(M1130-03-19) Targeting LSD1 Results in the Upregulation of ASGPR1, Offering a Promising Avenue to Improve the Therapeutic Potential of LYTAC-Driven Extracellular Protein Degradation in Liver Cancer

Purpose: Lysosomal targeting chimeras (LYTAC) represent a novel therapeutic platform designed to mediate the degradation of extracellular proteins. In hepatocytes, LYTACs function through the asialoglycoprotein receptor 1 (ASGPR1). However, in hepatic tumor cells, ASGPR1 expression is often downregulated, limiting the therapeutic efficacy of this approach. In this study, we aim to identify regulatory proteins that control ASGPR1 expression, with the goal of uncovering targets that could enhance LYTAC-based therapies through combination strategies.

Methods: To identify candidate genes regulating ASGPR1 expression, we analyzed gene expression profiles from The Cancer Genome Atlas (TCGA) using the GEPIA2 and TIMER2 web-based platforms. Subsequently, we treated HEP3B cells with LSD1 inhibitor GSK-LSD1 and assessed ASGPR1 expression levels via western blot analysis.

Results: Initially, we analyzed ASGPR1 expression across various tumor types using TCGA data accessed through the GEPIA2 platform. Our analysis revealed that ASGPR1 is highly expressed in normal hepatic and bile duct cells. Notably, its expression was significantly downregulated in hepatocellular carcinoma (LIHC) and cholangiocarcinoma (CHOL). To identify potential regulators of ASGPR1 expression, we conducted a screen of epigenetic modulators. Among them, LSD1 (KDM1A) emerged as a candidate, displaying a negative correlation with ASGPR1 expression in both LIHC and CHOL. While the correlation in CHOL was not statistically significant (p = 0.081), a strong and highly significant inverse relationship was observed in LIHC (p = 1.14e-08). Based on these findings, we treated HEP3B liver cancer cells with GSK-LSD1, a highly potent and selective LSD1 inhibitor, for 48 hours. Western blot analysis revealed a marked, dose-dependent upregulation of ASGPR1 expression following LSD1 inhibition. These results suggest that LSD1 negatively regulates ASGPR1 and that pharmacological inhibition of LSD1 may enhance ASGPR1 expression. This upregulation has potential therapeutic relevance by improving the efficacy of LYTAC-mediated delivery of extracellular proteins for lysosomal degradation.

Conclusion: Our findings demonstrate that LSD1 negatively regulates ASGPR1 expression in liver cancer, particularly in hepatocellular carcinoma. Pharmacological inhibition of LSD1 using GSK-LSD1 significantly upregulates ASGPR1 in a dose-dependent manner. These results suggest that targeting LSD1 may be a viable strategy to enhance ASGPR1-mediated LYTAC efficacy, offering a promising approach for improving extracellular protein degradation in hepatic tumors through combination therapy.

References: Liu, C., Chen, K., & Chen, P. (2015). Treatment of liver cancer. Cold Spring Harbor Perspectives in Medicine, 5(9), a021535.  https://doi.org/10.1101/cshperspect.a021535
Mamun, M. a. A., Zhang, Y., Zhao, J., Shen, D., Guo, T., Zheng, Y., Zhao, L., & Liu, H. (2023). LSD1: an emerging face in altering the tumor microenvironment and enhancing immune checkpoint therapy. Journal of Biomedical Science, 30(1). https://doi.org/10.1186/s12929-023-00952-0
Ramadas, B., Pain, P. K., & Manna, D. (2021). LYTACS: an emerging tool for the degradation of Non‐Cytosolic proteins. ChemMedChem, 16(19), 2951–2953. https://doi.org/10.1002/cmdc.202100393
Sun, Y., Sha, Y., Cui, G., Meng, F., & Zhong, Z. (2022). Lysosomal-mediated drug release and activation for cancer therapy and immunotherapy. Advanced Drug Delivery Reviews, 192, 114624. https://doi.org/10.1016/j.addr.2022.114624
Banik, S. M., Pedram, K., Wisnovsky, S., Ahn, G., Riley, N. M., & Bertozzi, C. R. (2020). Lysosome-targeting chimaeras for degradation of extracellular proteins. Nature, 584(7820), 291–297. https://doi.org/10.1038/s41586-020-2545-9

Acknowledgements: The authors declare no conflicts of interest.


Figure 1: A) Schematic presentation of LYTAC B) Schematic presentation of LYTAC mediated drug or protein delivery to lysosome for drug delivery or protein degradation


Figure 2: A) Expression of ASGPR across various tumor and normal tissue B) Comparative analysis of LSD1 and ASGPR1 expression in LIHC C) Comparative analysis of ASGPR1 expression in CHOL.


Figure 3: Western blot data presenting the expression of ASGPR1 in response to LSD1 inhibitor in a dose dependent manner