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HIV Resistance Testing Clinical Implications. Cyril K. Goshima, M.D. Director, AIDS Education Project June, 2009. I am a?. Physician Nurse Pharmacist Dentist Student Patient Other. Resistance Testing will tell us what meds will work for a patient?. True False.

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HIV Resistance Testing Clinical Implications

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HIV Resistance TestingClinical Implications

Cyril K. Goshima, M.D.

Director, AIDS Education Project

June, 2009


I am a?

  • Physician

  • Nurse

  • Pharmacist

  • Dentist

  • Student

  • Patient

  • Other


Resistance Testing will tell us what meds will work for a patient?

  • True

  • False


Resistance Testing is recommended before starting HAART.

  • True

  • False


Resistance Testing can be done on patients with viral load less than 500.

  • True

  • False


The K103N mutation is an NNRTI Mutation

  • True

  • False


M184V is a common PI Mutation

  • True

  • False


When to Use Resistance Testing

  • DHHS Guidelines

    • Recommend testing: acute infection, suboptimal virologic suppression after treatment is initiated, treatment failure, prior to the initiation of therapy.

    • Consider: chronic infection < 2 yrs.

    • Which test is not recommended


Resistance Testing

  • Genotypic Resistance Testing

  • Phenotypic Resistance Testing

  • Combined Geno/Pheno

  • “Virtual Phenotype” Testing

  • Trofile (HIV Tropism Assay)

  • PhenoSense Entry (FI Resistance Testing)

  • Integrase Resistance Testing

  • Replication Capacity


Requirements for Resistance Testing

  • Viral Load must be greater than 500 for genotype & phenotype resistance testing, integrase resistance testing

  • Viral load must be greater than 1000 for Trofile and PhenoSense Entry


Genotypic Resistance Testing

  • Detects mutations in the HIV genome associated with resistance to specific drugs.

  • Advantages

    • Adequate turn-around time (1-2 wks)

    • Less expensive

    • Detect mutations that may precede phenotypic resistance

    • Widely available

    • More sensitive in detecting mixtures of resistant and wild type viruses


Genotypic Resistance Testing

  • Disadvantages

    • Indirect measure of resistance

    • Relevance of some mutations unclear

    • Unable to detect minority variants (<20 – 25% of viral sample)

    • Complex patterns may be difficult to interpret

    • Genotypic correlates of resistance not well defined for non-B subtypes.


Phenotypic Resistance Testing

  • Measures the patient’s HIV isolates ability to replicate in the presence of varying concentration of specific drugs.

  • Advantages

    • Direct and quantitative measure of resistance

    • Method can be applied to any agent incl. new where genotypic correlates are unclear

    • Can assess interactions among mutations

    • Accurate with non-B HIV subtypes.


Phenotypic Resistance Testing

  • Disadvantages

    • Susceptibility cut-offs not standard between assays

    • Clinical cut-offs not defined for some drugs

    • Unable to detect minority species

    • Complex technology

    • More expensive

    • Longer turn-around time.


Other Tests

  • Geno/Phenotype Resistance Testing

    • e.g. Phenosense GT from Monogram

    • Both tests are performed

    • The discordance is reported

  • “Virtual” Phenotype

    • Genotyping is performed and the phenotype is determined by looking at all the matched pairs of genotype with phenotype in a data set to give the best estimate


Other Tests

  • Fusion Inhibitor Resistance Testing

    • Resistance to Enfuvirtide

  • Replication Capacity

    • How weak is your patient’s virus?

  • Chemokine Receptor Identification

    • CCR5, CXCR4, or mixed virus present

  • Integrase Inhibitor Resistance Testing


How We Identify a Mutation

  • How do we identify a resistance mutation?

M 184 M

“M” is the “wild type” amino acid

“184” is the codon position


How We Identify a Mutation

  • How do we identify a resistance mutation?

M 184 V

“M” is the “wild type” amino acid

“184” is the codon position

“V” is the mutant amino acid


How We Identify a Mixture

M 184 M/V

“M” is the “wild type” amino acid

“184” is the codon position

“M/V” is the mixture of wild type & mutant amino acid


Definitions for Phenotypic Resistance Testing

  • IC50 = Concentration of drug required to inhibit replication by 50%

  • Fold Change = IC50 pt./IC50 reference

  • Cut Off = Fold change or concentration below which the virus is considered susceptible, above which non-susceptible

  • Biological Cut Off = Fold change based on variations in clinical samples from treatment naïve individuals.


Definitions for Phenotypic Resistance Testing

  • Clinical Cut Off = Fold change based on virologic response to ARV in Clinical Trials

  • Replication Capacity: The ability of a pt’s virus to replicate in the absence of drug


NRTIs


NRTI Mutations

  • Single point mutation can result in high level resistance e.g. M184V (3TC, FTC), K65R (TDF)

  • TAMS pattern of mutations e.g. codons 41, 67, 70, 210, 215, 219 (AZT, D4T)

  • 2 other patterns that are selected for by AZT/DDI & DDI/D4T

    • Q151M:resist. all NRTI except TDF

    • T69insertion + 1 or more TAMS @ 41, 210, 215: resist. all NRTI


Common Mutations: NRTIs

  • TAMS = thymidine analog mutations (aka ZDV mutations): M41L, D67N, K70R, L210W, T215F/Y, K219E/Q

  • NAMS = nucleoside analog mutations: TAMS plus E44A/D, A62V*, K65R, T69D, T69ins, L74I/V, V75A/I*/M/S/T, V77L*, Y115F, F116Y*, V118I, Q151M, M184I/V

    *Secondary mutations seen with Q151M


NRTI Signature Mutations

*TAMS=Thymidine analog mutations.


NNRTIs


Common Mutations: NNRTIs

  • Delavirdine (DLV)

    • L100I, K103N, V106M, Y181C, I; Y188L, G190E/Q

    • P236L(rare), Y318F

  • Efavirenz (EFV)

    • L100I, K103N, V106M, Y181C, I; Y188L, G190A, S, E, Q…; P225H

  • Nevirapine (NVP)

    • L100I, K103N, V106A, M; Y181C, I; Y188C, L, H; G190A, E, S, Q…,F227L


NNRTI Multi-Drug Resistance

Class Resistance

  • L100I, K101E or P, K103N or S, V106A or M, Y188C, H, or L, M230L

  • Resistance to one NNRTI usually confers cross resistance to all other agents (exceptions: 181 and EFV, 190A/S and DLV)

  • Continued viral replication in the presence of NNRTI results in accumulation of additional resistance mutations

    • May impact clinical utility of future NNRTIs


  • NNRTI Novel Mutations

    • Those exhibiting a > 10 fold change:

      • K103R and V179D (in combination)

      • K101P


    NNRTI: Etravirine

    • K103N NNRTI mutation is not associated with resistance to Etravirine

    • Multiple Resistance Associated Mutations (RAMS)

    • Scoring of the number of RAMS determines resistance to Etravirine

    • Similar to Protease Inhibitor Scoring


    PIs


    PI Resistance

    • Cross resistance is common

    • PI mutations are uncommon in boosted PI regimens

    • Multiclass experienced pts. may have been exposed to unboosted regimens

    • The number of primary PI mutations may predict the response to therapy e.g. TPV score 0-3 good, 4-7 intermediate, >8 poor or Kaletra


    PI Common Mutations


    Hypersusceptibility


    NRTI Increased Susceptibility


    NNRTI Increased Susceptibility


    PI Increased Susceptibility


    PI Hypersusceptibility

    • Mutation I50V, selected by LPVr and APV, increased susceptibility to ATV, TPV.


    Integrase and Entry Inhibitor Resistance

    • Resistance has been seen against the Entry Inhibitors and Integrase Inhibitors

    • There are resistance tests that can be ordered

    • For CCR5 it may just be a repeat of the Trophile test to determine change to mixed or dual tropic viruses


    Case Discussion

    • Patient CB, 42 y/o, homosexual male

    • Current Regimen (05/31/06): CBV/TDF/EFV

    • Past Drugs: CBV/ IDV, CBV/NFV

    • CD4/VL

      • Date: 09/08/05 349/8,810

      • Date: 03/07/06 192/10,300

      • Date: 06/02/06 186/9,400

      • Date: 09/18/06 92/6,610

      • Date: 10/17/06 /12,000


    Case Discussion

    • NRTI

      • M184V present (3TC/FTC resist, TDF hs)

      • Multiple TAMs

      • No K65R (TDF sens despite 41 & 215 mut)

    • NNRTI

      • No significant mutations

    • PI

      • 4 TPV assoc mut (intermediate response)

      • DRV sens


    Case Discussion

    • Was the CBV/TDF/EFV regimen a reasonable one?

    • There has been no response to this therapy after 3 mos.

    • What should you do?

    • Any suggestions on a possible new regimen?


    Discordance

    • Inaccurate genotype interpretation algorithm that does not account for novel or previously unknown mutation effect

    • Mixtures of wild type and resistant strains. Phenotype underestimates resistance

    • Variability in phenotypic susceptibility with specific mutations

    • Believe the genotype. Genotypic change may precede phenotypic resistance.


    Clinical Implications

    • Is there evidence for sequencing of NRTIs?

    • Should the initial regimen be a boosted PI or a NNRTI?

    • Is 3TC = FTC as far as resistance is concerned?


    Clinical Implications

    • Try to use at least 2 new potent agents to switch from a failing regimen.

    • The longer a failing regimen is continued, the more mutations accumulate. If there is no new agent, better to cont. the same regimen unless compelled to do otherwise.

    • Resistance is relative. 3TC cont. to have virological effect despite M184V mutation. Boosted PIs may have more of a response than an unboosted PI evidenced by a lower fold change.


    Clinical Implications

    • NRTI

      • TAMs can prevent K65R mutation. K65R is associated with multiple NRTI resistance and TDF resistance. ? Add ZDV to failing regimen

      • Continue 3TC or FTC despite a M184V mutation (hypersusc. ZDV, TDF, D4T; RC)

    • NNRTI

      • DC NNRTI as soon as mutations develop. There is no virological or RC advantage.


    Clinical Implications

    • PI

      • Never use an unboosted PI.

    • Antiretroviral susceptibility is on a continuum. Using drugs with the most activity (lower fold change) is a reasonable choice.


    Clinical Implications

    • In initial therapy, a boosted PI regimen may have an advantage over a NNRTI regimen because of fewer HIV mutations. (J. Bartlett, et al, JAIDS, 4(3): 323-331; Swiss HIV Cohort Study, oral abstract 72, XV International HIV Drug Resistance Workshop) Possible explanations maybe lower genetic barrier and pharmacokinetics with missed doses.


    Clinical Implications

    • Replication Capacity

      • Lower RC with certain NRTI (3TC) and PI (NFV).

      • No change in RC with NNRTI


    Resistance Testing will tell us what meds will work for a patient?

    • True

    • False


    Resistance Testing is recommended before starting HAART.

    • True

    • False


    Resistance Testing can be done on patients with viral load less than 500.

    • True

    • False


    The K103N mutation is an NNRTI Mutation

    • True

    • False


    M184V is a common PI Mutation

    • True

    • False


    Acknowledgements

    • Monogram Bioscience, Sharon Martens, MN, ARNP/FNP

    • Dr. Joel Gallant, MD, MPH from Clinical Care Options HIV LLC, “Use and Interpretation of Resistance Tests in Multi-Class Experienced Patients,” September 2, 2005.


    Thank You

    Questions?


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