Medical Biochemistry

Medical Biochemistry Cancer and Oncogenes Lecture 74

Properties of cancer cells • Diminished or unrestrained control of growth • benign tumors also show diminished growth control • invasion of local tissues • spread, or metastasis to other parts of the body

Agents causing cancer • Three broad groups of agents • radiant energy • chemical compounds • viruses • Act by causing mutations or introducing genes into cells • Familial conditions also cause cancer • mutations in specific genes (e.g., tumor suppressor genes) • Spontaneous mutations (may predispose to cancer) • frequency ~10-7 to 10-6 per cell per generation • will increase in tissues subject to high rate of proliferation, increasing generation of potential cancer cells from affected stem cells • oxidative damage to DNA (e.g., release of OH• from mitochondria) may be factor  mutation rate

Radiant Energy • Ultraviolet rays, x-rays, and g-rays are mutagenic and carcinogenic • Result in DNA damage • formation of pyrimidine dimers (elimination of corresponding bases may form apurinic or apyrimidinic sites • single- and double-strand breaks or cross-linking strands may occur • x-rays and g-rays also cause free radicals to form in tissues • resultant OH•, superoxide, and other radicals can interact with DNA leading to molecular damage and contribute to carcinogenic effects

Chemical Carcinogens • Estimated that ~80% of human cancers caused by environmental factors, principally chemicals • Exposure by: • occupation (e.g., benzene, asbestos) • diet (e.g., aflatoxin B1, mold sometimes contaminant of peanuts • life-style (e.g., cigarette smoking) • other ways (e.g., some therapeutic drugs can be carcinogenic)

Chemical Carcinogens • Both organic and inorganic molecules may be carcinogenic - no common structural feature • Some compounds interact directly with target molecules (direct carcinogens)

Chemical Carcinogens • Others may require metabolic activation procarinogen  proximate carcinogen  ultimate carcinogen • procarcinogen not chemically reactive • ultimate carcinogen often highly reactive • usually electrophiles (i.e., molecules deficient in electrons) • readily attack nucleophilic (electron-rich) groups in DNA, RNA, and proteins • form adducts (most common site of attack is guanine)

Chemical Carcinogens • principally species of cytochrome P450 (in ER) responsible for metabolic activation of procarcinogens • normal function to detoxify noxious chemicals (xenobiotics) • therapeutic drugs, insecticides, polycyclic hydrocarbons • many of these compounds so fat-souble they would accumulate continually in fat cells and lipid membranes, not be excreted from body • detoxification by converting to water-soluble derivatives that can be excreted

Mutagens • Most chemical carcinogens are mutagens • Ames assay - used to detect mutagenicity of chemical carcinogens • Salmonella typhimurium auxotroph • strain with mutation (His-) in gene involved in synthesis of histidine • Grow on medium lacking His inpresence of test compound • screen for mutations that cause reversion to His+ • Compound may be activated by incubation with postmitochondrial supernatant (contains microsomes)

Initiation and Promotion • Chemical Carcinogen Experiment on Skin: • Paint skin of mice with benzo[a]pyrene  no tumors • After benzo[a]pyrene, apply croton oil several times  many tumors develop • Apply croton oil alone  no skin tumors • Conclusions: • stage of carcinogenesis caused by benzo[a]pyrene called initiation • rapid and irreversible (modification of DNA?) • second, slower stage (months or years) is promotion • promoters incapable of causing initiation • Most carcinogens capable of acting as both initiators and promoters

Initiation and Promotion • Large number of compounds can act as promoters in various organs • phenobarbital, saccharin • Active agent of croton oil is a mixture of phorbol esters • most active phorbol esters is12-O-tetradecanoylphorbol-13-acetate (TPA) • protein kinase C (PKC) can act as a receptor for TPA • activation of PKC with TPA may lead to phosphorylation of signaling molecules, loss of growth control • may increase proliferation of stem cells

DNA and RNA Viruses are Carcinogenic • Transforming activity of tumor virus resides in particular gene(s) carried in viral genome • Response of cell to infection by DNA tumor virus depends on whether cells are permissive or nonpermissive • permissive cells productively infected  virus lytic cycle (e.g., adenovirus in humans) • in nonpermissive cells, viral replication abortive, viral DNA integrates, may transforms cell (EBV associated with Burkitt’s lymphoma)

DNA and RNA Viruses are Carcinogenic • RNA tumor viruses (retroviruses) • during life cycle, RNA genome converted to DNA by viral reverse transcriptase, can be integrated into host genome • Transforming retrovirus carries copy of cellular sequence in place of some of its own gene(s) • usually replication defective • expression of transduced cellular gene(s) may alter phenotype of infected cell (may stimulate growth of infected cells, enhance proliferation of virus)

Oncogenes of retroviruses • c-onc (cellular oncogene) - oncogene present in tumor cells • proto-oncogene - gene present in normal cells • v-onc (viral oncogene) - oncogene present in virus • viral oncogenes represent wide variety of signaling molecules (mostly protein tyrosine kinases)

Oncogenes of retroviruses • Two theories may explain difference between v-onc and c-onc genes • quantitative model - viral genes functionally the same as cellular genes, but oncogenic because expressed in much greater amounts, in inappropriate cell types, or at inappropriate times • qualitative model - c-onc genes intrinsically lack oncogenic properties, but can be converted by mutation into oncogenes (acquire, or lose, important property)

Activation of Proto-oncogenes • Five mechanisms alter the expression or structure of proto-oncogenes and participate in their conversion to oncogenes • Promoter insertion • Enhancer insertion • Chromosomal Translocations • Gene Amplification • Point Mutation Quantitative model Qualitative model

Promoter Insertion • Certain retroviruses lack oncogenes (e.g., avian leukemia viruses) but may cause cancer over longer period of time • Infect cell, reverse transcriptase synthesizes cDNA of RNA genome, integrates into host genome (provirus) • cDNA of provirus flanked by long-terminal repeats (LTRs) that can promote transcription • provirus integration near c-myc gene leads to increased constitutive expression and formation of B cell tumor

Enhancer Insertion • Provirus inserted downstream of myc gene, or upstream in opposite orientation • myc gene activated because enhancer sequences in LTR increase rate of transcription

Chromosomal Translocations • Many tumor cells exhibit chromosomal abnormalities • One type seen in cancer cells is translocation • piece of one chromosomes is split off and joined to another chromosome • if second chromosome donates material to the first, translocation is “reciprocal” • Burkitt’s lymphoma is fast-growing cancer of human B lymphocytes • In certain cases, chromosomes 8 and 14 involved in reciprocal translocation • Segment of chromosome 8 that moves to 14 contains c-myc • Transposition places inactive c-myc under influence of enhancers from immunoglobulin heavy chain genes

Gene Amplification • Amplification of certain genes is found in a number of tumors • Methotrexate, an inhibitor of dihydrofolate reductase (DHFR), is administered as an anticancer drug • Tumor cells that become resistant to the action of this drug amplify the gene for DHFR resulting in increased enzyme activity • Appear either as homogeneously staining regions (HSR) or as self-replicating double-minute chromosomes (lacking centromeres)

Point Mutation • DNA sequence of c-ras proto-oncogene from normal human cells and c-ras oncogene from human bladder cancer showed difference in only one base (resulting in amino acid substitution at Gly12) • Mutation results in loss of GTPase activity, may lead to persistent activation of MAPK pathway (mitogenic pathway)

Medical Biochemistry