Vivek Misra B.Tech. (Genetic Engg), M.S. (Neuroscience)

Research AssociateTS Srinivasan Institute Neurokrish Of Neuroscience Consulting Pvt. Ltd

Tumor BiomarkersFor

Screening, Progression and Prognosis

vivek@uberbrain.netwww.UberBrain.net

What Are Tumour Markers ?

Tumour markers are indicators of cellular, biochemical, molecular, or genetic alterations by which neoplasia can be recognized.

These surrogate measures of the biology of the cancer provide insight into the clinical behaviour of the tumour.

How Does They Help ?

Diagnostic and distinguish benign from malignant disease

• Correlate with the amount of tumour present (so-called tumour burden)

• Allow subtype classification to more accurately stage patients

• Be prognostic, either by the presence or absence of the marker or by its concentration

• Guide choice of therapy and predict response to therapy

Characteristics Of Ideal Tumour Marker

The marker is expressed exclusively by the particular tumour.

• Collection of the specimen for the tumour marker assay is easy.

• The assay itself is reproducible, rapid, and inexpensive.

Currently, there is no one marker that fulfils all these criteria for any cancer.

Classification of Markers

Tumour markers fall into three broad categories:

• Proteins Markers ;• Genetic Mutations Markers ; • Epigenetic Markers

Proteins were the first type of tumour marker identified and hence are considered the so-called classic tumour markers. For Eg:

Protein Tumour Markers

• Carcinoembryonic antigen (CEA)• α-Fetoprotein • Carbohydrate Antigen 19-9• Prostate-Specific Antigen• Carbohydrate Antigen 125• Human Chorionic Gonadotropin in Testicular Germ Cell Tumors

Carcinoembryonic Antigen (CEA)

• CEA used clinically in patients with cancer of the colon and rectum.

• It is an oncofetal protein that is normally present during fetal life but can be seen in low concentration in healthy adults.

• it is a glycoprotein

• CEA itself is secreted into the circulation and is also found in the mucous secretions of the stomach, small intestine, and biliary tree.

• Although its exact function is unknown, CEA has been shown to be involved in cell adhesion and is able to inhibit apoptosis induced by loss of anchorage to the ECM

CEA contd…

• Normal serum levels are less than 2.5 ng/ml,• Borderline if 2.5 to 5.0 ng/ml, • Elevated if greater than 5.0 ng/ml.

Borderline levels occur with benign disorders such as:• inflammatory bowel disease,• pancreatitis, • cirrhosis, and• chronic obstructive pulmonary disease, and

smoking can also increase CEA—(the upper limit of normal in smokers is considered 5 ng/Ml)

CEA - Testing

CEA - Prognosis

• Elevated CEA levels reflect the burden of tumour present.

• The degree of CEA elevation correlates with increasing stage of disease, and therefore CEA levels have prognostic value.

• Preoperative serum CEA is an independent predictor of survival—the higher the preoperative serum level, the poorer the prognosis.

CEA contd…

• 5-year survival is higher in patients whose elevated preoperative CEA normalized postoperatively.

• Finally, patients with elevated preoperative CEA levels have higher recurrence rates than do those with normal CEA levels.

CEA - Monitoring

• The most common application of CEA is to monitor patients for recurrent disease.

• CEA is most sensitive for hepatic or retroperitoneal metastasis and relatively insensitive for local, pulmonary, or peritoneal involvement.

• Patients with advanced cancer whose CEA levels fall during chemotherapy survive significantly longer

Verdict ??

• CEA is not useful as a screening test because of its low sensitivity in early-stage disease.

• Elevated CEA levels occur in only 5% to 40% of patients with localized disease.

α-Fetoprotein

• α-Fetoprotein (AFP) is used for the detection and management of HCC.

• It is an oncofetal antigen

• It is synthesized by hepatocytes and endodermally derived gastrointestinal tissues.

AFP - Testing

• The upper limit of normal for a healthy, non-pregnant adult is less than 25 ng/Ml

• Levels are also raised in non-seminomatous testicular cancer, for which it is a valuable tumour marker.

• Elevated levels are also seen in hepatitis, inflammatory bowel disease, and cirrhosis.

AFP - Screening

• AFP has an estimated sensitivity of 25% to 75%, a specificity of 76% to 94%.

• The combination of AFP and ultrasound improves the efficacy of screening

• In the United States, recent studies suggest that surveillance of patients with HCV-related cirrhosis with a combination of AFP and an imaging modality (either ultrasound or computed tomography) would gain quality-adjusted life years at acceptable cost.

AFP - Prognosis

• The AFP concentration reflects tumour size, with levels higher than 400 ng/mL being associated with larger tumours.

• As a result, it has been shown that AFP correlates with stage and prognosis of disease.

AFP - Monitoring

• AFP has been shown to decline after resection or ablation.

• In patients whose AFP levels do normalize postoperatively, a subsequent rise in AFP over the course of serial serum measurements has been found to be the best indicator of recurrent disease.

• Tumour re-growth after chemoembolization does not correlate with rate of increase in AFP or tumour burden.

Contd…

• AFP levels usually decline in response to effective chemotherapy.

• Monitoring of AFP therefore avoids prolonged use of ineffective and potentially toxic chemotherapy.

Carbohydrate Antigen 19-9

• Carbohydrate antigen 19-9 (CA 19-9) is widely used as a serum marker for pancreas cancer

• But its use is limited to monitoring response to therapy, not as a diagnostic marker.

• Because CA 19-9 epitope is normally present within the biliary tree. Biliary tract disease, both acute and chronic, can elevate serum CA 19-9 levels.

• It is a mucin-type glycoprotein expressed on the surface of pancreatic cancer cells.

CA 19-9 Testing

• CA 19-9 is detected with an immunoassay.

• The normal range of CA 19-9 in the blood of a healthy individual is 0-37 U/mL

• CA 19-9 associated antigen levels are elevated in the blood of many patients with pancreatic cancer.

CA19-9 - Limitation

• Negative Lewisa blood group antigen cannot synthesize CA 19-9.

• Patients with benign biliary tract disease can have levels up to 400 U/mL.

• High CA 19-9 levels can also be caused by non-cancerous conditions such as gall stones, pancreatitis, cystic fibrosis, and liver disease.

CA19-9 - Screening

• CA 19-9 is not useful as a screening modality because of its low sensitivity in early-stage disease. With increasing levels of CA 19-9, the diagnosis of pancreatic cancer becomes more accurate.

• Levels higher than 1000 U/mL are almost diagnostic of pancreatic cancer.

CA19-9 - Prognosis

• In patients with pancreatic cancer who have CA 19-9 detectable in their serum , the level has been shown to correlate with tumour size.

• Of patients who undergo curative resection, those whose CA 19-9 levels returned to normal survived longer than those whose levels fell but never normalized.

CA19-9 - Monitoring

• Serial measurement of CA 19-9 is used to monitor response to therapy

• Besides pancreatic cancer, CA 19-9 levels are also elevated in patients with other cancers, including those of the biliary tree (95%), stomach (5%), colon (15%), liver (HCC, 7%) and lung (13%).

Prostate-Specific Antigen

• Prostate-specific antigen (PSA) is a serine protease that is formed in the prostatic epithelium and secreted into the prostatic ducts.

• Its function is to digest the gel that is formed in seminal fluid after ejaculation. Under normal circumstances, only small amounts of PSA leak into the circulation.

Contd…

• With enlargement of the gland (e.g., in patients with benign prostatic hyperplasia [BPH]) or distortion of its architecture, serum PSA levels increase.

• Thus, PSA is considered a tissue-specific rather than a prostate cancer–specific marker—patients who have undergone curative radical prostatectomy, as well as females, have no detectable PSA.

PSA - Testing

• PSA is detected with an immunoassay.

• serum PSA levels may be elevated include bph , prostatitis, prostatic massage, prostatic biopsy, and digital rectal examination.

Contd….

• it has been found that the upper limit of the normal range of PSA increases with age. The limit is 2.5 ng/mL for men aged 40 to 49 years, 3.5 ng/mL for those 50 to 59, 4.5 ng/mL for those 60 to 69, and 6.5 ng/mL for men 70 years and older.

• The rate of increase in PSA in a normal 60-year-old is 0.04 ng/mL/yr.

PSA - Density

• PSA density is defined as the ratio of PSA to prostatic volume, as measured by transrectal ultrasound or magnetic resonance imaging.

• Higher PSA densities are more suggestive of malignancy than BPH because the amount of PSA released per gram of prostate cancer is significantly greater than that released from normal prostatic tissue.

PSA - Screening

• PSA is widely used as a screening tool for prostate cancer because it enables early detection and diagnosis of this disease,

PSA - Monitoring

• After operative resection, the PSA level is expected to normalize after 2 to 3 weeks.

• Normally it takes 3 to 5 months for PSA to normalize after radiotherapy in ca prostate.

• In patients with advanced disease, PSA levels are also used to monitor response to systemic therapy.

Carbohydrate Antigen 125

• Carbohydrate antigen 125 (CA 125) is a carbohydrate epitope on a glycoprotein carcinoma antigen.

• It is present in the fetus and in derivatives of the coelomic epithelium, including the peritoneum, pleura, pericardium, and amnion.

Contd…

• In healthy adults, CA 125 has been detected by immunohistochemistry in the epithelium of the fallopian tubes, endometrium, and endocervix.

• However, neither adult nor fetal ovarian epithelium expresses CA 125.

CA 125 - Testing• CA 125 levels are measured with an immunoassay,

with the upper limit of normal set at 35 U/mL.• Elevated levels are detected in ovarian cancer.• cancer of the fallopian tube, endometrium, and

cervix.• as well as in nongynecologic malignancies of the

pancreas, colon, lung, and liver.• In Benign conditions CA 125 is elevated which are

endometriosis, adenomyosis, uterine fibroids, pelvic inflammatory disease, cirrhosis, and ascites

CA 125 - Screening

• CA 125 is not useful as a screening tool for ovarian cancer because of its poor specificity.

CA 125 - Prognosis

• Patients with elevated CA 125 levels at the time of diagnosis have a worse prognosis than patients with normal levels do.

• CA 125 levels—• 50% of stage I patients, • 70% of stage II patients, • 90% of stage III patients, and • 98% of stage IV patients

CA 125 - Monitoring

• Partial or complete response to therapy is associated with a decrease in CA 125 levels.

• Increasing levels of CA 125 correlate with disease recurrence and precede clinical or imaging evidence of recurrence by a median of 3 months.

• CA 125 levels in peritoneal fluid may be more sensitive than serum levels.

Contd…

• The upper limit of normal for peritoneal fluid CA 125 is 200 U/mL.

α-Fetoprotein and Human Chorionic Gonadotropin in Testicular Germ Cell Tumors

• Nonseminomatous testicular cancers comprise several different histologic types:

– embryonal carcinoma;– syncytiotrophoblasts (choriocarcinoma);– yolk sac tumors; and – teratomas.

Contd…

• Marker expression can be predicted on the basis of the predominant histologic type:

– human chorionic gonadotropin (HCG) is detected in more than 90% of choriocarcinomas,

– whereas AFP is expressed by 90% to 95% of yolk sac tumors,

– 20% of teratomas, and – 10% of embryonal carcinomas.

Diagnosis

• The presence of a testicular tumour in combination with an elevated level of AFP or HCG is suggestive of testicular cancer, without being diagnostic.

• Elevated levels of these markers in a man younger than 40 years without signs of a testicular tumour may indicate extra-testicular germ cell cancer.

Prognosis

• An absolute AFP concentration greater than 500 ng/mL or an HCG level higher than 1000 ng/mL is predictive of a poor prognosis.

• These tumour markers are useful in identifying biologically distinct categories of morphologically similar tumours

Monitoring

• In the majority of patients with nonseminomatous germ cell tumors, tumor marker levels correlate with response to chemotherapy.

• rate of marker decline can be used for early identification of patients who will respond poorly to chemotherapy.

• Half-lives longer than 3.5 days for HCG or longer than 7 days for AFP suggest that very aggressive therapy is required.

• HCG level in urine is similar to that of serum .

DNA - Based Markers• Specific mutations in oncogenes, tumor-suppressor

genes, and mismatch repair genes can serve as biomarkers.

• These mutations may be:• – germline. e.g. ret proto-oncogene of MEN 2– APC gene of FAP, – somatic mutations such p53 mutations.– Chromosomal abnormalities such as 9:22 translocation

that creates the bcr-abl oncogene (biomarkers).

Epigenetic Changes• Testing for epigenetic changes is still at an early

discovery stage. But it has great potential because –

1. DNA assays for aberrant methylation are easier and more sensitive than those for point mutations.

2. cancer-specific DNA methylation patterns can be detected in tumour-derived free DNA in the bloodstream and in epithelial tumour cells shed into the lumen.

Contd…

• DNA-methylation profiles are more chemically and biologically stable than RNA or most proteins. As a result, they may be more reliably detected in diverse biologic fluids.

Advances In Assays

• DNA Methylation biomarker studies have been performed in a variety of cancers, including breast, esophageal, gastric, colorectal, and prostate cancer.

• Combining DNA methylation assays may complement existing screening methods with high sensitivity but low specificity, such as PSA in prostate cancer.

• The use of panels of methylation targets in these studies improved the clinical sensitivity of the assay.

Biomarkers and Biologically Targeted Therapies

• Biomarker expression is increasingly being used, independent of formal staging criteria, to decide which patients receive biologically targeted therapies.

Contd…

CANCER BIOMARKER THERAPYBreast Estrogen receptor,

progesterone receptorTamoxifen/aromatase inhibitors

Lymphoma CD20 Rituximab

Chronic myelogenous leukemia (CML)

bcr-abl Imatinib

Gastrointestinal stromal tumor (GIST)

c-kit Imatinib

Non–small cell lung cancer EGFR mutation Gefitinib

Breast HER2/neu Trastuzumab

From Ludwig JA, Weinstein JN: Biomarkers in cancer staging, prognosis and treatment selection. Nat Rev Cancer 5:845-856, 2005.

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