Carcinogens

Introduction and meaning

Carcinogens are substances capable of causing cancer. By this definition, it means that not all substances labelled to be carcinogens would cause cancer in every person as there are other factors that play great roles in carcinogenesis. Both genetic and environmental factors contribute to cancer. Occupational cancers generally develop slowly.

Factors that makes carcinogens cause cancer

The development of cancer from substances labelled carcinogens depend on so many factors such as:

1. how they come in contact with us (inhalation, contact, oral),

2. the nature of the carcinogens exposed

3. The contact or exposure period

4. the amount of the carcinogen taken or exposed to

5. The duration or the time it spends in the body system before being getting rid of

6. Variation of individual compounds to give rise to tumor in different species

7. Individual differences (physiology, genetic, occupation)

Types of Carcinogens

Carcinogens may be grouped into the following

1. Genetic carcinogens

2. Physical agents: environmental, behavioural, occupational

3. Chemical agents: which generally is referred to as carcinogens

Mechanism of action of carcinogens

Mechanism of action of carcinogens discusses how they actually lead to cancer.

• The common features of parent compound or a metabolite is its ability to combine with nuclear DNA and with protein component of the cell.
• The neoplastic state may then arise either through a somatic mutation or an alteration in the surface properties of the cell.
• Then the end result is the development of a clone of cells that have lost the ability to respond to the complex growth controlling mechanism, characteristic differentiated tissues and cellular multiplication.
• These steps make cells become unrestricted in growth and differentiation thus becoming cancerous.

Sources of carcinogens

Physical agents:

• UV light,

• X-rays,

• gamma rays,

• α and β particles given off by radioactive atoms.

The UV component of sunlight penetrates the skin and modifies DNA in the nucleus of the germinal epithelium.

Examples of cancer caused by physical carcinogens are:

• X-ray therapy of an enlarged thymus in children resulting in thyroid cancer

• therapy of ankylossing spondylitis caused leukemia,

• radium and mesothorium sequestered by bone cause sarcomata.

• radioactive strotinum sequestered by bone cause bone tumor and

• leukemia transformation of bone marrow.

• Radioactiveiodine 131 for treatment of hyperthyroidism results in the development of cancer of the thyroid.

• Physical injury is another possible cause of tumor development because the repair of tissues after damage is associated with increased mitosis and therefore probability of spontaneous mutation.

• It is thought that burns and bone injuries are particularly carcinogenic.

Chemical agents

These are substances that have been shown to cause or promote the occurrence of tumor and are generally referred to as oncogenic chemicals. Most of them give rise to carcinomata and are commonly called carcinogenic chemicals or carcinogens. For the fact that they can also cause sarcomata or other tumors, they are generally described as carcinogenic substances.

Mechanism of action of Chemical carcinogens

Carcinogens act by suppressing immune mechanism or by reactivating dormant oncogenic viruses. Others act by exerting their action directly on the cells The incidence of tumor produced by a given carcinogens differs widely between different species and strains of animals, sex, hormonal balance, diet, environmental factors.

The chemical induction of tumors involves two procedures

1. Initiation and

2. Promotion.

Initiation is the production of an irreversible cellular change that is necessary but by itself insufficient to induce tumor growth.
Promotion is the process whereby a tumor develops in a tissue in which initiation has already occurred.
Carcinogens may be an initiating or promoting agents or they may posses both properties.

Initiating agents

Initiating agents are thought to act either at the cell surface whereby they may interfere with the cells immunological response or with nuclear DNA to produce a somatic mutation that result in loss of properties of content inhibition and responsiveness to other controlling mechanism.

Promoting agents

Promoting agents or co-carcinogens are generally non-specific irritant agents that produce all damage in the area where malignancy will eventually arise. Promoting agents act by inhibiting DNA repair mechanism or altering cellular environment in such a way that a clone of transformed neoplastic cells is allowed to develop.

Classes of Chemical carcinogens

The main classes of chemical carcinogens are:

• Polycyclic hydrocarbons

e.g. 1,2:5,6-dibenzanthracene and benzo(a) pyrene

• Alkylating agents

Although these agents are among the drugs used in the chemotherapy of cancer, like X-ray, they are also carcinogenic, mutagenic, teratogenic and immunosuppressive.

Examples of Alkylating Agents

• nitrogen mustard,
• ethyleneimines
• alkylsulphonates
• nitromycin
• antibiotics others include ethionine-analogue of amino acid (methionine)

Classes of Chemical carcinogens

Pyrrolizidine alkaloids

These occur in various species of plants and they produce acute hepatotoxic reaction but chronic exposure lead to liver damage.

Examples of pyrrolizidine alkaloids

• Lactones and β-propiolactone,
• parasorbic acid and aflatoxin (fungus)
• Safrole – oil of sassafras,
• nutmeg and cinnamon,
• shikimic acid-bracken and many other plants (liver cancer)

Amines and amides-associated with workers in dye industry.

2-naphthylamine

This which is used extensively in dye industry is not itself carcinogenic but its principle metabolites 2-amino-1-naphthol and 2-naphthyhydroxylamine are potent carcinogens (bladder cancer).

Certain azo dyes

An example is butter yellow (p-dimethylaminoazo benzene) which was formerly used in food industry as coloring agents (liver tumor).

Nitrosamine-alkyamino compounds

This was used as preservatives and colour fixatives in fish and meat products (liver and kidney tumors)

Metallic elements

Examples of these include

• arsenic
• nickel
• beryllium
• cadmium
• cobalt
• chromium
• zinc
• lead
• iron
• chloroform and
• trichloroethylene (liver, kidney and thyroid tumors)

Individuals are exposed to chemical carcinogens as a result of their occupation or particular behavioral pattern.

Occupational factors

Types of tumors and their occupational implication

• Scrotal skin cancer:  chimney sweepers
• Pleural/peritoneum cancer: asbestors
• Bladder cancer: rubber workers, cable & drug manufacturer
• Cancer of the nasal/ mucosa/antrum:  furniture makers
• Angiosarcoma of liver: plastic industry
• Stomach cancer/lung cancer: coal miners

Behavioural factors

Carcinogens and types of cancer likely cause

• cigarette smoking: benzo(a)pyrene likely causing lung cancer
• Dibenzylpyrene: likely causing buccal cavity cancer
• Nitrosamines: nasal/mucosa cancer
• 1,2:5,6-dibenzathracene: skin cancer
• Radioactive Polonium (210Po): antrum cancer
• Ethanol: likely causing mouth, larynx and oesophagus cancer
• Sexual intercourse (via viral carcinogens contraction): uterine cervix carcinoma in women and Prostate cancer in men

NB: Smegma/secretion that collects under prepuce of penis contain carcinogens. Carcinoma of the cervix is common among women whose regular partner are uncircumcised than those who are circumcised.

Mutagens

Introduction and meaning

Mutagenesis and carcinogenesis are different expressions of the same mechanism.
Most carcinogens can induce mutations and many mutagens have been shown to induce tumors.
Mutagens are substances that have the ability to cause gene mutation or chromosome aberration (anomaly).
Mutations that are produced in germ cells of individual prior to or during the reproductive period may be capable of transmission to later generations.
Changes confined to the genetic material of somatic (i.e. non germinal) cells cannot be transmitted to the next generation but may give rise to teratogenic effects if induced in the embryo or tumor growth if they occur at a later stage of development.

Test for mutagenicity

No single test or battery of tests is likely to detect and characterize all mutagenic agents, a variety of in-vitro and in-vivo non mammalian test systems are used.
Other techniques include

• human cell culture tests and
• chromosome examination of somatic and germ cells.

Test for mutagenicity involve

1. Examination of chromosomal aberrations and

2. Examination for gene mutations

Test for chromosomal aberration usually make use of in-vitro preparation of human peripheral lymphocytes and in-vivo studies of effects on the testicular cells of a mammalian laboratory animal.
Gene mutations are usually studied in bacterial cultures (S. typhimurium or E.coli), yeasts, Drosophile and mammalian cell cultures. Many chemicals that are mutagenic owe their action to a metabolite produced in the liver.
Mutagenicity tests have value as a prescreening procedure for tumor.

Types of Mutagenicity tests

PCR Based Methods

These methods are:

• Site-directed mutagenesis

• Mismatched mutagenesis

• 5’ add on mutagenesis

• Cassette Mutagenesis

Insertional Mutagenesis

• Trasposon mutagenesis

In vivo Mutagenesis

• Direct Mutagenesis

Mismatched Mutagenesis

• This is similar to Site-Directed Mutagenesis but only focuses on a single amino acid.
• It Important when trying to determine a particular missense mutation in known gene of a disease.
• Or when just trying to evaluate the contribution of the single amino acid to the function of the protein.

5’ add on Mutagenesis

This involves adding on a new sequence or chemical group to the 5’end of a PCR product.
This involves a particular way of designing the primers when the

• 3’ end of the primer matches the sequence of PCR product.
• 5’ end contains the novel sequence.
• Suitable restriction site
• Addition of a functional sequence (promoter sequence)
• Modified nucleotide that contains labeled group, biotinlyated, or fluorophore.

Uses and Limitations

• PCR based methods are useful in making specific mutations in the DNA which is useful when studying different aspects of protein function.
• With PCR based methods it is hard to replicate the mutated DNA.
• In order for replication to occur, super competent cells must be used and these are expensive!
• Screening can be tedious, usually requires sequencing to confirm if mutation occurred.

Cassette Mutagenesis

• This is used to introduce multiple mutations into the DNA sequence.
• It uses blunt ended DNA for insertion site of mutation.
• Where mutation is inserted, a 3 base pair direct terminal repeat is created.
• The mutagenic codon cassette has two head to head SapI sites allowing for removal of all DNA except for mutation

Uses and Limitations

• It is typically used for protein structure but possibly used for gene function
• It is less expensive than site directed mutagenesis to create several mutations because there is no need for primers
• It requires the SapI restriction enzyme cut sites, and other cut sites flanking the target region for removal of DNA
• It works best when target region is contained in a small DNA fragment

Transposon Mutagenesis

• Transposon is a piece of short DNA that replicates by inserting into other pieces of DNA (plasmids, chromosomes, etc)
• It useful for studying gene function because when the transposon moves into different location in the DNA, it may cause a disruption in a gene or a set of genes.

Transposons also have many useful properties for mutagenesis as listed below

• Cause clean mutations
• Can be random or specific mutations
• Typically encode for antibiotic resistance or some other advantageous gene.
• Can use a transposon that inserts at a high frequency
• When used in bacteria, it causes selectable phenotypes
• Recognize specific sequence that is ~2-12 base pairs long

Uses and Limitations

• Primary use of transposon is for the study of gene function, though can be used to create gene fusions
• It’s usually easy to see a change in phenotype due to gene knockout
• Because the transposon inserts at a specific sequence, helps in determining where insertion occurred
• Not useful in large plasmids because many recognition sites could be contained in the single plasmid
• Suicide vectors are used, though some may have limited replication, so further screening is needed