Jin-An Jiao

A soluble single-chain T-cell receptor IL-2 fusion protein retains MHC-restricted peptide specificity and IL-2 bioactivity.

Antibody-based targeted immunotherapy has shown promise as an approach to treat cancer. However, many known tumor-associated antigens are not expressed as integral membrane proteins and cannot be utilized as targets for antibody-based therapeutics. In order to expand the limited target range of antibodies, we have constructed a soluble single-chain T-cell receptor (TCR) fusion protein designated 264scTCR/IL-2. This fusion protein is comprised of a three-domain HLA-A2-restricted TCR specific for a peptide epitope of the human p53 tumor suppressor protein, which is overexpressed in a broad range of human malignancies. The 264scTCR/IL-2 fusion protein has been expressed at high levels in mammalian cells, and milligram quantities have been purified. MHC-restricted antigen-specific binding properties are maintained in the single-chain, three-domain TCR portion of the fusion protein, and the IL-2 portion retains bioactivity similar to that of free recombinant IL-2. Moreover, this fusion protein is capable of conjugating target and effector cells, remains intact in the blood and substantially increases the half life of the IL-2 portion of the molecule. Finally, the 264scTCR/IL-2 fusion protein can be used to stain tumor cells and is capable of reducing lung metastases in an experimental model of metastasis. Thus, TCR-based fusion proteins may provide a novel class of targeted immunotherapeutics for cancer.

In vitro and in vivo characterization of a novel antibody-like single-chain TCR human IgG1 fusion protein.

We have constructed a protein composed of a soluble single-chain TCR genetically linked to the constant domain of an IgG1 H chain. The Ag recognition portion of the protein binds to an unmutated peptide derived from human p53 (aa 264–272) presented in the context of HLA-A2.1, whereas the IgG1 H chain provides effector functions. The protein is capable of forming dimers, specifically staining tumor cells and promoting target and effector cell conjugation. The protein also has potent antitumor effects in an in vivo tumor model and can mediate cell killing by Ab-dependent cellular cytotoxicity. Therefore, single-chain TCRs linked to IgG1 H chains behave like Abs but possess the ability to recognize Ags derived from intracellular targets. These fusion proteins represent a novel group of immunotherapeutics that have the potential to expand the range of tumors available for targeted therapies beyond those currently addressed by the conventional Ab-based approach.

Inhibition of Acute Vascular Thrombosis in Chimpanzees by an Anti-Human Tissue Factor Antibody Targeting the Factor X Binding Site

Tissue factor (TF) antagonists targeting the factor VII (FVII) binding domain have been shown to interrupt acute vascular thrombus formation without impairing haemostasis in non-human primates. In this study, we evaluate whether a human/mouse chimeric monoclonal antibody (ALT-836, formerly known as Sunol-cH36) blocking the factor X/factor IX (FX/FIX) binding site of tissue factor could achieve similar clinical benefits in an arterial thrombosis model induced by surgical endarterectomy in chimpanzees. In this model, sequential surgical endarterectomies on right and left superficial femoral arteries were performed 30 days apart in five chimpanzees. A bolus (1 mg/kg) of ALT-836 was injected intravenously immediately preceding the restoration of flow in the endarterectomised femoral artery. Pre-surgical labelling of autologous platelets using (111)In-Oxine and post-surgical gamma camera imaging of (111)In-platelet deposition at endarterectomy sites was performed. The manipulated arterial segments were harvested for patency analysis 30 days following surgery. The results indicate that ALT-836 was highly effective at reducing acute vascular thrombosis, with no significant variations in surgical blood loss and template-bleeding time in the treated group compared to the control animals. These data suggest that ALT-836 is an effective and safe antithrombotic agent in preventing TF-initiated vascular thrombogenesis without compromising haemostasis.