IDH1 mutations are common in low-grade gliomas and secondary glioblastomas and cause overproduction of (R)-2HG. (R)-2HG modulates the activity of many enzymes, including some that are linked to transformation and some that are probably bystanders. Although prior work on (R)-2HG targets focused on 2OG-dependent dioxygenases, we found that (R)-2HG potently inhibits the 2OG-dependent transaminases BCAT1 and BCAT2, likely as a bystander effect, thereby decreasing glutamate levels and increasing dependence on glutaminase for the biosynthesis of glutamate and one of its products, glutathione. Inhibiting glutaminase specifically sensitized IDH mutant glioma cells to oxidative stress in vitro and to radiation in vitro and in vivo. These findings highlight the complementary roles for BCATs and glutaminase in glutamate biosynthesis, explain the sensitivity of IDH mutant cells to glutaminase inhibitors, and suggest a strategy for maximizing the effectiveness of such inhibitors against IDH mutant gliomas.

The electrophilic natural product parthenolide has generated significant interest as a model for potential chemotherapeutics. Similar to other α,β-unsaturated carbonyl electrophiles, parthenolide induces the heat shock response in leukemia cells, potentially through covalent adduction of heat shock proteins. Other thiol-reactive electrophiles have also been shown to induce the heat shock response as well as to covalently adduct members of the heat shock protein family, such as heat shock protein 72 (Hsp72). To identify sites of modification of Hsp72 by parthenolide, we used high-resolution tandem mass spectrometry to detect 10 lysine, histidine, and cysteine residues of recombinant Hsp72 as modified in vitro by 10 and 100 μM parthenolide. To further ascertain that modification of Hsp72 by parthenolide occurs inside cells and not simply as an in vitro artifact, an alkyne-labeled derivative of parthenolide was synthesized to enable enrichment and detection of protein targets of parthenolide using copper-catalyzed [3 + 2] azide–alkyne cycloaddition. The alkyne-labeled parthenolide derivative displays an half maximal inhibitory concentration (IC50) in undifferentiated acute monocytic leukemia cells (THP-1) of 13.1 ± 1.1 μM, whereas parthenolide has an IC50 of 4.7 ± 1.1 μM. Concentration dependence of protein modification by the alkyne–parthenolide derivative was demonstrated, as well as in vitro adduction of Hsp72. Following treatment of THP-1 cells in culture by the alkyne–parthenolide, adducted proteins were isolated with neutravidin resin and detected by immunoblotting in the enriched protein fraction. Hsp70 proteins were detected in the enriched proteins, indicating that Hsp70 proteins were adducted intracellularly by the alkyne–parthenolide derivative.

Inactivation of the retinoblastoma gene () product, pRB, is common in many human cancers. Targeting downstream effectors of pRB that are central to tumorigenesis is a promising strategy to block the growth of tumors harboring loss-of-function mutations. One such effector is retinoblastoma-binding protein 2 (RBP2, also called JARID1A or KDM5A), which encodes an H3K4 demethylase. Binding of pRB to RBP2 has been linked to the ability of pRB to promote senescence and differentiation. Importantly, genetic ablation of RBP2 is sufficient to phenocopy pRB's ability to induce these cellular changes in cell culture experiments. Moreover, germline deletion significantly impedes tumorigenesis in mice. The value of RBP2 as a therapeutic target in cancer, however, hinges on whether loss of RBP2 could block the growth of established tumors as opposed to simply delaying their onset. Here we show that conditional, systemic ablation of RBP2 in tumor-bearing mice is sufficient to slow tumor growth and significantly extend survival without causing obvious toxicity to the host. These findings show that established -null tumors require RBP2 for growth and further credential RBP2 as a therapeutic target in human cancers driven by inactivation.