Post on 11-Feb-2022
Prodrugs of Nucleoside and Nucleotide Analogs for the Treatment of HCV:
An Overview of Concepts and Chemistry
Michael J. Sofia, Ph.D. Gilead Sciences
7th International Workshop on Hepatitis C – Resistance and New Compounds
Cambridge, MA June 28-29, 2012
What is a Prodrug?
• A compound that undergoes biotransformation prior to eliciting its pharmacological effect – The administered therapeutic agent is inactive but is
transformed into one or more active metabolites – Usually implemented to address a deficiency
associated with the active agent • Chemical or biotransformation of a structural molecular
functionality that produces the actual active agent • Attachment of a promoiety to a pharmacologically active
agent such that the prodrug can overcome the barrier that hinders the optimal use of the active agent
Albert, A., Nature, 1958, 182, 421 Bundgaard, H., In A Textbook of Drug Design and Development , Harwood, Reading, UK, 19921, 113
The Prodrug Concept
Biologial B
arrier
Drug
Drug
Promoiety
Drug
Promoiety
Drug Promoiety +
Biotransformation
Non-toxic & rapidly cleared
Key Issues Addressed by Prodrugs
• Bioavailability – Aqueous solubility – Poor passive intestinal absorption - lipophilicity – Leveraging transporter-mediated intestinal absorption
• Peptide transporters • Nucleoside transporters • Ion transporters • Bile acid transporters • Vitamin transporters
– Fast metabolism • Tissue Selectivity (“magic bullet”)
– Passive enrichment – Transporter targeting – Targeting tissue- or cell-specific enzymes – Targeting surface antigens
Marketed Prodrugs
HO
OO
HO O
Lovastatin
O
OO
NH2
HN
O
Oseltamivir
N
OHO
POO
OOO
Fosinopril
N
N N
N
O
P
NH2
O
O OOO
O O
Adefovir Dipivoxil
NH
NN
N
O
O
O
NH2
O
NH2H
Valaciclovir
Guidances for Prodrug Development
• A comprehensive understanding of the ADME/PK of the prodrug and the parent drug is critical.
• Depending on the rate of conversion and the site of metabolism, prodrugs may alter the tissue distribution, efficacy, and toxicity of the parent drug.
• The promoiety should ideally be safe and rapidly excreted from the body.
• The choice of promoiety should be considered with respect to the disease state, dose, and duration of therapy
Unique Characteristics of Nucleos(t)ides
O
HO
Kinase 1 Kinase 2
Kinase 3
BASE
O
HO
HOBASE
O
HO
BASE O
HO
BASE
= Phosphate
X
Y
X X
Y Y
X
Y
The triphosphate metabolite of a nucleoside is the pharmacologically active agent A nucleoside must be a substrate for each of the kinases needed to produce the active triphosphate metabolite The first phosphorylation event is the most substrate selective. The triphosphate must be a substrate for the viral polymerase and it must get incorporated into the growing RNA or DNA chain – eliciting a chain termination event
Polymerase
O O OO O O
OOOO O
Ade CytUraGua
3'
5'
AdeUraGuaAde UraUraAde
O
HO
BASE
O
Ade
ChainTermination
Y
XHO HO HO HO HO
OHOHOHOHOHOHOH
RNA
O
HO
OBASE
XY
PO
O-OP
O
O-OP
O-
-OO
Poor Drug-like Characteristics of Nucleos(t)ides
• Nucleosides: – Highly polar – Require multiple enzyme mediated steps for activation – Can be enzymatically degraded – nucleosidases &
nucleobase metabolizing enzymes • Nucleotides
– Highly polar – Chemically unstable – Require enzyme mediated activation – Can be enzymatically degraded – nucleosidases,
nucleobase metabolizing enzymes and phosphatases
Ideal Characteristics of a Nucleos(t)ide Prodrug for the Treatment of Hepatitis
• Must have sufficient chemical stability and physical properties to be formulated for oral administration
• Must be stable to conditions of the gastrointestinal tract such that the prodrug reaches the site of absorption intact
• Prodrug must have good absorption properties • Prodrug must not undergo appreciable enzymatic degradation
during the absorption phase • Nucleotides: Once absorbed the prodrug must have sufficient
stability in the blood in order to reach the target organ • Nucleotides: The prodrug must be transported into hepatocytes
and release the free 5’-phosphorylated nucleoside • Achieving a high liver-plasma ratio (liver targeting) is desirable. • The intact prodrug must be biologically inert • The promoiety byproducts should be non-toxic and rapidly
cleared.
Issues Addressed by Nucleoside Prodrugs
• Poor oral bioavailability • Formation of inactive metabolites
O
YHO
BaseHOZ X
Nucleoside Prodrugs: NM283 (Valopicitabine)
O
OHHO
NHON
O
NH2
NM107(EC50 = 1.23 µM)
O
OHO
NHON
O
NH2
O
NH2
NM 283 Valopicitabine
Issues • Poor oral bioavailability in rat • Formation of inactive uridine metabolite
Results • Leveraged peptide transporter • %F = 33% (rat) • No detection of uridine metabolite • Human studies
• 800 mg BID, 14 days gave -1.2 log10 IU/mL reduction in viral load • 800 mg BID + SOC, 28 days ->4 log10 IU/mL reduction in viral load • terminated for GI tox Pierra, C, et al, JMed Chem, 2006, 49, 6614
Nucleoside Prodrugs: RG7128 (Mericitabine)
O
FHO
NHON
O
NH2
PSI-6130 (EC90 = 4.5 µM)
O
FO
NON
O
NH2O
O
RG7128 Mericitabine
Issues • Modest oral bioavailability in humans (~25%) • Formation of inactive uridine metabolite in significant amounts • Liver to plasma ratio ~1
Results • Human oral bioavailablity increased to ~75% • Reduction in formation of uridine metabolite • No improvement in liver to plasma ratio • Human studies
• 1500 mg BID, 14 days, -2.7 log10 IU/mL reduction in viral load • 1000 mg BID + SOC, 28 days, -5 log10 IU/mL reduction in viral load
Sofia,M., et al., J. Med. Chem., 2012, 55, 2481
Nucleoside Prodrugs: R1626 (Balapiravir)
O
OHHO
NHON
O
NH2
N3
O
OO
NON
O
NH2
N3
O
O
O
R1479 (EC50 = 1.28 µM)
R1626 Balapiravir
Issues • Poor animal oral bioavailability • Poor exposure in humans
Results • %F >90% dog • >5-fold increase in oral bioavailability in rat and monkey • Human studies
•4500 mg BID, 14 days, -3.7 log10 reduction in viral load • 1500 mg BID + SOC, 28 days, -5.2 log10 reduction in viral load 81% RVR • Terminated do to hematological tox
Toniutto, P., et al., IDRUGS, 2008, 11, 738
Nucleoside Prodrugs: TMC647078
O
R2O
NR1ON
O
NH2
TMC647078 (EC50 = 7.3 µM)
O
HO
NHON
O
NH2
Issues • Exposure in animals poor • Formation of inactive uridine metabolite
OR1 and/or R2 =
Results • 3’-Monoester 10-fold increase in animal exposure • Diester 24-fold increase in animal exposure • Liver to plasma ratio ~1
Jonckers, T.H., et al., J. Med. Chem., 2010, 53, 8150
Issues Addressed by Nucleotide Prodrugs
O
YHO
BaseOZ X
PO
-OO-
• Highly polar – do not penetrate biological membranes • Chemically unstable • Enzymatically degraded
Kinase Bypass
OBaseHO
X
YOH
OBaseO
X
YOH
PO
HOOH
OBaseO
X
YOH
PO
OOH
PO
OHHO
OBaseO
X
YOH
PO
OOH
PO
OHOP
O
OHHO
OBaseO
X
YOH
PO
X Bypass the non-productive phosphorylation step
Phosphoramidates
PO
O
NH
OH
O R2
Ar
PO
O
HNR2
O
PO
O
NH
OH
O R2
H
PO
O
HO
H
OPO
O
NH
OR1
O R2
Ar
Nucleoside NucleosideO
NucleosideO
NucleosideO NucleosideO
Carboxyesterase
orCathepsin A
HINT-1
H2Ospontaneous
HepDirect
NucleosideOOP
OO
Ar cytochrome P450NucleosideO
OP
OO
Ar OH
spontaneous NucleosideOOP
HOO
Ar
O
spontaneous
NucleosideOOHPHOO
Ar
O
Designed for liver targeting
SATE
NucleosideOPO
OO
S
S
R
R
O
O
NucleosideOPO
OO
S
HS
R
O
esterase
SNucleosideOP
OO
HO
SR
O
S
esterase
NucleosideOPO
HOHO
Cyclic Phosphates
O
X
YO
BASEO
PO
O
O
X
YO
BASEO
PO
NH
EtO O
O
X
YO
BASEO
PO
OSO
O
n-Pr
O
X
YO
BASEO
PO
O
O
X
YO
BASEO
PO
HO
CYP450
CYP3A4
PDE
O
X
YHO
BASEOPO
OHHO
Double Prodrugs
O
YHOX
NO
N
NN
OR
NH2
PO
O
YHOX
NO
N
NN
OR
NH2
PO
HOOH
O
YHOX
NO
N
NNH
NH2
PO
HOOH
ORemoval of phosphate
promoiety ADAL1
Improves overall lipophilicity thus improving cellular uptake
Murakami, E., et al., J. Med. Chem., 2011, 54, 5902
Nucleotide Phosphoramidates: GS-7977
O
FHO
NONH
O
O
PO
ONHO
O
GS-7977 (PSI-7977) EC90 = 0.42 µM)
O
FHO
NHONH
O
O
PSI-6206
Issues • Nucleoside inactive in replicon • TP potent inhibitor of NS5B • Long TP half-life in hepatocytes • Not a substrate for DCK • MP readily converted to TP
Results • Potent inhibitor of HCV replication • High TP levels in hepatocytes in vitro • High TP levels in liver on oral admin. • High liver to plasma ratio – leverages first pass metabolism for liver targeting • Human studies
• Safe and well tolerated to 24 wks • 400 mg QD, 14 days,~5.0 log10 IU/mL reduction in viral load • + RBV, GT2,3 100% SVR12 • +Daclatasvir, GT1, 100% SVR12 Sofia, M.J., et al., J. Med. Chem., 2010, 53, 7202.
Lam, A., et al., ACC, 2010, 54, 3187.
Nucleotide Phosphoramidates: PSI-353661
O
FHO
NHO
N
NNH
O
NH2
O
FHO
NO
N
NN
OMe
NH2
PO
ONHO
O
PSI-353661 (EC90 = 0.008 µM)
(EC90 = 69.2 µM)
Issues • Weak replicon potency • TP is potent inhibitor of NS5B, IC50 = 5.9 µM • Polar molecule • Weak kinase substrate
Results • Potent inhibitor of HCV replication • Conversion to the G-MP derivative occurs readily in hepatocytes • Produced high levels of G-TP in human hepatocytes • Produced a favorable ~5:1 liver to plasma ratio in vivo - leverages first pass metabolism for liver targeting
Chang, W., et al., ACS Med Chem Lett., 2011,2, 130 Furman, P., et al., Antiviral Res., 2011, 91, 120
Nucleotide Phosphoramidates: INX-8189
O
OHHO
NO
N
NN
OCH3
NH2
PO
OHN
O
O
CH3
INX-8189 (EC50 = 0.01 µM)
O
OHHO
NHO
N
NNH
O
NH2
(EC50 = 3.5 µM)
Issues • Low levels of TP detected in cells • G-TP potent inhibitor: IC50 = 0.13 µM • Natural guanosine derivative poor cellular uptake
Results • > 100-fold improvement in replicon potency • Conversion to the G-MP derivative occurs readily in hepatocytes • Substantial levels of G-TP produced in cells. • Human studies
• Monotherapy: 25 mg QD, -1.3 log10 IU/mL decline in viral load
Vernachio, J.H., et al., ACC, 2011, 55, 1843
Nucleotide Phosphoramidates: IDX184
O
OHHO
NO
N
NNH
O
NH2
PO
OHN
SO
OHIDX184
(EC50 = 0.4 µM)
O
OHHO
NHO
N
NNH
O
NH2
(EC50 = 3.5 µM)
Issues • Low levels of TP detected in cells • G-TP potent inhibitor: IC50 = 0.13 µM • Natural guanosine derivative poor cellular uptake
Results • P450 dependent and independent cleavage • High liver TP levels observed in cynomolgus monkeys on PO dosing • 10 mg/kg PO dose to HCV infected chimpanzees resulted in -2.3 log10 IU/mL decline in viral load after 3 days • Human studies
• 25-100mg QD , 3days, max -0.74 log10 IU/mL decline in viral load • 50 – 200 mg QD + SOC, 14 days, -2.7 to -4.1 log10 decline in viral load
Zhou, X.J., et al., AAC, 2011, 55, 76
Cyclic Phosphates: PSI-352938
O
FO
NO
N
NN
OEt
NH2PO
O
PSI-352938 (EC90 = 1.37 µM)
O
FHO
NHO
N
NNH
O
NH2
(EC90 = 69.2 µM)
Issues • Weak replicon potency • TP is potent inhibitor of NS5B, IC50 = 5.9 µM • Polar molecule • Weak kinase substrate
Results • High live G-TP levels in rat and dog after PO dosing • Observe circulating levels of intact prodrug • CYP3A4 mediated cleavage mech. • Human studies
• 100 – 300 mg QD, 7 days, -4.0 -4.6 log10 IU/mL decline in viral load • 300 mg, QD, +SOC, 14 days gave -5.5 log10 IU/mL viral load decline • Clinical hold – liver enzyme elevation Reddy, G.P., et al., BMCL, 2010, 20, 7376
Lam, A., et al., J. Virol., 2011, 85, 12334
HepDirect:
O
OHHO
NHON
O
NH2
O
OHHO
NOPO
O
O
F
F N
NH2
O
(EC50 = 1.23 µM)
Issues • Poor oral bioavailability in rat • Low levels of TP after i.p. dosing • Formation of inactive uridine metabolite
Results • 10-fold increase in TP levels on i.p. dosing • HCV-infected chimpanzees
• 10 mg/kg QD, -1.3 to -1.5 log10 IU/mL decline in viral load vs -1.0 log10 IU/mL decline with 16 mg/kg NM283
Carroll, S.S., et al., AAC, 2011, 55, 3854
Other Nucleotide Prodrugs
O
OHHO
NON
O
NH2
N3
PO
ONH
CH3
O
OPhO
OHHO
NONH
O
O
N3
PO
ONHO
OPh
Sofia, M.J., et al., J. Med. Chem., 2012, 55, 2481
O
OHHO
NON
O
NH2
PO
ONHO
O O
XY
NOPO
O
O
S
S
O
O
N
NH2
O
O
OHO
NON
O
NH2
PO
NH
EtO O
O
OHO
NO
N
NN
N
PO
O
HN SO2CH3
SO
O
n-Pr
O
OO
NO
N
NN
NH2
PO
O
O
Cl
O
N
N N
N
NH2
O P
HO
O
OHO (CH2)2O(CH2)17CH3
Developing a Prodrug: In Vitro and In Vivo Characterization
• Physico-chemical characterization – solubility, chemical stability, and lipophilicity
• Hepatic/microsomal/tissue homogenate/ intestinal fluid/ plasma stability
• Caco-2 and PAMPA permeability • Absorption mechanism(s) • Are the active drug or prodrug substrates for efflux
pumps? • Understand the disposition of both the active parent
and prodrug compounds
Summary
The application of a prodrug strategy is a well established approach to improve the physicochemical, biopharmaceutical or pharmacokinetic properties of a pharmacologically potent compound increasing its developability. In no therapeutic field has the application of prodrug technology seen as wide and impactful an application as has been seen in the development of nucleoside and nucleotide HCV therapies
References
• Prodrugs: Challenges and Rewards, Parts I and II, Biotechnology: Pharmaceutical Aspects, Vol 5, Editors: Stella, V., et al., Publisher: Springer
• Prodrugs: Design and Clinical Applications, Rautio, J., et al., Nature Reviews Drug Discovery, 2008, 7, 255.
• Prodrugs: Some Thoughts and Current Issues, Stella, V., J. Pharma. Sci., 2010, 99, 4755.
• Lessons Learned from Marketed and Investigational Prodrugs, Ettmayer, P., et al., J. Med. Chem., 2004, 47, 2393.
• Prodrugs of Phosphates and Phosphonates, Hecker, S., et al., J. Med. Chem., 2008, 51, 2328.
• Nucleotide Prodrugs for HCV Therapy, Sofia, M.J., Antiviral Chem and Chemother., 2011, 22, 23.
• Nucleoside, Nucleotide, and Non-Nucleoside Inhibitors of Hepatitis C Virus NS5B RNA-Dependent RNA-Polymerase, Sofia, M.J., et al., J. Med. Chem., 2012, 55, 2481.