Genotoxicity Testing of Nucleside Analogues

Abstract

Remdesivir and molnupiravir are two antiviral nucleoside analogues that were approved for emergency usage to treat SARS-COV-2 during the COVID-19 pandemic. However, concerns were raised surrounding he genotoxicity of these drugs. Genotoxicity refers to the ability of a compound to cause damage to the genomic DNA of cells. Genotoxicity often goes hand in hand with mutagenesis because damage to the DNA can result in mutations caused by damage such as deletions or mismatched bases as a result of incorrect or lack of repair. As such,
there is concern that these antiviral nucleoside analogues are genotoxic and could therefore lead to mutations. Gemcitabine is a cancer therapeutic that functions by causing genotoxicity in cancer cells, leading to cell death. Gemcitabine was originally developed as an antiviral, but was repurposed as a cancer drug due to its genotoxicity. It has been established that gemcitabine is integrated into genomic DNA, and is removed by MRE11. This suggests other antiviral nucleoside analogues may also be integrated into the genomic DNA, leading to genotoxicity, and that MRE11 may be able to remove them. The role of the meiotic recombination 11 (MRE11) protein in rescuing damage caused by nucleoside analogues was assessed. MRE11 appeared to rescue damage caused by nucleoside analogues in the DT40 MRE11+/- cell line. This suggests MRE11 removes nucleoside analogues from the DNA and is therefore important for maintaining genomic stability when nucleoside analogues are administered to patients. Using mass spectrometry we tested if the nucleoside analogues gemcitabine, GS-441524 and EIDD-1931 were integrated into the genomic DNA. EIDD-1931 and gemcitabine were identified as being incorporated into the genomic DNA of TK6 cells. Presence of nucleoside analogues in the genomic provided basis for further investigation of genotoxicity. To establish any genotoxic effects of the nucleoside analogues gemcitabine, GS-441524 and EIDD-1931, INDUCE-seq was used to identify the amount and locations of recurrent double strand break sites in the DNA. Gemcitabine appeared to increase the number of recurrent double strand breaks (DSBs) in TK6 cells, suggesting gemcitabine causes DSBs as a genotoxic effect.

Date of Award	27 Sept 2024
Original language	English
Awarding Institution	Bangor University
Sponsors	The Knowledge Economy Skills Partnership 2 (KESS 2) programme, Bangor University, ESF, and Tenovus Cancer Care
Supervisor	Edgar Hartsuiker (Supervisor)