Prof. A. Angeloni

 Biological substances synthesized and released by cancer cells themselves or  Produced by the host in response to the presence of tumor  Most tumor markers are proteins  Detected in a solid tumor, in circulating tumor cells in peripheral blood, in serum, lymph nodes, in bone marrow, or in other body fluid (urine, stool, ascites)

What a Tumor Marker should reveal?  Is the patient affected by cancer?  Which organ is affected?  Cancer is localized or metastatic?  What is the growth rate of the cancer?  Is there a recurrence of cancer before symptoms?  Which treatment will be more effective?

Screening Early Detection DiagnosisMonitor

1. Screening To identify early cancer risk 2. Diagnosis To confirm the diagnosis 3. Staging To assess & stratify the risk 4. Prognosis To predict the outcome 5. Localization To locate the primary site 6. Therapy To target the therapy 7. Surveillance To detect recurrence in F-Up 8. Monitoring To evaluate response to Th

1. Cancer heterogeneity 2. Lack of Specificity – false positives 3. Lack of Sensitivity - false negatives 4. Benign diseases - positive CA 125 or CEA 5. Smokers have raised CEA 6. Normal persons also have small amounts 7. Higher levels only with large tumor volume 8. Some cancers never have higher levels

AntigensHormones EnzymesTissue Specific TUMO R

 Tumour-Associated Proteins (TAP)  Cell membrane receptors  Hormones  Immunoglobulins / Cellular antigens  Polyamines  Protein clusters and fragments  Chromosomal material  Genes (single, clusters)  Genetic material (DNA, RNA, mRNA)  Cell modulators (transducers / suppressors)

1. Viral Antigen : a- Viral proteins and glycoproteins b- New antigens produced by virally infected host cells under control of viral nucleic acid 2. Tumor specific antigens : - Tumor cells develop new antigens specific to their carcinogens 3. Tumor specific transplantation antigens : - Tumor cells express new MHC antigens due to alteration of normally present MHC antigens

4. Oncofetal antigens: a- Carcino-embryonic antigens (CEA) - Normally expressed during fetal life on fetal gut - Reappearance in adult life: GIT, pancreas, biliary system and cancer breast b- Alpha fetoprotein: - Normally expressed in fetal life - Reappearance in adult life; hepatoma

A. Hormones : Human Chorionic Gonadotrophins (HCG) are secreted in Chorioncarcinoma, Ovarian Carcinoma; Thyroxin is secreted in thyroid cancer B. Enzymes : Acid phosphatase in prostate cancer; Alkaline phosphatase, lipase and amylase enzymes in cases of pancreatic cancer

 Enzymes (PSA, NSE, VMA, HVA)  Cell membrane receptors (ER, PR)  Tumor antigens (CEA, AFP)  Antibodies (IgA, IgG, IgM, IgD)  Antigens (p53, ki-62)  CA-specific proteins (CA 19-9, CA 125)  Gene mutation products (BRCA 1, 2)

 Tissue-specific proteins (PSA, hCGH)  Special hormones (b-hCGH, h-CGH)  Catecholamines (VMA, HVA, ACTH)  Polyamines  Cytoplasmic / Nucleic material (DNA)  Products of cell turn-over (TNF)  Cellular modulators (ki-62, c-erb-2)

 ELISA  Immuno-histochemistry (IHC)  Polymerase chain reaction (PCR)  Fluorescence in situ hybridization (FISH)  Cluster Kits ( All-in-One Kit)  Detects profiles  Patterns  Prototypes  Constellations

 Expression of single proteins  Expression of multiple proteins  Chip analysis – “All-in-One”  Expression of protein profiles (Proteomics)  Gene methylation at DNA level  Genes / mutations (Genomics)  G-scan (genome ID scan)

1. hCGH (prostate and germinal specific) 2. beta-hCGH (prostate and germinal specific) 3. CEA (NS) 4. AFP (NS) 5. Bence-Jones (MM) 6. Beta-2-M (MM staging) 7. Bladder Tumor Antigen (Bladder) (S) 8. CgA Chromogranin-A (neuroendocrine) 9. CA-15-3 (NS) 10. CA-19-9 (NS) 11. CA-72-4 (NS) 12. CA (NS) 13. CA-125 (NS) 14. ER / PR (Breast) 15. HER-2 neu (c-erbB-2) 16. BRCA-1 / BRCA-2

1. Lipid Associated Sialic Acid-P (S-ovarian) 2. Nuclear Matrix Protein- 22 (S-bladder) 3. PSA (Prostate-S) 4. Prostate Specific Membrane Antigen (Prostate-S) 5. S-100 (Melanoma) 6. TA-90 (early stage Metastatic melanoma NS) 7. TPA (colon cancer NS) 9. Alk. p’tase (mets) 10. Alpha Amylase 11. Galactosyl Transferase-II (pancreatic NS) 12. VMA, HVA (neuroendocrine S) 13. Polyamines (NS) 14. Genes (k-ras, ki-62) 15. Chromosome (p53)

 Prostate Specific Antigen(PSA) is a glycoprotein  Ideal as a tumor marker, high tissue specificity  High sensitivity for prostate cancer  Also elevated in BPH & prostatitis  Useful in  Follow up of prostate Ca, Prognostic factor  To monitor recurrence & response to treatment  ? For screening of prostate cancer along with Digital Rectal Exam

 Free PSA ： PSA not bound to the plasma anti proteases α1-antichymotrypsin & α2-macroglobulin  An ↑in ratio of free/total PSA is associated with increased probability of prostate cancer  97% specificity, 96% sensitivity for prostate Ca  For population screening and diagnosis an increase of 0.75 ng/ml per year in any given patient has high sensitivity and specificity for prostate cancer vs BPH, especially when combined with DRE and Trans Rectal Ultra Sound

 80% of non mucinous ovarian cancer detected by the monoclonal antibody to CA-125  Elevated in Ovarian, Endometrial, Pancreatic, Lung, Breast, Colon cancers and also in  Menstruation, Pregnancy, Endometriosis and other gynecological and non gynec conditions.  Useful in monitoring ovarian Ca recurrence & Rx.  Screening of high risk population (BRCA1-2 Carriers); Not useful for routine screening

 Cell surface glycoprotein, present during embryonic development of coelomic epithelium & is present in adult structures derived from it  For follow up, an increase may predict recurrent disease, precedes clinical recurrence by months  >80% of epithelial ovarian cancer, cell types ： serous > endometriod, clear cell > mucinous  Correlates with tumor bulk,  In Endometriosis most common   levels also found in PID, 1st trimester

 Alfa Feto Protein is a serum fetal protein synthesized by the liver, yolk sac, gastrointestinal tract – a glycoprotein  In Hepatocellular Cancer: It is diagnostic (>500) and also useful for screening of high risk population (HBV, HCV)  Benign conditions: hepatic parenchymal inflammation, hepatic necrosis, pregnancy, primary biliary cirrhosis, extra hepatic biliary obstruction give positive test.  Testicular germ cell tumor (embrional or endodermal):  Diagnosis, Prognosis, to monitor recurrence & response  The absolute AFP level correlates with tumor bulk  Cancers of pancreas, colon, stomach & bronchogenic Ca

 Complex glycoprotein that is associated with the plasma membrane of tumor cells, from which it may be released in to the blood  Elevated specially in Colon cancer, Adeno. Ca uterus  Normal pre Rx CEA indicates no metastasis  Also in Pancreatic, Gastric, Lung, breast & Ovarian Ca  Also in cirrhosis, inflammatory bowel disease, chronic lung disease, pancreatitis, fibrocystic breast disease  19% of smokers, 3% of healthy population  Not satisfactory for screening for a healthy population  Good for monitoring recurrence & to monitor Rx.

CA 19-9 is elevated in  In 21-42% patients of gastric Ca  In 20-40% patients of colonic Ca  In 71-93% patients of pancreatic Ca  For DD of benign from malignant disease Dx, FU, Relapse, 70% specificity & 90% sensitivity  It is a mucin, does not increase during pregnancy  Monitor patients who do not express CA 125, mucinous (76%) > serous (27%)

 Human chorionic gonodotropin (βHCG)  Glycoprotein synthesized by syncytio trophoblastic cells of normal placenta  Serum and urine HCG ↑ in early gestation and peak in the first trimester (60~90 days)  Elevated in Gestational trophoblastic disease (a progressive rise in after 90 days of gestation → highly suggestive), choriocarcinoma  Elevated in testicular cancer, βHCG after surgery  Monitor treatment response, relapse & recurrence

Estrogen Receptor (ER)  2 isoforms ： ERa and ERb  ERa → better prognosis, predictor of relapse  useful when deciding on adjuvant hormone treatment  As diagnostic marker when it is a primary unknown tumor  ERb → Good prognostic factor, correlates with low grade and negative axillary LN status

 HER-2/neu oncogene (using monoclonal antibody) - over expression related to poor prognosis in breast cancer  Oncogene c-erbB-2 gene ： over expressed in 30% of breast cancers, correlation between c-erbB-2 gene positivity, positive axillary node status, reduced time to relapse and reduced overall survival  BRCA1 gene on chromosome 17q ： familial breast-ovarian cancer syndrome, and breast cancer in early-onset breast cancer families → high risk screening

- To monitor Rx. & to detect recurrence BR Ca  ↑ in 20% with localized breast cancer, ~80% with metastatic disease, esp. if with bone involvement  Specificity of 86%, sensitivity of 30%  Also ↑ in gastric, pancreatic, cervical & lung cancer  c-erbB-2 overexpression should be evaluated on every primary breast cancer either at the time of diagnosis or at the time of recurrence.

 Tyrosinase  Use RT-PCR to detect hematogenous spread of melanoma cells from a solid tumor in peripheral blood  S100B protein  For confirmation of amelanotic malignant melanoma by immunohistology  ↑in 70% with stage IV metastasized melanoma  MIA (melanoma inhibitory activity)  Preoperative: 59% at stage III, 89% at stage IV

 Thyroglobulin  Tissue-specific, glycoprotein produced by thyroid follicular cells  Also increased in breast or lung cancer  Thyrocalcitonin  From thyroid C cells & medullary thyroid cancer  Effective to screen patients with 1st degree relatives affected by medullary thyroid cancer and multiple endocrine neoplasia type 2 (MEN2)

 Burkitt’s type lymphoma and leukemia  T (8;14) due to juxtaposition and activation of the c- myc gene  CD 25 most sensitive serum marker for tumor burden  CD 44 high concentration indicates poor prognosis  Lactate dehydrogenase (LDH)  Normal: 100~250 IU/L  High-grade lymphomas, blood levels correlate closely with disease activity and response to therapy

 Neuron-specific enolase (NSE)  A neuronal isoenzyme of cytoplasmic enzyme enolase, in neuroendocrine cells  As a prognostic factor in neuroblastoma  Occurs in neuroendocrine tumors: medullary carcinoma of the thyroid, pheochromocytoma, carcinoid tumors; immature teratoma, small cell carcinoma of lung, non-small-cell cancer, melanoma. Correlate with stage and bulk of disease  N-myc oncogene in neuroblastoma N-myc copy number is associated with stage and prognosis

 Expressed only in tumor cells  Example: an oncogene is translocated and fused to an active promoter of another gene → fusion proteins → constant active production → development of malignant clone  Philadelphia chromosome in CML, t(9;22)  (q34;q11) bcr/abl translocation  t(8;21) acute non-lymphocytic leukemia,  t(15;17) in APL  Hematological malignancies

 Abnormal Chromosome  Due to translocation  t (9;22) – Ph short Chrom  bcr/abl fusion gene  This takes place in a single bone marrow cell  Creating fusion proteins  Detected by FISH technique  Philadelphia chromosome in ALL - poor prognosis  Abnormal Chromosome  Due to translocation  t (9;22) – Ph short Chrom  bcr/abl fusion gene  This takes place in a single bone marrow cell  Creating fusion proteins  Detected by FISH technique  Philadelphia chromosome in ALL - poor prognosis