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Oncometabolic mutation IDH1 R132H confers a metformin-hypersensitive phenotype.

Cuyàs E, Fernández-Arroyo S, Corominas-Faja B, Rodríguez-Gallego E, Bosch-Barrera J, Martin-Castillo B, De Llorens R, Joven J, Menendez JA - Oncotarget (2015)

Bottom Line: A mitochondria toxicity-screening modality confirmed a severe inability of IDH1-mutated cells to use various carbon substrates that are fed into the electron transport chain at different points.Additionally, metabolic reprogramming of IDH1-mutant cells increased their sensitivity to metformin in assays of cell proliferation, clonogenic potential, and mammosphere formation.Moreover, synergistic interactions occurred when metformin treatment was combined with the selective R132H-IDH1 inhibitor AGI-5198.

View Article: PubMed Central - PubMed

Affiliation: Metabolism and Cancer Group, Translational Research Laboratory, Catalan Institute of Oncology (ICO), Girona, Catalonia, Spain.

ABSTRACT
Metabolic flexibility might be particularly constrained in tumors bearing mutations in isocitrate dehydrogenase 1 (IDH1) leading to the production of the oncometabolite 2-hydroxygluratate (2HG). To test the hypothesis that IDH1 mutations could generate metabolic vulnerabilities for therapeutic intervention, we utilized an MCF10A cell line engineered with an arginine-to-histidine conversion at position 132 (R132H) in the catalytic site of IDH1, which equips the enzyme with a neomorphic α-ketoglutarate to 2HG reducing activity in an otherwise isogenic background. IDH1 R132H/+ and isogenic IDH1 +/+ parental cells were screened for their ability to generate energy-rich NADH when cultured in a standardized high-throughput Phenotype MicroArrayplatform comprising >300 nutrients. A radical remodeling of the metabotype occurred in cells carrying the R132H mutation since they presented a markedly altered ability to utilize numerous carbon catabolic fuels. A mitochondria toxicity-screening modality confirmed a severe inability of IDH1-mutated cells to use various carbon substrates that are fed into the electron transport chain at different points. The mitochondrial biguanide poisons, metformin and phenformin, further impaired the intrinsic weakness of IDH1-mutant cells to use certain carbon-energy sources. Additionally, metabolic reprogramming of IDH1-mutant cells increased their sensitivity to metformin in assays of cell proliferation, clonogenic potential, and mammosphere formation. Targeted metabolomics studies revealed that the ability of metformin to interfere with the anaplerotic entry of glutamine into the tricarboxylic acid cycle could explain the hypersensitivity of IDH1-mutant cells to biguanides. Moreover, synergistic interactions occurred when metformin treatment was combined with the selective R132H-IDH1 inhibitor AGI-5198. Together, these results suggest that therapy involving the simultaneous targeting of metabolic vulnerabilities with metformin, and 2HG overproduction with mutant-selective inhibitors (AGI-5198-related AG-120 [Agios]), might represent a worthwhile avenue of exploration in the treatment of IDH1-mutated tumors.

No MeSH data available.


Related in: MedlinePlus

Metabolic synthetic lethality between metformin and the oncometabolic mutation IDH1 R132HIDH1 mutant cells exhibit unique metabolic properties that specifically distinguish them from the IDH1-wild type counterparts. We substantiate experimentally the fact that the cancer-driving R132H mutation drastically increases cancer cells’ vulnerability to the biguanide metformin. The ability of metformin to impact on the dependency on glutamine anaplerosis of the TCA cycle is synthetically lethal in IDH1 mutant cells, which are defective in reductive glutamine metabolism. The exacerbated metabolic vulnerability of cells bearing the 2HG-overproducing R132H mutation to clinically available biguanides may represent a new avenue in the treatment of IDH1-mutated tumors, combining clinically available metabolic drugs targeting mitochondria (e.g., metformin) and the aberrant gain-of-function of IDH1 mutant protein (e.g., AG-120).
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Figure 6: Metabolic synthetic lethality between metformin and the oncometabolic mutation IDH1 R132HIDH1 mutant cells exhibit unique metabolic properties that specifically distinguish them from the IDH1-wild type counterparts. We substantiate experimentally the fact that the cancer-driving R132H mutation drastically increases cancer cells’ vulnerability to the biguanide metformin. The ability of metformin to impact on the dependency on glutamine anaplerosis of the TCA cycle is synthetically lethal in IDH1 mutant cells, which are defective in reductive glutamine metabolism. The exacerbated metabolic vulnerability of cells bearing the 2HG-overproducing R132H mutation to clinically available biguanides may represent a new avenue in the treatment of IDH1-mutated tumors, combining clinically available metabolic drugs targeting mitochondria (e.g., metformin) and the aberrant gain-of-function of IDH1 mutant protein (e.g., AG-120).

Mentions: Because glutaminase catalyzes the conversion of glutamine to glutamate, and the increased activity of this enzyme is at least partially responsible for elevated glutamine metabolism in cancer, including IDH1-mutated tumors [43, 45, 49, 60-64], the fact that BPTES-induced blockade of glutamine flux augmented the sensitivity of IDH1-mutated cells to metformin was consistent with the changes in glutamine metabolism as an adaptive response following metformin treatment. Accordingly, metformin-induced inhibition of the ETC has been reported to result in an adaptive increase in glutamine metabolism that involves a switch from oxidative to reductive pathways [58], viz. the conversion of glutamine to acetyl-CoA for biosynthetic processes, which ultimately leads to a dramatic decrease in ATP production by the TCA cycle. Given that mutations in IDH1 alter TCA metabolism by targeting a critical step in reductive glutamine metabolism to drastically reduce the ability of IDH1-mutant cells to fully induce this pathway, it is tempting to suggest that the ability of metformin to interfere with the anaplerotic entry of glutamine into the TCA cycle results in a synthetically lethal interaction in these cells (Figure 6). Of note, these cells are exquisitely sensitive to deficiencies in the production of glutamine-derived metabolites but are unable to mount an adaptive response to metformin because of impaired glutamine metabolism [50, 58, 59]. Moreover, an enhanced conversion of glutamine to glutamate drives the serine biosynthetic pathway (SBP), which has been shown to confer a growth advantage to tumor cells beyond providing serine and glycine for biosynthesis reactions. The SBP intersects glutaminolysis and provides not only an alternate pathway for αKG production for mitochondrial metabolism, but it also provides substrates for production of reduced glutathione (GSH), a key cellular antioxidant [76-79]. The significant and concurrent depletion of glutamate and serine provoked by metformin suggests that the metformin-induced metabolic switch might drastically and distinctively alter a response against oxidative stress. Zaprinast, a recently identified glutaminase inhibitor that is highly effective against glutamine-addicted and IDH1-mutant cells, induced significant reductions in cellular pools of glutamate, increased reactive oxygen species formation, and increased susceptibility to oxidative damage [43]. Future studies should determine whether the reported ability of metformin to inhibit glutaminase activity [80], can translate into an increased reliance on wild-type IDH1 TCA activity in IDH1-mutant cells to impair the glutamate/serine-driven maintenance of redox balance. Nonetheless, serine is also involved in folate-mediated one carbon metabolism by acting as a methyl group donor to convert tetrahydrofolate (THF) to methylene-THF, a key intermediate that fuels nucleotide metabolism and methylation reactions [81-83]. Given that metformin-impaired THF-driven one carbon metabolism [84] has been shown to strongly deplete nucleotide triphosphates and impede nucleotide synthesis in mammosphere-derived breast CSCs [85], it might be pertinent to evaluate whether metformin-induced starvation of serine underlies the strong ability of metformin to suppress the intrinsically enhanced self-renewal activity of mammosphere-initiating IDH1-mutant cells. Moreover, glutamine is a required substrate for three enzymes involved in the de novo synthesis of purine nucleotides and two enzymes involved in the de novo synthesis of pyrimidine nucleotides [86]. Therefore, glutamine-addicted IDH1-mutant cells will be expected to exhibit increased levels of nucleotides when compared to IDH1-WT cells. Preliminary experiments in our laboratory have likewise confirmed that a significant overproduction of pyrimidines takes place in R132H/+ cells, a phenomenon that was fully prevented in the presence of metformin (unpublished observations). Although we acknowledge that the current study does not address the efficacy of metformin against cancer cell models naturally bearing clinically relevant IDH1 mutations, our findings provide a direct mechanistic link to the association between metformin use and a significant reduction in incidence of intrahepatic cholangiocarcinoma (IHCC) in patients with diabetes [87]. Because IHCC is a deadly liver malignancy in which highly prevalent IDH1 mutations subvert the hepatocyte differentiation/quiescence program to create a persistent pre-neoplastic state [27], it might be tempting to suggest that chronic exposure to metformin might create a state of intolerable metabolic stress for stem-like pre-malignant cells, and thereby stopping IHCC before it starts.


Oncometabolic mutation IDH1 R132H confers a metformin-hypersensitive phenotype.

Cuyàs E, Fernández-Arroyo S, Corominas-Faja B, Rodríguez-Gallego E, Bosch-Barrera J, Martin-Castillo B, De Llorens R, Joven J, Menendez JA - Oncotarget (2015)

Metabolic synthetic lethality between metformin and the oncometabolic mutation IDH1 R132HIDH1 mutant cells exhibit unique metabolic properties that specifically distinguish them from the IDH1-wild type counterparts. We substantiate experimentally the fact that the cancer-driving R132H mutation drastically increases cancer cells’ vulnerability to the biguanide metformin. The ability of metformin to impact on the dependency on glutamine anaplerosis of the TCA cycle is synthetically lethal in IDH1 mutant cells, which are defective in reductive glutamine metabolism. The exacerbated metabolic vulnerability of cells bearing the 2HG-overproducing R132H mutation to clinically available biguanides may represent a new avenue in the treatment of IDH1-mutated tumors, combining clinically available metabolic drugs targeting mitochondria (e.g., metformin) and the aberrant gain-of-function of IDH1 mutant protein (e.g., AG-120).
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Related In: Results  -  Collection

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Figure 6: Metabolic synthetic lethality between metformin and the oncometabolic mutation IDH1 R132HIDH1 mutant cells exhibit unique metabolic properties that specifically distinguish them from the IDH1-wild type counterparts. We substantiate experimentally the fact that the cancer-driving R132H mutation drastically increases cancer cells’ vulnerability to the biguanide metformin. The ability of metformin to impact on the dependency on glutamine anaplerosis of the TCA cycle is synthetically lethal in IDH1 mutant cells, which are defective in reductive glutamine metabolism. The exacerbated metabolic vulnerability of cells bearing the 2HG-overproducing R132H mutation to clinically available biguanides may represent a new avenue in the treatment of IDH1-mutated tumors, combining clinically available metabolic drugs targeting mitochondria (e.g., metformin) and the aberrant gain-of-function of IDH1 mutant protein (e.g., AG-120).
Mentions: Because glutaminase catalyzes the conversion of glutamine to glutamate, and the increased activity of this enzyme is at least partially responsible for elevated glutamine metabolism in cancer, including IDH1-mutated tumors [43, 45, 49, 60-64], the fact that BPTES-induced blockade of glutamine flux augmented the sensitivity of IDH1-mutated cells to metformin was consistent with the changes in glutamine metabolism as an adaptive response following metformin treatment. Accordingly, metformin-induced inhibition of the ETC has been reported to result in an adaptive increase in glutamine metabolism that involves a switch from oxidative to reductive pathways [58], viz. the conversion of glutamine to acetyl-CoA for biosynthetic processes, which ultimately leads to a dramatic decrease in ATP production by the TCA cycle. Given that mutations in IDH1 alter TCA metabolism by targeting a critical step in reductive glutamine metabolism to drastically reduce the ability of IDH1-mutant cells to fully induce this pathway, it is tempting to suggest that the ability of metformin to interfere with the anaplerotic entry of glutamine into the TCA cycle results in a synthetically lethal interaction in these cells (Figure 6). Of note, these cells are exquisitely sensitive to deficiencies in the production of glutamine-derived metabolites but are unable to mount an adaptive response to metformin because of impaired glutamine metabolism [50, 58, 59]. Moreover, an enhanced conversion of glutamine to glutamate drives the serine biosynthetic pathway (SBP), which has been shown to confer a growth advantage to tumor cells beyond providing serine and glycine for biosynthesis reactions. The SBP intersects glutaminolysis and provides not only an alternate pathway for αKG production for mitochondrial metabolism, but it also provides substrates for production of reduced glutathione (GSH), a key cellular antioxidant [76-79]. The significant and concurrent depletion of glutamate and serine provoked by metformin suggests that the metformin-induced metabolic switch might drastically and distinctively alter a response against oxidative stress. Zaprinast, a recently identified glutaminase inhibitor that is highly effective against glutamine-addicted and IDH1-mutant cells, induced significant reductions in cellular pools of glutamate, increased reactive oxygen species formation, and increased susceptibility to oxidative damage [43]. Future studies should determine whether the reported ability of metformin to inhibit glutaminase activity [80], can translate into an increased reliance on wild-type IDH1 TCA activity in IDH1-mutant cells to impair the glutamate/serine-driven maintenance of redox balance. Nonetheless, serine is also involved in folate-mediated one carbon metabolism by acting as a methyl group donor to convert tetrahydrofolate (THF) to methylene-THF, a key intermediate that fuels nucleotide metabolism and methylation reactions [81-83]. Given that metformin-impaired THF-driven one carbon metabolism [84] has been shown to strongly deplete nucleotide triphosphates and impede nucleotide synthesis in mammosphere-derived breast CSCs [85], it might be pertinent to evaluate whether metformin-induced starvation of serine underlies the strong ability of metformin to suppress the intrinsically enhanced self-renewal activity of mammosphere-initiating IDH1-mutant cells. Moreover, glutamine is a required substrate for three enzymes involved in the de novo synthesis of purine nucleotides and two enzymes involved in the de novo synthesis of pyrimidine nucleotides [86]. Therefore, glutamine-addicted IDH1-mutant cells will be expected to exhibit increased levels of nucleotides when compared to IDH1-WT cells. Preliminary experiments in our laboratory have likewise confirmed that a significant overproduction of pyrimidines takes place in R132H/+ cells, a phenomenon that was fully prevented in the presence of metformin (unpublished observations). Although we acknowledge that the current study does not address the efficacy of metformin against cancer cell models naturally bearing clinically relevant IDH1 mutations, our findings provide a direct mechanistic link to the association between metformin use and a significant reduction in incidence of intrahepatic cholangiocarcinoma (IHCC) in patients with diabetes [87]. Because IHCC is a deadly liver malignancy in which highly prevalent IDH1 mutations subvert the hepatocyte differentiation/quiescence program to create a persistent pre-neoplastic state [27], it might be tempting to suggest that chronic exposure to metformin might create a state of intolerable metabolic stress for stem-like pre-malignant cells, and thereby stopping IHCC before it starts.

Bottom Line: A mitochondria toxicity-screening modality confirmed a severe inability of IDH1-mutated cells to use various carbon substrates that are fed into the electron transport chain at different points.Additionally, metabolic reprogramming of IDH1-mutant cells increased their sensitivity to metformin in assays of cell proliferation, clonogenic potential, and mammosphere formation.Moreover, synergistic interactions occurred when metformin treatment was combined with the selective R132H-IDH1 inhibitor AGI-5198.

View Article: PubMed Central - PubMed

Affiliation: Metabolism and Cancer Group, Translational Research Laboratory, Catalan Institute of Oncology (ICO), Girona, Catalonia, Spain.

ABSTRACT
Metabolic flexibility might be particularly constrained in tumors bearing mutations in isocitrate dehydrogenase 1 (IDH1) leading to the production of the oncometabolite 2-hydroxygluratate (2HG). To test the hypothesis that IDH1 mutations could generate metabolic vulnerabilities for therapeutic intervention, we utilized an MCF10A cell line engineered with an arginine-to-histidine conversion at position 132 (R132H) in the catalytic site of IDH1, which equips the enzyme with a neomorphic α-ketoglutarate to 2HG reducing activity in an otherwise isogenic background. IDH1 R132H/+ and isogenic IDH1 +/+ parental cells were screened for their ability to generate energy-rich NADH when cultured in a standardized high-throughput Phenotype MicroArrayplatform comprising >300 nutrients. A radical remodeling of the metabotype occurred in cells carrying the R132H mutation since they presented a markedly altered ability to utilize numerous carbon catabolic fuels. A mitochondria toxicity-screening modality confirmed a severe inability of IDH1-mutated cells to use various carbon substrates that are fed into the electron transport chain at different points. The mitochondrial biguanide poisons, metformin and phenformin, further impaired the intrinsic weakness of IDH1-mutant cells to use certain carbon-energy sources. Additionally, metabolic reprogramming of IDH1-mutant cells increased their sensitivity to metformin in assays of cell proliferation, clonogenic potential, and mammosphere formation. Targeted metabolomics studies revealed that the ability of metformin to interfere with the anaplerotic entry of glutamine into the tricarboxylic acid cycle could explain the hypersensitivity of IDH1-mutant cells to biguanides. Moreover, synergistic interactions occurred when metformin treatment was combined with the selective R132H-IDH1 inhibitor AGI-5198. Together, these results suggest that therapy involving the simultaneous targeting of metabolic vulnerabilities with metformin, and 2HG overproduction with mutant-selective inhibitors (AGI-5198-related AG-120 [Agios]), might represent a worthwhile avenue of exploration in the treatment of IDH1-mutated tumors.

No MeSH data available.


Related in: MedlinePlus