Cell–cell communication is crucial for ensuring the health of organisms and cells. This communication occurs at the cell surface and is facilitated by the glycans observed on proteins and lipids. N-glycan processing is typical process for the construction of such glycans. During this process, Golgi mannosidase (GM) plays an important role in exposing a core glycan structure to aid N-glycan maturation. When GM is inhibited, the transportation of immature N-glycans to the cell surface may damage the cell–cell communication. The known GM inhibitor interferes with the cell–cell communication by inhibiting N-glycan maturation. Hence, the anticancer activity can be observed. However, attempts to use the inhibitor for anticancer drug applications were unsuccessful. We considered two issues that may have caused this failure based on the inhibitor properties, including low hydrophilicity and the number of asymmetric centers. To address these issues, we screened GM inhibitors from natural compound libraries and modified the obtained inhibitors. Consequently, four compounds, including three approved drugs, were observed to be GM inhibitors. Two among these, tamoxifen and raloxifene, were estrogen receptor antagonists, whereas the remaining drug, sulindac, was a non-steroidal anti-inflammatory drug. Diphenylpropylamine derivatives, called the AR500 series compounds, were also observed to be GM inhibitors possessing low hydrophilicity and one asymmetric center (R. Koyama, et al., Chem. Pharm. Bull. (Tokyo). 66, 678, 2018).
To develop suitable GM inhibitors for inhibiting cell–cell communication, the structure–activity relation should be analyzed. However, the bioactivity and synthesis methods for the AR500 series are not reported. Thus, a method suitable for synthesizing the structurally diverse derivatives of the AR500 series compounds was established and used to clarify the structure–activity relationship. Based on this method, 27 structurally diverse derivatives were synthesized. These were subsequently evaluated in the monolayer HeLa cell-based assays that determined the GM inhibitory activity, high-mannose glycan accumulation, and morphological changes based on the inhibition of cell adsorption with respect to the plastic flasks. All the compounds were observed to cause accumulation of high-mannose glycans based on the GM inhibition at sub- to 10-µM levels. Similarly, morphological changes were observed in the cells based on high-mannose glycan accumulation. Finally, we evaluated the inhibition activity of the spheroid formation of HeLa cells to achieve cell–cell communication inhibition based on GM inhibition. A spheroid is commonly considered to be a cancer tumor model. Consequently, some compounds exhibited objective activity at the sub- to 10-µM level; however, the amide derivatives exhibited significantly reduced activity. These biological results indicated that some AR500 series compounds exhibited cell–cell communication inhibition based on high-mannose glycan accumulation by GM inhibition without cytotoxicity. Also, the analysis of the structure–activity relations indicated that almost all the AR500 series derivatives exhibited targeted activity except for the amide derivatives.
In the future, continuing the aforementioned evaluation of the AR500 series derivatives and analyzing the detailed mechanism of bioactivity expression are necessary to apply these compounds as anticancer drugs. Particularly, the relation between the cell–cell communication inhibition and the maturation rate of N-glycans on the cell surface should be clarified by analyzing the cells separated from a GM-inhibitor-treated spheroid using flow cytometry.