ScienceWeek

2. ORIGINS OF CANCER

ON MUTATIONS AND THE ODDS OF DEVELOPING CANCER

K.C. Quon and A. Berns (The Netherlands Cancer Institute, NL) discuss mutations and cancer, the authors making the following points:

1) During the course of cancer development, a normal cell progresses toward malignancy by acquiring a specific series of mutations. These include mutations that activate otherwise innocuous proto-oncogenes, and other mutations that inactivate recessive tumor suppressor genes. By acquiring these mutations, a cell progressively alters its phenotype, and thereby eludes the various controls that normally prevent malignant growth in an organism.

2) Based on epidemiological data, and consistent with in-vitro experimental data, it is estimated that between 4 and 8 rate-limiting mutations occur during the development of most human cancers. But this raises a conundrum. The incidence of cancer should be proportional to the number of rate-limiting events necessary for tumorigenesis, the frequency of these events, and the size of the target-cell population for these events. Therefore, given that somatic mutations arise at a frequency of less than 6 x 10^(-6) per locus, an overly simplistic calculation would suggest that even a tumor requiring only 4 mutations would only arise at a frequency of approximately 1 in 10^(-21) cells, a vanishingly low frequency even in an organism composed of approximately 10^(14) cells, as humans are. Why, then, are the odds of developing cancer during one's lifetime approximately 1 in 3, and what does this tell us about the mechanisms that operate during tumorigenesis?

Genes & Development 2001 15:2917

Related Background:

MEDICAL BIOLOGY: ON MUTATIONS AND CANCER

The term "cancer", which means "crab" in Latin, was introduced by Hippocrates (460-370 B.C.) to describe diseases in which tissues grow and spread unrestrained throughout the body, eventually causing death. Cancers can originate in almost any tissue of the body, including nerve, muscle, blood, connective tissue, etc. Depending on the cell type involved, cancers are grouped into 3 main categories: a) carcinomas, the most common types of cancer, arise from the *epithelial cells that cover external and internal body surfaces, with lung, breast, and colon cancers the most frequent cancers of this type; b) sarcomas originate in supporting tissues of *mesodermal origin, such as bone, cartilage, fat, connective tissue, and muscle; c) lymphomas and leukemias arise from cells of blood and *lymphatic origin, the term "leukemia" used when such cancer cells circulate in large numbers in the bloodstream rather than growing mainly as solid masses of tissue. Cancer is a disease of the genomic apparatus of the cell, in particular of the growth-regulation apparatus, and considering the vast number of activities that must be coordinated and regulated by the genomic apparatus during the lifetime of each cell, it is not surprising that malfunctions arise. In general, cancer is the most prominent of the many diseases arising from aberrations in cell function, with more than 25 percent of people in the US now expected to develop cancer in their lifetime.

C.R. Boland and L. Ricciardiello (2 installations, US) present a review of current research on the genomic basis of cancer, the authors making the following points:

1) It has been known during most of this century that cancer is often associated with visible derangements in the nucleus of the cell. The cells of solid tumors commonly exhibit chromosome duplications, deletions, and rearrangements, but before the organization of the human cell nucleus was understood, these chromosome aberrations were difficult to categorize and were of little help in understanding the biological basis of cancer.

2) Within a few decades after the discovery of the structure of DNA, cancer-related genes (oncogenes) were isolated, and these were frequently found to be mutant versions of normal cellular genes in which an activating *point mutation or an aberrant *genetic amplification process resulted in a gain of function for that gene product, and a growth advantage for that aberrant cell. But as more and more oncogenes were identified, researchers realized that tumor growth was also associated with loss of function of certain "tumor suppressor genes". These tumor suppressor genes were often inactivated by their deletion from the nucleus, and the phrase "loss of heterozygosity" (LOH) was applied to genetic loci in which both *alleles were present in normal tissues, but one copy was lost in tumor tissue. In many instances, tumor suppressor genes were first identified by virtue of germ-line mutations that were present at a high frequency in a rare tumor, e.g., retinoblastoma, but it soon became apparent to researchers that many tumor suppressor genes were associated with a variety of different tumors, many of which were not rare at all.

3) There are no oncogenes or tumor suppressor genes that are activated or deleted in and from all cancers. Even tumors of a single organ rarely have uniform genetic alterations, although tumor types from one specific organ do have a tendency to share mutations in certain genes or in different genes within a single growth-regulatory pathway.

4) At the present time, it is not known how many critical mutations are required to convert a single normal cell into a malignant cell. Human cells have been difficult to transform in vitro, and the basis for this difficulty is not yet understood. The simplest model of tumorigenesis is as follows:

a) Human cells experience a certain number of mutations each day as a result of exposure to carcinogens or as a result of ordinary biological degradation, both of which can alter nucleotide sequences. Errors will also occur during new DNA synthesis and in the process of disentangling the chromosomes during *mitosis. Most of these errors would be either irrelevant to the life of the cell or deleterious because of the loss of a gene critical for cellular viability.

b) By chance, an occasional genomic mutation might create a growth advantage for a cell, permitting increased net cellular growth, because of increased proliferation or a reduction in programmed cell death (reduction in apoptosis), with a resulting *clonal expansion of that lineage. A second genomic alteration might then occur within this expanding clone, again by chance, providing an additional growth advantage for that cell and its progeny. By virtue of these two advantages, the cells of this clone would eventually overgrow neighboring cells, creating yet another expanding clone. This scenario would repeat as a consequence of each new mutation that provided an additional growth advantage. The accumulation of these growth promoting mutations is the basis of the current view of "multistep carcinogenesis".

Proc. Nat. Acad. Sci. 1999 96:14675

Notes:

*epithelial cells: In animals, "epithelial cells" compose the cell layers that form the interface between a tissue and the external environment, for example, the cells of the skin, the lining of the intestinal tract, and the lung airway passages.

*mesodermal: In the embryos of higher animals, there occurs the transformation of a single-layer "blastula" into a 3-layered "gastrula" consisting of ectoderm (outermost layer), mesoderm (middle layer), and endoderm (innermost layer) surrounding a cavity with one opening. The 3 layers are called the "germ layer", and these layers, via further cell differentiation and proliferation, determine the development of all the major body systems and organs.

*lymphatic: The lymphatic system is a complex network for the distribution of lymph fluid (which is similar to blood plasma --blood without red cells). Lymph is collected by drainage from the tissues throughout the body, flows in the lymphatic vessels through the lymph nodes, and is eventually added to the venous blood circulation.

*point mutation: A minor changes in the genome; a single base-pair substitution.

*genetic amplification process: The production, by various means, of additional copies of a stretch of genomic DNA.

*alleles: One of two or more forms of a given gene that control a particular characteristic, with the alternative forms occupying corresponding loci on homologous chromosomes.

*mitosis: Programmed division of the nucleus during cell replication.

*clonal expansion: This refers to the expansion of a population of cells all derived from repeated replications of progeny of a single cell.

Related Background:

ON GENETICS AND HUMAN CANCERS

The current consensus is that cancer results from the accumulation of mutations in the genes that directly control the birth and death of biological cells. But the mechanisms through which these mutations are generated are the subject of continuing debate and much research. It has been argued that an underlying genetic instability is absolutely essential for the generation of the multiple mutations that underlie cancer. On the other hand, it has also been suggested that normal rates of mutation, coupled with waves of *clonal expansion, are sufficient for the cancer process to occur in humans.

C. Lengauer et al (Johns Hopkins University, US) present a review of observations concerning the stability of the genome of human cancer cells, the authors making the following points:

1) Numerous genetic alterations that affect growth-controlling genes have been identified in neoplastic cells over the past 15 years, and these observations provide persuasive evidence for the genetic basis of human cancer. The alterations can be divided into 4 major categories:

a) Subtle sequence changes: These changes involve nucleotide base substitutions or deletions or insertions of a few nucleotides in the genome, and unlike the alterations described below, they cannot be detected via cytogenetic analysis. Such mutations, for example, occur in over 80 percent of pancreatic cancers.

b) Alterations in chromosome number: Such alterations involve losses or gains of whole chromosomes. Such changes are found in nearly all major human tumor types.

c) Chromosome translocations: These alterations can be detected cytogenetically as fusions of different chromosomes or of normally non-contiguous segments of a single chromosome. At the molecular level, such translocations produce fusions between two different genes, endowing the fused genetic entity with tumorigenic properties. Such translocations are known to occur in the *chronic myelogenous leukemias.

d) Gene amplifications: These are seen at the cytogenetic level as homogeneously stained regions, and at the molecular level they involve multiple copies of a gene. An example of gene amplification occurs in advanced *neuroblastomas.

2) All 4 of the alterations described above occur commonly in specific tumor types but are rarely or never observed in normal cells. However, the existence of genetic alterations in a tumor, even when frequent, does not mean that the tumor is genetically unstable. By definition, instability is a matter of rate, and the existence of a mutation provides no information about the rate of its occurrence. The higher prevalence of mutations in tumor cells compared with normal cells still requires explanation.

The authors conclude: "One can argue persuasively that all chemotherapeutic compounds used at present are more toxic to cancer cells than to normal cells only and specifically because of the defective *checkpoints that occur in cancer cells. This line of reasoning suggests that, although instability may be essential for neoplasia to develop, it may also prove to be its Achilles' heel when the tumor is attacked by the right agents. Further research to define the molecular and physiological bases of instability may, therefore, yield entirely new approaches to treating common forms of cancer."

Nature 1998 396:643

Notes:

*clonal expansion: In this context, this refers to the expansion of a population of cells all deriving from a single mutated cell.

*chronic myelogenous leukemias: (granulocytic leukemias) These leukemias are characterized by an uncontrolled proliferation of myelopoietic cells (blood cells derived from bone marrow).

*neuroblastomas: Neuroblastomas are malignant neoplasms characterized by only slightly differentiated immature nerve cells of embryonic type.

*checkpoints: In this context, the term "checkpoint" refers to a point in the eukaryotic cell division cycle where the cycle can be halted until conditions are suitable for the cell to proceed to the next stage. (eukaryotic = containing membrane-bound organelles such as a nucleus.)

Related Background:

ANEUPLOIDY AND GENETIC INSTABILITY OF CANCER CELLS

In general, germ cells (egg cells and sperm cells) and somatic cells (non-germ cells) carry different numbers of chromosomes, with germ cells carrying exactly half the number (haploid number) of somatic cell chromosomes (diploid number). The term "aneuploidy" (heteroploidy) refers to a condition in which the number of chromosomes in a cell is not an integer multiple of the haploid number typical for that cell or organism. For example, the haploid human chromosome number is 23; the normal somatic cell contains 46 chromosomes; a somatic cell with 47 or 44 chromosomes is aneuploid. Some authors, however, use the term "aneuploidy" to indicate merely an abnormal number of chromosomes. In cell biology, the term "karyotype" refers to the characteristics profile (number, size, and shape) of a set of chromosomes of a cell or organism. In this context, the term "phenotype" refers to the total appearance of a cell as determined by the interaction during development between its genetic constitution (genotype) and the cell's environment. Genetic and phenotypic instability are hallmarks of cancer cells, but the cause of the instability is not clear. The leading hypothesis suggests that a poorly defined gene mutation generates genetic instability and that one or more of the many subsequent mutations then cause cancer [*Note #1].

P. Duesberg et al (2 installations, DE US) report an investigation of the hypothesis that genetic instability of cancer cells is caused by aneuploidy, which they define as "an abnormal balance of chromosomes". The authors point out that because symmetrical segregation of chromosomes during mitosis depends on exactly two copies of the genes involved in mitosis ("mitosis genes"), aneuploidy involving chromosomes bearing mitosis genes will destabilize the karyotype. The authors propose that the aneuploidy hypothesis predicts that the degree of genetic instability should be proportional to the degree of aneuploidy, and it should thus be difficult to maintain the particular karyotype of a highly aneuploid cancer cell on *clonal propagation. The authors report this prediction is confirmed with clonal cultures of chemically transformed aneuploid Chinese hamster embryo cells. Defining the "ploidy factor" as the quotient of the modal chromosome number divided by the normal number of the species, it was found that the higher the ploidy factor of a clone, the more unstable was its karyotype. The authors point out that work by others has established an exact correspondence between the karyotype instability of human colon cancer cell lines and the degree of aneuploidy. The present authors suggest that, independent of gene mutation, aneuploidy is sufficient to explain genetic instability and the resulting karyotypic and phenotypic heterogeneity of cancer cells. The authors further suggest that because aneuploidy has also been proposed to cause cancer, their hypothesis "offers a common, unique mechanism of altering and simultaneously destabilizing normal cellular phenotypes."

Proc. Nat. Acad. Sci. 1998 95:13692

Notes:

*Note #1: In 1976, Peter Nowell postulated that a precancerous mutation generates exceptional "genetic instability" or "mutability", and that the highly mutable "premalignant" cell then suffers many further gene mutations, including those that cause cancer (P.C. Nowell, Science 194:21 1976).

*clonal propagation: In general, in this context, a "clone" is a line of identical cells produced from one or a few originating cells.

Related Background:

MEDICAL BIOLOGY: TELOMERES, TELOMERASE, AND CANCER

Telomeres are defined ends of chromosomes that contain specific repeated DNA sequences. They are essential for normal chromosome replication, and since their length shortens a bit with each replication, they are believed to be involved in the aging of the cell. Telomerase is an enzyme that repairs damage to telomeres, and it is thought by some researchers that cancerous cells may have mutant telomerase, the mutant enzyme conferring immortality on the cancer cell.

Charles H. Buys (University of Groningen, NL) presents a short review of current research on telomere targeting in the treatment of cancer, the author making the following points:

1) Before any biological cell can divide, it must first replicate the double-stranded DNA in its chromosomes. Each cell, however, has a problem replicating the DNA at the telomeres, where there are over 1000 short base sequences, thymine-thymine-adenine-guanine-guanine (TTAGGG), repeated again and again, and a variety of attached DNA-binding proteins. In a normal cell, the replication machinery is unable to copy the last few bases of the telomeres on one of the strands of DNA in the chromosome. As a consequence, the telomeres shorten with each round of DNA replication.

2) Telomeres are essentially molecular caps, protecting the ends of chromosomes against degradation and preventing ligation of the ends of DNA by DNA repair enzymes. These functions are crucial to the cell, but the wearing away of telomeres with each cell division, when repeated during many cell cycles, eventually eliminates the protective functions, and chromosomes become unstable, fused, or lost. Cells with such chromosome defects are not able to divide and may not survive. Attrition of telomeres thereby limits the lifespan of many types of biological cells.

3) Two distinctive types of cells -- *germ cells and early embryonic cells -- solve the problem of truncated telomeres by means of a complex of proteins and RNA called "telomerase". The RNA component of this complex contains a template sequence on which the TTAGGG repeated groups ("repeats") at the ends of DNA can by synthesized.

4) Unlike germ cells and early embryonic cells, most other cells (somatic cells) switch off the activity of telomerase after birth. In contrast, many types of cancer cells, perhaps as much as 90 percent of various types of cancer cells, reactivate telomerase. This essentially "rewinds the clock" on run-down telomeres and contributes to the growth of the malignant clone population of cancer cells.

5) With these considerations in mind, several recent studies have explored the possibility of inhibiting telomerase as a way of arresting the growth of tumor cells. Although the results have been interesting and have suggested new possibilities for the treatment of cancer, there are several cautionary points that must be noted:

a) Experiments have been conducted in vitro in cultured cell lines, with efficient uptake of inhibitors ensured by means unavailable in in vivo treatment.

b) The inhibitors apparently work best in cultured tumor cells when telomeres are shortest, but little is known about the length of telomeres in *primary human tumors, and there is some evidence that certain types of tumor cells may actually have shorter telomeres than benign cells of the same type of tissue.

c) Up to 20 percent of human tumors do not have telomerase activity and may use other mechanisms to preserve their telomeres.

6) Concerning the above cautionary points, the author concludes: "Even though the therapeutic potential of telomerase inhibitors may be limited by these considerations, further investigation of this approach is certainly worthwhile."

New Engl. J. Med. 2000 342:1283

Notes:

*germ cells: A germ cell is any cell from which gametes (sperm cells and egg cells) are derived. All other cells are called "somatic" cells.

*primary human tumors: The term "primary tumor" refers to the original tissue malignancy. Malignant cells from a primary lung cancer, for example, may relocate ("metastasize") to the brain, where they replicate as malignant lung-tissue cells, causing a "secondary tumor".

Related Background:

MEDICAL BIOLOGY: NEW DATA ON RETROVIRUSES AND LUNG CANCER

Whether viruses are classified as "living" or "non-living" is arbitrary, but in either case one is dealing with a remarkable entity. As a group, viruses are in the size range 20 to 300 nanometers, compact bundles of genetic information that have apparently existed for billions of years. They are unable to replicate except inside a living host cell, and outside the host cell they are inert. Viruses pass through filters that trap bacteria, and they can be seen only be electron microscopy. Like other biological systems, each virus contains molecular genetic information in the form of nucleic acid, but in viruses this information comes in the form of DNA or RNA, never both. The viral genome is transcribed and replicated only with a host cell, with variations in process dependent on the type of virus and the type of host cell.

When viruses are categorized in terms of their genomes, there are two general types: a) viruses with a DNA genome (DNA viruses), and b) viruses with an RNA genome (RNA viruses). RNA viruses are unique: only in these viruses do we find genomes consisting of RNA; all other biological entities, DNA viruses, bacteria, plant cells, animal cells, etc., contain DNA genomes.

There are more than 2500 groups of different viruses now recognized and at least partially characterized. In each case, for both DNA and RNA viruses, once it enters the host cell, the general challenge for the virus is the same: directly or indirectly, the viral genome must bring about the production of the *messenger RNAs needed by the host *ribosomes to produce the specific proteins necessary for replication of the complete virus.

With DNA viruses, the DNA viral genome acts as the template for the production of messenger RNA. With RNA viruses, however, the process is more complicated.

In general, with some types of RNA viruses, the RNA genome ("plus-sense"; "positive-strand") can itself act as messenger RNA for host ribosomes; while other types of RNA viruses, the RNA genome ("minus-sense"; negative-strand) must first produce a complementary RNA, which then acts as messenger RNA for the host ribosomes. The replication process in minus-sense RNA viruses is complex, since host cells do not carry enzymes that can polymerize complementary RNA from an RNA template, and such viruses therefore must carry their own special enzymes ("RNA-dependent transcriptases") to achieve this synthesis.

A third and special type of RNA virus is the so-called "retrovirus", of which there are many versions. Retroviruses are single-stranded RNA viruses that have an enzyme called reverse transcriptase, and with this enzyme the viral RNA is used as a template to produce viral DNA from host-cellular material. This DNA is then incorporated into the host cell's genome, where it codes for the production of messenger RNA and the ultimate synthesis of viral components. The HIV virus, for example, is a retrovirus.

As a class, retroviruses are usually spherical, 80 to 110 nanometers in diameter, with an RNA genome of approximately 7000 to 10,000 nucleotide bases. An outstanding characteristic of such viruses is that if they kill host cells at all it is usually only after a long latent period (although there are certain important exception). In addition, these viruses are apparently capable of altering, or affecting the expression of, host cell genes involved in cancer (oncogenes). The three primary genes of the retrovirus genome are called "gag", "pol", and "env". The gag gene encodes the protein of the virus capsid, the protein coat directly encapsulating the viral genome; the pol gene encodes a reverse transcriptase involved in replication of the genome; the env gene encodes the protein of the membrane envelope of the virus when it is outside a host cell (the membrane envelope of the "virion"). (These 3 genes actually produce more than 3 different proteins; what these genes encode for are precursor proteins, each of which is a precursor for several varieties of proteins with different viral functions. In addition, different proteins can be produced by splices of elements from the 3 primary genes.

Concerning the general structure of the retrovirus, the internal nucleic acid genome is encapsulated by a protein coat (the capsid), and the capsid in turn is surrounded by the external lipoprotein envelope. Beyond this general scheme, there is no single morphology for retroviruses.

The protein encoded by the env gene (called the Env protein) is important in recognition of host-cell surface receptors, with which it interacts to secure entry of the virus into the host cell. In addition, after infection the host-cell expresses Env protein on its own surface, and this evidently prevents reinfection of that host cell after new viruses are released.

Perhaps the most important property of retroviruses is their ability to splice into the host-cell genome pieces of their own genome, the result either the introduction of new genes into the host genome or the activation or inactivation of specific nearby genes of the host genome. In principle, either of these scenarios can lead to corruption of the growth regulation process of the host genome, and thus to a line of malignant cells.

Viruses have been implicated in the etiology of several types of human cancers, including cervical cancer and liver cancer. The viruses that have been strongly associated with human cancers include human papillomaviruses, Epstein-Barr virus, hepatitis B virus, and a human retrovirus. Many viruses can cause tumors in animals, either as a result of natural infection or after experimental inoculation. Animal viruses are studied to learn how a limited amount of genetic information (only one or a few viral genes) can profoundly alter the growth behavior of host cells, and ultimately convert a normal cell into a malignant cell. For example, classical studies of RNA tumor viruses first revealed the involvement of cellular *oncogenes in cancer, and similar studies with DNA tumor viruses first implicated a role for *tumor suppressor genes in cancer. In the 1980s, these discoveries revolutionized thinking about the molecular mechanisms of carcinogenesis.

Alveolar cell cancer (bronchiolo-alveolar cancer, broncho-alveolar cancer) is a human lung cancer, a subtype of *adenocarcinoma, and apparently a lung cancer unrelated to cigarette smoking. The cause of this disease is unknown.

"Jaagsiekte" is a contagious lung cancer (ovine pulmonary carcinoma) of sheep, sometimes also of goats and guinea pigs, the disease resembling the more benign forms of human alveolar cell cancer. Jagsiekte in sheep is apparently caused by a retrovirus (jagsiekte sheep retrovirus), and infected sheep secrete large amounts of lung fluids from which copious quantities of the virus may be obtained.

The term "fibroblasts" refers to a type of connective tissue cell that secretes the structural proteins (e.g., collagen) that form certain tissue components. Fibroblasts are easy to maintain in tissue culture, and they are often used as experimental cell systems.

In this context, the term "transformation" refers to the transformation of a normal cell into a malignant (cancer) cell.

N. Maeda et al (4 authors at University of California Irvine, US) present a report on cancerous transformation of mouse fibroblasts by jaagsiekte sheep retrovirus DNA, the authors making the following points:

1) The authors point out that animal retrovirus-induced cancers have played fundamental roles in understanding the molecular basis of cancer. Jaagsiekte sheep retrovirus is the causative agent of ovine pulmonary carcinoma, a contagious lung cancer of sheep (also known as sheep pulmonary adenomatosis). This retrovirus-induced cancer consists of transformed secretory *epithelial cells of the lungs, and a characteristic feature of the tumors is the production of large amounts of fluid secreted from tumor cells containing infectious virus. The disease closely resembles human alveolar carcinoma, and is thus an important model for understanding the latter.

2) The authors point out that oncogenic retroviruses induce tumors by two mechanisms. a) Acutely transforming retroviruses capture a normal host-cell gene (a "protooncogene") and convert it into a viral oncogene. Such retroviruses typically induce rapid neoplasms in vivo, and they frequently can transform cells in culture. b) Retroviruses that lack oncogenes (non-acute retroviruses) also can induce tumors, although they typically require longer incubation periods and multiple rounds of infection in vivo. An important molecular mechanism for these viruses is insertion in the host-cell genome of viral genetic material in the vicinity of a protooncogene. This results in overexpression of the protooncogene.

3) The authors report their experiments indicate that jaagsiekte sheep retrovirus DNA (i.e., DNA produced by transcription of the viral genome) can induce transformation in a line of mouse fibroblasts (NIH 3T3 cells), and that additional experiments have localized the transforming activity to the viral env gene. The authors suggest their results indicate that the envelope gene carries the transforming potential in this virus, an unusual case of a transformation potential carried by a viral structural protein.

In a contiguous independent report, S.K. Rai et al (6 authors at 3 installations, US) report that the envelope (env) gene of jaagsiekte sheep retrovirus "has the unusual property that it can induce transformation in rat fibroblasts, and thus is likely to be responsible for oncogenesis in animals."

In a commentary on the above work, Naomi Rosenberg (Tufts University, US) points out that more than 50 oncogenes now known to be involved in human cancers were first discovered and studied in retroviral models. "Nonetheless, only one retrovirus, human T cell lymphotrophic virus (HTLV), is oncogenic in humans, and, as we enter the 21st century, retroviral oncogenesis models may seem to be of largely historical interest. However, analyses of jaagsiekte sheep retrovirus by Maeda et al and Rai et al... reveal that oncogenic retroviruses still hold important secrets that may be directly relevant to human cancer."

Proc. Nat. Acad. Sci. 2001 98:4285,4443,4449

Notes:

*messenger RNAs: (mRNAs) The ribonucleic acid molecules transcribed from DNA that carry the coded information specifying the sequence of amino acids in proteins.

*ribosomes: A ribosome (not to be confused with riboZYME) is a small particle, a complex of various ribonucleic acid component subunits and proteins that functions as the site of protein synthesis. In general, ribosomes read the messenger RNA template to produce specific polypeptide sequences by polymerizing amino acids.

*oncogenes: There are two general meanings of this term in current use. The first meaning refers to any of a family of cellular genes that normally code for proteins involved in cell growth or regulation, but which may produce malignant processes when mutated or activated by viruses. The second meaning of the term "oncogene" refers to viral genes found in certain DNA tumor viruses, genes that are required for viral replication, but whose activation produces malignant transformations.

*tumor suppressor genes: Tumor suppressor genes code for proteins that apparently either prevent cell division or provoke cell death in defective cells. Thus, deletion or inactivation of tumor suppressor genes can result in malignant cell replication.

*adenocarcinoma: In general, a tumor of epithelial cells (see below) in which the cells are in a glandular or gland-like pattern.

*epithelial cells: In animals, "epithelial cells" compose the cell layers that form the interface between a tissue and the external environment, for example, the cells of the skin, the lining of the intestinal tract, and the lung airway passages.

Related Background:

TUMOR VIRUSES AND ONCOGENES

Viruses, first discovered in 1892 [*Note #1], are infectious agents that are smaller than biological cells such as bacteria, their size ranging from about 20 nanometers to about 900 nanometers. Beginning in the 1930s, there was much controversy about classification, with the central question whether viruses are a "life" form. The answer, of course, depends on the definition of "life", and since there are various definitions possible, the question is ill-defined. In general, a virus is a self-organizing molecular system capable of using living cells to replicate itself, the sequence of events in many cases altering or destroying the cells and thus causing a disease process.

In addition to disease processes caused by the destruction of cells, viruses are now known to be etiologic factors in the development of several types of human tumors, including two of worldwide significance: cervical cancer and liver cancer. Viruses have also been strongly associated epidemiologically with other human cancers, and these viruses include *human papillomavirus, *Epstein-Barr virus, and *hepatitis B virus.

Like other viruses, tumor viruses are classified among different virus families according to the nucleic acid of their genome and the biophysical characteristics of their virions (virion = the complete virus particle as it exists outside the host cell). All known tumor viruses either have a DNA genome or an RNA genome. In the case of an RNA genome, the RNA genome generates a DNA "provirus" (a preliminary virus entity) after infection of cells. All RNA tumor viruses (i.e., tumor viruses with an RNA genome) belong to the "retrovirus" family. Retroviruses carry an RNA-directed polymerizing enzyme (reverse transcriptase) that constructs a DNA copy of the RNA genome of the virus. This DNA copy (the provirus) becomes integrated into the DNA of the infected host cell, and it is from this integrated DNA that all proteins of the virus are translated.

Tumor viruses are of two general types with respect to tumor induction, distinguished by (among other things) whether or not they carry "oncogenes" (i.e., any gene associated with the causation of cancer). All DNA tumor viruses carry oncogenes, these oncogenes an integral part of the viral genome and not derived from host cells. Of RNA tumor viruses, there are two types: a) the highly oncogenic ("direct-transforming") RNA tumor viruses (class I RNA tumor viruses) carry an oncogene of host-cell origin; b) the weakly oncogenic ("slowly transforming") RNA tumor viruses (class II RNA tumor viruses) do not contain an oncogene but induce leukemias after long incubation periods by indirect mechanisms. In this context, the term "transforming" refers to the transformation of a normal host-cell into a cancer cell.

George Klein (Karolinska Institute, SE) presents an essay on the history of the idea of tumor viruses, the author making the following points:

1) In 1911, Peyton Rous (1879-1970) demonstrated that fowl *sarcomas could be transmitted with cell-free filtrates, and the rapid consensus was that cancer was a viral disease. But when similar experiments with mouse and rat tumors failed soon after, it was concluded that tumor viruses occurred only in birds, and the field fell into disrepute.

2) The discovery in the 1920s of the Shope papilloma virus, which causes warts in rabbits, produced little enthusiasm among researchers because the tumors were largely benign.

3) In the 1930s, the mouse mammary tumor virus was discovered, and this virus was called "*milk factor" rather than "milk virus" to avoid a negative reaction from people in the medical science community who had relegated tumor viruses to the cabinet of freaks.

4) The great change in the climate of opinion concerning tumor viruses came in the 1950s when Ludvik Gross discovered the mouse leukemia virus, and Sarah Stewart and Bernice Eddy identified the *polyomavirus. Within a few years the pendulum had swung to the opposite extreme. After decades of failed attempts, viruses that could induce tumors in mammals were now isolated in quick succession. Tumor virology rapidly became favored by grant-giving agencies, and the oncogene concept -- that tumor viruses carry "cancer genes" that can transform some of their target cells into a cancerous or precancerous state -- was formulated in the context of this enthusiasm.

5) The class I RNA tumor viruses can induce tumors because they have accidentally incorporated from host cells genes that regulate growth. After entering a new host cell, the viral enzyme reverse transcriptase copies the viral RNA into provirus DNA which integrates randomly into the host cell DNA. When the virus starts to reproduce itself, and the proviral DNA is transcribed back into RNA, some of the new virus particles may also carry additional cellular sequences from regions adjacent to the random integration site of the proviral DNA. These newly replicated virus particles have the potential to corrupt the DNA of other host cells when these host cells are subsequently infected.

6) Class II RNA tumor viruses do not themselves contain oncogenes, but contribute to malignant tumor development relatively infrequently when their proviral DNA happens to integrate into the host DNA near host oncogenes.

7) By discovering cellular genes that regulate growth and that can contribute to cancer development after illegitimate viral activation, the virologists demonstrated the existence of host oncogenes that when mutated can promote cancer formation independent of viruses. This discovery once again relegated tumor virology to a less prominent place and reaffirmed the sovereignty of cell biology. Cancer is essentially a disease of cellular DNA; when viruses are involved in cancer, they are involved as one of many possible DNA-corrupting agents.

8) All oncogenes have turned out to be *highly conserved housekeeping genes that participate in the regulation of the *cell cycle. Their potentially tumorigenic forms drive the cell towards proliferation. For overt tumor development, additional genetic changes are required, including loss of *cell-cycle check-point controls, inhibition of programmed cell death (apoptosis), and *up-regulation of blood supply (*angiogenesis). The current oncogene field, therefore, emerged from erroneous concepts concerning the etiology of cancer, but these concepts, when combined with several decades of diligent experimentation, finally produced a valuable outline of the events that cause the formation of cancerous tumors.

Nature 1999 400:515

Notes:

*Note #1: In 1892, a Russian botanist named Dmitri Ivanovsky (1864-1920) became interested in the cause of a disease of the valuable tobacco plant, a disease called "mosaic disease". The name was due to the mosaic patterns the disease produced on the leaves of the plant. Ivanovsky devised an experiment, mashed up infected leaves, and forced them through filters designed to remove all bacteria. He discovered that the liquid that passed through the filters could still infect healthy tobacco plants. Ivanovsky concluded, in error, that the infectious agent was still a bacterium, but that his filters were somehow defective, and it was this conclusion that he published in a Russian scientific journal. Six years later, a Dutch botanist named Martinus Beijerinck (1851-1931) repeated the experiments and concluded the infectious agent was not a bacterium but the liquid itself, a poison, and he called it a filterable "virus", the word "virus" being the Latin word for poison. In the 1930s, an American biochemist named Wendell Meredith Stanley (1904-1971) became interested in this mysterious "virus" liquid. He believed the poison to be a protein. He repeated the filtration experiments with diseased tobacco leaves, and he isolated a crystalline substance in high concentration that had all the infective properties of the so-called virus liquid. In the report which Stanley published in 1935, he concluded: "Tobacco mosaic virus is regarded as an autocatalytic protein which, for the present, may be assumed to require the presence of living cells for multiplication." For this work, Stanley received the Nobel Prize in Chemistry in 1946. Although the tobacco mosaic virus, like all viruses, is much more than just a simple autocatalytic protein, the first understanding of the true nature of viruses was now in place. The experiments that had begun in Russia in 1892 culminated 43 years later in the startling realization that an entire class of infectious agents much smaller than bacteria existed.

*human papillomavirus: The papillomavirus causes benign tumors called "warts". The virus in its extracellular form (virion) is 55 nanometers in diameter, with a circular double-stranded DNA genome of approximately 8000 nucleotide base pairs. The virus primarily infects surface *epithelia. There are more than 70 different types of human papillomaviruses.

*epithelia: In animals, epithelial cells compose the cell layers that form the interface between a tissue and the external environment, for example, the cells of the skin, the lining of the intestinal tract, and the lung airway passages.

*Epstein-Barr virus: The Epstein-Barr virus is a large ubiquitous herpesvirus that is the causative agent of acute infectious mononucleosis and a factor in the development of nasopharyngeal carcinoma, Burkitt's lymphoma, and other disorders. The virion is approximately 100 nanometers in diameters, and contains a double-stranded DNA genome consisting of approximately 172,000 nucleotide base pairs.

*carcinoma: In general, a carcinoma is any malignancy derived from epithelial tissue.

*hepatitis B virus: The hepatitis B virus is the causative agent of *serum hepatitis. The virion is 42 nanometers in diameter, the genome double-stranded DNA with 3200 nucleotide base pairs.

*serum hepatitis: In general, "hepatitis" is any inflammation of the liver. "Serum hepatitis" is usually transmitted by injection of infected blood or blood derivatives, or by the use of contaminated needles or other instruments.

*sarcomas: A sarcoma is a connective tissue neoplasm, usually highly malignant.

*milk factor: The prototype of RNA tumor viruses is the "mouse mammary tumor virus", which occurs in high-mammary-cancer strains of inbred mice, and is found in particularly large amounts in lactating mammary tissues and milk. The virus is readily transferred to suckling mice, in whom the incidence of subsequent development of carcinoma of the breast is high. The work of J.J. Bittner (1904-1961) on this virus is classic: In the 1930s, carefully inbred strains of mice were kept for research on cancer. Some strains were highly resistant to cancer and rarely developed it, while other strains were so prone to cancer that almost every individual animal developed the disease. In 1936, Bittner established that if young mice of a cancer-resistant strain were transferred to the breast of a foster mother of a cancer-prone strain, the young mice developed cancer in the course of their lives. If, on the other hand, young mice of a cancer-prone strain were fed at the breast of a foster mother of a cancer-resistant strain, they did not usually develop cancer. The Bittner "milk factor" was isolated in 1949 and was found to consist of virus-like particles containing nucleic acid.

*polyomavirus: Papillomaviruses and polyomaviruses are the two classes of papovaviruses. Like papillomaviruses, polyomaviruses have a double-stranded DNA genome, but with only 5000 nucleotide base pairs. The polyoma virion is 45 nanometers in diameter, and the target tissues are internal organs. An important research tool, the SV40 monkey virus, is a polyomavirus.

*highly conserved housekeeping genes: The term "highly conserved" refers to a gene sequence maintained across evolutionary time. So-called "housekeeping" genes are genes coding for proteins involved in essential functions such as metabolic cycles.

*cell cycle: The "cell cycle" is the name given to the ordered sequence of phases through which a cell passes from one mitotic cell division to the next.

*cell-cycle check-point controls: So-called "checkpoints" are points in the cell division cycle where the cycle can be halted until conditions are suitable for the cell to proceed to the next stage.

*up-regulation: In general, the term "up-regulation" refers to an increase in the activity of some entity or process.

*angiogenesis: Angiogenesis, the origin and development of blood vessels, is an important consideration in the growth of cancerous tumors, since the tumor provokes directed angiogenesis into itself (up-regulation of a blood supply) with the end result that the tumor is supplied with oxygen and nutrients. Without angiogenesis, tumors can attain only a small size before becoming self-inhibiting.

Related Background:

xCELLULAR SENESCENCE, CANCER, AND AGING

A. Krtolica et al (Lawrence Berkeley National Laboratory, US) discuss cellular senescence, the authors making the following points:

1) Multicellular organisms have evolved mechanisms to prevent the unregulated growth and malignant transformation of proliferating cells. One such mechanism is "cellular senescence", which arrests proliferation (essentially irreversibly) in response to potentially oncogenic events. Cellular senescence appears to be a major barrier that cells must overcome to progress to full-blown malignancy.

2) Cellular senescence was first described as a process that limits the proliferation of cultured human fibroblasts ("replicative senescence"). Proliferating cells progressively lose telomere DNA, and short telomeres, which are potentially oncogenic, elicit a senescence response. In addition, DNA damage, expression of oncogenes, and supraphysiological mitogenic signals also cause cellular senescence. Cellular senescence is controlled by tumor suppressor genes and seems to involve a checkpoint that prevents the growth of cells at risk for neoplastic transformation. In this regard, cellular senescence is similar to apoptosis. However, whereas apoptosis kills and eliminates damaged or potential cancer cells, cellular senescence involves a stable arrest of growth.

3) Cellular senescence is also thought to contribute to aging, although how it does so is poorly understood. In addition to arresting growth, senescent cells show changes in function. Because senescent cells accumulate with age, they may contribute to age-related declines in tissue function. If so, cellular senescence may be an example of "antagonistic pleiotropy". Aging phenotypes are thought to result from the declining force of natural selection with age. Consequently, traits selected to maintain early life fitness can have unselected deleterious effects late in life, a phenomenon termed "antagonistic pleiotropy". The senescence-induced growth arrest may suppress the development of cancer in young organisms. The functional changes, by contrast, may be unselected consequences of the growth arrest and thus compromise tissue function as senescent cells accumulate.

Proc. Nat. Acad. Sci. 2001 98:12072

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