Retinoblastoma

Retinoblastoma is the prototype of diseases caused by a pathogenic variant in a TSG. It is a rare malignant tumor of the retina in infants, with an incidence of ~1 in 20,000 births. It is the classic example put forth by Knudson, illustrating the role of a germline event leading to earlier age of disease and wider extent (unilateral vs bilateral). Treatment of a retinoblastoma may require removal of the affected eye; however, the advent of intraarterial chemotherapy has allowed many tumors to be effectively treated by local therapy so that vision can be preserved.

Approximately 40% of cases of retinoblastoma are of the heritable form, in which the child has one germline pathogenic variant in RB1, either inherited from a heterozygous parent or which occurred de novo, or from a parent with germline mosaicism for the RB1 pathogenic variant. In these children, retinal cells, which like all other cells of the body, are already carrying one defective RB1 allele, suffer a somatic mutation in the other allele, leading to loss of function from both copies of RB1 and initiating tumor development (Fig. 1).

Fig1. Chromosomal mechanisms that could lead to loss of heterozygosity (LOH) for DNA markers at or near a tumor suppressor gene in an individual heterozygous for a germline pathogenic variant. The figure depicts the events that constitute the second hit that leads to retinoblastoma with LOH. Local events such as mutation, gene conversion, or transcriptional silencing by promoter methylation, however, could also cause loss of function of both RB1 genes without producing LOH. +, Normal allele; rb, mutant allele.

The disorder appears to be inherited as a dominant trait because the large number of primordial retinoblasts and their rapid rate of proliferation make it very likely that a somatic mutation will occur as a second hit in one or more of the retinoblasts already carrying a heterozygous RB1 pathogenic variant. Because the chance of a second hit is so great, heterozygotes for the disorder often have tumors arising at multiple sites, which may be multifocal tumors in one eye or both eyes (bilateral retinoblastoma), as well as less commonly in the pineal gland (referred to as trilateral retinoblastoma). The occurrence of a second hit is, however, a matter of chance and does not occur 100% of the time; thus the penetrance of heritable retinoblastoma is high (>90%) but not complete.

The other 60% of cases of retinoblastoma are sporadic; in these cases, both RB1 alleles in a single retinal cell have been mutated or inactivated independently by chance, and the child does not carry a pathogenic RB1 variant in the germline. Because two hits in the same cell is a statistically rare event, there is usually only a single clonal tumor: the retinoblastoma is found at one location (unifocal) in one eye only. However, a unilateral tumor does not guarantee that the child does not have the heritable form of retinoblastoma because 15% of patients with unilateral retinoblastoma have a germline pathogenic RB1 variant. Another difference between hereditary and sporadic tumors is that the average age at onset of the sporadic form is in early childhood— later than in infants with the heritable form— reflecting the longer time typically needed for two somatic mutations, rather than one, to occur.

In a few patients with retinoblastoma, the variant responsible is a cytogenetically detectable deletion or translocation of the portion of chromosome 13 containing the RB1 gene. Such chromosomal changes, if they also disrupt genes adjacent to RB1, may cause a contiguous gene deletion syndrome involving varying degrees of developmental delay, congenital anomalies, and dysmorphic features.

Nature of the Second Hit. Typically, for retinoblastoma as well as for the other hereditary cancer syndromes, the first hit is an inherited pathogenic variant; that is, a change in the DNA sequence. The second hit, however, can be caused by a variety of genetic, epigenetic, or genomic mechanisms (see Fig. 1). Although it is most often a somatic mutation, loss of function without mutation, such as occurs with epigenetic silencing, has been observed. Although a number of mechanisms have been documented, the common theme is loss of function of RB1. The RB1 gene product, p110 Rb1, is a phosphoprotein that normally regulates entry of the cell into the S phase of the cell cycle. Thus loss of the RB1 gene and/ or absence of the normal RB1 gene product (by any mechanism) deprives cells of an important checkpoint and allows uncontrolled proliferation.

Loss of Heterozygosity. In addition to mutation and epigenetic silencing, a novel and important genomic mechanism was uncovered by geneticists who compared DNA polymorphisms at the RB1 locus in DNA from normal cells to those in the retinoblastoma tumor from the same patient. Individuals with retinoblastoma who were informative by being heterozygous at polymorphic loci flanking the RB1 locus in normal tissues (see Fig. 1) frequently had tumors with alleles from only one of their two chromosome 13 homologues. This reflected a loss of heterozygosity (LOH) in tumor DNA in and around the RB1 locus. Furthermore, in familial cases, the retained chromosome 13 markers were the ones inherited from the affected parent. Thus, in these cases, LOH represents the second hit. LOH may occur by interstitial deletion, or by mechanisms such as mitotic recombination or monosomy 13 due to nondisjunction (see Fig. 1).

LOH is the most common mutational mechanism by which the function of the remaining normal RB1 allele is disrupted in heterozygotes, although each of the mechanisms shown in Fig. 1 has been documented. LOH is a feature of a number of cancers, both heritable and sporadic, and is often considered evidence for a TSG in the region of LOH.

Hereditary Breast Cancer due to Pathogenic Variants in BRCA1 or BRCA2

 Breast cancer is common, with 250,000 women diagnosed annually in the United States alone. It is estimated that ~5% of breast cancer cases are due to a highly penetrant dominantly inherited mendelian predisposition that increases the risk for female breast cancer four- to sevenfold over the 12% lifetime risk observed in the general female population. In these families, one often sees features characteristic of hereditary (as opposed to sporadic) cancer: multiple affected individuals, earlier age at onset, frequent multifocal or bilateral disease or a second independent primary breast tumor, and additional primary cancers in other tissues such as ovary and pancreas.

Although a number of genes in which pathogenic variants cause highly penetrant mendelian forms of breast cancer have been discovered from family studies, the two genes responsible for the majority of all hereditary breast cancers are BRCA1 and BRCA2. Together, these two TSGs account for approximately one- half and one- third, respectively, of autosomal dominant familial breast cancer. Thousands of pathogenic variants in both genes have now been catalogued. Pathogenic variants in BRCA1 and BRCA2 are also associated with a significant increase in the risk for ovarian and fallopian duct cancer. Moreover, pathogenic variants in BRCA2 and, to a lesser extent, BRCA1 also account for 10% to 20% of all male breast cancer and increase the risk for male breast cancer 10- to 60- fold over the 0.1% lifetime risk in the general population (Table 1). BRCA2 is also the most commonly mutated gene observed in men with metastatic prostate cancer.

Table1. Lifetime Cancer Risks in Carriers of BRCA1 or BRCA2 Pathogenic Variants Compared to the General Population

The gene products of BRCA1 and BRCA2 are nuclear proteins contained within the same multiprotein complex. This complex has been implicated in the cel lular response to double- stranded DNA breaks, such as those occur ring during homologous recombination or because of damage to DNA. As might be expected for any TSG, tumor tissue from heterozygotes for BRCA1 and BRCA2 pathogenic variants frequently demonstrates LOH with loss of the normal allele. Moreover, germline pathogenic variants in BRCA1/ 2 also lead to tumor- specific phenotypes in breast and ovarian cancer as reflected by signature 3 or BRCAness characterized by high mutation burden of multiple types.

Penetrance of BRCA1 and BRCA2 Pathogenic Variants. Presymptomatic detection of women at risk for breast cancer as a result of any of these susceptibility genes relies on detecting clearly pathogenic variants. For the purposes of patient management and counseling, it would be helpful to know the lifetime risk for development of breast cancer in individuals, whether male or female, carrying particular variants in BRCA1 and BRCA2, compared with the risk in the general population (see Table 1). Initial studies showed a greater than 80% risk for breast cancer by the age of 70 years in women heterozygous for BRCA1 pathogenic variants, with a somewhat lower estimate for BRCA2 variant carriers. These calculations relied on estimates of cancer risk in female relatives within families ascertained because breast cancer had already occurred many times in the family (i.e., families in which the particular BRCA1 or BRCA2 pathogenic variant was highly penetrant).

When similar risk estimates were made from population- based studies, however, in which women carrying BRCA1 and BRCA2 pathogenic variants were not selected because they were members of families in which many cases of breast cancer had already developed, the risk estimates were lower and ranged from 40% to 50% by the age of 70 years. The discrepancy between the penetrance of pathogenic variants in families with multiple occurrences of breast cancer and the penetrance in women identified by population screening and not by family history suggests that other genetic or environmental factors must play a role in the ultimate penetrance of BRCA1 and BRCA2 pathogenic variants.

In addition to pathogenic variants in BRCA1 and BRCA2, pathogenic variants in other genes can also cause autosomal dominantly inherited breast cancer syndromes, albeit less commonly. These include the LFS, hereditary diffuse gastric and lobular breast cancer, Peutz- Jeghers syndrome, and Cowden syndrome. These conditions have lifetime breast cancer risks that approach those seen in carriers of BRCA1 or BRCA2 pathogenic variants, as well as risks for other cancers such as sarcomas, brain tumors, and carcinomas of the stomach, thyroid, and small intestine.

Clinicians faced with a family with multiple affected individuals with breast cancer often look for distinguishing signs in the patient and family history to help guide the choice of which genes to test. However, the rapid decline in the cost of gene and exome sequencing has allowed the development of gene panels in which multiple genes can be simultaneously analyzed, often at a cost that is equivalent to or even less than what was charged previously to analyze just one or two genes. Many breast and ovarian cancer panels include genes associated with moderately increased risk of breast and ovarian cancer (i.e., ATM, CHEK2, PALB2, BRIP1, RAD51C, and RAD51D).

Hereditary Colon Cancer

Colorectal cancer, a malignancy of the epithelial cells of the colon and rectum, is one of the most common forms of cancer. It affects ~1.3 million individuals worldwide per year (150,000 of whom are in the United States) and is responsible for ~10% to 15% of all cancer. Most cases are sporadic, but a small proportion of colon cancer cases are familial, among which are two autosomal dominant conditions: FAP and LS, along with their variants.

Familial Adenomatous Polyposis. FAP and its sub variant, Gardner syndrome, together have an incidence of ~1 per 10,000. In FAP, benign adenomatous polyps numbering in the many hundreds develop in the colon during the first 2 decades of life. In almost all cases, one or more of the polyps become malignant. Surgical removal of the colon (colectomy) prevents the development of colorectal malignancy.

FAP is caused by autosomal dominantly inherited heterozygous loss- of- function variants in a TSG known as APC (so- named because the condition used to be called adenomatous polyposis coli). Gardner syndrome is also due to pathogenic variants in APC and is therefore allelic to FAP. Patients with Gardner syndrome have, in addition to the adenomatous polyps with malignant transformation seen in FAP, extracolonic anomalies, including osteomas of the jaw and desmoids, which are tumors arising in the muscle of the abdominal wall. Although the relatives of an individual affected with Gardner syndrome who also carry the same APC pathogenic variant tend to also show the extracolonic manifestations of Gardner syndrome, the same variant in unrelated individuals has been found to cause only FAP. Thus whether an individual has FAP or Gardner syndrome is not simply due to which pathogenic variant is present in the APC gene but is likely affected by variation elsewhere in the genome.

Lynch Syndrome. Approximately 2% to 4% of cases of colon cancer are attributable to LS. LS is characterized by autosomal dominant inheritance of colon cancer in association with a small number of adenomatous polyps that begin during early adulthood. The number of polyps is generally quite small in com parison to the hundreds to thousands of adenomatous polyps seen with FAP. Nonetheless, polyps in LS have high potential to undergo malignant transformation. Heterozygotes for pathogenic variants in MLH1, one of the most penetrant LS genes, have an ~80% lifetime risk for colon cancer; female heterozygotes also have a ~40% risk for endometrial cancer. There are also additional risks of 10% to 20% for cancer of the biliary or urinary tract and the ovary. Sebaceous gland tumors of the skin may be the first presenting sign in LS (in which case it is a variant called Muir- Torre syndrome); thus the presence of such tumors in a patient should raise suspicion of a possible hereditary colon cancer syndrome.

LS results from loss- of- function variants in one of four DNA repair genes (MLH1, MSH2, MSH6, and PMS2) that encode MMR proteins. Although all four of these genes have been implicated in LS in different families, MLH1 and MSH2 are together responsible for the majority of LS, whereas MSH2 and PMS2 are often associated with a lesser degree of MMR deficiency and lower penetrance. Like BRCA1 and BRCA2, the LS MMR genes are TSGs involved in maintaining the integrity of the genome. Unlike BRCA1 and BRCA2, however, the LS genes are not involved in double- stranded DNA break repair. Instead, their role is to repair incorrect DNA base pairing (i.e., pairing other than A with T or C with G) that can arise during DNA replication.

At the cellular level, the most striking phenotype of cells lacking MMR proteins is an enormous increase in both point mutations and mutations occurring during replication of simple DNA repeats, such as segments containing a string of the same base, for example (A)n , or a microsatellite, such as (TG)n. Microsatellites are believed to be particularly vulnerable to mismatch because slippage of the strand being synthesized on the template strand can occur more readily when a short tandem repeat is being synthesized. Such instability, referred to as the microsatellite instability- positive (MSI+) phenotype, occurs at two orders of magnitude higher frequency in cells lacking both copies of an MMR gene. The MSI+ phenotype is easily seen in DNA as three, four, or even more alleles of a micro satellite polymorphism in a single individual’s tumor DNA (Fig. 2). It is estimated that cells lacking both copies of an MMR gene may carry 100,000 mutations within simple repeats throughout the genome.

Fig2. Gel electrophoresis of three different microsatellite polymorphic markers in normal (N) and tumor (T) samples from a patient with a germline pathogenic variant in MSH2 and microsatellite instability. Although marker 2 shows no difference between normal and tumor tissues, genotyping at markers 1 and 3 reveals extra alleles (blue arrows), some smaller, some larger, than the alleles present in normal tissue.

Because of the increased mutation rate in these classes of sequence, loss of function of MMR genes will lead to somatic mutations in other driver genes. Two such driver genes have been isolated and characterized. The first is APC, whose normal function and role in FAP were described previously. The second is the gene TGFBR2, in which heterozygous germline pathogenic variants primarily cause a connective tissue disorder called Loeys- Dietz syndrome; however, cases of early- onset colon cancer have also been reported. TGFBR2 encodes trans forming growth factor β receptor II, a serine- threonine kinase that inhibits intestinal cell division. Somatically, TGFBR2 is particularly vulnerable to mutation when MMR proteins are lost because it contains a stretch of 10 adenines encoding three lysines within its coding sequence; deletion of one or more of these As results in a frameshift and loss- of- function mutation. LS is an excellent example of how a gene, like MLH1, which has a global effect on mutation rate throughout the genome, can be a driver gene through its effect on other genes, such as TGFBR2, that are more specifically involved in driving the development of a cancer.

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مواضيع ذات صلة

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Monogenic Diseases Show One of Three Patterns of Inheritance

Activated Oncogenes in Hereditary Cancer Syndromes

Genetic Basis of Cancer

Conditional Mutations Can Be Used to Study Essential Genes in Yeast

Segregation of Mutations in Breeding Experiments Reveals Their Dominance or Recessivity

Recessive and Dominant Mutant Alleles Generally Have Opposite Effects on Gene Function

Genes and Environment in Development

Gene Therapy

The Genetics of mtDNA Disease

The APOE Gene Is an Alzheimer Disease Susceptibility Locus

The Genetics of Alzheimer Disease