M2-PK Research Articles

Stool DNA testing for early detection of colorectal cancer: systematic review using the HTA Core Model® for Rapid Relative Effectiveness Assessment.

Stool DNA testing for early detection of colorectal cancer (CRC) is a non-invasive technology with the potential to supplement established CRC screening tests. The aim of this health technology assessment was to evaluate effectiveness and safety of currently CE-marked stool DNA tests, compared to other CRC tests in CRC screening strategies in an asymptomatic screening population. The assessment was carried out following the guidelines of the European Network for Health Technology Assessment (EUnetHTA). This included a systematic literature search in MED-LINE, Cochrane and EMBASE in 2018. Manufacturers were asked to provide additional data. Five patient interviews helped assessing potential ethical or social aspects and patients' experiences and preferences. We assessed the risk of bias using QUADAS-2, and the quality of the body of evidence using GRADE. We identified three test accuracy studies, two of which investigated a multitarget stool DNA test (Cologuard®, compared fecal immunochemical test (FIT)) and one a combined DNA stool assay (ColoAlert®, compared to guaiac-based fecal occult blood test (gFOBT), Pyruvate Kinase Isoenzyme Type M2 (M2-PK) and combined gFOBT/M2-PK). We found five published surveys on patient satisfaction. No primary study investigating screening effects on CRC incidence or on overall mortality was found. Both stool DNA tests showed in direct comparison higher sensitivity for the detection of CRC and (advanced) adenoma compared to FIT, or gFOBT, respectively, but had lower specificity. However, these comparative results may depend on the exact type of FIT used. The reported test failure rates were higher for stool DNA testing than for FIT. The certainty of evidence was moderate to high for Cologuard® studies, and low to very low for the ColoAlert® study which refers to a former version of the product and yielded no direct evidence on the test accuracy for ad-vanced versus non-advanced adenoma. ColoAlert® is the only stool DNA test currently sold in Europe and is available at a lower price than Cologuard®, but reliable evidence is lacking. A screening study including the current product version of ColoAlert® and suitable comparators would, therefore, help evaluate the effectiveness of this screening option in a European context.

Evaluation of plasma M2-Pyruvate Kinase as a novel biomarker in the differentiation between Functional and Organic Colonic Disorders

Abstract Background Differentiating between functional bowel disorders and organic colonic disorders can often be difficult as they may be present with similar symptoms. Aim of the Work to evaluate the diagnostic value of plasma M2-pyruvate kinase level in differentiating functional colonic disorders (e.g: IBS) from organic colonic disorders (e.g: IBD, colorectal polyps and colorectal cancer) and to assess its use as screening tools for inflammatory bowel disease, premalignant and malignant colorectal lesions. Patients and Methods This study was conducted on 80 patients who divided into four groups: Group I included 20 patients, representing functional bowel disorders (as a control group). Group II included 20 patients with inflammatory bowel diseases; Group III included 20 patients with colorectal polyps; Group IV included 20 patients with colorectal cancer. Results The current study showed that plasma M2 - PK can be used to differentiate functional from organic colonic lesions at a cut-off point > 3 U/mL, with 93.33% sensitivity, 100% specificity, 100% PPV and 83.3% NPV. Also, plasma M2-PK can be used to discriminate between benign (colorectal polyp) and malignant colonic lesions (CRC) with a cut-off level of > 12 U/ml, with 100% sensitivity, 100% specificity, 100% PPV and 100% NPV. Also, In the present study there was a highly significant + ve correlations between Plasma M2 PK level and CEA as (r = 0.787, p value = 0.000) and their combination can give higher sensitivity and specificity. Conclusion Plasma M2-PK can differentiate between functional and organic colonic disorders as it is more elevated in organic than functional colonic disorders. Also, it is considered a promising rapid noninvasive biomarker for organic colonic disorders early detection and screening to reduce unnecessary endoscopic intervention.

The Diagnostic Value of Pyruvate Kinase Isoenzyme Type M2 for Biliary Tract Carcinoma. A Systematic Review and Meta-Analysis.

Growing evidence has shown that M2-PK is involved in cancer diagnosis and prognosis. The overall diagnostic accuracy of the pyruvate kinase isoenzyme type M2 (M2-PK) in biliary tract carcinoma (BTC) remains controversial. We performed a meta-analysis to evaluate the diagnostic value of M2-PK for BTC. The online PubMed, Cochrane, Web of Science, and Embase databases were searched for eligible studies published until August 8th, 2017. The Quality Assessment for Diagnostic Accuracy Studies 2 (QUADAS-2) was used to evaluate study quality. All statistical analyses were conducted with Stata 12.0. We included 7 studies from 5 articles with 410 patients with BTC and 438 controls. The pooled sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR), diagnostic odds ratio (DOR), and AUC for M2-PK in the diagnosis of BTC were 0.79 (95%CI 0.70-0.86), 0.81 (95%CI 0.71-0.88), 4.1 (95%CI 2.5-6.8), 0.26 (95%CI 0.16-0.41), 17.159 (95%CI 5.468-54.071), and 0.87 (95%CI 0.83-0.89), respectively. The same indicators assessed for CA19-9 were as follows: 0.70 (95%CI 0.62-0.77), 0.71 (95%CI 0.45-0.87), 2.38 (95%CI 1.2-4.73), 0.43 (95%CI 0.34-0.53), 6.28 (95%CI 2.4-16.44) and 0.73 (95%CI 0.69-0.77), respectively. Additionally, the diagnostic value of M2-PK varied based on characteristics of golden methods and different cut-off values. This meta-analysis showed that M2-PK had a better diagnostic accuracy for BTC compared with CA19-9, with moderate diagnostic performance. However, prospective studies are required to confirm its diagnostic value.

Abstract B19: LKB1-AMPK axis regulates the switch of Pyruvate Kinase M isoforms to tolerate nutritional stress

Abstract Background: Cancer cells are known to acquire a unique metabolic strategy of aerobic glycolysis (Warburg effect), where they rely more on glycolysis rather than oxidative phosphorylation, regardless of ample amount of oxygen. Aerobic glycolysis is presumed as an inefficient way to generate energy, yet believed to sustain equilibrium between catabolism and anabolism to achieve limitless proliferative competence in cancer cells. Conversely, glucose deprivation is an inevitable characteristic feature of tumors due to their rate of proliferation and poor vasculature. Thus, how cancer cells with an often encountered microenvironment within the tumor of hypoglycemia co-ordinate their growth and proliferates with an abnormal bio-energetics remains ambiguous. AMP-activated protein kinase (AMPK) is a serine/threonine protein kinase, known to get activated by sensing bioenergetic stress and remodels the core of cellular metabolism to preserve energy homeostasis. Recent studies have appreciated the critical importance of Liver kinase B1 (LKB1) – AMPK pathway in tumorigenesis, where the axis of LKB1-AMPK has been shown to provide tolerance to nutrient deprived cancer, but how AMPK contributes to cancer cell metabolism is largely unknown, except for the established role in enzymes, like PFK2 (phospho fructokinase 2); and more work is needed to define the role of AMPK in regulation of other essential glycolytic enzymes: Hexokinase (HK), Pyruvate Kinase (M2 PK) and Lactate Dehydrogenase (LDH), associated with aerobic glycolysis. Methodology: Lung cancer (H1299 & A549) and breast cancer (MCF-7 & MDA MB 231) cell lines were used in this study. qPCR and Immunoblotting analysis were performed to measure the gene expression status. Oligomeric status and the distribution of PKM isoforms was assessed using density gradient ultracentrifugation followed by immunoblotting. PKM1 and PKM2 knockdowns were generated by lentiviral transduction. Metabolites levels were assayed using enzyme coupled assays. Cell proliferation was assessed using MTB assay; and apoptosis measured by developing immunoblot for Cleaved PARP and Cleaved Caspase 3. Results: We show how activation of AMPK in cultured cancer cells through relative glucose deprivation (25, 5 & 1 mM) increased the relative activity of PKM, but not of HK and LDH. Further, we demonstrate AMPK facilitates PK activity by enhancing their subunit association, which is the consequential outcome of PKM isoform switch from a catalytically less active M2 isoform to more active M1 isoform. Other than the response in glucose deprivation, the role of AMPK in the PKM isoform switch is confirmed by experimenting with AMPK agonist, inhibitor and constitutively active mutant of AMPK in cell culture. We also show the coordinated axis between LKB1 and AMPK in mediating alternative splicing of PKM isoforms, where transfection of LKB1 cDNA was carried out in cell lines devoid of LKB1. We for the first time report the indirect involvement of AMPK in regulation of alternative splicing of PKM isoforms, independent of c-Myc. Moreover, we show either impairment of AMPK activation through pharmacological agents or knockdown of PKM1 isoform in cancer cells cultured in glucose deprived medium (5mM glucose), diminishes the rate of glucose uptake, lactate production and cellular pyruvate and ATP level, as a result of attenuated cell proliferation and enhanced onset of apoptosis. Together our findings suggest that AMPK –PKM1 axis enhances the aerobic glycolysis and favors cancer cells in culture by conserving the energy homeostasis to endure nutritional stress.” Conclusion: The study highlights the integral role of LKB1-AMPK axis in dynamically regulating the preferential expression of either PKM2 or PKM1 in response to nutritional status in lung cancer and breast cancer cell line models, drawing an inference with regard to real life tumor microenvironment. Citation Format: Gopinath Prakasam, Rameshwar N.K. Bamezai. LKB1-AMPK axis regulates the switch of Pyruvate Kinase M isoforms to tolerate nutritional stress. [abstract]. In: Proceedings of the AACR Special Conference: Metabolism and Cancer; Jun 7-10, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(1_Suppl):Abstract nr B19.

The Diagnostic Accuracy of the M2 Pyruvate Kinase Quick Stool Test--A Rapid Office Based Assay Test for the Detection of Colorectal Cancer.

BackgroundM2 pyruvate kinase (M2PK) is an oncoprotein secreted by colorectal cancers in stools. This the first report on the accuracy of a rapid stool test in the detection of colorectal cancer (CRC).ObjectiveTo determine the sensitivity, specificity and positive and negative predictive value of a rapid, point of care stool test M2 PK- the M2PK Quick.MethodsConsecutive cases of endoscopically diagnosed and histological proven CRC were recruited. Stools were collected by patients and tested with the immunochromatographic M2PK Quick Test (Schebo Biotech AC, Giessen, Germany). Controls were consecutively chosen from patients without any significant colorectal or gastrointestinal disease undergoing colonoscopy. CRC was staged according to the AJCC staging manual (7th Edition) and location of tumor defined as proximal or distal.ResultsThe sensitivity, specificity, positive predictive value, negative predictive value and overall accuracy were: 93%, 97.5%, 94.9%, 96.5% and 96.0% respectively. The positive predictive value for proximal tumors was significantly lower compared to distal tumors. No differences were seen between the different stages of the tumor.ConclusionsThe M2-PK Quick, rapid, point-of-care test is a highly accurate test in the detection of CRC. It is easy and convenient to perform and a useful diagnostic test for the detection of CRC in a clinical practice setting.

A systematic review and meta-analysis of the diagnostic accuracy of pyruvate kinase M2 isoenzymatic assay in diagnosing colorectal cancer.

BackgroundScreening programmes exist in many countries for colorectal cancer. In recent years, there has been a drive for a non-invasive screening marker of higher sensitivity and specificity. Stool-based pyruvate kinase isoenzyme M2 (M2-PK) is one such biomarker under investigation. The aim of this systematic review and meta-analysis is to determine the diagnostic accuracy, sensitivity and specificity of M2-PK as a screening tool in colorectal cancer.MethodsA literature search of Ovid Medline, EMBASE and Google Scholar was carried out. The search strategy was restricted to human subjects and studies published in English. Data on sensitivity and specificity were extracted and pooled. Statistical analysis was conducted using summary receiver operating characteristic (SROC) curve methodology.ResultsA total of eight studies were suitable for data synthesis and analysis. Our analysis showed a pooled sensitivity and specificity for M2-PK to be 79% (CI 73%–83%) and 80% (CI 73%–86%), respectively. The accuracy of M2-PK was 0.85(0.82–0.88).ConclusionFaecal M2-PK assay has a relatively good sensitivity and specificity and high accuracy for screening colorectal cancer.

INHIBITION OF EHRLICH ASCITES TUMOUR (EAT) CELLS PROLIFERATION THROUGH CHITOSAN-MEDIATED REGULATION OF ACTIVITY OF THE AKT PATHWAY

Isoenzyme M2 pyruvate kinase, which is a marker of cancer transformation, can take both tetramer (cytosol) and dimer (nucleus) forms. The former is responsible for ATP synthesis, and the latter demonstrates histone H1 kinase activity. Regulation of the expression of pyruvate kinase through which Akt controls the expression of genes involved in Ehrlich ascites tumour (EAT) cell proliferation, migration and death, also involves cross-talk with the other signalling pathways, transcription factors and co-regulatory proteins such as β-catenin and c-Myc. Treatment of EAT cells with chitosans significantly reduced their proliferation (by 45-60%), expression of nuclear β-catenin, c-Myc as well as cell migration. After 48–72 hours of treatment of the cell with oligochitosans, lower levels of p-Akt were detected. Simultaneously, decreased expression of isoenzyme M2 PK protein levels was observed. The dimeric form (nucleus) can participate in H1 histone phosphorylation, which contributes to increased EAT cell proliferation.

Comparison of fecal pyruvate kinase isoform M2 and calprotectin in acute diarrhea in hospitalized children.

Fecal concentrations of pyruvate kinase isoform M2 (M2-PK) and calprotectin (FC) serve as biomarkers of inflammation of gastrointestinal mucosa. The value of M2-PK in discriminating between patients with viral and bacterial acute diarrhea (AD) is currently unknown. We analyzed M2-PK and FC concentrations in fifty hospitalized children with AD (29 of which were caused by rotavirus and 21 by Salmonella enteritidis) as well as 32 healthy subjects. There was no difference in the areas under the receiver operating characteristic curves plotted for the two tests in differentiating rotaviral from bacterial AD. The sensitivity and specificity of M2-PK at optimal cut-off (20 U/g) were 75.9% and 71.4%, respectively. M2-PK and FC had similar values in distinguishing between children with AD caused by rotavirus and Salmonella enteritidis. The performance of both tests in hospitalized patients did not meet the needs of everyday clinical practice. Moreover, no advantage of fecal tests over the measurement of CRP was documented.

The value of expression of M2-PK and VEGF in patients with advanced gastric cancer.

Glycolytic pyruvate kinase isoenzyme type M2 (M2-PK) plays a key role in tumor metabolism and energy production. Vascular endothelial growth factor (VEGF) is critical in regulating angiogenesis which is an essential process required for tumor growth and metastasis. These two genes may function in accordance with tumor development. The purpose of this study was to investigate the relationship between the expression of M2-PK and VEGF, and their association with clinicopathological features in patients with advanced gastric cancer. Expression of M2-PK and VEGF were examined in 142 cases of paraffin-embedded tissue blocks from patients with advanced gastric cancer. M2-PK expression was found to strongly correlate with that of VEGF (r = 0.718). In addition, expression of M2-PK and VEGF correlates with tumor size (p = 0.0001, and p = 0.0017, respectively), depth of invasion (p = 0.0024, and p = 0.0261, respectively), and lymph node metastasis (p = 0.036, and p = 0.028, respectively). The high expression levels of M2-PK and VEGF may indicate poor prognosis in patients with advanced gastric cancer.

P166 Prospective evaluation of faecal tumour pyruvate kinase type M2 (M2-PK) in comparison to calprotectin in IBD patients

P166 Prospective evaluation of faecal tumour pyruvate kinase type M2 (M2-PK) in comparison to calprotectin in IBD patients

Sequential Combination of Serum Pyruvate Kinase Isoenzyme M2 and Colonoscopy-A Promising Screening Protocol for Colorectal Cancer Early Diagnosis

Background: Early diagnosis and treatment through mass screening is practical against colorectal cancer (CRC). Low compliance for current screening tests affects the effectiveness of CRC mass screening. An efficient screening protocol with high compliance is needed for CRC mass screening. Methods: Systematic searches were done through Medline and Cochrane Library databases - initial Medline searches were in February 2010 and additional searches in March/April 2010. Search terms included [(“Colorectal cancer” AND “screening program” AND “incidence”) OR (“Colorectal cancer” AND “screening program” AND “mortality”)] AND [“fecal occult blood test” OR “sigmoidoscopy” OR “colonoscopy” OR “Double-contrast barium enema”]. Results: This review explored the current CRC mass screening protocols to find a more efficient and practical mass screening protocol and problems suitable for further research. Considering the current economic crisis and limited available resources, combination of high risk factor questionnaire and immunochemical fecal occult blood test approach as primary CRC mass screening can currently be used as a risk stratification tool to identify high-risk populations from the community, especially for medically and economically underserved areas/countries before a new better test comes. Using serum Pyruvate Kinase Isoenzyme M2 (M2-PK) as primary and colonoscopy as secondary screening test sounds more efficient with higher compliance than current CRC mass screening protocols. Conclusion: Recommendations for CRC mass screening are suggested for each risk population based on risk stratification. Serum M2-PK- may be developed as a promising CRC primary mass screening test. Sequential combination of serum biomarker such as Pyruvate Kinase Isoenzyme M2 (M2-PK) and colonoscopy can be a promising CRC mass screening protocol.

Faecal pyruvate kinase isoenzyme type M2 for colorectal cancer screening: A meta-analysis

To present a critical discussion of the efficacy of the faecal pyruvate kinase isoenzyme type M2 (faecal M2-PK) test for colorectal cancer (CRC) screening based on the currently available studies. A literature search in PubMed and Embase was conducted using the following search terms: fecal Tumor M2-PK, faecal Tumour M2-PK, fecal M2-PK, faecal M2-PK, fecal pyruvate kinase, faecal pyruvate kinase, pyruvate kinase stool and M2-PK stool. Stool samples from 704 patients with CRC and from 11 412 healthy subjects have been investigated for faecal M2-PK concentrations in seventeen independent studies. The mean faecal M2-PK sensitivity was 80.3%; the specificity was 95.2%. Four studies compared faecal M2-PK head-to-head with guaiac-based faecal occult blood test (gFOBT). Faecal M2-PK demonstrated a sensitivity of 81.1%, whereas the gFOBT detected only 36.9% of the CRCs. Eight independent studies investigated the sensitivity of faecal M2-PK for adenoma (n = 554), with the following sensitivities: adenoma < 1 cm in diameter: 25%; adenoma > 1 cm: 44%; adenoma of unspecified diameter: 51%. In a direct comparison with gFOBT of adenoma > 1 cm in diameter, 47% tested positive with the faecal M2-PK test, whereas the gFOBT detected only 27%. We recommend faecal M2-PK as a routine test for CRC screening. Faecal M2-PK closes a gap in clinical practice because it detects bleeding and non-bleeding tumors and adenoma with high sensitivity and specificity.

In vivo factors influencing tumour M2-pyruvate kinase level in human pancreatic cancer cell lines

In tumour cells, the tetramer/dimer ratio of the pyruvate kinase isoenzyme type M2 (M2-PK) determines whether glucose carbons are degraded to lactate with production of energy (tetrameric form) or are channelled into synthetic processes (dimeric form). The influence of different tumour microenvironment conditions on the tetramer/dimer ratio of M2-PK and cell doublings were investigated in a non-metastatic and metastatic pancreatic cancer cell line. The metastatic Colo357 cells contained about fourfold more M2-PK protein and about 3.5-fold more dimeric M2-PK than the non-metastatic Panc-1 cells. In Colo357 cells hypoxia, glucose starvation as well as acidification induced an increase of the dimeric form of M2-PK, whereas in Panc-1 cells no effect on M2-PK was observed. Under hypoxia in Colo357 cells, the dimerization and inactivation of M2-PK results in an inhibition of cell proliferation, whereas under glucose starvation and acidification the dimerization of M2-PK allowed further cell doublings. M2-PK expression and the quaternary structure of M2-PK are influenced by the tumour metastatic potential. The quaternary structure of M2-PK may be differently affected by hypoxia, glucose starvation and acidification with severe consequences on cell doublings.

Pyruvate kinase type M2: A key regulator of the metabolic budget system in tumor cells

Cell proliferation only proceeds when metabolism is capable of providing a budget of metabolic intermediates that is adequate to ensure both energy regeneration and the synthesis of cell building blocks in sufficient amounts. In tumor cells, the glycolytic pyruvate kinase isoenzyme M2 (PKM2, M2-PK) determines whether glucose is converted to lactate for regeneration of energy (active tetrameric form, Warburg effect) or used for the synthesis of cell building blocks (nearly inactive dimeric form). This review discusses the regulation mechanisms of pyruvate kinase M2 expression by different transcription factors as well as the regulation of pyruvate kinase M2 activity by direct interaction with certain oncoproteins, tyrosine and serine phosphorylation, binding of phosphotyrosine peptides, association with other glycolytic and non glycolytic enzymes, the promyelocytic leukemia tumor suppressor protein, as well as metabolic intermediates. An intervention in the regulation mechanisms of the expression, activity and tetramer to dimer ratio of pyruvate kinase M2 has severe consequences for metabolism as well as proliferation and tumorigenic capacity of the cells which makes this enzyme a promising target for potential therapeutic approaches. The quantification of the dimeric form of pyruvate kinase M2 (Tumor M2-PK) in plasma and stool allows early detection of tumors and therapy control. Several different mechanisms may induce a translocation of pyruvate kinase M2 into the nucleus. The role of pyruvate kinase M2 in the nucleus is complex as witnessed by evidence of its effect both as pro-proliferative as well as pro-apoptotic stimuli.

Antigen Presentation by Dendritic Cells in Tumors Is Disrupted by Altered Metabolism that Involves Pyruvate Kinase M2 and Its Interaction with SOCS3

Dendritic cell (DC) function is negatively affected by tumors and tumor-derived factors, but little is known about the underlying mechanisms. Here, we show that intracellular SOCS3 in DCs binds to pyruvate kinase type M2 (M2-PK), which plays a critical role in ATP production through glycolysis. The interaction of SOCS3 with M2-PK reduced ATP production and impaired DC-based immunotherapy against tumors. Thus, SOCS3, which has been shown to be upregulated by tumor-derived factors, interacts with M2-PK to decrease ATP production, causing DC dysfunction. These dysfunctional DCs have a reduced ability to present antigens. Alteration of DC metabolism mediated by SOCS3 represents a novel mechanism for DC dysfunction in the tumor microenvironment.

Glucose transporter GLUT1 expression is an stage-independent predictor of clinical outcome in adrenocortical carcinoma

Owing to the rarity of adrenocortical carcinoma (ACC) no prognostic markers have been established beyond stage and resection status. Accelerated glycolysis is a characteristic feature of cancer cells and in a variety of tumour entities key factors in glucose metabolism like glucose transporter 1 and 3 (GLUT1 and -3), transketolase like-1 enzyme (TKTL1) and pyruvate kinase type M2 (M2-PK) are overexpressed and of prognostic value. Therefore, we investigated the role of these factors in ACC. Immunohistochemical analysis was performed on tissue microarrays of paraffin-embedded tissue samples from 167 ACCs, 15 adrenal adenomas and 4 normal adrenal glands. Expression was correlated with baseline parameters and clinical outcome. GLUT1 and -3 were expressed in 33 and 17% of ACC samples respectively, but in none of the benign tumours or normal adrenals glands. By contrast, TKTL1 and M2-PK were detectable in all benign tissues and the vast majority of ACCs. GLUT1 expression was strongly associated with prognosis in univariate and multivariate analysis (P<0.01), whereas GLUT3, TKTL1 and M2-PK did not correlate with clinical outcome. Patients with strong GLUT1 staining showed a considerably higher overall mortality (hazard ratio (HR) 6.34 (95% confidence interval 3.10-12.90) compared with patients with no GLUT1 staining. When analysing patients in their early stages and advanced disease separately, similar results were obtained. HR for survival was 5.31 (1.80-15.62) in patients with metastatic ACC and in patients after radical resection the HR for disease-free survival was 6.10 (2.16-16.94). In conclusion, GLUT1 is a highly promising stage-independent, prognostic marker in ACC.

The SUMO‐E3 ligase PIAS3 targets pyruvate kinase M2

Pyruvate kinase M2 (M2-PK) controls the rate-limiting step at the end of the glycolytic pathway in normal proliferating and tumor cells. Other functions of M2-PK in addition to its role in glycolysis are little understood. The aim of this study was to identify new cellular interaction partners of M2-PK in order to discover novel links between M2-PK and cellular functions. Here we show that the SUMO-E3 ligase protein PIAS3 (inhibitor of activated STAT3) physically interacts with M2-PK and its isoenzyme M1-PK. Moreover, we demonstrate that endogenous SUMO-1-M2-PK conjugates exist in mammalian cells. Furthermore, we show that transient expression of PIAS3 but not the RING domain mutant PIAS3 (C299S, H301A) is consistent with nuclear localization of M2-PK and PIAS3 and M2-PK partially co-localize in the nucleus of these cells. This study suggests a link between PIAS3 and nuclear pyruvate kinase.

Sensitivity and specificity of faecal tumour M2 pyruvate kinase for detection of colorectal adenomas in a large screening study

The measurement of faecal tumour M2 pyruvate kinase (tumour M2 PK) has been proposed as a novel approach for early detection of colorectal cancer (CRC). However, as regards the potential of the test to detect precursors to CRC, an issue that is highly relevant to estimate its use in reducing CRC incidence and mortality, the available evidence is scant and controversial. The aim of our study was to determine the performance characteristics of the tumour M2 PK test with respect to colorectal adenomas in the target population of screening. Among 1082 participants of screening colonoscopy in Germany, of whom 30% had any adenoma and 10% had an advanced adenoma, the median (interquartile range) tumour M2 PK level in the whole study population was 1.3 U ml−1 (0.3–3.3). At a cutoff value of 4 U ml−1, sensitivity was 22 and 23% for detection of advanced and other adenomas, respectively, whereas specificity was 82%. The area under the receiver-operating characteristics curve (95% confidence interval) was 0.54 (0.51–0.58) and 0.56 (0.52–0.59) for advanced and other adenomas, respectively. In conclusion, the tumour M2 PK test has only very limited potential to distinguish between people bearing precursors to CRC and people with no finding at colonoscopy.

Comparison of an Established Simple Office-Based Immunological FOBT With Fecal Tumor Pyruvate Kinase Type M2 (M2-PK) for Colorectal Cancer Screening: Prospective Multicenter Study

The immunological fecal occult blood test (IFOBT) has established itself as a more precise marker for colorectal cancer (CRC) screening than traditional guaiac-based FOBT. The simpler, cheaper, and more convenient newer office-based IFOBTs have been validated for diagnosing CRC. Dimeric isoenzyme of pyruvate kinase, M2-PK, expressed by tumor cells, has as well been proposed as a screening tool for CRC. This is the first study comparing fecal M2-PK as a screening biomarker for CRC against previously evaluated office-based IFOBT and colonoscopy. Six hundred forty consecutive subjects (symptomatic, as well as for CRC screening) referred for colonoscopy for various indications across five centers in Germany provided the stool samples for performing M2-PK and an immunochemical FOB strip test. The IFOBT used was a rapid immunochromatographic assay for detection of fecal hemoglobin. For M2-PK, a commercially available sandwich enzyme-linked immunosorbent assay (ELISA) was used. The M2-PK test needs 6 h, while the office-based test can be read in just 10 min and is five times cheaper. Office-based IFOBT had sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and positive and negative likelihood ratios (LR) of 64.5, 96.3, 72.0, 94.9, 17.5, and 0.4 for diagnosing colorectal neoplasia (CRN), while the above performance characteristics for M2-PK at a cutoff value of 4 U/mL were 72.4, 73.8, 29.0, 94.8, 2.8, and 0.8 respectively. This office-based IFOBT was found to have significantly higher specificity, PPV, and positive LR as compared with M2-PK. IFOBT proved to be a convenient, noncumbersome, quick, and cheap tool in patients with above-average risk for detection of CRN.

Isotype‐specific inhibitors of the glycolytic key regulator pyruvate kinase subtype M2 moderately decelerate tumor cell proliferation

Tumor cells express the glycolytic regulator pyruvate kinase subtype M2 (M2-PK), which can occur in a tetrameric form with high affinity to its substrate phosphoenolpyruvate (PEP) and a dimeric form with a low PEP affinity. The transition between both conformations contributes to the control of glycolysis and is important for tumor cell proliferation and survival. Here we targeted M2-PK by synthetic peptide aptamers, which specifically bind to M2-PK and shift the isoenzyme into its low affinity dimeric conformation. The aptamer-induced dimerization and inactivation of M2-PK led to a significant decrease in the PK mass-action ratio as well as ATP:ADP ratio in the target cells. Furthermore, the expression of M2-PK-binding peptide aptamers moderately reduced the growth of immortalized NIH3T3 cell populations by decelerating cell proliferation, but without affecting apoptotic cell death. Moreover, the M2-PK-binding peptide aptamers also reduced the proliferation rate of human U-2 OS osteosarcoma cells. In the present study, we developed the first specific inhibitors of the pyruvate kinase isoenzyme type M2 and present evidence that these inhibitors moderately decelerate tumor cell proliferation.