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Methods and compositions for inhibiting HIV infection

A compound, pseudovirus technology, applied in the field of new compounds that inhibit HIV infection, can solve the problems of toxicity or adverse side effects, incompatibility of antiviral activity, unsatisfactory drugs, etc.

Inactive Publication Date: 2008-12-03
IRM
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Drugs currently used to treat HIV infection and AIDS are unsatisfactory
Common drugs used to treat HIV infection, such as AZT or HIV protease inhibitors, have toxicity or adverse side effects that are incompatible with their antiviral activity when used at pharmaceutically effective concentrations

Method used

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  • Methods and compositions for inhibiting HIV infection
  • Methods and compositions for inhibiting HIV infection
  • Methods and compositions for inhibiting HIV infection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Example 1. General Materials and Methods

[0087]Cell Lines and Maintenance: HeLaCD4βgal cells were obtained from Dr. Michael Emerman through the AIDS Research and Reference Reagents Program of the NIH, NIAID AIDS Division (Kimpton, J. Virol. 66:2232-9, 1992). Cells were cultured in DMEM medium supplemented with 10% fetal bovine serum, 1× penicillin / streptomycin / L-glutamine, 0.2 mg / mL G418 and 0.1 mg / mL hygromycin B. Jurkat cells were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum and 1× penicillin / streptomycin L-glutamine. All cell culture reagents were purchased from Invitrogen.

[0088] Screening of cDNA in HeLaCD4βgal cells: High-throughput cDNA reverse transfection of HeLaCD4βgal cells was performed essentially according to the method described in Chanda et al., Proc. Briefly, individual cDNAs from a subgenome library containing 15,000 genes (see http: / / function.gnf.org for details), the negative control Sport6GFP cDNA, and the positive con...

Embodiment 2

[0094] Example 2. Identification of novel HIV-interacting host factors from siRNA screens

[0095] We performed a subgenomic siRNA screen for host proteins involved in HIV infection by monitoring the expression of a reporter gene under the control of the HIV LTR promoter in HeLaCD4Bgal cells (Kimpton et al., J Virol 66:2232-2239, 1992). HeLa-CD4-Bgal cells were obtained from Dr. Michael Emerman through the AIDS Research and Reference Reagents Program of the NIH, NIAID AIDS Division. Cells were transfected with siRNA against Tat as a positive control using a reverse transfection protocol and challenged with HIV-IIIb 24 hours after transfection (Huang et al., Proc. Natl. Acad. Sci. U.S.A. 101:3456-61 , 2004). In order to be able to observe effects on all stages of infection (from entry, to release, to spread in cell culture), the infection was continued for three days. By monitoring reporter gene expression in HeLaCD4βgal cells, this system can detect any modulation of HIV inf...

Embodiment 3

[0097] Example 3 Identification and Characterization of HIV Interacting Host Factors from cDNA Screening

[0098] We performed a high-throughput screen of a cDNA library containing 15,000 unique genes to find novel proviral factors whose overexpression would lead to enhanced HIV-IIIb infection. Since HeLaCD4βgal cells are easy to transfect, we used them for selection and challenge with replication competent HIV-IIIb. Negative control (Sport6-gfp) and positive control (Tat-Sport6) cDNAs were added to the wells of a 384-well plate containing library cDNA (one gene per well), followed by LTR-luciferase reporter gene containing HIVTat response Construct transfection reagent solution. Cells were added after complex formation, and 24 hours later, each well was infected with HIV-IIIb. In order to be able to observe the effect on all stages of the infection (from entry, to release, to spread in the cell culture), the infection was continued for three days. Infection was then assess...

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Abstract

This invention provides novel HIV-interacting host factors. The invention also provides methods of using the HIV-interacting host factors to screen for compounds that inhibit HIV infection. The methods comprise first screening test compounds for modulators of an HIV-interacting host factor disclosed herein, and then further screening the identified modulating compounds for ability to inhibit HIV infection. The invention further provides methods and pharmaceutical compositions for treating diseases and conditions associated with HIV infection.

Description

[0001] Cross References to Related Applications [0002] This application claims priority under U.S.C. 35 Section 119(e) to U.S. Provisional Patent Application No. 60 / 748,759, filed December 8, 2005. The disclosure of the priority application is hereby incorporated by reference in its entirety for all purposes. field of invention [0003] The present invention generally relates to the inhibition of HIV infection. More specifically, the present invention relates to the identification of novel HIV-interacting host factors and to methods of using these host factors to identify new compounds that inhibit HIV infection. Background of the invention [0004] Human immunodeficiency virus (HIV) is a lentivirus belonging to the Retroviridae family. It is estimated that as many as 90% of AIDS cases in the world are transmitted through sexual contact and pregnancy. This transmission is initiated by crossing the mucous membrane barrier of the sexual organs or the placenta upon exposu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/50C12Q1/70
CPCG01N2333/161C12Q1/6897G01N2500/00G01N33/56988C12Q1/18A61P31/18A61P43/00G01N33/50
Inventor D·源K·L·库亨J·考德威尔
Owner IRM
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