*3.9. Molecular Dynamics Study*

The stability of the interactions between ligand **11** and the proteins was assessed using Yasara Dynamic, following the previously described methodology [29]. In summary, the optimal binding between ligand **11** and the proteins was selected for further analysis, incorporating the AMBER14 force field with standard parameters. The simulation was conducted at a temperature of 298 K and a pH of 7.4 for 15 ns. Two RMSD parameters were employed: RMSD ligand conformation and RMSD ligand movement. The evaluation utilized the default scripts: md\_run.mcr and md\_analyse.mcr. The RMSD plots were generated using the Rstudio program (https://posit.co/products/open-source/rstudio/, accessed on 17 March 2023).

#### **4. Conclusions**

Here, 4- -pyridinecarbonyl-substituted renieramycin-type derivatives were prepared from compound **4** using light-induced intramolecular photoredox reactions and Steglich esterification. The optimal conditions for the photoredox reaction were achieved by employing 4 W blue light and dichloromethane. The natural renieramycins (**4**–**6**) were successfully transformed into their corresponding derivatives (**7** and **8**, which contained a 1,3-dioxolebridged phenolic moiety on ring A) in excellent–moderate yields (yields: 81%–37%). The attachment of the 4- -pyridinecarbonyl ester at the C-5 and C-22 positions was accomplished smoothly. Cytotoxicity study revealed that compounds **4** and **9** exhibited the most potent cytotoxicity against the human H292 and H460 NSCLC cell lines among the natural renieramycins and semisynthesized renieramycin-type derivatives, respectively. Interestingly, compound **11**, which contained both the 4- -pyridinecarbonyl ester at C-5 and the 1,3-dioxole-bridged phenolic motifs on ring A, showed better cytotoxicity than its parent compound (**7**). In addition, the renieramycin-type derivatives with a hydroxyl group at C-5 and C-22 lost their cytotoxicity against the NSCLC cells. Thus, the conformation of ring A featuring the 4- -pyridinecarbonyl ester at C-5 is crucial for the cytotoxic activity of renieramycins. Due to the limited quantity of the compound, a virtual network pharmacology study was conducted to simulate the target genes and molecular pathways. Molecular docking and dynamics studies predicted that the target proteins of compound **11** were MAPK1 and MAPK3. In future studies, the renieramycin-type derivatives will be further explored for their potential as anti-lung cancer agents.

**Supplementary Materials:** The following supporting information can be downloaded at: https:// www.mdpi.com/article/10.3390/md21070400/s1, Figures S1–S15: 1H NMR, 13C NMR and 2D NMR spectra of compounds **7**–**9** and **11**–**12**, Figures S16 and S17: Molecular docking of Ravoxertinib and the protonated **10** and **11** with both MAPK1 (ERK2) and MAPK3 (ERK1), Tables S1 and S2: Theoretical level of minimization of the 3D structures of **10** and **11**.

**Author Contributions:** Conceptualization, S.C.; methodology, S.S., K.B., K.P., B.I., K.S., M.Y., N.S., P.C. and S.C.; software, V.P., I.I. and S.C.; validation, K.S., M.Y., N.S. and S.C.; formal analysis, S.S., K.B., I.I., K.P., B.I., C.C. and S.C.; investigation, S.S., K.B., I.I., K.P., B.I. and S.C.; resources, P.C. and S.C.; data curation, S.C.; writing—original draft preparation, S.S., I.I. and K.B.; writing—review and editing, S.C.; visualization, S.C.; supervision, S.C.; project administration, S.C.; funding acquisition, S.C. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was funded by the Thai Research Fund, Thailand (RSA 6280009) to S.C.

**Data Availability Statement:** The data described and analyzed in this study are available from the corresponding author upon reasonable request.

**Acknowledgments:** This research is supported by the 90th anniversary of Chulalongkorn university fund No. GCUGR1125661018M (Ratchadaphiseksomphot Endowment Fund) for S.S. and the Second Century Fund (C2F), Chulalongkorn University for postdoctoral research fellowship to K.B. We gratefully acknowledge the Natural Products and Nanoparticles Research Unit (NP2) and the Department of Pharmacognosy and Pharmaceutical Botany and the Department of Pharmacology and Physiology, Faculty of Pharmaceutical Sciences, Chulalongkorn University for their scientific support.

**Conflicts of Interest:** The authors declare no conflict of interest.

#### **References**


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