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R A Koup

Publications and source records attributed to R A Koup.

88 records · Page 5Linked to original sources
PubMed1994

Studies with monoclonal antibodies to the V3 region of HIV-1 gp120 reveal limitations to the utility of solid-phase peptide binding assays.

Using human monoclonal antibodies (HuMAbs) r(1)-447 (L-736,523) and 19b to the V3 region of HIV-1 gp120, we have explored epitope presentation on V3-peptides and on the corresponding gp120 proteins. HuMAb r(1)-447 binds strongly to the MN and SF-2 peptides and gp120 proteins. In contrast, while this HuMAb binds equally avidly to both the HxB2 and the BRU/BH10 peptides, it binds but weakly to the HxB2 V3 loop on gp120 and fails to bind at all to BH10 gp120. Thus, the solid-phase peptide binding assay can falsely predict reactivity of an MAb with a gp120 protein. Conversely, HuMAb 19b fails to bind to a peptide from the V3 loop of HIV-1 AD-6 in solid-phase assays, but binds to the same peptide in solution and also to AD-6 gp120. Thus, the solid-phase peptide binding assay can fail to predict reactivity of an MAb with a gp120 protein. Furthermore, serum antibodies from individual AD-6 do not react well with the AD-6 V3-peptide in a solid-phase assay, but react strongly with the corresponding MN V3-peptide. On the basis of peptide binding assays, we had assumed that the AD-6 virus was "MN-like" with a prototypic North American/European subtype B GPGR motif at the crown of the V3 loop. However, direct sequencing demonstrates that the AD-6 V3 loop contains a variant GPGK motif. This highlights a limitation of V3-peptide-based assays for serotyping viruses.

Amino Acid Sequence↗
PubMed1993

Genotypic and phenotypic characterization of HIV-1 patients with primary infection.

Better characterization of human immunodeficiency virus-type 1 (HIV-1) in patients with primary infection has important implications for the development of an acquired immunodeficiency syndrome (AIDS) vaccine because vaccine strategies should target viral isolates with the properties of transmitted viruses. In five HIV-1 seroconverters, the viral phenotype was found to be uniformly macrophage-tropic and non-syncytium-inducing. Furthermore, the viruses were genotypically homogeneous within each patient, but a common signature sequence was not discernible among transmitted viruses. In the two cases where the sexual partners were also studied, the sequences of the transmitted viruses matched best with minor variants in the blood of the transmitters. There was also a stronger pressure to conserve sequences in gp120 than in gp41, nef, and p17, suggesting that a selective mechanism is involved in transmission.

Amino Acid Sequence↗
PubMed1993

Human peripheral blood xenografts in the SCID mouse: characterization of immunologic reconstitution.

Immune reconstitutions (hu-PBL-SCID mice) resulting from adoptive transfer of human peripheral blood mononuclear cells into 1800 C.B-17 scid-/scid-mice were characterized. Over 90% of reconstitutions were successful as evidenced by human immunoglobulin production. Variability was noted with donor, cell number, and cell type. Human cells (T lymphocytes, few B cells) could be recovered by 5 days after engraftment. High levels of soluble CD8 and interleukin-2 receptors were detected in sera of hu-PBL-SCID mice. Cells recovered from 17 mice proliferated in response to antigens to which the donor had been primed; responses to nonboosted antigen also increased in some animals. After reconstitution, lymphocytes were found in the spleen and lymph nodes without full restoration of normal architecture. The hu-PBL-SCID mouse shows promise as a model system for a variety of immunologic studies. The inherent variation in the system must be minimized for appropriate use of the model.

Age Factors↗
PubMed1993

hu-PBL-SCID mice can be protected from HIV-1 infection by passive transfer of monoclonal antibody to the principal neutralizing determinant of envelope gp120.

OBJECTIVE: To determine whether passive transfer of a monoclonal antibody specific for the principal neutralizing determinant in the V3 region of HIV-1IIIB gp120 can protect mice with severe combined immunodeficiency (SCID) transplanted with normal human peripheral blood leukocytes (hu-PBL), designated hu-PBL-SCID mice, from subsequent challenge with the homologous viral strain. DESIGN AND METHODS: hu-PBL-SCID mice were given intraperitoneal injections of an anti-HIV-1 neutralizing murine monoclonal antibody (BAT123), its mouse-human chimeric form (CGP 47 439), or a control murine antibody (PNTU), at a dose of 40 mg/kg. The mice were then challenged intraperitoneally with 10 mouse infectious doses of HIV-1IIIB. Three weeks later the mice were killed, and spleen cells and peritoneal lavage collected for determination of infection by coculture for viral isolation and by detection of HIV-1 DNA using polymerase chain reaction (PCR). RESULTS: All three antibodies had similar serum half-lives of 9-12 days. No toxicity was observed in the animals. HIV-1 was recovered by coculture from five out of the six mice given PNTU, and by PCR from two out of the six mice given PNTU, but was not recovered by either technique from any of the 12 mice given BAT123 or CGP 47 439. CONCLUSION: BAT123 and CGP 47 439, which are specific for the principal neutralizing determinant of HIV-1IIIB, protect hu-PBL-SCID mice from infection by this viral strain. Our findings support the use of the hu-PBL-SCID mouse as an in vivo model for studying protection against HIV-1 infection by passive immunization with anti-HIV-1 neutralizing antibodies.

Animals↗
PubMed1993

Nevirapine synergistically inhibits HIV-1 replication in combination with zidovudine, interferon or CD4 immunoadhesin.

OBJECTIVE: To determine whether synergistic inhibition of HIV-1 replication would result from the in vitro use of nevirapine in combination with zidovudine, interferon (IFN)-alpha 2C, and CD4 immunoadhesin. DESIGN AND METHODS: The non-nucleoside reverse transcriptase inhibitor nevirapine (formerly BI-RG-587) was tested in combination with the other antiretrovial agents. Assays were performed on stimulated peripheral blood lymphocytes infected with HIV-1. Virus replication was assessed by the release of viral p24 antigen into culture supernatants. The median-effect principle was used to assess for synergistic interactions of the combined agents. RESULTS: Zidovudine, IFN-alpha 2C and CD4 immunoadhesin, when used in combination with nevirapine, synergistically inhibited HIV-1 replication in human peripheral blood lymphocytes compared with each agent used alone. Some analyses were also consistent with additive effects, but antagonism was not noted. CONCLUSION: These in vitro findings provide a scientific basis for future trials with similar drug combinations.

Antiviral Agents↗
PubMed1993

Pharmacokinetics of nevirapine: initial single-rising-dose study in humans.

Nevirapine, a nonnucleoside inhibitor of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase, was administered for the first time to humans in a pilot study designed to investigate the pharmacokinetics and tolerance of the drug following single-dose administration to 21 HIV-1-infected individuals. The study followed a parallel design. Different groups of three subjects each were given one of seven dose levels (2.5 to 400 mg) in sequential order, starting with the lowest dose. Each subject received only one dose. Nevirapine was rapidly absorbed at all doses from a tablet formulation. Peak concentrations in plasma were generally achieved within 90 min of dose administration. Secondary peaks were also noted between 3 and 12 h or between 24 and 28 h, the latter being noted mainly in subjects receiving the higher doses. After 24 h, concentrations in plasma declined in a log-linear fashion. The terminal half-life and mean residence time exceeded 24 h in all but one subject, indicating a prolonged disposition time in this population. Both peak concentrations in plasma and areas under the plasma concentration-time curves increased proportionally with increasing dose from 2.5 to 200 mg; however, the increase in the peak concentration in plasma and the area under the plasma concentration-time curve appeared to be less than proportional at the 400-mg dose level in this small number of subjects. This observation may be due to increased clearance or decreased absorption at the highest dose or population differences in absorption or clearance between doses. Studies with a cross-over design are planned to resolve these issues. The pharmacokinetic characteristics of nevirapine are appropriate for once-daily administration. A daily 12.5-mg dose is predicted to achieve trough concentrations in plasma in the range required to totally inhibit replication of wild-type HIV-1 in human T-cell culture.

Acquired Immunodeficiency Syndrome↗
PubMed1991

Limiting dilution analysis of cytotoxic T lymphocytes to human immunodeficiency virus gag antigens in infected persons: in vitro quantitation of effector cell populations with p17 and p24 specificities.

The presence of cytotoxic T lymphocytes (CTL) to the gag antigens of human immunodeficiency virus (HIV) has been described in infected populations. We found that the majority of this immune response as measured in bulk CTL assays of unstimulated peripheral blood mononuclear cells (PBMC) is directed against the p24 component of the p55 gag precursor protein. Using limiting dilution analysis of this effector cell population we confirm that the majority of activated gag-specific CTL circulating in the PBMC of infected hemophilic patients are directed at p24 determinants and are present at frequencies of 1/36,000 to 1/86,000 lymphocytes. By performing in vitro stimulation after limiting dilution, the precursor population of gag-specific CTL are characterized and quantitated. HIV gag-specific CTL precursors are identified at frequencies of 1/1700 to 1/17,000 lymphocytes and are made up of cells with both p17 and p24 specificities. No HIV gag-specific CTL precursor cells are identified in the PBMC of HIV-uninfected individuals. These studies demonstrate that CTL directed at both p17 and p24 determinants make up the cellular immune repertoire in HIV-infected individuals but that only the p24-specific CTL are routinely found in an activated state in the circulation.

CD8 Antigens↗
PubMed1991

Deficient human immunodeficiency virus type 1-specific cytotoxic T cell responses in vertically infected children.

Cytotoxic T lymphocyte (CTL) responses to human immunodeficiency virus type 1 (HIV-1) gag proteins were studied prospectively in 17 children (12 infected) born of mothers with HIV-1 seropositivity and in five pediatric patients with hemophilia infected by transfusion of HIV-1-contaminated factor VIII concentrate. B lymphoblastoid cells infected with vaccinia virus vectors expressing HIV-1 gag gene products were combined with autologous peripheral blood mononuclear cells to detect circulating CTLs. Effector cells were defined by monoclonal antibody-mediated, complement-dependent cytolysis. Circulating HIV-1 gag-specific cytotoxic responses were detectable in 4 of 5 HIV-1-infected pediatric hemophilic patients, and were similar in magnitude to those previously described in adults. In contrast, circulating HIV-1 gag-specific cytolysis was detectible in only 3 of 12 vertically infected children. Depletion data revealed that the majority of detectible gag-specific cytolysis was CD8 T cell-mediated. No apparent relationships between CD4 T cell counts, CD8 T cells counts, or serum p24 antigen levels and CTL responses were seen. Deficient CTL development may, in part, explain the more rapid onset of symptomatic disease following vertical HIV infection.

Acquired Immunodeficiency Syndrome↗
PubMed1991

Broadly reactive antibody-dependent cellular cytotoxic response to HIV-1 envelope glycoproteins precedes broad neutralizing response in human infection.

To determine if and when the antibody-dependent cell-mediated cytotoxic (ADCC) response of human serum exhibits broad reactivity across HIV-1 strains, multiple sera were tested for their ability to mediate ADCC against target cells infected with recombinant vaccinia vectors expressing envelope genes of HTLV-IIIB or HTLV-IIIRF. These vectors were found to express the envelope glycoproteins of the two HIV-1 strains and so were appropriate targets for ADCC assays. All the HIV-1-positive sera were able to mediate ADCC against both HTLV-IIIB and HTLV-IIIRF envelope-expressing targets at similar titer. In sera from early seroconverters, the ADCC response was again broadly reactive, even in those sera that exhibited strain-specific neutralizing antibody responses. The ADCC response to natural infection with HIV-1 is therefore broadly reactive and precedes the development of a broad neutralizing antibody response. The broad reactivity of HIV-1-specific ADCC responses may be important for protection against cell-associated virus in vaccine development.

Antibody Specificity↗
PubMed1991

Inhibition of human immunodeficiency virus type 1 (HIV-1) replication by the dipyridodiazepinone BI-RG-587.

The dipyridodiazepinone human immunodeficiency virus type 1 (HIV-1)-specific reverse transcriptase (RT) inhibitor BI-RG-587 was tested for its ability to inhibit HIV-1 replication in both acutely and chronically infected cell lines. The ability of BI-RG-587 to inhibit steps in the virus replicative cycle other than reverse transcription was also assessed. BI-RG-587 was found to be a potent inhibitor of HIV-1 replication in acutely infected cells (50% inhibitory concentration [IC50] = 37.2 nM), and the sensitivity and kinetics of that inhibition was similar to the known RT inhibitor zidovudine (AZT). Even at 100x IC50, BI-RG-587 had no effect on gp120/CD4 interaction, syncytia formation, or envelope glycoprotein processing. In addition, no inhibition of viral replication or protein production was noted in a chronically infected cell line that produces viral products in an RT-independent manner. Finally, no inhibition of acute HIV-2 replication was noted, even with very high (2500x IC50 for HIV-1) concentrations of BI-RG-587. These results demonstrate that BI-RG-587 is a potent inhibitor of HIV-1 replication and that this inhibition occurs at the point of reverse transcription.

Antiviral Agents↗
PubMed1991

Antibody-dependent cell-mediated cytotoxicity directed by a human monoclonal antibody reactive with gp120 of HIV-1.

We used a human monoclonal antibody (MAb; 15e) to identify an antibody-dependent cell-mediated cytotoxicity (ADCC) epitope on HIV-1 gp120. 15e has been shown to recognize a conformation-dependent epitope on gp120 which is important in both CD4 binding and neutralizing of HIV-1 infection. 15e binds to gp120 of HIV-1IIIB but not HIV-1RF. Using a standard ADCC assay, 15e was found to mediate ADCC against cells infected with HIV-1IIIB but not HIV-1RF. 15e did not mediate ADCC against cells with recombinant gp120 bound to surface CD4, indicating that 15e does not mediate innocent bystander ADCC against uninfected CD4 cells. To better define the 15e epitope, we performed ADCC against target cells infected with a vaccinia vector which expresses processed HIV-1IIIB gp160 from which the third variable region was deleted (amino acids, 312-328). MAb 15e efficiently mediated ADCC against cells expressing this altered form of gp120, indicating that this region is not contributing to the conformational epitope defined by 15e. 15e defines an important epitope in the human immune response to HIV-1 infection. Antibodies with 15e-like activity may be useful in immunoprophylaxis or immunotherapy of HIV-1 infection.

Antibodies, Monoclonal↗
PubMed1989

Detection of major histocompatibility complex class I-restricted, HIV-specific cytotoxic T lymphocytes in the blood of infected hemophiliacs.

Major histocompatibility (MHC)-restricted, human immunodeficiency virus type one (HIV-1)-specific, cytotoxic T lymphocytes (CTLs) were detected in the peripheral blood mononuclear cells (PBMCs) of HIV-1-infected individuals. Using a system of autologous B and T lymphoblastoid cell lines infected with recombinant vaccinia vectors (VVs) expressing HIV-1 gene products, we were able to detect HIV-1-specific cytolytic responses in the PBMCs of 88% of HIV-1-seropositive hemophiliac patients in the absence of in vitro stimulation. These cytolytic responses were directed against both HIV-1 envelope and gag gene products. The responses were resistant to natural killer (NK) cell depletion and were inhibited by monoclonal antibodies (MoAbs) to the T cell receptor, CD8 surface antigens, and MHC class I antigens, suggesting a classical MHC class I restricted, virus-specific CTL response.

Antibodies, Monoclonal↗
PubMed1989

Antigenic specificity of antibody-dependent cell-mediated cytotoxicity directed against human immunodeficiency virus in antibody-positive sera.

Antibody-dependent cell-mediated cytotoxicity (ADCC) specific for human immunodeficiency virus (HIV) has been described for HIV-infected individuals. To determine the antigenic specificity of this immune response and to define its relationship to the disease state, an ADCC assay was developed using Epstein-Barr virus-transformed lymphoblastoid cell line targets infected with vaccinia virus vectors expressing HIV proteins. The vaccinia virus vectors induced appropriate HIV proteins (envelope glycoproteins gp160, gp120, and gp41 or gag proteins p55, p40, p24, and p17) in infected lymphoblastoid cell lines as demonstrated by radioimmunoprecipitation and syncytia formation with c8166 cells. Killer cell-mediated, HIV-specific ADCC was found in sera from HIV-seropositive but not HIV-seronegative hemophiliacs. This HIV-specific response was directed against envelope glycoprotein but was completely absent against target cells expressing the HIV gag proteins. The ADCC directed against gp160 was present at serum dilutions up to 1/316,000. There was no correlation between serum ADCC titer and the stage of HIV-related illness as determined by T-helper-cell numbers. These experiments clearly implicated gp160 as the target antigen of HIV-specific ADCC activity following natural infection. Vaccines which stimulate antibodies directed against gp160, which are capable of mediating ADCC against infected cells, could be important for protection against infection by cell-associated virus.

Antibody-Dependent Cell Cytotoxicity↗
PubMedDate not supplied

Reduced antiretroviral drug susceptibility among patients with primary HIV infection.

CONTEXT: The transmission of drug-resistant human immunodeficiency virus (HIV) has been documented, but the prevalence of such transmission is unknown. OBJECTIVE: To assess the spectrum and frequency of antiretroviral susceptibility among subjects with primary HIV infection. DESIGN, SETTING, AND PATIENTS: Retrospective analysis of 141 subjects identified from clinical research centers in 5 major metropolitan areas, enrolled from 1989 to 1998, with HIV seroconversion within the preceding 12 months and no more than 7 days' prior antiretroviral (ARV) therapy. MAIN OUTCOME MEASURES: Phenotypic and genotypic ARV susceptibility of HIV from plasma samples. RESULTS: The transmission of drug-resistant HIV as assessed by a greater than 10-fold reduction in ARV susceptibility to 1 or more drugs was observed in 3 (2%) of 141 subjects, including to a nonnucleoside reverse transcriptase inhibitor in 1 patient and to a nucleoside reverse transcriptase inhibitor and a protease inhibitor in 2 patients. Population-based sequence analysis of these 3 samples identified multidrug-resistance mutations in reverse transcriptase (M184V, T215Y, K219K/R) and protease (L101/V, K20R, M361, M46I, G48V, L63P, A71T, V771, V82T, 184V, L90M) in the 2 latter patient samples, along with numerous polymorphisms. A reduction in susceptibility of greater than 2.5- to 10-fold to 1 or more drugs was observed in viral isolates from 36 patients (26%). Sequence analysis of these 36 samples identified well-characterized drug resistance mutation in reverse transcriptase and protease in only 1 of these patients. CONCLUSIONS: Reductions in drug susceptibility of more than 10-fold were rare among this cohort of recently HIV-infected subjects and were distributed among each of the 3 major classes of ARV drugs tested. Reductions in susceptibility of more than 2.5- to 10-fold to certain ARV drugs of unknown clinical significance were highly prevalent among newly infected patients. Resistance testing may be warranted to monitor the frequency of drug resistance over time and to assess the potential for geographic variability.

Adolescent↗