Background:  In a retrospective analysis of a phase I/II study of the CXCR4 antagonist AMD3100, several subjects with X4 and dual-tropic virus at baseline developed R5 virus upon treatment. The total viral RNA in some patients remained high over the course of treatment despite receiving concomitant antiretroviral medications, suggesting that the virus in these patients was multi-drug resistant but sensitive to inhibition by chemokine receptor antagonists.

Methods:  Virus populations at baseline (day 0), on treatment (day 11), and off treatment (day 18) were characterized by determining the co-receptor tropism of 30 to 40 envelope clones per time-point using an envelope pseudo-virus infection assay. Gp160 sequences were determined for at least 10 clones per time-point. Sequence analysis of individual clones distinguished R5 viruses from X4 and dual-tropic viruses, based on differences in the V3 region. Viral stocks from patient samples at baseline were prepared by mixing patient lymphocytes with PHA-stimulated peripheral blood mononuclear cells (PBMC) from uninfected donors. The viruses were tested for their sensitivity to inhibition by the concomitant antiviral medications they were receiving during AMD3100 treatment both in MT-4 cells and PBMC, and the IC₅₀ values were compared with those obtained with reference X4 and R5 strains. Viruses were also tested for sensitivity to inhibition by a combination of the novel CXCR4 antagonist AMD070 and CCR5 antagonists in PBMC.

Results:  In 1 subject, the co-receptor utilization was 62.5% R5, 7.5% X4, and 30% dual clones at baseline and predominantly R5 clones at day 11 (85%) and day 18 (92%). This patient was receiving concomitant treatment with amprenavir (APV), lopinavir (LPV), didanosine (ddI), and stavudine (d4T). Compared with the inhibition of NL4.3 in MT-4 cells, the IC₅₀ for inhibition of patient virus with APV and LPV were about 80- and 130-fold higher, suggesting that the virus was resistant to these medications. In contrast, NL4.3 and patient viruses were equally sensitive to inhibition by ddI and d4T. Patient virus replication was completely inhibited in PBMC by a combination of AMD070 and CCR5 antagonists. Several other examples will be presented.

Conclusions:  Thus, 10-day treatment with AMD3100 suppressed replication of X4 and dual-tropic variants, resulting in a predominance of R5 variants. The X4 virus in these patients was in some cases resistant to their concomitant antiretroviral medications, but remained sensitive to inhibition with CXCR4 antagonists.

Keywords: resistance; CXCR4; antagonist