Associação Portuguesa de Investigação em Cancro
A new approach to predict the presence of MYCN gene amplification in pediatric brain tumors by analyzing RNA molecules
A new approach to predict the presence of MYCN gene amplification in pediatric brain tumors by analyzing RNA molecules
Since 2018, the Barretos Cancer Hospital, Barretos, SP, Brazil, has carried out molecular tests to subclassify a brain tumor called medulloblastoma that occurs most often in children, but can also occur in adults. This classification is important to direct more appropriate treatments. The method used to classify pediatric brain tumors involves the use of a state-of-the-art digital technology called NanoString that can detect and quantify hundreds of "RNA" molecules simultaneously and identify exactly which subtype of medulloblastoma.
In addition to this molecular classification, the analysis of other tumor characteristics, such as oncogene amplification, may also be important for medical management and for optimizing the treatment of patients. One oncogene, MYCN, known to stimulate cell proliferation, may be one such factor associated with patients' prognosis. Typically, when there are multiple copies of this oncogene (gene amplification), the tumors are more aggressive and require more intensive treatment.
Most laboratories use a test called fluorescence in situ hybridization (FISH) to detect MYCN amplification. However, this methodology is laborious, expensive, requires high quality biological material, a special microscope to visualize the results and trained professionals for analysis.
In this work, led by Dr. Daniel Moreno and Dr. Rui Manuel Reis, the scientists identified a relationship between the expression of the MYCN mRNA and cases with gene amplification. Subsequently, they established a formula capable of predicting cases with amplification in the gene from the counts of RNA molecules. The method showed 100% specificity and sensitivity, that is, this relationship was verified in all cases evaluated in the study! Subsequently, and through international collaboration involving six research centers in Brazil, Portugal and Argentina, it was possible to analyze more than 200 cases of medulloblastomas, where the molecular classification of the tumor subtype was performed simultaneously with the prediction of the MYCN amplification.
This approach can modify the molecular diagnostic routine of medulloblastomas, without additional cost and without the use of additional biological material, also providing a fast and accurate result.
Institutions involved in the research: Barretos Cancer Hospital, Barretos, SP, Brazil; Ribeirão Preto School of Medicine, Ribeirão Preso, SP, Brazil; AC Camargo Hospital, São Paulo, Brazil; Federal University of São Paulo (UNIFESP), São Paulo, Brazil; Italian Hospital of Buenos Aires, Argentina; Hospital São João, Porto, Portugal.
Funding: Pio XII Foundation, Barretos Cancer Hospital, Public Ministry of Labor (MPT), Campinas (Research, Prevention and Education of Cancer, Brazil), National Support Program for Oncology Care (PRONON), Brazil.
Authors and Affiliations:
Daniel Antunes Moreno 1, Luciane Sussuchi da Silva 1, Maicon Fernando Zanon 2, Murilo Bonatelli 2, Flávia Escremim de Paula 2, Marcus de Medeiros Matsushita 3, Gustavo Ramos Teixeira 3, 4, Iara Viana Vidigal Santana 3, Fabiano Saggioro 5, Luciano Neder 5, João N Stavale 6, Suzana Maria Fleury Malheiros 6, Matheus Lima 7, Glaucia Noeli Maroso Hajj 7, Hernan Garcia-Rivello 8, Silvia Christiansen 8, Susana Nunes 9, Maria João Gil da Costa 9, Maria José Soares 10, Jorge Pinheiro 11, Carlos Almeida Junior 12, Bruna Minniti Mançano 1, 12, Rui Manuel Reis 13, 14, 15, 16
1 Molecular Oncology Research Center Dr. Paulo Prata, Barretos Cancer Hospital, Barretos, Brazil.
2 Molecular Diagnosis Laboratory Dr. Paulo Prata, Barretos Cancer Hospital, Barretos, Brazil.
3 Pathology Department Dr. Paulo Prata, Barretos Cancer Hospital, Barretos, Brazil.
4 Barretos School of Health Sciences Dr. Paulo Prata, Barretos Cancer Hospital, Barretos, Brazil.
5 Department of Pathology and Forensic Medicine, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil.
6 Federal University of São Paulo (UNIFESP), São Paulo, Brazil.
7 AC Camargo Hospital, São Paulo, Brazil.
8 Pathology Department, Italian Hospital of Buenos Aires, Buenos Aires, Argentina.
9 Pediatric Oncology Department, Hospital São João, Porto, Portugal.
10 Clinical Hematology Department, Hospital São João, Porto, Portugal.
11 Department of Pathology, Hospital São João, Porto, Portugal.
12 Pediatric Oncology Department, Pediatric Neurosurgery Department, Barretos Cancer Hospital, Barretos, Brazil.
13 Molecular Oncology Research Center Dr. Paulo Prata, Barretos Cancer Hospital, Barretos, Brazil.
14 Molecular Diagnosis Laboratory Dr. Paulo Prata, Barretos Cancer Hospital, Barretos, Brazil.
15 ICVS/3B's-PT Government Associate Laboratory, Braga/Guimarães, Portugal.
16 School of Medicine, Life and Health Sciences Research Institute (ICVS), University of Minho, Braga, Portugal.
Abstract:
Purpose: Medulloblastoma is the most frequent pediatric malignant brain tumor, and is divided into four main subgroups: WNT, SHH, group 3, and group 4. MYCN amplification is an important medulloblastoma prognostic biomarker. We aimed to molecular classify and predict MYCN amplification in a single assay. Methods: It was included 209 medulloblastomas from 205 patients (Brazil, Argentina, and Portugal), divided into training (n=50) and validation (n=159) sets. A nCounter assay was carried out using a custom panel for molecular classification, with additional genes, including MYCN. nSolver 4.0 software and the R environment were used for profiling and MYCN mRNA analysis. MYCN amplification by FISH was performed in 64 cases. Results: The 205 medulloblastomas were classified in SHH (44.9%), WNT (15.6%), group 3 (18.1%) and group 4 (21.4%). In the training set, MYCN amplification was detected in three SHH medulloblastomas by FISH, which showed significantly higher MYCN mRNA counts than non-FISH amplified cases, and a cutoff for MYCN amplification was established (X + 4σ = 11,124.3). Applying this threshold value in the validation set, we identified MYCN mRNA counts above the cutoff in three cases, which were FISH validated. Conclusion: We successfully stratified medulloblastoma molecular subgroups and predicted MYCN amplification using a single nCounter assay without the requirement of additional biological tissue, costs, or bench time.
Journal: Journal of Neuro-Oncology
Link: https://pubmed.ncbi.nlm.nih.gov/35166989/
