Susceptibility of primary HTLV-1 isolates from patients with HTLV-1-associated myelopathy to reverse transcriptase inhibitors

Abstract

Since human T-lymphotropic virus type 1 (HTLV-1)-associated diseases are associated with a high HTLV-1 load, reducing this load may treat or prevent disease. However, despite in vitro evidence that certain nucleoside/nucleotide analogue reverse transcriptase inhibitors (NRTIs) are active against HTLV-1, in vivo results have been disappointing. We therefore assayed the sensitivity of HTLV-1 primary isolates to a panel of RT inhibitors. HTLV-1 primary isolates were obtained, pre- and post- NRTI treatment, from patients with HTLV-1-associated myelopathy. Sensitivity to azidothymidine (AZT), lamivudine (3TC), tenofovir (TDF) and three phosphonated carbocyclic 2'-oxa-3'aza nucleosides (PCOANs) was assessed in a RT inhibitor assay. With the exception of 3TC, HTLV RT from primary isolates was less sensitive to all tested inhibitors than HTLV-1 RT from MT-2 cells. HTLV-1 RT from primary isolates and from chronically infected, transformed MT-2 cells was insensitive to 3TC. Sensitivity of primary isolates to RT inhibitors was not reduced following up to 12 months of patient treatment with AZT plus 3TC. The sensitivity of HTLV-1 primary isolates to NRTIs differs from that of cell lines and may vary among patients. Failure of NRTIs to reduce HTLV-1 viral load in vivo was not due to the development of phenotypic NRTI resistance. AZT and the three PCOANs assayed all consistently inhibited primary isolate HTLV-1 RT.

Keywords: Human T-cell Lymphotropic Virus type 1; azidothymidine; drug sensitivity; lamivudine; phosphonated carbocyclic 2′-oxa-3′aza nucleosides; primary isolates; reverse transcriptase; tenofovir.

Figure 1

RT activity in human T-lymphotropic virus type 1 (HTLV-1) isolates from peripheral blood mononuclear cell (PBMC) cultures of HTLV-1-associated myelopathy (HAM) patients. Centrifuge-enriched isolates from PBMC cultures of 5 patients with HAM (TAY, TAR, TAU, TBI, TAQ), before (T0) and after (T48) therapy with 3TC plus AZT, or from MT-2 cells, were assayed for p19. Serial dilutions of p19-equivalent isolates were added to the RT assay and amplified products of the template were visualized on a 2% agarose gel and quantitated by densitometry, as a measure of RT activity.

Figure 2

p19 release in culture supernatants from PBMCs of HAM patients before and after therapy. The p19 concentration in supernatants from PBMC cultures of 5 patients with HAM (TAY, TAR, TAU, TBI, TAQ), before (T0) and after (T48) therapy with 3TC plus AZT, was evaluated by ELISA. Values are expressed as the mean of quadruplicate samples ± standard deviation. Production of p19 (black column) in the supernatant of chronically infected cells MT-2 was used as a positive control.

Figure 3

In vivo HTLV-1 viral load (HTLV DNA copies /100 PBMCs) during treatment with tenofovir.