A combination of inhibitors of glycolysis, glutaminolysis and de novo fatty acid synthesis decrease the expression of
Alejandro Schcolnik-Cabrera1, Guadalupe Dominguez-Gómez1, Alma Chávez-Blanco1
1Basic Research Division, National Cancer Institute, Mexico City 14080, Mexico.
Abstract:
Lonidamine, 6-Diazo-5-oxo-L-norleucine (DON) and orlistat are well known inhibitors of glycolysis, glutaminolysis and of de novo fatty acid synthesis, respectively. Although their antitumor effects have been explored in detail, the potential inhibition of the malignant metabolic phenotype and its influence on the expression of chemokines and growth factors involved in colon cancer, have not been previously reported to the best of our knowledge. In the present study, dose-response curves with orlistat, lonidamine or DON were generated from cell viability assays conducted in SW480 colon cancer cells. In addition, the synergistic effect of these compounds was evaluated in SW480 human colon cancer cells. The determination of the doses used for maximum synergistic efficacy led to the exploration of the mRNA levels of the target genes hexokinase-2 (HK2), glutaminase-1 (GLS-1) and fatty acid synthase (FASN) in human SW480 and murine CT26.WT colon cancer cells. The cell viability was evaluated following transfection with small interfering (si)RNA targeting these genes and was assessed with trypan blue. The expression levels of chemokines and growth factors were quantified in the supernatant of SW480 cells with LEGENDplex™. The combination of lonidamine, DON and orlistat resulted in a synergistic cytotoxic effect and induced the transcription of the corresponding gene targets but their corresponding proteins were actually downregulated. The downregulation of the expression levels of HK2, GLS-1 and FASN following transfection of the cells with the corresponding siRNA sequences decreased their viability. The treatment significantly reduced the expression levels of 9 chemokines [interleukin-9, C-X-C motif chemokine ligand (CXCL) 10, eotaxin, chemokine ligand (CCL) 9, CXCL5, CCL20, CXCL1, CXCL11 and CCCL4] and one growth factor (stem cell factor). These changes were associated with decreased phosphorylated-nuclear factor κB-p65. The data demonstrate that lonidamine, DON and orlistat in combination reduce the expression levels of chemokines and growth factors in colon cancer cells. Additional research is required to investigate the exact way by which both tumor and stromal cells regulate the expression levels of chemokines and growth factors.
Insights
Lonidamine, 6-Diazo-5-oxo-L-norleucine (DON), and orlistat synergistically reduce colon cancer cell viability. This combination therapy downregulates key metabolic genes and significantly decreases the expression of crucial chemokines and growth factors.
Area of Science:
- Oncology
- Cancer Metabolism
- Molecular Biology
Background:
- Lonidamine, DON, and orlistat are known inhibitors of key metabolic pathways: glycolysis, glutaminolysis, and de novo fatty acid synthesis.
- While their individual antitumor effects are studied, their combined impact on the malignant metabolic phenotype and associated signaling in colon cancer remains largely unexplored.
Purpose of the Study:
- To investigate the synergistic cytotoxic effects of lonidamine, DON, and orlistat in colon cancer cells.
- To determine the influence of this combination on the expression of genes involved in crucial metabolic pathways (HK2, GLS-1, FASN).
- To assess the impact on the secretion of chemokines and growth factors and their underlying regulatory mechanisms.
Main Methods:
- Dose-response and synergy assays were performed on SW480 colon cancer cells.
- mRNA levels of HK2, GLS-1, and FASN were analyzed in SW480 and CT26.WT cells.
- Cell viability was assessed after siRNA-mediated gene silencing of HK2, GLS-1, and FASN.
- Chemokine and growth factor levels in cell supernatants were quantified using LEGENDplex™.
Main Results:
- The combination of lonidamine, DON, and orlistat exhibited synergistic cytotoxicity against colon cancer cells.
- While gene transcription of HK2, GLS-1, and FASN was induced, their protein levels were downregulated.
- Silencing of HK2, GLS-1, and FASN using siRNA further reduced cell viability.
- The treatment significantly downregulated 9 chemokines and 1 growth factor, associated with decreased p-NF-κB-p65.
Conclusions:
- Combined lonidamine, DON, and orlistat treatment demonstrates a synergistic antitumor effect in colon cancer.
- This combination effectively modulates key metabolic pathways and reduces the expression of critical chemokines and growth factors.
- The findings highlight a novel therapeutic strategy targeting metabolic reprogramming and associated signaling in colon cancer.
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