Method and composition for altering a B cell mediated pathology
View Patent ↗The present invention provides a method for altering a B cell mediated pathology in a patient. This method comprises administering a composition comprising at least one and/or two chimeric proteins. Each chimeric protein comprises at least a portion of either the V H or V L region of a immunoglobulin molecule from particular B cells from a patient having a B cell mediated pathology, and an immunoglobulin constant region. The genes encoding V H and/or V L regions and the genes encoding immunoglobulin constant regions are isolated and inserted into an expression vector. The chimeric proteins are produced by introducing the expression vectors into insect cell lines. The chimeric proteins are purified using antibody affinity columns, and then chemically conjugated to an immunogenic carrier, keyhole-limpet hemocyanin (KLH). Since the conjugates comprise chimeric proteins made specifically from particular B cells from a patient having B cell mediated pathology, when it is administered to such a patient, with or without a cytokine, such as granulocyte-macrophage-CSF, or a chemokine, it can induce immune responses to alter such a B cell mediated pathology.
1. A method for treating a B cell lymphoma in a patient, said method comprising: administering a composition comprising two different proteins to said patient and eliciting an active immune response; wherein (1) the first said protein comprises a full length V H region from said patient and at least a portion of an immunoglobulin constant region, (2) the second said protein comprises a full length V L region from said patient and at least an antigenic portion of an immunoglobulin constant region, and (3) wherein (a) the nucleotide sequences of said V H and said V L region are isolated from a malignant B cell clone from said patient having said B cell lymphoma and wherein said proteins are produced in insect cells wherein V H and said V L sequences are inserted into an expression vector to allow for separate expression of said proteins by said insect cells: and (b) said expression vector has two separate expression cassettes each having a baculovirus promoter and a heterologous secretory signal sequence, wherein (i) said promoters are selected from the group consisting of a p10 promoter and a polyhedrin promoter, and (ii) said secretory signal sequences are selected from the group consisting of human placental alkaline phosphatase secretory signal sequence, honey bee melittin secretory signal sequence, and the endogenous secretory signal sequence associated with the immunoglobulin genes isolated from said patient; and (c) said proteins are produced by introducing said expression vector in insect cells.
2. The method of claim 1 wherein said second protein comprises an immunoglobulin constant region comprising a human kappa or lambda constant region.
3. The method of claim 1 wherein said first protein comprises an immunoglobulin constant region selected from the group consisting of a human IgG γ1 constant region, a human IgG γ2 constant region, a human IgG γ3 constant region, a human IgG γ4 constant region, a human IgA 1 constant region, a human IgA 2 constant region, a human IgM constant region, a human IgD constant region, and a human IgE constant region.
4. The method of claim 3 wherein said first protein comprises an immunoglobulin constant region comprising a human IgG γ1 constant region.
5. The method of claim 1 wherein said different proteins are conjugated to a carrier protein.
6. The method of claim 5 wherein said carrier protein is a keyhole-limpet hemocyanin (KLH).
7. The method of claim 1 wherein said composition is further co-administered with a cytokine or chemokine.
8. The method of claim 7 wherein said cytokine is granulocytemacrophage-colony stimulating factor (GM-CSF).
9. The method of claim 1 wherein said first and second different proteins comprise a protein comprising said V H region and a human IgG γ1 constant region and a protein comprising said V L region and a human kappa or lambda chain constant region.
10. The method of claim 1 wherein said insect cells are the Trichoplusia ni cell line or the Spodoptera frugiperda cell line.
11. The method of claim 1 wherein said different proteins are analyzed for expression by ELISA.
12. The method of claim 1 wherein said different proteins are isolated using a protein selected from the group consisting of protein A, protein G, protein L and an anti-immunoglobulin antibody.
13. The method of claim 1 wherein said B cell lymphoma is refractory low grade lymphoma or follicular B cell lymphoma.