Transcript Transmitted drug resistance (Pat Cane)

Transmitted drug resistance
Pat Cane
Questions
• What is the level of TDR and is it
changing?
• Are we measuring TDR accurately?
• Are more sensitive methods of detecting
TDR useful?
• What are the consequences of TDR?
ART status by year of sample
12000
Number of tests
10000
8000
6000
4000
2000
0
1997-9
2000
2001
2002
2003
2004
2005
2006
Year of sample
naive
experienced
unclassified
2007
2008
Linking to HPA databases
(Sam Lattimore’s work)
•
64,888 records in HIVDR database had sufficient information for linkage
–
–
–
–
•
•
•
Clinic ID & Region
Soundex, DOB & region
Clinic ID (modified), site, DOB
Clinic ID alone
40,609 (62.6%) linked to new dx database
48,826 (75.3%) linked to SOPHID (accessing treatment + care)
Overall, 51,033 records linked to either new dx or SOPHID
New dx
2,207
SOPHID & New dx
38,402
SOPHID
10,424
Prevalence of HIV drug resistance in
ART-experienced patients (IAS 2009)
Percentage of tests
80
60
NRTI
PI
nNRTI
any class
40
20
0
2000
2001
2002
2003
2004
2005
Year of sample
2006
2007
2008
Prevalence of HIV drug resistance in
ART-naive patients (WHO list)
14
Percentage of tests
12
10
NRTI
PI
nNRTI
any class
8
6
4
2
0
2000
2001
2002
2003
2004
2005
Year of sample
2006
2007
2008
Misclassification of TDR
(Hannah Castro’s work)
• Estimates of TDR will be distorted if some ARTexperienced patients are misclassified as ART-naïve
• Aim: to quantify the potential extent of misclassification
of treatment status bias
• The effect of misclassification depends on the number of
naïve tests relative to the number of experienced tests,
as well as the rate of resistance
• Explore possibility of distinguishing primary (in ARTnaïve patients) and secondary (in ART-experienced
patients) resistance based on patterns of mutations.
.1
0
.02 .04 .06 .08
Rate of THDR
.12 .14 .16
Trend in TDR over time in UK
1997
1998
1999
2000
2001
2002
2003
2004
2005
2006
Year
Observed THDR
Adjusted THDR m = 0.05
Adjusted THDR m = 0.02
Adjusted THDR m = 0.07
Number of
observed naïve tests
326
365
409
537
622
617
994
1867
3268
3520
Number of
observed experienced tests
113
214
568
847
819
1004
1163
1253
1550
1712
Ratio of observed naive to
experienced tests
2.88
1.71
0.72
0.63
0.77
0.61
0.85
1.49
2.11
2.06
Effect of TDR on pre-therapy viral load
(Linda Harrison’s work)
• In vitro studies indicate that most drug
resistance mutations reduce replication
capacity of HIV
• Patients with TDR might be expected to
have lower HIV RNA viral load (VL) than
patients infected with wild-type virus
• Epidemiological studies to date have been
inconclusive (patients with TDR: n=9-77)
Viral load by class of TDR
Group
No. (%)
Mean log10 VL
(SD)
Adjusted*
difference
No TDR
7285 (91)
4.60 (0.82)
REF
Any TDR
709 (9)
4.58 (0.83)
-0.04
0.12
350 (4)
164 (2)
90 (1)
105 (1)
4.60 (0.85)
4.59 (0.86)
4.71 (0.75)
4.44 (0.86)
-0.05
0.00
0.09
-0.21
0.21
0.96
0.27
0.004
NRTI only
NNRTI only
PI only
≥2 classes
P
Adjusted for: CD4 count, viral subtype, ethnicity, exposure group,
sex, age, calendar year, clinical centre, VL assay
NRTI
215 variants (n=250)
M41L (n=152)
K219N/Q/E/R (n=112)
D67N/G/E (n=73)
M184V/I (n=61*)
L210W (n=42)
T215Y/F (n=42)
K70R/E (n=33)
other NRTI (n=46)
NNRTI
K103N/S (n=154)
Y181C/V (n=48)
G190A/E (n=37)
K101E/P (n=23)
other NNRTI (n=35)
PI
L90M (n=55)
M46L/I (n=48)
V82A/L/T/F (n=39)
other PI (n=67)
-.5
-.4
-.3 -.2 -.1 0
.1 .2 .3
Difference in HIV RNA log10(c/ml)
*55V, 4I, 2V/I mixture
.4
.5
Eurocord analysis of impact of TDR on
virological response up to 16 months
(Linda Wittcop)
TDR categories
(WHO and Stanford)
N*
%
2: at least one mutation of the WHO and resistant
to at least one of their prescribed drugs
(classified as 3,4,5 using Stanford)
477
4.6
1: at least one mutation of the WHO list but no
drug resistance to their prescribed regimen
(classified 1,2 using Stanford)
476
4.5
9505
90.9
0: no detected mutations of the WHO list 2009
* UK provided 22% of data
VF according to TDR categories
%
VF
months
Group 2: at least one mutation of WHO and receiving at least one resistant drug (3,4,5)
Group 1: at least one mutation of WHO but receiving a fully active treatment (1,2)
Group 0: no mutations on WHO list
Summary of Eurocord study
• Treatment response is poorer in patients having
transmitted rug resistance mutations only when the
ARV regimen is based upon drugs to which the virus
has lost susceptibility
– At least intermediate/low-level resistance to at least one
drug
– Gradient effect among intermediate/low (Stanford 3+4)
and fully resistant (Stanford 5) levels
– Less than three active drugs
• Having transmitted drug resistance mutations is not
predictive of failure when the regimen has not lost
susceptibility
– Exception patients receiving 2NRTI + 1NNRTI (just sig.)
Minority TDR testing
• Allele specific PCR and deep sequencing can detect
mutations present as minority populations.
CAVEATS!
• Sensitivity: only to ~1% reliably due to error rates in
enzymes used in testing process (Shafer et al.)
• Background incidence of each mutation in vivo (due to
inherent variability of quasispecies due to RT and RNA
polymerase errors) can be assumed to be about 0.010.1%.
• Sensitivity cannot exceed input number of molecules ie
need 1 ml with a viral load of ~10,000 cp/ml to get a
sensitivity of 1% due to inefficiencies of RNA extraction
etc.
Prevalence of drug resistance mutations in
undiagnosed MSM
14
12
10
8
Standard Genotyping
% Prevalence 6
minority species
4
2
0
-2
K103N
Y181C
M184V
Drug Resistance Mutation
K103N by 30% (p=0.25, 95% CI: 1.2-67%)
M184V by 13-fold (p=0.0005, 95% CI: 2-85-fold increase)
(Buckton et al.)
SENSE Study
Tim Conibear’s (RF) data
Study Design
• Several study sites with patients in Europe and Russia
• Baseline plasma screened for drug resistance mutations by
population sequencing – all positive patients excluded
• 157 ART drug naïve HIV-1 infected patients eligible for study
• Randomised into EFV or ETV + 2 NRTIs
• Low-frequency drug resistance mutations to be detected in:
– plasma RNA by sensitive real-time PCR (CDC method)
Target codons: 100I, 101E, 103N, 181C, 184V, 188L, and
190A
– matched PBMC DNA pol gene sequences using both bulk and
low-frequency assay
SENSE Study: Results
Results from sensitive mutation detection (detection limit ranges: 0.1-10% mutant virus)
•
•
•
•
HIV-1
Subtype
n
157
100I
101E
103N
181C
184V
188L
190A
B
99
0
2
1
0
1
0
1
Non-B
58
N/A
N/A
0
1
3
N/A
0
%
0
2
1
0.6
2.5
0
0.6
≥1 drug resistance mutation (101E/103N/181C/184V/190A) in 5.1% (8/157) patients
One patient showed both Y181 and M184 mutation, and no patients showed 100I or 188L.
All borderline result amplicons also sequenced – all non-significant mutations
All PBMC samples sequenced. No significant mutations detected.
Conclusions
•
•
•
Population sequencing on PBMC does not appear to increase detection of resistance
relative to plasma sequencing
Clinical significance not yet known (study still blinded)
Cutoffs are optimised for surveillance. Adjustment for clinical significance considered.
CHAIN proposal
Clinical multicenter study
“Virological efficacy of first-line NNRTI-based
antiretroviral therapy in antiretroviral-naïve
subjects with minority HIV-1 mutants
resistant to reverse transcriptase inhibitors”
Roger Paredes, Karin Metzner
Objectives of the minority study
• To assess the risk of virological failure to first-line
NNRTI-including ART in antiretroviral naïve HIV-1infected subjects with pre-existing minority NNRTIresistant HIV-1 variants
• To establish a clinically relevant pre-ART threshold
of minority NNRTI-resistant HIV-1 variants
• To investigate if linkage of NNRTI-resistant
mutations improves the prediction of virological
failure in antiretroviral-naïve HIV-1-infected
subjects initiating NNRTI-based ART
Study design
• Case control study within observational multicohort studies with retrospective testing of
plasma samples collected prior to first-line ART
• To achieve 80% power 211 patients failing firstline NNRTI-containing ART and 633 matched
controls will be included
Inclusion criteria
• Adult patients (defined as age ≥ 18 years)
• Chronic HIV-1 infection
• Initiation of first-line cART with a NNRTI plus at least 2
NRTIs
• Available resistance test (population sequencing): No
evidence for NNRTI-resistance
• Available clinical data (e.g., CD4 count, viral load, ART
changes) during follow-up
• Pre-treatment plasma sample available for 454 ultra-deep
sequencing fulfilling the following criteria
– Collected within 6 months before ART initiation
– HIV-1 RNA levels ≥10.000 copies/mL plasma
– 1 mL plasma available for UDS testing
Definition of cases and controls
• Cases = Patients who experienced virological failure
• Controls = Patients of the same cohort/clinical center (i.e.,
cases extracted from EuroSIDA will be matched with
controls selected from EuroSIDA, etc) who did not
experienced virological failure
• Three controls per case will be identified
• At analysis time (i.e., the time point of failure (or matching
time for controls)) patients have to be still receiving a firstline NNRTI, while patients are allowed to switch NRTIs
Research questions
• What is the impact of M184V present as a minority on
treatment outcomes?
• Should treatment where any TDR detected by standard
methods include a boosted PI?
• How long does it take majority mutations to go to
minority?
• Can minority TDR be onward transmitted?
• If a significant proportion of treated patients who show
no resistance mutations when failing boosted PIs are
phenotypically resistant, will this resistance be
transmitted?