# The Impact of the S-Adenosylmethionine Analogue Sinefungin on Viral Life Cycles
## 4 Discussion  
Epigenetic-mechanism-targeting drugs represent promising new therapeutic strategies for the treatment of a broad spectrum of diseases, including malignancies, developmental disorders, and both acute and chronic viral infections [23,24]. Recent evidence has highlighted the critical role of methylation as a key epigenetic modification in viral genomes, with significant implications for the regulation of viral pathogenesis [25,26]. In both RNA and DNA viruses, methylation events, whether they occur on viral genomes or are mediated by the host's epigenetic machinery, modulate critical aspects of the viral life cycle. These include control of viral gene expression, latency, replication efficiency, and the ability to evade the host's immune surveillance [27,28]. Furthermore, methylation can influence virus-host interactions by altering the host's transcriptional landscape or mimicking host epigenetic marks, thereby facilitating persistent or chronic infections. The important role of viral genome methylation in regulating the replication cycle and contributing to pathogenic mechanisms highlights the need to direct the development of new therapeutic agents towards the enzymes and proteins that drive these epigenetic processes. Targeting these components could provide an innovative strategy for altering viral replication and modulating virus-host interactions, thus opening new avenues for antiviral therapy [29,30]. Sinfungin has emerged as a potential antiviral agent. This adenosine derivative, particularly the delta-(5'-adenosyl) derivative of ornithine, exhibits potent inhibition of SAM-dependent methyltransferases. By interfering with the activity of these enzymes, sinefungin offers a promising approach to modulating viral pathogenesis [31]. Although the antiviral effects of sinefungin are still limited, our study aimed to evaluate its activity against HSV-1 and SARS-CoV-2, representing enveloped and non-enveloped viral models with DNA and RNA genomes, respectively. Initially, the cytotoxic impact of sinefungin was evaluated on VERO-76 cells by exposing them to concentrations ranging from 12.5 to 200 \(\upmu\)g/mL for 24 h. The MTT results indicated that sinefungin reduced the cell viability by approximately 21.7% at the highest concentration tested, with CC\({}_{50}\) greater than 200 \(\upmu\)g/mL. In support of our findings, [PERSON] et al. reported that sinefungin did not significantly affect the cell viability of CRFK cells at a concentration of 100 \(\upmu\)g/mL [17]. In contrast, higher cytotoxicity was observed in the 3T3, HepGG2, and A549 cell lines. [PERSON] et al. reported CC\({}_{50}\) values of 38.7 \(\upmu\)g/mL, higher than 38 \(\upmu\)g/mL and 28 \(\upmu\)g/mL, respectively [32]. After evaluating the cytotoxicity used to establish a non-toxic concentration range, the antiviral activity of sinefungin was subsequently evaluated against HSV-1 and SARS-CoV-2. Our results demonstrated that sinefungin showed strong antiviral activity against HSV-1, reaching 81% inhibition at 200 \(\upmu\)g/mL and maintaining \(>\)50% inhibition up to 50 \(\upmu\)g/mL. In the case of SARS-CoV-2, sinefungin induced 70% and 54% inhibition at 200 and 100 \(\upmu\)g/mL, respectively, with a residual antiviral activity of 28% at 50 \(\upmu\)g/mL. The IC\({}_{50}\) values were 49.5 \(\pm\) 0.31 and 100.1 \(\pm\) 2.61 \(\upmu\)g/mL for HSV-1 and SARS-CoV-2, respectively. Considering the known inhibitory activity of sinefungin against SAM-dependent methyltransferases and the critical role of these enzymes in viral replication, we evaluated the effect of this compound on the viral yield in HSV-1- and SARS-CoV-2-infected VERO-76 cells at different times (6-30 h) and doses (25-200 \(\upmu\)g/mL). Sinfungin treatment at 10, 20, and 30 h post-infection resulted in dose- and time-dependent reductions in the viral yield. At 10 hpi, treatment with 200 \(\upmu\)g/mL induced a reduction in the viral titers of approximately 1.23 \(\log_{10}\), while 100 \(\upmu\)g/mL reduced the titers by \(\sim\)0.48 \(\log_{10}\).