开发基于cetuximab抗体靶向工程酶A4H363A的ADEPT肿瘤靶向疗法

前期，我们已经建立并验证了突变半乳糖苷酶A4H363A/6-甲基半乳糖苷前药(MeGal prodrug)的专一性激活体系(Nature Chemical Biololgy. 2019,15(2):151-160)。MeGal保护前药不被小鼠内源性酶识别，而被经过凸凹互补改造的A4H363A特异性激活。本申请将使用cetuximab抗体或其重组蛋白与改造的半乳糖苷酶A4H363A偶联(cetuximab-A4H363A)实现酶对A549(人非小细胞肺癌细胞)异源裸鼠成瘤的靶向递送。合成并筛选geldanamycin，nannocystin A(1e)，nannocystin A(1e)内酰胺类似物以及10-hydroxycamptothecin (10-HCPT)的MeGal糖苷前药，联合cetuximab-A4H363A评价此抗体导向酶－前药肿瘤治疗(ADEPT)的靶向和抑瘤效果。

Based on the "bump-and-hole" strategy, which has been proven to be powerful for generating high-affinity protein-ligand variant pairs, we developed a novel enzyme/prodrug activation system (Nature Chemical Biololgy. 2019,15(2):151-160). First, methylation at the O-6 position of a galactosyl molecule introduced a “bump”that completely disrupts its recognition by the endogenous galactosidase (β-gal). The selectivity of an engineered β-gal (A4H363A) for the 6-methylated derivative (MeGal-drug) was achieved by introducing a“hole” at its active pocket in order to accommodate the bumpy 6-methyl group. .In this proposal, a series of prodrugs including MeGal modified geldanamycin, nannocystin A (1e), nannocystin A (1e) lactam derivative and 10-hydroxycamptothecin will be synthesized. These prodrugs will be activated by the tumor-targeted β-gal (A4H363A) coupled with cetuximab antibody for treating Non-small-cell lung carcinoma (NSCLC).