Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Peptide containing doai
Reexamination Certificate
2000-02-15
2001-09-11
Leary, Louise N. (Department: 1623)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Peptide containing doai
C536S023500, C530S300000, C530S325000, C530S326000, C530S327000, C530S328000, C530S330000, C424S184100
Reexamination Certificate
active
06288035
ABSTRACT:
The present invention relates generally to the field of immunotherapy based on the identification and isolation of human tumor antigens. In particular embodiments, the invention also relates to the use of human monoclonal antibodies to isolate the epitopic sequence of a tumor-associated antigen. Further embodiments relate to synthetic peptides derived from tumor-associated proteins and their uses for therapy, diagnosis and prognosis of human cancer.
The ideal tumor vaccine is one that induces anti-tumor immunity without adverse effects. Ideally, such a vaccine would be stable, inexpensive to produce and easily administered. Early attempts with whole cell vaccines or cellular extracts, have improved the survival of cancer patients significantly. However, problems still exist with the production, storage and delivery of the vaccines. For example, there still exists the possibility of contamination by cellular molecules that may induce side effects in patients.
Because of these problems, synthetic or recombinant antigens are contemplated to be more effective vaccines, provided the antigen delivery system, carrier molecules, and adjuvants are optimized for the induction of specific anti-tumor immunity. What is needed is the identification and characterization of tumor antigens that play important roles in tumor destruction in vivo, in particular those capable of inducing T-cell immunity in humans.
Because elevated titers of autoantibodies are known to be present in hosts with various autoimmune diseases, neuropathological diseases and cancers, they have been used to screen cDNA clones from expression libraries in order to discover pathogenic antigens of such diseases (Tan E M, 1991; Amagai M et al, 1991; Dropcho E J et al., 1987; Szabo A et al., 1991; Hayashibe K et al., 1991). The difficulties of using this approach to identify human tumor associated antigens (TAA) include the unavailability of high titer human anti-TAA antibodies and the interference of serum antibodies unrelated to TAA.
Melanoma associated antigens (AU and FD) unique to autologous melanoma cells have been detected using autologous sera (Carey T E et al., 1979). The FD antigen is a 90 kD cell-surface glycoprotein. The antigenic determinant is present on an ion-binding protein with amino acid homology to transferrin (Real F X et al., 1984). The same molecule was identified by murine monoclonal antibodies developed against the human melanoma-associated antigen p97 (Brown J P et al, 1982). Although the sequence of p97 is known, the sequence of the unique FD epitope has not yet been determined. The Hellstrom laboratory has investigated the 97,000 MW glycoprotein antigen, p97, that is predominantly expressed by human melanoma cells (Woodbury R G et al., 1980; Brown J P et al., 1981). The p97 gene has been inserted into a recombinant vaccinia virus and has induced strong anti-tumor immunity against mouse melanoma cells transfected to express the p97 antigen (Brown J P et al., 1982). A clinical trial with this vaccine is ongoing by this group (Hellstrom I et al., 1992).
Vlock and colleagues used autologous melanoma sera after dissociating serum immune complexes via acidic treatment and found an antigenic 66 kD acidic glycoprotein (Vlock D R et al., 1988). Subsequent epitope analysis has shown that the carbohydrate moiety of the glycoprotein represents its antigenic determinant. The sequence of the core protein has not been reported.
Allogeneic polyclonal sera from melanoma patients have also been used to identify immunogenic melanoma associated antigens. These antigens are shared by more than one melanoma. Bystryn and colleagues detected immunogenic melanoma associated antigens (200+, 150, 110, 75, and 38 kD) using sera from patients who received immunotherapy with melanoma cell supernatant (Li J et al., 1990).
Gupta and associates have defined a urinary tumor associated antigen (UTAA), which is a glycoprotein antigen originally found in the urine of melanoma patients and also found on the melanoma cell surface, including the M14 cell line. The UTAA is comprised of several subunits that are linked together by disulfide bonds. The total molecular weight is approximately 300 kD, with immunogenic subunits of 45 kD, 65 kD, 90 kD, 120 kD and 150 kD (Euhus D M et al., 1990).
Ferrone and colleagues used pooled sera from melanoma patients to identify a 50 kD glycoprotein antigen (D-1) in melanoma (Hayashibe K et al., 1991). Using cDNA libraries of a melanoma cell line, the immunoreactive clone was isolated and sequenced. The successful cloning was possible after extensive absorption of non-specific antibodies with
E. coli
proteins and blocking of cDNA plaques on nitrocellulose with IgG isolated from the pooled sera of healthy donors. While the exact tumor specificity of this protein (D-1) is not known, northern blot hybridization has shown that peripheral blood lymphocytes (PBL) and normal fibroblasts do not express this antigen.
Several other studies have identified protein molecules from other human cancer cells that migrate at or around the 43 kD position in SDS-PAGE Western blot analysis. These studies have used murine or human monoclonal antibodies. HGP43 is a human glycoprotein of 43 kDa, which was identified by murine monoclonal antibody and was originally reported as a protective antibody against lethal
Listeria monocytogenes
infection in mice (Fontan E et al, 1992). HGP43 is detected in the urine of healthy, normal individuals as well as in cancer patients. HGP43 has also been shown to stimulate mouse monocytes to induce cytotoxicity against the Lewis lung tumor (Fontan E et al., 1993). The amino acid sequence from HGP43 that is reactive to the monoclonal antibody has not been reported.
The human monoclonal antibodies 16.88 and C-OU 1 react with another 43 kD protein molecule in human cancer cells (Erb K et al., 1991). The antigen is most strongly expressed in melanoma and less strongly in colon cancer cells. The amino acid sequence of this protein is partially known and exhibits about 70% identity with cytokeratin 18.
Another HuMAb, MS2B6, has been used to detect a cytoplasmic antigen present at high density in ovarian carcinoma and is less prevalent in a variety of human cancers and in certain types of normal tissues (Smith L H et al., 1992). On SDS-PAGE Western blot analysis, MS2B6 reacted to proteins with a broad range of molecular weights, 33-44 kD and 60 kD. Competitive inhibition studies have shown no crossreactivity between the antigen identified with MS2B6 and the cytokeratin identified with HuMAbs 16.88 and C-OU 1.
Another approach to developing anti-tumor vaccines is the development of anti-idiotype vaccines. In this method, MAbs are developed against an antigen of interest. These antibodies are then used to screen an epitope library to find a peptide that mimics the target antigen. This anti-id epitope is then used as a vaccine. This is particularly important in developing antibodies to glycosidic antigens which cannot be easily synthesized. Dr. Ferrone's laboratory has pursued an investigation of anti-id vaccines mimicking high molecular weight melanoma associated antigens (HMW-MAAs) for the active specific immunotherapy of melanoma (Kusama M et al., 1989). The HMW-MAAs are expressed in high density by melanoma cells, but have a restricted distribution on normal tissues (Reisfeld R A and Cheresh D A, 1987). Anti-ids mimicking HMW-MAAs were developed using syngeneic murine monoclonal antibodies to MAAs as immunogens. When these anti-id vaccines were injected in mice and rabbits, specific immune responses were demonstrate (Kusama M et al., 1989; Challopadhyay P et al., 1991). Injection of these vaccines into patients during a Phase I clinical trial has shown that the anti-id vaccine is safe and has induced clinical responses in patients with melanoma, including 2 complete remissions (Mittelman A et al., 1992). However, the high anti-tumor antibody or T-cell responses observed in animal models have not been replicated in clinical trials. The weak immune responses seen in patients by immunizati
Irie Reiko F.
Oka Takanori
Fulbright & Jaworski LLP
John Wayne Cancer Institute
Leary Louise N.
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