Study of the integrative systems biology (transcriptomics, proteomics and metabolomics) of placentas from pregnant Wistar rats carrying Walker 256 tumor. Evaluation of the effects of tumor growth and exposure to chemotherapy treatment during pregnancy

• Grant number: 25/00594-5
• Support Opportunities: Scholarships in Brazil - Master
• Start date: March 01, 2025
• End date: February 28, 2027
• Field of knowledge: Biological Sciences - Physiology - Physiology of Organs and Systems
• Principal Investigator: Laís Rosa Viana
• Grantee: Geisiéllen Moreira da Cunha
• Host Institution: Instituto de Biologia (IB). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil
• Associated research grant: 21/08931-0 - Cancer and pregnancy: proteomic, transcriptomic and metabolic alterations in placenta from pregnant women with cancer and Wistar pregnant Walker 256 tumour-bearing rats, AP.JP

Abstract

Cancer during pregnancy represents a major therapeutic challenge, since maternal and fetal/neonatal health must be considered. Women diagnosed with cancer during pregnancy may present alterations in the placenta, the main organ responsible for ensuring the success of the pregnancy and fetal health. Studies found in the literature combining the terms "placenta", "proteomics" and "cancer" presented a lack of results. Thus, the application of proteomics can contribute to the mapping of protein pathways potentially altered in the placenta, due to cancer and/or oncological treatment, especially chemotherapy, thus enabling a search for new therapeutic strategies that may be beneficial for mother and fetus. Preclinical experiments using animal models are necessary, as they allow greater manipulation/intervention and control of variables. In this sense, Walker 256 carcinosarcoma is a tumor model widely used in research. Thus, the objective of this study is to identify and compare changes in the proteomic profile of placentas of pregnant rats with Walker 256 carcinosarcoma, treated or not with the chemotherapeutic agents Doxorubicin and Cyclophosphamide. For this purpose, pregnant Wistar rats will be distributed into four experimental groups: Control/Healthy (C); Walker 256 Tumor (W); Chemotherapy treatment (Q); Walker 256 Tumor and Chemotherapy treatment (WQ). On the second day after pregnancy is identified, the Walker 256 tumor will be inoculated into the right flank of the rats. Regarding the chemotherapeutic agents, they will be administered intraperitoneally on the fourteenth and seventeenth days of pregnancy, with doses of 0.1 mg/kg of Doxorubicin and 1 mg/kg of Cyclophosphamide. Euthanasia will occur on the twentieth day of pregnancy, that is, one day before the date of delivery, considering that the usual gestation course of these animals lasts on average approximately 21 days. The placentas will then be weighed, collected (considering the sexes independently) and stored at -80º C for proteomic analyses. Such analyses will be performed using a two-dimensional nano UPLC (2D-RP/RP) in an Acquity UPLC M-Class System (Waters Corporation, MA, USA), which is connected in line to a Synapt G2-Si mass spectrometer (Waters Corporation). The mass spectrometer performs Independent Data Acquisition, specifically employing Ultra High Definition Data Independent Mass Spectrometry (UDMSE). The data obtained through the proteomic analysis will be compared between the four experimental groups in order to identify changes in protein expression, seeking to map possible mechanisms of increased or decreased regulation, or even proteins found exclusively in one of the groups. Protein identifications will be based on the detection of more than two fragmented ions per peptide and at least two peptides per protein, with only proteins modulated with p ¿ 0.05 considered significant. After protein identification by LC-MS/MS, Progenesis software will be used for label-free quantitative analysis, using the integrated relative abundance intensity considering all identified peptides for normalization. Cytoscape and STRING will be used for protein interaction analysis. And for pathway enrichment analyses, Gene Ontology, DAVID, Reactome and KEEG will be used as databases.