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Therapeutic targeting of PFKFB3 with a novel glycolytic inhibitor PFK158 promotes lipophagy and chemosensitivity in gynecologic cancers.

ABSTRACT: Metabolic alterations are increasingly recognized as important novel anti-cancer targets. Among several regulators of metabolic alterations, fructose 2,6 bisphosphate (F2,6BP) is a critical glycolytic regulator. Inhibition of the active form of PFKFB3^ser461 using a novel inhibitor, PFK158 resulted in reduced glucose uptake, ATP production, lactate release as well as induction of apoptosis in gynecologic cancer cells. Moreover, we found that PFK158 synergizes with carboplatin (CBPt) and paclitaxel (PTX) in the chemoresistant cell lines, C13 and HeyA8MDR but not in their chemosensitive counterparts, OV2008 and HeyA8, respectively. We determined that PFK158-induced autophagic flux leads to lipophagy resulting in the downregulation of cPLA2, a lipid droplet (LD) associated protein. Immunofluorescence and co-immunoprecipitation revealed colocalization of p62/SQSTM1 with cPLA2 in HeyA8MDR cells uncovering a novel pathway for the breakdown of LDs promoted by PFK158. Interestingly, treating the cells with the autophagic inhibitor bafilomycin A reversed the PFK158-mediated synergy and lipophagy in chemoresistant cells. Finally, in a highly metastatic PTX-resistant in vivo ovarian mouse model, a combination of PFK158 with CBPt significantly reduced tumor weight and ascites and reduced LDs in tumor tissue as seen by immunofluorescence and transmission electron microscopy compared to untreated mice. Since the majority of cancer patients will eventually recur and develop chemoresistance, our results suggest that PFK158 in combination with standard chemotherapy may have a direct clinical role in the treatment of recurrent cancer.

SUBMITTER: Mondal S

PROVIDER: S-EPMC6261695 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Metabolic dysregulations have emerged as a major mediator of cardiovascular disorders and fibrotic diseases. Metabolic reprogramming contributes a lot to cardiac fibroblast activation and cardiac fibrosis post-myocardial infarction (MI), yet the mechanism remains incompletely understood. Our work aimed to determine whether or not glycolytic reprogramming, regulated by phosphofructokinase-2/fructose-2,6-bisphosphatase 3 (PFKFB3), is a therapeutic target for alleviating post-MI cardiac fibrosis. Here, we showed that cardiac fibroblasts displayed cell energy phenotype toward augmented glycolysis in response to transforming growth factor-beta 1 (TGF-β1), evidenced by significant extracellular acidification rate (ECAR) increase and lactate accumulation. The expression of glycolytic enzyme PFKFB3, a master activator of glycolysis, was up-regulated in TGF-β1-treated cardiac fibroblasts and in cardiac fibroblasts of post-MI mice. Pharmacological inhibition of PFKFB3 by 3PO diminished TGF-β1-mediated profibrotic phenotypes, attenuated cardiac fibrosis, and preserved cardiac functions in post-MI mice. Meanwhile, the genetic inhibition of PFKFB3 decreased the cardiac fibroblast activation and reversed the differentiated phenotypes in vitro and in vivo. Mechanistically, we identified deubiquitinase OTUD4 as a new binding protein of PFKFB3, and their interaction blocked PFKFB3 degradation via OTUD4-mediated deubiquitylation. Taken together, this work characterized a key role for PFKFB3 in cardiac fibroblast activation and suggested that inhibiting PFKFB3-involved glycolysis is an alternative way to alleviate post-MI cardiac fibrosis. KEY MESSAGES: PFKFB3, a master activator of glycolysis, was highly expressed in ischemic cardiac fibroblasts to enhance cardiac fibrosis The deubiquitinase OTUD4 was identified as a new binding protein of PFKFB3 TGF-β1 blunted the ubiquitination-mediated degradation of PFKFB3 via OTUD4-mediated deubiquitylation Blockade of PFKFB3 contributed to ameliorating ischemia-induced cardiac fibrosis.

Project description:BackgroundHepatocellular carcinoma (HCC) remains a leading life-threatening health challenge worldwide, with pressing needs for novel therapeutic strategies. Sphingosine kinase 1 (SphK1), a well-established pro-cancer enzyme, is aberrantly overexpressed in a multitude of malignancies, including HCC. Our previous research has shown that genetic ablation of Sphk1 mitigates HCC progression in mice. Therefore, the development of PF-543, a highly selective SphK1 inhibitor, opens a new avenue for HCC treatment. However, the anti-cancer efficacy of PF-543 has not yet been investigated in primary cancer models in vivo, thereby limiting its further translation.MethodsBuilding upon the identification of the active form of SphK1 as a viable therapeutic target in human HCC specimens, we assessed the capacity of PF-543 in suppressing tumor progression using a diethylnitrosamine-induced mouse model of primary HCC. We further delineated its underlying mechanisms in both HCC and endothelial cells. Key findings were validated in Sphk1 knockout mice and lentiviral-mediated SphK1 knockdown cells.ResultsSphK1 activity was found to be elevated in human HCC tissues. Administration of PF-543 effectively abrogated hepatic SphK1 activity and significantly suppressed HCC progression in diethylnitrosamine-treated mice. The primary mechanism of action was through the inhibition of tumor neovascularization, as PF-543 disrupted endothelial cell angiogenesis even in a pro-angiogenic milieu. Mechanistically, PF-543 induced proteasomal degradation of the critical glycolytic enzyme 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3, thus restricting the energy supply essential for tumor angiogenesis. These effects of PF-543 could be reversed upon S1P supplementation in an S1P receptor-dependent manner.ConclusionsThis study provides the first in vivo evidence supporting the potential of PF-543 as an effective anti-HCC agent. It also uncovers previously undescribed links between the pro-cancer, pro-angiogenic and pro-glycolytic roles of the SphK1/S1P/S1P receptor axis. Importantly, unlike conventional anti-HCC drugs that target individual pro-angiogenic drivers, PF-543 impairs the PFKFB3-dictated glycolytic energy engine that fuels tumor angiogenesis, representing a novel and potentially safer therapeutic strategy for HCC.

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Therapeutic targeting of PFKFB3 with a novel glycolytic inhibitor PFK158 promotes lipophagy and chemosensitivity in gynecologic cancers.

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