Defined by monoclonal antibody GA733, the GA733-2 antigen is a cell surface 40-kDa glycoprotein associated with human carcinomas of various origins. Molecular clones for the GA733-2 antigen were isolated from a colorectal carcinoma cell line cDNA library using the high-efficiency COS cell expression system. A 1.4-kilobase cDNA species was enriched by immunoselection with monoclonal antibody. The authenticity of individual clones was established by immunologic and sequence criteria. At the amino acid sequence level, GA733-2 was found to be >99% identical to the previously described KSA antigen defined by monoclonal antibody KS1/4. The amino acid sequence derived from the previously described GA733-related gene, GA733-1, was found to be 49% identical to GA733-2. The positions of 12 cysteine residues in the extracellular domains of the two GA733 antigens are conserved, as is the overall distribution of hydrophobic and hydrophilic residues. A 1.45-kilobase transcript of the GA733-2/KSA gene was found to be expressed in cell lines derived from colorectal and pancreatic carcinoma.in a colorectal carcinoma cell line. Both GA733-2 and GA733-1 sequences have significant homology to an exon-encoded repeat unit in thyroglobulin and the HLA-DR-associated invariant chain.Here we report the isolation of cDNA clones for the GA733-2 antigen using the high-efficiency COS cell expression system (7-9) and the sequence of the GA733-2 cDNA.t Molecular clones for the GA733-2 antigen were enriched from a colorectal carcinoma cell line cDNA library by iterating cycles of expression of the plasmid library in COS cells, immunoselection of antigen-positive cells by "panning" with the mAb GA733, and transfer of plasmid DNA from COS cells into Escherichia coli. The sequence of GA733-2 was compared to that of the carcinoma-associated antigen KSA (10, 11) and to the related GA733-1 antigen (6). A group of related mAbs (1, 12, 13) were assayed for reactivity to the antigen expressed by the GA733-2 clone. The size and expression of the GA733-2 mRNA in tumor cell lines were also studied.Monoclonal antibodies (mAbs) defining tumor-associated cell surface antigens are currently being evaluated for use in the diagnosis and therapy ofhuman cancer. Initial studies of mAbs C017-1A and GA733 (1), demonstrating cytotoxic activity in vitro and tumoricidal activity in experimental animals (for review, see ref.2), led to an evaluation of their immunotherapeutic potential in cancer patients. Idiotypic cascades have been demonstrated in cancer patients treated with mAb C017-1A, providing support to the hypothesis that antigen-specific anti-anti-idiotypic antibody (Ab3) responses may explain some of the observed delayed clinical responses (3).The independently derived mAbs GA733 and C017-1A define the same 40-kDa cell surface glycoprotein (4), hereafter referred to as the GA733-2 antigen. It is expressed by carcinomas of several histologic types and to some extent by normal tissues (5). The molecular cloning of cDNA for the tumor-associated GA733-2 an...

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