Tumor Biomarkers

1. Tumor Markers

2. Table of Contents
Types of tumor
markers
Application of tumor
markers detection
Frequently ordered
tumor markers
Methodology of
testing
01 02
04
03

3. Introduction
“ Cancer refers to the uncontrolled growth of cells that can develop
into a solid mass or tumor and spread to other areas of the body”

4. Types of tumor
markers
01

5. Types of tumor markers
● Serum proteins
● Antigens
● Hormones
● Receptors
● Enzymes
● Metabolites

6. Serum proteins
■ Beta2-Microglobulin
● Found on the surface of all nucleated cells.
● Used as a nonspecific marker of the high cell turnover that is often
observed in tumors.
■ Immunoglobulins
● Specific measure of plasma cell production of monoclonal proteins
observed in hematologic malignancies such as multiple myeloma.

7. Antigens
■ Oncofetal antigens
● Examples: carcinoembryonic antigen (CEA) and Alpha-fetoprotein
(AFP).
● Expressed transiently during normal development and are then
turned on again in the formation tumors.
■ Monoclonal defined antigens
● Directed toward specific carbohydrate or cancer antigens and are
best used for monitoring treatment of tumor that secrete these
epitopes
● Examples: CA 15-3, CA 27-29, CA 19-9, CA-125.

8. Hormones
● Widely used as specific markers of secreting tumors.
● Valuable in diagnosing:
○ Neuroblastomas
○ Pituitary tumor
○ Adrenal adenomas.

9. Receptors
● Used to classify tumors for therapy.
● Examples: estrogen and progesterone receptors.
○ When solid tumor biopsies are positive for estrogen and
progesterone markers, tamoxifen chemotherapy is more
likely to be effective.

10. Enzymes
■ Elevated non-specifically in tumors.
■ Enzyme levels tend to correlate with tumor burden
■ Useful for monitoring the success of therapy.

11. Ideal tumor marker characteristics
Absent in healthy
individuals
Specific Readily detectable
in body fluids

12. Application of tumor
markers detection
02

13. Timeline of tumor marker use
Screening Diagnosis Prognosis Monitoring
treatment
Detection of
recurrence
• PSA
• AFP
• Metanephrines
• HVA/VMA
• Prolactin
• PTH
• Chromogranin A
• Cortisol
• ACTH
• β2-microglobulin
• CA 125
• CEA
• LD
• Her-2/neu
• ER
• PR
• CA 125
• CA 19-9
• CEA
• AFP
• hCG
• PSA
• SPE
• CA 15-3
• CA 125
• CEA
• AFP
• hCG
• PSA

14. Screening
● Tumor markers are not used to screen asymptomatic populations except PSA.
● A few tumor markers are used for screening in populations with high incidence
( e.g AFP).

16. Prognosis
● Concentration of tumor markers is increases with tumor progression.
● Highest levels is reached when tumors metastasize.
● High serum tumor markers at diagnosis might indicate the presence of
malignancy and possible metastasis associated with a poorer prognosis.

17. Monitoring of therapy and Recurrence
● By routine serial follow-up after:
○ surgical resection.
○ radiation.
○ chemotherapy.

18. Testing methods
03

19. Testing methods
Enzyme assay
Immunoassay
High-performance
liquid
chromatography
Immunohistochemistry

20. ● Most commonly used method to measure tumor markers .
● Important factors in interpreting tumor marker immunoassays:
○ Linearity of the assay.
○ Hook effect.
○ Heterophile Antibodies.
Immunoassay

21. Linearity
§ The linear range is the range of analyte concentrations in which a linear relationship
exists between the analyte and signal.
§ Linearity is determined by analyzing (in replicates) specimens spanning the reportable
range.

22. High-dose hook effect
• When analyte concentrations exceed the analytical range
excessively, both the capture and label antibodies can be
saturated, resulting in a lack of “sandwich” formation, which
results in a significant decrease in signal.
• Avoided by:
○ increasing the quantity of the reagent antibodies
○ Reducing the amount sample required for analysis
○ Sample dilution
• Samples displaying hook effect will yield higher values on
dilution.

23. High-dose hook effect
A
B

24. Heterophile Antibodies
• Known as human antianimal antibodies (HAAAs) or HAMAs.
• Causes:
○ Receiving mouse monoclonal antibodies for therapeutic reasons
○ Exposure to mice.
○ Idiopathic.
• Solutions:
○ Dilution
○ Directly detecting the presence of anti-animal immunoglobulins (such as
antimouse)
○ Nonimmune animal serum is often added to immunoassays to minimize the
effects of heterophilic antibodies
○ Using blocking reagent
○ Many monoclonal therapeutic agents are now derived to include only fragments
of an antibody to avoid the development of heterophilic antibodies.

25. Common Concerns Applied to
Tumor Marker Immunoassays
§ Antibody cross-reactivity
§ Carryover

26. ● HPLC is the most widely used methodology to detect catecholamines and their metabolites in
plasma and urine.
● Useful in the following cases:
○ Neuroblastoma
■ High levels of plasma catecholamines (epinephrine, norepinephrine, and dopamine).
○ Pheochromocytoma
■ Elevated plasma metanephrines (along with urine VMA and free catecholamines).
○ Carcinoid tumors arise from the small intestine, appendix, or rectum
■ Detection of 5-hydroxyindoleacetic acid (5-HIAA), which is a serotonin metabolite.
High performance liquid chromatography

28. ● There are a few tumor markers that are detected directly within solid tissue.
● They are present in the cancer cells themselves in a different manner than the
surrounding tissue.
Immunohistochemistry (IHC)

29. ● Elevation of circulating enzymes are generally not specific for a specific tumor,
except the prostate specific antigen (PSA).
● Enzymes that have been used as tumor markers include:
○ Alkaline phosphatase (bone, liver, leukemia, sarcoma).
○ Creatine kinase–BB (prostate, small cell lung, breast, colon, ovarian).
○ Lactate dehydrogenase (liver, lymphomas, leukemia, others).
○ PSA (prostate).
Enzyme assay

30. Frequently ordered
tumor markers
04

31. ● Alpha-Fetoprotein (AFP) is an abundant serum protein normally synthesized by the fetal
liver that is re-expressed in certain types of tumors.
● Often elevated in:
○ Hepatocellular carcinoma (HCC).
○ Germ cell tumors.
■ Non-seminomatous testicular cancer.
○ Pregnancy
○ Liver disease
● It is a glycoprotein related to albumin that normally functions as a transport protein and is
involved in regulating oncotic pressure in the fetus.
● The upper normal limit for serum AFP is approximately 15 ng/mL in healthy adults.
● Infants initially have high serum AFP values that decline to adult levels at an age of 7–10
months.
Alpha-Fetoprotein

32. ● Classification of testicular cancer:
○ Seminomatous tumors.
○ Non-seminomatous tumors.
§ Embryonal carcinoma.
§ Teratoma.
§ Choriocarcinoma.
§ Yolk sac tumors (endodermal sinus tumor).
● AFP is used in combination with beta-human chorionic gonadotropin (B-hCG) to classify nons-
eminomatous tumors
Alpha-Fetoprotein in testicular cancer
Germ cell tumor AFP hCG
Nonseminomatous
tumor
Yolk sac tumors Increased No
Choriocarcinoma No Increased
Embryonal
carcinoma
Increased
Teratoma No No
Seminomatous Not
elevated
+
-
+
-

33. ● hCG is a 45-kD glycoprotein consisting of Alpha and Beta sub-units.
● Normally secreted by trophoblasts in the placenta to maintain the corpus luteum during
pregnancy.
● Clinical applications as a tumor marker:
○ Ovarian cancer.
○ Testicular cancer.
○ Gestational trophoblastic diseases (GTDs).
● Methodology
○ Immunoassays using monoclonal capture and tracer antibodies targeted toward epitopes in
the beta subunit and intact hCG.
Human Chorionic Gonadotropin (hCG)

34. ● Useful for detecting ovarian tumors at an early stage and for monitoring treatments.
● The CA-125 gene encodes a high-molecular-weight (200,000–1,000,000 kD) mucin
protein.
● Elevated in:
○ Endometriosis.
○ First trimester of pregnancy.
○ Menstruation.
● Methodology:
○ Immunoassays using OC 125 and M11 antibodies.
Cancer Antigen 125 (CA 125)

35. ● Application:
○ Monitoring therapy and to distinguish benign masses from ovarian cancer.
○ For therapy monitoring, CA-125 is useful both for predicting the success of
surgery and for determining efficacy of chemotherapy.
○ Elevated CA-125 following either treatment modality have a poor prognosis.
○ Prognosis is also associated with CA-125 half-life; a CA-125 half-life of less
than 20 days is associated with longer survival; the average half-life of CA-125
is 4.5 days.14,
Cancer Antigen 125 (CA 125)

36. ● Is an oncofetal antigen .
● Used for :
○ colorectal cancer.
○ Lung tumors.
○ Breast tumors.
○ gastrointestinal tumors.
● The upper normal range for serum
CEA is 2.5–5 ng/mL depending on
the assay.
Carcinoembryonic Antigen (CEA)
● Function:
○ It is part of the immunoglobulin superfamily, and is
involved in apoptosis, immunity, and cell adhesion.
■ Because of its role in cell adhesion, CEA has
been postulated to be involved in metastasis.
● Elevated in:
○ heavy smokers
○ following radiation treatment and chemotherapy.
○ liver damage due to prolonged clearance.

37. ● It is a serine protease of the kallikrein gene family
● There are two major forms of PSA that are found circulating in the blood:
○ free form
○ Complexed to Alpha1-antichymotrypsin or Alpha2-macroglobulin.
● Patients with malignancy have a lower percentage of free PSA.
Prostate Specific Antigen (PSA)

38. References
● Bishop, M., 2012. Clinical Chemistry: Techniques, Principles, Correlations, Bishop, 6Th
Edition.
● Mingfang, J.,et.al 2018. Mass screening for liver cancer: results from a demonstration
screening project in Zhongshan City, China. Nature.
● Sturgeon, C. and Viljoen, A., 2011. Analytical error and interference in immunoassay:
minimizing risk. Annals of Clinical Biochemistry, 48(5), pp.418-432.

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