Abstract B13: BLM, FOXO3, FOXK2, FOXM1, FOXR1 genes as therapeutic targets to neuroblastoma
Authors: Gimenez Thamiris Magalhaes , Neves Nathalia Halley , Santos Andreia Rangel , Marchi Fabio A. , Kulikowski Leslie , Cristofani Lilian M. , Novak Estela M. , Filho Vicente Odone 
Published: May 2019

Due to heterogeneity of the neuroblastoma (NB) tumor, next-generation personalized medicine may identify therapeutic targets. Gene expression alterations of BLM (Bloom Syndrome RecQ Like Helicase) and FOX (Forkhead-box) family have been associated with an increased risk for cancer and resistance to chemotherapy drugs. Considering the potential implication of these genes on the clinical management of NB, we correlated clinical parameters, such as survival and treatments with genetic profiles of 11 NB patients stage 3 and 4 with MYCN-amplified (4 patients) or MYCN-non-amplified (7 patients) and one patient stage 2B and MYCN-non-amplified. Patients were treated with doxorubicin, etoposide, topotecan, cyclophosphamide, vincristine, and carboplatin. Targeted DNA sequencing, methylation array, and mRNA expression were performed on tumor and blood peripheral of these patients. Multiplex ligation-dependent probe amplification (MLPA). All variants considered important were validated by Sanger sequencing. Polymorphisms were identified using dbSNP138, 1000 genomes Project Consortium and NHLBI Exome Sequencing Project (ESP) databases with minor allele frequencies recorded from each database.

In this study, we observed in a patient with 17 months, diagnosed with MYCN-non amplified (11q-,+17p,+9p) stage 3 NB, a variant in FOXO3 gene (rs201947198) and BLM gene (rs28384991) with low frequency in the population (0.001% and 1%, respectively). FOXO3 and BLM gene expressions were decreased while FOXM1 gene expression was increased. This patient received chemotherapy and 2 years after the end of therapy, NB recurred locally. Six years after the diagnosis, the patient is still alive and receives chemotherapy.

In other case, we found a patient of 13 months, diagnosed with MYCN amplified (+2p,+3p,+17q) stage 3 NB, three synonymous variants in BLM gene (rs2227933,rs2227934,rs1063147). These variants have been shown to affect protein levels and function. We also identified an important variant in FOXO3 gene (rs 111556510) with low frequency in the population. FOXM1 mutation was not found in this patient and FOXM1 RNAm was increased. This patient received chemotherapy and despite the treatment, died 6 months after the diagnosis.

As relapse with a second event, we found two variants in BLM gene (rs2227935 and rs7167216), in a patient diagnosed at 6 months, stage 2B, MYCN-non-amplified (+2p;+3p;-11q;+17) NB, who received chemotherapy but has recurred three times at the same local site, despite the treatment. These variants are polymorphism low frequency and databases of the prediction algorithms, considered the SNP p.P868L damage. The variant identified in FOXR1 gene (c.C517G:p.R173G). This variant has not been described to NB. Further, hypermethylation in the promoter region of FOXK2 was correlated with decreased FOXK2 RNAm. Many studies have shown that epigenetic silencing of FOXK2 gene increases resistance to chemotherapy drugs. Other patients (3 MYCN-amplified and 6 MYCN-non-amplified) were found only polymorphisms synonymous in the BLM gene (rs2227933, rs2227934, and rs1063147). Moreover, variants or alterations in FOXO3, FOXR1, FOXK2 expression were not found. However, FOXM1 expression was increased in these patients.

Conclusion: Many studies have shown that personalized oncology plays an important role in treatment of cancer patients. In this study, we demonstrated that variants in FOXO3, FOXR1 and BLM genes, FOXM1 expression and silencing epigenetic of FOXK2 found in 3/12 patients with refractory relapse disease may be candidate targets for personalized therapy in neuroblastoma.

Citation Format: Thamiris Magalhaes Gimenez, Nathalia Halley Neves, Andreia Rangel Santos, Fabio A. Marchi, Leslie Kulikowski, Lilian M. Cristofani, Estela M. Novak, Vicente Odone Filho. BLM, FOXO3, FOXK2, FOXM1, FOXR1 genes as therapeutic targets to neuroblastoma [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr B13.