Abstract
Bifunctional degraders, also referred to as proteolysis-targeting chimeras (PROTACs), are a recently developed class of small molecules. They were designed to specifically target endogenous proteins for ubiquitin/proteasome-dependent degradation and to thereby interfere with pathological mechanisms of diseases, including cancer. In this study, we hypothesized that this process of acute pharmacologic protein degradation might increase the direct MHC class I presentation of degraded targets. By studying this question, we contribute to an ongoing discussion about the origin of peptides feeding the MHC class I presentation pathway. Two scenarios have been postulated: peptides can either be derived from homeostatic turnover of mature proteins and/or from short-lived defective ribosomal products (DRiPs), but currently, it is still unclear to what ratio and efficiency both pathways contribute to the overall MHC class I presentation. We therefore generated the intrinsically stable model antigen GFP-S8L-F12 that was susceptible to acute pharmacologic degradation via the previously described degradation tag (dTAG) system. Using different murine cell lines, we show here that the bifunctional molecule dTAG-7 induced rapid proteasome-dependent degradation of GFP-S8L-F12 and simultaneously increased its direct presentation on MHC class I molecules. Using the same model in a doxycycline-inducible setting, we could further show that stable, mature antigen was the major source of peptides presented, thereby excluding a dominant role of DRiPs in our system. This study is, to our knowledge, the first to investigate targeted pharmacologic protein degradation in the context of antigen presentation and our data point toward future applications by strategically combining therapies using bifunctional degraders with their stimulating effect on direct MHC class I presentation.
Highlights
Small molecules targeting endogenous proteins for proteolytic degradation have been suggested as a novel therapeutic strategy against diseases, including cancer
These experiments demonstrate that we generated an expression system that could be used to study the impact of degradation tag (dTAG)-7-mediated antigen degradation on its MHC class I presentation
We could show in various experimental setups and murine cell types, including a fibrosarcoma, a dendritic, and a macrophage cell line, that the induction of antigen degradation by dTAG-7 strongly increased the direct MHC class I presentation of the degraded target
Summary
Small molecules targeting endogenous proteins for proteolytic degradation have been suggested as a novel therapeutic strategy against diseases, including cancer. Bifunctional degraders act by linking the protein of interest to a specific E3 ligase complex that in turn induces E2-mediated ubiquitination and proteasomal degradation of the recruited target This strategy has recently been utilized to induce the degradation of BRD4, a transcriptional co-activator involved in cancer and other diseases [6,7,8, 10]. Apart from their potent effect on protein stability, which leads to the anticipated interference with the biological function of a given target, we hypothesized that bifunctional degraders might alter the immunological visibility of degraded proteins by increasing their presentation on MHC class I molecules. This study highlights a novel feature of therapies based on bifunctional degraders that could be used to increase CD8+ T-cell immunity towards cancer and intracellular infections
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