1 hiv drug resistance training module 1: introduction to hiv drug resistance
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Topics
Impact of HIV Drug Resistance Factors that Influence Development of Drug
Resistance How to Minimize Drug Resistance How to Respond to Detection of Drug
Resistance Drug Resistance Assays
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Objectives
Understand importance of measuring drug resistance for large-scale treatment programs.
Identify factors that influence drug resistance.
Understand the methods available for the measuring HIV drug resistance.
Identify strategies for minimizing development of drug resistance.
Identify strategies for responding to detection of moderate to high levels of drug resistant HIV.
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impact of drug resistance
Why is it important to measure drug resistance?How does it impact the success of large-scale treatment programs?
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What is Antiviral Drug Resistance?
Drugs no longer block virus replication Cause:
– Mutations in the viral genome One or more:
– Specific for an antiviral drug OR – Affecting related drugs (cross-resistance)
How much resistance? Which drugs?– Depends on type and number of mutations
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Total: 33 million (30 – 36 million)
Western & Central Europe
730 000730 000[580 000 – 1.0 million][580 000 – 1.0 million]
Middle East & North Africa
380 000380 000[280 000 – 510 000][280 000 – 510 000]Sub-Saharan Africa
22.0 million22.0 million[20.5 – 23.6 million][20.5 – 23.6 million]
Eastern Europe & Central Asia
1.5 million 1.5 million [1.1 – 1.9 million][1.1 – 1.9 million]
South & South-East Asia
4.2 million4.2 million[3.5 – 5.3 million][3.5 – 5.3 million]Oceania
74 00074 000[66 000 – 93 000][66 000 – 93 000]
North America1.2 million
[760 000 – 2.0 million]
Latin America1.7 million1.7 million
[1.5 – 2.1 million][1.5 – 2.1 million]
East Asia740 000740 000
[480 000 – 1.1 million][480 000 – 1.1 million]Caribbean230 000
[210 000 – 270 000]
Adults and Children Estimated to be Living with HIV, 2007
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Towards Universal Access – Scaling up priority HIV/AIDS interventions in the health sector. WHO/UNAIDS/UNICEF, June 2008
Number of People Receiving ARV Therapy In Low- and Middle-income Countries 2002-
2007
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Towards Universal Access – Scaling up priority HIV/AIDS interventions in the health sector. WHO/UNAIDS/UNICEF, June 2008
Median Price (USD) of First-line ARV Regimens in Low-income Countries, 2004-
2007
51% 12% 14% 9%
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Towards Universal Access – Scaling up priority HIV/AIDS interventions in the health sector. WHO/UNAIDS/UNICEF, June 2008
Median Price (USD) of second-line ARV regimens in Low-income Countries, 2004-
2007
AZT/3TC/EFV AZT/3TC/NVP d4T/3TC/EFV
d4T/3TC/NVP
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HIV Drug Resistance is Inevitable
HIV DR is an inevitable consequence of ART, influenced by:– Ability of regimens to suppress replication
completely– Adherence and tolerability of regimens – "Genetic barrier" to resistance– Relative fitness of resistant variant(s)– Pharmacokinetics (IQ)– Availability/continuity of drug supply– Removal of barriers to access to care
Therefore, efforts to minimize HIVDR should be focused on these factors
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HIV DR Testing in Resource Rich Settings
Prevalence of HIVDR at baseline
Utility of resistance testing before
initiating therapy
Individualization of 1st line regimen
Resistance developing on therapy
Resistance testing before switching
therapy (SOC)
Individualization of 2nd line and subsequent
regimens
24+ drugs from 6 classes
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HIV DR Testing in Resource Limited Settings
Prevalence of HIVDR at baseline
Utility of resistance testing before
initiating therapy
Individualization of 1st line regimen
Resistance developing on therapy
Resistance testing before switching
therapy (SOC)
Individualization of 2nd line and subsequent
regimens
24+ drugs from 6 classes
~6 drugs from 3 classes
Resistance developing on therapy
Prevalence and patterns of resistance
in population
Determination of standard 2nd line
regimens
Prevalence of HIVDR at baseline
Will standard 1st line regimens be effective?
Determination of standard 1st line
regimen
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Drug Resistance and HIV
HIV… evolves rapidly within human body has a high replication rate has a high mutation rate
Resistant strains can emerge within days if
drug pressure is not sufficient to suppress replication.
Resistant strains persist indefinitely and can re-emerge if same drugs are stopped and restarted (even if they are not detected by standard resistance assays).
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Review
Why is it important to measure drug resistance?
How does it impact the success of large-scale treatment programs?
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factors that influence development of drug resistanceWhat regimens influence drug resistance?What patient factors influence drug resistance?What public health approaches influence drug resistance?
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In Which Conditions is DR More Likely?
Treatment with <3 drugs Inappropriate selection of drugs Adding one drug to a failing regimen Interruption of treatment (even for a few
days) Prolonging a failing regimen
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In Which Conditions is DR Less Likely?Medication Factors
All patients treated with 3 or more drugs Use of appropriate drug regimens Can reliably suppress HIV replication to
levels of <50 copies/ml Use of fixed-dose combinations to support
adherence
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In Which Conditions is DR Less Likely?Systems Factors
Limited number of regimens Trained personnel, low turnover Supervision and monitoring Adequate lab services Drug supply and delivery systems
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In Which Conditions is DR Less Likely?Patient Factors
Adherence to treatment regimen Avoiding interruption of treatment, even if
only a few days Regular follow-up (going to clinic) Staying on uninterrupted first-line ART as
long as possible
CONTINUITY!
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Programmatic Factors Affecting Patient Compliance
Cost of treatment to patient Distance patient must travel to get
treatment Supply interruptions Availability of second-line regimens for
patients whose first-line regimens fail Timing of use of second-line regimens
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Discussion
What regimens influence drug resistance? What patient factors influence drug
resistance? What public health approaches influence
drug resistance?
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Reflection
What regimens do we use in our country? What systematic and programmatic
challenges do we face?
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Elements of a National HIVDR Prevention and Assessment Strategy
A. Development of a national HIVDR Working Group, five year plan and budget
B. Regular assessment of HIVDR early warning indicators (EWI) from all ART sites (or representative sites)
C. Surveys to monitor HIVDR and associated factors in sentinel ART sites
D. HIVDR transmission threshold surveys in geographic areas where ART has been widespread for > 3 years
E. HIVDR database developmentF. Designation of an in-country or regional WHO-accredited
HIVDR genotyping laboratoryG. HIVDR minimization activities review and supportH. Preparation of annual HIVDR report and
recommendations; use of data for ART and planning
Bennett, Bertagnolio, Sutherland and Gilks, Antiviral Therapy 13 Suppl 2:1–13, 2008
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Database, analysis
Transmission of DR HIV to uninfected individuals
Threshold Surveys(Surveillance)
Genotyping Laboratory
DR HIV prevalence < or ≥ 5%, 15%
To which drugs?
*Surveys to monitor HIV DR prevention and associated factors in sentinel ARV treatment sites
Emergence of HIVDR in treated
patients
Sentinel ART Site Monitoring Surveys*
Genotyping, VL Laboratory
HIVDR Prevention at 12 months; prevalence and
patterns of DR; associated factors
ART site factors associated with minimizing
HIVDR
Early Warning Indicators
Areas to directly target for improvement
PUBLIC HEALTH ACTION
Non-Laboratory Data collection
Relationship of Different Assessment Factors
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General WHO Strategies (1 of 2)
Strengthen existing programmes that minimize HIVDR
Form national HIV drug resistance working group
Monitor Early Warning Indicators Survey to monitor HIVDR prevention and
associated factors in sentinel ART sites Watch for transmitted HIVDR in recently
infected individuals
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General Strategies (2 of 2)
Designate one or more genotyping labs for HIVDR surveillance and monitoring
Maintain a national database to hold the HIV sequences collected through surveillance, monitoring, and research projects
Produce an annual report summarizing results from all the above
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Strengthen Existing Programmes That Minimize HIVDR
Support for adherence and follow-up Removal of barriers to ART access Drug supply continuity at the individual,
ART site, and national levels
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Form National HIV Drug Resistance Working Group
Develop HIVDR prevention, surveillance and monitoring plan
Make evidence-based recommendations for HIVDR prevention
Includes epidemiologists, clinicians, researchers, and laboratorians
Collect indicators and implement surveys Develop evaluation strategies
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Monitor Early Warning Indicators
Routine collection of medical and pharmacy records
Monitor for factors associated with HIVDR prevention or emergence– Extent to which prescribing practices meet
national and international guidelines– % of patients still on first-line; % lost to follow-up– % patients with timely medication pick up and
clinical follow-up– Drug supply continuity at site– Adherence and viral load, if feasible
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HIVDR Early Warning Indicators (EWI)
Prescribing
practices
Proportion lost to follow-up during
the first 12 months of ART
Patient retention on first-line ART
On-time ARV drug pick up
ART appointment-keeping
Drug supply
continuity
Site-level ART Program
Function
Viral load suppressio
n @12 months
Pill count/ adherence
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HIVDR EWI Site-Based Report Example
Site
Months with no
ARV drug stockoutsTarget: 12
% appropriate Initial ART regimen
prescriptionsTarget: 100%
% starting first line ART lost to follow up at 12
monthsTarget: < 20%
% on ART keeping all clinical
appointments on time
Target: > 80%
% on ART picking up all ART drugs on
time Target: ≥ 90%
1 12 94/94 (100%) 4/96 (4%) 182/209 (87%) 184/192 (96%)
2 10 81/81 (100%) 9/74 (12%) 342/402 (85%) 176/220 (80%)
3 9 31/40 (78%) 12/37 (32%) 122/244 (50%) 144/206 (70%)
4 12 104/104 (100%) 10/99 (10%) 891/993 (90%) 483/508 (95%)
5 12 112/112(100%) 13/105 (12%) 262/305 (85%) 184/202 (91%)
6 11 98/101 (97%) 2/90 (2%) 416/442 (95%) 254/359 (71%)
7 12 98/98 (100%) 9/88 (10%) 602/683 (88%) 369/402 (95%)
8 12 203/203 (100%) 43 /195 (22%) 292/356 (82%) 254/284 (86%)
9 12 304/305 (99.7%) 117/260 (45%) 753/1506 (50%) 829/1202 (69%)
10...
12 94/94 (100%) 12/90 (13%) 271/305 (89%) 269/290 (93%)
152 12 33/33(100%) 4/31 (13%) 147/180 (82%) 143/159 (90%)
153 10 26/34 (76%) 7/35 (20%) 148/224 (66%) 129/182 (71%)
154 12 73/73(100%) 9/69 (16%) 178/203 (87% ) 146/154 (95%)
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For More Information
See Bennett, Bertagnolio, Sutherland and Gilks (2008). The World Health Organization’s global strategy for prevention and assessment of HIV drug resistance. Antiviral Therapy 13 Suppl 2:1-13.
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Implementation of the WHO HIVDR Strategy – June 2009
Implementation of at least 1 component of the HIVDR strategy in 41 countries
HIVDR transmission surveys completed in 11 countries
EWI piloted in 23 countries HIVDR training workshops:
– Africa (3), Asia, Caribbean 22 accredited HIVDR Genotyping labs HIVDR laboratory training package Annual external Quality Assurance (supported
by NIH, through the VQA)
Countries implementing or planning a national HIVDR strategy
Countries with a national HIVDR working group implementing > 1 assessment element as of 2/2009
Countries setting up a working group and planning to implement > 1 assessment element in 2009
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Other
Designate one or more genotyping labs for HIVDR surveillance and monitoring– The WHO offers accreditation, support, and
training through the WHO Global HIVDR laboratory network
Maintain a national database to hold the HIV sequences collected through surveillance, monitoring, and research projects
Produce an annual report summarizing results from all the above
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how to respond to detection of drug resistanceWhat can be done if widespread drug resistance is found?
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Response to Detection of Transmitted Drug Resistance
If the prevalence of transmitted resistance to important drugs or drug classes is 5-15% or > 15%, surveys should be repeated in the original areas annually and extended to additional areas
Consider changing standard 1st line regimen
Monitor Early Warning Indicators for clues about what can be improved at a programmatic level
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Response to Results from Monitoring Surveys
If the patterns of resistance indicate reduced susceptibility to 2nd line drugs, surveys should be repeated in the original areas annually and extended to additional areas.
Consider changing standard 2nd line regimen.
Monitor Early Warning Indicators for clues about what can be improved at a programmatic level.
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hiv drug resistance assays
What are the different types of resistance assays?What are the advantages and limitations of these assays?What results can we expect from these tests?What if the results from one type of test are inconsistent with results from another?
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Clinical Use of Resistance Data
Resistance tests are most accurate in assessing resistance to current regimen
Absence of resistance to previously used drug does not rule out reservoirs of resistant virus that might emerge after re-initiation of that drug
If resistance to given drug has ever been detected, that drug should probably not be used again, even if current test results suggest viral susceptibility
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Conditions for Drug Resistance Testing:Individual Patient Management
When to use:– When there are treatment options– To indicate drug exposure– While patient is on therapy or before starting for
the 1st time What to consider:
– Treatment history, other reasons for therapy failure
– Lab-to-lab variability of results and interpretations– Requires expert advice for optimal use– Only test single agents, not combinations
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Conditions for Drug Resistance Testing: Public Health Applications
When to use:– When making decisions for national treatment
guidelines• 1st line regimens (based on results of surveillance
for transmitted drug resistant HIV)• 2nd line regimens (based on patterns of resistance
in treated patients)– To monitor program effectiveness minimizing
HIVDR What to consider:
– Early Warning Indicators, programme and site factors
– Treatment history, other reasons for therapy failure
– Lab-to-lab variability of results and interpretations
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Limitations of Resistance Testing:
Only possible in case of a detectable viral load
Lack of consistent quality control (see ENVA)
Current assays do not pick up “minority species;” information is given on predominant strain
Assays have mostly been studied in “late” failures; what is their value in early failures?
Susceptibility is not equal to activity (clinical efficacy)
Clinical validation: more data necessary
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Reflection
What benefits do I hope to see through DR testing in our country?
What are my concerns right now about this type of testing?
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Types of Resistance Assays
Genotypic Testing: Prediction of phenotype based on sequence
Phenotypic Testing: Measure of susceptibility to specific drugs– Recombinant Assays: RT/PCR portion of patient
virus and transfer into a vector• Several different versions commercialized,
automated and regulated– PBMC Assay: Culture virus from patient
• Largely replaced by recombinant assays due to difficulties in reproducibility and throughput
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Genotype Assays: Generic Procedure
Patient virus
PR-RT DNA
RT-PCR
Protein Sequence
Sequencing
Resistance Mutations
Selection
Prediction of Drug
Susceptibility
Interpretation
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Sources and Types of Genotype Assays
Reference Labs (North America and Europe)– Send blood sample in, receive results on report
• LabCorp, Quest, AML, Specialty, Mayo, etc.• Monogram Biosciences, Virco
Commercial kits– Can be performed on site (in laboratory)
• TruGene (Siemens)• ViroSeq (Abbott/Celera)
In-house assays– "home brew" assay performed at one site
(clinical, hospital or research laboratory)– Must be validated and approved if used for
patient management
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Genotyping: Pros and Cons
Advantages
Can be performed rapidly (days)
Relatively inexpensive ($100 - $400)
Available in many labs
Disadvantages
Does not directly measure susceptibility
Sometimes difficult to interpret results
Not all patterns of resistance mutations are known (esp. for new drugs and combinations)
Generally qualitative Subjectivity in mixture
detection
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Phenotype Assays: Generic Procedure
Patient virus
PR-RT DNA
RT-PCR
(Resistance Test Vector)
(Vector Assembly)
Recombinant Virus
Transfection
Measure of Drug
Susceptibility
Infection
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Commercially Available Phenotypic Assays
All are recombinant virus phenotypic assays
Antivirogram (Virco)– Homologous recombination used to generate
vector– Replication occurs over multiple cycles
PhenoSense (Monogram)– Vector generated directly by digestion and
ligation– Replication limited to single cycle– High precision and reproducibility
Phenoscript (VIRalliance)– Homologous recombination used to generate
vector– Uses VSV-G protein for virus entry– Replication limited to single cycle
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Phenotyping: Pros and Cons
Advantages
Direct measure of drug susceptibility
Quantitative Can immediately test
new RT and PR inhibitors
Disadvantages
Longer time to obtain results (weeks)
Relatively complex technology
More expensive than genotypic assays
Available in fewer labs
Clinical cutoff values for drug resistance not clearly defined for all drugs
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GENO vs PHENO?
A gross oversimplification:– Utility of genotypic testing greatest earlier in
treatment continuum– Utility of phenotypic testing increases with
subsequent treatment rounds
Genotypic testing Genotypic testing
Phenotypic testing
Treatment rounds
Utility
Increasing Genetic Complexity
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Phenotype/Genotype Discordance
GenotypePatient virus
PR-RT DNA
RT-PCR
Protein Sequence
Sequencing
Resistance Mutations
Selection
Prediction of Drug
Susceptibility
Interpretation
PhenotypePatient virus
PR-RT DNA
RT-PCR
(Resistance Test Vector)
(Vector Assembly)
Recombinant Virus
Transfection
Measure of Drug
Susceptibility
Infection
?
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3 Types of Phenotype-Genotype Discordance
Genotype predicts resistance, not reflected in phenotype ("PT-S, GT-R"); mixtures of resistant and sensitive viruses are present in the specimen
Genotype predicts resistance, not reflected in phenotype ("PT-S, GT-R"); not explained by mixtures
Genotype predicts susceptibility, but phenotype detects reduced susceptibility or resistance ("PT-R, GT-S")
Parkin et al. JAIDS 2002
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Genotype/Phenotype “Discordance”
Previously unrecognized cross-resistance (or lack of expected cross-resistance).
Previously unrecognized resistance-associated mutations (“RAMs”).
New amino acids at known positions.– e.g. N88S, I84A, K103S, I47A, K101P, V106M…
Effect of recognized RAMs tempered by other mutations.– e.g. L90M, V82A, K103N, suppression of ZDV by
184, etc. Particular combinations of polymorphisms
or “secondary” mutations– e.g. K103R/V179D
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Discussion
What are the different types of resistance assays?
What are the advantages and limitations of these assays?
What results can we expect from these tests?
What if the results from one type of test are inconsistent with results from another?
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Reflection
What type of DR assay is most appropriate for our situation?
What results can we expect? What factors should we keep in mind about
this type of assay?
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