Pyruvate kinase modulators as a therapy target: an updated patent review 2018-2023.

INTRODUCTION: Cancer cells adopt a glycolytic phenotype to fulfill their energy needs in unfavorable conditions. In metabolic rewiring, cancer cells upregulate the expression of glycolytic pathway regulators including glucose transporter 1, hexokinase 2, and PKM2 (pyruvate kinase) into its M2 splice form. Among these regulators, PKM2 plays a major role in metabolic reprogramming and is overexpressed in various diseases, including cancer. Dimerization of PKM2 causes the generation of synthetic precursors from glycolytic intermediates, which are essential for cellular growth and cancer cell proliferation. COVERED AREAS: This article is focused on examining recent patents (2018-2023) on PKM2 activators, inhibitors and their biological and synthesis properties by using the advanced search service of the European Patent Office (EPO). Moreover, other databases including PubMed, Google Scholar and Elsevier were also examined for scientific data. On basis of their chemical structures, PKM2 activators and inhibitors are classified into pyrazole, pyrolidine-pyrazole, phenol, benzoxazine, isoselenazolo-pyridinium, phthalazine, and propiolylamide derivatives. EXPERT OPINION: Activating PKM2 reduces proliferation and development of cells by reducing the quantity of biomolecules needed for cell formation. PKM2 activators and inhibitors are highly effective in treating many cancer pathogens. It is important to find new, more potent and selective molecules for PKM2 activation and inhibition.

Estrogen treatment in combination with pyruvate kinase M2 inhibition precipitate significant cumulative antitumor effects in colorectal cancer.

It is well established that pyruvate kinase M2 (PKM2) activity contributes to metabolic reprogramming in various cancers, including colorectal cancer (CRC). Estrogen or 17ß-estradiol (E2) signaling is also known to modulate glycolysis markers in cancer cells. However, whether the inhibition of PKM2 combined with E2 treatment could adversely affect glucose metabolism in CRC cells remains to be investigated. First, we confirmed the metabolic plasticity of CRC cells under varying environmental conditions. Next, we identified glycolysis markers that were upregulated in CRC patients and assessed in vitro mRNA levels following E2 treatment. We found that PKM2 expression, which is highly upregulated in CRC clinical samples, is not altered by E2 treatment in CRC cells. In this study, glucose uptake, generation of reactive oxygen species (ROS), lactate production, cell viability, and apoptosis were evaluated in CRC cells following E2 treatment, PKM2 silencing, or a combination of both. Compared to individual treatments, combination therapy resulted in a significant reduction in cell viability and enhanced apoptosis. Glucose uptake and ROS production were markedly reduced in PKM2-silenced E2-treated cells. The data presented here suggest that E2 signaling combined with PKM2 inhibition cumulatively targets glucose metabolism in a manner that negatively impacts CRC cell growth. These findings hold promise for novel therapeutic strategies targeting altered metabolic pathways in CRC.

Discovery of Cell-Permeable Allosteric Inhibitors of Liver Pyruvate Kinase: Design and Synthesis of Sulfone-Based Urolithins.

Metabolic dysfunction-associated fatty liver disease (MAFLD) presents a significant global health challenge, characterized by the accumulation of liver fat and impacting a considerable portion of the worldwide population. Despite its widespread occurrence, effective treatments for MAFLD are limited. The liver-specific isoform of pyruvate kinase (PKL) has been identified as a promising target for developing MAFLD therapies. Urolithin C, an allosteric inhibitor of PKL, has shown potential in preliminary studies. Expanding upon this groundwork, our study delved into delineating the structure-activity relationship of urolithin C via the synthesis of sulfone-based urolithin analogs. Our results highlight that incorporating a sulfone moiety leads to substantial PKL inhibition, with additional catechol moieties further enhancing this effect. Despite modest improvements in liver cell lines, there was a significant increase in inhibition observed in HepG2 cell lysates. Specifically, compounds 15d, 9d, 15e, 18a, 12d, and 15a displayed promising IC50 values ranging from 4.3 µM to 18.7 µM. Notably, compound 15e not only demonstrated a decrease in PKL activity and triacylglycerol (TAG) content but also showed efficient cellular uptake. These findings position compound 15e as a promising candidate for pharmacological MAFLD treatment, warranting further research and studies.

The search for pyruvate kinase-R activators; from a HTS screening hit via an impurity to the discovery of a lead series.

Pyruvate kinase (PK) is an essential component of cellular metabolism, converting ADP and phosphoenolpyruvate (PEP) to pyruvate in the final step of glycolysis. Of the four unique isoforms of pyruvate kinase, R (PKR) is expressed exclusively in red blood cells and is a tetrameric enzyme that depends on fructose-1,6-bisphosphate (FBP) for activation. PKR deficiency leads to hemolysis of red blood cells resulting in anemia. Activation of PKR in both sickle cell disease and beta-thalassemia patients could lead to improved red blood cell fitness and survival. The discovery of a novel series of substituted urea PKR activators, via the serendipitous identification and diligent characterization of a minor impurity in an High Throughput Screening (HTS) hit will be discussed.

Polyphyllin II Induces Apoptosis in Fibrosarcoma Cells via Activating Pyruvate Kinase M2.

Aerobic glycolysis is a metabolic reprogramming of tumor cells that is essential for sustaining their phenotype of fast multiplication by continuously supplying energy and mass. Pyruvate kinase M2 (PKM2) has a vital role in this process, which has given it high interest as a target for anticancer drug development. With potent toxicity to many types of cancer cells, polyphyllin II (PP2), a steroidal saponin isolated from the herbaceous plant Rhizoma paridis, brought to our attention that it might interfere with the PKM2 activity. In this study, we discovered that PP2 was a novel agonist of PKM2. PP2 activated recombinant PKM2 and changed the protein's oligomeric state to activate intracellular PKM2. At the same time, PP2 suppressed its protein kinase function by decreasing the content of nuclear PKM2. The mRNA levels of its downstream genes, such as Glut1, LDHA, and MYC, were inhibited. In addition, PP2 induced oxidative stress by downregulating the expression and activity of antioxidant proteins such as NQO1, TrxR, and Trx in HT-1080 cells, which in turn led to mitochondrial dysfunction and ultimately induced apoptosis. Moreover, PP2 reduced the proliferation and migration of HT-1080 cells. Thus, targeting the glycolysis pathway offers an unprecedented mode of action for comprehending PP2's pharmacological impacts and advances PP2's further development in fibrosarcoma therapy.

Design and synthesis of novel JNK inhibitors targeting liver pyruvate kinase for the treatment of non-alcoholic fatty liver disease and hepatocellular carcinoma.

Non-alcoholic fatty liver disease (NAFLD) comprises a broad range of liver disease including hepatocellular carcinoma (HCC) with is no FDA-approved drug. Liver pyruvate kinase (PKL) is a major regulator of metabolic flux and ATP generation in liver presenting a potential target for the treatment of NAFLD. Based on our recent finding of JNK-5A's effectiveness in inhibiting PKLR expression through a drug repositioning pipeline, this study aims to improve its efficacy further. We synthesized a series of JNK-5A analogues with targeted modifications, guided by molecular docking studies. These compounds were evaluated for their activities on PKL expression, cell viability, triacylglyceride (TAG) levels, and the expressions of steatosis-related proteins in the human HepG2 cell line. Subsequently, the efficacy of these compounds was assessed in reducing TAG level and toxicity. Compounds 40 (SET-151) and 41 (SET-152) proved to be the most efficient in reducing TAG levels (11.51 ± 0.90 % and 10.77 ± 0.67 %) and demonstrated lower toxicity (61.60 ± 5.00 % and 43.87 ± 1.42 %) in HepG2 cells. Additionally, all synthesized compounds were evaluated for their anti-cancer properties revealing that compound 74 (SET-171) exhibited the highest toxicity in cell viability with IC50 values of 8.82 µM and 2.97 µM in HepG2 and Huh7 cell lines, respectively. To summarize, compounds 40 (SET-151) and 41 (SET-152) show potential for treating NAFLD, while compound 74 (SET-171) holds potential for HCC therapy.

A non-canonical role for pyruvate kinase M2 as a functional modulator of Ca2+ signalling through IP3 receptors.

Pyruvate kinase isoform M2 (PKM2) is a rate-limiting glycolytic enzyme that is widely expressed in embryonic tissues. The expression of PKM2 declines in some tissues following embryogenesis, while other pyruvate kinase isozymes are upregulated. However, PKM2 is highly expressed in cancer cells and is believed to play a role in supporting anabolic processes during tumour formation. In this study, PKM2 was identified as an inositol 1,4,5-trisphosphate receptor (IP3R)-interacting protein by mass spectrometry. The PKM2:IP3R interaction was further characterized by pull-down and co-immunoprecipitation assays, which showed that PKM2 interacted with all three IP3R isoforms. Moreover, fluorescence microscopy indicated that both IP3R and PKM2 localized at the endoplasmic reticulum. PKM2 binds to IP3R at a highly conserved 21-amino acid site (corresponding to amino acids 2078-2098 in mouse type 1 IP3R isoform). Synthetic peptides (denoted 'TAT-D5SD' and 'D5SD'), based on the amino acid sequence at this site, disrupted the PKM2:IP3R interaction and potentiated IP3R-mediated Ca2+ release both in intact cells (TAT-D5SD peptide) and in a unidirectional 45Ca2+ flux assay on permeabilized cells (D5SD peptide). The TAT-D5SD peptide did not affect the enzymatic activity of PKM2. Reducing PKM2 protein expression using siRNA increased IP3R-mediated Ca2+ signalling in intact cells without altering the ER Ca2+ content. These data identify PKM2 as an IP3R-interacting protein that inhibits intracellular Ca2+ signalling. The elevated expression of PKM2 in cancer cells is therefore not solely connected to its canonical role in glycolytic metabolism, rather PKM2 also has a novel non-canonical role in regulating intracellular signalling.

Shikonin alleviates choroidal neovascularization by inhibiting proangiogenic factor production from infiltrating macrophages.

Choroidal neovascularization (CNV), a feature of neovasular age-related macular degeneration (AMD), acts as a leading cause of vision loss in the elderly. Shikonin (SHI), a natural bioactive compound extracted from Chinese herb radix arnebiae, exerts anti-inflammatory and anti-angiogenic roles and also acts as a potential pyruvate kinase M2 (PKM2) inhibitor in macrophages. The major immune cells macrophages infiltrate the CNV lesions, where the production of pro-angiognic cytokines from macrophage facilitates the development of CNV. PKM2 contributes to the neovascular diseases. In this study, we found that SHI oral gavage alleviated the leakage, area and volume of mouse laser-induced CNV lesion and inhibited macrophage infiltration without ocular cytotoxicity. Moreover, SHI inhibited the secretion of pro-angiogenic cytokine, including basic fibroblast growth factor (FGF2), insulin-like growth factor-1 (IGF1), chemokine (C-C motif) ligand 2 (CCL2), placental growth factor and vascular endothelial growth factor (VEGF), from primary human macrophages by down-regulating PKM2/STAT3/CD163 pathway, indicating a novel potential therapy strategy for CNV.

Silencing of Pyruvate Kinase M2 via a Metal-Organic Framework Based Theranostic Gene Nanomedicine for Triple-Negative Breast Cancer Therapy.

Triple-negative breast cancer (TNBC) is typically associated with poor prognosis due to its only partial response to chemotherapy and lack of clinically established targeted therapies coupled with an aggressive disease course. Aerobic glycolysis is a hallmark of reprogrammed metabolic activity in cancer cells, which can be repressed by small-interfering RNA (siRNA). However, the lack of effective carriers to deliver vulnerable siRNA restricts the clinical potentials of glycolysis-based gene therapy for TNBC. Herein, we develop a tumor-targeted, biomimetic manganese dioxide (MnO2)-shrouded metal-organic framework (MOF) based nanomedicine to deliver siRNA against pyruvate kinase muscle isozyme M2 (siPKM2), wherein PKM2 is a rate-limiting enzyme in glycolysis, to inhibit the reprogrammed glycolysis of TNBC. This MOF-based genetic nanomedicine shows excellent monodispersity and stability and protects siPKM2 against degradation by nucleases. The nanomedicine not only substantially blocks the glycolytic pathway but also improves intracellular hypoxia in TNBC cells, with a resultant O2-enhanced anticancer effect. In the mice orthotopic TNBC model, the nanomedicine shows a remarkable therapeutic effect. Meanwhile, the Mn2+ ions released from acid microenvironment-responsive MnO2 enable in vivo monitoring of the therapeutic process with magnetic resonance imaging (MRI). Our study shows great promise with this MRI-visible MOF-based nanomedicine for treating TNBC by inhibition of glycolysis via the RNA interference.

PKM2 Is a Potential Diagnostic and Therapeutic Target for Retinitis Pigmentosa.

Retinitis pigmentosa (RP) is a major cause of blindness that is difficult to diagnose and treat. PKM2, a subtype of pyruvate kinase, is strongly associated with oxidative stress and is expressed in photoreceptors. We investigated whether PKM2 reduces photoreceptor cell apoptosis and evaluated possible antiapoptotic mechanisms in RP. We established RP models by exposing 661W cells to blue light and modulated PKM2 activity using a PKM2 inhibitor. We measured the apoptosis rates using calcein-acetoxymethyl ester/propidium iodide double staining and Cell Counting Kit-8, the oxidative stress levels using a reactive oxygen species assay, and the changes in protein expression by western blotting. Photodamage increased PKM2 expression, cellular oxidative stress, and apoptosis of 661W cells. PKM2 inhibition significantly reduced the levels of apoptosis and oxidative stress induced by photodamage. Our data suggest that PKM2 is a potential disease marker and therapeutic target for RP.

Rationalization of the activity Profile of Pyruvate Kinase Isozyme M2 (PKM2) Inhibitors using 3D QSAR.

INTRODUCTION: Pyruvate kinase isozyme M2 (PKM2) was observed to be overexpressed and play a key role in cell growth and cancer cells' metabolism. During the past years, phytochemicals have been developed as new treatment options for chemoprevention and cancer therapy. Natural resources, like shikonin (naphthoquinone) and its derivatives, have emerged to be high potential therapeutics in cancer treatment. METHODS: Our study aimed to design novel anti-tumour agents (PKM2 inhibitors) focusing on the shikonin scaffold with a better activity using computational methods. We applied a three-dimensional quantitative structure-activity relationship (3D-QSAR) approach using Field-based QSAR. RESULTS: The Comparative Molecular Field Analysis (CoMFA) and Comparative Molecular Similarity Indices Analysis (CoMSIA) were performed on a series of forty shikonin derivatives, including shikonin, to develop robust models and rationalize the PKM2 inhibitory activity profile by building a correlation between structural features and activity. CONCLUSION: These predictive computational models will further help the design and synthesis of potent PKM2 inhibitors and their fast biological assessment at a low cost.

Specific Pyruvate Kinase M2 Inhibitor, Compound 3K, Induces Autophagic Cell Death through Disruption of the Glycolysis Pathway in Ovarian Cancer Cells.

Ovarian cancer is a common cause of death among gynecological cancers. Although ovarian cancer initially responds to chemotherapy, frequent recurrence in patients remains a therapeutic challenge. Pyruvate kinase M2 (PKM2) plays a pivotal role in regulating cancer cell survival. However, its therapeutic role remains unclear. Here, we investigated the anticancer effects of compound 3K, a specific PKM2 inhibitor, on the regulation of autophagic and apoptotic pathways in SK-OV-3 (PKM2-overexpressing human ovarian adenocarcinoma cell line). The anticancer effect of compound 3K was examined using MTT and colony formation assays in SK-OV-3 cells. PKM2 expression was positively correlated with the severity of the tumor, and expression of pro-apoptotic proteins increased in SK-OV-3 cells following compound 3K treatment. Compound 3K induced AMPK activation, which was accompanied by mTOR inhibition. Additionally, this compound inhibited glycolysis, resulting in reduced proliferation of SK-OV-3 cells. Compound 3K treatment suppressed tumor progression in an in vivo xenograft model. Our findings suggest that the inhibition of PKM2 by compound 3K affected the Warburg effect and induced autophagic cell death. Therefore, use of specific PKM2 inhibitors to block the glycolytic pathway and target cancer cell metabolism represents a promising therapeutic approach for treating PKM2-overexpressing ovarian cancer.

Metabolic but not transcriptional regulation by PKM2 is important for natural killer cell responses.

Natural Killer (NK) cells have an important role in immune responses to viruses and tumours. Integrating changes in signal transduction pathways and cellular metabolism is essential for effective NK cells responses. The glycolytic enzyme Pyruvate Kinase Muscle 2 (PKM2) has described roles in regulating glycolytic flux and signal transduction, particularly gene transcription. While PKM2 expression is robustly induced in activated NK cells, mice lacking PKM2 in NK cells showed no defect in NK cell metabolism, transcription or antiviral responses to MCMV infection. NK cell metabolism was maintained due to compensatory PKM1 expression in PKM2-null NK cells. To further investigate the role of PKM2, we used TEPP-46, which increases PKM2 catalytic activity while inhibiting any PKM2 signalling functions. NK cells activated with TEPP-46 had reduced effector function due to TEPP-46-induced increases in oxidative stress. Overall, PKM2-regulated glycolytic metabolism and redox status, not transcriptional control, facilitate optimal NK cells responses.

Upregulated PKM2 in Macrophages Exacerbates Experimental Arthritis via STAT1 Signaling.

Recent studies indicate that glucose metabolism is altered in rheumatoid arthritis. We hypothesize that Pkm2, as a key regulatory enzyme of glycolysis pathway, triggers the activation of macrophages (Mφ), which results in proinflammatory cytokine production during the arthritis progress. In this study, Pkm2 was found to be overexpressed in ED1-positive Mφ in spleens and synovial tissues from arthritic rats via immunofluorescence, Western blotting, and quantitative RT-PCR. To reveal the role of Pkm2, Dark Agouti rats were treated with either Pkm2 enzyme inhibitor shikonin or the RNA interference plasmids of Pkm2 and negative control plasmids, respectively, via i.p. injection. Pkm2 intervention could alleviate the severity of pristane-induced arthritis in aspects of the macroscopic arthritis score, perimeter changes of midpaw, and the synovitis and destruction of the bone and cartilage as well as reduce the ED1 and p-Stat1-positive cell population in rat synovial tissues. Silencing Pkm2 by RNA interference in classical activated rat and mouse Mφ resulted in less Tnf-α, Il-1ß production via Stat1 signaling. Collectively, Pkm2 is highly expressed in ED1-positive Mφ of spleens and synovial tissues from arthritic rats and promotes Mφ activation via Stat1 signaling. Pkm2 might be a promising selective metabolic target molecule for rheumatoid arthritis treatment.

Identification of Gliotoxin isolated from marine fungus as a new pyruvate kinase M2 inhibitor.

Pyruvate kinase M2 (PKM2) functions as an important rate-limiting enzyme of aerobic glycolysis that is involved in tumor initiation and progression. However, there are few studies on effective PKM2 inhibitors. Gliotoxin is a marine-derived fungal secondary metabolite with multiple biological activities, including immunosuppression, cytotoxicity, and et al. In this study, we found that Gliotoxin directly bound to PKM2 and inhibited its glycolytic activity in a dose-dependent manner accompanied by the decreases in glucose consumption and lactate production in the human glioma cell line U87. Moreover, Gliotoxin suppressed tyrosine kinase activity of PKM2, leading to a dramatic reduction in Stat3 phosphorylation in U87 cells. Furthermore, Gliotoxin suppressed cell viability in U87 cells, and cytotoxicity of Gliotoxin on U87 cells was obviously augmented under hypoxia condition compared to normal condition. Finally, Gliotoxin was demonstrated to induce cell apoptosis of U87 cells and synergize with temozolomide. Our findings identify Gliotoxin as a new PKM2 inhibitor with anti-tumor activity, which lays the foundation for the development of Gliotoxin as a promising anti-tumor drug in the future.

The synthetic oleanane triterpenoid CDDO-Me binds and inhibits pyruvate kinase M2.

BACKGROUND: The M2 isoform of the glycolytic enzyme pyruvate kinase (PKM2) is one of the key components in the Warburg effect, and an important regulator of cancer cell metabolism. Elevated PKM2 expression is a hallmark of numerous tumor types, making it a promising target for cancer therapy. METHODS: Migration of H1299 lung tumor cells treated with synthetic oleanane triterpenoid derivatives CDDO-Me and CDDO-Im was monitored using scratch and transwell assays. Direct binding and inhibition of PKM2 activity by CDDO-Me was demonstrated by pull-down and activity assays. PKM2 localization in the absence and presence of CDDO-Me or CDDO-Im was determined by subcellular fractionation and immunofluorescence microscopy. Involvement of PKM2 in tumor cell migration was assessed using a stable PKM2 knockdown cell line. RESULTS: We demonstrate that migration of H1299 lung tumor cells is inhibited by CDDO-Me and CDDO-Im in scratch and transwell assays. CDDO-Me binds directly and specifically to recombinant PKM2, leading to a reduction of its catalytic activity. PKM2 knockdown cells exhibit significantly lower migration compared to control cells when subjected to glucose and oxygen deprivation, but not under regular conditions. CONCLUSIONS: The results suggest that PKM2 expression in a tumor-like environment contributes to cell migration, and that PKM2 activity can be down regulated by synthetic triterpenoid derivatives.

Identification of Parthenolide Dimers as Activators of Pyruvate Kinase M2 in Xenografts of Glioblastoma Multiforme in Vivo.

Herein we detail the discovery of a series of parthenolide dimers as activators of PKM2 and evaluation of their anti-GBM activities. The most promising compound 5 showed high potency to activate PKM2 with an AC50 value of 15 nM, inhibited proliferation and metastasis, and induced apoptosis of GBM cells. Compound 5 could promote tetramer formation of PKM2 and reduce nucleus translocation of PKM2 in GBM cells without influence on the expression of total PKM2, thereby inhibiting the STAT3 signal pathway in vitro and in vivo. PKM2 knockdown assay demonstrated that the anti-GBM effect of 5 mainly depended on the expression of PKM2 in vitro and in vivo. Compound 16, a prodrug of 5, markedly suppressed U118 tumor xenograft growth and reduced the weight of tumor. On the basis of these investigations, we propose that 16 might be considered as a promising lead compound for discovery of anti-GBM drugs.

OSU-A9 induced-reactive oxygen species cause cytotoxicity in duodenal and gastric cancer cells by decreasing phosphorylated nuclear pyruvate kinase M2 protein levels.

Pyruvate kinase M2 (PKM2) is a key enzyme responsible for the final step of glycolysis. It is still unclear whether PKM2 is involved in reactive oxygen species (ROS)-mediated cytotoxicity in gastrointestinal cancer, and what mechanisms are involved. One duodenal (AZ521) and two gastric (NUGC and SCM-1) cancer cell lines were treated with an indole-3-carbinol derivative OSU-A9, which caused cytotoxicity in acute myeloid leukemia through ROS generation. OSU-A9 caused a dose- and time-dependent cytotoxicity and induced apoptosis in duodenal and gastric cancer cells through ROS generation. Pretreatment with ROS scavengers rescued cancer cells from apoptosis and concomitant poly (ADP-ribose) polymerase cleavage, implying a key role of ROS in OSU-A9-induced cell death. Moreover, OSU-A9-induced ROS generation decreased protein levels of pTyr105-PKM2, and this effect was rescued by pretreatment with ROS scavengers. Interestingly, pTyr105-PKM2 protein levels decreased in the cell nucleus rather than in the cytoplasm. PKM2 overexpression partially rescued the survival of duodenal and gastric cancer cells treated with OSU-A9. Furthermore, the anticancer activity of OSU-A9 extended in vivo, as OSU-A9 administered by oral gavage suppressed the growth of AZ521 xenograft tumors in nude mice without obvious toxicity. In conclusion, OSU-A9 inhibited duodenal and gastric cancer cell proliferation through ROS generation and caused a subsequent decrease in nuclear pTyr105-PKM2 protein. These findings provide evidence for the non-canonical activity of PKM2 in cancer cell survival. Furthermore, they highlight the potential role of PKM2 as a future therapeutic target for duodenal and gastric cancer.

Synthesis and Target Identification of Benzoxepane Derivatives as Potential Anti-Neuroinflammatory Agents for Ischemic Stroke.

Benzoxepane derivatives were designed and synthesized, and one hit compound emerged as being effective in vitro with low toxicity. In vivo, this hit compound ameliorated both sickness behavior through anti-inflammation in LPS-induced neuroinflammatory mice model and cerebral ischemic injury through anti-neuroinflammation in rats subjected to transient middle cerebral artery occlusion. Target fishing for the hit compound using photoaffinity probes led to identification of PKM2 as the target protein responsible for anti-inflammatory effect of the hit compound. Furthermore, the hit exhibited an anti-neuroinflammatory effect in vitro and in vivo by inhibiting PKM2-mediated glycolysis and NLRP3 activation, indicating PKM2 as a novel target for neuroinflammation and its related brain disorders. This hit compound has a better safety profile compared to shikonin, a reported PKM2 inhibitor, identifying it as a lead compound in targeting PKM2 for the treatment of inflammation-related diseases.

PKM2-dependent glycolysis promotes the proliferation and migration of vascular smooth muscle cells during atherosclerosis.

Increased glycolysis is involved in the proliferation and migration of vascular smooth muscle cells (VSMCs). Pyruvate kinase isoform M2 (PKM2), a key rate-limiting enzyme in glycolysis, accelerates the proliferation and migration of tumor cells. Although the intracellular mechanisms associated with oxidized low-density lipoprotein (oxLDL)-stimulated VSMC proliferation and migration have been extensively explored, it is still unclear whether oxLDL promotes the proliferation and migration of VSMCs by enhancing PKM2-dependent glycolysis. In the present study, we detected PKM2 expression and pyruvate kinase activity in oxLDL-treated VSMCs and explored the regulation of PKM2 in oxLDL-treated VSMCs and apoE-/- mice. The results showed that PKM2 expression in VSMCs was higher in the intima than in the media in plaques from atherosclerotic rabbits. Moreover, PKM2 level in VSMCs was increased during atherosclerosis progression in apoE-/- mice. Both PKM2 expression and pyruvate kinase activity were found to be upregulated by oxLDL stimulation in VSMCs. Shikonin (SKN), a specific inhibitor of PKM2, was found to inhibit the oxLDL-induced proliferation and migration in VSMCs, in addition to delaying the atherosclerosis progression in apoE-/- mice. More importantly, oxLDL increased glucose uptake, ATP and lactate production, and the extracellular acidification rate in VSMCs, which could be reversed by SKN. Meanwhile, oxygen consumption rate was unchanged after oxLDL stimulation, suggesting that glycolysis is the main contributor to the energy supply in oxLDL-treated VSMCs. Our results suggest that oxLDL induces VSMC proliferation and migration by upregulating PKM2-dependent glycolysis, thereby contributing to the atherosclerosis progression. Thus, targeting PKM2-dependent glycolysis might provide a novel therapeutic approach for the treatment of atherosclerosis.