Abstract 293: Non-metabolic regulatory functions of nuclear pyruvate kinase M2 induce a MAPK mediated phenotypic switch in invasive cancers
Authors: Vanaja Kiran , Apostolidi Maria , Levchenko Andre , Rinehart Jesse 
Published: April 2023

Abstract
Accumulating evidence suggests that PKM2, the differentially spliced pyruvate kinase isoform that is expressed in most cancers and embryonic tissue, is linked with epithelial-mesenchymal transition (EMT). However, how PKM2 may drive this process of tumor invasiveness remains unclear. Here we demonstrate that in metastatic hormone-sensitive and ovarian cancer cells PKM2 undergoes modification by phosphorylation on its Serine 37 residue (pS37) and that this modified protein displays distinct localization patterns in response to pro-migratory cues emanating from an artificial nanopatterned substrate mimicking the tumor associated extracellular matrix. We find that in epithelial-mesenchymal (EM) hybrid cohorts, migrating cells undergo phenotypic switching from followers to leaders and the PKM2-pS37 signals correlate with different cell phenotypes (less/more invasive and leaders vs followers). This phenotypic switching occurs as a function of cell distance from the leading edge in an expanding epithelial sheet. We found that the putative novel non-metabolic functions of PKM2 are attributed to its autoregulation that results in not only a PKM2-pS37 upregulation but also an induction in PKM2 expression via ERK signaling. Cues emanating from either the extracellular matrix (mediated via Integrin/FAK) or through growth factor signaling (EGFR signaling), culminating in ERK activation induce PKM2 phosphorylation at S37. Thus, our data suggest that an increase in PKM2pS37 is accompanied by a switch to less enzymatically active molecular complexes which can translocate to the nucleus and increase PKM2 expression through an auto-positive feedback loop resulting in increased total PKM2 concentration and function in the leader cells. To further confirm the existence of this positive feedback loop we developed a mathematical model for PKM2 regulation. This model allows for the evaluation of inputs emerging from the extracellular matrix, such as the integrin/FAK signaling, and growth factor receptor signaling both of which regulate PKM2 expression through ERK activation in a switch-like fashion. The mathematical model includes non-linearity in the phosphorylation of S37 on PKM2 and the non-linearity in the nuclear localization and upregulation of PKM2. Further experimental analysis supports the functional role of the switch, as the observed PKM2 expression states correlate with the expression of characteristic EMT markers and invasion-promoting pseudo-hypoxic stress in the leader cells. Importantly, the small molecule activator of PKM2, TEPP 46, can restore a more epithelial-like state, decrease invasion, and increase glucose uptake in these EM hybrid ovarian cancer cells. Taken together, our results highlight a link between PKM2 and cancer aggressiveness driven by novel regulatory mechanisms that support both metabolic rewiring and invasive spread of tumor cells.
Citation Format: Kiran Vanaja, Maria Apostolidi, Andre Levchenko, Jesse Rinehart. Non-metabolic regulatory functions of nuclear pyruvate kinase M2 induce a MAPK mediated phenotypic switch in invasive cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 293.