Brazilian study finds therapeutic target and promising diagnostic for the treatment of melanoma -
A Brazilian study shows that the inhibition of RNA called RMEL3, which is encoded by a gene uncharacterized (also the name RMEL3), can reduce the viability of cultured melanoma cells up to 95%.
Although RMEL3 is a non-coding RNA and therefore does not contain information for protein synthesis, it appears to modulate the major signaling pathways related to cell proliferation and survival. How it does this is not fully understood.
"Our research suggests RMEL3 is expressed in most cases of melanoma. Furthermore, this RNA is rarely found in other tumor types, or even in healthy cells, so it is a diagnostic and therapeutic target very specific considerable promise for development, "said Enilza Espreafico, professor at the University of Ribeirão Preto medical school of São Paulo (FMRP-USP) and principal investigator of the FAPESP-funded study.
These results are from the doctoral research by two beneficiaries of FAPESP scholarships, Lucas Goedert and Cristiano Gonçalves Pereira, in collaboration with Cibele Cardoso and other researchers in Brazil and abroad. the existence RMEL3 of and association with melanoma, however, had already been identified in earlier research by Espreafico group.
"at that time, our goal was to identify genes expressed only in cases of melanoma . We used bioinformatics tools to explore the databases created from sequencing projects of the tumor, "said Espreafico.
first analyzes revealed the existence of 29 RNA sequences transcribed only in melanoma cells. three of them appeared most important for the group :. RMEL1 Ribeirao Preto, RMEL2 and RMEL3
the study showed that the three non-coding RNA are proteins present both in melanoma cell lines and patients. RMEL3 of tumor samples even appears in precancerous skin lesions considered.
Next, the researchers conducted in vitro experiments to study how the presence or the absence of these three RNAs changed the cell phenotype. the first results have shown that inhibition of RMEL3 provided the most intense reduction in the viability of melanoma cells in culture, and this RNA has become their favorite target.
To silence RMEL3 in cultured cells, the group deployed RNA interference, a technique of using small molecules noncoding RNA can bind to the RNA transcribed from the target gene (in this case, RMEL3) and inducing its degradation. The effects of this procedure were compared in five different cell lines. The first three were melanoma cell lines with a mutation known to be associated with cancer in a gene called BRAF. The fourth line, also a melanoma cell line, lacking the BRAF mutation. The fifth, regarded as a kind of control group was an ovarian cancer line that did not express RMEL3 and also lacked the BRAF mutation.
"BRAF is the principal proto-oncogene associated with the development of melanoma. About 60% of cases of this type of cancer involves a BRAF mutation, which encodes a protein kinase that initiates the signaling pathway MAPK , an important trigger of cell proliferation, "said Espreafico.
" the mutation changes a single letter of the genetic code in BRAF, which results in the substitution of an amino acid in the polypeptide chain , which is enough to create the protein BRAF V0E proto-oncogene. in its mutant form, this enzyme is intrinsically active, so that the cell enters the replication cycle, even without receiving any external signal for proliferation. " He was in melanoma cells in culture with the BRAF V0E mutation that inhibition of RMEL3 had the most dramatic effect, which reduced cell survival and proliferation to 95%. In the melanoma cell line that did not have the mutation, cell viability decreased by about 40%. In the line of control cells, RNA interference had no effect, and the cells continued proliferating normally.
The researchers still can not say exactly what role that RNA plays in the cell or why it is often present in melanoma cells. However, they found evidence of what happens in the cell when the expression of this RNA is interrupted.
"When we shut RMEL3, there is a decrease in levels of the oncogene BRAF protein kinase and Akt (pAkt), a key protein in the survival of PI3K signaling cells. the opposite effect is observed for PTEN protein, the main inhibitor of this pathway, "said Espreafico.
They also observed increased levels of ACC -pS79, a substrate of AMPK enzyme which is a nutrient deficiency sensor.
"This suggests the absence of RMEL3 induces a state that mimics nutrient deprivation, similar to that reported for pharmacological inhibition of BRAF V0E. In accordance with such changes, cell cycle effectors are altered. There are decreased levels of cyclin B1 protein, important in the activation of cell mitosis. secondly, there are increased levels of proteins that inhibit the cell cycle, such as p27 and p21. These and other changes are consistent with the fact that we have observed in the cells: an increase in cell death and cell cycle arrest, "said Espreafico
To better understand the role of RMEL3 in cells, the. researchers are conducting new experiments in vitro in which its expression is induced artificially in both melanoma cells and healthy cells, where it is normally expressed. The results are due for publication soon.
The group also put on how often RMEL3 is expressed in melanoma cases by analyzing nearly 500 samples from patients with this cancer of The Cancer Genome Atlas (TCGA), a consortium linked to the National Institute of the United States of cancer that collects genomic, epigenomic and clinical patients in many countries.
"We observed that RMEL3 was expressed in a greater or lesser extent over 0% of the melanoma samples available. Together with the fact that it is not present in healthy tissue in the rest of the body, which makes a very interesting therapeutic target, "Espreafico said.
RNA interference itself could have therapeutic uses, she said, noting that many technical barriers should be overcome first.
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