Targeting Aberrantly Activated Glycolysis in Natural Killer T Cell Lymphomas

Background and aims: With advances in the diagnosis and treatment of malignant lymphoma, improvements in long-term survival have been achieved for most common lymphoma types. However, natural killer/T-cell lymphoma (NKTL), which is rare in Europe and North America (0.3%) but relatively common in Asia and Latin America, has been left behind as a subtype of lymphoma with a high relapse rate and a poor prognosis. In particular, the outcome of patients with advanced or recurrent NKTL is dismal, with an overall survival of only a few months. This is mainly attributed to the intrinsic resistance of NKTL cells to anthracycline due to the high expression of P-glycoprotein. Molecular-targeted drugs used for other hematological malignancies, such as anti-CD30 and anti-CD52 monoclonal antibodies, have only limited effects on NKTL, and conversely, these drugs can cause severe immunodeficiency and severe autoimmune diseases, often with a fatal course. The development of NKTL-specific and effective molecular-target therapies is expected to improve the prognosis of NKTL. Methods: We performed a gene expression array targeting genes involved in glucose cell metabolism to compare the gene signatures of NK cell lines (NKL, NK92, YT, and KYHG1) and freshly isolated NK cells from three healthy donors (FNK). Results: The expression of various glucose metabolism genes was higher in NK cell lines than in FNK cells, and in particular, the PKM gene was highly enriched in NK cell lines. The PKM gene encodes the pyruvate kinase muscle isozyme 2 (PKM2), which is responsible for ATP production within the glycolytic sequence and plays an essential role in aerobic glycolysis (the Warburg effect), a dominant metabolic pathway used by cancer cells. Immunohistochemistry studies revealed high PKM2 protein expression in 17 out of 20 tumor tissues derived from patients with extranodal NK/T cell lymphoma. Treatment of NKL, NK92, YT, and KYHG1 cells with Benserazide or Shikonin, two known PKM2 inhibitors, robustly induced cell apoptosis in a time- and dose-dependent manner and inhibited colony formation in in vitro clonogenic assays. Western blotting studies showed a dose-dependent inhibition of STAT1 and STAT3 in NK cell lines exposed to PKM2 inhibitors. A colorimetric assay showed that PKM2 glycolytic activity correlated with the PKM2 gene expression in NK cells, with significantly higher glycolytic activity in malignant NK cell lines than in normal FNK cells. Further mechanistic studies showed that in normal FNK cells, PKM2 is mainly in the tetrameric form, where glucose is converted to pyruvate to generate ATP. In contrast, in malignant NK cell lines, PKM2 is mainly expressed in its dimeric form, with a low affinity for pyruvate and a higher affinity for glycolytic intermediates that promote the synthetic processes and thereby lead to persistent cell proliferation. Conclusions: These results substantiate the importance of aerobic glycolysis in malignat NK cells and indicate that targeting this pathway by blocking PKM2 activity has therapeutic potential for NKTL.