Drug – bio-affecting and body treating compositions – Antigen – epitope – or other immunospecific immunoeffector – Amino acid sequence disclosed in whole or in part; or...
Reexamination Certificate
1999-02-25
2001-06-19
Nelson, Brett L. (Department: 1648)
Drug, bio-affecting and body treating compositions
Antigen, epitope, or other immunospecific immunoeffector
Amino acid sequence disclosed in whole or in part; or...
C424S184100, C424S208100, C530S395000, C530S350000, C530S324000, C530S325000, C530S326000, C530S327000, C530S328000, C530S329000, C435S005000, C435S007100, C435S007200
Reexamination Certificate
active
06248328
ABSTRACT:
The invention relates to new HIV-1 virus isolates of a subtype, vaccines against HIV-1 virus infections of this subtype, methods of producing same, and the use of the virus isolates for the production of vaccines and for differential diagnostics.
Hitherto, efforts to develop agents for the prophylaxis and stherapy of HIV infections have focused on the inhibition of the reverse transcriptase or some other enzyme of the HIV-1 virus such as, for example, protease. Thereby a specific inhibition of viral replication in contrast to cellular replication is intended to be accomplished. Nevertheless, the antiviral agents of prior art suffer from the drawback that they are accompanied by a relatively high toxicity towards the cells, i.e. they do not only affect the virus.
Those groups of substances which have been identified as being active against HIV viruses include, inter alia, various nucleoside analogues (azidothymidine, di-deoxyinosine and di-deoxycytidine). However, a massive formation of resistence of the virus has been detected after some relatively short periods of therapy with these substances {Zimmermann et al., Abstract No. 3656, IV International Conference on AIDS, Stockholm, 1988, Rübsamen-Waigmann et al., Infection 19, Suppl. 2, 77-82, 1991}. Furthermore, all of the substances mentioned produce considerable side-effects, at least when applied in higher doses.
Another inhibitor of reverse transcriptase is the substance Suramin. However, this substance, due to its toxicity towards the mammal organism, is also not suitable for prophylaxis or therapy of HIV-virus infections {H. Mitsuya et al. “Suramin Protection of T-Cells in vitro against Infectivity and Cyto-pathic Effect of HTLV-III, Science 226 (1984), pages 172-174}.
Further development has led to reverse transcriptase inhibitors which are less toxic towards the mammal organism. These include substances such as dextran sulfate and pentosan polysulfate, which have proven to display a HIV-1-inhibiting effect in vivo, as has been described in DE 36 01 136 and in EP 0 293 826.
In addition to a chemotherapeutic treatment of HIV infections there is basically the possibility of a gene therapy or immunotherapy. Gene therapy comprises the incorporation of parts of viral nucleic acids in human cells, especially in the target cells of the HIV virus, for example the CD4-positive cells of the immune system. Then, the viral messenger RNA (m-RNA) can be neutralized by the formation of an “antisense” RNA, i.e. an RNA complementary to m-RNA, and hence the virus proliferation can be terminated. In some other form, oligonucleotides or structures chemical related thereto that are complementary to m-RNA can be employed. In immunotherapy, antigens are administered after the infection which are expected to support the immune response to HIV.
Meanwhile, the World Health Organization (WHO) estimates a number of at least 13 million of HIV-infected people worldwide; no country has been left free from infections with HIV-1 or HIV-2 or both viruses at the same time. It is urgently necessary to develop a vaccine for protection from HIV infections in order to prevent the epidemic from farther spreading. Here, and also in immunotherapy, the main problem is the high variability of the HIV viruses: a prophylactic inoculation should and must include all of the possible virus variants. Epidemiologic investigations in combination with the genetic characterization of the viruses produced the result that several subtypes do already exist in both virus families {Myers et al., Human Retroviruses and AIDS, Los Alamos, 1991; Dietrich et al., Nature 342, 948-950, 1989}, which subtypes are significantly distinguished from one another with respect to their hereditary information to a degree such that one single vaccine will not be capable to be active against all variants. Moreover, these subtypes are also subject to different geographic distributions.
The present invention provides three new HIV 1 virus isolates HIV-1
D757
, HIV-1
D747
, HIV-1
D760
, which, surprisingly, form a further independent subtype of the HIV-1 family and have been recovered from three Indian patients who, at the time of virus isolation, did not show any typical AIDS symptoms. Furthermore, the invention also includes virus isolates of the HIV-1 type which are up to 30%, and preferably up to 5%, divergent from HIV-1
D757
, HIV-1
D747
, and HIV-1
D760
.
The virus isolates, in accordance with the regulations according to the Budapest Treaty, have been deposited with the European Collection of Animal Cell Cultures (ECACC), Porton Down, Salisbury, Wiltshire, United Kingdom, on Aug. 27, 1992, under the following Accession Numbers:
HIV-1
D747
—ECACC V92082718
HIV-1
D757
—ECACC V92082719
HIV-1
D760
—ECACC V92082720
The virus isolates proliferate on fresh mononuclear cells of peripheral blood, i.e. lymphocytes and macrophages.
The invention further provides a vaccine having an activity spectrum against this specific HIV-1 subtype.
The vaccine contains, as the active ingredient, the peptides
1. MPNGTKSNS SEQ ID NO:7, MPNGTKGNS SEQ ID NO:8, MPNGTKSNL SEQ ID NO:9,
2. RNEKDLLALDSWKN SEQ ID NO:10,
combinations comprising said peptides or longer or shorter subpeptides thereof being up to 7 amino acids in length.
Said peptides are derived from the env region of the strains according to the invention (cf. FIG.
1
). The nucleotide sequences of these viruses in the env gene over a range of 1.8 kb are only to 79.4-81.6% homologous with viruses of the North American/European subtype and to 78.9-81.2% homologous with prototype viruses of the Central African subtype. The homology on the amino acid level is 72.1-75.9% and 71.2-74.2%, respectively (cf. Table). Thus, these viruses genetically are to be classified between these two subtypes and form some further independent subtype which is also distinguished from the subtypes found in Rwanda/Uganda and in Northern Thailand by genetic equidistance.
Upon comparison of the amino acid sequences of the env proteins of HIV-1
D757
, HIV-1
D747
, HIV-1
D760
with the consensus sequences as derived for all of the HIV env sequences already published, there result for the env region two short ranges, in which the three Indian sequences are different from all of the other HIV-1 sequences in 7 of 9 and in 7 of 14 amino acids, respectively (FIG.
1
). Peptides from these regions are preferably suitable for a vaccine having a specific spectrum of activity against the Indian subtype, and particularly so, if the vaccine consists of a combination of these peptides.
The peptides according to the invention are synthetically produced and, if advantageous, are chemically modified. These peptides or longer as well as shorter subpeptides, down to 7 amino acids in length, thereof are also a constituent of this invention, and so are their modified forms.
Furthermore, the invention also relates to a vaccine which contains, as the active ingredient, peptides from variable regions of other genes of HIV-1
D757
, HIV-1
D747
, HIV-1
D760
, if the deviation from the corresponding sequence of HIV-1 viruses of other subtypes is more than 30%.
In the vaccine, there may further be used antigens from the viruses according to the invention or peptide combinations or combinations of peptides and antigens. More specifically, the use of such antigens or peptides is preferred which, within the loop of the coat glycoprotein (amino acids 423-450, relative to HIV-1
Lai
), is responsible for binding to the CD4-receptor, contain a glycosylation site (NXT/S) immediately in front of cysteine 450 (HIV-1
Lai
). This glycosylation site is typical for the Indian viruses HIV-1
D757
, HIV-1
D747
and HIV-1
D760
. The amino acid regions of the HIV-1
D757
, HIV-1
D747
and HIV-1
D760
that correspond to the amino acids 423-450 of HIV-1
Lai
are amino acids 294-321 of SEQ ID NO:2, amino acids 296-323 of SEQ ID NO:4, or amino acids 304-331 of SEQ ID NO:6 respectively. This glycosylation site has not been encountered among 29 HIV-1 env-genes that have been sequenced worldwide.
One preferred emb
Dietrich Ursula
Grez Manuel
Rubsamen-Waigmann Helga
Von Briesen Hagen
Chemotherapeutisches Forschungsinstitut
Jacobson & Holman PLLC
Nelson Brett L.
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