Understanding how carcinogenesis can expose cancers to synthetically lethal vulnerabilities has been an essential underpinning of development of modern anticancer therapeutics. Isocitrate dehydrogenase wild-type (IDHWT) glioblastoma multiforme (GBM), which is known to have upregulated branched-chain amino acid transaminase 1 (BCAT1) expression, has not had treatments developed to the same extent as the IDH mutant counterpart, despite making up the majority of cases. In this issue, Zhang and colleagues utilize a metabolic screen to identify α-ketoglutarate (AKG) as a synthetically lethal treatment in conjunction with BCAT1 inhibition in IDHWT GBM. These treatments synergize in a multipronged approach that limits substrate catabolism and disrupts mitochondrial homeostasis through perturbing the balance of NAD+/NADH, leading to mTORC1 inhibition and a reduction of nucleotide biosynthesis. Based on these results, the authors propose combination treatment targeting branched chain amino acid catabolism as a potential option for patients with IDHWT GBM.

See related article by Zhang et al., p. 2388

Targeted, personalized therapies serve as a landmark of modern cancer treatment. While early treatments utilized powerful anticancer drugs with broad, systemic side effects, modern treatments identify key vulnerabilities in an individual cancer that are highly effective with minimal off-target effects. One concept critical in the development of these therapies is synthetic lethality where mutations or other changes that occur during carcinogenesis can expose a vulnerability distinct from the healthy progenitors. This occurs when cancers become reliant on specific proteins and pathways to support distinct functional programs or compensate for losses. Uncovering these vulnerabilities is unfortunately challenging as they are often context dependent and emerge from the behavior of the biological system as a whole.

Glioblastoma multiforme (GBM), the most common and aggressive cancer in the brain, is a tragically potent disease with limited treatments. Current options include only the standards of surgery, radiotherapy, and chemotherapy, but prognosis for these treatments remains poor. While significant progress has been made in designing synthetically lethal treatments for isocitrate dehydrogenase (IDH) mutant gliomas, limited approaches have been identified for targeting their more common IDH wild-type (IDHWT) counterparts. Thus, finding an appropriate target has been a key goal of GBM research. Previous work has linked the metabolism of branched chain amino acids (BCAA) through branched-chain amino acid transaminase 1 (BCAT1) to proliferation in GBM, making BCAAs a promising target for investigation (1).

BCAAs are essential amino acids including leucine, isoleucine, and valine, which can not only be incorporated into proteins but also catabolized for energy or biosynthesis. The first step in BCAA catabolism is reversible transamination through either cytosolic BCAT1 or mitochondrial BCAT2, which produces branched chain α-ketoacids (BCKA), including α-ketoisocaproate (KIC), α-ketoisovalerate (KIV), and α-keto-β-methylvalerate (KMV), by donating the amino-group to α-ketoglutarate (AKG) to form glutamate. Following this step, BCKAs are oxidized irreversibly by the BCKA dehydrogenase (BCKDH) complex in the mitochondrion, eventually leading to the production of acetyl-CoA and succinyl-CoA, which can enter the tricarboxylic acid (TCA) cycle and produce 10, 6, and 5 NADH molecules per molecule of KIC, KIV, and KMV, respectively (2).

Notably, IDHWT GBMs depend on upregulated BCAA catabolism rather than the reductive metabolism featured in some other cancers. When reductive BCAA metabolism is high such as in chronic myeloid leukemia, BCKAs acquired from the tumor microenvironment accumulate as BCAAs in the cancer cells, activating mTORC1 to promote cancer proliferation (3). The BCAA oxidation of IDHWT GBM instead consumes BCAAs while converting AKG into glutamate, promoting cancer proliferation through hypoxia-inducible factor-1α and DNA hypermethylation (4, 5).

Zhang and colleagues identified oxidative BCAA metabolism as a targetable vulnerability for IDHWT GBM (6). Although, unsurprisingly, BCAT1 knockout (KO) alone was unable to inhibit GBM proliferation, the authors hypothesized that metabolites could synergize to strengthen the inhibition. Through a metabolic screen, the authors classified the effects of a spectrum of metabolites on cell proliferation, finding that AKG alone could potently and specifically inhibit the growth of BCAT1 KO GBM. Unexpectedly, the authors provided evidence that, although AKG can be directly incorporated into the TCA cycle, its inhibitory effect relies on an increase of the NAD+/NADH ratio and disruption of mitochondrial homeostasis.

To reinforce the synergistic role of loss of BCAT1 coupled with AKG, the pharmacologic BCAT1 inhibitor gabapentin was administered in tandem with AKG, which replicated the results of BCAT1 KO. The authors further validated these results in patient-derived tumorspheres and human tumors implanted in mice, demonstrating an absence of toxicity while effecting potent anticancer activity.

The mechanism behind this observation was explored through metabolic experiments including BCKA treatment, examination of the NAD+/NADH ratio, and monitoring of mitochondrial activity and morphology. BCKAs—in particular KIC—rescued the cells and restored the balance between NAD+ and NADH. The increase in NAD+/NADH ratio proved to be a major mechanism by which cell death was orchestrated, which significantly altered mitochondrial morphology and limited their activity. Furthermore, the authors noted that levels of aspartate and accompanying purine metabolism were specifically downregulated by both inhibition of BCAT1 and AKG treatment, and AKG inhibited mTORC1 through disruption of mitochondrial ATP production. Supplementation of KIC was able to partially rescue both of these effects. In total, these results identify a synthetic lethal combination therapy to specifically target IDHWT GBM while sparing healthy tissues. IDHWT GBM is critically reliant on BCAT1 activity to reduce AKG levels, generate NADH, and drive mitochondrial homeostasis. Targeting BCAT1 and increasing AKG levels together inhibit mTORC1 and ultimately reduce cancer proliferation (Fig. 1).

Figure 1.

Synthetic lethality of coadministration of gabapentin and AKG in IDHWT GBM. BCAAs can be catabolized reversibly through BCAT1 into BCKAs, which are then irreversibly processed further into acetyl- and succinyl-CoA for incorporation into the TCA cycle. The electron transport chain (ETC) can utilize NADH to regenerate NAD+ and produce ATP. Inhibition of BCAT1 with gabapentin synergizes with addition of AKG to increase the ratio of NAD+/NADH, to inhibit mitochondrial activity, and to hamper the activity of mTORC1. This culminates in death of tumor cells following loss of proliferative signaling and synthesis of essential biomolecules.

Figure 1.

Synthetic lethality of coadministration of gabapentin and AKG in IDHWT GBM. BCAAs can be catabolized reversibly through BCAT1 into BCKAs, which are then irreversibly processed further into acetyl- and succinyl-CoA for incorporation into the TCA cycle. The electron transport chain (ETC) can utilize NADH to regenerate NAD+ and produce ATP. Inhibition of BCAT1 with gabapentin synergizes with addition of AKG to increase the ratio of NAD+/NADH, to inhibit mitochondrial activity, and to hamper the activity of mTORC1. This culminates in death of tumor cells following loss of proliferative signaling and synthesis of essential biomolecules.

Close modal

Through their thorough analysis, Zhang and colleagues have uncovered some new questions surrounding the role of BCAA metabolism in IDHWT GBM. The authors have shown convincingly that loss of BCAT1 activity has profound effects on GBM metabolism and viability, but it remains unclear where unmodified GBM directs BCAAs and to what end. While it is known that BCAT1 is needed for excitotoxic secretion of glutamate in GBM (1), further work including metabolic isotope tracing is warranted to conceptualize the flux through central carbon and BCAA metabolism and how these fluxes could be altered by varying environmental conditions. This would help to ascertain in more detail the vital role that BCAAs play in GBM and what makes this pathway indispensable for growth despite the presence of other analogous pathways for energy and biosynthesis. Among BCKAs, KIC provides the most robust rescue as well as the greatest yield of NADH per molecule consumed. Still, BCKDH KO failed to replicate the synthetic lethality with AKG, leaving it unclear to what extent energy metabolism contributes rather than regulatory effects like epigenetics.

In addition, the mechanism by which AKG synergizes with BCAT1 inhibition is not yet well-defined. Previous studies have shown that lactate dehydrogenase A could convert AKG to L-2-hydroxyglutarate (2-HG; ref. 7), which could exacerbate the increased NAD+/NADH ratio caused by BCAT1 deficiency. However, AKG but not 2-HG has been shown to inhibit BCAT1 (1). Zhang and colleagues observed increased levels of 2-HG following BCAT1 inhibition and AKG treatment. It is currently not clear whether more AKG is being directed towards 2-HG or accumulation is being promoted through reduced clearance from the tissue. If AKG is being converted to 2-HG in the tumors, potential off-target effects must also be considered. 2-HG is known to act as an oncometabolite with inhibitory effects on AKG-dependent dioxygenases (8). This could potentially result in hypermethylation, which may limit the efficacy of AKG as a tumor suppressor. Further experiments would be necessary to validate this mechanism and to understand how it might be subverted during treatment.

The context of a tumor can have dramatic and unpredictable effects on the outcome of treatments, which must be addressed before clinical application is assured. The broad mutational landscape of GBM means that patient response to this treatment will likely be variable. In addition, GBM resides in a unique tumor niche within the central nervous system and may rely on the support of stromal cells for survival. In other cancers like pancreas, stromal cells support the proliferation of cancer cells by supplying BCKAs to fuel BCAA production through BCAT2 or TCA cycle activity through BCKDH (2). Likewise in GBM, secretion of BCKAs metabolized from BCAAs has been shown to alter macrophage functional activity (9). Interference from other cells in the tumor microenvironment could impact the systemic effects of BCAA treatment or contribute to unexpected results. It is still unclear whether combination treatment with BCAT1 inhibition and AKG will be sufficient to improve patient survival rather than just mitigating progression, especially in the complete context of the tumor microenvironment. With answers to these questions it may be viable to take advantage of this synthetically lethal vulnerability not just in patients with GBM but potentially also in other cancers with similar BCAA dependence such as leukemia, lung, breast, endometrial, bone, liver, or ovarian cancer (10).

No disclosures were reported.

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