Tumor M2-PK
Encyclopedia
Tumor M2-PK is a synonym for the dimeric form of the pyruvate kinase
isoenzyme type M2 (PKM2
), a key enzyme within tumor metabolism
. Tumor M2-PK can be elevated in many tumor types, rather than being an organ-specific tumor marker
such as PSA
. Increased stool (fecal) levels are being investigated as a method of screening for colorectal tumors
(= bowel cancer), and EDTA plasma
levels are undergoing testing for possible application in the follow-up of various cancer
s.
Sandwich ELISA’s
based on two monoclonal antibodies
which specifically recognize Tumor M2-PK (the dimeric form of M2-PK) are available for the quantification of Tumor M2-PK in stool and EDTA-plasma samples respectively. As a biomarker, the amount of Tumor M2-PK in stool and EDTA-plasma reflects the specific metabolic status of the tumors.
, is a potential tumor marker for colorectal cancer
. When measured in feces
with a cutoff value of 4 U/ml, its sensitivity
has been estimated to be 85% (with a 95% confidence interval
of 65 to 96%) for colon cancer and 56% (confidence interval 41–74%) for rectal cancer. Its specificity
is estimated to be 79% (95% confidence interval 76–81%).
Most people are more willing to accept non-invasive preventive medical check-ups. Therefore, the measurement of Tumor M2-PK in stool samples, with follow-up by colonoscopy
to clarify the Tumor M2-PK positive results, may prove to be an advance in the early detection of colorectal carcinomas. More research is needed before the test can be recommended as a screening procedure.
.
The combination of Tumor M2-PK with the appropriate classical tumor marker, such as CEA
for bowel cancer, CA 19-9 for pancreatic cancer and CA 72-4 for gastric cancer, significantly increases the sensitivity to detect various cancers.
An important application of the Tumor M2-PK test in EDTA-plasma is for follow-up during tumor therapy, to monitor the success or failure of the chosen treatment, as well as predicting the chances of a “cure” and survival.
If Tumor M2-PK levels decrease during therapy and then remain low after therapy it points towards successful treatment. An increase in the Tumor M2-PK values during or after therapy points towards relapse and/or metastasis.
Increased Tumor M2-PK values can sometimes also occur in severe inflammatory diseases
, which must be excluded by differential diagnosis.
, the dephosphorylation of phosphoenolpyruvate to pyruvate and is responsible for net energy production within the glycolytic pathway. Depending upon the different metabolic functions of the tissues, different isoenzymes of pyruvate kinase are expressed.
M2-PK (PKM2
) is the predominant pyruvate kinase isoform in proliferating cells, such as fibroblasts, embryonic cells and adult stem cells and most human tissue, including lung, bladder, kidney and thymus; M2-PK is upgregulated in many human tumors.
M2-PK can occur in two different forms in proliferating cells:
The tetrameric form of M2-PK has a high affinity to its substrate, phosphoenolpyruvate (PEP), and is highly active at physiological PEP concentrations. Furthermore, the tetrameric form of M2-PK is associated with several other glycolytic enzymes within the so-called glycolytic enzyme complex. Due to the close proximity of the enzymes, the association within the glycolytic enzyme complex leads to a highly effective conversion of glucose to lactate.
When M2-PK is mainly in the highly active tetrameric form, which is the case in most normal cells, glucose is mostly converted to lactate, with the attendant production of energy.
In contrast, the dimeric form of M2-PK has a low affinity for phosphoenolpyruvate, being nearly inactive at physiological PEP concentrations. When M2-PK is mainly in the dimeric form, which is the case in tumor cells, all phosphometabolites above pyruvate kinase accumulate and are channelled into synthetic processes which branch off from glycolytic intermediates, such as nucleic acids, phospholipids and amino acids, important cell building blocks for highly proliferating cells such as tumor cells.
As a consequence of the key position of pyruvate kinase within glycolysis, the tetramer : dimer ratio of M2-PK determines whether glucose carbons are converted to pyruvate and lactate, along with the production of energy (tetrameric form), or channelled into synthetic processes (dimeric form). In tumor cells M2-PK is mainly in the dimeric form. Therefore, the dimeric form of M2-PK has been termed Tumor M2-PK.
The dimerization of M2-PK in tumor cells is induced by the direct interaction of M2-PK with different oncoproteins.
However, the tetramer : dimer ratio of M2-PK is not constant.
Oxygen starvation or highly accumulated glycolytic intermediates, such as fructose 1,6-bisphosphate (fructose 1,6-P2) or the amino acid serine, induce the reassociation of the dimeric form of M2-PK to the tetrameric form. Consequently, due to the activation of M2-PK, glucose is converted to pyruvate and lactate under the production of energy until the fructose 1,6-P2 levels drop below a certain threshold value, which allows the dissociation of the tetrameric form of M2-PK to the dimeric form. Thereafter, the cycle of oscillation starts again when the fructose 1,6-P2 levels reach a certain upper threshold value which induces the tetramerization of M2-PK.
When M2-PK is mainly in the less active dimeric form, energy is produced by the degradation of the amino acid glutamine to aspartate, pyruvate and lactate, which is termed glutaminolysis
.
In tumor cells the increased rate of lactate production in the presence of oxygen is termed the Warburg effect.
patients, prone to develop cancer. Results show that despite the presence of mutations in the inter-subunit contact domain, the K422R and H391Y mutant proteins maintained their homotetrameric structure, similar to the wild-type protein, but showed a loss of activity of 75 and 20%, respectively. Interestingly, H391Y showed a 6-fold increase in affinity for its substrate phosphoenolpyruvate and behaved like a non-allosteric protein with compromised cooperative binding. However, the affinity for phosphoenolpyruvate was lost significantly in K422R. Unlike K422R, H391Y showed enhanced thermal stability, stability over a range of pH values, a lesser effect of the allosteric inhibitor Phe, and resistance toward structural alteration upon binding of the activator (fructose 1,6-bisphosphate) and inhibitor (Phe). Both mutants showed a slight shift in the pH optimum from 7.4 to 7.0. The co-expression of homotetrameric wild type and mutant PKM2 in the cellular milieu resulting in the interaction between the two at the monomer level was substantiated further by in vitro experiments. The cross-monomer interaction significantly altered the oligomeric state of PKM2 by favoring dimerisation and heterotetramerization. In silico study provided an added support in showing that hetero-oligomerization was energetically favorable. The hetero-oligomeric populations of PKM2 showed altered activity and affinity, and their expression resulted in an increased growth rate of Escherichia coli as well as mammalian cells, along with an increased rate of polyploidy. These features are known to be essential to tumor progression.
Pyruvate kinase
Pyruvate kinase is an enzyme involved in glycolysis. It catalyzes the transfer of a phosphate group from phosphoenolpyruvate to ADP, yielding one molecule of pyruvate and one molecule of ATP.-Reaction:The reaction with pyruvate kinase:...
isoenzyme type M2 (PKM2
PKM2
Pyruvate kinase isozymes M1/M2 also known as pyruvate kinase muscle isozyme , pyruvate kinase type K, cytosolic thyroid hormone-binding protein , thyroid hormone-binding protein 1 , or opa-interacting protein 3 is an enzyme that in humans is encoded by the PKM2 gene.PKM2 is an isoenzyme of the...
), a key enzyme within tumor metabolism
Tumor metabolome
The term tumor metabolome describes the characteristic metabolic phenotype of tumor cells.Comparison of the metabolic characteristics of different tumor cell lines and tumors revealed that during tumorigenesis, the same alterations always occur in distinct metabolic parameters regardless of the...
. Tumor M2-PK can be elevated in many tumor types, rather than being an organ-specific tumor marker
Tumor marker
A tumor marker is a substance found in the blood, urine, or body tissues that can be elevated in cancer, among other tissue types. There are many different tumor markers, each indicative of a particular disease process, and they are used in oncology to help detect the presence of cancer...
such as PSA
Prostate specific antigen
Prostate-specific antigen also known as gamma-seminoprotein or kallikrein-3 is a glycoprotein that in humans is encoded by the KLK3 gene. KLK3 is a member of the kallikrein-related peptidase family that are secreted by the epithelial cells of the prostate gland...
. Increased stool (fecal) levels are being investigated as a method of screening for colorectal tumors
Colorectal cancer
Colorectal cancer, commonly known as bowel cancer, is a cancer caused by uncontrolled cell growth , in the colon, rectum, or vermiform appendix. Colorectal cancer is clinically distinct from anal cancer, which affects the anus....
(= bowel cancer), and EDTA plasma
Blood plasma
Blood plasma is the straw-colored liquid component of blood in which the blood cells in whole blood are normally suspended. It makes up about 55% of the total blood volume. It is the intravascular fluid part of extracellular fluid...
levels are undergoing testing for possible application in the follow-up of various cancer
Cancer
Cancer , known medically as a malignant neoplasm, is a large group of different diseases, all involving unregulated cell growth. In cancer, cells divide and grow uncontrollably, forming malignant tumors, and invade nearby parts of the body. The cancer may also spread to more distant parts of the...
s.
Sandwich ELISA’s
ELISA
Enzyme-linked immunosorbent assay , is a popular format of a "wet-lab" type analytic biochemistry assay that uses one sub-type of heterogeneous, solid-phase enzyme immunoassay to detect the presence of a substance in a liquid sample."Wet lab" analytic biochemistry assays involves detection of an...
based on two monoclonal antibodies
Monoclonal antibodies
Monoclonal antibodies are monospecific antibodies that are the same because they are made by identical immune cells that are all clones of a unique parent cell....
which specifically recognize Tumor M2-PK (the dimeric form of M2-PK) are available for the quantification of Tumor M2-PK in stool and EDTA-plasma samples respectively. As a biomarker, the amount of Tumor M2-PK in stool and EDTA-plasma reflects the specific metabolic status of the tumors.
Early detection of colorectal tumors and polyps
M2-PK, as measured in fecesFeces
Feces, faeces, or fæces is a waste product from an animal's digestive tract expelled through the anus or cloaca during defecation.-Etymology:...
, is a potential tumor marker for colorectal cancer
Colorectal cancer
Colorectal cancer, commonly known as bowel cancer, is a cancer caused by uncontrolled cell growth , in the colon, rectum, or vermiform appendix. Colorectal cancer is clinically distinct from anal cancer, which affects the anus....
. When measured in feces
Feces
Feces, faeces, or fæces is a waste product from an animal's digestive tract expelled through the anus or cloaca during defecation.-Etymology:...
with a cutoff value of 4 U/ml, its sensitivity
Sensitivity and specificity
Sensitivity and specificity are statistical measures of the performance of a binary classification test, also known in statistics as classification function. Sensitivity measures the proportion of actual positives which are correctly identified as such Sensitivity and specificity are statistical...
has been estimated to be 85% (with a 95% confidence interval
Confidence interval
In statistics, a confidence interval is a particular kind of interval estimate of a population parameter and is used to indicate the reliability of an estimate. It is an observed interval , in principle different from sample to sample, that frequently includes the parameter of interest, if the...
of 65 to 96%) for colon cancer and 56% (confidence interval 41–74%) for rectal cancer. Its specificity
Sensitivity and specificity
Sensitivity and specificity are statistical measures of the performance of a binary classification test, also known in statistics as classification function. Sensitivity measures the proportion of actual positives which are correctly identified as such Sensitivity and specificity are statistical...
is estimated to be 79% (95% confidence interval 76–81%).
Most people are more willing to accept non-invasive preventive medical check-ups. Therefore, the measurement of Tumor M2-PK in stool samples, with follow-up by colonoscopy
Colonoscopy
Colonoscopy is the endoscopic examination of the large bowel and the distal part of the small bowel with a CCD camera or a fiber optic camera on a flexible tube passed through the anus. It may provide a visual diagnosis and grants the opportunity for biopsy or removal of suspected...
to clarify the Tumor M2-PK positive results, may prove to be an advance in the early detection of colorectal carcinomas. More research is needed before the test can be recommended as a screening procedure.
Cancer follow-up
Studies from various international working groups have revealed a significantly increased amount of Tumor M2-PK in EDTA-plasma samples of patients with renal, lung, breast, cervical and gastrointestinal tumors (oesophagus, stomach, pancreas, colon, rectum), as well as melanoma (= skin cancer), which correlated with the tumor stageCancer staging
The stage of a cancer is a description of the extent the cancer has spread. The stage often takes into account the size of a tumor, how deeply it has penetrated, whether it has invaded adjacent organs, how many lymph nodes it has metastasized to , and whether it has spread to distant organs...
.
The combination of Tumor M2-PK with the appropriate classical tumor marker, such as CEA
Carcinoembryonic antigen
Carcinoembryonic antigen is a glycoprotein involved in cell adhesion. It is normally produced during fetal development, but the production of CEA stops before birth. Therefore, it is not usually present in the blood of healthy adults, although levels are raised in heavy smokers...
for bowel cancer, CA 19-9 for pancreatic cancer and CA 72-4 for gastric cancer, significantly increases the sensitivity to detect various cancers.
An important application of the Tumor M2-PK test in EDTA-plasma is for follow-up during tumor therapy, to monitor the success or failure of the chosen treatment, as well as predicting the chances of a “cure” and survival.
If Tumor M2-PK levels decrease during therapy and then remain low after therapy it points towards successful treatment. An increase in the Tumor M2-PK values during or after therapy points towards relapse and/or metastasis.
Increased Tumor M2-PK values can sometimes also occur in severe inflammatory diseases
Inflammation
Inflammation is part of the complex biological response of vascular tissues to harmful stimuli, such as pathogens, damaged cells, or irritants. Inflammation is a protective attempt by the organism to remove the injurious stimuli and to initiate the healing process...
, which must be excluded by differential diagnosis.
Tetrameric and dimeric PKM2
Pyruvate kinase catalyzes the last step within the glycolytic sequenceGlycolysis
Glycolysis is the metabolic pathway that converts glucose C6H12O6, into pyruvate, CH3COCOO− + H+...
, the dephosphorylation of phosphoenolpyruvate to pyruvate and is responsible for net energy production within the glycolytic pathway. Depending upon the different metabolic functions of the tissues, different isoenzymes of pyruvate kinase are expressed.
M2-PK (PKM2
PKM2
Pyruvate kinase isozymes M1/M2 also known as pyruvate kinase muscle isozyme , pyruvate kinase type K, cytosolic thyroid hormone-binding protein , thyroid hormone-binding protein 1 , or opa-interacting protein 3 is an enzyme that in humans is encoded by the PKM2 gene.PKM2 is an isoenzyme of the...
) is the predominant pyruvate kinase isoform in proliferating cells, such as fibroblasts, embryonic cells and adult stem cells and most human tissue, including lung, bladder, kidney and thymus; M2-PK is upgregulated in many human tumors.
M2-PK can occur in two different forms in proliferating cells:
- a tetrameric form, which consists of four subunits
- a dimeric form, consisting of two subunits.
The tetrameric form of M2-PK has a high affinity to its substrate, phosphoenolpyruvate (PEP), and is highly active at physiological PEP concentrations. Furthermore, the tetrameric form of M2-PK is associated with several other glycolytic enzymes within the so-called glycolytic enzyme complex. Due to the close proximity of the enzymes, the association within the glycolytic enzyme complex leads to a highly effective conversion of glucose to lactate.
When M2-PK is mainly in the highly active tetrameric form, which is the case in most normal cells, glucose is mostly converted to lactate, with the attendant production of energy.
In contrast, the dimeric form of M2-PK has a low affinity for phosphoenolpyruvate, being nearly inactive at physiological PEP concentrations. When M2-PK is mainly in the dimeric form, which is the case in tumor cells, all phosphometabolites above pyruvate kinase accumulate and are channelled into synthetic processes which branch off from glycolytic intermediates, such as nucleic acids, phospholipids and amino acids, important cell building blocks for highly proliferating cells such as tumor cells.
The dimerization of M2-PK in tumor cells is induced by the direct interaction of M2-PK with different oncoproteins.
However, the tetramer : dimer ratio of M2-PK is not constant.
Oxygen starvation or highly accumulated glycolytic intermediates, such as fructose 1,6-bisphosphate (fructose 1,6-P2) or the amino acid serine, induce the reassociation of the dimeric form of M2-PK to the tetrameric form. Consequently, due to the activation of M2-PK, glucose is converted to pyruvate and lactate under the production of energy until the fructose 1,6-P2 levels drop below a certain threshold value, which allows the dissociation of the tetrameric form of M2-PK to the dimeric form. Thereafter, the cycle of oscillation starts again when the fructose 1,6-P2 levels reach a certain upper threshold value which induces the tetramerization of M2-PK.
When M2-PK is mainly in the less active dimeric form, energy is produced by the degradation of the amino acid glutamine to aspartate, pyruvate and lactate, which is termed glutaminolysis
Glutaminolysis
Glutaminolysis is a series of biochemical reactions by which the amino acid glutamine is degraded to glutamate, aspartate, CO2, pyruvate, lactate, alanine and citrate.- The glutaminolytic pathway:...
.
In tumor cells the increased rate of lactate production in the presence of oxygen is termed the Warburg effect.
Mutations
For the first time pyruvate kinase M2 enzyme was reported with two missense mutations, H391Y and K422R, found in cells from Bloom syndromeBloom syndrome
Bloom's syndrome , also known as Bloom–Torre–Machacek syndrome, is a rare autosomal recessive chromosomal disorder characterized by a high frequency of breaks and rearrangements in an affected person's chromosomes. The condition was discovered and first described by dermatologist Dr...
patients, prone to develop cancer. Results show that despite the presence of mutations in the inter-subunit contact domain, the K422R and H391Y mutant proteins maintained their homotetrameric structure, similar to the wild-type protein, but showed a loss of activity of 75 and 20%, respectively. Interestingly, H391Y showed a 6-fold increase in affinity for its substrate phosphoenolpyruvate and behaved like a non-allosteric protein with compromised cooperative binding. However, the affinity for phosphoenolpyruvate was lost significantly in K422R. Unlike K422R, H391Y showed enhanced thermal stability, stability over a range of pH values, a lesser effect of the allosteric inhibitor Phe, and resistance toward structural alteration upon binding of the activator (fructose 1,6-bisphosphate) and inhibitor (Phe). Both mutants showed a slight shift in the pH optimum from 7.4 to 7.0. The co-expression of homotetrameric wild type and mutant PKM2 in the cellular milieu resulting in the interaction between the two at the monomer level was substantiated further by in vitro experiments. The cross-monomer interaction significantly altered the oligomeric state of PKM2 by favoring dimerisation and heterotetramerization. In silico study provided an added support in showing that hetero-oligomerization was energetically favorable. The hetero-oligomeric populations of PKM2 showed altered activity and affinity, and their expression resulted in an increased growth rate of Escherichia coli as well as mammalian cells, along with an increased rate of polyploidy. These features are known to be essential to tumor progression.
Stool
- Koss K, Maxton D, Jankowski JAZ. The potential use of fecal dimeric M2 pyruvate kinase (Tumor M2-PK) in screening for colorectal cancer (CRC). Abstract from Digestive Disease Week, May 2005; Chicago, USA.
- Mc Loughlin R, Shiel E, Sebastian S, Ryan B, O´Connor HJ, O´Morain C. Tumor M2-PK, a novel screening tool for colorectal cancer. Abstract from Digestive Disease Week, May 2005, Chicago/USA