Drug – bio-affecting and body treating compositions – Immunoglobulin – antiserum – antibody – or antibody fragment,... – Monoclonal antibody or fragment thereof
Reexamination Certificate
2001-01-26
2003-12-02
Housel, James (Department: 1648)
Drug, bio-affecting and body treating compositions
Immunoglobulin, antiserum, antibody, or antibody fragment,...
Monoclonal antibody or fragment thereof
C424S133100, C530S388300
Reexamination Certificate
active
06656467
ABSTRACT:
BACKGROUND OF THE INVENTION
The present invention relates to novel ultra high affinity neutralizing antibodies.
The current incidence of infection caused by resistant or difficult to control microbes has created a need for newer approaches to controlling such organisms, as well as to treating those already infected.
Among the more difficult infectious agents to control and treat are the viruses. For example, respiratory syncytial virus (RSV) is the major cause of acute respiratory illness in young children admitted to hospitals and the major cause of lower respiratory tract infection in young children. A major obstacle to producing an effective vaccine against such agents as RSV has been the issue of safety. Conversely, the use of immunoglobulins against such viral agents has proven of some value. For example, studies have shown that high-titred RSV immunoglobulin was effective both in prophylaxis and therapy for RSV infections in animal models.
An alternative approach has been the development of antibodies, especially neutralizing monoclonal antibodies, with high specific neutralizing activity. One drawback to this route has been the need to produce human antibodies rather than those of mouse or rat and thus minimize the development of human anti-mouse or anti-rat antibody responses, which potentially results in further immune pathology.
An alternative approach has been the production of human-murine chimeric antibodies in which the genes encoding the mouse heavy and light chain variable regions have been coupled to the genes for human heavy and light chain constant regions to produce chimeric, or hybrid, antibodies.
In some cases, mouse CDRs have been grafted onto human constant and framework regions with some of the mouse framework amino acids being substituted for correspondingly positioned human amino acids to provide a “humanized” antibody. [Queen, U.S. Pat. Nos. 5,693,761 and 5,693,762]. However, such antibodies contain intact mouse CDR regions and have met with mixed effectiveness, producing affinities often no higher than 10
7
to 10
8
M
−1
.
A humanized anti-RSV antibody with good affinity has been prepared and is currently being marketed. [See: Johnson, U.S. Pat. No. 5,824,307]
The production of ultra high affinity antibodies would be desirable from the point of view of both the neutralizing ability of such an antibody as well as from the more practical aspects of needing to produce less antibody in order to achieve a desirable degree of clinical effectiveness, thereby cutting costs of production and/or allowing a greater degree of clinical effectiveness for administration in the same volume.
BRIEF SUMMARY OF THE INVENTION
The present invention relates to high affinity neutralizing antibodies and active fragments thereof exhibiting affinity constants of at least 10
10
M
−1
, and even 10
11
M
−1
, and more specifically to such a neutralizing monoclonal immunoglobulin, including antibodies and/or active fragments thereof, wherein the antibody and/or fragment has human constant regions.
The present invention solves the above-mentioned problems by providing high affinity neutralizing antibodies without the presence of intact mouse CDR regions that cause human anti-mouse antibody reactions (HAMA) and with sufficiently high affinity neutralizing activity to reduce cost and efficacy of overall production.
One aspect of the present invention relates to high affinity neutralizing antibodies with affinity constants of at least 10
10
M
−1
, and even 10
11
M
−1
, and with specificity towards specific antigenic determinants, such as those exhibited by virus-expressed proteins.
One object of the present invention is to provide such high affinity neutralizing antibodies with specificity toward antigens produced by viruses, such as where such antigens are expressed by virus-infected cells in a mammal, especially a human.
In one such embodiment, the high affinity neutralizing immunoglobulin, including antibodies and/or active fragments thereof, of the present invention, and active fragments thereof, are specific for respiratory syncytial virus (RSV), most especially for the F antigen expressed by said RSV and also expressed on the surfaces of cells infected with RSV (the presence of which antigen on the cell surface causes fusion of the cells into syncytia).
Thus, in one embodiment, a high affinity neutralizing immunoglobulin, including antibodies and/or active fragments thereof, of the present invention binds to the same epitope on RSV as the antibody whose light chain variable chain has the sequence of SEQ ID NO: 1 (shown in
FIG. 1A
) and whose heavy chain variable chain has the sequence of SEQ ID NO: 2 (shown in FIG.
1
B).
It is an object of the present invention to provide ultra high affinity neutralizing antibodies having substantially the framework regions of the immunoglobulin disclosed in
FIG. 1
(with the same specificity of that immunoglobulin, which is an anti-RSV antibody structure) but wherein the immunoglobulins, including antibodies and active fragments thereof, of the present invention contain one or more CDRs (complementarity determining regions) whose amino acid sequences are independent of those in the so-called reference antibody, although said sequences may, in some cases, differ by no more than one amino acid and this may be limited to a difference in only one of said CDR regions.
In a preferred embodiment of the present invention, the novel immunoglobulins of the present invention will differ from the antibody of
FIG. 1
(hereafter, the “basic antibody” or “reference antibody” or “reference immunoglobulin”) only in the sequences of one or more of the CDRs and, in a most preferred embodiment these differences occur only in CDRs L2, L3, H1, and H3.
Especially preferred embodiments of the present invention have the framework sequences depicted in
FIG. 1
, thus having the heavy and light chain variable sequences depicted in
FIGS. 3
,
4
,
5
,
6
, and
7
.
In one embodiment, the high affinity neutralizing antibodies of the invention include a human constant region.
In a preferred embodiment, a high affinity RSV-neutralizing antibody of the invention, including active fragments thereof, with an affinity constant (K
a
) of at least as high as 10
10
M
−1
, and even 10
11
M
−1
, is a recombinant immunoglobulin, such as an antibody or active fragment thereof, that includes a human constant region and framework regions for the heavy and light chains wherein at least a portion of the framework is derived from a human antibody (or from a consensus sequence of a human antibody framework), an example of said framework regions depicted for the antibody sequences of FIG.
1
.
In one embodiment, all of the framework is derived from a human antibody (or a human consensus sequence).
In another highly specific embodiment, a high affinity RSV-neutralizing antibody, with an affinity of at least 10
10
M
−1
, is a recombinant antibody having a human constant region, one or more CDRs that are derived from a non-human antibody in which at least one of the amino acids in at least one of the CDRs is changed and in which all or a portion of the framework is derived from a human antibody (or a consensus sequence of a human antibody framework).
In a separate embodiment, a humanized neutralizing immunoglobulin that binds to the same epitope as the basic or reference antibody or immunoglobulin whose variable chains are shown in
FIG. 1
, and that has an affinity of at least 10
11
M
−1
, includes at least one of the following amino acids at the following positions of the CDRs: an alanine at position 2 of CDR H1, a phenylalanine at position 6 of CDR H3, a phenylalanine at position 3 of CDR L2, and a phenylalanine at position 5 of CDR L3. Other embodiments comprise other single amino acid substitutions at these locations.
It is another object of the present invention to provide compositions comprising the immunoglobulins disclosed herein wherein said structures are suspended in a pharmacologically accepta
Huse William D.
Johnson Leslie S.
Watkins Jeffry D.
Wu Herren
Young James F.
Foley Shanon
Grant Alan J.
Housel James
MedImmune, Inc.
Olstein Elliot M.
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