The targeted next-generation sequence revealed SMAD4, AKT1, and TP53 mutations from circulating cell-free DNA of breast cancer and its effect on protein structure – A computational approach

Abstract

Breast cancer biomarkers that detect marginally advanced stages are still challenging. The detection of specific abnormalities, targeted therapy selection, prognosis, and monitoring of treatment effectiveness over time are all made possible by circulating free DNA (cfDNA) analysis. The proposed study will detect specific genetic abnormalities from the plasma cfDNA of a female breast cancer patient by sequencing a cancer-related gene panel (MGM455 – Oncotrack Ultima), including 56 theranostic genes (SNVs and small INDELs). Initially, we determined the pathogenicity of the observed mutations using PredictSNP, iStable, Align-GVGD, and ConSurf servers. As a next step, molecular dynamics (MD) was implemented to determine the functional significance of SMAD4 mutation (V465M). Lastly, the mutant gene relationships were examined using the Cytoscape plug-in GeneMANIA. Using ClueGO, we determined the gene’s functional enrichment and integrative analysis. The structural characteristics of SMAD4 V465M protein by MD simulation analysis further demonstrated that the mutation was deleterious. The simulation showed that the native structure was more significantly altered by the SMAD4 (V465M) mutation. Our findings suggest that SMAD4 V465M mutation might be significantly associated with breast cancer, and other patient-found mutations (AKT1-E17K and TP53-R175H) are synergistically involved in the process of SMAD4 translocate to nuclease, which affects the target gene translation. Therefore, this combination of gene mutations could alter the TGF-β signaling pathway in BC. We further proposed that the SMAD4 protein loss may contribute to an aggressive phenotype by inhibiting the TGF-β signaling pathway. Thus, breast cancer’s SMAD4 (V465M) mutation might increase their invasive and metastatic capabilities.

Communicated by Ramaswamy H. Sarma

Keywords:

• cfDNA
• breast cancer
• TP53
• SMAD4
• AKT1
• mutation
• molecular dynamics simulations
• circulating-tumor DNA

Acknowledgments

The authors would like to take this opportunity to thank the management of Vellore Institute of Technology (VIT), Vellore, India, and Sri Ramachandra Institute of Higher Education and Research (SRIHER), Porur, Chennai, India, for providing the necessary facilities and encouragement to carry out this work.

Author’s contributions

AB, UKS and CGPD were involved in the study’s design. AB and UKS were involved in the data collection and experiment. AB, UKS, TKD, AAD, RG, SSK, MSSD and CGPD were involved in the results’ acquisition, analysis, and interpretation. AB, UKS and AAD drafted the manuscript. CGPD supervised the entire study and was involved in study design, the acquisition, analysis, and understanding of the data, and critically reviewed the manuscript. All authors edited and approved the submitted version of the article.

Disclosure statement

The authors have declared that no conflicts of interest exist.

Additional information

Funding

No funding agency was involved in the present study.