Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Ester doai
Reexamination Certificate
1997-06-25
2001-09-18
Marx, Irene (Department: 1651)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Ester doai
C514S546000, C514S721000, C514S734000
Reexamination Certificate
active
06291524
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to the isolation, purification and characterization of derivatives of 1,4-bis-(3,4-dihydroxyphenyl)-2,3-dimethylbutane (nordihydroquaiaretic acid, NDGA) The derivatives were isolated from leaf and flower extracts of the creosote bush (Larrea tridentata, Zygophyllaceae) and together with NDGA can be used to suppress Tat trans- activation in lentiviruses, including the HIV virus. Other broader aspects of the invention, including use of the disclosed compounds for the treatment of Herpes simplex virus, will also be hereinafter apparent.
2. Description of the Related Art
Tat is a transactivator of human immunodeficiency virus (HIV) gene expression and is one of the two or more necessary viral regulatory factors (Tat and Rev) for HIV gene expression. Tat acts by binding to the TAR RNA element and activating transcription from the long terminal repeat (LTR) promoter.
The Tat protein stabilizes elongation of transcription and has also been shown to be involved in transcription initiation. Previous studies have shown that Tat mediates reduction of antibody-dependent T cell proliferation, contributing substantially to the failure of the immune response. Tat also directly stimulates Kaposi's cell growth.
Since Tat has no apparent cellular homologs, this strong positive regulator has become an attractive target for the development of anti-AIDS drugs (see FIG.
1
). In contrast to currently available HIV reverse transcriptase inhibitors (AZT, DDI) or potential protease inhibitors that prevent new rounds of infection, an inhibitor which suppresses viral gene Tat regulated expression of integrated proviral DNA will arrest the virus at an early stage (Hsu et al.,
Science
254:1799-1802, 1991).
Efforts aimed at the elucidation of factors which control gene expression at transcriptional and post-transcriptional levels in host eukaryotes have recently made possible quantitative assessment of Tat function (Sim,
Ann. N.Y. Acad. Sci.
616: 64-70, 1990, the entire contents of which are hereby incorporated by reference and relied upon). To screen for inhibitors for Tat regulated transactivation (Tat-TRS), the secreted placental alkaline phosphatase (SEAP) reporter gene is put under the control of HIV-1 LTR promoter in the plasmid pBC12/HIV/SEAP. The Tat-coded activity is supplied by a second plasmid construct pBC12/CMV/t2. Transient cotransfection of COS cells with these two plasmids leads to secretion of alkaline phosphatase into the culture medium which is analyzed by a simple colorimetric assay (Berger et al.,
Gene
66: 1-10, 1988, the entire contents of which are hereby incorporated by reference and relied upon) The SEAP assay, therefore, provides an indirect determination of Tat transactivation. An inhibitor should cause reduction of the SEAP activity which is due to inhibition of the expression of the SEAP mRNA via transactivation of the HIV-1 LTR promoter by Tat protein (Tat-TRS).
SUMMARY OF THE INVENTION
In the present application, we disclose Tat-TRS inhibitory activity of the desert plant
Larrea tridentata.
Among several plant extracts prepared from rain forest and desert medicinal plants used in traditional medicinal against viral affections, only the total extract from the leaves and flowers of the creosote bush (
Larrea tridentata
) showed Tat-TRS inhibitory activity. This extract also inhibits HIV cytopathic effects on human lymphoblastoid cells chronically infected with the virus as assessed by the newly developed soluble-formazan assay (Weislow et al.,
JNCI
81: 577-586, 1989, the entire contents of which are hereby incorporated by reference and relied upon).
The present invention discloses compounds of the structural formula:
wherein R
1
, R
2
, R
3
and R
4
are each selected from the group consisting of HO—, CH
3
O— and CH
3
(C═O)O—, provided that R
1
, R
2
, R
3
and R
4
are not each HO- simultaneously.
Each compound was isolated from leaf-flower extracts of the creosote bush
Larrea tridentata
and is a derivative of 1,4-bis-(3,4-dihydroxyphenyl) -2,3-dimethylbutane (nordihydroquaiaretic acid, NDGA).
In addition, NDGA and derivatives can be used to suppress Tat transactivation of a lentivirus, including the HIV virus, in a cell by administering NDGA or a derivative thereof to the cell.
It is also contemplated according to the invention that the compounds disclosed herein can be used to inhibit Herpes simplex virus and other undesired virus growth in a host as later described.
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Giza et al., A self-inducing runaway-replication plasmid expression system utilizing the Rop protein, Elsevier Science Publishers B.V. (Biomedical Division), 1989, pp. 73-84.
Staal et al., Antioxidants Inhibit Stimulation of HIV Transcription, Aids Research and Human Retroviruses, 1993, vol. 9, No. 4, pp.299-306.
Gnabre et al., Isolation of anti-HIV-1 lignans fromLarrea tridentataounter-current chromatography, Journal of Chromatography A., 719, 1996, pp. 353-364.
Gnabre et al., Characterization of Anti-HIV Lignans fromLarrea tridentata, Tetrahedrom,1995, vol.. 51, No. 45, pp. 12203-12210.
Weislow et al., New Soluble-Formazan Assay for HIV-1 Cytopathic Effects: Application to High-Flux Screening of Synthetic and Natural Products for AIDS-Antiviral Activity, Journal of the National Cancer Institute, 1989, vol. 81, No. 8, pp. 577-586.
Gnabre et al., Inhibition of human immunodeficiency virus type 1 transcription and replication by DNA sequence-selective plant lignans, Proc. Natl. Acad. Sci. USA, 1995, vol. 92, pp. 11239-11243.
Gisvold et al., Lignans fromLarrea divaricata,Journal of Pharmaceutical Sciences, 1974, vol. 63, No. 12, pp. 1905-1907.
Perry et al., Synthesis of Lignans, I. Nordihydroguaiaretic Acid1, J. Org. Chem., 1972, vol. 37, No. 26, pp. 4371-4376.
Gnabre John N.
Huang Ru Chih C.
Hobbs Ann S.
Johns Hopkins University
Marx Irene
Venable
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