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

Mo.A.I 105 - Mo.A.I 110

Monday, July 8, 1996

Mo.A.I 105
QUANTITATIVE NASBA AND COMPETITIVE PCR AND RT-PCR CAN RAPIDLY
MONITOR "IN VITRO" THE EFFECT OF ANTIRETROVIRAL DRUGS ON HIV
REPLICATION.
Debuia Ci, arrlev r M., Brerra R., Zuara F., "Bruno R., "Achll G., Gaco-.,e E., Romero
E, "Fil e, (. istiituto di MiCrobiologia, Universita Studi di Pavia, 'Servizio Ar alls
Microbiologiche IRCCS S.Matteo, Divisione Malattie Infettive e Tropicali, Universit, Studi
sir Pavia.
Objective: We have applied a quantitative NASBA and competitive PCR to examine the
kinetics of HIV replcation in vitro" and to assess the effect of antiviral drugs on the accumulation of HIV gag RNA and DNA.
Methods: H9 cells ( ix 106/mli) were infected with HTLV IIB (100 TCID50). Cell cultures
were treated with AZ1, ddl or AZT + ddi (2pg/ml and I pg/ml) at different timres before
and after rinfection At 3, 6. I0 and 13 days after infection supernatant and cell samples were
collected.( Genoric RNA in supernatant and proavirl DNA and trascrnptional RNA in cells
were subjected to quanrtitative analysi, by competitive PCR, RT PCR and NASBA.
Results: An incualum size of 100 TCID50/106 cells provide measurable DNA and RNA signals in a 7 day per iod that could be used effectively to compare the effect of drugs and
drug a;sociations.Vral lord measurement demonstrates that HIV replication was significantly inhibited by AZT and AZ FT+ddl as long as DNA and RNA transcripts accumulation when
drugs were added up to 24 hours after infection.Virus replication was significantly less
inhibited by ddl 'when drug was added to cultures from simultaneously to 24 hours after
infection asr measured by accumulation of RNA and DNA.
Conclusion: The use of gag RNA and DNA markers to measure the replication of HIV at a
relatively low multiplicity of infection (I 00TCID50/106 cells) within a short time period (<7
days) demonstrated the rephicative capacity of the virus anrid the overall effect of antiretroviral drugs. Moreover qurntitation values of genornic RNA obtained by competitive RT.PCR
and NASBA have a good correlation.
Maurizia Debiggi - Istituto di Microbiologia, via Brambilla, 74 27100 PAVIA - ITALY
University of PAVIA
Mo.A. I 106
COMPARATIVE STUDY WITH DDC PLUS AZT IN HIV INFECTION: EARLY VS LATER:
PRELIMINAR REPORT
Dominguez Ca rlos CanoVillarreal C,Torres R, Robles M.,Terrazas J. BlannoV: Hospital de
Infectologa, Centro Medico National La Raza IMSS, Mexico DF.
Objective: to comrpar efficacy and tolerance with a double drug regimen (AZT plus DDC)
in HI V patients infection: early vs later Gruop A included DDC 0.750 mg tid plus AZT 100
my tid, lymphocites CD4 counts >300/mm. Group B DDC plus AZT at same doses but
lymphocites CDT counts <300/nmm3.
Methods: Patient- of both sex, older than 18 years old, HIV positive by ELISA and WesternBlot, asymptonatics or with history of clirical disorders of AIDS, and non pregnant women
were included. The following tests hemartic biornetry, CD4, CD8 lymphocytes, were done
brisal, every two weeks the first month, aftereward every two months. Oportunistic infections episodes and mortality were enregistered to month 20.
Results: Group A (GA) 4 7 cases 38 males and 9 women; group B (GB) 100 cases 88 males
and 12 wormen; GA: average age 32 ~ 7 years old (range 21-50), GB: 31 + 12 years old
(range 20- 5 1): A HIeroglobin level of I.8 +t 1.5 gr/dL (range I 1I 7), GB: hemoglobin
level of 14.2 + 1.56 gr/dl ('rnge I -18); GA: neutrophils GA: 2791 + 892 mr3 (range
1400 -5500), GB: 2582 + 920 mis3 (range 1200-5600), GA: CD4 lymphocites 401 ~ 87
(ringe 303 700) B (iB: /4.8 + 79 (range 4-398. After a follow up of 20 months, GA: one
death vs 22 )in GB GA: 17 oporturnistic infection episodes vs 181 I in GB. GA hemoglobin
14.7 ~ 1. / (range I I- 9) vs GB 13.6 1 1.8 (range 10- 17), GB neutrophils 3136 + 1255
(range 1200-6100) vs 2268 ~ 1176 (range 100-5400) GB. AZT Toxicity GA one patient vs
42 GB, DDC toxicity GA 0 patient vs 12 GB.
Conclusions: we consider is better to begin the double regimen early, because the efficacy
and tolerance are superior:
Carlos Canoe Donainguez. Avenida 5 de Mayo, 152 I Col., Lormas de Tarango Ap. 19-132
Mexico D.F. Tel: [phone]
Mo.A. I 107
FACTOR(S) ACTIVE AGAINST HUMAN IMMUNODEFICIENCY VIRUS,
CYTOMEGALOVIRUS AND HERPES SIMPLEX VIRUS
Nosik Dmitr N V.',Vainbeg Yu.'' Nossik N. Kaplina.", Kalnina L.'D.I.Iva iovsky
Institute or Viroloy Ru Iirn Academy of Medical Sciences, "Joint Stock Company "Medicia',
Moscow, Russ.
Objective: To study an antiviral activity of agents against Human Irnmunodeficiency Virus,
Cytomegalovirus (CMV) nd Herpes Suimplex Virus (HSV)
Methods: The antivira l activity of soluble low weight factor(s) (m.rn. around 5 kD) (named
AVF) produced n T- cels was studied in the model system of human PBMCs and T-cell lines
rinfected with HIV I isolates, diploid fibrobiast cells infected with CMV and on tie animal
model - herpetic ereplitis in mice.
Results: The prelirminary experiments showed that frctor(s) efficiently blocked HIIV - I producton in naturally infected PBMCs and 3Tcell lines infected with HIV. The EC50 value for
fhe factor(s) wis lb 1:20 drlution in p24 antienr and syncitiun-reduction asays.The
degree of su,,ceptibility to AVE varied among 7 Russian HIV - I isolates but the reduction of
viral yield was observed in all cases. The AVF was also active against CMV and reduced in
50 100 folds viius infectiity the survival rate of letally HSV-infected animals increased on
53% when the factor(s) was administer ed. It was non toxic and stimulated cell proliferation.
Conclusions: It may be concluded that nducible soluble factor(s) had antiviral activity
against Human Immunodefiiercy Virus Cytomegalovirus and Herpes Simplex Virus and
could be considered a. potential antiviral agent for the cormbned therapy of AIDS and
ccompanied oppor tunistis viral nfections.
D.N.Nosk, 16, Garmaleyn St., Moscow, 123098, Russia Telephone: [phone]
lax: (095) i90-74-85

Mo.A.I 109
CYSTAMINE INHIBITS HIV-I REPLICATION IN CORD BLOOD-DERIVED
MONONUCLEAR PHAGOCYTES AND LYMPHOCYTES.
Ho, We-Zhe. K.r mr. D r d, Song, Li, Douglas, Steven D. The Children's Hospital of
Philadelphia, Jr e sity of Pnsylvania School of Medicine, PA. U.S.A.
Objective: To determine the effects of cystamine on HIV expression in cord blood monocytes derived macrophages (CBMDM) and lymphocytes in vitro.
Methods: Cord blood were obtained from the umbilical vein of healthy term newborn
infants after uncomplihcated pregnancies and deliveries. Cord monocytes and lymphocytes
were isolated according to our previously described technique. Cord monocytes were cul
tured in vitro for 10 days before HIV infection. Both laboratory-adopted and primary isolates of HIV were applied to the infection study.
Results: Cystamine potently suppressed in a concentration dependent fashion HIV- I
expression in CBMDM and lymphocytes as determined by HIV-I RT activityTThis inhibitory
effects of cystarnine were applicable to all 5 HIV- I strains (both laboratory adopted and
fresh isolate) tested in the studyThe addition of cystamine into HIV I chronically infected
CBMDM (I 2 days after infection was established) also suppressed 80-90% of RT activity in
comparison to the untreated controls. Cystamine also decreased HIV- I protein expression
in CBMDM as determined by indirect immunofluorescence assayThe inhibitory effects of
cystanrmine on HIV- I did not appear to be caused by toxicity to CBMDM or lymphocytes
because there was no change in cell viability or cellular DNA synthesis as evaluated by trypan blue dye exclusion and [3H]-thymidine incorporation, at doses of cystamine that inhibit
the virus. HIV- I infected CBMDM or lymphocyte cultures (without cystamine treatment)
demonstrated giant syncytium formation or cytopathic effect (CPE), respectively, whereas
cystamine treated cultures lacked the giant syncytia or CPE induced by HIV- I infection.
Conclusion: We demonstrated that cystamine inhibited HIV- I replication in CBMDM and
lymphocytes, suggesting that cystamine has the potential to limit HIV- I replication in vivo
and may represent potentially useful compounds in the treatment of pediatric HIV I infection and AIDS.
Wen-Zhe Ho, Immunology Dept. The Children's Hospital of Philadelphia 34th St. and Civic
Cental Blvd.Tel: [phone]; Fax: [phone]: email: Ho@D email. CHOPEdu
Mo.A.I 110
DETECTION OF MUTANT VIRUSES AT CODON 215 IN THE HIV REVERSE
TRANSCRIPTASE GENE IN UNTREATED PATIENTS.
Picard H*, Binet D*, Burghoffer B*, Kirstetter M*, Frottier J:, Petit J C*, Morand-Joubert
L*. * Service des Maladies infectieuses: * Service de Bactnerologie-Virologie, H6pital
Saint-Antoine, Paris, France
Objective: to determine the prevalence of mutant viruses at codon 21 5 in the reverse
transcriptase gene, in HIV- I positive patients who have never received antiretroviral drugs.
Methods: forty-three HIVinfected patients were included in this study between April 1994
and January 1996, in Saint-Antoine hospital.The patients hac never received antiretroviral
drugs. specially zdovudine.
Genotypic resistance to zdovudine was detected by differential PCR to discern mutations
at codon 215 of the HIV I reverse transcriptase (RT) gene, directly in the perpheral blood
mononuclear cells (PBM(Cs) of infected persons (as previously described by Larder BA).
Results: Seven (I6%) had mutant sequence or mixed viral population of wild-type and
mnutant sequence, corresponding to genotypic resistance. Only one of these 7 patients had
a mutant sequence.Thirly-six (84%) had a wild-type RT gene sequence at codon 215.The
presence of mutant sequence was found more frequently in the latest samples than in the
samples obtained at the beginning of the study:
Conclusions: We found that mutant sequence at codon 215 can be detected in the PBMCs
of infected persons who have never received zdovudine.This finding requires a molecular
epidemiological study of RT gene sequence of untreated patients to determine the evolution of their viral population.
Morand--joubert L. Service de Bacternologie-Virologe; H6pital Saint-Antoine;
184 rue du Faubourg Saint-Antoine 75012 Paris, France.Telephone: [phone]
Fax: [phone]

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