Chapter 32 Circulating Tumor Markers Basic Concepts and
Chapter 32: Circulating Tumor Markers: Basic Concepts and Clinical Applications By Christopher R. Mc. Cudden, Monte S. Willis Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Introduction • Cancer – 2 nd leading cause of death in developed countries – Accounts for more than 2. 7 million deaths annually – Is uncontrolled growth of cells that can develop into a tumor & spread to other areas of body – Formation & spreading (metastasis) of tumors is caused by a complex combination of inherited & acquired genetic mutations. – In formation, mutations include activation of growth factors & oncogenes & inhibition of apoptosis, tumor suppressor, & cell cycle regulation genes. – Cancer is staged based on tumor size, histology, regional lymph node involvement, & presence of metastasis; 4 stages (I–IV). Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Introduction (cont’d) • Cancer staging and progression Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Types of Tumor Markers • Function and Origin – Detect & monitor cancer – Produced by tumor directly or as an effect of tumor on tissue • Types – Enzymes: levels of certain enzymes correlate with tumor burden – Serum proteins: 2 -microglobulin & immunoglobulins – Hormones & metabolites: specific markers of secreting tumors – Oncofetal antigens: carcinoembryonic, alpha-fetoprotein – Receptors: non-serologic; estrogen & progesterone receptors Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Types of Tumor Markers (cont’d) • Timeline of tumor marker use Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Applications of Tumor Marker Detection • Screening – Most tumor markers are found in normal cells, not just cancer cells. – Therefore, screening asymptomatic populations would result in detection of false positives, causing undue alarm & cost. – Few tumor markers are used to screen populations. – Susceptibility to breast, ovarian, & colon cancer can be determined by identifying germline mutations in patients with a family history of these diseases. – Breast & ovarian cancers are associated with germline BRCA 1 & BRCA 2 mutations, colon cancer with adenomatous polyposis coli gene (APC). Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Applications of Tumor Marker Detection (cont’d) • Prognosis – Tumor marker concentration gradually increases with tumor progression, reaching highest levels when tumors metastasize. – Tumor marker levels at diagnosis can reflect presence of malignancy & aggressiveness of tumor & help predict outcome. • Monitoring Therapy Effectiveness & Disease Recurrence – After surgical resection, radiation, or chemotherapy, tumor markers are observed. – Effective therapy can result in decrease in tumor markers. – Appearance of tumor markers after effective therapy can be used as a highly sensitive marker of recurrence. Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Laboratory Considerations for Tumor Marker Measurement • Multiple tests should be performed, using same commercial kits. • Lack of standardization • Serially evaluate tumor markers, because they increase with time, whereas high normal values will not. Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Methods • Standardization found for other common clinical assays generally does not exist for cancer assays. • Comparisons of results from different assays for a single patient can be treacherous due to differences in: – Antibody specificity – Analyte heterogeneity – Assay design – Lack of standard reference material – Calibration & kinetics – Variation in reference ranges Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Methods (cont’d) • Immunoassays – Most commonly used method to measure tumor markers – Have many advantages, including ability to automate testing – Factors in interpreting tumor marker immunoassays: linearity, hook effect, & heterophile antibodies – Linearity • Linear range: range of analyte concentrations in which a linear relationship exists between analyte & signal • Measured by analyzing specimens spanning reportable range Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Methods (cont’d) • Immunoassays – Hook effect • Falsely low measurements as a result of excessively high tumor marker concentrations • Capture & label antibodies are saturated, resulting in a lack of a “sandwich” formation, resulting in decrease in signal. • Samples exceeding linear range should be diluted & retested. – Heterophile antibodies • Circulating antibodies against animal immunoglobulins can cause significant interference in immunoassays. • Occur in patients given mouse monoclonal antibodies Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Methods (cont’d) • High-Performance Liquid Chromatography (HPLC) – Most widely used method to detect catecholamines & their metabolites in plasma & urine – Analytes of interest are separated from plasma or urine, run over a column, & separated by physical characteristics. – Used to detect neuroblastoma, pheochromocytoma, carcinoid tumors • Immunohistochemistry (IHC) – Tests tumor markers that are detected directly within solid tissue – Slice of tissue is placed on glass slide & incubated with specific antibodies in solution to detect presence of antigens. Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Methods (cont’d) • Enzyme Assays – Detection of elevated circulating enzymes generally cannot be used to identify a specific tumor or site of tumor. – Exception: prostate specific antigen (PSA), found exclusively in diseased & benign prostate glands – Before use of immunoassays & oncofetal antigens was common, enzyme detection was widely used. – Examples of enzymes used as tumor markers: alkaline phosphatase (bone, liver, leukemia, sarcoma), creatine kinase. BB (prostate, small-cell lung, breast, colon, ovarian), lactate dehydrogenase (liver, lymphomas, leukemia), & PSA (prostate) Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Frequently Ordered Tumor Markers • Alpha-Fetoprotein (AFP) – Introduction/Description • An abundant serum protein synthesized by fetal liver & reexpressed in certain types of tumors • Often elevated in patients with hepatocellular carcinoma & germ cell tumors – Regulation/Physiology • A 70 -kd glycoprotein related to albumin; normally functions as a transport protein & is involved in regulating oncotic pressure in fetus • Upper limit of normal for serum AFP: ~15 ng/m. L in adults Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Frequently Ordered Tumor Markers (cont’d) • Alpha-Fetoprotein (AFP) – Clinical Application/Interpretation • Used for diagnosis, staging, prognosis, & treatment monitoring of hepatocellular carcinoma • Also used for classification & monitoring therapy for testicular cancer & for tumor staging – Methodology • Measured by a variety of immunoassays Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Frequently Ordered Tumor Markers (cont’d) • Cancer Antigen 125 (CA-125) – Introduction/Description • May be useful for detecting ovarian tumors at an early stage & for monitoring treatments without surgical restaging – Regulation/Physiology • Expressed in ovary, other tissues of müllerian duct origin, & ovarian carcinoma cells – Clinical Application/Interpretation • Only clinically accepted serologic marker of ovarian cancer – Methodology • Immunoassays using “OC 125” & “M 11” antibodies Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Frequently Ordered Tumor Markers (cont’d) • Carcinoembryonic Antigen (CEA) – Introduction/Description • Discovered in 1960 s; prototypical example of oncofetal antigen • Expressed during development & re-expressed in tumors • Most widely used tumor marker for colorectal cancer; also elevated in lung, breast, & GI tumors – Regulation/Physiology • Large heterogenous glycoprotein; molecular weight: ~200 k. Da • Is involved in apoptosis, immunity, & cell adhesion Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Frequently Ordered Tumor Markers (cont’d) • Carcinoembryonic Antigen (CEA) – Clinical Application/Interpretation • A tumor marker for colorectal cancer • Used for prognosis, post-surgery surveillance, & to monitor response to chemotherapy – Methodology • Historically used polyclonal antibodies; now uses monoclonal anti-CEA antibodies • Due to high heterogeneity of CEA, it is essential that same assay be used for serial monitoring. Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Frequently Ordered Tumor Markers (cont’d) • Human Chorionic Gonadotropin (h. CG) – Introduction/Description • A dimeric hormone secreted by trophoblasts in placenta to maintain corpus luteum during pregnancy – Regulation/Physiology • A 45 -kd glycoprotein consisting of alpha & beta subunits – Clinical Application/Interpretation • Prognosis of ovarian cancer, diagnosis of testicular cancer, most useful marker for gestational trophoblastic diseases – Methodology • Immunoassays w/ monoclonal capture, tracer antibodies Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Frequently Ordered Tumor Markers (cont’d) • Prostate Specific Antigen (PSA) – Introduction/Description • A 28 -kd glycoprotein produced only in epithelial cells of acini & in prostatic ducts • Regulates seminal fluid viscosity • Dissolves cervical mucous cap, allowing sperm to enter – Regulation/Physiology • Low levels of PSA can be detected in serum of healthy men. • Two major forms found circulating in blood: free & complexed Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
Frequently Ordered Tumor Markers (cont’d) • Prostate Specific Antigen (PSA) – Clinical Application/Interpretation • Annual screening of prostate cancer in men over 50 years old & in younger men at high risk (family history) • Normal range for PSA: <4 ng/m. L • Factors to take into account when testing for PSA: age, PSA velocity, free PSA/total PSA ratios – Methodology • Measured by immunoassay using enzyme, fluorescence, or chemiluminescence on an automated immunoassay platform Copyright © 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins
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