Elucidating the cellular determinants of targeted membrane protein degradation by lysosome-targeting chimeras

Abstract

Targeted protein degradation can provide advantages over inhibition approaches in the development of therapeutic strategies. Lysosome-targeting chimeras (LYTACs) harness receptors, such as the cation-independent mannose 6–phosphate receptor (CI-M6PR), to direct extracellular proteins to lysosomes. In this work, we used a genome-wide CRISPR knockout approach to identify modulators of LYTAC-mediated membrane protein degradation in human cells. We found that disrupting retromer genes improved target degradation by reducing LYTAC recycling to the plasma membrane. Neddylated cullin-3 facilitated LYTAC-complex lysosomal maturation and was a predictive marker for LYTAC efficacy. A substantial fraction of cell surface CI-M6PR remains occupied by endogenous M6P-modified glycoproteins. Thus, inhibition of M6P biosynthesis increased the internalization of LYTAC-target complexes. Our findings inform design strategies for next-generation LYTACs and elucidate aspects of cell surface receptor occupancy and trafficking.

Targeted Extracellular Protein Degradation by Dendronized DNA Chimeras

“…Three branches and T10 were discovered as the optimal components in the DNA dendron, and the linker property might also affect DENTAC performance. In addition, cellular factors determining recycling of extracellular protein degraders and lysosomal maturation also deserve in-depth investigations in the future . We validated the anticancer application of DENTAC by targeting oncogenic MMP-9.…”

Lysosome‐targeting chimera (LYTAC): A silver bullet for targeted degradation of oncogenic membrane proteins

“…Blockage of M6P biosynthesis genes (e.g., ALGO12, GNPTAB) upregulated the ratio of unoccupied receptors, increasing LYTAC-receptor internalization and the degradation of cell surface proteins, including EGFR and c-Met. 1 Overall, this work discovered a series of critical cellular regulators that modulated LYTAC-mediated degradation of EGFR and c-Met (Figure 1B, right), giving important support for understanding LYTAC mechanism and developing nextgeneration LYTAC with improved clinical potential.…”

“…Recently, the group of Prof. Carolyn Bertozzi, a laureate of the Nobel Prize in chemistry 2022, reported the detailed mechanism of lysosome-targeting chimera (LY-TAC) in the journal of Science, 1 after the publication of their first LYTAC molecule in Nature in 2020. 2 The establishment of LYTAC, a subtype of targeted protein degradation technology, expands the scope of protein degradation to extracellular and membrane-associated targets, and Bertozzi group's new discovery is expected to accelerate the development of LYTAC in cancer therapy.…”

Targeted protein degradation directly engaging lysosomes or proteasomes

“…50 Using CRISPR/Cas9-knockout screening, it was found that IGF2R (insulin like growth factor 2 receptor, an alternative gene name for CI-M6PR ) and several genes involved in protein neddylation and activation of the E3 ligase cullin-3 ( CUL3 ) are essential for the cargo uptake and LYTAC recycling. 115 These results provided not only an unbiased validation of the underlying molecular mechanism of LYTACs but also a potential indicator ( e.g. , neddylation of CUL3) to predict whether and to what extent LYTAC therapy will be effective in vivo .…”