Lin Aifu and other teams of Zhejiang University jointly developed a post-translational modification based on immune checkpoint molecules...

In recent years, the immune checkpoint blockade therapy with PD-1/PD-L1 inhibitors as the typical representative has made important progress
in tumor treatment.
However, due to the individual heterogeneity and high complexity of anti-tumor immune regulation, currently approved immune checkpoint inhibitors are only effective in a small number of patients, and in the vast majority of unselected solid tumors, the efficacy rate of PD-1 or PD-L1 inhibitors alone is only 10%-30%.
Related therapies generally have bottlenecks
such as low response rate and limited treatment scenarios in some patients.
In-depth revelation and elucidating of the molecular regulatory mechanism of related tumor immune checkpoints and tumor immune escape mechanism may bring new hope
to tumor patients.

Post-translational modification of proteins is one of the important cell biological events that determine their functional diversity and maintain the precision and complexity of
cell activity regulation.
As the core node molecule of tumor immune evasion, post-translational modifications such as glycosylation, ubiquitination, palmitoylation, acetylation and phosphorylation of immune checkpoints such as PD-L1 play an important regulatory role in their maturation, degradation and translocation, and in-depth search for post-translational modification regulators of immune checkpoint molecules and elucidating their specific regulatory mechanisms may provide new targets and strategies for tumor immunotherapy, which has important scientific research significance and clinical transformation value
。

Recently, Professor Lin Aifu, Professor Zhou Tianhua, Professor Zhou Ruhong of Zhejiang University, Professor Wenqi Wang of the University of California, Irvine, and Professor Li Xu of the School of Life Sciences of Westlake University jointly published a joint paper on Nature Communications published a publication titled " Promoting anti-tumor immunity by targeting TMUB1, a modulator of PD-L1 polyubiquitination and glycosylation", revealing targeting The PD-L1 post-translational modification regulator TMUB1 can effectively reverse the inhibitory tumor immune microenvironment, and the development of related immunotherapy combined with targeted strategies has important clinical transformation value
.

Through in-depth investigation and analysis of clinical big data, the researchers found that important post-translational modification pathways such as glycosylation and ubiquitination were highly enriched in tumor patients with high expression of PD-L1, and were significantly correlated with low response rate of PD-1/PD-L1 monoclonal antibody.

The further discovery of PD-L1 has a higher level of post-translational modification in tumor tissues, suggesting that the regulatory mechanism of PD-L1 post-translational modification is revealed in depth or a potential targeting strategy
for related immunotherapies.
Based on the above preliminary findings, the team successfully screened a batch of PD-L1 candidate interaction molecules by combining mass spectrometry and bioinformation analysis, among which transmembrane and ubiquitin-like domain protein 1 (TMUB1), which is a quality control element of endoplasmic reticulum protein, is highly expressed in gastric cancer, breast cancer and other tumor tissues and was highly negatively correlated with CD8+ T cell infiltration, suggesting that it has a potential regulatory function
in PD-L1-mediated tumor immune escape.
In-depth mechanistic studies confirm that TMUB1 plays an important role
in the regulation of multiple post-translational modifications of PD-L1.
On the one hand, TMUB1 localized by the endoplasmic reticulum is greatly weakened by competitively binding PD-L1 to the E3 ubiquitin ligase HUWE1 The ubiquitination pathway of PD-L1 is degraded, which in turn enhances the protein stability of PD-L1.
On the other hand, TMUB1 promotes PD-L1 glycosylation modification, maturation, cell membrane translocation and tumor microenvironment regulation by recruiting related glycosyltransferases, which ultimately leads to immune evasion
of tumors.

Figure 1: Schematic diagram of the core mechanism of clinical analysis of TMUB1 and its participation in PD-L1 post-translational modification in breast cancer patients

The above findings on the regulatory mechanism of TMUB1 on PD-L1 post-translational modification suggest the clinical potential
of TMUB1 as a combination target of tumor immunotherapy.
With the help of multi-team cross-advantages, through computational structure simulation, binding segment screening, molecular design of peptide precursor drugs and in vivo and in vivo series of functional verification, the researchers designed and developed a targeted polypeptide PTPR, which inhibits the binding of TMUB1 to PD-L1.
Successfully blocked the multiplex post-translational modification regulation of PD-L1 by TMUB1, and achieved significant inhibition of tumor growth in various tumor-bearing mouse models such as breast cancer and gastric cancer, and verified the in vivo biosafety
of the peptide inhibitor.
Furthermore, the combination with αCTLA4 and other immune checkpoint molecular monoclonal antibody drugs successfully reversed the inhibitory tumor immune microenvironment, greatly enhanced the infiltration of cytotoxic T cells and the specific killing of tumor cells, and finally achieved tumor suppression
.

Figure 2: Structural docking simulation of targeted peptide PTPR and efficacy in a 4T1-tumor-bearing mouse model

In summary, this study revealed the regulatory mechanism of PD-L1 post-translational modification and the molecular mechanism of related tumor immune escape, and preclinical evidence showed that TMUB1 could be used as a reliable joint target for immunotherapy, and the developed peptide procursor drug molecule further demonstrated the targeting of PD-L1 Post-translational modification modulates molecular strategies with important clinical translational value
.
This research is the research direction of Lin Aifu's team in the regulation of tumor microenvironment, and after revealing the mechanism of tumor immune microenvironment remodeling mediated by signals such as calcium ions (Molecular cell, 2018; Nature Metabolism, 2021) and the mechanism of tumorigenesis regulated by metabolic signals of lipids such as cholesterol (Cell Research, 2021; Nature Metabolism, 2022).

Drs.
Chengyu Shi, Ying Wang and Minjie Wu of Zhejiang University are co-first authors
of the paper.
The research work has also been strongly supported by collaborators such as Professor Shao Jianzhong, Professor Xu Pinglong, Professor Dong Xiaowu, Professor Wang Liangjing, Professor Yan Qingfeng and Professor Yu Luyang
。 The work was supported
by the National Key Research and Development Program of the Ministry of Science and Technology, the National Natural Science Foundation of China, the Natural Science Foundation of Zhejiang Province, the Star Science Foundation of the Shanghai Advanced Research Institute of Zhejiang University, the Scientific Research Project Fund of the Qilu Joint Research Institute of Zhejiang University, and the **** of Zhejiang University.
Professor Lin Aifu's research group welcomes young talents who are interested in tumor biology and RNA biomedicine to join us (providing postdoctoral fellows, distinguished researchers, associate researchers and other positions).

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