Background: Despite the success of HAART, cells which are latently infected with HIV remain in treated individuals. These cells are predominately CD45RO⁺, but some reports suggest that CD45RA⁺ cells can also act as an HIV reservoir. We have reported that latently-infected cells can be eliminated with an immunotoxin (IT) directed against CD45RO. The present study was designed to determine whether any latently-infected cells could be eliminated with an IT against CD45RA.

Methods: We evaluated the effect of an anti-CD45RA-IT (RA IT), an anti-CD45RO-IT (RO IT ) or a combination of the two on in vitro-generated latently infected cells. These ITs were also evaluated ex vivo on latently-infected CD4⁺ T-cells obtained from 7 HIV-infected individuals with detectable plasma viremia . A limiting dilution assay (5 x 10⁴ to 10⁶ cells/ml) was used to measure the frequency of infected cells. After treatment with one or both ITs, cells were stimulated with PHA or with PHA-activated PBMCs and p24 production was measured by ELISA. Full-length (FL) HIV DNA transcripts were quantitated using real time PCR.

Results: Treatment with the RO IT reduced mean levels of p24 by 87% whereas RA IT reduced it by 42%. Treatment with both ITs reduced p24 production by > 95%. Real time PCR analysis demonstrated reductions of FL HIV DNA transcripts of 96%, 75% ,and 98%, respectively, in the same cells. Ex vivo treatment of CD4⁺ T-cells obtained from 7 different HIV-infected individuals with the RO IT consistently reduced p24 levels by 95%–100%; the RA IT had a variable effect on p24 production. The combination of the 2 ITs reduced levels of p24 by 99%–100%. Limiting-dilution analysis demonstrated that RO IT reduced the mean frequency of HIV latently-infected cells from 8/10⁶ to 4/10⁶ cells (50% reduction) while the combination of both ITs reduced the frequency to 1.3/10⁶ cells (84% reduction).

Conclusions: These results suggest that a significant number of HIV latently-infected cells express both CD45RA and CD45RO. In comparing the effects of the 2 ITs on the latently-infected cells prepared in vitro vs those obtained from patients, it appears that the former may contain more RA⁺RO⁺ transitional cells than the latter.