Test 4

Other Potential Pathogen Cancer Links

Are there other pathogens that may cause cancer? The answer is that we really aren't sure. There are many studies that have very tentatively linked other pathogens to some cancers but the evidence is not terribly compelling: Chlamydia trachomatis, bacteria. This is actually one of the most common sexually spread bacterial infections. In women it has been linked to an increased risk of cervical cancer, perhaps by a synergistic action with HPV. This is probably one of the stronger associations with cancer in this entire list. Chlamydia pneumoniae, bacteria. A very common cause of pneumonia in humans, it has been suggested that infection may be linked to an increased risk of lung cancer. Chlamydia psittaci, bacteria. Sometimes called "parrot fever" as it is typically spread from birds to humans. There is some evidence that certain lymphomas may be linked to infection. Borrelia burgdorferi, bacteria. The cause of Lyme disease. May possibly be linked to some cases of cutaneous lymphoma. Salmonella typhi (typhoid fever) and S. enteritidis, bacteria. Linked to cancers of the gall bladder and colon, respectively. Mouse Mammary Tumor Virus. This is a virus that causes breast cancers in mice. Surprisingly, MMTV sequences are found in some human breast cancers but it is unknown if it is driving tumor development or not. Cytomegalovirus, aka Human beta-Herpes Virus 5. A very common virus, probably infecting 80 - 90% of the world population. Causes no symptoms in most individuals. Various studies have implicated CyMV infection in a variety of tumors including brain, colon, breast, prostate, sarcomas, and salivary gland. An emerging field of research with no clear answers yet. Feline Leukemia Virus. A common virus among domestic cats, there has been some concern that humans could become infected, potentially leading to leukemia. Most studies show that there are significant barriers to human infection and this probably isn't a real concern.

Hepatitis B (HBV) And Hepatitis C (HCV)

Both of these viruses are linked to liver cancer. In the United States, about 50% of liver cancers are linked to Hepatitis C while 15% are linked to Hepatitis B. In other parts of the world, especially sub-Saharan African and parts of Asia, over 50% of liver cancers are linked to Hepatitis B; in fact, about 80% of worldwide liver cancer cases are found in those areas. Hepatocellular carcinoma, most induced by Hepatitis B or C infection, is the second leading cause of cancer deaths on a worldwide basis. Hepatitis B. HBV (Hepatitis B Virus) is spread via contaminated blood products (not a risk in developed countries due to testing), unprotected sex, sharing of needles, improperly sterilized medical equipment, and from mother to child at birth. It is not spread via casual contact or breast feeding. Chronic infection with HBV increases the risk of liver cancer later in life. Most adults -90% or more - who are exposed to HBV will clear the virus. Children, on the other hand, are very likely to become chronically infected. 90% of children who exposed under the age of 6 (childbirth, infected sibling) will go on to develop lifelong, chronic infections. It is these individuals, the chronically infected, who are most at risk of developing cirrhosis of the liver and / or hepatocellular carcinoma. It is estimated that over 290 million individuals, worldwide, are chronically infected with HBV. Not surprisingly, it is those parts of the world with the highest chronic infection rates that also see the highest rates of liver cancer - particularly China, Mongolia, and sub-Saharan Africa. In the U.S., approximately 2 million are chronically infected. Despite the introduction of a vaccine in the 1980's, 80,000 people in the U.S. are estimated to become infected each year. HBV vaccination is required by almost all states for children and is recommended for adults if they are at high risk - for example, sexually active individuals not in a monogamous relationship, family members of HBV positive individuals, incarcerated individuals, victims of sexual assault, and health care workers at risk of exposure to blood and / or body fluids. Of course, anyone can request, and will be given, the vaccine. Carcinogenesis by HBV seems to occur by one of two mechanisms: Indirect / cirrhosis. In this case, chronic infection leads to inflammation of the liver and increased ROS production. This will contribute to cirrhosis and ultimately hepatocellular carcinoma. Cirrhosis is a condition in which normal liver tissue is replaced by fibrous scar tissue. As this occurs, the liver function begins to decrease, potentially resulting in liver failure and can be a precursor to full fledged hepatocellular carcinoma. Below are two images - on the left is the depiction of cirrhosis compared to a normal liver and on the right is a micrograph in which the fibrous scar tissue shows up with a blue stain: Direct. In this case, HBV proteins appear to directly influence cellular metabolism and physiology leading to hepatocellular carcinoma. In this case, cirrhosis DOES NOT APPEAR. A key viral protein seems to be the HBx protein. Phisiologically, HBx is believed to inhibit the activity of p53 AND promote telomere maintenance. Both of these could contribute to cancer development. Hepatitis D Virus can infect a person BUT ONLY IF THEY ARE ALREADY INFECTED WITH HBV. There is no good evidence as to whether co-infection with HDV increases the risk of liver cancer. Hepatitis C. Unlike HBV, HCV (Hepatitis C Virus) is not easily cleared by the immune system - up to 80% of those infected will become chronically infected. Like HBV, HCV is spread through unsafe sex practices or contact with contaminated blood. In the U.S. most new infections occur in intravenous drug users, but many were infected via blood transfusions prior to testing - HCV wasn't discovered until 1989 and testing of donated blood commenced in 1990. While there is no vaccine for HCV, there are effective drug therapies than can clear over 90% of those infected. HCV carcinogenesis appears to proceed exclusively through an inflammation / cirrhosis pathway virtually identical to the indirect pathway for HBV. You can see the pathway below: Up to 25% of chronically infected individuals will eventually develop cirrhosis. Of those that do, 1 - 4% of them will progress to hepatocellular carcinoma yearly. The 2020 Nobel Prize in physiology or medicine was awarded to three researchers - Harvey Alter, Michael Houghton and Charles Rice - whose work not only identified the Hepatitis C virus but proved it was the cause of both hepatitis and liver cancer. Believe it or not, this research began back in the 1960's when it was noted that many patients receiving blood transfusions went on to develop hepatitis. Dr. Alter, working at the U.S. National Institute of Health (NIH), showed that there was an unknown infectious disease causing these illnesses. Hepatitis A and B were ruled out so it was called non-A, non-B hepatitis. In 1989 Dr. Houghton, working for the pharmaceutical firm Chiron, used novel analytical techniques to isolate genetic material from this unknown agent. Analysis showed it was related to both Hepatitis A & B viruses so it was named Hepatitis C. Dr. Rice, working at Washington University in St. Louis in 1997, provided the last piece of the puzzle when he showed that injections of purified Hepatitis C could cause hepatitis in chimpanzees. As a result of their work blood transfusions became safer as tests for Hepatitis C were developed and, ultimately, the drug therapies we see today evolved. Note that their work spanned decades and their recognition (via the Nobel Prize) is probably long overdue.

Chemical Carcinogens

Cigarettes And PAH Humanity has had a relationship with tobacco for 1000's of years. Below are some questions and answers of the important and interesting milestones with regard to tobacco and smoking: How long has tobacco been around? Tobacco has been growing wild in the Americas for nearly 8000 years. Around 2,000 years ago tobacco began to be chewed and smoked during cultural or religious ceremonies and events. Who discovered tobacco and where? The first European to discover smoking was Christopher Columbus In 1531 tobacco was cultivated for the first time in Europe (at Santo Domingo). By 1600 tobacco use had spread across Europe and England and was being used as a monetary standard, a practice that continued throughout the following century By the 1700s smoking had become more widespread and a tobacco industry had developed When was tobacco first considered to be dangerous to health? In 1602 an anonymous English author published an essay titled Worke of Chimney Sweepers which stated that illnesses often seen in chimney sweepers were caused by soot and that tobacco may have similar effects. This is the earliest known instancesof smoking being linked to ill health and was made more than a century before Dr. Pott's official link of coal tar and cancer in chimney sweeps. In 1795 Samuel Thomas von Soemmering of Maine (Germany) reported that he was becoming more aware of cancers of the lip in pipe smokers In 1798 the US physician Benjamin Rush wrote on the medical dangers of tobacco During the 1920s the first medical reports linking smoking to lung cancer began to appear. Many newspaper editors refused to report these findings as they did not want to offend tobacco companies who advertised heavily in the media A series of major medical reports in the 1950s and 1960s confirmed that tobacco caused a range of serious diseases. When were cigarettes developed? Cigarette making machines were developed in the latter half of the 1800s. The first such machines produced about 200 cigarettes per minute (today's machines produce about 9,000 per minute). Cheap mass production and the use of cigarette advertising allowed tobacco companies to expand their markets during this period. What caused the growth and later decline of smoking in traditional markets? The prevalence of cigarette smoking continued to grow in the early 20th Century mainly as a result of: successful advertising programs, effective lobbying of lawmakers by the tobacco industry, the inclusion of cigarettes in ration packs for soldiers in WWI and WWII. Later in the twentieth century, smoking became less popular due to a rapid increase in knowledge of the health effects of both active and passive smoking. People also became aware of the tobacco industry's efforts to mislead the public about the health effects of smoking and to manipulate public policy for the short-term interests of the industry. The first successful lawsuits against tobacco companies over smoking-related illness happened in the latter part of the 20th Century. What are current global smoking trends? As smoking prevalence rates have declined in the traditional markets of North America and Western Europe, the tobacco industry has re-focused its promotional efforts onto the less developed and emerging nations in Africa, Asia, the Middle East, the former Soviet Union and Latin America. The often weak regulatory environment in these countries has further encouraged the industry to target populations in these nations. If current patterns continue, tobacco use will kill approximately 10 million people every year throughout the world by 2020; 70% of these deaths will occur in less developed and emerging nations. On January 11, 1964, Luther L. Terry, M.D., Surgeon General of the U.S. Public Health Service, released the first report of the Surgeon General's Advisory Committee on Smoking and Health. On the basis of more than 7,000 articles relating to smoking and disease already available at that time in the biomedical literature, the Advisory Committee concluded that cigarette smoking is— • A cause of lung cancer and laryngeal cancer in men • A probable cause of lung cancer in women • The most important cause of chronic bronchitis For several days, the report furnished newspaper headlines across the country and lead stories on television newscasts. Later it was ranked among the top news stories of 1964. Health-warning labels have been required to appear on all cigarette packages sold in the United States for more than 50 years. The Cigarette Labeling and Advertising Act of 1965 required the following health warning, prescribed by Congress, to be placed on all cigarette packages sold in the United States: CAUTION: CIGARETTE SMOKING MAY BE HAZARDOUS TO YOUR HEALTH. This warning appeared on cigarette packs from Jan. 1, 1966, through Oct. 31, 1970. In 1969, the Public Health Smoking Act of 1969 required all cigarette packaging contain the statement: WARNING: THE SURGEON GENERAL HAS DETERMINED THAT CIGARETTE SMOKING IS DANGEROUS TO YOUR HEALTH. This warning label appeared on cigarette packaging from Nov. 1, 1970, through Oct. 11, 1985. The Comprehensive Smoking Education Act of 1984 amended the Cigarette Labeling and Advertising Act by requiring cigarette manufacturers and importers to display on a quarterly rotating basis the following four health warnings on all cigarette packages: SURGEON GENERAL'S WARNING: Smoking Causes Lung Cancer, Heart Disease, Emphysema, and May Complicate Pregnancy SURGEON GENERAL'S WARNING: Quitting Smoking Now Greatly Reduces Serious Risks to Your Health SURGEON GENERAL'S WARNING: Smoking by Pregnant Women May Result in Fetal Injury, Premature Birth, and Low Birth Weight SURGEON GENERAL'S WARNING: Cigarette Smoke Contains Carbon Monoxide In addition to dried tobacco leaves, the manufacture of cigarettes adds multiple compounds to make them smoother and alter taste (think menthol). The smoke from these products is a complex mixture of chemicals produced by burning the tobacco and its additives. The combustion of tobacco results in over 7000 distinct chemical compounds of which about 70 are recognized carcinogens. Most of these carcinogens result from the burning (combustion) of the tobacco leaves. One of the single worst is a compound called benzo{a}pyrene, an example of a class of compounds called polyaromatic hydrocarbons (PAH): Based upon the above chemical structure, you can see why they are called PAHs - they are composed of multiple aromatic rings. PAHs such as benzo{a}pyrene are not unique to cigarette smoke - they are produced anytime something with carbon is burned (fuel, paper, diesel, wood, pot, etc.). Remember Percival Pott's link of coal tar and cancer? Coal tar contains a complex mix of PAHs. With cigarette smoke, however, this PAH is delivered straight to the lung tissue. Benzo{a}pyrene is not mutagenic in and of itself, rather, it is metabolized by enzymes in your cells into what is called an epoxide diol derivative: This is what is actually mutagenic and can ultimately become covalently bound to the bases of DNA resulting in a variety of mutations. Smokeless tobacco products include snuff and chewing tobacco that is put into the mouth or nose but is not burned like cigarettes or cigars. Still, smokeless products in the United States contain a variety of potentially harmful chemicals. These products are linked to cancers of the oral cavity. Smokeless tobacco products are less deadly than cigarettes. On average, they kill fewer people than cigarettes. Smokeless products are often promoted as a less harmful alternative to smoking, but they are still linked with cancer and can still be deadly. E-cigarettes and other electronic nicotine delivery systems (ENDS) are often used as substitutes for cigarettes or other tobacco products. Marketers of e-cigarettes and other ENDS often claim the ingredients are safe but the aerosols these products produce can contain addictive nicotine, flavorings, and a variety of other chemicals, some known to be toxic or to cause cancer. The levels of many of these substances appear to be lower than in traditional cigarettes, but the amounts of nicotine and other substances in these products can vary widely because they are not standardized. The long-term health effects of these devices are not known, but they are being studied. Certainly, some of the additives are damaging - it is believed that vitamin E additives were the cause of the severe lung damage many e-cigarette users suffered in 2019 - 2020. The IARC has declared cigarette smoking a class 1 carcinogenic agent. It has also declared "second hand smoke" to be a class 1 carcinogen. Second hand smoke is defined as smoke that has been both inhaled and exhaled by a smoker and is now in the environment. Non-smokers, in this environment, are now exposed to this second hand smoke and can inhale the same mutagenic agents as smokers. Recently, the topic of "third hand smoke" has arisen. Essentially, third hand smoke is defined as outgassed volatile compounds, into the environment, from cigarette smoke that has permeated clothing, fabrics, or other surfaces. For example, you can smell smoke on a smokers clothing and these compounds can volatilize into a closed environment potentially exposing other individuals. This topic is not well studied but you can read about "third hand smoke" in this article. Heterocyclic Amines (HCAs) Heterocyclic amines are a group of chemical compounds that can be formed during cooking meats by the reaction of amino acids and sugars. They are similar to PAHs except in addition to the aromatic rings, they also have at least one amine (-NH2) group. They are found in meats that are cooked to the "well done" stage, in pan drippings and in meat surfaces that show a brown or black crust. Below you can see a steak - HCA compounds will be present in the black, charred, grill marks where the meat achieved its highest temperature: The effects of HCAs and well-done cooked meat on humans are a bit ambiguous. Meat consumption, especially of well-done meat and meat cooked at a high temperature, can be used as an indirect measure of exposure to HCAs. Out of 22 studies looking at meat consumption, 18 showed a relationship between either meat intake or HCA exposure and some form of cancer. HCA exposure was measured in 10 of the studies and of those, 70% showed an association with cancer. It has been concluded that high intake of well-done meat and/or high exposure to certain HCAs may be associated with cancer of the colon, breast, prostate, pancreas, lung, stomach and esophagus. The IARC has labeled heterocylic amines as class 2B compounds - likely to cause cancer. Below are images of two HCAs: PhIP on the left and MelQ on the right: PhIP is one of the most abundant HCAs in cooked meat but MelQ seems to be a more potent carcinogen. Daily exposure to increasing levels of MelQ was linked to an increased risk of cancer but the relationship wasn't as strong for PhIP. Some Perspective We have discussed just a handful of exogenous carcinogenic agents. I wan't to point out, that especially with respect to chemicals, we are literally AWASH in potentially carcinogenic compounds, most of them occur naturally to some degree. Sure, smoking cigarettes produces PAHs but so does running a car and even forest fires. Below is a list of known or suspected carcinogens and where, naturally, you can find them: Acetaldehyde (apples, bread, coffee) Acrylamide (bread, rolls) Aflatoxin (nuts) Allyl isothiocyanate (arugula, broccoli) Aniline (carrots) Benzaldehyde (apples, coffee, tomatoes) Benzene (butter, coffee, roast beef) Benzo(a)pyrene (bread, coffee) Benzofuran (coffee) Benzyl acetate (jasmine tea) Caffeic acid (apples, carrots, celery,) Catechol (coffee) Coumarin (cinnamon in pies) 1,2,5,6-dibenz(a)anthracene (coffee) Estragole (apples, basil) Ethyl alcohol (bread, red wine, rolls) Ethyl acrylate (pineapple) Ethyl benzene (coffee) Ethyl carbamate (bread, rolls, red wine) Furan and furan derivatives (bread, onions, celery, mushrooms, sweet potatoes) Furfural (bread, coffee, nuts, rolls, sweet potatoes) Heterocyclic amines (roast beef, turkey) Hydrazines (mushrooms) Hydrogen peroxide (coffee, tomatoes) Hydroquinone (coffee) d-limonene (black pepper, mangos) 4-methylcatechol (coffee) Methyl eugenol (basil, cinnamon & nutmeg in pies) Psoralens (celery, parsley) Quercetin glycosides (apples, onions, tea, tomatoes) Safrole (nutmeg in apple and pumpkin pies, black pepper) Just how important are these exposures? Let's consider furfural, produced from flour when it is baked into bread. The average slice of bread contains 0.167 mg of furfural. To produce cancer in laboratory animals, you have to give rats about 197 mg per kg of body weight per day. For a human to get an equivalent does, they would have to eat about 82,000 slices of bread per day! Not realistic. The point I'm trying to make here is that yes, we are exposed to many agents in a given day, many of them naturally occurring. Stop worrying about what you can't prevent and think about what you can - protect yourself from UV, if you have a home in an at-risk area then test for radon, stop smoking, etc. This is where you can have some significant health effects. Another thing about carcinogens: often they do not cause cancer at all times, under all circumstances. In other words, a carcinogen does not always cause cancer in every person, every time there is any kind of exposure. Some may only be carcinogenic if a person is exposed in a certain way (for example, ingesting it as opposed to touching it). Some may only cause cancer in people who have a certain genetic makeup. Years ago there was an artificial sweetener called cyclamate that was pulled as a suspected carcinogen. Turns out it was only a POTENTIAL carcinogen if someone had a specific liver enzyme variant that would metabolize the sweetener in a specific manner. Some of these agents may lead to cancer after only a very small exposure, while others might require intense exposure over many years. Even if a substance or exposure is known or suspected to cause cancer, this does not necessarily mean that it can or should be avoided at all costs. For example, sunlight is a major source of ultraviolet (UV) rays, which are a known cause of skin cancer, but it's not practical (or advisable) to completely avoid the sun; you need at least some UV to manufacture vitamin D. Many commonly used medicines, particularly some hormones and drugs used to treat cancer, are listed as carcinogens. For example, tamoxifen increases the risk of certain kinds of uterine cancer, but it can be very useful in treating some breast cancers, which may be more important for some women.

Epstein-Barr Virus (EBV)

Epstein Barr virus, also called Human Herpes virus 4, is one of the most widespread viruses in the world. By the age of 30, probably 90% of the world's population will have been exposed. In developed countries, EBV infection may cause mononucleosis although many infections have no symptoms at all. Mononucleosis, also called "kissing disease" is spread by saliva: kissing OR sharing of utensils. It can also be spread by sexual activity. EBV infection has been linked to several cancers. EBV and Burkitt's Lymphoma Burkitt's lymphoma is a B-cell neoplasm. It is currently divided into 3 clinical sub-types: The endemic variant (also called "African variant") most commonly occurs in children living in malaria endemic regions of the world (e.g., equatorial Africa, Brazil, and Papua New Guinea); it is relatively rare in adults. Epstein-Barr virus (EBV) infection is found in nearly all patients. Chronic malaria is believed to reduce resistance to EBV, allowing the tumor to develop. The link between these two conditions can be seen in the image below. On the left is the distribution of Burkitt's lymphoma in Africa; darker blue represents more cases. On the right is the distribution of malaria (causal organism Plasmodium falciparum). Note that the greatest incidence of Burkitt's lymphoma mirrors the incidence of malaria: The disease characteristically involves the jaw or other facial bone, small intestine, ovaries, kidney, or breast. In parts of Africa, it may account for 30 - 50% of all childhood tumors. Below is an image of an African child with Burkitt's lymphoma: The sporadic type of Burkitt's lymphoma (also known as "non-African") is the most common variant found in places where malaria is not endemic. The tumor cells have a similar appearance to the cancer cells of classical endemic Burkitt's lymphoma. Only about 15% of these sporadic Burkitt's lymphoma are associated with EBV. The jaw is less commonly involved, compared to the endemic variant. Lymph nodes in the small intestine is the common site of involvement. Immunodeficiency-associated Burkitt's lymphoma is usually associated with HIV infection or occurs in the setting of post-transplant patients who are taking immunosuppressive drugs. Burkitt's lymphoma can be one of the diseases associated with the initial manifestation of AIDS. About 30% of these cases are linked to EBV infection. Treatment outcomes for Burkitt's lymphoma are generally good: up to 90% of patients are cured and survive long term. This assumes that treatment is available - a given in the developed world but not so in Africa, where this disease is common. Survival rates in Africa are substantially worse. EBV in other cancers Hodgkins lymphoma. In many cases, EBV DNA can be found in the Reed-Sternberg cell but the role of EBV is still uncertain. Some studies have suggested that EBV infection may be the root cause of up to 40% of cHL. It's role non-classic Hodgkins lymphoma is even less certain. Non-Hodgkins lymphoma. EBV is linked to a small number of B-cell NHL cases in both immunocompromised and non-immunocompromised individuals. Nasopharyngeal cancer. Occurring in the nasopharynx (see diagram in the section regarding HPV), some studies have indicated that EBV may be involved in most, if not all, nasopharyngeal cancers. These same studies have suggested that EBV infection, along with environmental exposures to other carcinogens, such as smoking, collectively work to trigger nasopharyngeal cancer. Gastric adenocarcinoma. Most studies looking at EBV and gastric carcinoma have been small and inconclusive. At best, maybe 10% of stomach cancers may involve EBV. Studies are ongoing. Carcinogenesis by EBV is complex and involves many different viral proteins - in fact, the type of cancer caused seems to depend upon the differential activity of several different viral proteins. One of the more important is EBN1A; this protein promotes carcinogenesis by inhibiting the activity of p53: In this way it is a bit different than HPV which promotes the degradation of p53 rather than inhibition.

Exogenous DNA Damage

Exogenous mutations are caused by the exposure to some agent that is capable of causing DNA mutations. These agents are thus mutagens AND, since most cancers are due to mutations, carcinogens. Carcinogenic agents are either radiation, of which there are two basic types, or some chemical compound. Whether or not something is declared carcinogenic is determined by studies conducted by many different regulatory bodies. Almost every government has its own agency to review safety of chemicals - in the United States it is called the National Toxicology Program, or NTP. In addition to agencies specific to a government, there is a body, The International Agency For Research On Cancer, or IARC, that will make carcinogen determinations. It is organized under the WHO (World Health Organization) and operates independently of any and all other governmental agencies. What this means is that one or perhaps several bodies may declare a compound carcinogenic while one or more others will not. For example, the active ingredient in the herbicide Roundup, called glyphosphate, has been deemed carcinogenic by the IARC but NOT by the NTP or the European Union regulatory body. How can divergence such as this happen? There are many possible reasons: One particular body may have studied the compound in question while another may not. The IARC acts independently from all other bodies and sometimes it may have studied compounds when the NTP (or some other governments agency) may not have. These regulatory agencies will only issue determinations on compounds they have studied - they won't just accept another agencies decision. Sometimes one regulatory body may be willing to accept a lower level of certainty, thus declaring a compound carcinogenic, while another agency may want a higher level of evidence. It is important to understand that all of these regulatory agencies - IARC, NTP, and all other governmental agencies (EU, Australia, etc.) act independently of one another. You can read about the glyphosate controversy in this paper. Anyway, for the purposes of this course, we will focus on carcinogens on the IARC list - the vast majority of them will be on same lists of other governmental agencies as well. For the purposes of this class, we will be looking at carcinogens as determined by the IARC. IARC Carcinogens Within the IARC carcinogen classification system, there are four different groups: Class 1: carcinogenic to humans. There are 120 agents currently on this list, including several infectious organisms, different forms of radiation, and a variety of chemicals. For these agents there is sufficient evidence, at least according to the IARC, to declare it carcinogenic in humans. Class 2A: probably carcinogenic to humans. There are 88 different agents in this group. This designation is applied when there is limited evidence of carcinogenicity in humans as well as sufficient evidence of carcinogenicity in experimental animals Class 2B: possibly carcinogenic to humans. There are 313 agents in this list. This category is used for agents for which there is limited evidence of carcinogenicity in humans and less than sufficient evidence of carcinogenicity in experimental animals. It may also be used when there is inadequate evidence of carcinogenicity in humans but there is sufficient evidence of carcinogenicity in experimental animals Class 3: not classifiable as carcinogenic. 499 agents fit this category. This category contains agents for which we have inadequate evidence for a determination OR the agent may be relatively new and don't have any historical evidence to make a determination. Below are two different links that will introduce you to the IARC classified agents. IARC Complete List: this document lists ALL the IARC agents that have been investigated and which category they fit in. I obviously don't expect you to know this but it is interesting to look through these agents. Some you will undoubtedly recognize and others will be new. IARC poster: this is a really interesting poster. It includes just class 1 agents BUT illustrates what part(s) of the body their cancers are associated with. Class 1 agents are in red. In green are things you can do to REDUCE your chances of cancer of that body part. You will probably have to download this pdf poster so you can blow up parts of it and clearly see everything. We will be discussing several of the class 1 agents in this module: radiation (both types; I'll explain shortly), one of the worst agents found in cigarette smoke, and a class of compounds called aromatic amines found in grilled meat. Each of you will also have to pick ANOTHER class 1 agent, do a little research on it, and present this info in the Discussion post for "DNA Damage And Cancer".

Intro to Radiation

Generally speaking, as the wavelength of radiation decreases, the energy possessed by that radiation increases. In other words, the shorter the wavelength, the greater the energy. Gamma radiation (produced by nuclear reactions, such as the sun, and decay events) is the most powerful, followed by X-rays, then UV (ultraviolet), and finally the visible portion of the spectrum. Wavelengths longer than UV don't really concern us as they are not energetic enough to damage DNA. Gamma and X-rays are collectively called ionizing radiation - they are powerful enough to create ions as they move through matter while UV is sometimes called non-ionizing because it lacks the energy to create ions upon passage through matter. Both ionizing and non-radiation are damaging to cells and DNA, just in different ways. We will focus upon UV, non-ionizing, radiation first.

Helicobacter pylori

Helicobacter pylori, or H. pylori, is a spiral-shaped bacterium that grows in the mucus layer that coats the inside of the human stomach. It is the only bacterial class 1 carcinogen. To survive in the harsh, acidic environment of the stomach, H. pylori secretes an enzyme called urease, which converts urea to ammonia. The production of ammonia around H. pylori neutralizes the acidity of the stomach, making it more hospitable for the bacterium. In addition, the helical shape of H. pylori allows it to burrow into the mucus layer, which is less acidic than the inside space, or lumen, of the stomach. H. pylori can also attach to the cells that line the inner surface of the stomach. You can see the bacteria, in a biopsy taken from stomach lining, in the image below. One of them is circled: Infection with this bacterium is common and, due to the location of the infection, the immune system really can't eliminate the bacteria - they are shielded by the mucus layer. The Centers for Disease Control and Prevention (CDC) estimates that approximately two-thirds of the world's population harbors the bacterium, with infection rates much higher in developing countries than in developed nations. Although H. pylori infection does not cause illness in most infected people, it is a major risk factor for peptic ulcer disease and is responsible for the majority of ulcers of the stomach and upper small intestine. Those carrying the bacteria have a 10 - 20% lifetime risk of peptic ulcer disease. The bacterial infection is amenable to antibiotic treatment, if diagnosed. In 1994, the International Agency for Research on Cancer classified H. pylori as a carcinogen in humans despite conflicting results at the time. Since then, it has been increasingly accepted that colonization of the stomach with H. pylori is an important cause of gastric cancer and of gastric mucosa-associated lymphoid tissue (MALT) lymphoma. Epidemiological studies indicated a 1 - 2% lifetime risk for gastric cancer and less than a 1% lifetime risk for gastric MALT. H. pylori is thought to spread through contaminated food and water and through direct mouth-to-mouth contact. In most populations, the bacterium is first acquired during childhood. Infection is more likely in children living in poverty, in crowded conditions, and in areas with poor sanitation. Infection with H. pylori generally leads to chronic infection and inflammation of the stomach - something termed "gastritis". A combination of this inflammation and potentially DNA damaging compounds produced by the bacteria can lead initially to a hyperplasia and ultimately to gastric cancer or MALT. You can see this in the image below:

HPV

Human papillomavirus (HPV) is the most common sexually transmitted infection in the United States. The relationship of cervical cancer and sexual behavior was suspected for more than 100 years and was established by epidemiologic studies in the 1960s. In the early 1980s, cervical cancer cells were demonstrated to contain HPV DNA. Epidemiologic studies showing a consistent association between HPV and cervical cancer were published in the 1990s. The first vaccine to prevent infection with four types of HPV was licensed in 2006. Below is an electron micrograph of HPV: Human papillomaviruses are small, DNA viruses that infect the epithelium; there are well over 100 known types of HPV, distinguished from one another by DNA sequence. Most HPV types infect the cutaneous epithelium (skin) and can cause common skin warts. About 40 types infect the mucosal epithelium (oral cavity, reproductive tract, etc.); these are categorized according to their epidemiologic association with, and risk of, cervical cancer. Since their initial association with cervical cancer, HPV types have also been linked to what are collectively called the anogenital cancers (penis, vagina, vulva, anus, and cervix) as well as cancers of the oral cavity and pharynx (oropharyngeal cancers). In the table below you can see what types of HPV are associated with different conditions: The table below shows you the annual numbers of the cancers that HPV is associated with AND the percentage of those cancers that are believed to be caused by HPV and not some other factor: The two highest risk strains of HPV, 16 & 18, collectively account for about 70% of all cervical cancers and probably most of the other ano-genital cancers; type 16 is believed to account for most of the oropharyngeal cancers too. The oropharynx is shown in the image below: essentially, it is at the back of the oral cavity: Infection with a high-risk HPV type is considered necessary for the development of cervical cancer, but by itself it is not sufficient to cause cancer (additional steps, and or factors are needed) because the vast majority of women with HPV infection do not develop cancer. The diagram below shows the timeline for development of cervical cancer IF HPV infection occurs. The steps in the development of the other HPV caused cancers are not as certain: Note that most infections are cleared by the immune system either almost immediately or over the course of a year or so. If a low-grade cervical abnormality does occur, it will regress if the immune system can clear the infection. Only in those women who remain chronically infected with HPV will develop cervical cancer and the process is believed to take, minimally 20 years or so. While no medical treatment exists for HPV, there are several vaccines that are approved to prevent HPV infection with some of the highest risk strains. While there are (3) vaccines currently approved for use in the United States, only one is actually available: Gardasil: The original vaccine approved in 2006. Protects against HPV strains 16 & 18 (most cervical cancers, other ano-genital cancers, and oropharyngeal cancer) and strains 6 & 11 (genital warts). Cervarix: Approved in 2009, Cervarix also protected against the same strains as Gardasil but appeared to provoke a better immune response and thus possibly better protection. Gardasil 9: Approved in 2018. Currently the only vaccine used in the U.S. Protects against 9 types of HPV (hence the name)For females 9 through 45 years of age for the prevention of cervical, vulvar, vaginal, anal, and oropharyngeal cancers caused by human papillomavirus (HPV) Types 16, 18, 31, 33, 45, 52, and 58; and genital warts caused by HPV Types 6 and 11.For males 9 through 45 years of age for the prevention of anal and oropharyngeal cancers caused by HPV Types 16, 18, 31, 33, 45, 52, and 58; and genital warts caused by HPV Types 6 and 11. This vaccine will NOT PROTECT against all HPV strains and, since some cancers are caused by other HPV strains, it will NOT PREVENT all HPV cancers. It will, however, prevent many if not most. The vaccine will not negate the need for regular cervical cancer screenings. Carcinogenesis via HPV infection appears to involve one of the viral proteins, E6, and its interaction with p53. So long as p53 is active within an epithelial cell, HPV reproduction cannot occur. Viral protein E6 complexes with and triggers the degradation of p53, allowing viral replication. This also triggers all the cellular physiological changes associated with the loss of p53 (review tumor suppressors if needed). It is believed that this loss of p53 is probably the first step in the progression to an HPV induced cancer. You can see this in the image below:

The Flukes

In humans, there are three species of "flukes", parasitic trematode flatworms, that are associated with cancer: Schistosoma haematobium, commonly called a blood fluke because, at some stage of its life, it can be found in blood. You can see its life cycle below: In humans, the adult flukes reside in the bladder. Eggs are shed via the urine. Juveniles require a snail host until free living cercariae are released into an aqueous environment. When humans come into contact with contaminated water, these cercariae penetrate the skin, go through several stages in the blood, and finally take up residence in the bladder, thus bringing us full circle. Clonorchis sinensis and Opisthorchis viverrini, called the liver flukes because they reside in the liver or adjacent tissue. You can see a life cycle for Clonorchis below, the life cycle for Opisthorchis is very similar. Adult Clonorchis take up residence in the bile duct and eggs are released in fecal material. Again, a snail is an intermediate host and the free living cercariae are released into water. Here is a difference between the blood and liver flukes - the liver flukes require a fish for the next part of their life cycle, becoming metacercariae. Humans are infected when they consume raw or undercooked fish, thus starting the cycle all over again. Infection results in a condition called schistosomiasis and chronic infection is strongly linked to bladder cancer. Egypt has one of the highest bladder cancer rates in the world it is thought to be due, at least in part, to S. haematobium. There are two other species of Schistosoma, S. japonicum (class 2B) and S. mansoni (currently class 3), that have been linked to cancer. The evidence is greater for S. japonicum (southeast Asia), as reflected in its class, than there is for S. mansoni (Africa, Asia, and South America). Both are tentatively linked to colon cancer. Both Clonorchis and Opisthorchis are found in Asia as indicated on the map below. C. sinensis is particularly common in Thailand while O. viverrini is a little more widespread through eastern and central Asia. Two locales, highlighted in yellow, have particularly high infections with O. viverrini. Both of these parasites are linked primarily to cholangiocarcinoma, cancer of the bile duct, as this where adults take up residence. C. sinensis is also linked to hepatocellular carcinoma. Note that I have indicated on the map another fluke - O. felineus. It is common through Russia and into central Europe. Many researchers are pushing for a reclassification of O. felineus from class 3 agent (what it is now) to a class 1 agent (known human carcinogen). This push is based upon research, primarily from Russia that suggests O. felineus is also responsible for cholangiocarcinoma.

Ionizing Radiation

Ionizing radiation is much more energetic and damaging to DNA, and cells in general, than is ultraviolet. Like ultraviolet, the IARC classifies ALL ionizing radiation as a class 1 carcinogen. As the name implies, ionizing radiation is powerful enough to produce ions from compounds it interacts with in cells. In humans, about 80% of the energy from ionizing radiation is absorbed by water, resulting two ionized species: oxoniumyl (water ion; positively charged ion) and a free electron (negatively charged). You can see this in the equation below: The water ion will directly produce hydroxyl while the free electrons indirectly produce it via the formation of peroxide and the Fenton reaction. Remember, the hydroxyl radical is the most damaging of all ROS and most studies suggest that about 65% of the damage done by ionizing radiation is a result of the hydroxyl radical. There are really two different forms of ionizing radiation, electromagnetic and particulate: Gamma rays and X-rays are examples of electromagnetic ionizing radiation. Gamma rays have the shortest wavelength (remember shortest wavelength means highest energy).and are produced by atomic decay events. X-rays are typically produced when high energy electrons impact metal. They are also electromagnetic but have longer wavelengths (and therefore less energy) than gamma rays. Particulate ionizing radiation consists of free atomic particles such as electrons, neutrons, and alpha particles (a helium nucleus; two protons and two neutrons). They are also produced by atomic decay events. Cosmic radiation is a type of particulate ionizing radiation - free protons and heavy metal nuclei. They are produced by stars, our sun and every other star in the universe. While both types are capable of doing damage, they vary in their ability to penetrate biological tissues. In the image below, you can see a visual depiction of this: Alpha particles are stopped by the skin. While potentially quite damaging to biological systems, unless they are ingested or inhaled, this lack of penetrating ability limits the damage they can do. Beta particles (free electrons) are a little more penetrating ability, requiring a sheet of tin foil to stop them so do pose a little more threat to humans. Neutrons. May require several meters of concrete to fully stop. Neutron radiation is quite damaging to biological systems. Gamma rays and X-rays. Both of these are highly penetrating and what is required to stop them depends upon the wavelength and thus energy. Medical grade X-rays require a thin layer of lead to stop them. That is why, when you have dental X-rays done, they cover your chest with the thin lead apron - to stop exposure to other body parts. Gamma rays, depending on their wavelength, may require several METERS of lead to stop. Ionizing radiation is capable of inducing, via the hydroxyl radical, all sorts of damage to DNA. The worst is what are called double strand breaks - when the DNA molecule is broken in two. You can see these breaks in the image below (the arrows indicate broken DNA molecules in the chromosomes): The newly broken strands of DNA have ends THAT ARE NOT PROTECTED BY TELOMERES; these broken, unprotected ends, are prone to fuse together producing a variety of structural aberrant chromosomes. Below is a diagram of just one possibility: the fusion of two different broken chromosomes into a structure called a di-centric (two centromeres) chromosome: These broken and structurally abnormal chromosomes disrupt the process of mitosis and can end up killing the cell or causing multiple mutations. This is how ionizing radiation can end up causing cancer. There are essentially two types of exposure to ionizing radiation possible: an acute, whole body dose (large amounts all at once), or chronic low-level exposure. Short of an atomic blast or nuclear accident, nobody receives acute doses but we are all exposed to chronic, low levels of radiation. In order to discuss its effects, we need to understand how ionizing radiation is measured: The rad, radiation absorbed dose. This reflects how much radiation is absorbed by a material. Today, the preferred unit is the SI unit the Gray (Gy). One Gy is one joule of absorbed energy per kilogram and equals 100 rad. While the Gy may be the preferred unit, a lot of biologically relevant work was done decades ago so you still see rad quite frequently. The rem, roentgen equivalent man. This takes the rad and multiplies it by a quality factor (Q) that reflects the biological damage potential of a type of radiation. For example, by definition, one rad of X-rays equals one rem but one rad of neutrons may equal anywhere from 5 to 20 rems because they have more potential to damage biological tissues. Today, the preferred unit is the Sievert (Sv) and one Sv equals 100 rem. Because rad and rem are still used throughout the U.S. government regulatory agencies, we will use those units in our discussion. Outside of the U.S., Gy and Sv are the norm. In the United States, the average person is exposed to about 620 millirems (mrem = one one thousandth of a rem) annually. Of course, depending on your circumstances you may get more or less. Below is a breakdown of where this comes from: Natural background sources - 310 mrem. Yep, half our annual exposure comes from natural sources, mostly from radon gas (we discussed this previously). Smaller amounts come from cosmic radiation and what is called internal radiation. All foods contain naturally radioactive isotopes of potassium, carbon, and other elements. When we consume them, they may be incorporated into our tissues, thus the concept of internal radiation. Did you know that bananas and potato chips are some of the most radioactive foods? This is due to their relatively high levels of potassium-40. The amounts are, in reality, quite tiny and internal radiation only contributes about 30 mrem of the 610 receive annually. Man-made sources - 310 mrem. This includes commercial and medical sources; by far the largest contributor here are medical procedures. If you actually have some of these medical procedures, you may get much more than the annual average 620 mrem . Below is a list of medical procedures and the amount of radiation you may receive: Other sources of man-made radiation include consumer products such as luminous watch dials (tritium), smoke detectors (some use small amounts of americium), granite counter tops (granite is igneous and is naturally enriched for radioactive isotopes of many elements), televisions, some ceramics, and cigarette smoke (polonium-210). The effects of all this exposure are debatable. Epidemiological studies indicate that exposure to one rem, at one time, is associated with about a 0.05% increased risk of cancer but chronic exposure is a little more tricky. It is generally assumed that any and all exposure may increase cancer risk but how much is unsure. Part of the issue is that the time period between any given exposure and cancer development may be years. Additionally, whatever proportion of cancers attributable to radiation as opposed to another cause may be so small that it is statistically impossible to determine. Acute Exposures So what about people who may receive much larger than average doses of radiation all at once. Studies on the atomic survivors in Japan have shown that leukemia rates increased for about 30 years post exposure then began to decline. This included all leukemias EXCEPT chronic lymphocytic leukemia (CLL). The Chernobyl disaster in 1986 saw a significant increase of thyroid cancer in children due to the release of radioactive iodine (among other radiation). Children are at particular risk because their thyroids are still developing and tend to take up a lot of iodine. Over 5000 extra cases of thyroid cancer were identified in the those exposed to Chernobyl fallout. Surprisingly, CLL seems to be occurring at an increased rate among personnel involved in the Chernobyl clean up. I say surprisingly as the work on atomic survivors indicated this one particular leukemia DID NOT increase in frequency. The WHO estimates that Chernobyl may ultimately cause 6000 deaths due to cancer; other organizations put the toll much higher - Greenpeace estimates at least 90,000 extra deaths. All in all, we may never know Chernobyl's effects on cancer rates not only in the surrounding area but in other parts of Western Europe contaminated by the fallout. Nuclear accidents are rated on a scale of 1 - 7 based upon severity and potential health effects. Until the Fukushima nuclear accident (Japan) in 2011, Chernobyl had been the only other "7" in history. The Fukushima accident was triggered by an earthquake and resulting tsunami. Due to far better construction techniques compared to the Russians, the Fukushima event released only about 20% as much radiation as did Chernobyl. While no one was killed as a direct result of the accident (Chernobyl had about 30 deaths as a result of the accident), almost 1600 died due to accidents and stress during the resulting evacuation. Based upon epidemiological models, it is estimated that Fukushima may result in 130 excess deaths due to cancer, a number we will never be able to detect statistically. Slightly increased levels of radiation were detected in pacific tuna but the amount is actually less than you would get in one banana. In fact, it was estimated that eating such fish, daily for one year, would be equivalent to spending about 23 seconds in a basement with average radon levels.

Human T-Lymphotropic Virus Type 1 (HTLV-1)

It has been nearly 40 years since human T-lymphotropic virus-1 (HTLV-1; also called Human T-cell leukemia virus), the first oncogenic virus in humans and the first demonstrable cause of cancer by an infectious agent, was discovered. HTLV-1 is spread by transfusion (rare since testing), unsafe sex practices, sharing of needles, or mother to child primarily via breastfeeding. As you can see from the map below, HTLV-1 infection is most common in Japan, parts of Africa and South American, and the Caribbean. In the United States, HTLV-1 is most common in the southeast and is often associated with Caribbean immigrant communities. Diseases associated with HTLV-1 seem to depend upon how it is acquired. Those infected as adults are more likely to develop a neurological manifestation called myelopathy / tropical spastic paraparesis. Individuals infected as children from their mothers are at risk of developing adult T-cell leukemia / lymphoma (ATL), an aggressive, almost always fatal, hematopoietic cancer. The risk of ATL may be as high as 25% among those infected as young children but does require a 30 - 50 year lag time (exposure to ATL). There are two viral genes that seem to be of importance in carcinogenesis, the tax gene and HBz gene. It is not clear which one is of primary importance BUT HTLV-2 (Human T-lymphotropic Virus Type 2), which also has a tax gene, is not generally considered carcinogenic. This would suggest that HBz is more important. The closely related HTLV-2 has, at various times, been associated with cancers. Decades ago, it was tentatively linked to hairy cell leukemia and more recently it has been suggested as the cause of some cases of cutaneous lymphoma (mycosis fungoides). Currently, it is not listed as a class 1 or even class 2 carcinogen. What we do know would indicate that those infected with HTLV-2 rarely, if ever, have symptoms.

HIV, KSHV, And Kaposis Sarcoma

Kaposis sarcoma is not a true sarcoma but is a neoplasm arising from cells lining lymphatic vessels (lymphatic endothelium). These neoplastic cells, called spindle cells (elongated shape), become highly vascularized masses, usually in the epidermal layers, that give the appearance of raised reddish - purplish lesions. The image below shows the multiple lesions of Kaposis sarcoma (left) and on the right the histology of a Kaposis sarcoma. A spindle cell is circled in yellow and a mass of blood vessels and red blood cells is circled in blue. Epidemiologically, there are four sub-types: Classic: Affects older males typically in the Mediterranean area, particularly Italy. The lesions often appear on the feet. The disease, while not curable, does respond to localized treatments and tends to proceed very slowly. Endemic: Usually affecting younger males in sub-Saharan Africa, this type tends to be more aggressive than classic Kaposis and infiltrates the skin extensively. Immunosuppression associated Kaposis: Found in organ transplant recipients who's immune system must be medically suppressed to avoid transplant rejection HIV associated: Kaposis sarcoma was one of the first illnesses first described in HIV-AIDS patients. Also due to immunosuppression because of HIV infection, it is about 300 times more common in HIV-AIDS patients than organ transplant recipients. There is one common thread between all types of Kaposis sarcoma - infection with Kaposis Sarcoma Herpes Virus (KSHV), also called Human Herpes Virus 8 (HHV-8). KSHV infection is quite common in the Mediterranean area and Africa, though it is unclear what contributing factors, along with KSHV, may exist for classic and endemic Kaposis. It is clear, though, for the last two types, immunosuppression allows KSHV to trigger Kaposis. So, why do HIV-AIDS patients have such a higher chance of Kaposis than transplant recipients? The HIV virus - one of its proteins, tat, seems to act synergistically with KSHV to promote Kaposis sarcoma development. It is generally believed that HIV type 1 is more likely than HIV type 2 to promote Kaposis due to slight differences in the tat protein. Generally, treatment of AIDS patients with anti-HIV drugs can effectively control Kaposis sarcoma. While Kaposis sarcoma is the only cancer linked to KSHV, HIV-1 has been determined to be a class 1 carcinogen and HIV-2 is listed as a class 2B carcinogen. These classifications are based upon the fact that HIV-AIDS patients are more likely to develop cancers, in addition to Kaposis sarcoma, than uninfected individuals. There are actually three cancers that are considered defining cancers for AIDS. This means that a person goes from "HIV infected" to a true diagnosis of "AIDS" if one of these cancers is diagnosed (there are other conditions that can also define AIDS, such as pneumocystis pneumonia, a fungal infection): Kaposis sarcoma Non-Hodgkins lymphoma Cervical cancer Note that all of these are linked to other viruses: Kaposis and KSHV, Non-hodgkins lymphoma and EBV, cervical cancer and HPV. It is believed that the immune suppression allows these other viruses to trigger the cancers. HIV-AIDS patients also seem to be at higher of other cancers, such as lung, Hodgkins lymphoma, melanoma and non-melanoma skin cancers, anal cancer (HPV infection?), testicular cancer, and liver cancer, although it is unclear what role HIV infection may play.

The Polyoma Viruses

Polyoma viruses are widespread and affect 80% of the human population. They are known to cause tumors in rodents but their carcinogenicity in humans is poorly understood at best. Only one polyoma virus, Merkel Cell Polyoma Virus, is considered a class 2A agent - probably carcinogenic in humans. This virus is believed to be responsible for an aggressive form skin cancer called Merkel Cell Carcinoma. In actuality, Merkel Cell Carcinoma isn't skin cancer as we may think of it - it involves cells of the nervous system that are embedded in the epidermal layers. They serve to sense pressure changes to the skin. You see this below: Merkel Cell Carcinoma looks a lot like basal cell carcinoma but, has a very poor survival rate - only about 40% at 5 years. I should point out that Merkel Cell Carcinoma is quite uncommon with only about 1500 cases per year. Compare this to over 60,000 cases of melanoma and over a million cases of non-melanoma skin cancers per year. Only two other polyoma viruses are suspected carcinogens (class 2B agents), the JC and BK virus. The JC virus is the causative agent of a central nervous system disorder and has been tenuously linked to colon cancer. The BK virus is mostly known for its role in nephritis in kidney transplant recipients but has also tentatively been linked to bladder cancer in immune suppressed individuals. There is one other polyoma virus that merits a brief discussion - SV40, simian virus 40 from Rhesus macaques. It is a class 3 agent and has been intensively studied for more than 50 years. The interest in SV40 goes back to the polio epidemic of the 1950's. Some batches of polio virus were inadvertently grown in monkey cells infected with SV40. Ironically, researchers didn't want to grow the vaccine virus in human cells because they were worried about contamination with unknown human pathogens. Between 1955 and 1962, thousands were vaccinated with contaminated doses. Then SV40 was found to cause cancer in hamsters. Later, SV40 infection in humans was tentatively linked to mesotheliomas (cancer of the lung lining), brain tumors, and bone cancers when researchers identified SV40 DNA in some of these cancers. Today, there is really no evidence supporting SV40 as a carcinogen - it is currently a class 3 agent. A number of epidemiological studies have found: Mesotheliomas are actually least frequent in those most likely to have been given the contaminated vaccine Those given the contaminated vaccine actually have fewer cancers, of all types, than those who weren't vaccinated In India, SV40(+) monkeys are quite common but there are very few of the suspected cancers that have any detectable SV40 DNA The original studies that linked SV40 to the cancers relied upon the detection of SV40 DNA in the tumors. It is not clear if it is really SV40 DNA or DNA from the JC or BK viruses, both common in humans and closely related to SV40.

Replication Errors And Driver Mutations

So, by now you should have gotten the idea that there are several sources of mutations that may be involved in cancer: Endogenous mutations; primarily replication errors Exogenous mutations caused by exposure to environmental agents. Heredity - remember the previous module discussing the inheritance of genetic damage and familial predispositions. So just how important are these sources? Do they contribute equally or is one more important than another? The short answer is the majority of driver mutations are the result of endogenous mutations, with DNA replication errors being the most important. Below is a table that, based upon a lot of epidemiological work, estimates the percentage of driver mutations due to replication errors (R), environmental exposure to exogenous agents (E), and inherited mutations (H): Note that, overall, two-thirds of driver mutations are the result of replication errors, 5% are due to heredity, and about 30% are due to environmental exposures to exogenous agents. This came as a surprise to most oncologists who had just assumed that most driver mutations would be the result of exogenous carcinogen exposure. Of course, this can vary tremendously based upon the precise type of cancer. Note that most driver mutations in melanoma are environmental in origin, specifically UV exposure, while in brain tumors almost 100% of driver mutations are due to replication errors. Cervical cancer's high environmental component is due to exposure to the HPV virus. Of all the tumors, only "eye" (read retinoblastoma) has a high inherited component, due to inherited defects in the pRB gene. Collectively, all this means that, on average, 66% of driver mutations are UNAVOIDABLE. This DOES NOT MEAN that 66% of cancers are unavoidable because we also have to consider the interaction between environmental agents and replication errors (hereditary plays such a small role in most cancers we can pretty much ignore it). Remember, most cancers require multiple driver mutations to become true neoplasms. Epidemiological studies suggest that perhaps as much as 42% of cancers could be avoided if we tried to minimize exposure to environmental agents, especially smoking and UV exposure. This would mean that at least some driver mutations will occur - due to replication errors - but we could minimize the occurrence of others, thereby preventing all necessary driver mutations from occurring.

DNA Damage Mitigation

So, we have talked about a lot of ways to damage DNA but can cells avoid or repair damage BEFORE it leads to a mutation. The answer is yes - remember the initial diagram that showed the potential outcomes of DNA damage; one of them was repair. Your cells have a number of mechanisms to repair genetic damage BEFORE it can lead to a mutation: Mis-match repair. We have already looked at this in the context of colorectal cancer Base excision repair. This system will repair damage done by ROS, hydrolytic damage, alkylation, and some other types of damage. In base excision repair (BER), the damaged base is removed in a two step process and replaced with an undamaged base. BER is capable of recognizing damage that does not change the geometry of the DNA helix. Nucleotide excision repair. This system can repair UV induced damage, damage done by PAHs, HCAs, and other DNA damaging events, SO LONG AS THE GEOMETRY OF THE HELIX IS DISTURBED - it won't be able to repair damage that doesn't alter the helix geometry. Another key difference between nucleotide excision repair (NER) and BER is that, in NER, the damaged bases AND SOME SURROUNDING UNDAMAGED BASES ARE REMOVED. This creates a gap that will have to be filled. In humans, about 30 total bases are removed and replaced during NER. As we have already looked at mis-match repair so will briefly discuss BER and NER. Base Excision Repair This repair system has several steps: Recognition of damaged base. Specialized enzymes called DNA glycosylases recognize a variety of damaged bases. They sever the bond between the base and the sugar in the DNA nucleotide, releasing it. The DNA phosphodiester backbone remains intact Removal of nucleotide remains. An enzyme called AP endonuclease will remove the remnants of the nucleotide - the sugar and phosphate. The result is a one nucleotide gap in the DNA strand Replacement of the nucleotide. Two enzymes are involved in the replacement of the nucleotide: DNA polymerase, which inserts an undamaged nucleotide, and DNA ligase which creates a phosphodiester bond to complete the replacement process. As I stated before, BER involves the removal of ONLY the damaged nucleotide. It is a very efficient process and can repair many types of DNA damage. Below is an image that shows the steps I detailed above: Nucleotide Excision Repair NER is a more complex process than BER and involves the removal of section the DNA strand, usually about 30 nucleotides in length, that contains damaged nucleotides. Additionally, NER is restricted to the repair of DNA damage that changes the geometry of the helix. This will include the dimers produced by UV (NER is the ONLY system we have for dimer repair) and some of the damage done by compounds like PAHs and HCAs. In NER, repair will begin by the recognition of the DNA damage by a protein called XPC and possibly another called XPE: Once the damage has been recognized, XPB and XPD are recruited. They unwind the DNA double helix in the area of the DNA damage. XPC and XPE will be replaced by XPA. Finally, XPG and XPF are recruited and create two incisions on the strand of DNA containing the damaged nucleotide(s). This results in the release of a short portion of the DNA strand (30 bases or so) that contains the damage but also surrounding undamaged nucleotides. This gap will be filled by DNA polymerase, restoring the DNA molecule to an undamaged state. You can see these steps below: You may be wondering why these proteins are called XP - this stands for Xeroderma Pigmentosum, an inherited condition in which patients are deficient in NER. Note there are seven different XP proteins involved in NER, thus there are seven sub-types of Xeroderma Pigmentosum (XP), one sub-type for each of the XP proteins. In other words, if you have inherited defects in the protein XPC, then you have Xeroderma Pigmentosum type C, if you have inherited defects in XPB, then you have Xeroderma Pigmentosum type B, etc. XP is inherited as a recessive condition, meaning that each parent must carry one defective allele for an XP protein; such offspring have a 25% chance of inheriting two recessive alleles and suffering from XP: Avoiding sun exposure (staying indoors, using sun-screen or heavy clothing, etc.) may help prolong life but the average life expectancy is less than 40. The most common cause of death is metastatic basal or squamous cell skin cancer. DNA Damage Avoidance In addition to the repair of damage, cells do have a few methods of avoiding DNA damage all together. For example, in epidermal tissue, melanin, produced by the melanocytes, can absorb UV light preventing damage to the skin cell. In the image below you can see a number of keratinocytes (a general term for a basal or squamous cell). I have circled the nucleus of two of them in red. Note the structure circled in yellow: it us a membrane bound vesicle containing melanin. See how it sits just atop the nucleus of the skin cell? In this position, UV will have to pass through the melanin first BEFORE impacting the DNA in the keratinocyte. Much of the UV will be absorbed by the melanin first, thus reducing the damage done to the keratinocyte's DNA. There are also detoxifying enzymes that can inactivate mutagens BEFORE they have a chance to do DNA damage. N-acteyl transferases. Using acetyl CoA, these enzymes transfer acetyl groups to HCAs, rendering them non-mutagenic. You can see the reaction below: It is known that natural variants of these enzymes may mediate this reaction "fast" or "slow". The so-called "slow" acetylators may allow HCAs to persist longer in cells and therefore do more DNA damage. People with "slow" variants may be more susceptible to bladder cancer if they smoke as the HCAs produced by tobacco linger longer in the urine. Epoxide hydrolases. These enzymes detoxify the epoxide derivatives produced from PAHs: Again, natural variants may mediate the reaction quickly or slowly; the slow variants have been implicated in increasing the risk of multiple types of cancer.

Endogenous DNA Damage

There are several processes by which endogenous mutations may occur. These include DNA replication errors, spontaneous damage to the bases of DNA, and metabolic damage via what are called reactive oxygen species (ROS). We will look at each in turn. Replication Errors We have looked at these in the context of familial colon cancer, namely Lynch Syndrome. It is estimated that UNCORRECTED replication errors (those that mismatch repair fails to correct) occur about once every billion bases replicated by DNA polymerase. I know that sounds like a very small number but think about this: there are about 6 billion base pairs of DNA per cell AND an estimated 30 trillion cells in an adult human body. Mathematically, this works out to 6 replication errors EVERY TIME A CELL DIVIDES and therefore, potentially, 180 trillion possible replication errors (6 errors per cell * 30 trillion cells) in our bodies - EVERY CELL CYCLE. Now, this is a vast over-estimate because not all our cells are actively dividing - many, like neurons, are arrested in G0 - but it does indicate that there is a large potential for replication errors to occur and lead to mutations. Spontaneous Base Damage We are talking here about hydrolytic damage and the spontaneous transfer of methyl groups to the bases of DNA. Given that water is the single most abundant compound in our cells, it is no surprise that the potential for hydrolytic damage (damage induced via base reaction with said water) exists. One of the most common types of hydrolytic damage is the deamination of cytosine: In this reaction, the amino group of cytosine is lost, via hydrolysis, and is replaced with the ketone group (double bonded oxygen). This reaction chemically converts cytosine to uracil. If this goes unrepaired, the potential exists for the uracil to lead to further base changes, and thus mutations, within the DNA molecule. While this type of deamination event is quite common, most of them can be efficiently repaired. The transfer of a methyl (-CH3) is a common event in biochemical reactions. The standard methyl donor is called S-adenosyl methionine, or SAM. You can see it below; the transferred methyl group is circled: Unfortunately, this methyl group can sometimes be spontaneously, and erroneously, transferred to the bases of DNA. Guanine is especially susceptible. Below is an image showing intact guanine and a methylated guanine: Like hydrolytic damage, if the methylation event is not corrected, the possibility exists for the damaged guanine to lead to further base changes and mutations. Our metabolism is oxygen based: mitochondria, using the electron transport chain to produce ATP, ultimately reduce molecular oxygen to water. One potential side effect of this is the production of reactive oxygen species, or ROS, that have the potential to lead to DNA damage. These ROS include the superoxide radical, hydrogen peroxide, and the hydroxyl radical: Of all of these, the hydroxyl radical is by far the worst. Please note, this IS NOT the same thing as the hydroxide ion. The hydroxide ion has a negative charge and typically doesn't last long in a cell - it picks up a proton to become water. The hydroxyl radical is an electrically neutral species with a single unpaired electron. It is HIGHLY reactive in a biological setting. There is a process called the Fenton reaction in which metallic cations catalyze the formation of hydroxyl from water: As you know, DNA stands for deoxyribonucleic acid. Acids are negatively charged and DNA is no exception. In the nucleus, DNA is closely associated with metallic cations (opposite charges attract) meaning the Fenton reaction can generate lots of hydroxyl radicals in close proximity to the DNA molecule. Interaction of DNA with hydroxyl radicals can lead to hundreds of different kinds of base damage. Some are lethal to the cell but others can lead to mutational events. The worst, i.e. the most mutagenic, type of hydroxyl induced base damage is probably 8-oxo-guanine: The circled oxygen is added by the hydroxyl radical and, if unrepaired, can lead to a lot of different mutations.

UV Radiation

UVC rays, with the shortest wavelength range, are the most powerful and, if they reached the surface of the earth, would essentially sterilize almost all life. Fortunately for us, they are 100% absorbed by the ozone layer BEFORE reaching earth's surface. UVB rays. Most UVB rays are absorbed by the ozone and only constitute about 5% of the total UV that reaches the earth's surface. They are powerful enough to directly damage DNA. These rays are the ones that produce a sunburn. UVA rays. 95% of the UV that reaches us is UVA. While UVA does not cause sunburns It is responsible for the aging and skin damage associated with sun exposure and can contribute to DNA damage. UVB interacts directly with DNA bases, specifically cytosine and thymine (the pyrimidine bases), creating what are called dimers. Basically, if two pyrimidines are next to one another ON THE SAME STRAND OF DNA, UVB can induce the formation of covalent bonds between them. Adjacent thymines are by far the most susceptible to dimer formation - they are called thymine - thymiine, or T-T, dimers, but you can also form (less frequently) C-T dimers and C-C dimers. The image below shows two of these dimers, specifically T-T dimers: The covalent bonds between the pyrimidines will distort the geometry of the helix potentially leading to mutations. UVA is not powerful enough to induce dimer formation - it appears to indirectly damage DNA by producing various ROS in tissues. Neither UVA or UVB is deeply penetrating and thus damage is limited to the skin. Both types of UV are linked to all types of skin cancer. There are a number of factors that can influence your UV exposure: Time of day: UV rays are strongest between 10 am and 4 pm. Season of the year: UV rays are stronger during spring and summer months. This is less of a factor near the equator. Distance from the equator (latitude): UV exposure goes down as you get farther from the equator. Altitude: More UV rays reach the ground at higher elevations. Clouds: The effect of clouds can vary, but what's important to know is that UV rays can get through to the ground, even on a cloudy day. Reflection off surfaces: UV rays can bounce off surfaces like water, sand, snow, pavement, or even grass, leading to an increase in UV exposure. To protect yourself from the damaging effects of UV exposure, the use of sunscreen is recommended. I'm sure you have all probably seen sunscreen before and recognize the acronym SPF, or Sun Protection Factor. SPF can be thought of as a guide as to how much time it would take for you to burn WITH the sunscreen as to opposed to WITHOUT it. Thus, SPF 30 means if it would take you 1 minute to burn without the sunscreen, it would take 30 minutes with it. In reality, many factors contribute to just how protective a sunscreen is: What type of skin do you have? Fair skinned people will burn faster than those with darker skin. What time of day is it? At noon, the sun is more intense than near sunset. Have you been swimming and washed off some of the sunscreen? Experts generally recommend re-applying sunscreen every two hours, sooner if you are in the water. Just FYI, you can purchase sunscreen with a maximum SPF of 50 in the United States. It is usually significantly more expensive than something with an SPF of 30 but provides only marginally better protection: SPF 30 blocks 97% of UVB while SPF 50 blocks 98%. Save some money and just apply a little more often. SPF refers only to protection against UVB. To ensure protection against UVA as well, you should buy a sunscreen that has a minimum of SPF 15 (UVB protection) AND ALSO SAYS BROAD SPECTRUM - this will provide protection against UVA. Last item: What about tanning beds? These usually emit mostly UVA but at an intensity about 20 times greater that what you are naturally exposed to. Both IARC and NTP classify the use of UV-emitting tanning devices (including sunlamps and tanning beds) as carcinogenic to humans; UV is thus a class 1 agent. The US Food and Drug Administration (FDA), which refers to all UV lamps used for tanning as "sunlamps," requires them to carry a label that states, "Attention: This sunlamp product should not be used on persons under the age of 18 years." The FDA also requires that user instructions and sales materials directed at consumers (including catalogs, specification sheets, descriptive brochures, and webpages) carry the following statements: Contraindication: This product is contraindicated for use on persons under the age of 18 years. Contraindication: This product must not be used if skin lesions or open wounds are present. Warning: This product should not be used on individuals who have had skin cancer or have a family history of skin cancer. Warning: Persons repeatedly exposed to UV radiation should be regularly evaluated for skin cancer. The FDA has also proposed a new rule to ban the use of indoor tanning devices by anyone under age 18, to require tanning facilities to inform adult users about the health risks of indoor tanning, and to require a signed risk acknowledgment from all users. Some US states have already banned indoor tanning by all people younger than 18, while others have banned use by younger teens and children. It is believed that the use of tannings beds is as dangerous (skin cancer) as smoking (lung cancer).

The Multi-Step Nature Of Cancer

We have talked a lot about DNA damage and its relationship to cancer. You need to understand that it takes more than one mutation in a critical gene to trigger a truly neoplastic growth. Remember the characteristics of a neoplastic cell and the concept of multiple driver mutations needed for a cancer. There is a model for this "multi-step" cancer development model and it was derived from studies of skin cancer development in mice. You can see a diagram illustrating these experiments below: Everything began with the application of a tumor "initiator" to the skin of mice. These initiators are mutagens and do some initial damage in critical genes. I arbitrarily picked ras but it could have been almost any proto-oncogene, tumor suppressor, or mutator gene. Even with this damage, unless the cells are stimulated, NOTHING WILL HAPPEN because normal growth controls can still over-ride this minimal damage. Next, a tumor promoter was applied. Tumor promoters are typically mitogens - they don't damage DNA but will stimulate a cell to divide. In normal, undamaged cells, growth circuits would prevent this artificially stimulated division BUT, in cells with the initial DNA damage, the promoter will stimulate mitosis; the initial genetic damage eliminates at least some normal control circuits such that the tumor promoter can induce some division. The result of the "promotion" was, in mice, the development of benign skin tumors. When the tumor promoter was removed, they regressed. Application of more mutagen - perhaps the same one used initially or another one - allows for the accumulation of more genetic damage; I arbitrarily picked p53. This additional damage will allow for the cells to divide EVEN IN THE ABSENCE OF A MITOGEN (promoter). The cells have undergone "progression" and now have the characteristics of a neoplastic cell, namely they don't respond to normal growth control circuits and can grow even without stimulatory signals. In a nutshell, cancer therefore has a three step development process: Initiation: exposure to a mutagen (or spontaneous mutation) causes some initial driver mutation Promotion: exposure to growth signals allows the damaged cell to begin dividing. Promoters DO NOT directly damage DNA but do stimulate cell division, possibly allowing the accumulation of more mutations via replication errors Progression: ultimately enough driver mutations have been accumulated that the cells can grow independently of stimulatory signals - a true neoplasm. In humans, initiation may occur via endogenous or exogenous mutagenic mechanisms. It is also believed that inflammation is a powerful and important human tumor promoter. You can review the link between inflammation and cancer in module 2 if you wish. Progression could then be the result of additional mutations caused by endo- or exogenous means. This multi-step model means that cancer is the end result of a years or decades long process and explains why most cancers occur in older patients. Various models have suggested that most breast and colon cancers begin developing at least 10 years before they are diagnosed; prostate cancers may take several decades to grow to the point of detection. As our knowledge of tumor development and genetics increases, we are beginning to build models that link specific genetic changes to specific stages of tumor development. We have one of the best models for colon cancer development: APC gene. Remember, the APC gene is lost in virtually all colorectal cancers. It is believed that colon cancer won't even start to develop until an epithelial cell loses APC expression. COX-2 overexpression. COX-2 is an enzyme involved in inflammation. It is believed that overexpression of COX-2 may act as a tumor promoter, stimulating the initiated epithelium (loss of APC) to become hyperproliferative - reproducing in excess. This then leads to an early stage, and benign, adenoma. Mutations in one of the ras proto-oncogenes (usually K-ras) are probably involved in the transition of an early adenoma to a mid stage adenoma (still benign). Transition to a late stage, but still benign adenoma, may involve mutations in the Smad4 gene or the DCC gene. DCC is a tumor suppressor that has been found to be mutated in about 70% of colorectal cancers. Loss of p53 appears to be involved in the transition of a late stage, benign adenoma to a truly neoplastic and malignant adenocarcinoma. It is unclear what mutation(s) may occur to confer metastatic potential on the now neoplastic adenocarcinoma. All of these mutations take time to accumulate, hence the decade (or longer) developmental time frame. This also explains why, if you have a clear colonoscopy, you typically don't have to get another one for a number of years.