Oxoisoaporphine Alkaloid Iridium(III) Derivative: An Immunogenic Cell Death Inducer That Engages the Autophagy-Dependent Regulator Cathepsin D

Autophagy has been recognized as one of the pathways for eliciting immunogenic cell death (ICD). However, the specific molecular target responsible for autophagy-mediated ICD has not yet been elucidated. Here, we report that an oxoisoaporphine alkaloid-modified iridium(III) complex (2a) displays autophagy-inducing ICD activity. Through unbiased thermal proteome profiling (TPP), this new complex was found to interact with the lysosomal protease cathepsin D (Cat D). Subsequent cellular and biochemical assays─including the cellular thermal shift assay, isothermal dose-response assay, enzymatic assays, and molecular docking─confirmed that 2a binds to and inhibits Cat D. Further pathway analysis demonstrated that 2a triggers autophagy-dependent ICD via the LKB1-AMPK-ULK1 signaling pathway by inhibiting Cat D. Several other autophagy-dependent ICD inducers were tested and likewise found to inhibit Cat D. In contrast, an earlier reported analogue of 2a, complex 1a, was found to bind and destabilize binding immunoglobulin protein (BiP) and promote its ICD activity through an endoplasmic reticulum stress response. We believe that the findings reported here will enhance the understanding of the novel mechanisms of ICD agents and pave the way for the design of new ICD inducers with high specificity and efficacy.