Phenotyping

Phenotypic resistance tests involve direct quantification of drug sensitivity. Viral replication is measured in cell cultures under the selective pressure of increasing concentrations of antiretroviral drugs and is compared to viral replication of wildtype virus.

Drug concentrations are expressed as IC50 values (50% inhibitory concentration). The IC50 is the concentration of drug required to inhibit viral replication by 50%. The sensitivity of the virus is expressed as the IC50 compared to the so-called cutoff value. The cut-off value indicates by which factor the IC50 of an HIV isolate can be increased in comparison to that of the wild-type, whilst still being classified as sensitive. The determination of the cut-off is crucial for the interpretation of the results!

Three different cut-offs are currently being used. The technical cut-off is a measure of the methodological variability of the assay and is approximately 2.5 fold more than the IC50. The biological cut-off, for example the comparative value on an anti-virogram, involves the inter-individual variability of virus isolates from ART-naive

HIV patients and is slightly higher than the technical cut-off. The biological cut-off does not, however, allow prediction of the clinical response to a drug. The clinical cut-off indicates up to which levels of IC50 virological success can still be expected.

For protease inhibitors (PIs), one has to take into account whether or not the respective clinical cut-off values are derived from clinical studies using unboosted or boosted PIs. Through boosting with ritonavir, drug levels may overcome certain levels of resistance.

Disadvantages of phenotypic testing include the lengthy procedure and high expense of the assay.

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