Biochemistry of CancerGlimpse of chemistry, pathways & tumor markers

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Precis

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Introduction

▪ A disease caused by an uncontrolled division of abnormal cells in a part of the body.

▪ Cancer is characterized by ▪ Unrestrained cell growth

▪ Cell Immortality

▪ Local invasion & Distant metastasis

▪ Cancer is the second most important cause of death world-wide

▪ Can affect potentially any person with any demographic parameters (Age, gender, ethnicity, geographical location etc.).

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Causes & Characteristics▪ Exact cause is unknown.

▪ Found to be associated various factors, both internal & external

▪ External factors are called Carcinogens

▪ Association & Causation of external factors

▪ Carcinogens can be chemical, physical, infectious agents (viral & bacterial).

▪ Internal factors so far identified are inherited genetic changes

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Two types of growth

▪ Benign: ▪ Well-defined mass of tissue

▪ demarcated from normal tissue

▪ slow-growing

▪ Mostly differentiated cells

▪ no invasion

▪ no distant metastasis.

▪ Malignant▪ Ill-defined tissue growth,

▪ no demarcation from normal tissue,

▪ can be rapidly growing;

▪ poorly differentiated cells

▪ local invasion

▪ distant metastasis are usual.

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Two types of spread

▪ Local Invasion: ▪ Early stage of the disease

▪ Direct migration of cancer cells

▪ Penetration of basement membrane

▪ Distant metastasis: ▪ Later stage of the disease

▪ Direct migration of cancer cells or indirectly materials from cancer cells such as DNA

▪ Usually penetration and transport via lympho-vascular system

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External Factors (Carcinogens) ▪ Physical: Harmful Radiations (Ultraviolet, X and radiations)

▪ Chemical: from lifestyles (alcohol & cigarette smoking), diet (aflatoxins) etc.

Class CompoundPolycyclic aromatic hydrocarbon Benzo(a)pyrene, Dimethyl benzanthracene

Aromatic amines Acetylaminofluorene, aminobenzene

Nitrosamines Dimethyl and diethyl nitrosamines

Drugs Cyclophosphamide

Naturally occurring compounds Aflatoxin B1, Dactinomycin

Inorganic compounds Arsenic, Asbestose, beryllium, cadmium, chromium

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External Factors (Carcinogens)

▪ Biological factors: Oncogenic viruses

Virus Nucleic Acid Associated cancerEpstein Barr virus (EBV) dsDNA Burkitt lymphoma, Nasopharyngeal carcinoma

Hu papilloma virus (HPV) dsDNA Uterine, Cervical carcinoma

Hepatitis B virus (HBV) dsDNA - RT Hepatoma & Hepatocellular Carcinoma

Biochemical MechanismsWhat are the different cellular and molecular mechanisms involved in Cancer?

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Mechanisms for tumor cell formation

▪ What happens when a normal cell becomes tumor cell? ▪ External or tumor microenvironment changes. ▪ Internal acquired/inherited cellular growth mechanisms

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External factors or Tumor- Microenvironment or Extracellular- Matrix (ECM)

Cell 2011 144, 646-674DOI: (10.1016/j.cell.2011.02.013)

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Growth factors

▪ Abnormal amount of growth factors to sustain proliferative signaling or their increased susceptibility for certain types of cells. E.g. Insulin Like Growth Factor, Epidermal Growth Factor, Platelet Derived Growth Factor etc. Estrogen and Bread adenoma. Estrogen receptor hypersensitivity in Ca breast

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Mucopolysaccharides

▪ Stromal Contact inhibitors: Molecules which affect contact inhibition of cell proliferation such as Sialic acid and Hyaluronic acid residues. Due to higher content of these negatively charged long residues there is a loss of orientation of cells and repel apart.

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Cell::Cell interaction

▪ Abnormal cell adherence junction (tight junction) due to altered protein interaction due to structural modification

▪ Loss Apico-basal polarity due to abnormal regulation of distribution of protein

▪ Abnormal anchoring proteins to ECM e.g. integrins, fibronectin, vimentin etc. due to protein structure abnormality.

ERMNF2 / Merlin

Apical

LateralDynamic cell:cell adhesion Stable cell:cell adhesion

Brit. J. Cancer 2008; BBA 2007

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Acetate and SCFA produced by microbiota of gut cells regulate abnormal growth of colonic cells

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External factors

Altered immunity:

1. Abnormal recruitment Regulatory T-cells (T-regs) which controls the specific local infiltration of T-lymphocytes to weed out Cancer cells is checked.

2. Regulated Immuno “rheostat” or immunostat or Check-point inhibitors such as PD-1/PD-L1

Cancer Immunity 2013 39(1), 1-10 DOI: (10.1016/j.immuni.2013.07.012)

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CONFLUENCE OF EXTERNAL MECHANISMS IS THE RULE FOR MALIGNANT PROGRESSION

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Internal Factors

▪Acquired changes.▪Hereditary changes.

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Broad Cellular Mechanisms

Cell 2011 144, 646-674DOI: (10.1016/j.cell.2011.02.013)

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Internal Changes: Intracellular signaling cell viability circuits

Cell 2011 144, 646-674DOI: (10.1016/j.cell.2011.02.013)

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Cell 131, December 14, 2007 p1204.e1–1204.e2 DOI 10.1016/j.cell.2007.11.036

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Cell 131(5), December 30, p1018.e1–1018.e2 DOI 10.1016/j.cell.2007.11.013

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Cell 133 (7) June 21, 2008 p1292–1292.e1

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Cell 152 (3) June 21, 2008 p656–656.e1

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Cell 133, May 2, 2008 DOI 10.1016/j.cell.2008.04.023

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Snapshot: Tumor Angiogenesis

Cell 149(6) p1408–1408.e1 8 June 2012 DOI 10.1016/j.cell.2012.05.025

Cancer EnergeticsWhere does cancer cells get energy from?

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Warburg Effect

▪ Otto Heinrich Warburg (1883-1970)

▪ Identified that most cancer cells predominantly produce energy by high rate of glycolysis followed by lactic acid fermentation in cytosol even when plenty of oxygen is available.

▪ Nobel Prize in 1931

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Mechanism of Warburg Effect

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Uses of Warburg effectDiagnostic▪ 2-fluoro-2-deoxy glucose (FdG) (18F

replaced C2 of glucose.

▪ Cancer cells uptake 10x more, trapped in cancer cells as 6-phosoho FdG

▪ Decay of 18F gives positron which are detected by Positron Emission Tomography

Therapeutic

▪ Gleevec (Imatinib) inhibits Tyrosine Kinase enzyme preventing hexokinase activation is a clinically approved drug.

▪ Oxythiamine which inhibit transketolase enzyme in preclinical trials.

Changes in DNAWhat are the biochemical changes in the informational molecules?

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Categories of DNA modifications

Types of Genes involved in cancer

OncogenesTumor

Suppressor Genes

Stability Genes

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Oncogenes▪ Oncogenes are mutant forms of the genes for

proteins that regulate cell cycle.

▪ Are originally discovered from viruses, which was later found to be DNAs incorporated from animal hosts of their precursor viruses, called proto-oncogenes.

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Oncogenes▪ Several mechanisms of oncogene formation

▪ Viral incorporation Replication errors in virus Reincorporation into human cells failed regulation of cell division

▪ Selective dominance: Mutations in certain genes give characteristic dominance in specific proteins which trigger for cell proliferation. E.g. nuclear transcription factors which control cell division (Jun, Fos, etc.).

▪ Activating mutations: Oncogene mutations lead to spontaneously activating mutations in certain growth factor cell membrane receptor. E.g. Mutations of oncoprotein ErbB lead to spontaneous activation of the EGF receptor, without binding of ligand.

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Tumor suppressor gene

▪ Tumor suppressor genes encode proteins that normally restrain cell division.

▪ Mutations of one/ more of these genes can lead to tumor formation

▪ Usually genetically recessive. (unlike oncogenes).

▪ Second hit: If one copy of the gene is mutated congenitally, and if a second copy is mutated in any of the 1012 cells of the body, tumor could potentially start forming from that cell.

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Tumor suppressor genes

▪ Retinoblastoma gene (Rb): Second hit usually associated retinoblastoma and many types of tumors such as cancers of lung, prostate, breast etc. later in life.

▪ Von Hippel Lindau (VHL) gene: Congenital absence of p.VHL protein lead to uninhibited vasculogenesis in cerebellar hemisphere (called hemangioblastoma), also associated with multiple cysts in kidneys, liver, pancreas etc.

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Stability genes▪ Also known as Caretaker genes: e.g. ATM, BRCA1 gene family, XP

gene family, TP53 etc.

▪ The proteins encoded by these genes function in the repair of major genetic defects resulted from aberrant DNA replication, ionizing radiation, or carcinogens.

▪ TP53 : called “Guardian of human genome”. This gene▪ Activate DNA repair▪ Arrest growth by holding cell cycle at G1/S phase ▪ Can initiate apoptosis (or programmed cell death)▪ Senescence response to short telomeres

▪ Congenital defects of p53 protein lead to a rare disorder called Li-frumeni syndrome (associated with cancers in multiple organs).

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Overall

Cancer generally results from accumulated genetic changes or mutations to oncogenes, tumor suppressor genes and stability genes over a period of time.

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What are the chemical changes in DNA?▪ Point mutations: Substitutions (missense/nonsense),

Deletion/duplications one or two nucleotide bases in the DNA

▪ Deletions/duplications of short stretch of DNA: called indels, micro-indels.

▪ Gene Rearrangements: of large regions of chromosomes giving some peculiar proteins called fusion proteins which gives some cell proliferative properties. For e.g. BCR-ABL fusion of in Chronic Myeloid Leukemia (CML). Entire ABL gene in chromosome 9 is juxtaposed/fused onto entire BCR gene of chromosome 22 producing a hybrid protein which gives uncontrolled tyrosine kinase activity for cell growth in certain cells of bone-marrow which led to CML.

Tumor markersProteins that can portend!

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What are tumor markers? ▪ Factors released by tumor cells, detected in blood and other body fluids

and potentially indicate the presence of tumor in the body.

▪ Uses▪ Diagnostic: Mere presence could be a sign of tumor, but cautious of non-

malignant conditions▪ Prognostic: Serum levels may roughly indicate the tumor load, which can

predict the effect of treatment▪ Localization: Certain tumor markers are suggestive of tumors in certain

specific organs or part of organs. ▪ Therapeutic: Once the treatment is started, progressive determination of

serum levels could mirror tumor remission.

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Classical tumor markers

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Clinically important Tumor markers

1. Alpha-fetoprotein (AFP)

2. Carcinoembryonic antigen (CEA)

3. Beta Human Chronic Gonadotrophin (-HCG)

4. Cancer Antigen 125 (CA-125)

5. Tissue Polypeptide Antigen (TPA)

6. Prostate Specific Antigen (PSA)

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Alpha fetoprotein (AFP)

▪ Fetal albumin like protein, about 70 kDa molecular weight.

▪ In adult males and non-pregnant females, it can be upto 15 ng/ml

▪ A value of upto 300 ng/ml can occur nonmalignant liver disease

▪ More than 300 ng/ml is usually associated with cancer

▪ Associated with mostly hepatocellular carcinoma.

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Carcinoembryonic antigen (CEA)▪ Set of highly related glycoprotein closely belong to immunoglobulin

superfamily by 29 genes.

▪ Normally produced by fetal gastrointestinal system, stops production by birth

▪ Molecular weight: as large as 185 kDa.

▪ Mostly elevated in adenocarcinomas of colon, lung, breast, stomach & pancreas (above approximately 2.5 µg/L)

▪ Left sided colon cancers have higher levels

▪ Higher levels are associated with lymph node spread.

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Cancer Antigen 125 (CA-125)▪ A glycoprotein of more than 1 MDa molecular

weight.

▪ Normal level in the blood is < 35 U/ml

▪ Elevated levels are associated with 75% of ovarian cancers.

▪ Elevated levels can also be found in 20% of pancreatic and GI cancers.

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-HCG▪ Synthesized by syncytio-trophoblasts of placental villi

▪ Molecular weight of HCG is ~ 45 kDa.

▪ HCG has an -subunit shares common structure with FSH, LH & TSH.

▪ -subunit is unique to HCG.

▪ Increased in hydatidiform mole, choriocarcinoma & germ cell tumor

▪ Also associated with 60% of testicular cancers.

▪ Normal value is 20 IU/L (> 10,000 IU/L is trophoblastic tumor)

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Bence-Jones Proteins (BJP)▪ Abnormal production of

immunoglobulin when plasma cells proliferate.

▪ Plasmacytoma/Multiple myeloma

▪ Characterized by lytic bone lesions, anemia, para-proteinemia, proteinuria.

▪ BJP is seen in 20% cases of multiple myeloma

Alpha-I

Beta

Gamma

Albumin

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Newer tumor markers▪ Estrogen Receptor (ER)

▪ Progesterone Receptor (PR)

▪ Beta2-macroglobulin

▪ Bladder tumor Antigen (BTA)

▪ Calcitonin

▪ Her2/neu

▪ Neuron Specific Enolase (NSE)

▪ Thyroglobulin (Tg)

▪ Thyroid Transcription Factor-1 (TTF-1)

▪ Cytokeratin-7 (CK-7)

▪ Synaptophysin

Thank youDr. Prasanth. A. S.