Abstracts Vol. 1 [International Conference on AIDS (11th: 1996: Vancouver, Canada)]

Track A: Basic Science

Mo.A.1055 - Mo.A.1058

cyte-derived macrophages infected with HIV-I Ada-M.Tissue distribution and pharmacokinetic studies revealed that the entrapment of foscarnet in liposomes greatly improved the
targeting of lymphoid organs (tissues harboring HIV) and largely reduced the systemic clearance of the antiviral agent following intravenous administration. Of particular interest, we
demonstrated that the AUC of liposomal foscarnet in the eyes, lungs and spleen (tissues
harboring CMV) was 4, 40 and 1500 times greater than that of the unencapsulated drug.
Moreover, the plasma half-life of the entrapped drug was more than 4 times higher than the
free agent. Encapsulation of foscarnet in liposomes was shown to protect against abnormal
changes in calcium anrid phosphorus levels induced by the free agent in mice.
Conclusion: Liposome-based therapy represent a convenient approach to concentrate foscarnet at specific sites of infection. Such drug delivery system could improve the efficacy of
foscarnet against viral replication and reduce its toxicityThe use of liposomes could lead to
the development of new strategies which may improve the efficacy and safety of foscarnet
for the treatment of HIV and CMV infections.
Nathalie Dusserre, Centre de Recherche en Infectiologie, Centre Hospitalier de l'Universite
Laval, Ste-Foy Quebec, Canada G IV 4G2.Tel: [phone]; Fax: [phone]
Mo.A. 1055
INHIBITION OF HIV INFECTION BY PSEUDOPEPTIDES BLOCKING VIRAL ENTRY
INTO CD4+ CELLS
Ara G. Hovanessian, Etienne JacototI, Gilles Guichard2, Bernard KrustI, Julia BlancoI, MarieAnne Rey-Cuille, Sylviane Muller2, Jean-Paul Briand2, and Christian CallebautI. I UniteVIC,
UA 1 157 CNRS, Institut Pasteur Paris; 2IBMC, UPR 902 I CNRS, Strasbourg; France
Objective: The development of potent inhibitors of HIV entry into CD4+ T Lymphocytes.
Methods: The RP dipeptide motif is highly conserved in the V3 loop of the extracellular
envelope ylcoprotein of different types of HIV isolates. In view of this, we have designed
and synthesized a construction referred to as "template assembled synthetic peptide"
(TASP), in which a lysine-rich short polypeptide was used as a template to covalently
anchor arrays of tripeptides, such as RPR, RPK or KPR.
Results: The pentavalent presentation, 5(RPR)-, 5(RPK)- or 5(KPR)-TASP molecules manifested maximum inhibitory activity on HIV-infection with a 50% inhibitory concentration (IC
50) value of I -5 uM, respectively. Structure and inhibitory-activity relationship studies using
analogs of 5(KPR)-TASP indicated that the positively charged side chains of the K and R
residues in the tripeptide molecules are critical for the optimal inhibitory activity of the
pentavalent construct. Interestingly, replacement of L-amino acid residues by D-amino acids
or reduction of the peptide bond between the first two amino acids of the tripeptide, generated peptide-TASP analogs active at sub-uM concentrations.The peptide-TASP inhibitors
bind specifically to a still unidentified cell-surface protein referred to as TASP-Binding protein
(TASP BP), which is resistant to trypsin but sensitive to proteinase K digestion.The peptideTASP inhibitors inhibit infection of several types of CD4-expressing cells by HIV- I Lai and
HIV-2 EHO but not by the simian SIV-mac isolate, thus indicating the specific nature of
these inhibitors.
Conclusions: Our results suggest that the peptide-TASP inhibitors block three post-CD4
binding functions of the HIV envelope glycoproteins, to mediate viral entry syncytium formation and triggering cell death by apoptosis. As the peptide-TASP derivatives with unnatural amino acid sequences in the tripeptide moiety retain full inhibitory activity, they should
provide potent protease resistant peptide inhibitors as potential therapeutic agents for
treatment of AIDS patients.
A.G. Hovanessian, Unite VIC, UA I 157 CNRS, Institut Pasteur, Paris 15, France.Tel. [phone]
8776 Fax. 33.1.406 I 301 I2 email: arahovan@dpasteur:fr
Mo.A. 1056
NANOPARTICLES AS DRUG CARRIER FOR ANTIVIRAL AGENTS AGAINST HIV
Bender A*, Immelmann, Andreas*, Kreuter J**, Rubsamen-Waigmann H*, von Briesen H*.
*Georg-Speyer-Haus, Frankfurt/M., Germany; **J. W. Goethe University, Frankfurt/M.,
Germany
Objective: In this study, nanoparticles (NP) as drug carriers loaded with different antiviral
agents were tested in vitro for their activity in HIV-infected human monocytes/macrophages
(MO/MAC).
Methods: Polyhexylcyanoacrylate nanoparticles were prepared by emulsion polymerization
of the monomer in acid medium. For preparation of zalcitabine (ddC)-loaded or saquinavir
(Ro 31-8959)-loaded nanoparticles drugs were present during polymerization. Monocytes
were isolated from blood of healthy donors and after maturation to macrophages on
hydrophobic Teflon membranes were transferred to 24-well plates before in vitro infection
with the monocytotropic HIV- I strain DI 17111. Antiviral activity of drug-loaded nanoparticles
were determined under prophylactic conditions for zalcitabine or in acutely and chronically
infected MO/MAC for saquinavir, respectively
Results: Both zalcitabine and saquinavir nanoparticulate formulations showed a dosedependent reduction of HIV- I.While with nanoparticle-bound zalcitabine no superior effect
was obtained, a significantly higher efficacy compared to an aqueous solution of the drug
was observed when saquinavir-loaded na.noparticles were used. In acuitely infected cells free
saquinavir showed little antiviral activity in concentrations below 10 nM (IC50-4.23 nM)
whereas the nanoparticle formulation exhibited good antiviral effect at I nM and still a significant HIV antigen reduction at 0. I nM (IC50-0.39 nM). At a concentration of 100 nM
saquinavir was completely inactive in chronically HIV-infected MAC, but when bound to
nanoparticles caused a 35% decreased virus production.
Conclusions: Nanoparticles as a drug carrier system could improve the delivery of antiviral
agents which are compromised by disadvantageous physico-chemical properties leading to
an insufficient cellular up-take. Other promising classes of antiviral agents, which would on
their own not get into cells, but in combination with carrier systems could gain access into
target cells and thus become potential new therapeutics for the treatment of AIDS.
(The Georg-Speyer-Haus is supported by the Hessische Ministerium f/r Wissenschaft und
Kunst and the Bundesministeriun fli Gesundheit. Saquinavir was kindly provided by Roche
Products Ltd. UK.)
A. Immelmann, Analysis GmbH, Georg-Speyer-Haus, Paul-Ehrlich-St: 42-44, D-60596
Frankfurt, GerTelephone: [phone] I 2; Fax: [phone] I

Mo.A. 1057
BM 21.1290: IN VITRO EVALUATION OF A POTENTIAL NEW ANTI-AIDS
COMPOUND
Herrmann, Dieter B.I*, Kucera L.S**,. Zilch H*, Mertens A*, Opitz H.G.*. *Boehringer
Mannheim GmbH, Department of Molecular Pharmacology, Mannheim, Germany;
**Bowman Gray School of MedicineWake Forest University,Winston-Salem, NC
Objective: Determination of the anti-HIV activity and cytotoxicity of the new anti-AIDS
compound BM 21. 1290 (INN: Fozivudine tidoxil) in different in vitro cell systems.
Methods: 50% anti-HIV (IC50) and 50 % toxic (TC50) concentrations were determined in
several human T cell lines (CEM-SS, H9, H911IB) and in human peripheral blood leukocytes
(PBLs), acutely or persistently infected with HIV- I, for BM 21.1290 in comparison to the
standard compounds 3'-azido-3'-deoxythymidine (AZT) and 2,3'-dideoxyinosine (DDI).The
procedures to measure anti-HIV- I activity included syncytial plaque and reverse transcrip
tase (RT) assays as well as p24 antigen ELISA. Cytotoxicity was measured by using the
[3H]-thymidine assay and/or by the trypan blue exclusion methode. Differential selectivity
indices are expressed as TC50/IC50 ratios.
Results: Data indicate that BM 2 I. 1290, AZT and DDI inhibit virus infection and propagation concentration dependently, as measured by RT activity, p24 core antigen concentration
and infectious virus production.The IC50 values for BM 21.1290 ranged between 0.02-0.2
pM (15-150 ng/ml), dependent on the cell type studied, the HIV isolate used and the para
meters for anti-HIV activity measured. Anti-HIV activity of BM 2 I. 1290 was most pronounced in infected human T cells and PBLs. On the other side, BM 2 I. 1290 was generally
much less toxic to HIV-infected cells compared to AZT or DDI. Differential selectivity
indices for BM 2 I. 1290. AZT and DDI in CEM-SS cells were, for example, 1925, 4 II and
59, respectivelyThis finding is in accordance with the lack of bone marrow toxicity of BM
21.1290 in vitro and in vivo.
Conclusions: This is the first report on the anti-HIV activity of the new anti-AIDS compound BM 21.1290.The results indicate that BM 2 I. 1290 has a higher differential selectivity
index in vitro in comparison to standard therapeutics. After having successfully passed pharmacological/toxicological and clinical phase I development. BM 21.1290 has entered phase II
efficacy studies in ARC/AIDS patients in January 1996.
PD Dr Dieter Herrmann, Boehringer Mannheim GmbH, Dept. Molecular
Pharmacology/New Indications, Sandhofer Strasse 11 6, 68305 Mannheim, Germany
Mo.A. 1058
BLOCKING OF HIV-I ENVELOPE GLYCOPROTEIN (GPI20)-BINDINGTO CD4+
LYMPHOCYTES BY ANTI-HIV MONOCLONAL ANTIBODIES
Kroepelin, Marianne, Suesal C, Daniel V Opelz G. University of Heidelberg, 69 I 20
Heidelberg, Germany
Objective: Our research aim was to characterize HIV- I envelope glycoprotein (gp I 20)binding to CD4+ lymphocytes and to block this interaction with monoclonal antibodies as
a potential strategy for immunotherapy.
Methods: CD4+ lymphocytes of healthy volunteers were isolated by Ficoll-Hypaque gradient centrifugation followed by cell sorting and incubation with recombinant gp I 20 or FITClabeled rgp I 20 in the presence of different inhibitors. Reduction of median fluorescence
was quantitated after antigp l 20-FITC binding to rgp l 20 coated CD4+ cells or directly
measured (rgp I 20/-FITC: HIV- I IIIB-isolate generated in the baculovirus vector system) using
flow cytometry (FACScan, BD).
Results: Two epitopes of gp I 20 are major contact sites for CD4. Our data support an,induced fit" binding-mechanism, assuming also two binding sites on CD4 for the HIV- I
envelope. One of those sites is hidden and bound by gpl20 in a competitive way. In addi
tion to the known anti-viral and neutralizing activity of human mAb F I05 (Posner M. et al.)
which is directed against a discontinuous gp 120-binding region of C4, the mouse mnAb 87 -I 35/9 described by Niedrig M, et al. binds to a C I epitope at the NH2-temninal gp I 20 -region (109-123) and blocks rgp120/CD4-binding in a synergistic mannei:The blocking
capacity of the mouse mAb is concentration dependent and approx. 50% of that of F 105.
Conclusions: Our results showed that, in addition to the known activity of human mAb
F 105, the mouse mAb 87-135/9 has a similar gp I 20/CD4-blocking capacityThese findings
could improve concepts of passive immunotherapy using a combination of mAbs or a bispecific, human antibody
Dr M. Kroepelin, University of Heidelberg, Institute of Immunology Im Neuenheimer Feld
305, 69120 Heidelberg, GermanyTel.:([phone]; Fax: [phone] email:
marianne. [email protected]

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