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Abstract
Investigating cellular antiretroviral resistance: preliminary results of the “ICARE” study
Lafeuillade A.1, Hittinger G.1, Poggi C.2, Benech H.3, Cupo A.4
1CHITS, Toulon, France, 2General Hospital, Toulon, France, 3CEA, Gif sur Yvette, France, 4CNRS, Sofia Antipolis, France
Introduction: despite HAART, low levels of HIV-1 RNA remain detectable in blood and reservoirs of most patients. No in vivo data are currently available on antiretroviral diffusion in lymph node cells (LNMC) versus blood cells (PBMC), or their correlation with the expression of multi-drug resistance proteins (MDR).
Methods: ongoing study of patients on first-line effective HAART (plasma viremia <50 c/ml for >6 months). A blood sample and surgical lymph node biopsy are done simultaneously. HIV-1 RNA levels are measured using RT-PCR, intra-cellular levels of NRTI active metabolites are measured using LC/MS/MS or radio-immunology, PI levels in plasma and total lymph node tissue are measured with HPLC. mRNA expression of P-gp, MRP-4, MRP-5 in cells is measured by a semi-quantitative PCR.
Results: 15 patients have already been analyzed, including 8 on Nelfinavir and 7 on Lopinavir- containing regimens. Four patients were also on tenofovir, 5 on abacavir, 7 on d4T, 8 on AZT and 11 on 3TC. Mean PBMC residual HIV RNA and DNA levels were respectively 44 and 2940 copies/million cells; mean LNMC residual HIV RNA and DNA levels were respectively 8420 and 5630 copies/million cells. Mean AZT-TP levels in PBMC were 4384 fmol/ million cells, 3TC-TP: 9.4 pmol/ million cells, d4T-TP: 66 fmol/ million cells, TDF-DP: 239 fmol/ million cells, CBV-TP: 2100 fmol/ million cells. AZT-TP levels were 10 fold lower in LNMC versus PBMC; 3TC-TP levels were 15 fold higher, d4T-TP, TDF-DP and CBV-TP were undetectable. The lymph node/plasma ratio was 58% for Nelfinavir and 21% for Lopinavir. High MDR mRNA expression was found in PBMC in 26% cases and in LNMC in 73% (p<0.03).
Conclusions: huge differences in antiretroviral diffusion and metabolism are found in vivo in PBMC versus LNMC; these differences could be related to differences in MDR expression. These data may contribute to understanding the mechanisms of HIV-1 RNA persistence during effective
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