Abstract B024: PKM2 activation suppresses cellular ROS scavenging capacity and potentiates doxorubicin antitumor activity

Abstract

Abstract Pyruvate kinase (PK) is a key mediator of the metabolic reprogramming and the Warburg effect observed in tumorigenesis. PK catalyzes the rate-limiting conversion of phosphoenolpyruvate (PEP) to pyruvate in glycolysis. There are fourteen known transcript variants of the PKM gene. In normal adult tissues, the M1 (PKM1) isoform predominates, promoting oxidative phosphorylation and generation of ATP. The M2 (PKM2) isoform, however, and indeed many of the PKM variants other than M1, predominate in tumor tissues. Unlike PKM1, PKM2 exists largely as a less active dimer. The decreased activity of dimer PKM2, relative to PKM1, in this key conversion step leads to an accumulation and shunting of upstream glycolytic intermediates into synthetic pathways needed for the building blocks necessary to maintain continued tumor cell growth. Thus, PKM2 is the focus of intense research into its utility as a target for the treatment of cancer. It has been hypothesized that PKM2 activation should starve cells of necessary building blocks, slowing tumor cell proliferation. We have developed a PKM2 activator, TP-1454, that activates PKM2 in biochemical assays with an AC50 of 11 nM. In cell viability assays, TP-1454 inhibited A549 lung cell growth with an IC50 of 3.4 μM. In conditions where additional nutrients, such as serine, are withdrawn but not growth limiting, TP-1454 inhibited cell growth with an IC50 of 20.3 nM. Consistent with the notion that PKM2 activation leads to a reduction in upstream building blocks, a mass spectrometry metabolomics study showed that TP-1454-induced PKM2 activation modulated several key cellular metabolic components, including key amino acids and glycolytic and citric acid intermediates. However, the single highest-modulated metabolite was glutathione, which was reduced more than 10-fold in treated cells. This finding was also confirmed using a luciferase-based approach with the GSH-Glo glutathione assay. These data led us to hypothesize that PKM2 activation may cause multiple changes in cellular metabolism, but that the greatest vulnerability induced by PKM2 activation was to compromise the cell’s ability to combat reactive oxygen species (ROS) due to the loss of glutathione levels. Therefore, we reasoned that TP-1454 would combine well with drugs known to induce ROS-mediated cell death, such as the anthracyclines. Doxorubicin inhibited A549 cell viability with an IC50 of 173 nM. We observed a four-fold reduction in IC50 of A549 cells treated with 4 μM TP-1454. The antioxidant N-acetyl cysteine partially rescued the synergistic effect of TP-1454 on doxorubicin activity, suggesting that the combination effect was ROS driven. Studies investigating TP-1454-induced glutathione suppression in combination with doxorubicin, in vivo, are currently under way. Taken together, these data support the hypothesis that PKM2 activators may combine well with anthracycline drugs and warrant clinical investigation of PKM2 as a potential therapeutic target for the treatment of multiple cancer types. Citation Format: Peter Peterson, Clifford J. Whatcott, David J. Bearss, Steven L. Warner, Adam Siddiqui-Jain. PKM2 activation suppresses cellular ROS scavenging capacity and potentiates doxorubicin antitumor activity [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B024.