EHA Library - The official digital education library of European Hematology Association (EHA)

ALPHA-KETOGLUTURATE EXPOSES METABOLIC VULNERABILITIES IN B-CELL LYMPHOMAS
Author(s): ,
Daifeng Jiang
Affiliations:
Hematology/Oncology - Medicine,University of Texas Health Science Center,San Antonio,United States
,
An-Ping Lin
Affiliations:
Hematology/Oncology - Medicine,University of Texas Health Science Center,San Antonio,United States
,
Long Wang
Affiliations:
Hematology/Oncology - Medicine,University of Texas Health Science Center,San Antonio,United States
Ricardo Aguiar
Affiliations:
Hematology/Oncology - Medicine,University of Texas Health Science Center,San Antonio,United States
(Abstract release date: 05/18/17) EHA Library. Aguiar R. 06/23/17; 181413; S126
Ricardo Aguiar
Ricardo Aguiar
Contributions
Abstract

Abstract: S126

Type: Oral Presentation

Presentation during EHA22: On Friday, June 23, 2017 from 12:15 - 12:30

Location: Room N101

Background
Metabolic rewiring is a cancer hallmark. These metabolic changes can be secondary to a broader oncogenic-driven deregulation (e.g., MYC), or they may be more specific and result from mutations in enzymes (e.g., IDH1 and IDH2) that directly control energy flux in the cell. Mutant IDH acquires a neomorphic activity and aberrantly generates high levels of D2-hydroxyglutarate (D2-HG), a natural metabolite with marked structural similarity to alpha-ketoglutarate (α-KG). D2-HG functions as an oncometabolite by competitively inhibiting the activity of multiple α-KG-dependent dioxygenases, including TET DNA hydroxylases and JmJC HDMs. The role of a D2-HG/α-KG metabolic imbalance in cancer was expanded by our recent discovery of somatic loss-of-function mutations in D2HGDH in diffuse large B cell lymphomas (DLBCL) (Nat Commun. 2015 Jul 16; 6:7768). D2HGDH catalyzes the conversion of D2-HG into α-KG, and D2HGDH -mutant DLBCLs display deficiency of α-KG with attending impaired dioxygenase function. Together, these data suggest that while D2-HG is an oncometabolite, α-KG may function as tumor suppressive metabolite.

Aims
To explore the concept that α-KG has tumor suppressor activities, and to characterize the signaling nodes that mediate the anti-lymphoma activity of this intermediated metabolite.

Methods
We utilized a panel of 14 well-characterized DLBCL cell lines to test the growth suppressive activity of cell-permeable synthetic α-KG derivative, dimethyl-α-KG (DM-KG), using cell proliferation and apoptosis assays. The cell line data was expanded to primary mature B cell tumors as well as normal B cells. These in vitro assays were complemented by xenografts models of human DLBCL. Functional studies were performed in cell lines and primary tumors, and included the enzymatic quantification of ATP synthase activity, the effects of α-KG on cellular ATP levels, the measurement of AMPK activity and of the mTORC1 kinase.

Results
The cell-permeable DM-KG induced a marked dose-dependent growth suppression in a panel of DLBCL cell lines that encompasses the molecular heterogeneity of this disease (ABC, n= 5, GCB, n=5 and OxPhos, n=4). In most instances, the growth inhibition exceeded 80% of vehicle control exposed cells and it could be detected as early as 24h and reached its peak at 72-96h post-exposure (Figure 1A). In all cell lines examined, induction of apoptosis accounted for most of the anti-lymphoma effects of DM-KG. There was no clear segregation between the DLBLC molecular subtype and DM-KG-induced growth inhibition (e.g., the ABC cell lines Ly10 and Ly3 were the most sensitive and most resistant, respectively, Figure 1A). Remarkably, we found that normal mature B cells (murine and human) were resistant to growth inhibition and apoptosis induced by DM-KG (Figure 1C). Contrary to that, exposing viable primary CLL, FL and DLBCL cells to DM-KG significantly induced apoptosis (p<0.01) in all tumors examined (n=17). In xenograft models of GCB- or ABC-DLBCLs (Ly7 and Ly10, respectively), DM-KG dosed intra-peritoneally significantly inhibited tumor growth in comparison to vehicle treated mice (p<0.01, n=16) (Figure 1B). To determine how DM-KG may induce growth suppression/apoptosis in mature B cell tumors, we first showed that in DLBCL cell lines α-KG inhibited the activity of ATP synthase, a key enzyme in the mitochondrial electron transport chain that generates most of the cellular ATP. Accordingly, short exposure (8h) to DM-KG suppressed ATP levels in all 14 DLBCL cell lines examined (mean = 21%, range 3% to 50%). Next, we examined how the fuel stress generated by the α-KG-mediated ATP synthase inhibition influenced energy-saving cellular signals. We found that in cell lines and primary tumors, DM-KG consistently activated the kinase AMPK, with consequent marked inhibition of mTORC1. Importantly, these signals were also engaged in normal B cells, but they did not result in growth inhibition or apoptosis, thus highlighting the unique sensitivity of cancer cells to the modulation of energy metabolism.

Conclusion
We showed that α-KG induces growth suppression and apoptosis in mature B cell tumors, in vitro and in vivo. We demonstrated that proximally α-KG exerts its tumor suppressive effects by inhibiting ATP synthase activity and ATP generation. This energy stress activates AMPK and suppresses mTORC1 resulting in growth inhibition and apoptosis in malignant cells, but not in their normal counterparts. These data highlight a metabolic cancer dependency and vulnerability that can be exploited therapeutically.

Session topic: 18. Non-Hodgkin & Hodgkin lymphoma - Biology

Keyword(s): Targeted therapy, mTOR, Mitochondria, DLBCL

Abstract: S126

Type: Oral Presentation

Presentation during EHA22: On Friday, June 23, 2017 from 12:15 - 12:30

Location: Room N101

Background
Metabolic rewiring is a cancer hallmark. These metabolic changes can be secondary to a broader oncogenic-driven deregulation (e.g., MYC), or they may be more specific and result from mutations in enzymes (e.g., IDH1 and IDH2) that directly control energy flux in the cell. Mutant IDH acquires a neomorphic activity and aberrantly generates high levels of D2-hydroxyglutarate (D2-HG), a natural metabolite with marked structural similarity to alpha-ketoglutarate (α-KG). D2-HG functions as an oncometabolite by competitively inhibiting the activity of multiple α-KG-dependent dioxygenases, including TET DNA hydroxylases and JmJC HDMs. The role of a D2-HG/α-KG metabolic imbalance in cancer was expanded by our recent discovery of somatic loss-of-function mutations in D2HGDH in diffuse large B cell lymphomas (DLBCL) (Nat Commun. 2015 Jul 16; 6:7768). D2HGDH catalyzes the conversion of D2-HG into α-KG, and D2HGDH -mutant DLBCLs display deficiency of α-KG with attending impaired dioxygenase function. Together, these data suggest that while D2-HG is an oncometabolite, α-KG may function as tumor suppressive metabolite.

Aims
To explore the concept that α-KG has tumor suppressor activities, and to characterize the signaling nodes that mediate the anti-lymphoma activity of this intermediated metabolite.

Methods
We utilized a panel of 14 well-characterized DLBCL cell lines to test the growth suppressive activity of cell-permeable synthetic α-KG derivative, dimethyl-α-KG (DM-KG), using cell proliferation and apoptosis assays. The cell line data was expanded to primary mature B cell tumors as well as normal B cells. These in vitro assays were complemented by xenografts models of human DLBCL. Functional studies were performed in cell lines and primary tumors, and included the enzymatic quantification of ATP synthase activity, the effects of α-KG on cellular ATP levels, the measurement of AMPK activity and of the mTORC1 kinase.

Results
The cell-permeable DM-KG induced a marked dose-dependent growth suppression in a panel of DLBCL cell lines that encompasses the molecular heterogeneity of this disease (ABC, n= 5, GCB, n=5 and OxPhos, n=4). In most instances, the growth inhibition exceeded 80% of vehicle control exposed cells and it could be detected as early as 24h and reached its peak at 72-96h post-exposure (Figure 1A). In all cell lines examined, induction of apoptosis accounted for most of the anti-lymphoma effects of DM-KG. There was no clear segregation between the DLBLC molecular subtype and DM-KG-induced growth inhibition (e.g., the ABC cell lines Ly10 and Ly3 were the most sensitive and most resistant, respectively, Figure 1A). Remarkably, we found that normal mature B cells (murine and human) were resistant to growth inhibition and apoptosis induced by DM-KG (Figure 1C). Contrary to that, exposing viable primary CLL, FL and DLBCL cells to DM-KG significantly induced apoptosis (p<0.01) in all tumors examined (n=17). In xenograft models of GCB- or ABC-DLBCLs (Ly7 and Ly10, respectively), DM-KG dosed intra-peritoneally significantly inhibited tumor growth in comparison to vehicle treated mice (p<0.01, n=16) (Figure 1B). To determine how DM-KG may induce growth suppression/apoptosis in mature B cell tumors, we first showed that in DLBCL cell lines α-KG inhibited the activity of ATP synthase, a key enzyme in the mitochondrial electron transport chain that generates most of the cellular ATP. Accordingly, short exposure (8h) to DM-KG suppressed ATP levels in all 14 DLBCL cell lines examined (mean = 21%, range 3% to 50%). Next, we examined how the fuel stress generated by the α-KG-mediated ATP synthase inhibition influenced energy-saving cellular signals. We found that in cell lines and primary tumors, DM-KG consistently activated the kinase AMPK, with consequent marked inhibition of mTORC1. Importantly, these signals were also engaged in normal B cells, but they did not result in growth inhibition or apoptosis, thus highlighting the unique sensitivity of cancer cells to the modulation of energy metabolism.

Conclusion
We showed that α-KG induces growth suppression and apoptosis in mature B cell tumors, in vitro and in vivo. We demonstrated that proximally α-KG exerts its tumor suppressive effects by inhibiting ATP synthase activity and ATP generation. This energy stress activates AMPK and suppresses mTORC1 resulting in growth inhibition and apoptosis in malignant cells, but not in their normal counterparts. These data highlight a metabolic cancer dependency and vulnerability that can be exploited therapeutically.

Session topic: 18. Non-Hodgkin & Hodgkin lymphoma - Biology

Keyword(s): Targeted therapy, mTOR, Mitochondria, DLBCL

By clicking “Accept Terms & all Cookies” or by continuing to browse, you agree to the storing of third-party cookies on your device to enhance your user experience and agree to the user terms and conditions of this learning management system (LMS).

Cookie Settings
Accept Terms & all Cookies