Single-dose nevirapine resistance in over 75% of mothers

Although there has always been an element of concern over this strategy - and certainly the recent IAS conference in Paris saw something of a “nevirapine backlash” with much discussion around alternative approaches (see IAS report later in this issue) – single dose nevirapine (NVP), given to the mother at onset of labour followed by a dose to the infant is a low cost and simple strategy to reduce mother to child transmission (MTCT) of HIV in resource poor settings.

However, in HIVNET 012 early findings demonstrated NVP resistance was detected in 21/111 (19%) of women at 6-8 weeks and mutations were also detected in some women as early as seven days following the 200 mg maternal dose. Two posters reported further analysis of NVP resistance mutations and their rate of selection in women receiving NVP to reduce MTCT.

High frequency of mutations demonstrated in women with subtype C

To date, NNRTI-associated mutations have been shown in subtypes A and D and subtype B HIV-infected women. In a report from Kantor and colleagues the rate of selection of specific mutations and their persistence and fitness is described in a group of women with subtype C HIV. [1]

The investigators analysed reverse transcriptase sequences from 34 women from Chitungwiza, Zimbabwe, participating in the HPTN 023 – where women received single dose NVP at onset of labour. Samples were collected at weeks zero, two, eight and 20-32 post partum. Sequences were evaluated for subtype and recombination and assessed for NNRTI mutations at codons 98, 100, 101, 103, 106, 108, 179, 181, 188, 190, 225, 227, 230 and 238.

They reported that overall 25/33 (76%), of all samples were found to have NNRTI mutations at any time point, 21/28 (75%) of available samples at two weeks, 11/32 (34%) at eight weeks and 1/8 (13%) of available samples at 24 weeks. At two and eight weeks two or more mutations were found in 11/28 and 5/32 sequences, and a single mutation in 10/28 and 6/32 respectively.

Prevalence of common NVP associated mutations at two weeks included Y181C in 16/28 (57%), K103N in 4/28 (25%), V106A/M in 5/28 (19%) and Y188C in 4/28 (14%). The investigators report that reversion to wild type had occurred by week eight in 12/27 (44%) samples with week two mutations, where a week eight sample was available. K103N was detected in 9/32 (28%) and V106A and Y181C in 2/32(6%) each. Of seven women with mutations for whom late samples were available 6/7 with K103 had reverted to wild type.

The investigators concluded that samples taken within two weeks of receiving single dose NVP harboured a high frequency of NNRTI resistance mutations and that dual mutations may be mixtures of competing single mutants. There appeared to be rapid reversion to wild type occurring by eight and 24 weeks and although Y181C is predominant in early samples, K103N is the predominant mutation selected at eight weeks. They also noted: “Sequencing multiple clones from sequential time points will be useful for quantification and linkage of mutations. The change over time in the proportion of specific mutations may serve as an indication of in vivo fitness relative to wild type RT in subtype C HIV-1.”

Diverse mutations emerge rapidly: HIVNET 012

A further evaluation from Eshlemen and colleagues from the HIVNET 012 looked at NVP mutations detected by population sampling in early samples, collected at seven days. Some samples had more than one NVP mutation. In this study the investigators cloned and sequenced individual HIV-1 variants to examine their spectrum and generic linkage. [2]

The samples from five women had one to four nevirapine mutations. DNA amplified from those samples was cloned and inserted into plasmids that were subsequently isolated. At least ten plasmids were isolated and sequenced from each plasma sample.

Sequencing of cloned plasmids revealed diverse patterns of NVP mutations. The investigators reported some plasmids lacking NVP mutations (wild type) were isolated from each plasma sample. The other plasmids had detectable NVP mutations including K103N, V106A, V108I, V179D, Y181C, G190A, G190S, Y188c and Y188L.

Three of the five women had NVP mutations that were detected in plasmids but not by population sequencing and as many as seven different mutations were detected in plasmids from a single sample. The investigators noted that three women had plasmids with more than one NVP mutation, confirming that those mutations were genetically linked.

They concluded that cloning and characterisation of individual HIV-1 variants reveals rapid selection of diverse subpopulations, that some variants contained more than one NVP mutation, and many contained NVP mutations that were not detected by population sequencing. They added that detection of variants with NVP mutations only seven days after receiving single dose NVP suggests that these variants were present at low levels before receiving the NVP. The investigators also noted “Further studies of NVP resistance in women and children receiving single dose NVP may help optimise use of NVP for prevention of MTCT.”

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Both studies confirm the well-recognised fact that single doses of nevirapine given to antiretroviral naïve women will result in selection of resistant variants. The scale of this is startling. Three-quarters of the samples obtained by Kantor et al two weeks after the dose was given had evidence of mutations. It also highlights how fast these mutations can disappear from the plasma, but although not present in later plasma samples they will be archived in pro viral DNA.

Neither study addresses the crucial question of the longer-term consequences of briefly selecting out resistant virus. It has been argued that such variants arise by natural mutation every day in untreated, infected individuals. However, this is not the same as flooding the long-lived lymphoid cells and sanctuary sites, albeit briefly, by single dose intervention. Will this result in loss of efficacy in future pregnancies, or for those women able to access treatment for themselves? First line ART in many poorly resourced settings is with nevirapine-containing combinations, so this is not an academic question.

References:

1. Kantor R, Lee E, Johnston E et al. Rapid flux in non-nucleoside reverse transcriptase inhibitor resistance mutations among subtype C HIV-infected women after single dose nevirapine. Abstract 78.

2. Eshleman EH, Jones D, Guay et al. HIV-1 variants with diverse nevirapine resistance mutations emerge rapidly after single dose nevirapine: HIVNET 012. Abstract 79.

	HIV Treatment Bulletin Vol 4 No7 August / September 2003