TY - JOUR
T1 - Cytotoxicity of frutalin on distinct cancer cells is independent of its glycosylation
AU - Oliveira, Carla
AU - Freitas, Ana Isabel
AU - Campos, Nair
AU - Saraiva, Lucília
AU - Domingues, Lucília
N1 - Funding Information:
This research was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 and UIDB/50006/2020 (LAQV/REQUIMTE).
Publisher Copyright:
© 2021 by the authors. Licensee MDPI, Basel, Switzerland.
PY - 2021/8
Y1 - 2021/8
N2 - Frutalin is a plant lectin with beneficial immunobiological action, although the access to its active form is still restricted. Moreover, there is a knowledge gap on isoform activity and glycosylation impact on its bioactivity, and recombinant production protocols were seen as ineffective. Here, a simpler and faster production and purification protocol was developed, attaining a yield of purified frutalin 3.3-fold higher than that obtained previously. Hemagglutination assays confirmed that this frutalin isoform could not agglutinate rabbit erythrocytes, while maintaining the native tetrameric structure, as indicated by DLS analysis, and strong interaction with methyl-alpha-galactose, in fluorescence spectroscopy studies. The cytotoxicity of the recombinant frutalin isoform was shown in a broad panel of human cancer cells: colon (HCT116), melanoma (A375), triple-negative breast cancer (MDA-MB-231), and ovarian (IGROV-1). Treatment with 8.5–11.8 µM TrxFTL reduced proliferation of all cancer cells to half in 48 h. This anti-proliferative effect encompasses the p53 pathway since it was significantly reduced in p53-null colon cancer cells (HCT116 p53−/−; GI50 of 25.0 ± 3.0 µM), when compared to the isogenic p53-positive cells (HCT116 p53+/+; GI50 of 8.7 ± 1.8 µM; p < 0.002). This recombinantly produced frutalin isoform has relevant cytotoxic effect and its biological activity is not dependent on glycosylation. The developed E. coli production and purification protocol generates high yield of non-glycosylated frutalin isoform with potent cytotoxic activity, enabling the development of novel anticancer p53-targeting therapies.
AB - Frutalin is a plant lectin with beneficial immunobiological action, although the access to its active form is still restricted. Moreover, there is a knowledge gap on isoform activity and glycosylation impact on its bioactivity, and recombinant production protocols were seen as ineffective. Here, a simpler and faster production and purification protocol was developed, attaining a yield of purified frutalin 3.3-fold higher than that obtained previously. Hemagglutination assays confirmed that this frutalin isoform could not agglutinate rabbit erythrocytes, while maintaining the native tetrameric structure, as indicated by DLS analysis, and strong interaction with methyl-alpha-galactose, in fluorescence spectroscopy studies. The cytotoxicity of the recombinant frutalin isoform was shown in a broad panel of human cancer cells: colon (HCT116), melanoma (A375), triple-negative breast cancer (MDA-MB-231), and ovarian (IGROV-1). Treatment with 8.5–11.8 µM TrxFTL reduced proliferation of all cancer cells to half in 48 h. This anti-proliferative effect encompasses the p53 pathway since it was significantly reduced in p53-null colon cancer cells (HCT116 p53−/−; GI50 of 25.0 ± 3.0 µM), when compared to the isogenic p53-positive cells (HCT116 p53+/+; GI50 of 8.7 ± 1.8 µM; p < 0.002). This recombinantly produced frutalin isoform has relevant cytotoxic effect and its biological activity is not dependent on glycosylation. The developed E. coli production and purification protocol generates high yield of non-glycosylated frutalin isoform with potent cytotoxic activity, enabling the development of novel anticancer p53-targeting therapies.
KW - Escherichia coli
KW - Isoforms
KW - P53
KW - Recombinant frutalin
KW - Targeted anticancer therapy
UR - http://www.scopus.com/inward/record.url?scp=85112039156&partnerID=8YFLogxK
U2 - 10.3390/molecules26164712
DO - 10.3390/molecules26164712
M3 - Article
C2 - 34443300
AN - SCOPUS:85112039156
SN - 1420-3049
VL - 26
JO - Molecules
JF - Molecules
IS - 16
M1 - 4712
ER -