# Detection of XPO1E571K Gene Mutation from Cell-Free DNA in Blood Circulation of Lymphoma Patients by FAST-COLD PCR

## TL;DR

This paper introduces a new method to detect a specific gene mutation in lymphoma patients using DNA from blood, which could improve diagnosis and treatment.

## Contribution

The study develops an optimized FAST-COLD-PCR method for sensitive and specific detection of the XPO1E571K mutation in lymphoma patient cfDNA.

## Key findings

- FAST-COLD-PCR with a critical temperature of 73.3°C selectively amplifies XPO1E571K mutant DNA.
- The method enables high specificity and sensitivity for detecting XPO1E571K mutations in lymphoma patients.
- The approach allows rapid and straightforward detection of mutations from cell-free DNA in blood.

## Abstract

The XPO1 (exportin 1) gene encodes exportin 1 protein responsible for transporting proteins and RNA from the nucleus to the cytoplasm. It has been used as a biomarker for lymphoma detection. XPO1E571K mutation has been frequently observed and identified as a good prognostic indicator for lymphoma patients. The detection of a target molecule released by lymphoma cells into blood circulation (cell-free circulating tumor DNA, cfDNA) is a better method than tissue biopsy. However, cfDNA concentration in blood circulation is very low in cancer patients. Therefore, a precise and sensitive method is needed. In this study, cfDNA was extracted, and then the XPO1 gene was detected and amplified using conventional PCR. Sanger sequencing was employed to verify the DNA sequences. FAST-COLD-PCR was developed to detect XPO1E571K gene mutation using a CFX96 Touch Real-Time PCR System. The optimal critical temperature (Tc) was 73.3 °C, allowing selective amplification of XPO1E571K mutant DNA while wild-type XPO1 could not be amplified. XPO1E571K gene mutation can be detected by this method with high specificity and sensitivity in lymphoma patients. This approach facilitates rapid and straightforward detection in a timely manner after the diagnosis. Accordingly, the optimized FAST-COLD-PCR conditions can be used as a prototype for XPO1E571K mutant detection in lymphoma patients.