TY - JOUR
T1 - A Covalent Calmodulin Inhibitor as a Tool to Study Cellular Mechanisms of K-Ras-Driven Stemness
AU - Okutachi, Sunday
AU - Manoharan, Ganesh Babu
AU - Kiriazis, Alexandros
AU - Laurini, Christina
AU - Catillon, Marie
AU - McCormick, Frank
AU - Yli-Kauhaluoma, Jari
AU - Abankwa, Daniel
N1 - Funding Information:
the Academy of Finland (Key Project # 304638) and core funding support by the University of Luxembourg.
Funding Information:
We are thankful to Dominic Esposito and Vanessa Wall (Frederick National Laboratory for Cancer Research, Frederick, MD, United States) for providing the entry clones and destination vectors for multisite gateway cloning. Nina Sipari from the Viikki Metabolomics Unit (Helsinki Institute of Life Science, University of Helsinki; Biocenter Finland) is thanked for her expertise with the LC-MS analyses. Funding. This work was supported by the Finnish Funding Agency for Innovation, Tekes, 3iRegeneration, project 40395/13) and Jane and Aatos Erkko Foundation to JY-K. DA received funding from the Academy of Finland (Key Project # 304638) and core funding support by the University of Luxembourg.
Funding Information:
This work was supported by the Finnish Funding Agency for Innovation, Tekes, 3iRegeneration, project 40395/13) and Jane and Aatos Erkko Foundation to JY-K. DA received funding from
Publisher Copyright:
© Copyright © 2021 Okutachi, Manoharan, Kiriazis, Laurini, Catillon, McCormick, Yli-Kauhaluoma and Abankwa.
PY - 2021/7/8
Y1 - 2021/7/8
N2 - Recently, the highly mutated oncoprotein K-Ras4B (hereafter K-Ras) was shown to drive cancer cell stemness in conjunction with calmodulin (CaM). We previously showed that the covalent CaM inhibitor ophiobolin A (OphA) can potently inhibit K-Ras stemness activity. However, OphA, a fungus-derived natural product, exhibits an unspecific, broad toxicity across all phyla. Here we identified a less toxic, functional analog of OphA that can efficiently inactivate CaM by covalent inhibition. We analyzed a small series of benzazulenones, which bear some structural similarity to OphA and can be synthesized in only six steps. We identified the formyl aminobenzazulenone 1, here named Calmirasone1, as a novel and potent covalent CaM inhibitor. Calmirasone1 has a 4-fold increased affinity for CaM as compared to OphA and was active against K-Ras in cells within minutes, as compared to hours required by OphA. Calmirasone1 displayed a 2.5–4.5-fold higher selectivity for KRAS over BRAF mutant 3D spheroid growth than OphA, suggesting improved relative on-target activity. Importantly, Calmirasone1 has a 40–260-fold lower unspecific toxic effect on HRAS mutant cells, while it reaches almost 50% of the activity of novel K-RasG12C specific inhibitors in 3D spheroid assays. Our results suggest that Calmirasone1 can serve as a new tool compound to further investigate the cancer cell biology of the K-Ras and CaM associated stemness activities.
AB - Recently, the highly mutated oncoprotein K-Ras4B (hereafter K-Ras) was shown to drive cancer cell stemness in conjunction with calmodulin (CaM). We previously showed that the covalent CaM inhibitor ophiobolin A (OphA) can potently inhibit K-Ras stemness activity. However, OphA, a fungus-derived natural product, exhibits an unspecific, broad toxicity across all phyla. Here we identified a less toxic, functional analog of OphA that can efficiently inactivate CaM by covalent inhibition. We analyzed a small series of benzazulenones, which bear some structural similarity to OphA and can be synthesized in only six steps. We identified the formyl aminobenzazulenone 1, here named Calmirasone1, as a novel and potent covalent CaM inhibitor. Calmirasone1 has a 4-fold increased affinity for CaM as compared to OphA and was active against K-Ras in cells within minutes, as compared to hours required by OphA. Calmirasone1 displayed a 2.5–4.5-fold higher selectivity for KRAS over BRAF mutant 3D spheroid growth than OphA, suggesting improved relative on-target activity. Importantly, Calmirasone1 has a 40–260-fold lower unspecific toxic effect on HRAS mutant cells, while it reaches almost 50% of the activity of novel K-RasG12C specific inhibitors in 3D spheroid assays. Our results suggest that Calmirasone1 can serve as a new tool compound to further investigate the cancer cell biology of the K-Ras and CaM associated stemness activities.
KW - BRET
KW - calmodulin
KW - cancer stem cell (CSC)
KW - covalent inhibitor
KW - K-Ras
UR - http://www.scopus.com/inward/record.url?scp=85111017775&partnerID=8YFLogxK
U2 - 10.3389/fcell.2021.665673
DO - 10.3389/fcell.2021.665673
M3 - Article
AN - SCOPUS:85111017775
SN - 2296-634X
VL - 9
JO - Frontiers in Cell and Developmental Biology
JF - Frontiers in Cell and Developmental Biology
M1 - 665673
ER -