Biological Evaluation of a Series of Thiosemicarbazones Targeting the Large Subunit Ribosomal Protein eL42 from Human 80S Ribosomes.

Abstract

Background: Thiosemicarbazones are a family known as excellent chelators of enzyme cofactors or molecules with multiple biological activities including anti-tumor activity.1,2 Recent studies by Hountondji’s team has revealed that the large subunit ribosomal protein eL42 (molecular weight of 12 kDa, pHi: 10,59) plays a functional role during the elongation step of translation3 and is over expressed in most cancers.4–6

Method: The method used was the one developed by Hountondji et al.7 which converts the OH functional groups at the positions 2' and 3' of adenine sugar of the CCA tRNA into a dialdehyde group by oxidation with NaIO4. Each aldehyde function can form a Schiff base with the amino group of the side chain of K53 residue of eL42, which results in a covalent bond after reduction with NaBH4 (crosslinking).

Results and discussion: The results of different experiments (the radioactivity, and the Western Blot technique) performed in this study show that 4 molecules (citral-4-phenylthiosemicarbazone, piperitone-4-phenylthiosemicarbazone, the benzoin-4-phenylthiosemicarbazone and 4-chlorobenzophenone-4-phenylthiosemicarbazone) out of 29 thiosemicarbazones tested showed activity by inhibiting the crosslinking reaction between tRNAox and eL42. These results suggest that the inhibition produced by thiosemicarbazones consist in masking the amino group of the side chain of Lys-53 located in the catalytic site of protein eL42, preventing Schiff base formation and the crosslinking reaction. These results are in accordance with those of Schneider-Poetsch et al.8 who found that certain compounds for example cycloheximide bind to the eL42 protein through their carbonyl function. The weaker effects of benzoin-4-phenylthiosemicarbazone and 4-chlorobenzophenone-4-phenylthiosemicarbazone as compared with citral-4-phenylthiosemicarbazone and piperitone-4- phenylthiosemicarbazone are likely to reflect the inaccessibility of the NH2 of Lys-53 of eL42 to the thiocarbonyl groups (C=S) of these thiosemicarbazones because of steric hindrance. Finally, binding assays on Biacore 3000 confirmed interactions between active molecules and the human eL42 protein.

Conclusion: The inhibitory action of thiosemicarbazones on the interaction between the protein eL42 and tRNAox prevents formation of a Schiff base between the ε-amino group of Lys-53 and an aldehyde group of tRNAox. The Lys-53 residue of eL42 which contributes to the catalysis of peptide bond formation that represents the key step in the biosynthesis of proteins is a target of choice for small molecule inhibitors. Therefore, thiosemicarbazones can be used to reduce the rate of protein synthesis in order to block the hyper-proliferation of tumor cells (Figure 1).