理由：由于癌基因表达产物通常表现出上调或异常激活的活性，因此开发调节异常蛋白质水平的技术代表了治疗肿瘤和蛋白质异常相关疾病的可行方法。方法：首先筛选出具有高靶向降解效率的eMIATAC组分，并探讨eMIATAC诱导靶蛋白降解的机制，并通过蛋白印迹和流式细胞术验证靶蛋白的降解效率。接下来，我们将eMIATAC与一些可控元件重组，以验证目标蛋白的可调控降解性能。随后，我们构建了能够表达AKT1靶向降解的eMIATAC，并在体外和体内验证了其对GBM细胞发育的影响。最后，我们将eMIATAC与CAR序列串联，构建低BATF蛋白水平的CAR-T细胞，并验证其抗肿瘤功效的变化。结果：我们开发了一种基于内体-微自噬-溶酶体途径降解内源蛋白的系统：内体-微自噬靶向嵌合体（eMIATAC），依赖于Vps4A而不是溶酶体相关膜蛋白2A（LAMP2A）与伴侣Hsc70结合，并且感兴趣的蛋白质（POI）。然后该复合物通过晚期内涵体转运至溶酶体，在那里发生类似于微自噬的降解。 eMIATAC 在降解目标蛋白 EGFP 方面表现出准确性、效率、可逆性和可控性。此外，eMIATAC 在体内和体外靶向内源蛋白时在敲低 POI 方面表现出优异的性能。结论：eMIATAC不仅可以直接敲低胶质瘤治疗中的异常蛋白，而且可以通过敲低T细胞耗竭相关蛋白来增强CAR-T细胞治疗肿瘤的治疗效果。新开发的 eMIATAC 系统有望成为蛋白质敲除策略的新型工具。通过直接控制内源性蛋白质水平，eMIATAC 有潜力彻底改变癌症和遗传性疾病的治疗。© 作者。

Rationale: Since oncogene expression products often exhibit upregulation or abnormally activated activity, developing a technique to regulate abnormal protein levels represent a viable approach for treating tumors and protein abnormality-related diseases. Methods: We first screened out eMIATAC components with high targeted degradation efficiency and explored the mechanism by which eMIATAC induced target protein degradation, and verified the degradation efficiency of the target protein by protein imprinting and flow cytometry. Next, we recombined eMIATAC with some controllable elements to verify the regulatable degradation performance of the target protein. Subsequently, we constructed eMIATAC that can express targeted degradation of AKT1 and verified its effect on GBM cell development in vitro and in vivo. Finally, we concatenated eMIATAC with CAR sequences to construct CAR-T cells with low BATF protein levels and verified the changes in their anti-tumor efficacy. Results: we developed a system based on the endosome-microautophagy-lysosome pathway for degrading endogenous proteins: endosome-MicroAutophagy TArgeting Chimera (eMIATAC), dependent on Vps4A instead of lysosomal-associated membrane protein 2A (LAMP2A) to bind to the chaperone Hsc70 and the protein of interest (POI). The complex was then transported to the lysosome by late endosomes, where degradation occurred similarly to microautophagy. The eMIATACs demonstrated accuracy, efficiency, reversibility, and controllability in degrading the target protein EGFP. Moreover, eMIATAC exhibited excellent performance in knocking down POI when targeting endogenous proteins in vivo and in vitro. Conclusions: The eMIATACs could not only directly knock down abnormal proteins for glioma treatment but also enhance the therapeutic effect of CAR-T cell therapy for tumors by knocking down T cell exhaustion-related proteins. The newly developed eMIATAC system holds promise as a novel tool for protein knockdown strategies. By enabling direct control over endogenous protein levels, eMIATAC has the potential to revolutionize treatment for cancer and genetic diseases.© The author(s).