Several years ago it was shown that very high doses of an
entry inhibitor-based microbicide could protect macaques from infection with SHIV162-p3. A paper in J Virology
now raises some important caveats about this approach, including the first
reported example of a microbicide selecting for drug-resistant virus. The
microbicide in question is a CCR5 inhibitor called PSC-RANTES. The challenge
virus used in the studies is called SHIV162-p3, a lab-created SIV/HIV hybrid in
which an HIV envelope that uses CCR5 to gain entry into CD4 T cells is inserted
into an SIV genome instead of the SIV envelope.
The new paper, authored by Dawn Dudley and colleagues from
Case Western Reserve University and the Tulane Primate Research Center, is
based on an analysis of macaques given lower doses of PSC-RANTES that were not
protective against SHIV162-p3 infection. In most cases, viruses in these
animals showed mutations that were deemed unrelated to PSC-RANTES because similar
mutations were seen in controls that were challenged with the same virus in the
absence of the microbicide. But the virus in one macaque contained two
mutations, K315R in gp120 and N640D in gp41, which were very rare in the
challenge virus stock (after infection, compared to their frequency in the challenge
virus, these mutations were increased at least 25-fold and 75-fold). Importantly,
PSC-RANTES failed to completely inhibit this mutant SHIV, even at high
concentrations that inhibited the parent SHIV162-p3 and all other R5-using HIV
isolates tested.
Dudley and colleagues acknowledge that this apparent example
of resistance was only seen in 1/25 macaques studied, and there is an outlying
possibility that the mutations represent a random adaptation of the virus to
better replicate in macaque cells. However, they argue that the weight of
evidence supports their conclusion that the mutations arose from drug selection
pressure, and that this possibility is particularly concerning given that
SHIV162-p3 is a very homogenous virus stock from which the emergence of
resistance would have been considered unlikely.
Discussing the implications for microbicides generally, they
note that the positive results reported to date with entry inhibitor-based
microbicides (PSC-RANTES, BMS-378806, CMPD167, and C52L) have all involved administering relatively high
doses and the SHIV162-p3 challenge virus (which “is exquisitely sensitive to
many entry inhibitors”). As a
result, they believe that “the barrier for selecting resistance to an anti-HIV
microbicide (such as PSC-RANTES) has been set very high and possibly beyond
physiological relevance.” To address these concerns, the authors recommend the development
of macaque models that use multiple R5-SHIVs as a challenge in order to better
duplicate the diversity of viruses to which individuals are typically exposed. Because
the resistance described in their study was associated with a dose of
PSC-RANTES lower than that which conferred full protection, they also caution
that waning microbicide levels after application could conceivably provide the
conditions necessary for the emergence of drug resistance.
JVI Accepts, published online ahead of print on 11 March
2009
J. Virol. doi:10.1128/JVI.00055-09
Selection of SHIV resistant to a vaginal microbicide in
macaques
Dawn M. Dudley, Jennifer L. Wentzel, Matthew S. Lalonde,
Ronald S. Veazey, and Eric J. Arts
Department of Molecular Biology and Microbiology, Division
of Infectious Diseases, Department of Medicine, Department of Biochemistry,
Case Western Reserve University, Cleveland, OH, 44106; Department of Pathology,
Tulane National Primate Research Center, Tulane University Health Sciences
Center, Covington, LA
Abstract
PSC-RANTES binds to CCR5, inhibits HIV-1 entry, and has been
shown to protect rhesus macaques as a vaginal microbicide from SHIVSF162-p3
infection in a dose-dependent manner. In this study, env gene sequences from
SHIVSF162-p3-infected rhesus macaques treated with PSC-RANTES were analyzed for
possible drug escape variants. Two specific mutations located in the V3 region
of gp120 (K315R) and C-helical domain of gp41 (N640D) were identified in a
macaque (m584) pre-treated with a 100 µM dose of PSC-RANTES. These two env
mutations were found throughout infection (through week 77) but were only found
at low frequencies in the inoculating SHIVSF162-p3 stock and in the other
SHIVSF162-p3-infected macaques. HIV-1 env genes from the m584 macaque and from
inoculating SHIVSF162-p3 were cloned into an HIV-1 backbone. Increases in IC50
values to PSC-RANTES with envm584 was modest (7-fold) and most pronounced in
cells expressing rhesus macaque as compared to human CCR5. Nonetheless, virus
harboring the envm584, unlike inoculating envp3, could replicate even at the
highest tissue culture PSC-RANTES concentrations (100 nM). Dual virus
competitions revealed a dramatic increase in fitness of chimeric virus
containing the envm584 (K315R/N640D) over that containing the envp3, but again,
only in rhesus CCR5-expressing cells. This study is the first to describe the
immediate selection and infection of a drug-resistant SHIV variant in the face
of a protective vaginal microbicide, PSC-RANTES. This rhesus
CCR5-specific/PSC-RANTES-resistance selection is particularly alarming given
the relative homogeneity of the SHIVSF162-p3 stock as compared to the potential
exposure to a heterogeneous HIV-1 population in human transmission.
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