493P - Identifying genomic changes in breast tumours that metastasise to the brain

Background

The development of metastatic tumours after what appears to be a successful treatment of breast cancer remains a major problem. Many of these late-developing, secondary metastatic tumours occur specifically in the brain and metastatic brain tumours have very poor prognoses. Identifying genomic alterations in breast-to-brain metastatic tumours may provide new opportunities for treatment and prognosis.

Methods

Whole Exome Sequencing of 26 breasts to brain metastases was carried out on a high output V2 150 cycle flowcell (Illumina) as a 75 paired. Bioinformatic analysis was performed to identify recurrent mutations. Confirmation of mutated variants by Sanger sequencing. Functional analysis was conducted on a CRISPR_Cas9 knockout candidate gene in the MCF7 breast cancer cell line.

Results

Following analysis of Whole Exome Sequencing data, we identified > 13000 nonsynonymous variants. To eliminate potential germline polymorphisms, variants with a MAF ≤1% were maintained. All variants were screened for their consequence on the protein product (via SIFT and Polyphen2), and the variants with scores related to pathogenicity were retained. This screening generated a list of 286 variants found across the 26 tumours analyzed. Protein domain analysis showed that most mutated variants are found within conserved protein domains. This sequencing identified frequent mutations in a gene encoding a member of the ARFGEF family of proteins. Furthermore, Following CRISPR_Cas9 knockout of this ARFGEF protein, the expression of neurotransmitter subunits was observed. This suggests that this protein may be implicated in the regulation of neuronal signalling, presenting a rationale for brain colonization by breast to brain metastatic tumour cells.

Conclusions

We expect that the identification of these commonly mutated genes may lead to the development of prognostic biomarkers that could be used to predict the risk of brain metastases from breast tumours and provide essential molecular information relating to the existing metastatic tumour. Additionally, these genes can contribute to the development of novel therapeutic approaches.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

University of Wolverhampton.

Funding

Ministerio de Educación Ciencia y Tecnología (MESCyT).

Disclosure

All authors have declared no conflicts of interest.