BHIVA draft adult antiretroviral treatment guidelines: Resistance

INTRODUCTION

Drug resistance testing has been used for many years to support the treatment of infectious diseases. In virology, resistance testing has been constrained, until recently, by a lack of appropriate technology, and difficulty in routinely propagating relevant viruses. Gene sequencing was established as a research tool over 30 years ago and now academic and commercial groups have developed and modified research technologies to form ‘kits’ and/or systems for evaluation for clinical use.

The challenge is to develop a clinical ‘service’ that will detect or predict changes in susceptibility associated with antiretroviral drug resistance, using a system that is rapid, reliable, reproducible and quality assured.

Assuming that technologies available withstand this scrutiny we are left to determine how results of resistance testing are related to virological and clinical outcomes, and their relevance for improving patient care.

Intuitively it would seem obvious that an understanding of HIV-1 drug resistance profiles should help in the management of their therapy. However, within the setting of a chronic viral infection where drug responses are equally influenced by adherence, pharmacological tolerance and drug resistance it may be difficult to demonstrate this unequivocally.

Antiretroviral drug resistance has been described retrospectively, in vitro and in vivo, to be associated with poor virological and clinical outcomes (Table 1a) and short-term prospective, randomized controlled trials have, in some studies, demonstrated the short term benefits of resistance testing over standard of care (Table 1b). It remains to be clarified in which patient groups resistance testing is of long term benefit, or indeed if resistance testing without measures of adherence and/or drug pharmacokinetics, in the long term, is likely to be useful at all.

TABLE 1

Retrospective studies

Study

Resistance test

Significant outcomes

1. 1994: CONCORDE SUBSTUDY

Genotyping

ZDV MTs predicted virological and clinical outcome

2. 1995: ACTG 116B/117

Phenotyping

ZDV resistance associated AIDS/death

3. 1995: ACTG 116B/117

Genotyping

ZDV MTs associated with AIDS/death

4. 1998:FRANKFURT COHORT

Phenotyping & Genotyping

Resistance ZDV/3TC associated with failure

5. 1999: UNIVERSITY CLINIC

Genotyping

PI MTs associated with RIT-SAQ failure

6. 1999: HOSPITAL CLINIC

Phenotyping

Number of PI/NRTI sensitive associated with positive response

7. 1999:SWISS HIV COHORT

Genotyping

PI/NRTI MTs predict virological response

8. 2000: FRANKFURT COHORT

Phenotyping

PI/NRTI sensitivity

associated with virological response to Mega-HAART

9. 2000: HOSPITAL CLINIC?

Genotyping

MT at baseline to a new NRTI predicts virological failure of new regimen

F Brun-Vezinet,S Kaye, F.Ferchal, C Loveday,C, Buffet-Janvresse, R Tedder, J Darbyshire, JP Aboulker AZT Resistance:A Case Controlled Study from Concorde. Third Int. Workshop on HIV Drug Resistance.Kauai, Hawaii. 2-5 August 1994. Abstr.49

D’Aquila RT,Johnson VA,Welles SL et al.ZDV resistance and HIV-1 disease progression during antiretroviral therapy.Ann Int Med.1995, 122: 401-408

Japour AJ, Welles S, D’Aquila RT et al. Prevalence and clinical significance of ZDV resistance mutations in HIV isolates from patients after long term ZDV therapy. J Infect.Dis.1995. 171:1172-1179

Miller V, Phillips, Rottman C et al. Dual resistance to ZDV and 3TC in patients treated with ZDV/3TC combination therapy: association with therapy failure. J Inf ect Dis. 1998, 177: 1521-1532

Zolopa AR, Shafer R, Warford A et al. HIV-1 genotypic resistance patterns predict response to saquinavir/ritonavir therapy in patients in whom previous protease inhibitor therapy failed. Ann Int Med. 1999, 131: 813-821

Deeks SG, Hellman NS, Grant RM et al. Novel 4 drug salvage treatment regimens after failure of HIV-1 protease inhibitor-containing regimen: antiviral activity and correlation of baseline phenotype drug susceptibility to virological outcome. J Infect Dis, 1999, 179: 1375-1381

Lorenzi P, Opravil M, Hirschel B et al. Impact of drug resistance mutations on virological response to salvage therapy. AIDS, 1999, 13: F17-21

Miller V, Cozzi-Lepri A, Hertogs K et al. HIV drug susceptibility and drug response to Mega-HAART regimen in patients from the Frankfurt cohort. Antiviral Therapy 2000, 5:56-71

Van Vaerenbergh K, Van Leuthem K, Van Wijngaerden E et al. Baseline HIV-1 genotypic resistance to a newly added nucleoside analogue is predictive for virological failure of the new therapy. AIDS Res Hum Retroviruses 2000. 16: 529-537

Prospective Studies Resistance Test

Study

Resistance test

Significant outcomes

1. 1999: VIRADAPT (RCT)

Genotyping

Better virological response after 6 months (p<0.05)with genotype v SOC

2. 1999: VIRADAPT (open Study)

Genotyping

Benefits of genotyping maintained for 1 year

3. 1999: GART (RCT)

Genotyping

Better virological response

after 8 weeks (p<0.05) with genotyping+expert advice v SOC

4. 2000: VIRA 3001 (RCT)

Phenotyping

Better virological response after 16 weeks (p<0.05) with phenotyping v SOC

5. 2000: HAVANA (RCT)

Genotyping

Better virological response after 6months(p<0.05) with genotype v SOC

6. 2001: NARVAL (RCT)

Phenotyping

No difference for SOC & Genotyping, phenotyping but better virological response after 6 months (p<0.05) with genotyping v SOC

MTs = mutations; PI = protease inhibitors; NRTI = nucleoside reverse transcriptase inhibitors; SOC = standard of care; RCT = randomized controlled trial

Durant J, Clevenbergh F, Halfon F et al. Drug resistance genotyping in HIV-1 therapy: the VIRADAPT randomised controlled trial. Lancet. 1999, 353: 2195-2199

Clevenbergh F, Durant J, Halfon F et al. Persisting long-term benefit of antiretroviral genotypic guided treatment for HIV infe4cted patients failing HAART. Antiviral Ther. 1999 (Suppl 1) 42: Abstr 60

Baxter JD, Mayers DL, Wentworth DN et al. A randomised study of antiretroviral management based on plasma genotypic resistance testing in patients failing therapy.CPCRA 046 study team for the Terry Beirn Community Programs for Clinical Research on AIDS. AIDS 2000. 14: F83-93

Cohen C, Kessler H, Hunt S et al. Phenotypic resistance testing significantly improves response to therapy: final analysis of a randomized controlled trial (VIRA 3001). Antiviral Ther. 2000. 5 (Suppl 3): 67

Havana – CROI, 2001

Meynard JL, Wray M, Mourand-Joubert L et al. Impact of treatment guided by phenotypic and genotypic resistance tests on the response to antiretroviral therapy: a randomized controlled trial (NARVAL,ANRS 088). Antiviral Ther. 2000, (Suppl 3): 67-68

THE TESTS

Resistance tests available for determination of of HIV-1 drug susceptibility (phenotyping) and detection of mutations associated with drug resistance (genotyping) are amongst the most complex ever addressed for use in routine clinical care. In terms of reviewing performance and quality these issues divide naturally into:

Clinical sample and request. This is an important determinant of the ability to generate a useful result (Table 2)

The technical process. A range of different assays are available, summarised in Table 3

Validation of the technical process and printout. Assay quality control means the assimilation of extra samples at all stages of the test process, in parallel with clinical samples, in order to identify changes in sensitivity in the assay, and the generation of potentially false results. These extra controls increase the overall cost of the test. External quality assurance schemes test the quality of the overall process, from receipt of sample in the laboratory, to production of the final report. Such schemes are being developed for HIV drug resistance.

All these quality aspects of the test are assessed as part of laboratory accreditation procedures.

Generation of clinical report. Genotypic assays produce large volumes of detailed gene sequence information, from which must be identified those changes thought to be important in determining drug susceptibility. This comprises two steps. Firstly, the identification of any mutations which are different from the presumed "drug sensitive" virus, and secondly, providing an interpretation of these changes, with regard to specific drugs.. Software programmes are available to undertake these analyses, however they need regular updating, in the light of new knowledge on drug resistance, and differences between them still exist. A selection of programmes available for providing an interpretation of drug resistance associated mutations is given in Table 4. Table provides guidance on the association of specific mutations with resistance to specific drugs.

Clinical interpretation and advice. The factors involved in optimizing the clinical utility of resistance test results are given in Table 6.

Table 2

Practicalities in requesting resistance test

1. Request test on sample taken before starting first-line therapy, before stopping therapy or before changing therapy

2. In drug-experienced patients, do not request test when off therapy >2 weeks. In such circumstances, consider testing stored samples from the time of drug failure.

3. Request test on sample with viral load >1,000c/ml. EDTA.

4. Attempt to construct at least one specific question before requesting resistance tests, since this will assist in interpretation of results.

5. Discuss with the laboratory providing the test the most appropriate format for reporting.

6. Since testing of stored samples is likely to become increasingly requested in drug-experienced patients, close liaison with the laboratory undertaking viral load testing is required to ensure that relevant samples are stored appropriately.

Table 3

(a)

AVAILABLE TESTS

Genotype

Techniques

Advantages

Disadvantages

Examples and Availability

Identification of specific changes in relevant areas of the HIV genome associated with drug resistance. This is undertaken by gene sequencing or hybridisation

Rapid (turnaround time 2-3 weeks).

Detects minority population of virus (down to ~10-20%), so leads to early identification of resistance.

Moderately expensive (£200-£300).

Key mutations clearly associated with drug failure

Hybridisation based assays do not detect all possible mutations.

Complex pattern of mutations difficult to interpret.

Sequencing

Hybridisation

Applied Biosystems (Viroseq)

Visible Genetics

(Trugene)

VIRCO

Virologics

In-house

Innogenetics

(Innolipa)

Affymetrix

Clinical Virology Laboratory

Clinical Virology Laboratory

Commercial & Clinical Virology Laboratory ???

Commercial

Clinical Virology Laboratory

Clinical Virology Laboratory

Clinical Virology Laboratory

Table 3

(b)

Phenotype

Techniques

Advantages

Disadvantages

Examples and Availability

Classically undertaken by infection of PBMC, but in the main surpassed by producing a recombinant virus incorporating relevant genes from patient sample, which can be tested for drug susceptibility

Quantitative values for fold resistance against all drugs generated

Teminology to clinicians

Phenotypic resistance may only become apparent after appearance of specific mutations

Expensive (£400-£500)

Time consuming (>4 weeks)

Relevance of assay cut-offs, and biological cut-offs to clinical practice unclear

VIRCO

(Antivirogram)

Virologics

(Phenosense)

Viralliance

( Phenoscript)

Commercial Laboratory

Commercial Laboratory

Commercial Laboratory

Table 4

Some sources of information assisting in interpretation of resistance data

Guidelines by the International AIDS Society [137]

French guidelines (Error! Reference source not found.)

Guidelines by the US Dept of Health and Human Services (Error! Reference source not found.)

British Guidelines (Error! Reference source not found.)

Schinazi website table (Error! Reference source not found.]

Los Alamos website table (Error! Reference source not found.)

Stanford website database (Error! Reference source not found.]

Published specific genotypic algorithms [64,65,120]

Stand alone software based on specific genotyping algorithms (such as the Retrogramñ

software, Error! Reference source not found.)

Commercial interpretation systems (such as the VirtualPhenotype, Error! Reference source not found.)

Association of specific mutations with drug resistance

Table 5

Most common HIV-1 resistance mutations against currently available drugs

Name

(Abbreviation)

Time to resistance (1)

Most common resistance mutations in the target protein (2)

Nucleoside analogue reverse transcriptase inhibitors (NRTIs)

Zidovudine

(AZT)

Intermediate

M41L, D67N, K70R, L210W, T215Y/F, K219Q

(combined mutations needed for high level resistance)

Didanosine

(ddI)

Long

K65R, L74V, M184V

Zalcitabine

(ddC)

Long

K65R, T69D, V75T, M184V, (L74V)

Stavudine

(d4T)

Long (insufficient data)

V75T

M41L, D67N, L210W, T215Y/F, K219Q

Lamivudine

(3TC)

Fast

M184I/V, (K65R)

Abacavir

Intermediate

(not enough in vivo data)

K65R, L74V, Y115F, M184V

M41L, D67N, L210W, T215Y/F, K219Q

(combined ZAMS? mutations needed for high level resistance)

Most common NRTI cross resistance mutation patterns (3)

(A62V, S68G, V75I, F77L, F116Y, Q151M, combined mutations)

(T69S-XX, usually T69S-SS, in combination with other NRTI mutations)

Non-nucleoside analogue reverse transcriptase inhibitors (NNRTIs)

Nevirapine

Fast

A98G, L100I, K103N, V106A, V108I, Y181C/I, Y188C, G190A

Delavirdine

Fast

K103N/T, Y181C, P225H,P236L

Efavirenz

Fast

L100I, K101E/Q, K103N, V106A, V108I, G179N/E/D,Y188C/H/L, G190E/A/S, P225H (combined mutations needed for high level resistance)

Most common NNRTI cross resistance mutations (3)

K103N, Y181C

Protease inhibitors (PIs)

Saquinavir

Intermediate

L10I, G48V, I54L/M/V, L63P, G73S, V82A, L90M (combined mutations needed for high level resistance)

Ritonavir

Intermediate

L10I, K20R, V32I, L33F, M36I, M46I/L, I54L/M/V, L63P, A71V, V82A/F/S/T, I84V, L90M (combined mutations needed for high level resistance)

Indinavir

Intermediate

L10I/R/V, K20R/M, L24I, V32I, M46I/L, I54L/M/V, L63P, A71T/V, G73S, V82A/F/T, I84V, L90M (combined mutations needed for high level resistance)

Nelfinavir

Intermediate

D30N, M36I, M46I, L63P, A71V, V77I, I84V, N88D, L90M (combined mutations needed for high level resistance)

Amprenavir

Intermediate?

(not enough in vivo data)

L10F/I/V, V32I, L33F, M46I/L, I47V, I50V, I54V/L/M, V82I, I84V

(combined mutations needed for high level resistance)

Lopinavir

(not enough data)

Any 1 of M46I/L, V82A/F/L, I84V, L90M + any 7 of L10I/R/V, K20M/R, L24I/V, M46I/L, F53L, I54L/M/V, L63P, A71T/V, V82A/F/V, I84V, L90M

Notes

1. Key mutations, clearly associated with reduced drug susceptibility, are in bold

2. Our understanding of the impact of new mutations and interaction of mutations is ever changing. Therefore, the table above should be viewed as general guidance only, rather than a definitive list of all mutations.

3. The "time to resistance" column provides relative information for the particular drugs, rather that specific time periods. The actual time taken for resistance to develop to any one particular drug is determined by a multitude of factors such as co-exisiting therapies, adherence and pharmacological factors.

(1) Time to resistance is very variable and highly dependent on the efficacy of the combination therapy. Therapy choice should not primarily be based on a longer period to resistance since this is often associated with a less powerful drug. Fast: resistance generally develops within weeks of initiation of monotherapy, intermediate: resistance generally develops within a few months of initiation of monotherapy, long: resistance has either been poorly described in the literature or has developed only years after the initiation of monotherapy.

(2) Specific drug related mutations indicated are the ones observed under monotherapy. Mutations shown in bold are generally considered either primary or key mutations, their clinical use in patient follow-up is currently still under investigation. Mutations between brackets are cross-resistance mutations selected during therapy with another drug.

(3) Combination therapy can select for the mutations mentioned as cross-resistance mutations.

(4) There is increasing evidence for at least some degree of cross-resistance between AZT and d4T, as well as other nucleoside analogues.

(5) The phenotypic implications of these mutations have not been added, since assay cut-offs and their biological and clinical implications are still unclear. Clinically relevant cut-offs are only available for lopinavir, for which 10-fold reduced susceptibility is associated with an increased risk of non-response.

Aminoacid codes are: A (alanine), C (cysteine), D (aspartic acid), E (glutamic acid), F (phenylalanine), G (glycine), H (hystidine), I (isoleucine), K (lysine), L (leucine), M (methionine), P (proline), Q (glutamine), R (arginine), S (serine), T (threonine), V (valine), W (tryptophane), Y (tyrosine).

The data in this table are adapted from reference 1. [Vandamme A-M, Houyez F, Banheghi D et al. Laboratory guidelines for the practical use of HIV drug resistance tests in patient follow up. Antiviral Therapy 2001 (in press)].

Table 6

Aspects to consider when interpreting resistance results

1. For genotypic assays, the "weight" of specific mutations for causing resistance is dependent on the mutation and drug in question. In addition, mutations can interact to enhance or suppress resistance to specific drugs.

2. For phenotypic assays, the precise fold resistance required to confer a lack of drug response in vivo remains unclear for most drugs.

3. Adherence is very likely to impact on the resistance result; however, the precise manner in which differing degrees of adherence have an effect remains unclear.

4. Consider drug history. Previous drug failure may have been accompanied by mutations not detectable at a later time point. It should be assumed that these mutations remain archived and can rapidly re-emerge within the majority of virus population.

5. Discussion of results in the context of clinical picture with a virologist/clinician with expertise and experience in this area is strongly recommended. This will enhance the body of knowledge and experience within each clinical centre.

6. Geographical origin of virus. The interpretation of resistance mutations may be different between subtype B and other virus subtypes. If the virus is known to be of a non-B subtype, or this is suspected through the likely geographical origin of infection, then expert advice on interpretation should be sought.

TABLE 7 RECOMMENDATIONS

Patient Group

Recommendations

Comment

Primary HIV Infection (PHI)

Recommend

Within the UK, a national surveillance programme provides real-time resistance testing free of charge, to monitor transmission of drug resistant viruses (2). If therapy indicated, use results to guide therapy, or modify therapy if result obtained after initiation.

Drug naïve, chronic infection

Consider

Resistance result may not reflect presence of transmitted resistant virus due to ongoing virus evolution, so testing may underestimate presence of resistance. Also consider testing baseline specimen if poor initial response to therapy

Drug experienced- 2^nd line or salvage

Recommend

Interpret in light of clinical and therapeutic history

Pregnancy

Recommend

Do not delay therapy where late maternal presentation, and modify therapy when result obtained

Paediatrics

Recommend

Indicated prior to therapy if mother had detectable viral load while receiving therapy, and also at times of therapy failure

Post exposure prophylaxis

Recommend

Undertake if sample available from source patient, but use result to modify therapy rather than delay onset of PEP.

Notes:

1) Ensure that laboratory undertaking viral load assays store aliquots of plasma samples appropriately, in order that retrospective resistance testing can be undertaken from relevant time points e.g prior to therapy, at times of previous drug failure

2) UK Collaborative Group on Monitoring Transmission of HIV Drug Resistance, is a collaboration between the PHLS and UK HIV Seroconverter Register. Plasma samples should be sent to PHLS Antiviral Susceptibility Reference Unit, Birmingham Public Health Laboratory, Birmingham Heartlands Hospital, Birmingham B9 5SS