Amprenavir (APV) is a highly active and selective HIV protease inhibitor (PI) that is used for the treatment of HIV infection in adults and children. In this review we present data from extensive resistance studies undertaken during the development of amprenavir: These include in vitro and clinical studies where the phenotype and genotype of HIV protease was determined after treatment with amprenavir either as the single PI [alone or with nucleoside reverse transcriptase inhibitors (NRTIs)] or in combination with other PIs. In addition, cross-resistance with other PIs has been examined to help position use of amprenavir in the clinic. The key signature amino acid substitution associated with amprenavir resistance that was identified from in vitro and subsequent in vivo studies was isoleucine to valine at position 50 (I50V) in HIV protease. This was a new mutation not observed as a natural variant or in PI-experienced patients. Additional mutations including M46I/L and I47V were required to produce high level resistance. Cross-resistance was limited and observed only with ritonavir. In amprenavir/zidovudine/lamivudine combination therapy in PI-naive and lamivudine-naive patients, therapy failure is associated frequently with the reverse transcriptase M184V mutation, and not with amprenavir resistance. However, when amprenavir was added to NRTI therapy in NRTI-experienced and PI-naive patients where treatment was compromised by baseline NRTI resistance, failure was more frequently associated with the development of amprenavir resistance. From these studies, four pathways of amprenavir resistance were identified, with the I50V pathway associated with the highest levels of resistance. Alternative pathways to amprenavir resistance involved key substitutions, either V32I + I47V, or I54L/M, or more rarely I84V. Limited cross-resistance was observed to other PIs with each of these genetic mechanisms. Similarly, in PI-experienced patients, cross-resistance to amprenavir is markedly lower than for the other four approved PIs. Markers of cross-resistance to amprenavir include the above mutations but not L90M or V82A/T/Y as observed for other PIs. These data suggest that amprenavir may have an important part to play in rescue therapy regimens. In combination with other PIs, resistance development may be suppressed and key signature mutations, such as I50V (amprenavir), D30N (nelfinavir) and V82A/T/Y (ritonavir, indinavir) have not been observed in vitro. In addition, passage with saquinavir may resensitise amprenavir-resistant variants to amprenavir. Hypersensitivity to amprenavir has been reported for N88S variants occasionally observed after nelfinavir therapy. Differences in the resistance profile of amprenavir from that of other PIs suggest that amprenavir may add value to HIV combination therapy, particularly in PI combinations and in rescue therapy.