81P - miR-23b and -133a role on TRAIL-induced apoptosis pathway components expression and TRAIL sensitization in lung adenocarcinoma cells

Background

TRAIL-induced apoptosis pathway is a potential therapeutic target, but the majority of tumors are TRAIL-resistant. TRAIL receptors translocation is associated to TRAIL resistance. Besides TRAIL receptors, DR4 and DR5, other potentially TRAIL-induced apoptosis pathway regulatory proteins are: Apaf-1, CUL3, CLTA (clathrin chain A , involved in TRAIL receptors nuclear translocation) and KPNA-1 ( involved in TRAIL receptors internalization). Overcome TRAIL resistance is essential to therapeutic strategies development.

Methods

A549 cell line was used. Expression of miR-23b, miR-133a, DR4, DR5, Apaf-1, CUL3, CLTA/CLTC and KPNA-1 were estimated by RT-qPCR. Transfection effects of miR-23b inhibitor and Mimic-133a on TRAIL apoptosis pathway components expression. Western Blotting to TRAIL receptors compartmentalization after Mimic-133a and TRAIL stimulus. MTT cell viability assay to evaluate TRAIL-induced cytotoxicity of miR-23b inhibition and Mimic-133a transfection.

Results

A549 was TRAIL resistant. miR-23b was superexpressed and miR-23b inhibition upregulated Apaf-1 and CUL3 expression. miR-133a was undetectable. Mimic-133a transfection downregulated CLTA and DR5 expression. miR-23b inhibition and Mimic-133a transfection promoted TRAIL sensitization.TRAIL receptors were predominantly located in nuclear compartment. Mimic-133a downregulated CLTA expression but did not alter receptors compartmentalization. Mimic-133a and TRAIL associated stimulus leaded to predominant nuclear TRAIL receptors localization in comparison to control cells.

Conclusions

Both miR-23b and -133a are involved in TRAIL cells resistance. Apaf-1 and CUL3 expression downregulation by miR-23b can contribute to resistance. miR-133a role on TRAIL sensitization mechanisms must be further investigated. Mimic-133a downregulated CLTA expression but did not alter TRAIL receptors compartmentalization, suggesting that receptors translocation is clathrin independent.CLTA expression control by miR-133a, however, can contribute to drug-resistance research. Although more research is needed to fully clarify miRs roles on TRAIL-induced apoptosis, our findings deepen TRAIL resistance understanding.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

FAPESP.

Disclosure

All authors have declared no conflicts of interest.