Effect of melatonin treatment on the intratumor heterogeneity in breast cancer model


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André Lima Mota1,Bruna Victorasso Jardim Perassi1,Thaiz Ferraz Borin1,Nathália Martins Sonehara1,Marina Gobbe Moschetta Gobbe Moschetta1,Lorena Pozzo2,Emerson Soares Bernardes2,Bruno Cogliati3,Debora Aparecida Pires de Campos Zuccari1,Tialfi Bergamin de Castro4
1Faculdade de Medicina de São José do Rio Preto - FAMERP,2Instituto de Pesquisas Energéticas e Nucleares (IPEN),3Faculdade de Medicina Veterinária e Zootecnia - USP,4UNESP - Universidade Estadual Paulista "Júlio de Mesquita Filho"

Abstract

Background

Cancer progression is strongly influenced by intratumor heterogeneity, which can be resultant of acidification of tumor microenvironment by hypoxia and preferential anaerobic glycolysis. In this context, melatonin, has shown several antitumor actions, as immunomodulatory, antioxidant, pro-apoptotic, antiangiogenic and possibly can control the intratumor heterogeneity. The aims of this study were to evaluate the action of melatonin on intratumor heterogeneity in breast cancer by in vitro and in vivo studies.

Method

Triple-negative breast cancer cells (MDA-MB-231) were implanted in Balb/c nude mice (n=14), which were treated with melatonin (n=7) or vehicle (n=7). At the end, micro-PET/CT scanning was performed with 18F-FDG, which is an analogue of glucose, and with 18F-FAZA, a specific radiopharmaceutical for hypoxia detection. Then, markers of hypoxia (pimonidazole, HIF-1α, GLUT-1/3, CA-IX/XII) were evaluated by immunohistochemistry in mammary tumors. To in vitro studies MDA-MB-231 cells were treated with melatonin or vehicle under normal pH and low pH conditions (acidosis) and gene and protein expression of hypoxia markers were evaluated by qPCR and immunocytochemistry.

Results

The results showed that there was a lower tumor growth in animals treated with melatonin (p<0.05). The PET/CT images showed that in some tumors with high volume, there was no uptake of 18F-FDG in the tumor center, corresponding to areas of necrosis, and there was an intratumor 18F-FAZA uptake heterogeneity, indicating possible hypoxic areas. All markers of hypoxia evaluated by immunohistochemistry showed less expression in melatonin treated tumors (p<0.05). Also, the in vitro results were correlated with the in vivo study, showing that melatonin has effectiveness under acidosis conditions.

Conclusion

Taken together, our results showed an important action of melatonin in control the adverse conditions in tumor microenvironment and control the cancer progression.