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Construction, expression, and characterization of anticarcinoma sFv fused to IL-2 or GM-CSF
L Zhao1, S K Rai, L S Grosmaire
1Bristol-Myers Squibb Pharmaceutical Research Institute, Seattle, WA 98121, USA.
Abstract:
Local production of cytokines by genetically engineered tumor cells decreases their tumorigenicity and elicits protective immune responses against the parental tumor cells. An alternative approach to elicit a therapeutic immune response is to use fusion proteins that can target tumor cells and simultaneously activate effector cells. Fusion proteins between human IL-2, murine or human GM-CSF, and sFv of antihuman carcinoma antibody L6 have been constructed, expressed in both COS and Chinese hamster ovary (CHO) cells, and purified by affinity chromatography. The biologic activity of L6 sFV-hIL-2, L6 sFv-mGM-CSF, and L6 sFv-hGM-CSF was tested on human T cell blasts, factor-dependent FDCP-1, and TF-1 cells, respectively. The ability of soluble L6 sFv-hIL-2, L6 sFv-mGM-CSF, and L6 sFv-hGM-CSF to stimulate the proliferation of the indicator cells was found to be comparable to that of recombinant hIL-2, mGM-CSF, or hGM-CSF. Tumor cells coated with L6 sFV-mGM-CSF or L6 sFv-hGM-CSF were also tested in this way and were found to be potent stimulators, indicating that the cytokines were functionally active when bound to the tumor cell surface. This work demonstrates the feasibility of targeting sFv-cytokine fusion proteins for the activation of effector cells as an alternative to cytokine gene therapy.
Insights
Engineered fusion proteins targeting tumor cells with cytokines like IL-2 and GM-CSF show promise for cancer immunotherapy. These novel constructs activate immune cells, offering an alternative to traditional cytokine gene therapy.
Area of Science:
- Immunology
- Biotechnology
- Cancer Research
Background:
- Local cytokine production by engineered tumor cells can reduce tumor growth and enhance anti-tumor immunity.
- Fusion proteins offer an alternative strategy to activate immune effector cells by targeting tumor cells.
Purpose of the Study:
- To construct and evaluate fusion proteins combining tumor-targeting antibodies with cytokines (IL-2, GM-CSF) for cancer immunotherapy.
- To assess the biological activity and efficacy of these novel fusion proteins in activating immune responses.
Main Methods:
- Construction and expression of fusion proteins (L6 sFv-hIL-2, L6 sFv-mGM-CSF, L6 sFv-hGM-CSF) in COS and CHO cells.
- Purification via affinity chromatography and biological activity testing on relevant cell lines (T cell blasts, FDCP-1, TF-1).
- Evaluation of cytokine function when bound to tumor cell surfaces.
Main Results:
- Fusion proteins demonstrated comparable biological activity to recombinant cytokines in stimulating indicator cell proliferation.
- Cytokines remained functional when conjugated to tumor cells via the L6 sFv moiety.
- The fusion proteins effectively stimulated effector cells, indicating therapeutic potential.
Conclusions:
- Targeting sFv-cytokine fusion proteins represent a feasible and promising alternative to cytokine gene therapy for cancer treatment.
- These fusion proteins can activate effector cells by targeting tumor cells, enhancing anti-tumor immune responses.