IN-VITRO SELECTION AND MOLECULAR CHARACTERIZATION OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 WITH REDUCED SENSITIVITY TO 9-[2-(PHOSPHONOMETHOXY)ETHYL]ADENINE (PMEA)

9-[2-(Phosphonomethoxy)ethyl]adenine (PMEA) is an acyclic nucleotide w ith potent in vitro activity against human immunodeficiency virus type 1 (HIV-1). The present study was undertaken to determine whether HIV- 1 resistance to PMEA could be generated by in vitro selection and if s o, to determine which mutations in reverse transcriptase CRT) were res ponsible. HIV-1(LA1) was serially passaged for 10 months in the presen ce of increasing concentrations of PMEA up to a maximum of 40 mu M. Af ter 40 passages, the 50% inhibitory concentration (IC50) of PMEA had i ncreased almost 7-fold from 4.45 to 30.5 mu M. Some cross-resistance t o 2',3'-dideoxycytidine (ddC, zalcitabine), 2',3'-dideoxyinosine (ddI, didanosine), and 3'-thiacytidine (3TC, lamivudine) was also observed, but no cross-reactive resistance to 3'-azido-3'-thymidine (AZT, zidov udine). Sequencing of the RT encoding region of each of eight pol clon es from resistant isolates revealed a Lys-65 --> Arg (K65R) substituti on. HIV with the K65R mutation inserted by site-directed mutagenesis a lso had decreased sensitivity to PMEA in H9 cells and a similar cross- resistance profile. Thus, HIV can develop decreased sensitivity to PME A after long-term in vitro exposure and this change is associated with a K65R substitution. Additional studies will be needed to determine w hether a similar mutation in HIV RT develops in patients receiving PME A or its orally bioavailable prodrug adefovir dipivoxil (bis-POM PMEA) .