Methods of producing secreted receptor analogs and biologically active dimerized polypeptide fusions

A method for producing a secreted, biologically active dimerized polypeptide fusion, comprising:

• introducing into a eukaryotic host cell a first DNA construct comprising a transcriptional promoter operatively linked to a first secretory signal sequence followed downstream by and in proper reading frame with a first DNA sequence encoding a non-immunoglobulin polypeptide requiring dimerization for biological activity joined to an immunoglobulin light chain constant region;
• introducing into said host cell a second DNA construct comprising a transcriptional promoter operatively linked to a second secretory signal sequence followed downstream by and in proper reading frame with a second DNA sequence encoding an immunoglobulin heavy chain constant region domain selected from the group consisting of C.sub.H 1, C.sub.H 2, C.sub.H 3, and C.sub.H 4, and optionally encoding an immunoglobulin hinge or variable region joined in proper reading frame to said immunoglobulin heavy chain constant region;
• growing said host cell in an appropriate growth medium under physiological conditions to allow the secretion of a biologically active dimerized polypeptide fusion encoded by said first and second DNA sequences;
• and
• isolating said biologically active dimerized polypeptide fusion from said host cell.

A method for producing a secreted, biologically active dimerized polypeptide fusion, comprising:

• introducing into a eukaryotic host cell a first DNA construct comprising a transcriptional promoter operatively linked to a first secretory signal sequence followed downstream by and in proper reading frame with a first DNA sequence encoding a non-immunoglobulin polypeptide requiring dimerization for biological activity joined to an immunoglobulin heavy chain constant region domain selected from the group consisting of C.sub.H 1, C.sub.H 2, C.sub.H 3, and C.sub.H 4, and optionally encoding an immunoglobulin hinge or variable region joined in proper reading frame to said immunoglobulin heavy chain constant region;
• introducing into said host cell a second DNA construct comprising a transcriptional promoter operatively linked to a second secretory signal sequence followed downstream by and in proper reading frame with a second DNA sequence encoding an immunoglobulin light chain constant region, and optionally encoding immunoglobulin variable region joined in proper reading frame to said immunoglobulin heavy chain constant region;
• growing said host cell in an appropriate growth medium under physiological conditions to allow the secretion of a biologically active dimerized polypeptide fusion encoded by said first and second DNA sequences;
• and isolating said biologically active dimerized polypeptide fusion from said host cell.

A method for producing a secreted receptor analog, comprising:

• introducing into a eukaryotic host cell a DNA construct comprising a transcriptional promoter operatively linked to at least one secretory signal sequence followed downstream by and in proper reading frame with a DNA sequence encoding a ligand-binding domain of a receptor requiring dimerization for biological activity joined to a dimerizing protein heterologous to said ligand-binding domain, and optionally an immunoglobulin hinge or variable region to said dimerizing protein;
• growing said host cell in an appropriate growth medium under physiological conditions to allow the secretion of a receptor analog encoded by said DNA sequence;
• and
• isolating said receptor analog from said host cell.

A method for producing a secreted PDGF receptor analog, comprising:

• introducing into a cultured rodent myeloma cell a first DNA construct comprising a mouse V.sub.H promoter operatively linked to a PDGF receptor signal sequence followed downstream of and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIGS. 11A-11D (Sequence ID Numbers 35 and 36) from glutamine, number 24 to glutamic acid, number 524, joined to an immunoglobulin heavy chain constant region domain selected from the group consisting of C.sub.H 1, C.sub.H 2, C.sub.H 3, and C.sub.H 4 joined to an immunoglobulin hinge region;
• introducing into said cultured rodent myeloma cell a second DNA construct comprising a mouse V.sub..kappa. promoter operatively linked to a PDGF receptor signal sequence followed downstream of and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIGS. 1A-1D (Sequence ID Numbers 1 and 2) from isoleucine number 29, to lysine, number 531, joined to an immunoglobulin light chain constant region;
• growing said cultured rodent myeloma cell in an appropriate growth medium under physiological conditions to allow the secretion of a PDGF receptor analog encoded by said first and second DNA sequences;
• and
• isolating said PDGF receptor analog from said cultured myeloma cell.

A method for producing a secreted active multimerized polypeptide fusion, comprising:

• introducing into a eukaryotic host cell a DNA construct comprising a transcriptional promoter operatively linked to a secretory signal sequence followed downstream by and in proper reading frame with a DNA sequence encoding a non-immunoglobulin polypeptide requiring multimerization for biological activity joined to a multimerizing protein selected from the group consisting of:
• an immunoglobulin heavy chain or light chain constant region, and optionally encoding an immunoglobulin hinge or variable region joined to said multimerizing protein;
• growing said host cell in an appropriate growth medium under physiological conditions to allow the secretion of a multimerized polypeptide fusion encoded by said DNA sequence;
• and
• isolating said multimerized polypeptide fusion from said host cell.

A method for producing a secreted active heteromultimeric polypeptide fusion, comprising:

• introducing into a eukaryotic host cell a first DNA construct comprising a transcriptional promoter operatively linked to a secretory signal sequence followed downstream by and in proper reading frame with a first DNA sequence encoding a first polypeptide fusion comprising a first non-immunoglobulin polypeptide joined to a first multimerizing protein selected from the group consisting of:
• an immunoglobulin heavy chain or light chain constant region, and optionally encoding an immunoglobulin hinge or variable region joined to said first multimerizing protein, and a second DNA construct comprising a transcriptional promoter operatively linked to a secretory signal sequence followed downstream by and in proper reading frame with a second DNA sequence encoding a second polypeptide fusion comprising a second non-immunoglobulin polypeptide joined to a second multimerizing protein selected from the group consisting of:
• an immunoglobulin heavy chain or light chain constant region, and optionally encoding an immunoglobulin hinge or variable region joined to said second multimerizing protein;
• growing said host cell in an appropriate growth medium under physiological conditions to allow secretion of said heteromultimeric polypeptide fusion encoded by said DNA sequences;
• and
• isolating said heteromultimeric polypeptide fusion from said host cell.