Activity of the mTOR protein kinase and the NOS2 enzyme with colony formation capacity from human colorectal cancer and melanoma strains

• Grant number: 20/10675-9
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Effective date (Start): January 01, 2021
• Effective date (End): December 31, 2021
• Field of knowledge: Biological Sciences - Biochemistry - Molecular Biology
• Principal Investigator: Hugo Pequeno Monteiro
• Grantee: Letícia Torres Barboza
• Host Institution: Centro de Terapia Celular e Molecular. Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil
• Associated research grant: 18/15038-7 - Tumor development under the perspective of redox signaling: temporal modulation of the production of nitric oxide and reactive oxygen species, AP.TEM

Abstract

Colorectal cancer (CRC) and human melanoma are characterized by their aggressiveness and metastatic power, associated with a poor prognosis for patients. These properties are related to changes in cell homeostasis, characterized by genetic mutations, excessive production of reactive species, hypoxia, shortage of nutrients, among other factors. These changes enable metabolic reprogramming that promotes the malignant phenotype of cells. The PI3K / Akt / mTOR signaling pathway has its activity modulated by the supply of nutrients and is altered in most cancers, acting in an important way in tumorigenesis. It has recently been demonstrated that the imposition of nutrient deficiency on tumor cells for determined periods, promotes an adaptive survival response in these cells with production of pro-inflammatory cytokines (PUSCHELA, et al., 2020). In addition, it is well documented that the inducible isoform of the enzyme nitric oxide (NO) synthase inducible (NOS2) and NO, a free radical with signaling properties, produced by this enzyme, play very important roles in the development of various cancers, included there CCR and melanoma. NOS2 / NO stimulate phosphorylation and s-nitrosylation of proteins constituting oncogenic signaling pathways, activating these pathways. Another important observation is that the lack of nutrients (removal of fetal serum from the culture medium) stimulates the expression of NOS2 with a concomitant increase in intracellular levels of NO in cells of the metastatic lineage of triple-negative breast tumor, MDA-MB -231 (HEINECKE et al., 2014). Thus, we are proposing that the condition of nutrient restriction can establish a connection between the PI3K / Akt / mTOR signaling pathway and the induction of NOS2 expression and NO production. Furthermore, this connection could occur through the transcription factor NFkB and be responsible for the adaptive survival response of tumor cells when they are in this condition. In order to explore this hypothesis, we intend to determine the contributions of the protein kinase mTOR, the enzyme NOS2 and the transcription factor NFkB in the capacity of survival and colony formation in human strains of melanoma (SKMEL-28) and CCR (SW620) originating from metastases. The strains will be maintained in a condition of nutrient restriction and scarcity of intercellular contact, in the presence and absence of inhibitors of the two enzymes and the transcription factor. These variables will be assessed for their relevance to cell proliferation and the mechanisms by which the tumor cell develops its resistance in stressful situations. The in-depth knowledge about cell signaling mediated by mTOR and NOS2 in metastatic tumor cells can be extremely useful in elucidating the problems associated with the low effectiveness of therapies in the treatment of tumors in an advanced stage of development (metastasis). The information obtained from our study may also be important in the identification of new targets and specific therapeutic approaches, less invasive and more functional.