Method for preparation of single chain antibodies

A method for making a single chain antibody (scFv) directed against an antigen, wherein the selection of clones is made based upon interaction of those clones with an appropriate anti-idiotype, and heretofore inaccessible scFv so made.

A method for selecting cell lines that package and produce high titers of retroviral particles that carry the scFv gene for transfection into cells.

A method for inducing proliferation in a population of T cells comprising the steps of (a) introducing to the T cells an expressable gene sequence encoding a chimeric single chain antibody (scFv) coupled to a transmembrane and signaling domain;

• and (b) exposing the T cells to which the chimeric scFv has been introduced to an anti-idiotype directed to an antibody specific for a target such that the anti-idiotype will bind to scFv on the surface of cells expressing the chimeric scFv thereby inducing proliferation of the T cells.

A method for tagging cells to facilitate sorting or enrichment, comprising the steps of expressing a single chain antibody (scFv) in the cell and capturing or tagging the cells using anti-idiotype.

A method for isolating an antigen comprising the steps of preparing an antibody to the antigen, preparing an anti-idiotype directed to the antibody, using the anti-idiotype to select a single chain antibody (scFv), targeting the antigen from a scFv library, and using the selected scFv as an affinity probe to capture antigen, preferably with the scFv immobilized on a solid support.

A method for treating cancer in a patient suffering from cancer expressing an antigenic marker comprising the steps of removing lymphocytes from the patient, introducing to the lymphocytes an expressable gene sequence encoding a chimeric single chain antibody (scFv) coupled to a transmembrane and signaling domain;

• exposing the lymphocytes to which the chimeric scFv has been introduced to an anti-idiotype directed to an antibody specific for a target antigen to which the scFv is directed under conditions such that the anti-idiotype will bind to scFv on the surface of cells expressing the chimeric scFv and any necessary co stimulatory molecules to induce proliferation of the lymphocytes;
• and returning the expanded population of lymphocytes to the patient.

A method for treating cancer in a patient suffering from cancer expressing an antigenic marker comprising the steps of introducing to human cell lines an expressable gene sequence encoding a chimeric single chain antibody (scFv) coupled to a transmembrane and signaling domain (including zeta chain);

• exposing the lymphocytes to which the chimeric scFv has been introduced to an anti-idiotype directed to an antibody specific for a target antigen to which the scFv is directed under conditions such that the anti-idiotype will bind to scFv on the surface of cells expressing the chimeric scFv and any necessary co stimulatory molecules to immunoselect and stimulate clones with high density of scFv expression and efficient tumor cytotoxicity to produce a gene-modified cell line, and returning the expanded population of gene-modified cell line to the patient.

A method for enhancing in vivo survival and anti-tumor activity of infused lymphocytes gene-modified with scFv-Chimeric immune receptors by intravenous injection of anti-idiotype antibody.