Tarin-Loaded Nanoliposomes Activate Apoptosis and Autophagy and Inhibit the Migration of Human Mammary Adenocarcinoma Cells

Abstract

Raiane Vieira Cardoso,1 Patricia Ribeiro Pereira,1 Cyntia Silva Freitas,1 Érika Bertozzi de Aquino Mattos,1 Anna Victoria De Freitas Silva,2 Victor do Valle Midlej,2 Mauricio Afonso Vericimo,3 Carlos Adam Conte-Júnior,1 Vania Margaret Flosi Paschoalin1 1Departamento de Bioquímica, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brazil; 2Instituto Oswaldo Cruz, Rio de Janeiro, RJ, Brazil; 3Departamento de Imunobiologia; Universidade Federal Fluminense, Niterói, RJ, BrazilCorrespondence: Vania Margaret Flosi Paschoalin, Av. Athos da Silveira Ramos 149 – sala 545 - Cidade Universitária, Rio de Janeiro, 21941-909, RJ, Brazil, Tel [phone], Fax [phone], Email [email protected]: Tarin, a lectin purified from Colocasia esculenta, promotes in vitro and in vivo immunomodulatory effects allied to promising anticancer and antimetastatic effects against human adenocarcinoma mammary cells. This makes this 47 kDa-protein a natural candidate against human breast cancer, a leading cause of death among women. Tarin encapsulated in pegylated nanoliposomes displays increased effectiveness in controlling the proliferation of a mammary adenocarcinoma lineage comprising MDA-MB-231 cells.Methods: The mechanisms enrolled in anticancer and antimetastatic responses were investigated by treating MDA-MB-231 cells with nano-encapsulated tarin at 72 μg/mL for up to 48h through flow cytometry and transmission electron microscopy (TEM). The safety of nano-encapsulated tarin towards healthy tissue was also assessed by the resazurin viability assay, and the effect of nanoencapsulated tarin on cell migration was evaluated by scratch assays.Results: Ultrastructural analyses of MDA-MB-231 cells exposed to nanoencapsulated tarin revealed the accumulation of autophagosomes and damaged organelles, compatible with autophagy-dependent cell death. On the other hand, the flow cytometry investigation detected the increased occurrence of acidic vacuolar organelles, a late autophagosome trait, along with the enhanced presence of apoptotic cells, activated caspase-3/7, and cell cycle arrest at G0/G1. No deleterious effects were observed in healthy fibroblast cells following tarin nanoencapsulated exposition, in contrast to reduced viability in cells exposed to free tarin. The migration of MDA-MB-231 cells was inhibited by nano-encapsulated tarin, with delayed movement by 24 h compared to free tarin.Conclusion: The nanoliposome formulation delivers tarin in a delayed and sustained manner, as evidenced by the belated and potent antitumoral and anti-migration effects on adenocarcinoma cells, with no toxicity to healthy cells. Although further investigations are required to fully understand antitumorigenic tarin mechanisms, the activation of both apoptotic and autophagic machineries along with the caspase-3/7 pathway, and cell cycle arrest may comprise a part of these mechanisms. Keywords: tarin-controlled release, MDA-MDB-231 cells damage, kinetics of ultrastructural changes, cell cycle arrest, cell migration assay, caspase 3/7 pathway

Keywords

• tarin-controlled release
• mda-mdb-231 cells damage
• kinetics of ultrastructural changes
• cell cycle arrest
• cell migration assay
• caspase 3/7 pathway.