Transcript Resistance to HIV therapy

Antiretroviral Drug Resistance
Anna Maria Geretti
Host-related factors
•Adherence
•Tolerability
•Immunity
•Genetics
Drug-related factors
•Potency
•PK properties
•Genetic barrier
Virus-related factors
•Viral load
•Drug susceptibility
•Fitness
Persistent virus replication
during HAART
Drug pressure
Emergence
and evolution
of drug resistance
Attachment
Fusion
Release of RNA
Assembly
Reverse transcription
Integration
Transcription
Maturation and budding
Characteristics of HIV-1 infection
1. High virus replication rate (109-1010 virus particles / day)
2. Rapid virus clearance (T½ cells producing virus: <1 day;
T½ free virus in plasma: a few hrs)
3. Virus latency (1:106 resting CD4 T cells)
4. Chronic immune activation
5. CD4 T-cell depletion (108-109 cells lost daily)
6. Progressive immune deficient state
7. Continuous virus genetic evolution
Wong 1997; Chun 1997; Siliciano 2003; Strain 2003; Han 2007
Mechanisms of HIV genetic evolution
1. RT-driven mutagenesis
– Rate: ~1 wrong incorporation per genome round
– All possible point mutations generated daily
2. APOBEC-driven mutagenesis
– Deamination of cytosine residues in nascent DNA
– GA hypermutation
3. Recombination
– Rate: 7-30 per genome round
– Hybrid virus progeny produced from different strains
Dominant quasispecies
Escape
from pressure
Preserved
fitness
 rapid turnover
 rapid adaptation
Consequences of HIV genetic variability
 At the population level
 Continuous emergence of new variants
 At the patient level
 Escape from immune pressure
 Escape from drug pressure
 Increased fitness and pathogenicity
 Challenge for diagnostic and monitoring assays
Emergence and evolution of resistance
Emergence
Single mutant




Evolution
Double mutant
Triple mutant
Increasing number of mutations
Accumulation of mutations on the same viral genome
Initially reduced viral fitness
Compensatory changes restore fitness
Key principles of resistance
 Drug-resistant mutants are selected (not created)
by drug pressure if virological suppression is incomplete
 Ongoing virus replication under drug pressure leads
to the evolution of resistance and cross-resistance
 Resistant mutants often display reduced fitness
but compensatory changes emerge over time that
partially restore virus fitness
PCR
Viral gene (e.g., RT)
Sequencing
HIV RNA
Plasma
Mutations
RT M184V
Methionine  Valine
@ codon 184 of RT
ATG / AUG  GTG / GUG
PCR
Viral gene (e.g., RT)
HIV RNA
Plasma
Culture with
escalating drug
concentrations
Defective laboratory
HIV vector
(e.g., RT_)
Infectious HIV
Fold-changes in IC50
relative to wild-type
M184V = 100 FC for 3TC
Detection of resistant mutants
Mutation Frequency
100
10
1
Detected by
routine methods
Detected by
ultrasensitive
methods
0.1
0.01
0.001
Natural background
Low-frequency resistance in the FIRST study
Mutations
Resistance test
P
Standard
UDS
NNRTI
7%
15%
<0.001
NRTI
6%
14%
<0.001
PI
2%
5%
0.03
Any
14%
28%
<0.001
N=258
Risk of failure of first-line NNRTI-based ART in
patients with NNRTI resistance
 Bulk resistance: HR 12.4 [3.4-45.1]
 UDS resistance: HR 2.5 [1.2-5.4]
USD = Ultra deep sequencing
Siemen, JID 2009
Resistant
Drug
pressure
20-30%
Wild-type
Limit of detection
Key principles of resistance
 Once drug pressure is removed, resistant mutants
are outgrown by fitter wild-type virus and become
undetectable by routine tests
 Resistance test results obtained after therapy is
discontinued are not reliable
 Resistant mutants persist at low frequency in
plasma and are “archived” in latently infected cells
 Resistance is long-lasting
 Resistance test results must be interpreted in
the context of the patient’s treatment history
Ms S., 35 yr
Δ HIV+ Dec 1997
Resistant
Wild-type
M184V = 3TC, FTC
Y181C = NNRTIs
d4T
3TC
NVP
VL
d4T
3TC
NVP
M184V
100000
M184V
Y181C
Y181C
1000
<50
10
Dec-97 Jun-98 Dec-98 Jun-99 Dec-99 Jun-00 Dec-00 Jun-01 Dec-01 Jun-02 Dec-02
Transmitted drug resistance
Drug
pressure
Transmission
Stable after transmission
Gradual reversion over time, sometimes incomplete
Persistence at low frequency in plasma
Persistence in latently infected cells
Key concepts
Genetic barrier
Residual activity
Hypersusceptibility
Genetic barrier to resistance
Defined by:
Number of mutations required to compromise activity
Impact of each mutation on drug susceptibility
Interactions between mutations
Fitness cost of resistance
Drug concentration
Resistant
Wild-type
Genetic barrier – A simplified overview
Class
NRTIs
ARVs
Genetic barrier
ZDV/3TC, d4T/3TC
++
ABC/3TC, TDF/3TC
+
TDF/FTC
++
EFV, NVP, ETV, RPV
+
NNRTIs
ETV, (RPV)
+/++
PIs
Boosted
+++/++++
Fusion inhibitors
T20
+
CCR5 antagonists
MVC
++ (for R5 virus)
Integrase inhibitors
RAL, ELV
+
Common NRTI resistance patterns
NRTIs
Mutations
ZDV d4T
ABC ddI
TDF 3TC FTC
M184V
ZDV 3TC
d4T 3TC
TAMs
+ TAMs
d4T 3TC M184V
TDF 3TC
K65R
TDF FTC
M184V
ABC
3TC
L74V
K65R Y115F
TAMs = thymidine analogue mutations: M41L, D67N, K70R, L210W, T215Y/F, L219Q/E
Resistance with first-line HAART
3rd drug
Study
ART
wk Tests K65R L74V M184V TAMs
GS903 n=299
TDF 3TC EFV
48
29
24%
0
41%
0
EFVR 55%
GS934 n=244
TDF FTC EFV
96
14
0
0
14%
0
EFVR 71%
GS934 n=243
ZDV 3TC EFV
96
29
0
0
31%
3%
EFVR 62%
CNA30021 n=770 ABC 3TC EFV
48
38
3%
21%
47%
0
EFVR 58%
ABT418 n=190
TDF FTC LPV/r
96
23
0
0
17%
0
PIR 0
SOLO n=190
ABC 3TC FPV/r
48
32
0
0
12%
0
PIR 0
ARTEMIS n=343
TDF FTC DRV/r 96
31
0
0
6%
0
PIR 0
13
0
0
38%
0
RALR 33%
STARTMRK n=281 TDF FTC RAL
96
Margot, HIV Med 2006; Margot, JAIDS 2009; Moyle, JAIDS 2005; Molina, IAC 2004; Gathe, AIDS 2004;
Mills, AIDS 2009; Lennox, Lamcet 2009.
1st and 2nd generation NNRTIs
Nevirapine
Efavirenz
Etravirine
Major resistance mutations
L100I, K101E/P
K103N/S
V106A/M
E138K, V179F
Y181C/I/V, Y188L/H/C, G190A/S/E
F227C, M230L, K238T
Rilpivirine
Activity of ETV with a strong backbone
DUET studies: OBR (with DRV/r) + ETV or Placebo
Patients with VL <50 copies/ml at wk 48 (ITT-TLOVR)
Responders (%) ± 95% CI
100
ETV + OBR (n=599)
Placebo + OBR (n=604)
90
80
70
61%
60
50
40%
40
30
p<0.0001*
20
10
0
024
8 12 16 20 24
32
Time (weeks)
40
48
ART-experienced patients with documented NNRTI and PI resistance
Katlama, AIDS 2009
Activity of ETV with a weak backbone
Study TMC125-C227: 2 NRTIs + ETV or PI
ART-experienced, PI-naive patients with documented NNRTI resistance
Ruxrungtham, HIV Med 2008
The genetic barrier of PIs
in vitro
450
DRV (R41T, K70E)
400
350
TPV (L33V, M46L, V82T)
ATV (L10F, V32I, M46I, I62V, A71V, I84V, N88S)
300
LPV (L10F, L23I, M46I, I50V, I54V, L63P, V82A)
APV (L10F, V32I, L33F, M46I, I47V, I50V)
250
NFV (L10F, D30N, R41K, K45I, M46I, V77I, I84V, N88D)
SQV (G48V, A71V, G73S, I84V, L90M)
200
RTV (G16E, M46I, V82F, I84V)
150
100
50
0
0
100
300
500
700
900
1100
Time (days)
De Meyer, Antimicrob Agents Chemother 2005;
De Meyer, IHDRW 2006
Emergence of PI mutation with DRV/r vs LPV/r
TITAN study: OBR + DRV/r or LPV/r
TITAN 96 week analysis
p0.05*
VFs with developing
mutations (%)
40
p0.05*
24/72
(33%)
19/72
(26%)
30
20
DRV/r
LPV/r
6/39
(15%)
3/39
(8%)
10
0
Primary PI
mutations1
NRTI RAMs1
*Exact Chi-Squared Test; TITAN 96 week analysis
1Johnson
et al. Top HIV Med 2007;
ART-experienced, LPV- and DRV-naive patients
De Meyer, HIV-9 2008
Resistance as a continuum
Lower cut-off =
Level of resistance beyond
which response begins to fall off
Response
Upper cut-off =
Level of resistance beyond which
clinical response is lost
Zone of
intermediate
response
Resistance
How to calculate the GSS
Genotypic Susceptibility Score
 RT mutations: M41L T215Y K103N
 Protease mutations: None
 Regimen:
 Resistance:
 Score:
 GSS ?
TDF
3TC
LPV/r
Intermediate Susceptible Susceptible
0.5
1
1
Partial treatment interruption in patients
with resistance reveals residual activity
Week 2 change in VL
1.0
Change in 0.5
plasma
viral load
0.0
–0.5
NRTI
PI
NNRTI
T20
Discontinued treatment class
Deeks, CROI 2005
Mechanisms of NRTI resistance:
Primer unblocking

T215Y-mediated resistance

Hydrolytic removal of the chain-terminating
NRTI enables DNA synthesis to resume

The pyrophosphate donor in most cells is ATP

P
P
P
M184V antagonizes the process
P
 3TC partially restores susceptibility to ZDV,
d4T and TDF in the presence of TAMs
P
P
P
P
 3TC antagonizes the emergence of TAMs
P
P
P
P
P
P
P
P
Gotte, J Virol 2000
Key principles of resistance
 Resistance moves along a continuum and increasing
numbers of mutations lead to progressive loss of
responses
 Residual activity is possible despite the presence of
extensive resistance (best evidence for the NRTIs)
 Resistance carries a fitness cost that reduces viral
replication
 Antagonistic effects between mutations can
have beneficial effects
Clinical implications for patients
with treatment failure
 The likelihood of drug-resistance depends upon the
drug, the regimen and the level of adherence
 When selecting a new regimen, aim for a GSS ≥2
 Avoid functional monotherapy with drugs that have
a low genetic barrier
 If options are limited, exploit residual activity and
hypersusceptibility effects – continue the NRTIs
rather than stopping therapy
Are you curious about what to do when a
patient suddenly stops NNRTI-based ART?
What “undetectable” viral load really means?
How we can easily predict HIV-1 tropism?
Advanced Virology
Workshop
Thank you