Correlation of human and biorelevant media solubility for poorly soluble compounds
Patrick AugustijnsNottingham, September 12 2012
Low solubility: a growing problem
More lipophilic
- HTS
- Combinatorial chemistry
- More lipophilic targets
More solubility issues
- Higher risk of failing during early or late development.- Appropriate solubility measurements necessary
06/06/2012
Pharmaceutical profiling - Solubility
3
Van Herwaarden et al., Trends Pharmacol. Sci. 2009
Disintegration
Dissolution
Permeation
SolubilityAbsorption
Overview
• Intestinal drug behavior
Solubility
Supersaturation
Definition
• Solubility: The maximum quantity of a substance that can be completely dissolved in a given amount of solvent.
– Solvent used and its composition– Temperature– pH– Degradation– Solid state (polymorph type)– Aggregation– Counter ions (salts)– Ionic strength
Determination of thermodynamic solubility
• Thermodynamic solubility (equilibrium solubility):
– starting with crystalline material– equilibrium with solid compounds (time consuming)– 24-48h timescale
C
t
Disruption of Dissolution
crystal lattice
Determination of kinetic solubility
• Kinetic Solubility: precipitation from organic solvent
- Appropriate up to lead optimization- Solubility data are time-dependent- Likely to overpredict thermodynamic solubility
Supersaturation
Precipitation
C
t
- DMSO
- Ethanol
- Dimethoxyethane
06/06/2012
Thermodynamic solubility• Crystalline material
• Equilibrium with solid form
• Shake flask method
Pharmaceutical profiling - Solubility
Put excess ofcompound in Eppendorftube
Add solvent Shake for 24hat 37°C
Centrifugefor 15min
at14000rpm
Analysesupernatans
withHPLC
8
C
t
Key questions
• Which solvent systems?
0
1
2
3
4
5
6
7
8
9
10
µM
Phosphate Buffer pH = 6
SGF
pH = 1
FaSSIFFeSSIF
Fasted HIF
Solubility of Itraconazole in different solvent systems
Biorelevance of solubility assays
in vivo in vitro
BS
PL + LP DD
Intraluminal conditionsIntroduction
fasted ↔ fed state:
1)pH (↓)
2)Bile salts (↑)
3)Biliary Phospholipids (↑)
4)Dietary digestion products
• Lipolytic digestion products
=> Mixed micelles
Collection of HIF
Aspiration state (duodenum)
Fasted Fed Fat-enriched fed
Administration of liquid meal
250 ml water 400 ml Ensure Plus+ 250 ml water
300 ml Scandishake Mix+ 350 ml water
Fat content 30% 46%
Energy content 600 kcal 600 kcal
Collection period 2h 5h 5h
Used for:
characterizationin vitro solubility
0 30 60 90 120 150 180 210 240 270 3002
4
6
8
10
collection time (min)
pH
pH
fasted
fat-enriched fed
fed
0 30 60 90 120 150 180 210 240 270 3002
4
6
8
10
collection time (min)
pH
0 30 60 90 1202
4
6
8
10
S1S2S3S4S5
collection time (min)
pH
Bile salts
0 30 60 90 1200
5
10
15
20
25
30
35
S1S2S3S4S5
collection time (min)
tota
l bile
sal
t con
c (m
M)
0 30 60 90 120 150 180 210 240 270 3000
5
10
15
20
25
30
35
collection time (min)
tota
l bile
sal
t con
c (m
M)
0 30 60 90 120 150 180 210 240 270 3000
5
10
15
20
25
30
35
collection time (min)
tota
l bile
sal
t con
c (m
M)
fasted
fat-enriched fed
fed
Phospholipids
0 30 60 90 1200
2
4
6
8
10
S1S2S3S4S5
collection time (min)
tota
l pho
spho
lipid
con
c (m
M)
0 30 60 90 120 150 180 210 240 270 3000
2
4
6
8
10
collection time (min)
tota
l pho
spho
lipid
con
c (m
M)
0 30 60 90 120 150 180 210 240 270 3000
2
4
6
8
10
collection time (min)
tota
l pho
spho
lipid
con
c (m
M)
fasted
fat-enriched fed
fed
0 30 60 90 120 150 180 210 240 270 3000
5000
10000
15000
time (min)
solu
bilit
y (µ
M)
Solubility indomethacin
fasted
fat-enriched fed
fed
0 30 60 90 1200
5000
10000
15000
S1S2S3S4S5
time (min)
solu
bilit
y (µ
M)
0 30 60 90 120 150 180 210 240 270 3000
5000
10000
15000
time (min)
solu
bilit
y (µ
M)
Indomethacin: acid, pKa = 4.6, logP = 3.4
Correlations - pH
• Indomethacin (acid, pKa = 4.6, logP = 3.4)
0 2 4 6 80
1
2
3
4
5
fastedfedenriched
pH
log
(sol
ubili
ty) (
µM)
pKa
R² = 0.86
Correlations – BS, PL
• Danazol (neutral, logP = 4.2)
0 2 4 6 8 100
100
200
300
400
phospholipids (mM)
solu
bilit
y (µ
M)
0 10 20 30 400
100
200
300
400
fastedfedenriched
bile salts (mM)
solu
bilit
y (µ
M)
Correlations – LP
• Diazepam (basic, pKa = 3.3, logP = 2.9)
0 5 10 15 200
2000
4000
6000
8000
fastedfedenriched
lipolytic products (mg/ml)
solu
bilit
y (µ
M)
Biorelevant Conditions
• in-vitro surrogate medium
Composition Fassif Updated Fassif FessifNa Taurocholate (mM) 3 3 15Lecithine (mM) 0.75 0.2 3.75pH 6.5 6.5 5
Maleic Acid (mM) / 19.12 /
Sodium Hydroxide (mM) 8.7 34.8 101
Sodium Chloride (mM) 105.7 68.62 203
Osmolality (mosmol/kg) 270 180 ± 10 670
Intestinal drug solubility
solubility of 17 model drugs withdifferent physicochemical characteristics were investigated:
-1.0 -0.5 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0
ritonavirprednisolone
glipizide
furosemide
cinnarizine
sulfasalazine
carbamazepine
danazol
quinidine
diazepam
indomethacin
loviride
glibenclamide
fenofibrate
nifedipine
ketoconazole
hydrochlorothiazide
logP
in: - HIF- SIF- d-α-tocopheryl polyethyleneglycol 1000 succinate (TPGS)in phosphate buffer
Mean solubility in fasted HIF versus FaSSIF
y = 5,4x0,79
R2 = 0,91
1
10
100
1000
10000
1 10 100 1000 10000
solu
bilty
in fa
sted
HIF
(µM
)
solubility in FaSSIFc (µM)
Fold error between solubility in FaSSIF, corrected solubility in FaSSIF and solubility in fed HIF.
Mean solubility in fed HIF versus FeSSIF
y = 4,3x0,86
R2 = 0,86
1
10
100
1000
10000
1 10 100 1000 10000
solu
bilit
y in
fed
HIF
(µM
)
solubility in FeSSIFc (µM)
Fold error between solubility in FeSSIF, corrected solubility in FeSSIFc and solubility in fed HIF.
y = 5,2x0,79
R2 = 0,84
1
10
100
1000
10000
1 10 100 1000 10000
solubility in 0,1% TPGS (µM)
solu
bilit
y in
fast
ed H
IF (µ
M)
y = 0,098x1,20
R2 = 0,84
1
10
100
1000
10000
1 10 100 1000 10000
solubility in 2% TPGS (µM)so
lubi
lity
in fe
d H
IF (µ
M)
Overview
• Intestinal drug behavior
Solubility
Supersaturation
Beyond solubility: supersaturation
Supersaturation refers to a solution that contains more of the dissolved material than could be dissolved by the solvent under normal circumstances.
Concept of supersaturation
Con
cent
ratio
n
Time
Equilibrium solubility
Supersaturation zone
PH-Shift method
• Only useful for basic compounds (80%)
Stomach Low pH
GI Tract
Good Solubilitythrough ionization
+H3N
Poor solubilitydue to neutralization
NH2
Similar results compared to solvent shift method for itraconazole
Stomach Low pH
Wat
erba
th @
37°C
Test Medium
Super-saturated
Concentrated drug solution
0 30 60 90 1200
5
10
15
20
25
Time (min)
Degr
ee o
f su
pers
atur
atio
n (D
S)
medium test in solubilityt
tCDS =
Supersaturation Assay
5 Poorly soluble model drugs
Initial DS of 20
Itraconazole Supersaturation in FaSSIF
Supersaturation profile of Itraconazole
0,02,04,06,08,0
10,012,014,016,018,020,0
0 50 100 150 200 250 300Time (min)
Deg
ree
of s
uper
satu
ratio
n Fassif
Itraconazole Supersaturation in FeSSIF
Supersaturation profile of Itraconazole
0,0
5,0
10,0
15,0
20,0
25,0
0 50 100 150 200 250 300Time (min)
Deg
ree
of s
uper
satu
ratio
n
FessifFassif
Human intestinal Fluids?
What about supersaturation in human intestinal fluids ?
Itraconazole supersaturation in Fasted State Human Intestinal Fluids
Fasted HIF V1
0,0
2,0
4,0
6,0
8,0
10,0
12,0
14,0
0 20 40 60 80 100 120 140
Time (min)
Deg
ree
of S
uper
satu
ratio
n
Itraconazole supersaturation in Fed State Human Intestinal Fluids
Fed HIF V3
0,0
1,0
2,0
3,0
4,0
5,0
6,0
7,0
8,0
9,0
10,0
0 20 40 60 80 100 120
time (min)
Deg
ree
of s
uper
satu
ratio
n
0 20 40 60 80 100 1200
2
4
6
8
10
12
14
16
18
20
22
A
B
C
Time (min)
Degr
ee o
f sup
ersa
tura
tion
factoration Supersatur 2h-0 ABA+
=
DS 1 (solubility)
Impact of medium on precipitation
Etravirine Ritonavir Loviride Danazol Fenofibrate0
1
2
3
4
5
6
7
8
9
10
11
12 FaHIF
Supe
rsat
urat
ion
Fact
or0-
2h
Increasing solubilityBevernage et al, 2010, J Pha Sci 99:4525
Impact of medium on precipitation
Etravirine Ritonavir Loviride Danazol Fenofibrate0
1
2
3
4
5
6
7
8
9
10
11
12 FaHIF FeHIF
Supe
rsat
urat
ion
Fact
or0-
2h
Bevernage et al, 2010, J Pha Sci 99:4525
Impact of medium on precipitation
Etravirine Ritonavir Loviride Danazol Fenofibrate0
1
2
3
4
5
6
7
8
9
10
11
12 FaHIFFaSSIF
FeHIFFeSSIF
Supe
rsat
urat
ion
Fact
or0-
2h
Bevernage et al, 2010, J Pha Sci 99:4525
Impact of medium on precipitation
Etravirine Ritonavir Loviride Danazol Fenofibrate0
1
2
3
4
5
6
7
8
9
10
11
12 FaHIFFaSSIF
FeHIFFeSSIF
Buffer
Supe
rsat
urat
ion
Fact
or0-
2h
Bevernage et al, 2010, J Pha Sci 99:4525
Impact of medium on precipitation
Influence solubility on supersaturation
f
0
5
10
15
20
25
0 50 100 150 200 250 300Time (min)
Deg
ree
of s
uper
satu
ratio
n
ItraconazoleEtravirineLoviride
0,00
1,00
2,00
3,00
4,00
5,00
6,00
7,00
8,00
9,00
10,00
1
ItraconazoleEtravirineLoviride
Absorption assessment: Caco-2
OMS formulation (1)
crystalline itraconazole (2)
(1)
(2)
In vivo exposure (rats)
Itraconazole
0 4 8 12 16 20 240
400
800
Time (hrs)
Plas
ma
conc
entr
atio
n (n
M)
OH-itraconazole
0 4 8 12 16 20 240
400
800
1200
Time (hrs)Pl
asm
a co
ncen
trat
ion
(nM
) Crystalline ITZITZ:SBA-15 1:4Sporanox
Precipitation inhibition in human gastric fluid: excipient effects
Con
cent
ratio
n
Time
solubility
+ excipient
HPMC E5
Eudragit® E-PO
PVP K25
Effect of HPMC on ITZ precipitation in FaSSIF (pH 6.5)
3 µg/ml
0 60 120 180 2400
1
2
3 FaSSIFFaSSIF + 0.05% HPMC
Time (min)
Con
c (µ
g/m
l)
In vivo exposureEffect of HPMC
Itraconazole
0 4 8 12 16 20 240
400
800
Time (hrs)
Plas
ma
conc
entr
atio
n (n
M)
OH-itraconazole
0 4 8 12 16 20 240
400
800
1200
Time (hrs)
Plas
ma
conc
entr
atio
n (n
M) Crystalline
ITZ:SBA-15 1:4SporanoxITZ:SBA-15:HPMC 1:4:1.5ITZ:SBA-15:HPMC 1:4:6
Con
cent
ratio
n
Time
Intestine
Con
cent
ratio
n
Time
Plasma
solubility
Solubility-limited absorptionJ = P ⋅ C
Con
cent
ratio
n
Time
Intestine
Con
cent
ratio
n
Time
Plasma
solubility
Supersaturation J = P ⋅ C
Con
cent
ratio
n
Time
Intestine
Con
cent
ratio
n
Time
Plasma
solubility
Supersaturation + precipitationJ = P ⋅ C
Con
cent
ratio
n
Time
Intestine
Con
cent
ratio
n
Time
Plasma
solubility
Supersaturation + precipitation:low permeability J = P ⋅ C
Con
cent
ratio
n
Time
Intestine
Con
cent
ratio
n
Time
Plasma
solubility
J = P ⋅ CSupersaturation + precipitation:high permeability
06/06/2012 54
Solu
bilit
y(µ
M)
Solubility profiling of HIV-PI
55
Solubility profiling of HIV-PI
56
Solubility profiling of HIV-PI
Prediction of food-effects
57
Solubility profiling of HIV-PI
Conclusions
• High intra/inter subject variability
• FaSSIF & FeSSIF: adequate solvent systems for:• Solubility estimation• Supersaturation evaluation• Prediction of food effects
Acknowledgements
• Laboratory for Pharmacotechnology & Biopharmacy, KU Leuven
Jan BevernageMichiel Van SpeybroeckNico HolmstockRandy MellaertsJoachim Brouwers
• Center for Gastroenterology, University Hospitals LeuvenRita Vos, Jan Tack
• FundingResearch Foundation Flanders (‘FWO’); Agency for Innovation by
Science and Technology (‘IWT’); KU Leuven
Overview
• Intestinal drug behavior
Solubility
Supersaturation
• Gastric drug behavior:
Case study posaconazole
Posaconazole in the lab
• XLogP = 4.6• aqueous solubilty < 1 µg/ml• high permeability• BCS II• weak base (pKa 3.6 and 4.6)
Posaconazole: influence of gastric conditions
Posaconazole pharmacokinetics and gastrointestinal behavior
– crossover study (5 healthy volunteers)
– oral administration of a single dose (400 mg) of Noxafil®
– posaconazole concentration-time assessment
• plasma
• gastric fluid
• intestinal fluid
– 4 different conditions
• water
• PPI / water
• PPI / cola
• cola
Posaconazole & PPI
Stomach
0 30 60 90 120 150 180 210 240 270 3000
2
4
6
8
Water PPI - Water
Time (min)
pH
0 30 60 90 120 150 180 210 240 270 3000
100
200
300
400
500
Time (min)
Posa
cona
zole
(µM
)
Posaconazole & PPI
Plasma
0 6 12 18 24 30 36 42 480.00
0.05
0.10
0.15
0.20 WaterPPI - Water
Time (h)
Posa
cona
zole
(µM
)
Posaconazole & PPI & Cola
Stomach
0 30 60 90 120 150 180 210 240 270 3000
2
4
6
8
Water PPI - Water PPI - Cola
Time (min)
pH
0 30 60 90 120 150 180 210 240 270 3000
100
200
300
400
500
Time (min)
Posa
cona
zole
(µM
)
Posaconazole & PPI & Cola
Plasma
0 6 12 18 24 30 36 42 480.00
0.05
0.10
0.15
0.20 WaterPPI - WaterPPI - Cola
Time (h)
Posa
cona
zole
(µM
)
Posaconazole & Cola
Plasma
0 6 12 18 24 30 36 42 480.0
0.1
0.2
0.3
0.4 WaterCola
Time (h)
Posa
cona
zole
(µM
)
Posaconazole & Cola
Stomach
0 30 60 90 120 150 180 210 240 270 3000
2
4
6
8
Water Cola
Time (min)
pH
0 30 60 90 120 150 180 210 240 270 3000
200
400
600
800
1000
Time (min)
Posa
cona
zole
(µM
)
Posaconazole & Cola: effects of cola
Solubilization
0 30 60 90 120 150 180 210 240 270 3000
200
400
600
800
1000WaterCola
Time (min)
Posa
cona
zole
(µM
)
pH 1.2 pH 2.5 pH 6.50
20
40
60800
1600
2400 ColaSGFFaSSIF
Posa
cona
zole
sol
ubili
ty (µ
M)
Increased gastric dissolution time
normal gastric pH: dissolution-limited absorption
Summary: gastric conditions & fasted state posaconazole absorption
• Gastric dissolution affects posaconazole absorption
• Clinically: coadminister posaconazole with cola– with PPI: partially restored fasted state absorption– without PPI: improved fasted state absorption
• Posaconazole formulation improvements are desirable– dissolution rate / solubility
• Mechanistically: effect of cola– pH-dependent increase in solubility (in case of abnormal gastric pH
levels)– pH-independent increase in solubility– increased gastric dissolution time
y = 64,4x0,55
R2 = 0,75
1
10
100
1000
10000
0,1 1 10 100 1000 10000
solubility in blank FeSSIF (µM)so
lubi
lity
in fe
d H
IF (µ
M)
y = 34,3x0,56
R2 = 0,81
1
10
100
1000
10000
0,1 1 10 100 1000 10000
solubility in blank FaSSIF (µM)
solu
bilit
y in
fast
ed H
IF (µ
M)
Conclusions
• High intra/inter subject variability
• FaSSIF & FeSSIF: adequate solvent systems for:• Solubility estimation• Supersaturation evaluation
• Conditioning environment of the stomach
Preparation room Waiting room