7.013 Recitation 13 Notes

7.013 Recitation 13 (Note: The recitation summary should NOT be regarded as the substitute for lectures) Summary of Lectures 21 (4/6), 22 (4/9) and 23 (4/11) Cancer causing agents are called carcinogens and they are mostly mutagens or pro- mutagens (they do not cause cancer in their native form but can be metabolized by the enzymes, specially the enzymes in the liver into a cancer causing form). Ames test: Ames test is a widely employed method that uses bacteria (His–) or that cannot produce amino acid histidine and therefore need histidine in the growth medium to grow and divide) to test whether a given chemical can cause mutations in the DNA of the test organism (in this case the Hisbacteria). More formally it is the biological assay to test the mutagenic (and hence the carcinogenic) potential of a test compound. In Standard Ames test, you take your test compound and incubate it with the His- bacterial cells. Then you plate the treated bacterial cells on a plate in which the growth medium lacks the histidine. You look for the bacterial colonies that have reverted from the His- phenotype to His+ phenotype. The more the number of colonies the more is the mutagenic potential of the test compound. There are some chemicals, which are not mutagenic in their original (native state). But once they are injested by organisms they may be metabolized into metabolites that can be mutagenic. Such chemicals are called PROMUTAGENS and they can be detected by the modified Ames test. Here the test compound is first treated with the liver extract, which supposedly should have the enzymes needed for metabolizing the test compound. The His- bacterial cells are then treated with the liver extract treated mixture of test chemical and its metabolites are then plated on the His- plate to look for the number of His+ bacterial colonies. Please check the following link: testhttp://study.com/academy/lesson/the-ames-test-using-bacteria-to-test-for-carcinogens.html © source unknown. All rights reserved. Carcinogenicity assay: Here you inject the test chemical into mice regularly for a period of time and look for the development of tumor. You can also take the cells from the developed tumor, culture them in plates and see if they loose contact inhibition and grow on top of each other to foci. Ames test (original and modified) using bacterial mutants is often used to detect the carcinogenic/ mutagenic potential of different chemicals. Oncogenes, proto- oncogenes and Tumor suppressor genes: Tumor suppressors genes and proto- oncogenes are normal genes that work in a regulated fashion in a normal cell to properly control the cell cycle. The wild-type function of a tumor suppressor gene is to inhibit the cell cycle in any cell that is not supposed to be actively growing and dividing. Both homologous versions of a tumor suppressor gene must loose their function to transform a normal cell to a cancerous type. The wild-type function of an oncogene is to promote the cell cycle in any cell that is supposed to be actively growing and dividing. One of the two homologous versions of an oncogene must gain a function or increase its function for a cell to become cancerous. Normal cellular counterparts of the oncogenes are called the proto- oncogenes. Some of these genes are carried by oncogenic viruses and are designated as v-oncogenes. The v– oncogenes can be linked to potent promoters that lead to their inappropriate and high level expression, leading to deregulated cell division. One example is the Rous sarcoma virus (RSV). This retrovirus infects the chickens, thereby causing them to acquire tumors. Here the viral genome contains a gene that it has stolen at some point from a host cell. This gene is an oncogene called src that is involved in cell signaling. The virus carries a mutant version of src that produces an overactive form of the normal cellular kinase src. When RSV infects a cell, the mutant src is transcribed and translated, creating an overactive cell signaling protein that promotes growth and division in chicken cells to form tumors. Other examples include the avian leukemia virus that causes leukemia and human papilloma virus (HPV) responsible for cervical cancer. Retinoblastoma: This is a cancer of the retina. In Familial retinoblastoma, multiple tumors in the retinas of both eyes occur in the first weeks of infancy when the fetus inherits from one of its parents a chromosome that has its RB locus deleted or otherwise mutated. So in this form of the disease, a germline mutation plus a somatic mutation of the second allele leads to the disease. In sporadic retinoblastoma a single tumor appears in one eye sometime in early childhood before the retina is fully developed and mitosis in it ceases. In this form, both inherited RB genes are normal and a single cell must be so unlucky as to suffer a somatic mutation (often a deletion) in both in order to develop into a tumor. Such a double hit is an exceedingly improbable event, and so only rarely will such a tumor occur. Cancer Therapy: Most cancer patients are treated with some combination of surgery, radiation, chemotherapy or immunotherapy. Radiation and chemotherapy have the disadvantage of destroying healthy as well as malignant cells and thus can cause severe side- effects. Drug design is a very expensive process. Drugs must be specific (i.e. they can’t inhibit other proteins in addition to their targets or else they will cause side effects), must work at a low concentration (so that the amount that needs to be taken by the patient is feasible), and must not be metabolized by the patient either too quickly (so that taking the drug is ineffectual), too slowly, or into toxic byproducts. Gleevec is a drug that works against the type of cancer of the white blood cells called CML (chronic myelogenous leukemia). Almost all patients with CML contain cancerous cells that harbor a mutation that is a translocation between chromosomes #9 and #22. The genes at the two breakpoints of the translocation are Bcr and Abl (a kinase). The translocation causes a fusion of these genes to be created, such that a chimeric protein is made that is one continuous protein, half of which looks like part of BCR and half of which looks like part of ABL. The BCR-ABL fusion protein that is produced by this mutation functions aberrantly by phosphorylating proteins it normally shouldn’t, thereby activating the growth and division of white blood cells. The drug Gleevec specifically inhibits the kinase activity of Abl, thereby ceasing the aberrant phosphorylation in CML patients and sending their cancer into remission. The success of Gardasil is an example of preventive therapy for head and neck as well as cervical cancer. Heceptin is another example of immunotherapy for Her-2 positive breast cancer patients. Some less aggressive forms of breast cancer are not associated with an overexpression of the Her2 gene, so they are not sensitive to Herceptin. Often the aggressive forms of breast cancer, are associated with an amplification of the Her 2 gene. The amplification of the Her2 gene correlates with the increased expression of receptor on cell surface, which increases the proliferation signal that is critical for tumor development. Herceptin is a monoclonal antibody that works on both the extracellular and the intracellular domains of the HER2 receptor. It does so by binding to the Her2 receptors that are expressed on the surface of cancerous cells thereby flagging these cells for destruction by the immune system. At the same time herceptin blocks the downstream signaling by the Her2 receptors thereby preventing tumor proliferation.