Previous Article | Next Article 
Journal of Virology, January 2001, p. 1048-1053, Vol. 75, No. 2
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.2.1048-1053.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Susceptibility of the Porcine Endogenous Retrovirus
to Reverse Transcriptase and Protease Inhibitors
Shoukat H.
Qari,1
Saema
Magre,2
J. Gerardo
García-Lerma,1
Althaf I.
Hussain,1
Yasuhiro
Takeuchi,2
Clive
Patience,2
Robin A.
Weiss,2 and
Walid
Heneine1,*
HIV and Retrovirology Branch, Division of
AIDS, STD, and TB Laboratory Research, Centers for Disease Control
and Prevention, Atlanta, Georgia,1 and
Wohl Virion Centre, Windeyer Institute, University College
London, London, United Kingdom2
Received 28 August 2000/Accepted 25 October 2000
 |
ABSTRACT |
Porcine xenografts may offer a solution to the shortage of human
donor allografts. However, all pigs contain the porcine endogenous retrovirus (PERV), raising concerns regarding the transmission of PERV
and the possible development of disease in xenotransplant recipients.
We evaluated 11 antiretroviral drugs licensed for human
immunodeficiency virus type 1 (HIV-1) therapy for their activities
against PERV to assess their potential for clinical use. Fifty and
90% inhibitory concentrations (IC50s and
IC90s, respectively) of five nucleoside reverse
transcriptase inhibitors (RTIs) were determined enzymatically for PERV
and for wild-type (WT) and RTI-resistant HIV-1 reference isolates. In a
comparison of IC50s, the susceptibilities of PERV RT to
lamivudine, stavudine, didanosine, zalcitabine, and zidovudine were
reduced >20-fold, 26-fold, 6-fold, 4-fold, and 3-fold, respectively,
compared to those of WT HIV-1. PERV was also resistant to nevirapine.
Tissue culture-based, single-round infection assays using
replication-competent virus confirmed the relative sensitivity of PERV
to zidovudine and its resistance to all other RTIs. A Gag
polyprotein-processing inhibition assay was developed and used to
assess the activities of protease inhibitors against PERV. No
inhibition of PERV protease was seen with saquinavir, ritonavir,
indinavir, nelfinavir, or amprenavir at concentrations >200-fold the
IC50s for WT HIV-1. Thus, following screening of many
antiretroviral agents, our findings support only the potential clinical
use of zidovudine.
| |
TEXT |
The use of transplants from animal
origin offers a potential solution to the limited supply of human
organs and tissues. Pigs are presently the preferred source for
xenotransplantation for a variety of practical and safety reasons,
including availability, comparable organ size, and possibility of
genetic manipulation to overcome rejection by the recipient's immune
system. Clinical trials involving pig xenografts have included (i)
perfusion with pig livers or porcine hepatocytes as a bridging strategy
for hepatic failure, (ii) the use of pancreatic islet cells as a
treatment for chronic diabetes, and (iii) the implantation of fetal
neuronal tissue as a therapy for Parkinson's and Huntington's
diseases (5, 10, 11, 17, 47, 39). Proposed future
therapies include the use of solid organs from pigs.
A major concern surrounding porcine xenotransplantation is the exposure
of xenograft recipients to the porcine endogenous retrovirus (PERV).
PERV is a C-type retrovirus that is permanently integrated, in multiple
copies, in the pig genome (23, 33). Infectious PERV
particles are released from a variety of porcine cells, including
peripheral blood mononuclear cells, endothelial cells, pancreatic
islets, and some kidney cell lines (22, 26, 33, 50). PERV
has three envelope classes (PERV-A, -B, and -C) that have been shown to
have distinct receptor specificity and in vitro tropism (43,
51). However, their protease and reverse transcriptase (RT)
sequences are indistinguishable (1).
Concerns regarding transmission of PERV to humans were heightened when
PERV derived from either porcine cell lines or primary cells was shown
to infect some human cells (26, 33, 50). In a severe
combined immunodeficiency mouse model, Laan et al. demonstrated
recently the ability of porcine pancreatic islet cells to transmit PERV
infection, providing the first evidence for a xenogenic PERV infection
(22). However, evidence of PERV infection has not been
seen thus far in retrospective studies of patients who had received
either extracorporeal porcine splenic, liver, or kidney perfusion;
porcine skin grafts; or porcine pancreatic islet cell transplants
(19, 24, 31, 32, 39). While reassuring, these data do not
fully define risks of PERV transmission to exposed humans because of
our limited ability to extrapolate these findings to other types of
xenotransplants, particularly those involving whole organs. The risk
that any xenograft recipients may become infected with PERV is likely
to be a function of several factors associated with the xenograft
(e.g., cellular or solid organs, transgenic or nontransgenic animal,
duration of the xenograft, etc.), the xenotransplantation technique,
and the recipient's characteristics (e.g., immunosuppression, levels
of xenoantibody, etc.).
Infections with C-type retroviruses have been associated with different
outcomes, ranging from benign to neoplastic and neurologic diseases
(8). While the risk that PERV-infected persons will develop disease and will require antiretroviral therapy is still unknown, it is prudent to identify drugs that are active against PERV.
Such drugs will be available should a need to treat PERV-infected persons arises.
In this study, we have evaluated the susceptibility of PERV to 11 inhibitors licensed by the U.S. Food and Drug Administration for the
treatment of human immunodeficiency virus type 1 (HIV-1). These
inhibitors included six RT inhibitors (RTIs) and five protease inhibitors (PIs). We provide evidence of sensitivity of PERV to zidovudine and poor or no susceptibility to all other drugs.
RT susceptibility analysis by an enzymatic assay.
The
susceptibility of PERV RT to the RTIs was evaluated enzymatically in
the Amp-RT assay to determine the 50 and 90% inhibitory concentrations
(IC50s and IC90s, respectively) (20,
52). Results obtained in triplicate were compared with
those determined from reference wild-type (WT) and drug-resistant
HIV-1 isolates using methodologies previously described (15,
49). The RTIs tested were nevirapine; the triphosphorylated
nucleoside analogs of zidovudine (AZT-TP), lamivudine
(2',3'-deoxy-3'-thiacytidine; 3TC-TP), zalcitabine (dideoxycytosine
[ddC]-TP), stavudine (2',3'-didehydro-3'-deoxythymidine; d4T-TP), and
the active form of didanosine (dideoxyadenine [ddA]-TP). The ratios
of nucleoside analogs to their corresponding deoxynucleoside triphosphates (dNTPs) in the RT reaction of the Amp-RT assay varied, with 15 µM dTTP being used for reaction mixtures containing AZT-TP or
d4T-TP, 5 µM dCTP being used for mixtures containing 3TC-TP or
ddC-TP, and 5 µM dATP being used for mixtures containing dd-ATP along
with 20 µM concentrations of each of the other three dNTPs. For
nevirapine reactions, a 20 µM dNTP mixture was used. AZT-TP was
obtained from Moravek Biochemicals, Inc. (Brea, Calif.), ddC-TP and
ddA-TP were obtained from Sigma Chemical Co. (St. Louis, Mo.), d4T-TP
was kindly provided by Anne-Mieke Vandamme (Rega Institute for Medical
Research and University Hospitals, Leuven, Belgium), and 3TC-TP and
nevirapine were provided by R. F. Schinazi (Veterans Affairs Medical
Center and Emory University, Atlanta, Ga.).
Two sources of PERV were used, the first from culture supernatant of a
porcine embryonic kidney cell line (PK15) and the second from
PERV-infected human embryonic kidney cell line 293 (PERV-293) (33). PK15 and PERV-293 cells, which release both PERV-A
and -B (23), were maintained in minimal essential medium
by using standard tissue culture techniques. Three HIV-1 isolates
derived from molecular infectious clones containing WT RT,
HIV-1SUM9 (40), and
xxHIV-1LAI (30) or
HIV-1xxBRUpitt (37) were used as reference WT
HIV-1 isolates. HIV-1 isolates derived from three molecular infectious
clones containing one to five multidideoxynucleoside-resistant (MDR) mutations were used as reference viruses that have different levels of resistance: HIV-1SUM8 (Q151M) has low-level
resistance, and HIV-1SUM12 (F77L, F116V, Q151M) and
HIV-1SUM13 (A62V, V75I, F77L, F116Y, Q151M) have higher
levels of resistance (40, 48).
The IC
50s and IC
90s of RTIs for PERV, WT HIV-1,
and drug-resistant HIV-1 are shown in Table
1. As expected, all the MDR HIV-1
isolates had reduced susceptibility to all the nucleoside analogs.
The
level of resistance was lower with HIV-1
SUM8 containing
Q151M
than with HIV-1
SUM12 and HIV-1
SUM13,
which contained additional
MDR mutations. These data are consistent
with previous findings
on these viruses (
40,
48). Table
1
also shows that, in comparison
to WT HIV-1, both PERV isolates had
reduced susceptibilities to
all five nucleoside RTIs. However, the
level of resistance varied
among the drugs. No activity was seen with
lamivudine, while high-level
resistance was observed with stavudine.
The susceptibility of
PERV to both zalcitabine and didanosine was also
reduced compared
to that of WT HIV-1. The highest activity against PERV
RT was
observed with zidovudine, with which there was only a threefold
difference in IC
50 from that for WT HIV-1. This level of
resistance
was lower than that of HIV-1
SUM8 (Q151M).
View this table:
[in this window]
[in a new window]
|
TABLE 1.
Susceptibility of the enzymatic activity of PERV RT to
various RTIs and comparison with WT and
drug-resistant HIV-1a
|
|
No significant differences were found in the drug susceptibilities of
PERV-PK15 and PERV-293, except to d4T-TP, with which
a twofold
difference in IC
50s was observed. This likely reflected
assay variability rather than inherent phenotypic changes between
the
two viral preparations. Similarly, changes were also seen
for the WT
HIV-1 isolates, with IC
50s for 3TC-TP and ddA-TP differing
by 1.5-fold and 2.7-fold,
respectively.
We also tested PERV RT for susceptibility to nevirapine, a
nonnucleoside inhibitor of HIV-1 RT. One Amp-RT assay with 50 µM
nevirapine was used in this screening. This concentration of nevirapine
completely inhibits WT HIV-1 RT activity (IC
50, 4 µM)
(
49).
In contrast to what was observed with WT HIV-1, no
inhibition
of PERV RT was observed in this test, indicating lack of
activity
of nevirapine on PERV RT (data not
shown).
RT susceptibility analysis by cell culture-based infectivity
assays.
Susceptibility of PERV and HIV-1 to RTIs was evaluated on
the CD4-expressing human rhabdomyosarcoma cell line (RD-CD4)
(7). This cell line is able to support the infection and
replication of both PERV and HIV-1. In situ focus-forming assays were
carried out using PERV-B lacZ pseudotype (see below) and the
HIV-1LAI isolate. For IC50 and IC90
determinations for WT HIV-1, reduction in the number of foci producing
p24 was quantitated (41), while for PERV a lacZ
pseudotype was generated and reduction in the number of foci producing
-galactosidase was quantitated (44).
To generate PERV-B
lacZ pseudotypes, the human embryonic
kidney cell line 293/PERV-B, persistently infected with PERV-B
(
43),
was transduced with the MFGnlslacZ vector using a
helper-free
vector bearing gibbon ape leukemia virus envelopes
(
35). MFGnlslacZ
is a murine leukemia virus (MuLV)-based
vector containing functional
long terminal repeats and a
packaging
signal sequence, as well
as a
lacZ marker gene. It encodes
-galactosidase and no MuLV
gag,
pol, or
env gene products (
13). A cell population of
which
the majority of cells (>95%) contain MFGnlslacZ was established
and named 293/PERV-B/lacZ. These cells produce a mixture of
replication-competent
PERV-B and PERV pseudotypes which contain the
MFGnlslacZ genome
(
45). This viral mixture is referred to
as PERV-lacZ pseudotypes.
After overnight incubation of these cells,
the supernatant containing
PERV-lacZ was filtered (filter pore size,
0.45 µm) and used for
drug susceptibility
testing.
RD-CD4 cells were seeded in 24-well plates at 8 × 10
4
cells/well in Dulbecco modified Eagle medium with 10% fetal calf serum
and incubated overnight. The culture medium was replaced with
450 µl
of HIV-1 or PERV-lacZ pseudotype supernatants which had
been diluted in
Opti-MEM (Gibco-BRL) to have approximately 150
and 60 focus-forming
units of virus per well, respectively. After
incubation for 1 h,
virus inocula were removed and 1 ml of medium
containing RTIs was
added. Three days later, cells were fixed
and stained for expression of
-galactosidase (
44) or HIV-1
p24 (HIV-1
LAI)
(
41).
The data from the culture-based assays are shown in Table
2 and are in agreement with the data from
the enzymatic assays.
This testing also demonstrated that, with the
exception of zidovudine,
which had an IC
50 close to that
for WT HIV-1, all other drugs
had little or no activity against PERV.
The level of resistance
of PERV to zalcitabine and didanosine was
higher in these assays
than in the enzymatic assays.
Protease susceptibility analysis.
Susceptibility of PERV to
PIs was assessed in a Gag polyprotein-processing inhibition assay, as
previously described for MuLV (2), in which the presence
of cleaved PERV Gag proteins in PERV-293 cells was determined by
Western blot analysis following PI treatment. PIs tested included
indinavir, nelfinavir, saquinavir, ritonavir, and amprenavir. For WT
HIV-1 PI tests, OM-10.1 cells were used. These cells contain a single
integrated WT HIV-1 provirus which is induced after treatment with 20 U
of tumor necrosis factor alpha per ml (4). HIV-1
production in 3 × 106 OM-10.1 cells was induced at
the time of addition of PIs (0.001 to 10 µg/ml). Cells were incubated
for 24 h and harvested. One to 25 µg of each PI per ml was added
to 3 × 106 PERV-293 cells in 10 ml of growth medium.
After incubation for 3 days, PERV-293 cells were harvested by
trypsinization. Aliquots of harvested OM-10.1 and PERV-293 cells were
checked for viability by trypan blue exclusion. Protein concentration
of homogenized lysates was determined with a BCA protein assay kit
(Pierce Chemical Co., Rockford, Ill.).
Ten micrograms of OM-10.1 and 15 µg of PERV-293 whole-cell lysate
proteins were electrophoresed and electroblotted (
27).
HIV-1 p24 monoclonal antibody (
16), which reacts with p24,
p55,
and several intermediates, was used (1:600 dilution) to react
with
the OM-10.1 blots. Anti-simian sarcoma-associated virus p29
polyclonal
serum (Quality Biotech, Camden, N.J.), which cross-reacts
with the
processed PERV Gag p30 and its precursor p55, was used
(1:200 dilution)
to react with PERV-293 blots (
27). Proteins
were
visualized by chemiluminescence using an ECL Western blot
detection
reagent (Amersham Pharmacia Biotech, Piscataway, N.J.).
As expected, WT HIV-1 was found to be sensitive to all five PIs tested.
No p24 Gag protein was detected in OM-10.1 cell cultures
treated with 1 µg of PIs per ml, while untreated cells had a detectable
p24 band
(Fig.
1). In addition, accumulation of
the unprocessed
p55 and other intermediates (p48 and p42) was also
observed in
these lysates (Fig.
1). No p24 or its precursors (p55, p48,
and
p42) were detected in the cells which were not stimulated with
tumor necrosis factor alpha (data not shown). In contrast, no
difference in p30 reactivity was seen among PERV-293 cells that
were
treated with a concentration up to 25 µg/ml (the highest
concentration tested) of indinavir, nelfinavir, saquinavir, ritonavir,
or amprenavir, which represented a 233- to 6,520-fold increase
in the
IC
50s reported for WT HIV-1 (
3,
9).
Representative
results are shown for nelfinavir and indinavir in Fig.
1. Thus,
our results indicate that PERV is resistant to all five PIs.
View larger version (24K):
[in this window]
[in a new window]
|
FIG. 1.
Susceptibility of HIV-1 and PERV to PIs by a Gag
protein-processing inhibition assay. (A) Immunoblot of HIV-1-infected
OM-10.1 cells treated with different concentrations of indinavir and
nelfinavir (first lane, no treatment). Positions of Gag proteins are
indicated. (B) Immunoblot of PERV-293 cells untreated (first lane) and
treated with different concentrations of nelfinavir and indinavir.
|
|
Sequence comparison.
To better understand the basis of the
susceptibility of PERV to the antiretrovirals tested in this study, we
compared the amino acid sequences of the pol regions of PERV
derived from PK15 cells (GenBank accession numbers U77599 and AF038601)
and HIV-1 (subtype B) (accession number M38432). Alignment was performed with the multiple alignment construction and analysis workbench program (38), with introduction of minimal gaps
to facilitate optimal alignment. The PERV sequence analyzed
corresponded to the HIV-1 protease codons 29 to 99 and RT codons 1 to
291. A portion of the alignment of HIV-1 RT, which harbors codons
associated with drug resistance, with PERV RT is shown in Fig.
2. HIV-1 and PERV were found to share
only about 22.5% amino acid residues in protease or RT, indicating
that these proteins are structurally diverse. The high level of
divergence between both sequences restricts the comparison between RTI
resistance-related mutations of HIV-1 and PERV to only those which are
conserved among all retroviruses. Mutations at two such codons, Q151M
(in the conserved LPQG motif), and M184V (in the polymerase active-site
YMDD motif) are associated with resistance to multiple nucleoside
analogs and to lamivudine, respectively, in HIV-1 and other
lentiviruses. PERV RT, as in WT HIV-1, has a Q at the codon
corresponding to 151, which does not support a role of this residue in
the observed resistance of PERV to several nucleoside analogs. However,
PERV has a V instead of an M at the codon corresponding to 184, which
may likely explain the observed resistance to lamivudine. Verifying the
possible role of this residue in PERV's resistance to lamivudine will
require drug susceptibility analysis of site-specific mutants
containing either an M or a V residue at this site.
View larger version (18K):
[in this window]
[in a new window]
|
FIG. 2.
Alignment of portions of HIV-1 and PERV RT amino acid
sequences. Identical residues are indicated with an asterisk. Four D
residues (codons 110, 113, 185, and 186) and the LPQG motif (codons 149 to 152) conserved among the retroviruses are underlined
(34a). The HIV-1 polymerase active-site YMDD (codons 183 to 186) is in bold. Codons associated with resistance of HIV-1 to
multinucleoside analogs (Q151M), nevirapine (Y181C), lamivudine
(M184V), and zidovudine (T215Y/F) are shaded.
|
|
The observed susceptibility of PERV to zidovudine may not be surprising
since this nucleoside analog has been shown to have
a broad range of
activity against several retroviruses (
25,
28),
including MuLV and feline leukemia virus, two C-type iruses
which
are closely related to PERV (
36,
46). Our sequence
analysis
indicated that both MuLV (accession number
U13766) and feline
leukemia virus (accession number
AF052723) share significant
homology
(79 and 70% amino acid residues, respectively) with PERV
RT.
Accordingly, isolates with
lacZ pseudotypes containing
Moloney
MuLV Gag-Pol were tested by our culture-based infectivity assay
and showed a pattern of drug sensitivity similar to that of PERV
(data
not
shown).
As expected, PERV showed no susceptibility to HIV-1-specific PIs,
despite the structural diversity of these compounds and
the use of
concentrations that were several hundredfold higher
than those for WT
HIV-1 (
6,
12,
29). The lack of susceptibility
may be
explained by the structural differences between PERV and
HIV-1
protease, which were found to share only ~22% of their amino
acid
residues.
Structure-dependent binding may also explain the resistance of PERV to
nevirapine, which binds specifically to a hydrophobic
cavity adjacent
to the polymerase active site in HIV-1 RT and
requires close contact
with a tyrosine residue at codon 181 (
21,
42). The observed resistance of PERV RT to nevirapine was,
therefore,
expected and may very likely be due to the absence of this
pocket,
resulting from significant structural
differences.
Our data do, however, demonstrate that some nucleoside RTIs were active
against PERV RT. Zidovudine had the highest level
of activity against
PERV, with IC
50s only ~3-fold those for WT
HIV-1 but well
within the achievable concentration of zidovudine
in vivo, which has a
maximum concentration of drug in serum of
3.4 µM (
14).
These in vitro data are promising and support the
clinical use of
zidovudine in PERV-infected persons. Prophylactic
use of zidovudine has
also been successful in reducing transmission
of HIV-1 in infants born
to HIV-1-infected mothers as well as
in recipients of needle-stick
injuries from HIV-1-infected source
patients (reviewed in references
18 and
53). Therefore, our
data on zidovudine
may also support evaluating the use of this
drug as a prophylactic in
persons who will be transiently exposed
to pig tissues, such as in
extracorporeal perfusions with pig
livers or hepatocytes. The need for
such a prophylactic will,
however, depend on the risks of PERV
transmission from these procedures.
The assays developed and used in
this study also provide tools
for identifying novel PIs or RTIs that
are active against
PERV.
| |
ACKNOWLEDGMENTS |
We gratefully acknowledge Sal Butera (CDC, Atlanta, Ga.) for
providing OM-10.1 cells and Hiroaki Mitsuya (National Cancer Institute,
Bethesda, Md.) for HIV-1 clones (HIV-1SUM9,
HIV-1SUM8, HIV-1SUM12, and
HIV-1SUM13). HIV-1 p24 monoclonal antibody, some RTIs, and PIs were obtained from the AIDS Research and Reference Reagent Program, Division of AIDS, NIAID, NIH (Rockville, Md.).
The research at the Wohl Virion Centre was supported by the United
Kingdom Medical Research Council.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: HIV and
Retrovirology Branch, CDC, MS G-19, 1600 Clifton Rd., Atlanta, GA
30333. Phone: (404) 639-0218. Fax: (404) 639-1174. E-mail:
WMH2{at}CDC.GOV.
| |
REFERENCES |
| 1.
|
Akiyoshi, D. E.,
M. Denaro,
H. Zhu,
J. L. Greenstein,
P. Banerjee, and J. A. Fishman.
1998.
Identification of a full-length cDNA for an endogenous retrovirus of miniature swine.
J. Virol.
72:4503-4507[Abstract/Free Full Text].
|
| 2.
|
Black, P. L.,
M. B. Downs,
M. G. Lewis,
M. A. Ussery,
G. B. Dreyer,
S. R. Petteway, and D. M. Lambert.
1993.
Antiretroviral activities of protease inhibitors against murine leukemia virus and simian immunodeficiency virus in tissue culture.
Antimicrob. Agents Chemother.
37:71-77[Abstract/Free Full Text].
|
| 3.
|
Boden, D., and M. Markowitz.
1998.
Resistance to human immunodeficiency virus type 1 protease inhibitors.
Antimicrob. Agents Chemother.
42:2775-2783[Free Full Text].
|
| 4.
|
Butera, S. T.,
B. D. Roberts,
L. Lam,
T. Hodge, and T. M. Folks.
1994.
Human immunodeficiency virus type 1 RNA expression by four chronically infected cell lines indicates multiple mechanisms of latency.
J. Virol.
68:2726-2730[Abstract/Free Full Text].
|
| 5.
|
Chari, R. S.,
B. Collins,
J. Magee,
J. DiMaio,
A. Kirk,
R. Harland,
R. McCann,
J. Platt, and W. Meyers.
1994.
Treatment of hepatic failure with ex vivo pig-liver perfusion followed by liver transplantation.
N. Engl. J. Med.
331:234-237[Free Full Text].
|
| 6.
|
Chen, Z.,
Y. Li,
H. Schock,
D. Hall,
E. Chen, and L. Kuo.
1995.
Three-dimensional structure of a mutant HIV-1 protease displaying cross-resistance to all protease inhibitors in clinical trials.
J. Biol. Chem.
15:21433-21436.
|
| 7.
|
Clapham, P. R.,
D. Blanc, and R. Weiss.
1991.
Specific cell surface requirements for the infection of CD4-positive cells by human immunodeficiency virus types 1 and 2 and by simian immunodeficiency virus.
Virology
181:2703-2715.
|
| 8.
|
Coffin, J. M.
1996.
Retroviridae: the viruses and their replication, p. 1767-1830.
In
B. N. Fields, D. M. Knipe, and P. M. Howley (ed.), Fields virology, 3rd ed. Lippincott-Raven Publishers, Philadelphia, Pa.
|
| 9.
|
Craig, J. C.,
I. B. Duncan,
D. Hockley,
C. Grief,
N. Roberts, and J. S. Mills.
1991.
Antiviral properties of Ro 31-8959, an inhibitor of human immunodeficiency virus (HIV) proteinase.
Antivir. Res.
16:295-305[CrossRef][Medline].
|
| 10.
|
Cramer, D. V.
1995.
The use of xenografts for acute hepatic failure.
Transplant. Proc.
27:80[Medline].
|
| 11.
|
Deacon, T.,
J. Schumacher,
J. Dinsmore,
C. Thomas,
P. Palmer,
S. Kott,
A. Edge,
D. Penny,
S. Kassissieh,
P. Dempsey, and O. Isacson.
1997.
Histological evidence of fetal pig neural cell survival after transplantation into a patient with Parkinson's disease.
Nat. Med.
3:350-353[CrossRef][Medline].
|
| 12.
|
Erickson, J. W., and M. A. Eissenstat.
1999.
HIV protease as a target for the design of antiviral agents for AIDS, p. 1-45.
In
B. M. Dunn (ed.), Proteases of infectious agents. Academic Press, New York, N.Y.
|
| 13.
|
Ferry, N.,
O. Duplessis,
D. Houssin,
O. Danos, and J. M. Heard.
1991.
Retroviral-mediated gene transfer into hepatocytes in vivo.
Proc. Natl. Acad. Sci. USA
88:8377-8381[Abstract/Free Full Text].
|
| 14.
|
Flexner, C., and C. Hendrix.
1997.
Pharmacology of antiretroviral agents, p. 479-493.
In
V. T. DeVito, Jr., S. Hellman, and S. A. Rosenberg (ed.), AIDS: biology, diagnosis, and prevention. Lippincott-Raven Publishers, Philadelphia, Pa.
|
| 15.
|
Garcia-Lerma, J.,
R. F. Schinazi,
A. Juodawlkis,
V. Soriano,
Y. Lin,
K. Tatti,
D. Rimland,
T. M. Folks, and W. Heneine.
1999.
A rapid non-culture-based assay for clinical monitoring of phenotypic resistance of human immunodeficiency virus type 1 to lamivudine (3TC).
Antimicrob. Agents Chemother.
43:264-270[Abstract/Free Full Text].
|
| 16.
|
Gorny, M. K.,
V. Gianakakos,
S. Sharpe, and S. Zolla-Pazner.
1989.
Generation of human monoclonal antibodies to human immunodeficiency virus.
Proc. Natl. Acad. Sci. USA
86:1624-1628[Abstract/Free Full Text].
|
| 17.
|
Groth, C. G.,
O. Korsgren,
A. Tibell,
J. Tollemar,
E. Moller,
J. Bolinder,
J. Ostman,
F. P. Reinholt,
C. Hellerstrom, and A. Andersson.
1994.
Transplantation of porcine fetal pancreas to diabetic patients.
Lancet
344:1402-1404[CrossRef][Medline].
|
| 18.
|
Henderson, D. K.
1999.
Postexposure chemoprophylaxis for occupational exposures to the human immunodeficiency virus.
JAMA
281:931-936[Free Full Text].
|
| 19.
|
Heneine, W,
A. Tibell,
W. M. Switzer,
P. Sandstrom,
G. V. Rosales,
A. Mathews,
O. Korsgren,
L. E. Chapman,
T. M. Folks, and C. G. Groth.
1998.
No evidence of infection with porcine endogenous retrovirus in recipients of porcine islet-cell xenografts.
Lancet
352:695-699[CrossRef][Medline].
|
| 20.
|
Heneine, W.,
S. Yamamoto,
W. M. Switzer,
T. J. Spira, and T. M. Folks.
1995.
Detection of reverse transcriptase by a highly sensitive assay in sera from persons infected with human immunodeficiency virus type 1.
J. Infect. Dis.
171:1210-1216[Medline].
|
| 21.
|
Kohlstaedt, L. A.,
J. Wang,
J. M. Friedman,
P. A. Rice, and T. A. Steitz.
1992.
Crystal structure at 3.5 Å resolution of HIV-1 reverse transcriptase complexed with an inhibitor.
Science
256:1783-1790[Abstract/Free Full Text].
|
| 22.
|
Laan, L. J. W.,
C. Lockey,
B. C. Griffeth,
F. S. Frasier,
C. A. Wilson,
D. E. Onions,
B. J. Hering,
Z. Long,
E. Otto,
B. E. Torbett, and D. R. Salomon.
2000.
Infection by porcine endogenous retrovirus after islet xenotransplantation in SCID mice.
Nature
407:501-504.
|
| 23.
|
Le Tissier, P.,
J. P. Stoye,
Y. Takeuchi,
C. Patience, and R. A. Weiss.
1997.
Two sets of human-tropic pig retroviruses.
Nature
389:681-682[CrossRef][Medline].
|
| 24.
|
Levy, M. F.,
J. Crippin,
S. Sutton,
G. Netto,
J. McCormack,
T. Curiel,
R. M. Goldstein,
J. Newman,
T. Gonwa,
J. Banchereau,
L. Diamond,
G. Byrne,
J. Logan, and G. B. Klintmalm.
2000.
Liver allotransplantation after extracorporeal hepatic support with transgenic (hCD55/hCD59) porcine livers: clinical results and lack of pig-to-human transmission of the porcine endogenous retrovirus.
Transplantation
69:272-280[CrossRef][Medline].
|
| 25.
|
Macchi, B.,
I. Faraoni,
J. Zhang,
S. Grelli,
C. Favalli,
A. Mastino, and E. Bonmassar.
1997.
AZT inhibits the transmission of human T cell leukaemia/lymphoma virus type I to adult peripheral blood mononuclear cells in vitro.
J. Gen. Virol.
78:1007-1016[Abstract].
|
| 26.
|
Martin, U.,
V. Kiessig,
J. Blusch,
A. Haverich,
K. von der Helm,
T. Herden, and G. Steinhoff.
1998.
Expression of pig endogenous retrovirus by primary porcine endothelial cells and infection of human cells.
Lancet
352:692-694[CrossRef][Medline].
|
| 27.
|
Matthews, A. L.,
J. Brown,
W. Switzer,
T. M. Folks,
W. Heneine, and P. A. Sandstrom.
1999.
Development and validation of a Western immunoblot assay for detection of antibodies to porcine endogenous retrovirus.
Transplantation
67:939-943[CrossRef][Medline].
|
| 28.
|
Mitsuya, H., and S. Broder.
1998.
Inhibition of infectivity and replication of HIV-2 and SIV in helper T-cells by 2',3'-dideoxynucleosides in vitro.
AIDS Res. Hum. Retroviruses
4:107-113.
|
| 29.
|
Nascimbeni, M.,
C. Lamotte,
G. Peytavin,
R. Farinotti, and F. Clavel.
1999.
Kinetics of antiviral activity and intracellular pharmacokinetics of human immunodeficiency virus type 1 protease inhibitors in tissue culture.
Antimicrob. Agents Chemother.
43:2629-2634[Abstract/Free Full Text].
|
| 30.
|
Nguyen, M. H.,
R. F. Schinazi,
C. Shi,
N. M. Goudgaon,
P. M. McKenna, and J. W. Mellors.
1994.
Resistance of human immunodeficiency virus type 1 to acyclic 6-phenylselenenyl- and 6-phenylthiopyrimidines.
Antimicrob. Agents Chemother.
38:2409-2414[Abstract/Free Full Text].
|
| 31.
|
Paradis, K.,
G. Langford,
Z. Long,
W. Heneine,
P. Sandstrom,
W. M. Switzer,
L. E. Chapman,
C. Lockey,
D. Onions, and E. Otto.
1999.
Search for cross-species transmission of porcine endogenous retrovirus in patients treated with living pig tissue.
Science
285:1236-1241[Abstract/Free Full Text].
|
| 32.
|
Patience, C.,
G. S. Patton,
Y. Takeuchi,
R. A. Weiss,
M. O. McClure,
L. Rydberg, and M. E. Breimer.
1998.
No evidence of pig DNA or retroviral infection in patients with short-term extracorporeal connection to pig kidneys.
Lancet
352:699-701[CrossRef][Medline].
|
| 33.
|
Patience, C.,
Y. Takeuchi, and R. A. Weiss.
1997.
Infection of human cells by an endogenous retrovirus of pigs.
Nat. Med.
3:282-286[CrossRef][Medline].
|
| 34.
|
Pettit, S. C.,
R. Sanchez,
T. Smith,
R. Wehbie,
D. Derse, and R. Swanstrom.
1998.
HIV type 1 protease inhibitors fail to inhibit HTLV-I Gag processing in infected cells.
AIDS Res. Hum. Retroviruses
14:1007-1014[Medline].
|
| 34a.
|
Poch, O.,
I. Sauvaget,
M. Delarue, and N. Tordo.
1989.
Identification of four conserved motifs among the RNA-dependent polymerase encoding elements.
EMBO J.
8:3867-3874[Medline].
|
| 35.
|
Porter, C. D.,
M. K. Collins,
C. S. Tailor,
M. H. Parkar,
F. L. Cosset,
R. A. Weiss, and Y. Takeuchi.
1996.
Comparison of efficiency of infection of gene therapy target cells via four different retroviral receptors.
Hum. Gene Ther.
7:913-919[Medline].
|
| 36.
|
Powell, S. K.,
M. Artlip,
M. Kaloss,
S. Brazinski,
R. Lyons,
G. J. McGarrity, and E. Otto.
1999.
Efficacy of antiretroviral agents against murine replication-competent retrovirus infection in human cells.
J. Virol.
73:8813-8816[Abstract/Free Full Text].
|
| 37.
|
Schinazi, R. F.,
A. McMillan,
D. Cannon,
R. Mathis,
R. M. Lloyd,
A. Peck,
J. P. Sommadossi,
M. St. Clair,
J. Wilson,
P. Furman,
G. Painter,
W. Choi, and D. C. Liotta.
1992.
Selective inhibition of human immunodeficiency viruses by racemates and enantiomeres of cis-5-fluoro-1-[2-(hydroxymethyl)-1,3-oxathiolan-5-yl]cytosine.
Antimicrob. Agents Chemother.
36:2423-2431[Abstract/Free Full Text].
|
| 38.
|
Schuler, G. D.,
S. F. Altschul, and D. J. Lipman.
1991.
A workbench for multiple alignment construction and analysis.
Proteins Struct. Funct. Genet.
9:180-190[CrossRef][Medline].
|
| 39.
|
Schumacher, J. M.,
S. A. Ellias,
E. P. Palmer,
H. S. Kott,
J. Dinsmore,
P. K. Dempsey,
A. J. Fischman,
C. Thomas,
R. G. Feldman,
S. Kassissieh,
R. Raineri,
C. Manhart,
D. Penney,
J. S. Fink, and O. Isacson.
2000.
Transplantation of embryonic porcine mesencephalic tissue in patients with PD.
Neurology
54:1042-1050[Abstract/Free Full Text].
|
| 40.
|
Shirasaka, T.,
M. F. Kavlick,
T. Ueno,
W. Y. Gao,
E. Kojima,
M. L. Alcaide,
S. Chokekijchai,
B. M. Roy,
E. Arnold,
R. Yarchoan, and H. Mitsuya.
1995.
Emergence of human immunodeficiency virus type 1 variants with resistance to multiple dideoxynucleosides in patients receiving therapy with dideoxynucleosides.
Proc. Natl. Acad. Sci. USA
92:2398-2402[Abstract/Free Full Text].
|
| 41.
|
Simmons, G.,
A. McKnight,
Y. Takeuchi,
H. Hoshino, and P. R. Clapham.
1995.
Cell to cell fusion, but not virus entry in macrophages by T-cell tropic HIV-1 strains: a V3 loop determined restriction.
Virology
209:696-700[CrossRef][Medline].
|
| 42.
|
Smerdon, S. J.,
J. Jager,
J. Wang,
L. A. Kohlstaedt,
A. J. Chirino,
J. M. Friedman,
P. A. Rice, and T. A. Steitz.
1994.
Structure of the binding site for nonnucleoside inhibitors of the reverse transcriptase of human immunodeficiency virus type 1.
Proc. Natl. Acad. Sci. USA
91:3911-3915[Abstract/Free Full Text].
|
| 43.
|
Takeuchi, Y.,
C. Patience,
S. Magre,
R. A. Weiss,
P. T. Banerjee,
P. LeTissier, and J. P. Stoye.
1998.
Host-range and interference studies on three classes of pig endogenous retrovirus.
J. Virol.
72:9986-9991[Abstract/Free Full Text].
|
| 44.
|
Takeuchi, Y.,
F. L. Cosset,
P. J. Lachmann,
H. Okada,
R. A. Weiss, and M. K. L. Collins.
1994.
Type C retroviral inactivation by human complement is determined by both the viral genome and the producer cell.
J. Virol.
68:8001-8007[Abstract/Free Full Text].
|
| 45.
|
Takeuchi, Y.,
G. Simpson,
R. G. Vile,
R. A. Weiss, and M. K. L. Collins.
1992.
Retroviral pseudotypes produced by rescue of a Moloney murine leukemia virus vector by C-type, but not D-type, retroviruses.
Virology
186:792-794[CrossRef][Medline].
|
| 46.
|
Tavares, L.,
C. Roneker,
K. Johnston,
S. N. Lehrman, and F. de Noronha.
1987.
3'-Azido-3'-deoxythymidine in feline leukemia virus-infected cats: a model for therapy and prophylaxis of AIDS.
Cancer Res.
47:3190-3194[Abstract/Free Full Text].
|
| 47.
|
Tibell, A.,
C. G. Groth,
E. Moller,
O. Korsgren,
A. Andersson, and C. Hellerstrom.
1994.
Pig-to-human islet transplantation in eight patients.
Transplant. Proc.
26:762-763[Medline].
|
| 48.
|
Ueno, T.,
T. Shirasaka, and H. Mitsuya.
1995.
Enzymatic characterization of human immunodeficiency virus type-1 reverse transcriptase resistant to multiple 2',3'-dideoxynucleoside 5'-triphosphates.
J. Biol. Chem.
270:23605-23611[Abstract/Free Full Text].
|
| 49.
|
Vazquez-Rosales, G.,
J. G. Garcia Lerma,
S. Yamamoto,
W. M. Switzer,
D. Havlir,
T. M. Folks,
D. D. Richman, and W. Heneine.
1999.
Rapid screening of phenotypic resistance to nevirapine by direct analysis of HIV type 1 reverse transcriptase activity in plasma.
AIDS Res. Hum. Retroviruses
15:1191-1200[CrossRef][Medline].
|
| 50.
|
Wilson, C. A.,
S. Wong,
J. Muller,
C. E. Davidson,
T. M. Rose, and P. Burd.
1998.
Type C retrovirus released from porcine primary peripheral blood mononuclear cells infects human cells.
J. Virol.
72:3082-3087[Abstract/Free Full Text].
|
| 51.
|
Wilson, C. A.,
S. Wong,
M. VanBrocklin, and M. J. Federspiel.
2000.
Extended analysis of the in vitro tropism of porcine endogenous retrovirus.
J. Virol.
74:49-56[Abstract/Free Full Text].
|
| 52.
|
Yamamoto, S.,
T. M. Folks, and W. Heneine.
1996.
Highly sensitive qualitative and quantitative detection of reverse transcriptase activity: optimization, validation, and comparative analysis with other detection systems.
J. Virol. Methods
61:135-143[CrossRef][Medline].
|
| 53.
|
Zorrilla, C. D.
2000.
Mother-to-child HIV-1 transmission: state of the art and implications for public policy.
P. R. Health Sci. J.
19:29-34[Medline].
|
Journal of Virology, January 2001, p. 1048-1053, Vol. 75, No. 2
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.2.1048-1053.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
This article has been cited by other articles:
-
Shi, M., Wang, X., De Clercq, E., Takao, S., Baba, M.
(2007). Selective Inhibition of Porcine Endogenous Retrovirus Replication in Human Cells by Acyclic Nucleoside Phosphonates. Antimicrob. Agents Chemother.
51: 2600-2604
[Abstract]
[Full Text]
-
Miyagawa, S., Nakatsu, S., Nakagawa, T., Kondo, A., Matsunami, K., Hazama, K., Yamada, J., Tomonaga, K., Miyazawa, T., Shirakura, R.
(2005). Prevention of PERV Infections in Pig to Human Xenotransplantation by the RNA Interference Silences Gene. J Biochem
137: 503-508
[Abstract]
[Full Text]
-
Toro, C., Rodes, B., Mendoza, C. d., Soriano, V., Macchi, B., Balestrieri, E., Mastino, A.
(2003). Lamivudine Resistance in Human T-Cell Leukemia Virus Type 1 May Be Due to a Polymorphism at Codon 118 (V->I) of the Reverse Transcriptase. Antimicrob. Agents Chemother.
47: 1774-1775
[Full Text]
-
Bartosch, B., Weiss, R. A., Takeuchi, Y.
(2002). PCR-based cloning and immunocytological titration of infectious porcine endogenous retrovirus subgroup A and B. J. Gen. Virol.
83: 2231-2240
[Abstract]
[Full Text]
-
Blusch, J. H., Seelmeir, S., von der Helm, K.
(2002). Molecular and Enzymatic Characterization of the Porcine Endogenous Retrovirus Protease. J. Virol.
76: 7913-7917
[Abstract]
[Full Text]
-
Chapman, L. E., Bloom, E. T.
(2001). Clinical Xenotransplantation. JAMA
285: 2304-2306
[Full Text]
Top
Abstract
Text
