
46%
1.
NaOH, H2O
2. HNCy2, MTBE
85%
• Preparation of 2 using 1,4-dibromobutane gives lower yield in Grignard coupling.• Recycle of (R)-3 possible by racemization with Ac2O at reflux followed by resolutionas above (71% total yield after one recycle).• DCHA salt used for crystallinity. Process smoothly scalable to >100 Kg.
Process Chemistry, Ridgefield
ABCDSynthesis of Quinolone Unit
MeO NH2
AcCl, BCl3, AlCl3, CH2Cl2
-50 to 40 oCMeO NH2 HO NH2
O O
1, 35% 2, 10%
+
• Literature procedure: J. Org. Chem. 1979, 44, 578; J. Am. Chem. Soc. 1978, 100, 4842.• Reported yield: 40%. Obtained on 300g scale: 35%.• Difficulty in scale-up due to further reaction (decomposition) of 1 and emulsions during work-up.• m-Anisidine is a very attractive starting material.
Discovery Chemistry, Laval, and Process Chemistry, Ridgefield
Step 1: Friedel-Crafts Reaction
ABCDAn Alternative: Sugasawa Reaction
OMe
NH2O
MeO NH2
CH3CN, BCl3, AlCl3
MeO NH2
O
+
1 2
PhMe: CH2Cl2 = 1: 1.6
Solvent Ratio 1/2
CH2Cl2 1.4 : 1
PhMe 6 : 1
Highlight of solvent effects:
• Order of addition important: preform anisidine/BCl3 complex, then CH3CN, then AlCl3
• Optimum temperature: 40 oC. Optimum pH for work-up: 3.0• Purified by MTBE slurry at reflux.• Typical isolated yields: 42-47% on multikilo scale.
Discovery Chemistry, Laval, and Process Chemistry, Ridgefield
ABCDSynthesis of Quinolone Unit (cont.)
Discovery Chemistry, Laval, and Process Chemistry, Ridgefield
HN
S
NH2 +HO
O
O
Brdioxane , 100 oC
94%
NS
NHiPr
CO2H
. HBr
Oxalyl Chloride
CH2Cl2, NEt3, 20 oC
NS
NHiPr
COCl
MeO NH2
O
0 - 14 oC
89% isolated
3, crystalline solid
MeO NH
O
O
S
N NHiPr
tBuOK, tBuOH, THF
78-84 oC NH
O
MeO
S
NNHiPr
75% isol.
1 24
5
6 7
ABCDScale-up Problems
Process Chemistry, Ridgefield
MeO NH
O
O
S
N NHiPr
tBuOK, tBuOH, THF
78-84 oC NH
O
MeO
S
NNHiPr
6
7
NH
O
MeO
S
NNHiPr
HN
O
MeO
S
NNHiPr
NH
N
S NHiPr
OMeO
Early lots had only 64-78%purity (HPLC)Major impurities Isolated:
A: formed from CH2Cl2(incomplete removal)
B: Formed at lower temperatures.Ratio (Product/B) is solvent dependent
ABCDSolution to Scale-up Problems
MeO NH
O
O
S
N NHiPr
tBuOK, DME
89-90 oC NH
O
MeO
S
NNHiPr
6
7
• DME best solvent to minimize formation of B (probably minimizes deprotonation of NH-i Pr group).• Dichloromethane “chased” with DME prior to cyclization.• Crude product is purified by slurry in DME / water.• HPLC Assay yield 82%, isolated 77% .• Reproduced on multikilo scale.
Process Chemistry, Ridgefield
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