Development and evaluation of biological reagents targeting and inhibiting function of the EphA3 receptor on tumor cells

Eph受体及其配体调节胚胎的形态发生;它们在组织层和器官系统的形成过程中指导细胞的迁移和定位。几乎没有证据表明Ephs在成人组织中的功能。然而，它们在各种恶性肿瘤中大量、非计划性地出现，表明它们在癌症中起作用。人EphA 3是本提案的主要主题，在成人组织中未发现，但在肺、肾和脑肿瘤、白血病和恶性黑色素瘤中以高水平存在。高水平的EphA 3和相应的配体与黑素瘤进展相关，并且EphA 3刺激触发黑素瘤细胞的排斥和脱离。Eph A3可能参与黑色素瘤进展期间肿瘤细胞的释放和扩散。我们已经表征了试剂，可溶性EphA 3配体和单克隆抗EphA 3抗体，其以高亲和力和特异性结合EphA 3。我们将使用这两种蛋白质，或含有附加放射性化学物质或细胞毒素的修饰形式，靶向植入免疫缺陷小鼠作为动物模型系统的人类肿瘤。我们的研究将确定我们的试剂的特异性，从以前的体外研究中得到的建议，是否将允许含有EphA 3的肿瘤的成像，并影响它们的靶向杀伤。我们还将使用含有皮肤细胞的人工表皮和真皮层的组织培养模型来研究EphA 3配体的抑制形式是否会影响EphA 3阳性转移性黑素瘤细胞的侵袭性。此外，我们将确定这种配体的重要组成部分，以开发具有改进药理学特性的抑制剂。总之，我们的研究将确定EphA 3在癌症进展中的作用，并评估EphA 3靶向杀死肿瘤和预防转移的功效。我们设想，这将为在癌症治疗中具有抗转移作用的Eph特异性试剂提供基础。

Eph receptors and their ligands regulate morphogenesis in the embryo; they direct migration and positioning of cells during the formation of tissue layers and organ systems. There is little evidence for a function of Ephs in adult tissues. However, their abundant, un-scheduled occurrence in various malignant tumours, indicates a role in cancer. Human EphA3, the principle subject of this proposal, is not found in adult tissue but is present at high levels in lung, kidney and brain tumours, leukemia and malignant melanoma. High levels of EphA3 and corresponding ligands correlate with melanoma progression, and EphA3 stimulation triggers repulsion and detachment of melanoma cells. It is likely that Eph A3 is involved in release and spreading of tumour cells during melanoma progression. We have characterised reagents, the soluble EphA3 ligand and a monoclonal anti-EphA3 antibody, which bind EphA3 with high affinity and specificity. We will use these two proteins, or modified forms containing attached radiochemicals or cytotoxins, to target human tumours that were implanted into into immuno-deficient mice as animal model system. Our studies will determine if the specificity of our reagents, suggested from previous in-vitro studies, will allow imaging of EphA3 containing tumours, and effect their targeted killing. We will also use a tissue culture model, containing artificial epidermal and dermal layers of skin cells, to study if an inhibitory form of the EphA3 ligand will affect the invasiveness of EphA3 positive, metastatic melanoma cells. Furthermore, we will identify essential parts of this ligand to develop inhibitors with improved pharmacological properties. Together, our studies will establish the role for EphA3 in cancer progression and to assess the efficacy of EphA3 targeting for tumor killing and prevention of metastasis. We envision that this will provide the groundwork for Eph-specific reagents with anti-metastatic action in cancer therapy.