Учебники / Genetics and Auditory Disorders Keats 2002

Studying hearing loss in aneuploidies does not lend itself easily to a better understanding of hearing and deafness in general. Because of the wide spectrum of malformations in most patients with chromosomal aneuploidies, and the various stages of development that are affected, it is difficult to determine which genes on the extra or missing chromosome influence the normal development or function of any particular organ system. However, because of the prevalence of hearing loss in individuals with aneuploidy syndromes, it represents a significant aspect of health care management of such patients. Undiagnosed hearing loss adds another invisible burden for individuals who are already challenged with mental retardation or physical difficulties.

2.2.1.1 Trisomy 21 and Hearing Loss

Down syndrome occurs in about 1 in 700 newborns (Fig. 5.3). Individuals with Down syndrome have a host of clinical findings, including mental retardation, short stature, hypotonia, characteristic facial features, cataracts, heart defects, thyroid disorders, an increased incidence of leukemia, and premature aging (Jones 1997). Hearing loss is found in about 40% to 80% of patients, depending on the threshold level used and method of testing (Roizen et al. 1993). The hearing loss is generally mild, bilateral conductive; however, sensorineural hearing loss was detected in 34% of children evaluated in one study (Roizen et al. 1993). The external ears of Down syndrome patients tend to be simple and low set with an overfolded upper helix and small or absent lobes (Jones 1997). Children tend to have dysfunctional Eustachian tubes, leading to a high incidence of otitis media and accounting for some of the conductive hearing loss (Roizen et al. 1993). The pathogenesis of the sensorineural hearing loss in individuals with Down syndrome has not been determined, although a gene dosage effect of a gene on chromosome 21 must be considered a likely mechanism.

2.2.1.2 Trisomy 13 and Hearing Loss

With an incidence of 1 in 12,000 (Hook 1980), trisomy 13 is much rarer in liveborns than is trisomy 21, and the phenotype is much more severe. Ninety percent of trisomy 13 infants die before 6 months of age. The infants are severely retarded due to various types of forebrain defects. More than 50% of cases are noted to have eye defects ranging from anopthalmia to micropthalmia, cleft lip and/or cleft palate, polydactyly, microcephaly, heart defects, renal anomalies, and deafness (Jones 1997).

Individuals with trisomy 13 mosaicism may show a less severe phenotype than those with a full trisomy. Presumably, the presence of a normal cell line, especially in particular tissues and organs, mitigates the effects of the trisomic cells. There have been at least two reports of hearing loss or deafness in patients with mosaic trisomy 13 (Delatycki and Gardner 1997), but the mechanism of this pathology remains unclear.

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2.2.1.3 Turner Syndrome and Hearing Loss

Turner syndrome, as illustrated in Figure 5.4, is characterized by monosomy X (45,X) in 50% of cases and various mosaicisms or structural abnormalities of the other X chromosome in the remaining cases. The incidence is about 1 in 2,500 live births, but the frequency at conception is much higher. Greater than 95% of 45,X concepti result in spontaneous loss (Gardner and Sutherland 1996). Females with Turner syndrome are characterized by small stature, streak gonads, short or webbed neck, broad chest with widely spaced nipples, prominent ears, epicanthal folds, and a high-arched palate. Coarctation of the aorta and renal anomalies are common findings. Intelligence is usually within normal limits (Jones 1997). About half of Turner syndrome females have moderate sensorineural hearing loss, often combined with conductive hearing loss. Chronic or recurrent ear infections are common in childhood and may account for some of the conductive hearing loss (Gorlin et al. 1995). The nature of the sensorineural hearing loss is unknown.

2.2.2 Unique Chromosomal Rearrangements Associated with Hearing Loss

There are a number of reports in the literature of various chromosomal rearrangements and hearing loss. Unlike aneuploidies and microdeletion syndromes, these are not disorders that have been seen repeatedly. Instead, they are rearrangements that are either unique, or reported in only a few individuals. In each case, the chromosomal rearrangement was discovered either at amniocentesis, or in the evaluation of a dysmorphic, developmentally delayed child. Hearing loss is usually just one finding of many physical abnormalities. All the children have various degrees of mental retardation and dysmorphic features. Hearing loss is probably underdiagnosed among infants born with unusual chromosomal rearrangements, but given the severity of the physical problems present at birth a hearing test may not be considered a high priority.

2.2.2.1 An Unusual Marker Chromosome 15

Marker chromosomes are chromosomes that are structurally abnormal. They are often difficult to identify, and specific tests are required to determine the origin of the chromosomal material. The prevalence of marker chromosomes is less than 0.7 per 1,000 births (Gardner and Sutherland 1996). An inverted duplication of chromosome 15 is among the more common markers. Generally, the centromere and the proximal portion of the q arm are duplicated and inverted. Small inverted duplications may have no phenotypic effect, whereas larger ones produce characteristic mental retardation and dysmorphic features (Gardner and Sutherland 1996). Hearing loss among individuals with the “common” inverted duplication of chromosome 15 is not a typical finding. However, Huang et al.

(1998) reported an unusual inverted duplication of chromosome 15 involving the distal portion of chromosome 15q, rather than the proximal, from q25 to qter (Fig. 5.5A). The infant had severe hypotonia, cardiovascular defects, CNS anomalies, and dysmorphic facies. Severe hearing loss was also present, as determined by auditory evoked brainstem response. The child had such severe developmental anomalies that she died at 12 days of life.

2.2.2.2 Inversion 2

Pericentric inversions are the most frequent chromosomal rearrangements in humans, occurring in approximately 1% of the population (Therman and Susman 1993). The majority are inherited and clinically insignificant. Pericentromeric inversions of chromosome 2 are the second most commonly recognized inversion of human chromosomes, after pericentromeric inversion of chromosome 9. Usually, the breakpoints are in p11q13. Kozma et al. (1996) reported an unusual inversion of chromosome 2, with breakpoints in p13q11.2 (Fig. 5.5B). The child had craniofacial anomalies, significant hypotonia, developmental delay, and severe to profound bilateral hearing loss.

2.2.2.3 Partial Trisomy 6q

Conrad et al. (1998) reported a toddler with a partial trisomy of approximately the lower third of chromosome 6 (Fig. 5.5C). The child had microcephaly, facial anomalies, a webbed neck, congenital heart disease, renal hypoplasia, developmental delay, and bilateral hearing loss. The additional portion of chromosome 6 was translocated to the short arm of chromosome 14 and was inherited from the child’s father, who had an apparently balanced translocation between 6q22 and 14p13.

2.2.2.4 A Tandem Duplication and Deletion

Meschede et al. (1998) reported a translocation between two acrocentric chromosomes, 14 and 21, with essentially the entire chromosome 21 translocated to the telomeric end of 14q, resulting in a small deletion of 14q32.3 (Fig. 5.6). The phenotype included developmental delay, severe hypotonia, mild facial dysmorphism, growth retardation, hypospadias, palmar creases, marbled skin and a patent ductus arteriosus. Marked hearing loss required hearing aids. A few other deletions of the very terminal portion of 14q have been reported (Meschede et al. 1998). Although this is the only case in which hearing loss was documented, it is possible that hearing evaluations were not performed in all other cases.

2.2.3 Chromosomal Rearrangement Syndromes and Hearing Loss

The majority of chromosomal deletions result in partial monosomy for a particular chromosomal region. Deletions may result from the unbalanced segregation of a parental reciprocal translocation, or occur de novo.

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FIGURE 5.5. Novel chromosome rearrangements that cause hearing loss, among other physical findings. (A) Marker chromosome of an inverted duplication of 15q25–q26, on right, as compared with a normal chromosome 15 (left). The patient described (Huang et al. 1998) had two normal chromosomes 15 in addition to the marker 15. (B) Normal chromosome 2, on left, as compared with inverted chromosome 2 (Kozma et al. 1996). (C) Partial trisomy of 6q described by Conrad et al. (1998) as a result of a 6;14 translocation. Arrows indicate the chromosomal breakpoints. The child had two normal chromosomes 6 (on left), one normal chromosome 14 (middle) and one derivative 14, with additional 6q material attached to 14p.

FIGURE 5.6. Translocation between chromosomes 14 and 21. (A) The patient at three years of age (From Meschede et al. 1998, Submicroscopic deletion in 14q32.3 through a de novo tandem translocation between 14q and 21p, American Journal of Medical Genetics 80:443–7, Copyright 1998 John Wiley & Sons, Inc. Reprinted by permission of Wiley-Liss, Inc., a subsidiary of John Wiley & Sons, Inc.). Notice the posteriorly rotated, low-set ears and the need for a hearing aide. (B) Ideograms representing the patient’s translocation. Arrows indicate the chromosomal breakpoints. The child has one normal chromosome 14 (left), one normal 21 (middle), and one translocated chromosome, t(14;21) (right), which results in monosomy for the distal portion of 14q.

Terminal deletions, as well as many interstitial deletions, have been described (Therman and Susman 1993). Some deletions, such as those at 15q11-q13 or 22q11, associated with the Prader-Willi/Angelman syndromes or DiGeorge/velocardiofacial syndromes, respectively, are relatively common. Others, such as 5p- (cri-du-chat syndrome) or 4p- (Wolf– Hirschorn syndrome), are rarely seen.

Many deletions or smaller microdeletions have been observed often enough to be categorized into known clinical syndromes, i.e., similar deletions producing similar phenotypic patterns. The following sections will

the deletion may frequently be missed upon standard cytogenetic analysis. Shapira et al. (1997) estimate its incidence at more than 1 in 1,000, which would make 1p36 deletions one of the more common cytogenetic deletion syndromes. The physical findings in this syndrome can be somewhat vague and variable, depending on the size of the deletion. Hypotonia, moderate mental retardation, mild facial dysmorphism, abusive behavior, and hearing loss are all part of the spectrum of physical findings. Interestingly, a preponderance of maternally derived deletions has been observed (Wu et al. 1999).

By studying a series of deletions of various sizes, Wu et al. (1999) describe the smallest deletion in which sensorineural hearing loss is present.

2.2.3.2 DiGeorge Syndrome/Velocardiofacial Syndrome

DiGeorge syndrome/velocardiofacial syndrome (DGS1/VCFS) was originally thought to be two separate entities until both were found to be caused by deletions in 22q11. Thus, they are now viewed collectively with variable phenotypes, depending on the deletion size and genetic background. Ninety percent of deletions occur de novo (Gardner and Sutherland 1996). Clinical findings include thymus deficiency, conotruncal heart anomalies, mildly dysmorphic facies, hypoparathyroidism, palatal anomalies and deafness (Hong 1998). Heart disease is the leading cause of death among DGS1/VCFS infants. The phenotype may be highly variable, and members of the same family, presumably with identical deletions, have variable expressivity of DGS1/VCFS features (Gardner and Sutherland 1996). The majority of patients are growth delayed, and most also experience learning disabilities (Gorlin et al. 1995). The underlying embryological defect is thought to be improper development of the facial neural crest tissues, resulting in defective neural pouch derivatives (Lammer and Opitz 1986).

Cytogenetic analysis of DGS1/VCFS individuals reveals a deletion on one chromosome 22 at band q11 in approximately 33% of cases (Fig. 5.7B). Often the deletion is cryptic, being so small that it can only be observed with molecular probes. By molecular studies, at least 90% of DGS1/VCFS patients have been found to have deletions.

The hearing loss seen in DGS1/VCFS can range from mild to severe and is usually conductive, though it can be sensorineural. Defects detected are often structural, owing to interference of development of the 3rd and 4th pharyngeal pouches (Ohtani and Schuknecht 1984). Mondini deformity is found, as well as malformed ossicles, and external auditory canal anomalies.

An additional DGS2/VCFS deletion locus is located on 10p. It has been found de novo as an interstitial or terminal deletion of 10p (Fig. 5.7B)

(Schuffenhauer et al. 1998), or as the result of an inherited, unbalanced translocation (Hon et al. 1995). In addition to the spectrum of features found in DGS1/VCFS patients with the 22q11 deletion, patients with the 10p deletion can also demonstrate microcephaly, hand and foot anomalies, genitourinary defects, severe psychomotor retardation and sensorineural deafness (Shapira et al. 1994). Similar to individuals with 22q11 deletions,

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10p patients can have highly variable expressivity of any or all of these findings. Schuffenhaur et al. analyzed the deletions of 12 DGS2 patients, three of whom had sensorineural hearing loss, and determined that the hearing loss gene(s) within the DGS2 locus on 10p must reside just distal to marker D10S1705 (Schuffenhauer et al. 1998).

2.2.3.3 Smith-Magenis Syndrome

Smith-Magenis syndrome is a contiguous gene-deletion syndrome originally described in 1982 and resulting from interstitial deletions in 17p11.2 (Fig. 5.7C). The phenotype includes brachycephaly, midface hypoplasia, prognathism, deep hoarse voice, psychomotor and growth retardation, and behavioral problems that include self-injurious behavior and sleep disorders (OMIM 1999). Greenberg et al. (1996) found that 68% of Smith-Magenis patients have hearing loss, with approximately two-thirds of those having conductive loss and one-third having sensorineural hearing loss.

The nonsyndromic autosomal-recessive hearing loss locus DFNB3 was mapped to the pericentromeric region of chromosome 17 (Friedman et al. 1995) and then refined to 17p11.2, within the deletion interval for SmithMagenis syndrome (Liang et al. 1998). Later that same year, DFNB3 was shown to be caused by mutations in the gene encoding the unconventional myosin, MYO15 (Wang et al. 1998). Interestingly, several Smith–Magenis patients who have sensorineural hearing loss and deletion of one allele of MYO15 also have a point mutation in the remaining allele of MYO15 (Liang et al. 1998). Deletion of the Smith–Magenis region uncovered the recessive mutation, making the patients hemizygous for the mutated allele.

2.3 Use of Cytogenetics to Help Identify Candidate Genes

An extremely useful aspect of cytogenetics when studying any genetic disease is that it can lead to identification or refinement of a disease locus. Chromosomal rearrangements or deletions that disrupt critical genes can be the first clue to the locus of a disease gene. This approach was successful in studies of Waardenburg syndrome type 1, where linkage analyses had been unproductive, apparently due to locus heterogeneity (Mueller, Van Camp, and Lench, Chapter 4). In both Branchio-Oto-Renal syndrome and X-linked mixed deafness, deletions found in the chromosomes of affected individuals allowed refinement of the disease intervals, ultimately leading to cloning of the genes.

2.3.1 Waardenburg Syndrome Type 1

Waardenburg syndrome is an autosomal-dominant disorder that commonly manifests as deafness with pigmentary anomalies. Ishikiriyama et al. (1989) reported a Japanese child with a phenotype consistent with Waardenburg

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suggested that WS1 may be homologous to the Splotch mouse that maps to a region of homologous synteny on mouse chromosome 1. The Splotch mouse has pigmentary anomalies similar to those seen in humans with WS1, but, interestingly, does not have any hearing deficit. When the mouse Pax3 gene was cloned and shown to cause the Splotch phenotype (Epstein et al. 1991; Goulding et al. 1991), the human homolog, HuP2 or PAX3, was evaluated as a candidate gene in several WS1 families by heteroduplex analysis (Mueller, Van Camp, and Lench, Chapter 4). Band shifts were observed that showed complete concordance with the WS1 phenotype. The PAX3 gene was sequenced in these families and found to contain disease causing mutations (Tassabehji et al. 1992). To date, dozens of mutations in PAX3 have been found that cause WS1. Identification of PAX3 as the WS1 gene was facilitated greatly by the cytogenetic finding of the inv(2)(q35q37.3).

2.3.2 X-Linked Mixed Deafness

Loci for a number of disorders have been mapped to Xq21 by linkage analysis and translocation studies, including retinal dystrophy choroideremia, mental retardation, cleft lip and palate, and mixed deafness with stapes fixation and perilymphatic gusher (DFN3) (OMIM 1999). DFN3 is the most common form of X-linked hearing impairment. Physical mapping of the Xq21 region using patients with these disorders and with cytogenetically visible deletions of Xq21 allowed successive refinement of the locus for DFN3 (Bach et al. 1992; Cremers et al. 1989), until ultimately it was reduced to 500 kb (Fig. 5.9) (Huber et al. 1994). When the murine Pou3f4 gene was cloned and mapped to the mouse X chromosome in a region of homologous synteny with human Xq21 (Douville et al. 1994), POU3F4 became a positional candidate gene for DFN3. Radiolabled mouse Pou3f4 probes hybridized to Southern blots of genomic DNA from DFN3 males with cytogenetically visible deletions, failed to detect any restriction fragments, suggesting the orthologous POU3F4 gene was deleted in these individuals.The mouse sequence was used to make primers to amplify and clone the human POU3F4 gene. SSCP analysis showed frameshift mutations in POU3F4 in four DFN3 patients who did not have cytogenetically visible deletions (de Kok et al. 1995),confirming that loss of POU3F4 in the deletion patients was responsible for their hearing loss. Additional DFN3 patients have been identified subsequently who have deletions in Xq21 that do not encompass POU3F4,but delete regions centromeric to the gene. These patients may harbor deletions in unidentified POU3F4 regulatory sequences, or another gene whose product is similar to or interacts with the POU3F4 protein (de Kok et al. 1996).

2.3.3 Branchio-Oto-Renal Syndrome

The autosomal dominant Branchio-Oto-Renal (BOR) syndrome is the association of branchial arch anomalies, such as branchial cysts or fistulas,