Download Pigmented Rabbit Ocular PK Formulation Comparison Study

Comparison of AzaSite® and
Azithromycin 1% for Bacterial
Conjunctivitis
Christopher Crean1, Jason L. Vittitow1, Richard C. Zink1, Lori Richards1,
Rozemarijn S. Verhoeven1, Kendall D. Powell2 and RK Brazzell1
1 Inspire
Pharmaceuticals, Inc., Durham, NC
2 Enthalpy Analytical, Inc. Durham, NC
Research Sponsored by:
Purpose
To study the ocular distribution of 1 % azithromycin in
an aqueous formulation and 1 % azithromycin in
DuraSite® (AzaSite®).
To compare the tissue distribution and
pharmacokinetics of AzaSite® after a single
administration and after the approved seven-day
course treatment.
2
Methods
This study was designed as a terminal sample collection study in pigmented rabbits. At the indicated
times after drug administration, 40 µL of PBS were applied to the ocular surface of anesthetized
animals and 20µL was collected for estimation of tear drug concentrations. Ocular tissues were
removed in the following order: inferior and superior bulbar conjunctiva, inferior and superior
palpebral conjunctivas followed by cornea and eyelids. Samples were analyzed by LC/MS/MS.
Single administration:
Day 1
2
3
4
5
6
7
Sampling Time Period
Sampling times: 0, 0.083, 0.25, 0.5, 1, 2, 4, 8, 12, 24, 36, 48, 72, 96, 120 and 144 h
Multiple administration:
Day 1
2
3
4
5
6
7
8
9
10
11
12
13
Sampling Time Period
Sampling times: Days 1-6 : Predose (0), 0.5, 1 h
Day 7: 0, 0.083, 0.25, 0.5, 1, 2, 4, 8, 12, 24, 36, 48, 72, 96, 120 and 144 h
3
The azithromycin conjunctiva and cornea tissue concentrations are
higher for the mucoadhesive formulation.
Results
Eye Drop
Conjunctiva
Azithromycin ng/g
1,000,000
AzaSite
Aqueous Azithromycin
100,000
10,000
1,000
100
10
0
2
4
6
8 10 12 14 16 18 20 22 24
48
72
96
120
144
Time (hr)
Eye Drop
(…) The dotted line in the graph
represents the MIC50 of
azithromycin for typical
bacterial pathogens.
Cornea
1,000,000
Azithromycin ng/g
Azasite
Aqueous Azithromycin
100,000
10,000
1,000
100
10
0
2
4
6
8 10 12 14 16 18 20 22 24
48
Concentration of azithromycin
(mean ± SE) in conjunctiva
and cornea is plotted on a loglinear graph in nanograms of
drug per gram of tissue over
time, where 1000 nanograms is
equal to 1 microgram. The
conjunctiva plotted in this graph
is an average plot of the four
conjunctiva quadrants.
72
96
Time (hr)
4
120
144
Results
Seven days of treatment with azithromycin in DuraSite increases the
observed conjunctiva and cornea tissue concentrations.
Eye Drop
Conjunctiva
Azithromycin ng/g
1,000,000
AzaSite Day 1
Azasite Day 7
100,000
10,000
1,000
100
0
2
4
6
8 10 12 14 16 18 20 22 24
48
72
96
120
144
Time (hr)
Eye Drop
(…) The dotted line in the graph
represents the MIC50 of
azithromycin for typical bacterial
pathogens.
Azasite Day 1
Cornea
Azasite Day 7
Azithromycin ng/g
1,000,000
100,000
10,000
1,000
100
10
0
2
4
6
8 10 12 14 16 18 20 22 24
48
72
96
Concentration of azithromycin
(mean ± SE) in conjunctiva and
cornea is plotted on a log-linear
graph in nanograms of drug per
gram of tissue over time, where
1000 nanograms is equal to 1
microgram. The conjunctiva
plotted in this graph is an average
plot of the four conjunctiva
quadrants.
120
144
Time (hr)
5
Results
The azithromycin conjunctiva tissue concentrations reach a stable
minimum concentration by day 2.
AzaSite BID Days 1-2, QD Days 3-7
250,000
Azithromycin ng/g
200,000
Conjunctiva
150,000
MIC50 of azithromycin for
typical bacterial pathogens
100,000
Minimum observed
concentration at steady state
50,000
} 15 fold
0
0
24
48
72
96
120
144
168
192
216
Eye Drop
Application
Time (hr)
Concentration of azithromycin (mean ± SE) in conjunctiva is plotted in nanograms of drug per
gram of tissue over time, where 1000 nanograms is equal to 1 microgram. The conjunctiva
plotted in this graph is an average plot of the four conjunctiva quadrants. Azithromycin reaches
steady state tissue levels by day 2 that are a minimum of 15 fold higher than the typical minimum
inhibitory concentration (MIC50) level for bacterial pathogens. Azithromycin is cleared from
conjunctiva with a 40 hour average half-life.
6
240
264
288
Results
The azithromycin cornea tissue concentrations reach a stable
minimum concentration by the end of day 2.
AzaSite BID Days 1-2, QD Days 3-7
500,000
Azithromycin ng/g
400,000
Cornea
300,000
MIC50 of azithromycin for
typical bacterial pathogens
Minimum observed
concentration at steady state
200,000
100,000
}
0
Eye Drop
Application
0
24
48
30 fold
72
96
120
144
168
192
216
Time (hr)
Concentration of azithromycin (mean ± SE) in cornea is plotted in nanograms of drug per gram of
tissue over time, where 1000 nanograms is equal to 1 microgram. Azithromycin reaches steady
state tissue levels by day 2 that are a minimum of 30 fold higher than the typical minimum
inhibitory concentration (MIC50) level for bacterial pathogens. Azithromycin is cleared from cornea
with a 43 hour average half-life.
7
240
264
288
Results
The azithromycin eye lid tissue concentrations reach a stable
minimum concentration by the start of day 3.
AzaSite BID Days 1-2, QD Days 3-7
Azithromycin ng/g
300,000
Eye Lids
200,000
MIC50 of azithromycin for
typical bacterial pathogens
Minimum observed
concentration at steady state
100,000
}
0
Eye Drop
Application
0
24
48
72
40 fold
96
120
144
168
192
216
240
Time (hr)
Concentration of azithromycin (mean ± SE) in eye lids is plotted in nanograms of drug per gram
of tissue over time, where 1000 nanograms is equal to 1 microgram. Azithromycin reaches steady
state tissue levels by day 2 that are a minimum of 40 fold higher than the typical minimum
inhibitory concentration (MIC50) level for bacterial pathogens. Azithromycin is cleared from eye
lids with a 125 hour average half-life.
8
264
288
Results
Azithromycin resides on the ocular surface at higher levels and for a
longer period of time when formulated in DuraSite®.
AzaSite BID Days 1-2, QD Days 3-7
10,000
1,000
Azithromycin g/mL
Tears
100
MIC50 of azithromycin for
typical bacterial pathogens
10
1
Tears
0.1
0.01
Eye Drop
Application
0
24
48
72
96
120
144
168
192
216
Time (hr)
Concentration of azithromycin (mean ± SE) in tears is plotted in micrograms per milliliter.
9
240
264
288
Azithromycin Calculated Peak and Total Concentration Levels
AUC (h*ng/g or h*ng/mL)
AUC(t0-tlast)
Single Dose Azithromycin
Single Dose AzaSite
Multi Dose Azasite
10 7
10 6
10 5
s
a
C
on
ju
Ey
e
nc
lid
tiv
a
ne
or
C
Te
ar
10 4
Cmax
Single Dose Azithromycin
Single Dose AzaSite
Multi Dose Azasite
10 6
10 5
10 4
lid
Ey
e
C
on
j
s
a
un
ct
iv
or
ne
a
C
r
10 3
Te
a
Cm ax (ng/g or ng/mL)
10 7
10
Total exposure is reported as
mean±SE area under the curve
(AUC) in h*ng/g for the tissues and
h*ng/mL for tear in log format for
visual purposes; while peak
concentration is reported in
mean±SE ng/g for the tissues and
ng/mL for tear. The reported AUC
values for the single dose studies
are from 0-144h while the multi
dose AUC values are after the last
eye drop from 144-288h. Peak
concentrations (Cmax) are reported
after the initial eye drop from the
single dose studies and the last eye
drop (9th) on Day 7 for the multi
dose study. Within a given tissue
there was no appreciable difference
between the different quadrants of
conjunctiva or the inferior or
superior eye lids. The eye lids were
not analyzed during the single dose
studies. All tissues exhibit higher
concentrations of azithromycin in a
mucoadhesive formulation and after
multiple administrations.
Pharmacokinetic Parameters of Azithromycin
Treatment
Group
1%
Azithromycin
with DuraSite®
(AzaSite®)
Single Eye Drop
1%
Azithromycin
without
DuraSite®
Single Eye Drop
1%
Azithromycin
with DuraSite®
(AzaSite®)
Multiple Eye
Drops
a
Tissue Type
AUC0-t
(h*ng/g)a
Cmax (ng/g)a
Tmax (h)
T ½ (h)
CL/F (g/h)a
Vz/F (g)a
Cornea
837,319
40,425
0.08
67
0.28
27.17
Average
Conjunctiva
737,717
108,356
0.08
63
0.39
33.83
Tears
3,016,257
10,539,048
0.08
15
0.1
2.22
Cornea
196,486
8,775
0.08
91
0.90
118.31
Average
Conjunctiva
79,227
9,547
0.875
48
4.09
272.66
Tears
154,750
893,482
0.08
37
193
102.11
Cornea
16,402,463
252,000
1.0
43.3
0.015
0.935
Averaged
Conjunctiva
3,101,108
177,394
0.08
40
0.091
4.315
Tears
392,021
1,217,500
0.08
78.7
0.754
85.6
Average Eye
Lids
8,146,912
180,300
0.25
125
0.014
4.74
For tears and aqueous humor, units for Cmax (ng/mL), AUC0-t (h*ng/mL), CL/F (mL/h) and Vz/F (mL) were different
Noncompartmental pharmacokinetic parameters for the different matrices were calculated with the WinNonlin
pharmacokinetic software from Pharsight, Inc., utilizing the sparse sampling option. The time course of analysis varied
depending on the treatment paradigm; single administration time period was from 0- 144 h, while the multi-dose period was
evaluated from 144-288 h post initial eye drop or 0-144 h after the last eye drop.
11
Conclusions
Azithromycin differs pharmacokinetically from a number of antibiotics in its
ability to achieve and maintain high tissue concentrations after a single
and multiple eye drops and sustain the exposure within the tissue for
some time after the last eye drop due to its extended half-life.
Pharmacodynamically it differs from other macrolide antibiotics (e.g.
erythromycin) in its ability to kill both gram positive and negative bacteria.
Also, potential clinical effectiveness of azithromycin is further enhanced
by its formulation within Durasite®, which increases tissue exposure 5-18
fold over a non-mucoadhesive formulation.
This data provides evidence that azithromycin administered from a
mucoadhesive formulation has levels at steady state;
• in conjunctiva at least 15-fold above the average MIC,
• in cornea at least 30-fold above the average MIC,
• in eye lids at least 40-fold above the average MIC.
• persists in all tissues analyzed 6 days after the last eye drop
12