Bridging the Gap: Conference Record [Abstract book, International Conference on AIDS (12th: 1998: Geneva, Switzerland)]

12th World AIDS Conference

Abstracts 21192-21195 277

FACSCount. The Ly activation was detected by 3H-thymidine incorporation. IL-2
and slL-2R levels were measured by ELISA.
Results: Patient 1 showed the highest IL-2 level before therapy followed during
next two months by progressive decrease whereas slL-2R levels demonstrated
opposite dynamics. At the beginning of the 3rd month a recovery of IL-2 and slight
decrease of slL-2R were observed. The dynamic of SLP followed that of slL-2R.
Patient 2, 3 and 4 IL-2 levels increased slowly during the first two months and the
highest concentrations were detected in the 3rd month. Patient 2 expressed high
slL-2R level at the start of the treatment correlating with the lowest IL-2 level.
At the same time a steady-state SLP values were observed. In patients 3 and 4
slL-2R were undetectable at start point, despite of the high activation detected by
SLP.
Conclusions: IL-2 levels in supernatants from PHA stimulated lymphocytes
showed inverse correlation to slL-2R and SLP. The highest values were found
3 months after the onset of therapy. slL-2R levels correlated well with the SLP
dynamics. Additionally, no correlation between the levels of IL-2, sll-2R and CD4+,
CD8+ and CD3+ T counts was found.
495*/21192 CD8 + CTL induced in AIDS vaccine evaluation
group phase I trials using canarypox vectors
(ALVAC) encoding multiple HIV gene products
(vCP125, vCP205, vCP300) given with or without
subunit boost
Thomas Evans1, L. Corey2, M.L. Clements-Mann3, K. Weinhold4, R.B. Belshe5,
J.L. Excler6, A.M. Duliege7. 1601 Elmwood Avenue Box 689 Rochester New
York 14642; 2University of Washington, Seattle, WA; 3Johns Hopkins University,
Baltimore, MD; 4Duke University, Durham, NC; 5St. Louis University, St Louis,
MO; 7Chiron Vaccines, Emeryville, CA, USA; 7Pasteur-Merrieux Connaught,
Marnes La Coquett, France
Objectives: The use of live canarypox vectors provides a safe method for eliciting
CTL which are desirable for developing an effective HIV vaccine strategy. We
will present the frequency of CTL induced by canarypox vectors encoding gp160
(vCP125), the gp120 and gp41 TM portion of env, gag, and protease (vCP205),
or the vCP205 plus sequences of pol and nef encoding multiple CTL epitopes
(vCP300).
Methods: CD8 + CTL were measured in one laboratory by stimulating volunteer PBMC using autologous cells infected with matched vaccinia-HIV-encoding
constructs. After 14 days a standard chromium release assay was performed using EBV-transformed cells infected with appropriate gene targets. Positive assays
were defined by lysis that is 10% above control-infected targets and >50% reduced with CD8 + depletion. Assays in ALVAC recipients were performed in AVEG
012B (vCP125), AVEG 022, 022A, 029 (vCP205), and AVEG 026 (vCP300).
Results: Background rates of CTL in control volunteers across all protocols
(n = 73) were less than 8% per gene target tested. Comparative cumulative
frequencies of env/gag CD8 + CTL were: 012B-11/37 (30%, env only), 022-28/59
(47%), 022A-69/118 (58%), 029-14/22 (64%), 026-49/107 (46%). Positive CTL
have been measured for up to 2 years after initial vaccination. Pol and nef CTL
were also induced by vCP300. The CTL in AVEG 029 were induced rapidly using
an accelerated schedule. As protocols vary by the number and timing of assays
performed, the results will be presented according to the effects of the number
of ALVAC doses, the schedule of doses, the frequency of assays performed, and
the effect of subunit boosting.
Conclusion: Canarypox vectors encoding multiple gene products induced
durable CTL in a majority of human volunteers in phase I vaccine trials. The CTL
responses have broad cross-clade killing activity. Canarypox constructs using
different primary isolates env subtypes with improved in vitro expression will
begin phase I testing soon.
2119 A polytope vaccine strategy for the co-delivery of
multiple HIV CD8+ cytotoxic T cells epitopes
Andreas Suhrbier, S.A. Thomson, S.L. Elliot, J. Gardner, T. Woodberry,
L. Mateo. Queensland Institute of Medical Research, Post Office Royal Brisbane
Hospital, QLD 4029, Australia
Background: There is now a considerable body of compelling indirect evidence
that CD8 /i cytotoxic T lymphocytes (CTL) have a role in preventing or limiting
(or even clearing) initial HIV infection and/or progression to AIDS. An ideal HIV
CTL based vaccine would need to be able to effectively co-deliver a large number
of CTL epitopes to cover (i) multiple epitope variants, (ii) epitopes from different
antigens, and (iii) the HLA diversity of a target population. By simultaneously
generating CTL specific for multiple epitope variants and multiple antigens, such
a vaccine might protected against multiple HIV variants and/or may limit expansion of epitope escape variants by having induced CTL activity against other
unchanged epitopes.
Methods: A number of epitope-based CTL vaccines were constructed, in which
DNA coding for multiple minimal conjoined CTL epitopes were delivered using
recombinant vaccinia virus and DNA vaccination. These artificial polyepitope (or

polytope) vaccines thus contained a string of CTL epitopes, which were derived
from different antigens, different pathogens and/or were restricted by different
MHC. To illustrate that the CTL raised by the vaccines were functional in vivo,
polytope vaccinated mice were challenged in multiple challenge systems.
Results: In all the polytope vaccines made, each epitope in each construct
was capable of being processed and presented to CTL. Mice immunised with a
murine polytope-recombinant vaccinia or -DNA vaccine generated primary CTL

responses against each epitope, and were protected in multiple viral and tumour
challenge systems.
Conclusions: The ability of every CTL epitope in a protein containing only CTL
epitopes to be immunogenic, is likely to find application in HIV CTL based vaccine design. In a therapeutic setting, perhaps after drug therapy, a HIV polytope
vaccine may provide long term benefit by strengthening, restoring or generating
poly-specific CTL based immunity. A prophylactic mucosal HIV polytope vaccine
may also limit or even eliminate a primary HIV inoculum.
550*/21194 Potent neutralization of primary HIV-1 isolates by
antisera induced upon immunization with a
recombinant protein expressing the native V1/V2
domain of gp120
Abraham Pinter1, 0. Trochev1, W.J. Honnen1, Z. Wu2, S. Reiken3, M. Lewis4,
S.C. Kayman1. 'Public Health Research Institute, 455 First Avenue, New York,
New York 10016; 2Rockefeller University, New York NY; 3Columbia University
New York NY; 4Henry M. Jackson Foundation, Rockville MD, USA
Objectives: Whereas it is known that some human immune sera possess potent
neutralizing activities for primary viruses, the identity of the target epitopes mediating this neutralization is unknown, and currently available immunogens have
not been able to induce such activities. Recent evidence from our laboratory
suggests that the V1/V2 domain of HIV-1 gp120 contains epitopes that are potent
neutralization targets for macrophage-tropic HIV-1 isolates. The objectives of this
study were to elicit V1A/2-specific antibodies by immunization with a recombinant
protein that expressed the isolated V1/V2 domain and to determine the breadth
and potency of HIV-1 neutralization by these antibodies.
Design and Methods: The V1/V2 domain of a clinical HIV-1 isolate (Case A2)
was expressed as a fusion glycoprotein in CHO cells. The protein was purified by
affinity chromatography on Ni-NTA columns, utilizing a His6 tag incorporated into
the carrier sequence. A number of rats and pigtailed macaques were immunized
with the purified protein in the presence of Ribi RAS adjuvants at initial doses of
25 / cg/kg. followed by boosts with antigen concentrations of 5 / g/kg.
Results: Both rats and monkeys immunized with the purified Case A2-V1/V2
fusion protein produced antibodies that reacted with heterologous gp120s and
with V1/V2 domains derived from env sequences of a number of different clades.
The immune sera neutralized a number of primary, macrophage-tropic viruses.
including clade A, B, C, D and E isolates. Specific V1/V2-immunoglobulin fractions isolated from the sera of immunized rats by affinity chromatography on
columns containing the immobilized immunogen possessed potent neutralizing
activities, demonstrating that the neutralizing activities of the sera were mediated
by Vl/V2-specific antibodies.
Conclusion: These results indicate that the V1/V2 domain contains epitopes
that are broadly conserved across clades and that can mediate potent neutralization of primary viruses. While additional testing and development is needed, these
findings suggest that subunit vaccines that efficiently induce such antibodies may
provide protective humoral immunity against a broad range of clinically relevant
HIV-1 strains.
121195 CD86 confers and directs and MHC class I restricted CTL
induction
Michael Agadjanyan, J. Kim, D. Weiner. University of Pennsylvania, 505b Steller
Chance Bldg, 422 Curie Blvd., Philadelphia PA, USA
DNA plasmid based immunization has emerged recently as a promising novel
vaccine strategy and our laboratory has been investigating the potential of this
technique for the development of a vaccine against HIV since 1993. Muscle
tissue has previously been demonstrated to be a major site of antigen expression after intramuscular injection of DNA vaccines. Unlike professional antigen
presenting cells (APCs), however, muscle cells do not express the costimulatory
molecules CD80 (B7-1) or CD86 (B7-2) which provide critical secondary signals
for T-cell activation. Recently we the first time presented data demonstrating that
co-inoculation of mice with DNA plasmids encoding HIV-1 antigens with genes
for CD86 dramatically increased antigen-specific T-cell responses without a significant change in humoral immune response. To resolve whether muscle cells
or professional APCs are responsible for enhancement of anti-viral CTL following
CD86 co-inoculation, we constructed a set of bone marrow chimeric animals using
mice bearing a genetic disruption of /12-microglobulin (#/2 m). The relationship
between CTL induction and MHC class I-restriction after expression of both viral
antigen and CD80 or CD86 costimulatory molecule genes was determined. MHC
class I-restriction of CTL was determined to be under the providence of the B7-2
(CD86) molecule, even when this molecule and antigen were displayed on a
non-professional antigen presenting cell background.
This study defines a role of CD86 in CTL immune induction during DNA immunization. Our results demonstrated that non-hematopoeitic cells, most likely
muscle cells, can be engineered to present the viral antigen.

551*/21196 Multi-subtype envelope-based DNA and protein
immunization strategies for broadening immune
responses to HIV-1
Susan W. Barnett, Y. Sun, H.S. Legg, T.C. VanCott, J.R. Mascola, F.M. Sinangil.
Chiron Corporation, Emeryville, CA, USA
Background: HIV-1 genetic and phenotypic diversity presents a major challenge