amsterdam, the netherlands - apr 2013 1presidio ... · amsterdam, the netherlands - apr 2013...
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PPI-668 Dose
Bas
elin
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% Q
54H
Bas
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e !0
0% R
30Q
+L31
I+Y9
3H
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31M
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54D
/N
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L28M R30Q L31M L31V L31I P58S Y93H Y93C
0 hr 24 hr 72 hr Day 17
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N = 10
NS5A Resistance Substitutions
0102030405060708090
100
M28T M28V Q30R Q30H Q30E L31M L31V H58D H58N H58P Y93H Y93C Y93N
0 hr 24 hr 72 hr Day 17
Perc
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N = 22
NS5A Resistance Substitutions
VAST MAJORITY OF DETECTED NS5A RESISTANT VARIANTS ARE NOT AMPLIFIED IN HCV PATIENTS DURING 3-DAY MONOTHERAPY WITH THE OPTIMIZED NS5A INHIBITOR PPI-668
Qi Huang1, Ningwu Huang1, Anja Huq1, Miriam Lau1, Eric Peng1, Jay Lalezari2, Geoff Farrell3, Purvee Shah4, Eric Lawitz5, Pamela Vig1, Nathaniel Brown1 and Richard Colonno1
1Presidio Pharmaceuticals, San Francisco, CA, USA; 2Quest Clinical Research and UCSF, San Francisco, CA, USA; 3The Canberra Hospital, Canberra, Australia; 4West Coast Clinical Trials, Costa Mesa, CA, USA; and 5Alamo Medical Research, San Antonio, TX, USA
BACKGROUND: HCV NS5A inhibitor PPI-668 was assessed in a 3-day Phase 1b, placebo controlled, monotherapy trial in HCV gt-1 patients using QD oral doses of 40, 80, 160 and 240 mg, with 8/10 patients receiving PPI-668 per cohort. Mean maximal viral load reductions of 3.54-3.75 log10 IU/mL were observed with doses ≥80 mg QD. METHODS: Comprehensive monitoring of HCV resistant variants was performed during dosing and post-dosing periods. HCV RNA was extracted from patient serum samples, RT-PCR amplified and the resulting DNA subjected to population sequencing. Clonal sequencing was performed to determine if detected substitutions were genetically linked. PPI-668 susceptibility was assessed in transient transfection assays using HCV replicons encoding specific substitutions or population NS5A gene inserts from clinical samples. RESULTS: Known NS5A resistance substitutions at residues 28, 30, 31, 58 and 93 were detected early during PPI-668 monotherapy. Among the 40 enrolled gt-1 patients, eight had detectable NS5A resistance substitutions at baseline. One patient (gt-1b 240 mg) was a non-responder exhibiting high level resistance at baseline, with 100% of his circulating virus encoding genetically linked R30Q+L31I+Y93H substitutions. The other seven PPI-668 treated patients with baseline resistance substitutions responded well (RNA reductions of 2.23 to 3.95 log10 IU/mL). Resistance substitutions became detectable in all but one PPI-668 treated patient within 24-48 hr, as WT virus was rapidly eliminated. Importantly, these observed substitutions were not further amplified with continued monotherapy, suggesting that PPI-668 concentrations were sufficient to suppress these single-substitution variants. Susceptibility (EC90) of replicons encoding resistance substitutions detected in PPI-668 treated patient samples were generally at or below Cmin levels, confirming the advantageous PK profile of PPI-668 and its ability to cover single-substitution resistant variants. No significant differences were observed in the overall resistance patterns across the four PPI-668 treated gt-1 cohorts. CONCLUSIONS: NS5A resistance variants frequently pre-exist among HCV patients, emphasizing the need for combination therapy and use of optimized NS5A inhibitors, such as PPI-668, that achieve plasma/liver levels high enough to suppress single substitution HCV variants. Further studies of PPI-668 in combination with other DAAs are warranted.
Emerging Substitutions
Activity in Patients with Baseline Resistance
Summary
1192 48th Annual Mtg of the European
Association for the Study of the Liver Amsterdam, The Netherlands - Apr 2013
Improved Potency and Lead Optimization Abstract
Introduction
PPI-668 is a potent and selective HCV NS5A inhibitor, with pan-genotypic activity in replicon assays and an optimized PK profile
Phase 1b study was conducted in treatment-naïve HCV-g1 patients – Three-day treatment period with 40, 80, 160 or 240 mg of PPI-668 QD – Well tolerated at all dose levels, no AEs attributed to PPI-668 – Predictable and consistent PK profile in humans, with steady-state reached after a single dose – Rapid, marked HCV RNA responses seen in 31/32 HCV gt-1 patients – Nearly equivalent activity observed in both HCV gt-1a and HCV gt-1b patients
Comprehensive analysis performed on serum samples from all patients HCV RNA was RT PCR-amplified from patients’ samples and genotyped (population) at
Baseline and at several subsequent treatment time points Clonal sequencing was performed on patient samples when multiple substitutions were
present to determine if they were genetically linked Phenotypes (EC90) determined for individual and linked substitutions emerging on therapy
HCV Genotype
Dose (mg QD)
Maximum HCV RNA Reduction (Log10 IU/mL) Mean Range
gt-1a (n=7) 40 3.27 2.81 – 3.86 gt-1b (n=1) 40 2.89 Only 1 patient gt-1a (n=5) 80 3.26 2.82 – 3.96 gt-1b (n=3) 80 4.00 3.95 – 4.12 gt-1a (n=6) 160 3.33 2.23 – 3.69 gt-1b (n=2) 160 3.93 3.47 – 4.39 gt-1a (n=4) 240 3.71 3.14 – 4.33
gt-1b (n=3)* 240 3.80 3.71 – 3.83
Resistance Monitoring
*Excludes a baseline resistant patient with 100% of his circulating virus encoding 3 genetically-linked substitutions (R30Q+L31I+Y93H )
Individual Patients - Antiviral Efficacy and Genotypic Monitoring for Resistance Substitutions
PPI-668 was well tolerated and exhibited strong antiviral potency in a Phase 1b clinical trial in gt-1 patients involving 3-day QD monotherapy with 40 mg, 80 mg, 160 mg or 240 mg (AASLD 2012)
The optimized PK properties of PPI-668 enabled all seven patients with single resistance substitutions at Baseline to respond well to treatment
One gt-1b patient (240 mg), harboring a homogeneous population of virus containing 3 linked NS5A resistance substitutions, was highly resistant to PPI-668 and was the only non-responder (0.33 log10 IU/mL)
With WT virus rapidly eliminated, low level pre-existing resistant variants at Baseline became detectable in 31/32 patients, but did not show a significant increase on treatment, indicative of inhibition during the treatment period
Despite the appearance and enrichment of resistance substitutions by 24 -48 hr, a subsequent decline of ~0.4 log10 IU/mL in viral RNA levels was observed in patients treated with 160 mg and 240 mg PPI-668
No significant differences observed in resistance frequency or degree across the PPI-668 treatment cohorts evaluated
Like other classes of DAAs, combination therapy will be required to suppress the emergence of resistance
Resistance substitutions were present at Baseline in eight PPI-668 treated patients and became detectable within 24-48 hr in all patients except patient 2-29 (gt-1b 240 mg), who had no evidence of resistant substitutions With the exception of patient 2-30 (gt-1b 240 mg, with 100% of his Baseline circulating virus encoding genetically linked R30Q-L31I-Y93H substitutions), all patients exhibited 2.2 to 4.4 log10 IU/mL drops in their HCV RNA levels Despite the rapid appearance of resistance substitutions, patients in the 160 mg and 240 mg cohorts experienced a further mean decline of 0.41 and 0.36 log10 IU/mL, respectively, in viral RNA levels between 24 and 72 hr on therapy
Genotype Patient Dose (mg)
Baseline Substitutions (%)
RNA Reduction (log10 IU/mL)
gt-1b 2-47 40 R30Q (100%) 2.89 gt-1b 2-08 80 L31M (100%) 3.95 gt-1a 2-09 80 M28T (9%) 2.82 gt-1a 2-11 80 M28T (6%), L31M (11%) 3.56 gt-1b 2-26 160 R30Q (84%), L31M (25%) 3.47 gt-1a 2-36 160 H58D (69%), H58N (31%) 2.23 gt-1b 6-06 240 P58S (79%) 3.82 gt-1b 2-30 240 Linked R30Q+L31I+Y93H (100%) 0.33
PPI-668 was active in 7/8 gt-1 patients with pre-existing single NS5A substitutions at amino acid residues 28, 30, 31 and 58, with HCV RNA declines of 2.23-3.95 log10 IU/mL
HCV gt-1a
HCV gt-1b
Resistance substitutions observed at a high frequency in HCV gt-1a patients include M28T, Q30R, Q30H, L31M, Y93H and Y93C
High frequency of resistance substitutions L31M and Y93H observed in HCV gt-1b patients
Apart from the HCV gt-1a Y93H substitution, primary resistance substitutions tend to remain in circulation 2 wk post-treatment
Substitution Frequencies During Dosing
Once detectable following elimination of WT HCV population, the vast majority of resistant variants encoding single substitutions show no significant increase in population percentage during continued treatment with PPI-668
No obvious correlation with dose level, as all PPI-668 doses appear to provide exposure levels (peak and trough) needed to constrain expansion of single substitution variants
Phenotypes (EC90) of substitutions enriched on therapy were at (gt-1a) or below (gt-1b) PPI-668 plasma trough levels (blue bars)
Single substitutions Q30E and Y93H/N in gt-1a and Y93H in gt-1b confer the highest levels of resistance in each genotype
Ability to constrain replication of single substitution variants during monotherapy appears to be driven by the relatively high peak and trough levels achieved with all PPI-668 doses
Percent of Linked Substitutions
Cmin and Resistant Phenotypes
Clonal analysis (n=100) of 72 hr patient samples was performed to determine the frequency of genetically linked substitutions among amino acid residues 28, 30, 31, 58 and 93
Overall, percentage of linked substitutions was low at 72 hr if substitutions were not pre-existing at Baseline
No obvious trend observed over this relatively short dosing period (3 days) in patients without baseline substitutions
Resistant variants encoding double/triple linked substitutions likely needed to escape formidable PPI-668 concentrations (Cmax and Cmin) established with 160 mg and 240 mg doses
72 hr Samples
gt-1b
gt-1a
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1001a - M28T (EC90 19 nM)
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% o
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1a - Q30R (EC90 30 nM) 1a - Q30H (EC90 22 nM)40 mg80 mg160 mg240 mg
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24 48 72Treatment Hours
1a - Y93H (EC90 190 nM) 1a - Y93C (EC90 23 nM)40 mg80 mg160 mg240 mg
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1b - L31M (EC90 0.57 nM)
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1b - Y93H (EC90 4.28 nM)
% o
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0.01
0.1
1
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40 m
g80
mg
160
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240
mg
40 m
g80
mg
160
mg
240
mg
M28
TM
28V
Q30
RQ
30H
Q30
EL3
1ML3
1VH
58D
H58
NH
58P
Y93H
Y93C
Y93N
L28M
R30
QL3
1ML3
1VL3
1IP5
8SY9
3HY9
3CTr
iple
PPI-6
68 C
once
ntra
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(nM
)
HCV gt-1a Replicons - EC90Plasma Cmax Plasma Cmin HCV gt-1b Replicons - EC90
30Q
+31I
+93H
L28R
30QL31
Y93
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
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0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1b
Day 17
P5824 hr
2-47
48 hrDay 17
M28T
Q30R
/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
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0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a8-07
24 hr L31M
Y93C/N
H58 M
28Q
30R/H
L31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
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100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
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Day 17
8-10
24 hr
H58 M
28TQ
30RL31
Y93C
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
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% S
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0
3.0
4.0
5.0
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7.0
2.00 hr
1a8-14
24 hrDay 17
H58 M
28TQ
30R/H
L31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
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80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-44
H5824 hr M
28TQ
30R/H
L31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-52
24 hr
H58 M
28TQ
30R/H
L31M
Y93C
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
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100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-53
24 hr
H58 M
28T/V
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
80
100
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-54
24 hr Q30R
/HL31M
/V
Y93
0H58
L28R
30L31
Y93H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
80
100
0
4.0
5.0
6.0
7.0
8.0
3.00 hr
1b
Day 17
2-03
24 hr
P58 L28R
30L31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
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0
3.0
4.0
5.0
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7.0
2.00 hr
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Day 17
2-08
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P58 L28R
30L31M
Y93H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
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% S
ubst
itutio
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Pop
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4.0
5.0
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8.0
3.00 hr
1b
Day 17
2-17
24 hr
P58 M28T/V
Q30R
/HL31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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0
4.0
5.0
6.0
7.0
8.0
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Day 17
2-02
24 hr
H58P M
28TQ
30R/H
L31M
Y93C
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
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0
4.0
5.0
6.0
7.0
8.0
3.00 hr
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Day 17
2-05
24 hr
H58P M
28TQ
30R/H
L31M
Y93C/N
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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0
3.0
4.0
5.0
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7.0
2.00 hr
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Day 17
2-09
H5824 hr M
28TQ
30R/H
L31M
Y93C
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
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Pop
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0
4.0
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6.0
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8.0
3.00 hr
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Day 17
2-11
24 hr
H58
L28R
30L31
Y93H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1b
Day 17
2-20
24 hr
P58 L28MR
30QL31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
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0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1b2-26
24 hrDay 17
P58 M28T
Q30R
/HL31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
ion
20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
6-01
24 hr
H58D
/N
M28T
Q30R
/HL31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
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100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-18
24 hr
H58 M
28TQ
30R/H
L31
Y93C
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
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100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-21
24 hr
H58 M
28TQ
30R/H
L31M
Y93H/C
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
6 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
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20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-27
24 hr
H58 M
28TQ
30RL31M
Y93H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
6 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
80
100
0
4.0
5.0
6.0
7.0
8.0
3.00 hr
1a
Day 17
2-36
24 hr H58D
/N
M28T
Q30R
/HL31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
6 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
80
100
0
4.0
5.0
6.0
7.0
8.0
3.00 hr
1a
Day 17
2-40
24 hr
H58
L28R
30L31M
/V
Y93H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1b
Day 17
6-06
24 hr
P58S L28R
30L31
Y93
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1b
Day 17
2-29
P5824 hr L28R
30QL31I
Y93H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
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20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1b
Day 17
2-30
P5824 hr L28R
30L31V
Y93H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
ion
20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1b
Day 17
8-05
P5824 hr M28T
Q30R
/H/K
L31M
Y93C/N
/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
ion
20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
6-05
H5824 hr M
28TQ
30R/H
/EL31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
ion
20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-41
H58D
/P
24 hr M28T
Q30R
/H/E
L31M
Y93C/H
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
ion
20
40
60
80
100
0
4.0
5.0
6.0
7.0
8.0
3.00 hr
1a
Day 17
2-43
H5824 hr M
28Q
30R/H
/EL31
Y93C
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
ion
20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
8-21
H5824 hr
M28T
Q30R
/HL31M
Y93C/N
HC
V R
NA
Leve
ls (L
og10
IU/m
L)
8 hr 48 hr 72 hr
% S
ubst
itutio
ns in
Pop
ulat
ion
20
40
60
80
100
0
3.0
4.0
5.0
6.0
7.0
2.00 hr
1a
Day 17
2-15
24 hr
H58
40 m
g Co
hort
16
0 m
g Co
hort
80
mg
Coho
rt
240
mg
Coho
rt