3648 PART 16 Genes, the Environment, and Disease

intron 2

β-Globin Gene Cluster

–10 kb 0 kb 10 kb 20 kb 30 kb 40 kb 50 kb 60 kb

Promoter 5'UTR intron 1 Poly A

ε Gγ Aγ ψβ β δ

1

1 2 3

A *

FIGURE 466-9 Point mutations causing a thalassemia as example of allelic heterogeneity. The β-globin gene is

located in the globin gene cluster. Point mutations can be located in the promoter, the CAP site, the 5′-untranslated

region, the initiation codon, each of the three exons, the introns, or the polyadenylation signal. Many mutations

introduce missense or nonsense mutations, whereas others cause defective RNA splicing. Not shown here are

deletion mutations of the β-globin gene or larger deletions of the globin locus that can also result in thalassemia.

, promoter mutations; *, CAP site; , 5′UTR; 1 , initiation codon; , defective RNA processing; , missense and

nonsense mutations; A , Poly A signal.

Wild-type

A B

AA

DNA

A

GCA

Wild-type

F

T T C

T

A C C

D

G A C

F

T T C

I

A T A

C

T G C

F

T T C

T

A C C

D

G A C

T A C

Y

F

T T C

I

A T A

C

T G C

Heterozygous point mutation

F

T T C

T

A C C

Y

T A C

F

T T C

I

A T A

C

T G C

Homozygous point mutation

L

CTC

L

CTA

S

TCG

H

CAC

A

GCT

R

CGG

E

GAG

G

GGC

E

GAA

N

AAT

E

GAG

S

AGC

Silent mutation

AA

DNA

A

GCA

L

CTC

L

CTA

S

TCG

H

CAC

A

GCT

R

CGT

E

GAG

G

GGC

E

GAA

N

AAT

E

GAG

S

AGC

Missense mutation

AA

DNA

A

GCA

L

CTC

L

CTA

S

TCG

H

CAC

A

GCT

P

CCG

E

GAG

G

GGC

E

GAA

N

AAT

E

GAG

S

AGC

Nonsense mutation

AA

DNA

A

GCA

L

CTC

L

CTA

S

TCG

H

CAC

A

GCT

R

CGG

E

GAG

G

GGC

X

TAA AAT GAG AGC

1 bp Deletion with frameshift

AA

DNA

A

GCA

L

CTC

L

CTA

R

CGC

T

ACG

L

CTC

G

GGG

R

AGG

A

GCG

K

AAA

M

ATG

R

AGA GC

FIGURE 466-10 A. Examples of mutations (now commonly referred to as variations). The coding strand is shown with the encoded amino acid sequence. B. Chromatograms

of sequence analyses after amplification of genomic DNA by polymerase chain reaction.

a strong predisposition to develop aplastic anemia and acute myelogenous leukemia (Chap. 104). Cells from these patients are susceptible

to chromosomal breaks caused by a defect in genetic recombination. It

can be caused by mutations in the multiple genes forming the Fanconi’s

anemia pathway, which is involved in DNA repair and replication.

HNPCC (Lynch syndrome) is characterized by autosomal dominant

transmission of colon cancer, young age (<50 years) of presentation,

predisposition to lesions in the proximal large bowel, and associated

malignancies such as uterine cancer and ovarian cancer. HNPCC is

predominantly caused by mutations in one of several different mismatch repair (MMR) genes including MutS homologue 2 (MSH2),

MutL homologue 1 and 6 (MLH1, MLH6), MSH6, PMS1, and PMS2

(Chap. 81). These proteins are involved in the detection of nucleotide

mismatches and in the recognition of slipped-strand trinucleotide

repeats. Germline mutations in these genes lead to microsatellite

instability and a high mutation rate in colon cancer. Genetic screening

tests for this disorder are now being used for families considered to be

at risk. Recognition of HNPCC allows

early screening with colonoscopy and the

implementation of prevention strategies

using nonsteroidal anti-inflammatory

drugs.

UNSTABLE DNA SEQUENCES Trinucleotide repeats may be unstable and

expand beyond a critical number. Mechanistically, the expansion is thought to

be caused by unequal recombination

and slipped mispairing. A premutation

represents a small increase in trinucleotide copy number. In subsequent

generations, the expanded repeat may

increase further in length and result in

an increasingly severe phenotype, a process called dynamic mutation (see below

for discussion of anticipation). Trinucleotide expansion was first recognized as a

cause of the fragile X syndrome, one of

the most common causes of intellectual

disability. Other disorders arising from a

similar mechanism include Huntington’s

disease, X-linked spinobulbar muscular atrophy, and myotonic dystrophy.

Malignant cells are also characterized by genetic instability, indicating a

breakdown in mechanisms that regulate DNA repair and the cell cycle.

Functional Consequences of Mutations Functionally, mutations can be broadly classified as gain-of-function and loss-of-function

mutations. Gain-of-function mutations are typically dominant (e.g.,

they result in phenotypic alterations when a single allele is affected).

Inactivating mutations are usually recessive, and an affected individual

is homozygous or compound heterozygous (e.g., carrying two different

mutant alleles of the same gene) for the disease-causing mutations.

Alternatively, mutation in a single allele can result in haploinsufficiency,

a situation in which one normal allele is not sufficient to maintain

a normal phenotype. Haploinsufficiency is a commonly observed

mechanism in diseases associated with mutations in transcription

factors (Table 466-2). Remarkably, the clinical features among patients

with an identical mutation often vary significantly. One mechanism

underlying this variability consists in the influence of modifying

Principles of Human Genetics

3649CHAPTER 466

genes. Haploinsufficiency can also affect the expression of rate-limiting

enzymes. For example, haploinsufficiency in enzymes involved in

heme synthesis can cause porphyrias (Chap. 416).

An increase in dosage of a gene product may also result in disease, as

illustrated by the duplication of the DAX1 gene in dosage-sensitive sex

reversal (Chap. 390). Mutation in a single allele can also result in loss

of function due to a dominant-negative effect. In this case, the mutated

allele interferes with the function of the normal gene product by one

of several different mechanisms: (1) a mutant protein may interfere

with the function of a multimeric protein complex, as illustrated by

mutations in type 1 collagen (COL1A1, COL1A2) genes in osteogenesis

imperfecta (Chap. 413); (2) a mutant protein may occupy binding sites

on proteins or promoter response elements, as illustrated by thyroid

hormone resistance β, a disorder in which inactivated thyroid hormone

receptor β binds to target genes and functions as an antagonist of normal receptors (Chap. 382); or (3) a mutant protein can be cytotoxic as

in α1

 antitrypsin deficiency (Chap. 292) or autosomal dominant neurohypophyseal diabetes insipidus (Chap. 381), in which the abnormally

folded proteins are trapped within the endoplasmic reticulum and

ultimately cause cellular damage.

Genotype and Phenotype • ALLELES, GENOTYPES, AND HAPLOTYPES An observed trait is referred to as a phenotype; the genetic

information defining the phenotype is called the genotype. Alternative

forms of a gene or a genetic marker are referred to as alleles. Alleles

may be polymorphic variants of nucleic acids that have no apparent

effect on gene expression or function. In other instances, these variants

may have subtle effects on gene expression, thereby conferring adaptive advantages associated with genetic diversity. On the other hand,

allelic variants may reflect mutations that clearly alter the function of

a gene product. The common Glu6Val (E6V) sickle cell mutation in

the β-globin gene and the ΔF508 deletion of phenylalanine (F) in the

CFTR gene are examples of allelic variants of these genes that result in

disease. Because each individual has two copies of each chromosome

(one inherited from the mother and one inherited from the father), an

individual can have only two alleles at a given locus. However, there can

be many different alleles in the population. The normal or common

allele is usually referred to as wild type. When alleles at a given locus

are identical, the individual is homozygous. Inheriting identical copies

of a mutant allele occurs in many autosomal recessive disorders, particularly in circumstances of consanguinity or isolated populations. If the

alleles are different on the maternal and the paternal copy of the gene,

the individual is heterozygous at this locus (Fig. 466-10). If two different

mutant alleles are inherited at a given locus, the individual is said to be

a compound heterozygote. Hemizygous is used to describe males with a

mutation in an X chromosomal gene or a female with a loss of one X

chromosomal locus.

Genotypes describe the specific alleles at a particular locus. For

example, there are three common alleles (E2, E3, E4) of the apolipoprotein E (APOE) gene. The genotype of an individual can therefore

be described as APOE3/4 or APOE4/4 or any other variant. These designations indicate which alleles are present on the two chromosomes

in the APOE gene at locus 19q13.2. In other cases, the genotype might

be assigned arbitrary numbers (e.g., 1/2) or letters (e.g., B/b) to distinguish different alleles.

A haplotype refers to a group of alleles that are closely linked together

at a genomic locus (Fig. 466-4). Haplotypes are useful for tracking the

transmission of genomic segments within families and for detecting

evidence of genetic recombination, if the crossover event occurs

between the alleles (Fig. 466-6). As an example, various alleles of the

histocompatibility locus antigens (HLA) at the major histocompatibility complex (MHC) on chromosome 6 are used to establish haplotypes

associated with certain disease states. For example, 21-hydroxylase

deficiency, complement deficiency, and hemochromatosis are each

associated with specific HLA haplotypes. It is now recognized that

these genes lie in close proximity to the HLA locus, which explains why

HLA associations were identified even before the disease genes were

cloned and localized. In other cases, specific HLA associations with

diseases such as ankylosing spondylitis (HLA-B27) or type 1 diabetes

mellitus (HLA-DR4) reflect the role of specific HLA allelic variants

in susceptibility to these autoimmune diseases. The characterization

of common SNP haplotypes in numerous populations from different

parts of the world has provided the necessary tools for association studies designed to detect genes involved in the pathogenesis of complex

disorders (Table 466-1). The presence or absence of certain haplotypes

can also be relevant for the customized choice of medical therapies

(pharmacogenomics) or may have value for preventive strategies.

Genotype-phenotype correlation describes the association of a specific mutation and the resulting phenotype. The phenotype may differ

depending on the location or type of the mutation in some genes. For

example, in von Hippel–Lindau disease, an autosomal dominant multisystem disease that can include renal cell carcinoma, hemangioblastomas, and pheochromocytomas, among others, the phenotype varies

greatly and the identification of the specific mutation can be clinically

useful in order to predict the phenotypic spectrum.

ALLELIC HETEROGENEITY Allelic heterogeneity refers to the fact that

different mutations in the same genetic locus can cause an identical

or similar phenotype. For example, many different mutations of the

β-globin locus can cause β thalassemia (Table 466-3) (Fig. 466-9).

In essence, allelic heterogeneity reflects the fact that many different

mutations are capable of altering protein structure and function. For

this reason, maps of inactivating mutations in genes usually show a

near-random distribution. Exceptions include (1) a founder effect, in

which a particular mutation that does not affect reproductive capacity

can be traced to a single individual; (2) “hot spots” for mutations, in

which the nature of the DNA sequence predisposes to a recurring

mutation; and (3) localization of mutations to certain domains that are

particularly critical for protein function. Allelic heterogeneity creates a

practical problem for genetic testing because one must often examine

the entire genetic locus for mutations, because these can differ in each

patient. For example, ~2000 variants have been identified in the CFTR

gene to date, although some of them are very rare and some may not

be disease-causing (Fig. 466-3). Mutational analysis may initially focus

on a panel of mutations that are particularly frequent (often taking the

ethnic background of the patient into account), but a negative result

does not exclude the presence of a mutation elsewhere in the gene.

One should also be aware that mutational analyses tend to focus on the

coding region of a gene without considering regulatory and intronic

regions. Because disease-causing mutations may be located outside the

coding regions, negative results need to be interpreted with caution.

The advent of more comprehensive sequencing technologies now

greatly facilitates concomitant mutational analyses of several genes

after targeted enrichment or even mutational analysis of the whole

exome or genome. However, comprehensive sequencing can result in

significant diagnostic challenges because the detection of a sequence

alteration is not always sufficient to establish that it has a causal role

(variants of unknown significance [VUS]).

PHENOTYPIC HETEROGENEITY Phenotypic heterogeneity occurs when

more than one phenotype is caused by allelic mutations (e.g., by different mutations in the same gene) (Table 466-3). For example, laminopathies are monogenic multisystem disorders that result from mutations

in the LMNA gene, which encodes the nuclear lamins A and C. Multiple

autosomal dominant and recessive disorders are caused by mutations

in the LMNA gene. They include several forms of lipodystrophies,

Emery-Dreifuss muscular dystrophy, progeria syndromes, a form

of neuronal Charcot-Marie-Tooth disease (type 2B1), and a group

of overlapping syndromes. Remarkably, hierarchical cluster analysis

has revealed that the phenotypes vary depending on the position of

the mutation (genotype-phenotype correlation). Similarly, identical

mutations in the FGFR2 gene can result in very distinct phenotypes:

Crouzon’s syndrome (craniofacial synostosis) or Pfeiffer’s syndrome

(acrocephalopolysyndactyly).

LOCUS OR NONALLELIC HETEROGENEITY AND PHENOCOPIES Nonallelic or locus heterogeneity refers to the situation in which a similar

disease phenotype results from mutations at different genetic loci

(Table 466-3). This often occurs when more than one gene product

3650 PART 16 Genes, the Environment, and Disease

produces different subunits of an interacting complex or when different genes are involved in the same genetic cascade or physiologic

pathway. For example, osteogenesis imperfecta can arise from mutations in two different procollagen genes (COL1A1 or COL1A2) that

are located on different chromosomes and can involve multiple other

genes (Chap. 413). The effects of inactivating mutations in these two

genes are similar because the protein products comprise different

subunits of the helical collagen fiber. Similarly, muscular dystrophy

syndromes can be caused by mutations in various genes, consistent

with the fact that it can be transmitted in an X-linked (Duchenne or

Becker), autosomal dominant (limb-girdle muscular dystrophy type 1),

or autosomal recessive (limb-girdle muscular dystrophy type 2) manner (Chap. 449). Mutations in the X-linked DMD gene, which encodes

dystrophin, are the most common cause of muscular dystrophy. This

feature reflects the large size of the gene as well as the fact that the

phenotype is expressed in hemizygous males because they have only

a single copy of the X chromosome. Dystrophin is associated with a

large protein complex linked to the membrane-associated cytoskeleton

in muscle. Mutations in several different components of this protein

complex can also cause muscular dystrophy syndromes. Although

the phenotypic features of some of these disorders are distinct, the

phenotypic spectrum caused by mutations in different genes overlaps,

thereby leading to nonallelic heterogeneity. It should be noted that

mutations in dystrophin are also associated with allelic heterogeneity.

For example, mutations in the DMD gene can cause either Duchenne’s

or the less severe Becker’s muscular dystrophy, depending on the severity of the protein defect.

Recognition of nonallelic heterogeneity is important for several reasons: (1) the ability to identify disease loci in linkage studies is reduced

by including patients with similar phenotypes but different genetic

disorders; (2) genetic testing is more complex because several different

genes need to be considered along with the possibility of different

mutations in each of the candidate genes; and (3) novel information is

gained about how genes or proteins interact, providing unique insights

into molecular physiology.

Phenocopies refer to circumstances in which nongenetic conditions

mimic a genetic disorder. For example, features of toxin- or druginduced neurologic syndromes can resemble those seen in Huntington’s

disease, and vascular causes of dementia share phenotypic features with

familial forms of Alzheimer’s dementia (Chap. 431). As in nonallelic

heterogeneity, the presence of phenocopies has the potential to confound linkage studies and genetic testing. Patient history and subtle

TABLE 466-3 Selected Examples of Phenotypic Heterogeneity and Locus Heterogeneity

Phenotypic Heterogeneity

GENE, PROTEIN PHENOTYPE INHERITANCE OMIM

LMNA, Lamin A/C Emery-Dreifuss muscular dystrophy (AD) AD 181350

Familial partial lipodystrophy Dunnigan AD 151660

Hutchinson-Gilford progeria AD 176670

Atypical Werner’s syndrome AD 150330

Dilated cardiomyopathy 1A AD 115200

Familial atrial fibrillation 3 AD 607554

Charcot-Marie-Tooth type 2B1 AR 605588

KRAS Noonan’s syndrome AD 163950

Cardio-facio-cutaneous syndrome 1 AD 115150

Locus Heterogeneity

PHENOTYPE GENE CHROMOSOMAL LOCATION PROTEIN

Familial hypertrophic cardiomyopathy MYH7 14q11.2 Myosin heavy chain beta

Genes encoding sarcomeric proteins TNNT2 1q32.1 Troponin-T2

TPM1 15q22.2 Tropomyosin alpha

MYBPC3 11p11q Myosin-binding

protein C

TNNC1 19q13.4 Troponin 1

MYL2 12q24.11 Myosin light chain 2

MYL3 3p21.31 Myosin light chain 3

TTN 2q31.2 Cardiac titin

ACTC 15q14 Cardiac alpha actin

MYH6 14q11.2 Myosin heavy chain alpha

MYLK2 20q11.21 Myosin light-peptide kinase

CAV3 3p25 Caveolin 3

Genes encoding nonsarcomeric proteins MT-T1 Mitochondrial tRNA isoleucine

MT-TG Mitochondrial tRNA glycine

PRKAG2 7q36.1 AMP-activated protein kinase γ2 subunit

DMPK 19q13.32 Myotonin protein kinase (myotonic

dystrophy)

FRDA 9q21.11 Frataxin (Friedreich’s ataxia)

Polycystic kidney disease PKD1 16p13.3 Polycystin 1 (AD)

PKD2 4q22.1 Polycystin 2 (AD)

PKHD1 6p21.1-p12.2 Fibrocystin/polyductin (AR)

Noonan’s syndrome PTPN11 12q24.13 Protein-tyrosine phosphatase 2c

KRAS 12p12.1 KRAS

Abbreviations: AD, autosomal dominant; AR, autosomal recessive; OMIM, Online Mendelian Inheritance in Man.

Principles of Human Genetics

3651CHAPTER 466

differences in phenotype can often provide clues that distinguish these

disorders from related genetic conditions.

VARIABLE EXPRESSIVITY AND INCOMPLETE PENETRANCE The same

genetic mutation may be associated with a phenotypic spectrum in

different affected individuals, thereby illustrating the phenomenon of

variable expressivity. This may include different manifestations of a

disorder variably involving different organs (e.g., multiple endocrine

neoplasia [MEN]), the severity of the disorder (e.g., cystic fibrosis), or

the age of disease onset (e.g., Alzheimer’s dementia). MEN 1 illustrates

several of these features. In this autosomal dominant tumor syndrome,

affected individuals carry an inactivating germline mutation that is

inherited in an autosomal dominant fashion. After somatic inactivation of the alternate allele (loss of heterozygosity; Knudson two-hit

model), patients can develop tumors of the parathyroid gland, endocrine pancreas, the pituitary gland and dermatologic lesions (Chap.

388). However, the pattern of tumors in the different glands, the age

at which tumors develop, and the types of hormones produced vary

among affected individuals, even within a given family. In this example,

the phenotypic variability arises, in part, because of the requirement for

a second somatic mutation in the normal copy of the MEN1 gene, as

well as the large array of different cell types that are susceptible to the

effects of MEN1 gene mutations. In part, variable expression reflects

the influence of modifier genes, or genetic background, on the effects

of a particular mutation. Even in identical twins, in whom the genetic

constitution is essentially the same, one can occasionally see variable

expression of a genetic disease.

Interactions with the environment can also influence the course of

a disease. For example, the manifestations and severity of hemochromatosis can be influenced by iron intake (Chap. 414), and the course

of phenylketonuria is affected by exposure to phenylalanine in the diet

(Chap. 420). Other metabolic disorders, such as hyperlipidemias and

porphyria, also fall into this category. Many mechanisms, including

genetic effects and environmental influences, can therefore lead to

variable expressivity. In genetic counseling, it is particularly important

to recognize this variability, because one cannot always predict the

course of disease, even when the mutation is known.

Penetrance refers to the proportion of individuals with a mutant

genotype that express the phenotype. If all carriers of a mutant

express the phenotype, penetrance is complete, whereas it is said to

be incomplete or reduced if some individuals do not exhibit features

of the phenotype. Dominant conditions with incomplete penetrance

are characterized by skipping of generations with unaffected carriers

transmitting the mutant gene. For example, hypertrophic obstructive

cardiomyopathy (HCM) caused by mutations in the myosin-binding

protein C gene is a dominant disorder with clinical features in only a

subset of patients who carry the mutation (Chap. 259). Patients who

have the mutation but no evidence of the disease can still transmit the

disorder to subsequent generations. In many conditions with postnatal

onset, the proportion of gene carriers who are affected varies with age.

Thus, when describing penetrance, one has to specify age. For example, for disorders such as Huntington’s disease or familial amyotrophic

lateral sclerosis, which present later in life, the rate of penetrance is

influenced by the age at which the clinical assessment is performed.

Imprinting can also modify the penetrance of a disease. For example,

in patients with AHO, mutations in the Gs

α subunit (GNAS1 gene) are

expressed clinically only in individuals who inherit the mutation from

their mother (Chap. 410).

SEX-INFLUENCED PHENOTYPES Certain mutations affect males and

females quite differently. In some instances, this is because the gene

resides on the X or Y sex chromosomes (X-linked disorders and

Y-linked disorders). As a result, the phenotype of mutated X-linked

genes will be expressed fully in males but variably in heterozygous

females, depending on the degree of X-inactivation and the function

of the gene. For example, most heterozygous female carriers of factor

VIII deficiency (hemophilia A) are asymptomatic because sufficient

factor VIII is produced to prevent a defect in coagulation (Chap. 116).

On the other hand, some females heterozygous for the X-linked lipid

storage defect caused by α-galactosidase A deficiency (Fabry’s disease)

experience mild manifestations of painful neuropathy, as well as other

features of the disease (Chap. 418). Because only males have a Y chromosome, mutations in genes such as SRY, which causes male-to-female

sex reversal, or DAZ (deleted in azoospermia), which causes abnormalities of spermatogenesis, are unique to males (Chap. 390).

Other diseases are expressed in a sex-limited manner because of the

differential function of the gene product in males and females. Activating mutations in the luteinizing hormone receptor cause dominant

male-limited precocious puberty in boys (Chap. 391). The phenotype is

unique to males because activation of the receptor induces testosterone

production in the testis, whereas it is functionally silent in the immature ovary. Biallelic inactivating mutations of the follicle-stimulating

hormone (FSH) receptor cause primary ovarian failure in females

because the follicles do not develop in the absence of FSH action. In

contrast, affected males have a more subtle phenotype, because testosterone production is preserved (allowing sexual maturation) and

spermatogenesis is only partially impaired (Chap. 391). In congenital

adrenal hyperplasia, most commonly caused by 21-hydroxylase deficiency, cortisol production is impaired and ACTH stimulation of the

adrenal gland leads to increased production of androgenic precursors

(Chap. 386). In females, the increased androgen level causes ambiguous genitalia, which can be recognized at the time of birth. In males,

the diagnosis may be made on the basis of adrenal insufficiency at

birth, because the increased adrenal androgen level does not alter sexual differentiation, or later in childhood, because of the development of

precocious puberty. Hemochromatosis is more common in males than

in females, presumably because of differences in dietary iron intake

and losses associated with menstruation and pregnancy in females

(Chap. 414).

Chromosomal Disorders Chromosomal disorders and the techniques used for their characterization have been discussed in detail

in a chapter in previous editions of this textbook. Chromosomal or

cytogenetic disorders are caused by numerical (aneuploidy) or structural aberrations (deletions, duplications, translocations, inversions,

dicentric and ring chromosomes, Robertsonian translocations) in

chromosomes. They occur in ~1% of the general population, in 8%

of stillbirths, and in close to 50% of spontaneously aborted fetuses.

Indications for cytogenetic and cytogenomic chromosome analyses

are summarized in Table 466-4. Contiguous gene syndromes (e.g., large

deletions affecting several genes) have been useful for identifying the

location of new disease-causing genes. Because of the variable size of

gene deletions in different patients, a systematic comparison of phenotypes and locations of deletion breakpoints allows positions of particular genes to be mapped within the critical genomic region.

TABLE 466-4 Indications for Cytogenetic and Cytogenomic Analysis

across the Life Span

TIMING OF TESTING INDICATIONS FOR TESTING

Prenatal Advanced maternal age

Abnormalities on ultrasound

Increased risk for genetic disorder on maternal serum

screen

Neonatal and childhood Multiple congenital anomalies

Intellectual disability

Autism

Developmental delay

Failure to thrive

Short stature

Disorders of sexual development

History of familial chromosomal alteration

Cancer

Adult Infertility

Recurrent miscarriage

Familial cancer

3652 PART 16 Genes, the Environment, and Disease

Monogenic Mendelian Disorders Monogenic human diseases

are frequently referred to as Mendelian disorders because they obey

the principles of genetic transmission originally set forth in Gregor

Mendel’s classic work. The continuously updated OMIM catalogue

lists several thousand of these disorders and provides information

about the clinical phenotype, molecular basis, allelic variants, and

pertinent animal models (Table 466-1). The mode of inheritance for

a given phenotypic trait or disease is determined by pedigree analysis.

All affected and unaffected individuals in the family are recorded in

a pedigree using standard symbols (Fig. 466-11). The principles of

allelic segregation, and the transmission of alleles from parents to children, are illustrated in Fig. 466-12. One dominant (A) allele and one

recessive (a) allele can display three Mendelian modes of inheritance:

autosomal dominant, autosomal recessive, and X-linked. About 65% of

human monogenic disorders are autosomal dominant, 25% are autosomal recessive, and 5% are X-linked. Genetic testing is now available

for many of these disorders and plays an important role in clinical

medicine (Chap. 467).

AUTOSOMAL DOMINANT DISORDERS In autosomal dominant disorders, mutations in a single allele are sufficient to cause the disease. In

contrast to recessive disorders, in which disease pathogenesis is relatively straightforward because there is a biallelic loss of gene function,

dominant disorders can be caused by various disease mechanisms,

many of which are unique to the function of the genetic pathway

involved. Mechanistically, the mutation may confer constitutive activation (gain of function), exert a dominant negative effect, or result in

loss of function and haploinsufficiency.

In autosomal dominant disorders, individuals are affected in successive generations; the disease does not occur in the offspring of

unaffected individuals. Males and females are affected with equal frequency because the defective gene resides on one of the 22 autosomes

(Fig. 466-13A). Autosomal dominant mutations alter one of the two

alleles at a given locus. Because the alleles segregate randomly at meiosis, the probability that an offspring will be affected is 50%. Unless

there is a new germline mutation, an affected individual has an affected

parent. Children with a normal genotype do not transmit the disorder.

Due to differences in penetrance or expressivity (see above), the clinical manifestations of autosomal dominant disorders may be variable.

Because of these variations, it is sometimes challenging to determine

the pattern of inheritance.

It should be recognized, however, that some individuals acquire

a mutated gene from an unaffected parent due to de novo germline

mutations. They occur more frequently during later cell divisions in

gametogenesis, which explains why siblings are rarely affected. As

noted before, new germline mutations occur more frequently in fathers

of advanced age. For example, the average age of fathers with new

germline mutations that cause Marfan’s syndrome is ~37 years, whereas

fathers who transmit the disease by inheritance have an average age of

~30 years.

Male

Mating

Monozygotic twins Dizygotic twins

Consanguineous

union

Female

Heterozygous

male

Heterozygous

female

Female

carrier of

X-linked trait

Affected

male

Affected Proband

female

6

Unknown

sex

Multiple

siblings

Spontaneous

abortion

Deceased

male

1 2

1 2 3

I

II

FIGURE 466-11 Standard pedigree symbols.

Aa

50:50

aa

Aa aa

AA

100

aa

Aa Aa

Aa

25:50:25

Aa

AA Aa Aa aa

FIGURE 466-12 Segregation of alleles. Segregation of genotypes in the offspring

of parents with one dominant (A) and one recessive (a) allele. The distribution of

the parental alleles to their offspring depends on the combination present in the

parents. Filled symbols = affected individuals.

A Autosomal dominant

D Mitochondrial

C X-linked

B

Autosomal recessive

Autosomal recessive

with pseudodominance

FIGURE 466-13 A. Dominant, B. recessive, C. X-linked, and D. mitochondrial

(matrilinear) inheritance.

Principles of Human Genetics

3653CHAPTER 466

AUTOSOMAL RECESSIVE DISORDERS In recessive disorders, the

mutated alleles result in a complete or partial loss of function.

They frequently involve enzymes in metabolic pathways, receptors,

or proteins in signaling cascades. In an autosomal recessive disease,

the affected individual, who can be of either sex, is a homozygote or

compound heterozygote for a single-gene defect. With a few important

exceptions, autosomal recessive diseases are rare and often occur in the

context of parental consanguinity. The relatively high frequency of certain recessive disorders such as sickle cell anemia, cystic fibrosis, and

thalassemia, is partially explained by a selective biologic advantage for

the heterozygous state (see below). Although heterozygous carriers of

a defective allele are usually clinically normal, they may display subtle

differences in phenotype that only become apparent with more precise

testing or in the context of certain environmental influences. In sickle

cell anemia, for example, heterozygotes are normally asymptomatic.

However, in situations of dehydration or diminished oxygen pressure,

sickle cell crises can also occur in heterozygotes (Chap. 98).

In most instances, an affected individual is the offspring of heterozygous parents. In this situation, there is a 25% chance that the

offspring will have a normal genotype, a 50% probability of a heterozygous state, and a 25% risk of homozygosity for the recessive alleles

(Figs. 466-10 and 466-13B). In the case of one unaffected heterozygous and one affected homozygous parent, the probability of disease

increases to 50% for each child. In this instance, the pedigree analysis

mimics an autosomal dominant mode of inheritance (pseudodominance). In contrast to autosomal dominant disorders, new mutations

in recessive alleles are rarely manifest because they usually result in an

asymptomatic carrier state.

X-LINKED DISORDERS Males have only one X chromosome; consequently, a daughter always inherits her father’s X chromosome in addition to one of her mother’s two X chromosomes. A son inherits the Y

chromosome from his father and one maternal X chromosome. Thus,

the characteristic features of X-linked inheritance are (1) the absence

of father-to-son transmission and (2) the fact that all daughters of an

affected male are obligate carriers of the mutant allele (Fig. 466-13C).

The risk of developing disease due to a mutant X-chromosomal gene

differs in the two sexes. Because males have only one X chromosome,

they are hemizygous for the mutant allele; thus, they are more likely

to develop the mutant phenotype, regardless of whether the mutation

is dominant or recessive. A female may be either heterozygous or

homozygous for the mutant allele, which may be dominant or recessive. The terms X-linked dominant and X-linked recessive are therefore

only applicable to expression of the mutant phenotype in women. In

addition, the expression of X-chromosomal genes is influenced by X

chromosome inactivation.

Y-LINKED DISORDERS The Y chromosome has a relatively small

number of genes. One such gene, the sex-region determining Y factor

(SRY), which encodes the testis-determining factor (TDF), is crucial

for normal male development. Normally, there is infrequent exchange

of sequences on the Y chromosome with the X chromosome. The SRY

region is adjacent to the pseudoautosomal region, a chromosomal segment on the X and Y chromosomes with a high degree of homology. A

crossing-over event occasionally involves the SRY region with the distal

tip of the X chromosome during meiosis in the male. Translocations

can result in XY females with the Y chromosome lacking the SRY gene

or XX males harboring the SRY gene on one of the X chromosomes

(Chap. 390). Point mutations in the SRY gene may also result in individuals with an XY genotype and an incomplete female phenotype.

Most of these mutations occur de novo. Men with oligospermia/

azoospermia frequently have microdeletions on the long arm of the

Y chromosome that involve one or more of the azoospermia factor

(AZF) genes.

Exceptions to Simple Mendelian Inheritance Patterns • 

MITOCHONDRIAL DISORDERS Mendelian inheritance refers to the

transmission of genes encoded by DNA contained in the nuclear chromosomes. In addition, each mitochondrion contains several copies of

a small circular chromosome (Chap. 468). The mitochondrial DNA

(mtDNA) is ~16.5 kb and encodes transfer and ribosomal RNAs and

13 core proteins that are components of the respiratory chain involved

in oxidative phosphorylation and ATP generation. The mitochondrial

genome does not recombine and is inherited through the maternal line

because sperm does not contribute significant cytoplasmic components

to the zygote. A noncoding region of the mitochondrial chromosome,

referred to as D-loop, is highly polymorphic. This property, together

with the absence of mtDNA recombination, makes it a valuable tool for

studies tracing human migration and evolution, and it is also used for

specific forensic applications.

Inherited mitochondrial disorders are transmitted in a matrilineal

fashion; all children from an affected mother will inherit the disease,

but it will not be transmitted from an affected father to his children

(Fig. 466-13D). Alterations in the mtDNA that involves enzymes

required for oxidative phosphorylation lead to reduction of ATP supply, generation of free radicals, and induction of apoptosis. Several

syndromic disorders arising from mutations in the mitochondrial

genome are known in humans, and they affect both protein-coding

and tRNA genes. The broad clinical spectrum often involves (cardio)

myopathies and encephalopathies because of the high dependence of

these tissues on oxidative phosphorylation. The age of onset and the

clinical course are highly variable because of the unusual mechanisms

of mtDNA transmission, which replicates independently from nuclear

DNA. During cell replication, the proportion of wild-type and mutant

mitochondria can drift among different cells and tissues. The resulting

heterogeneity in the proportion of mitochondria with and without a

mutation is referred to as heteroplasmia and underlies the phenotypic

variability that is characteristic of mitochondrial diseases.

Acquired somatic mutations in mitochondria are thought to be

involved in several age-dependent degenerative disorders affecting

predominantly muscle and the peripheral and central nervous system (e.g., Alzheimer’s and Parkinson’s diseases). Establishing that an

mtDNA alteration is causal for a clinical phenotype is challenging

because of the high degree of polymorphism in mtDNA and the phenotypic variability characteristic of these disorders. Certain pharmacologic treatments may have an impact on mitochondria and/or their

function. For example, treatment with the antiretroviral compound

azidothymidine (AZT) causes an acquired mitochondrial myopathy

through depletion of muscular mtDNA.

MOSAICISM Mosaicism refers to the presence of two or more genetically distinct cell lines in the tissues of an individual. It results from

a mutation that occurs during embryonic, fetal, or extrauterine

development. The developmental stage at which the mutation arises

will determine whether germ cells and/or somatic cells are involved.

Chromosomal mosaicism results from nondisjunction at an early

embryonic mitotic division, leading to the persistence of more than

one cell line, as exemplified by some patients with Turner’s syndrome

(Chap. 390). Somatic mosaicism is characterized by a patchy distribution of genetically altered somatic cells. The McCune-Albright

syndrome, for example, is caused by activating mutations in the stimulatory G protein α (Gs

α) that occur early in development (Chap. 410).

The clinical phenotype varies depending on the tissue distribution

of the mutation; manifestations include ovarian cysts that secrete sex

steroids and cause precocious puberty, polyostotic fibrous dysplasia,

café-au-lait skin pigmentation, GH-secreting pituitary adenomas, and

hypersecreting autonomous thyroid nodules.

X-INACTIVATION, IMPRINTING, AND UNIPARENTAL DISOMY According to traditional Mendelian principles, the parental origin of a mutant

gene is irrelevant for the expression of the phenotype. There are,

however, important exceptions to this rule. X-inactivation prevents

the expression of most genes on one of the two X chromosomes in

every cell of a female. Gene inactivation through genomic imprinting

occurs on selected chromosomal regions of autosomes and leads to

inheritable preferential expression of one of the parental alleles. It is of

pathophysiologic importance in disorders where the transmission of

disease is dependent on the sex of the transmitting parent and, thus,

plays an important role in the expression of certain genetic disorders.

Two classic examples are the Prader-Willi syndrome and Angelman’s

3654 PART 16 Genes, the Environment, and Disease

syndrome. Prader-Willi syndrome is characterized by diminished fetal

activity, obesity, hypotonia, intellectual disability, short stature, and

hypogonadotropic hypogonadism. Deletions of the paternal copy of the

Prader-Willi locus located on the short arm of chromosome 15 result

in a contiguous gene syndrome involving missing paternal copies of

the necdin and SNRPN genes, among others. In contrast, patients with

Angelman’s syndrome, characterized by intellectual disability, seizures,

ataxia, and hypotonia, have deletions involving the maternal copy of

this region on chromosome 15. These two syndromes may also result

from uniparental disomy. In this case, the syndromes are not caused

by deletions on chromosome 15 but by the inheritance of either two

maternal chromosomes (Prader-Willi syndrome) or two paternal chromosomes (Angelman’s syndrome). Lastly, the two distinct phenotypes

can also be caused by an imprinting defect that impairs the resetting

of the imprint during zygote development (defect in the father leads

to Prader-Willi syndrome; defect in the mother leads to Angelman’s

syndrome).

Imprinting and the related phenomenon of allelic exclusion may be

more common than currently documented, because it is difficult to

examine levels of mRNA expression from the maternal and paternal

alleles in specific tissues or in individual cells. Genomic imprinting,

or uniparental disomy, is involved in the pathogenesis of several other

disorders and malignancies. For example, hydatidiform moles contain

a normal number of diploid chromosomes, but they are all of paternal

origin. The opposite situation occurs in ovarian teratomata, with 46

chromosomes of maternal origin. Expression of the imprinted gene

for insulin-like growth factor 2 (IGF-2) is involved in the pathogenesis

of the cancer-predisposing Beckwith-Wiedemann syndrome (BWS).

These children show somatic overgrowth with organomegalies and

hemihypertrophy, and they have an increased risk of embryonal malignancies such as Wilms’ tumor. Normally, only the paternally derived

copy of the IGF2 gene is active, and the maternal copy is inactive. BWS

can be caused by several genetic defects that result in overactivity of

IGF-2, or a missing active copy of CDKN1C, that result in inhibition

of cell proliferation. They include paternal uniparental disomy (UPD)

of chromosome 11, aberrant methylation of this region, maternal chromosomal rearrangements, or deletions within the locus.

Alterations of the epigenome through gain and loss of DNA methylation and altered histone modifications play an important role in the

pathogenesis of malignancies.

SOMATIC MUTATIONS Cancer can be considered a genetic disease at

the cellular level (Chap. 71). Cancers are monoclonal in origin, indicating that they have arisen from a single precursor cell with one or several mutations in genes controlling growth (proliferation or apoptosis)

and/or differentiation. These acquired somatic mutations are restricted

to the tumor and its metastases and are not found in the surrounding

normal tissue. The molecular alterations include dominant gain-offunction mutations in oncogenes, recessive loss-of-function mutations

in tumor-suppressor genes and DNA repair genes, gene amplification,

and chromosome rearrangements. Chromothripsis refers to a mutational process including multiple clustered chromosomal rearrangements in close vicinity, for example, after injury by ionizing radiation.

Rarely, a single mutation in certain genes may be sufficient to transform a normal cell into a malignant cell. In most cancers, however,

the development of a malignant phenotype requires several genetic

alterations for the gradual progression from a normal cell to a cancerous cell, a phenomenon termed multistep carcinogenesis. Genome-wide

analyses of cancers using deep sequencing often reveal somatic rearrangements resulting in fusion genes and mutations in multiple genes

(Table 466-1 and Fig. 466-14). Comprehensive sequence analyses, now

also possible through single-cell sequencing (SCS), provide insight into

the evolution and genetic heterogeneity within malignancies; these

include intratumoral heterogeneity among the cells of the primary

tumor, intermetastatic and intrametastatic heterogeneity, and interpatient differences. These analyses further support the notion of cancer

as an ongoing process of clonal evolution, in which successive rounds

of clonal selection within the primary tumor and metastatic lesions

result in diverse genetic and epigenetic alterations that require targeted

(personalized) therapies (precision medicine). The heterogeneity of

mutations within a tumor can also lead to resistance to targeted therapies because cells with mutations that are resistant to the therapy, even

if they are a minor part of the tumor population, will be selected as the

more sensitive cells are eliminated.

Telomeres, repeats of conserved sequences, protect the ends the

chromosomes from DNA damage or fusion with neighboring chromosomes. Telomere length shortens with age. Most human tumors express

telomerase, an enzyme formed of a protein and an RNA component,

which adds telomere repeats at the ends of chromosomes during

replication. This mechanism impedes shortening of the telomeres

and is associated with enhanced replicative capacity in cancer cells.

0

Histology

HPV clade HPV integration

APOBEC mutagenesis

UCEC-like

EMT score Purity

iCluster

PIK3CA (26%) EP300 (11%) FBXW7 (11%) PTEN (8%) HLA-A (8%) ARID1A (7%) NFE2L2 (7%) HLA-B (6%)

KRAS (6%) ERBB3 (6%) MAPK1 (5%)

CD274 (8%) PTEN (8%) YAP1 (16%)

BCAR4 (16%)

3q (66%)

Synonymous

log2[CN] ≤ –0.4 –0.4 < log2[CN] ≤ –0.1 log2[CN] ≥ 0.4

Gene-level SCNAs

0 40 80 120

Mutations

APOBEC

Non-APOBEC

Keratin-low Keratin-high Adeno.

1.15

No No

No

Other

Adeno. Adenosq. Squamous

Synonymous

Non-synonymous

A9A7 Negative

Yes

Yes High Low

0.96 –3.76

0.22

0 20 6040 80

Gain

Loss

0.1 ≤ log2[CN] < 0.4

In-frame indel Other non-synonymous Missense Splice site Frameshift Nonsense

CASP8 (4%)

TGFBR2 (3%)

SHKBP1 (2%) Mutations per Mb

2

4

A

B

C

D

6

8

10

FIGURE 466-14 Somatic alterations in cervical cancer. A. Cervical carcinoma samples ordered by histology and mutation frequency; B. clinical and molecular platform

features; C. significantly mutated genes (SMGs); and D. select somatic copy number alterations. SMGs are ordered by the overall mutation frequency and color-coded

by mutation type. Adeno, adenocarcinomas; Adenosq, adenosquamous cancers; CN, copy number; SCNAs, somatic copy number alterations; Squamous, squamous

cell carcinomas. (From The Cancer Genome Atlas Research Network. Integrated genomic and molecular characterization of cervical cancer. Nature 543:378–384, 2017.

Permission for reproduction granted through a Creative Commons CC-BY [CC BY 4.0] license.)

Principles of Human Genetics

3655CHAPTER 466

Telomerase inhibitors provide a strategy for treating advanced human

cancers.

In many cancer syndromes, there is frequently an inherited predisposition to tumor formation. In these instances, a germline mutation is

inherited in an autosomal dominant fashion inactivating one allele of

an autosomal tumor-suppressor gene. If the second allele is inactivated

by a somatic mutation or by epigenetic silencing in a given cell, this will

lead to neoplastic growth (Knudson two-hit model). Thus, the defective allele in the germline is transmitted in a dominant mode, although

tumorigenesis results from a biallelic loss of the tumor-suppressor gene

in an affected tissue. The classic example to illustrate this phenomenon

is retinoblastoma, which can occur as a sporadic or hereditary tumor.

In sporadic retinoblastoma, both copies of the retinoblastoma (RB)

gene are inactivated through two somatic events. In hereditary retinoblastoma, one mutated or deleted RB allele is inherited in an autosomal

dominant manner and the second allele is inactivated by a subsequent

somatic mutation. This two-hit model applies to other inherited cancer

syndromes such as MEN 1 (Chap. 388) and neurofibromatosis types

1 and 2 (Chap. 90). In contrast, in the autosomal dominant MEN 2

syndrome, the predisposition for tumor formation in various organs

is caused by a gain-of-function mutation in a single allele of the RET

gene (Chap. 388).

NUCLEOTIDE REPEAT EXPANSION DISORDERS Several diseases are

associated with an increase in the number of nucleotide repeats above

a certain threshold (Table 466-5). The repeats are sometimes located

within the coding region of the genes, as in Huntington’s disease or

the X-linked form of spinal and bulbar muscular atrophy (SBMA;

Kennedy’s syndrome). In other instances, the repeats probably alter

gene regulatory sequences. If an expansion is present, the DNA fragment is unstable and tends to expand further during cell division. The

length of the nucleotide repeat often correlates with the severity of the

disease. When repeat length increases from one generation to the next,

disease manifestations may worsen or be observed at an earlier age;

this phenomenon is referred to as anticipation. In Huntington’s disease,

for example, there is a correlation between age of onset and length of

the triplet codon expansion (Chap. 424). Anticipation has also been

documented in other diseases caused by dynamic mutations in trinucleotide repeats (Table 466-5). The repeat number may also vary in a

tissue-specific manner. In myotonic dystrophy, the CTG repeat may

be tenfold greater in muscle tissue than in lymphocytes (Chap. 449).

Complex Genetic Disorders The expression of many common

diseases such as cardiovascular disease, hypertension, diabetes, asthma,

psychiatric disorders, and certain cancers is determined by a combination of genetic background, environmental factors, and lifestyle. A trait

is called polygenic if multiple genes contribute to the phenotype or multifactorial if multiple genes are assumed to interact with environmental

factors. Genetic models for these complex traits need to account for

genetic heterogeneity and interactions with other genes and the environment. Complex genetic traits may be influenced by modifier genes

that are not linked to the main gene involved in the pathogenesis of the

trait. This type of gene-gene interaction, or epistasis, plays an important

role in polygenic traits that require the simultaneous presence of variations in multiple genes to result in a pathologic phenotype.

Type 2 diabetes mellitus provides a paradigm for considering a multifactorial disorder, because genetic, nutritional, and lifestyle factors are

intimately interrelated in disease pathogenesis (Table 466-6) (Chap.

403). The identification of genetic variations and environmental factors that either predispose to or protect against disease is essential for

predicting disease risk, designing preventive strategies, and developing

novel therapeutic approaches. The study of rare monogenic diseases

may provide insight into some of the genetic and molecular mechanisms important in the pathogenesis of complex diseases. For example,

the identification of the genes causing monogenic forms of permanent

neonatal diabetes mellitus or maturity-onset diabetes defined them

as candidate genes in the pathogenesis of diabetes mellitus type 2

(Tables 466-2 and 466-6) (Fig. 466-15). Genome scans have identified

numerous genes and loci that may be associated with susceptibility to

development of diabetes mellitus in certain populations (Fig. 466-16).

Efforts to identify susceptibility genes require very large sample sizes,

and positive results may depend on ethnicity, ascertainment criteria,

and statistical analysis. Association studies analyzing the potential

influence of (biologically functional) SNPs and SNP haplotypes on

a particular phenotype have revealed new insights into the genes

involved in the pathogenesis of these common disorders. Large variants ([micro]deletions, duplications, and inversions) present in the

human population also contribute to the pathogenesis of complex

disorders, but their contributions remain poorly understood.

Linkage and Association Studies There are two primary strategies for mapping genes that cause or increase susceptibility to human

disease: (1) classic linkage can be performed based on a known genetic

model or, when the model is unknown, by studying pairs of affected

relatives; or (2) disease genes can be mapped using allelic association

studies (Table 466-7).

GENETIC LINKAGE Genetic linkage refers to the fact that genes are

physically connected, or linked, to one another along the chromosomes. Two fundamental principles are essential for understanding the

concept of linkage: (1) when two genes are close together on a chromosome, they are usually transmitted together, unless a recombination

TABLE 466-5 Selected Trinucleotide Repeat Disorders

DISEASE LOCUS REPEAT

TRIPLET LENGTH

(NORMAL/DISEASE) INHERITANCE GENE PRODUCT

X-chromosomal spinobulbar muscular

atrophy (SBMA)

Xq12 CAG 11–34/40–62 XR Androgen receptor

Fragile X syndrome (FRAXA) Xq27.3 CGG 6–50/200–300 XR FMR-1 protein

Fragile X syndrome (FRAXE) Xq28 GCC 6–25/>200 XR FMR-2 protein

Dystrophia myotonica (DM) 19q13.32 CTG 5–30/200–1000 AD, variable penetrance Myotonin protein kinase

Huntington’s disease (HD) 4p16.3 CAG 6–34/37–180 AD Huntingtin

Spinocerebellar ataxia type 1 (SCA1) 6p22.3 CAG 6–39/40–88 AD Ataxin 1

Spinocerebellar ataxia type 2 (SCA2) 12q24.12 CAG 15–31/34–400 AD Ataxin 2

Spinocerebellar ataxia type 3 (SCA3);

Machado-Joseph disease (MD)

14q32.12 CAG 13–36/55–86 AD Ataxin 3

Spinocerebellar ataxia type 6 (SCA6, CACNAIA) 19p13 CAG 4–16/20–33 AD Alpha 1A voltage-dependent

L-type calcium channel

Spinocerebellar ataxia type 7 (SCA7) 3p14.1 CAG 4–19/37 to >300 AD Ataxin 7

Spinocerebellar ataxia type 12 (SCA12) 5q32 CAG 6–26/66–78 AD Protein phosphatase 2A

Dentatorubral pallidoluysian atrophy (DRPLA) 12p13.31 CAG 7–23/49–75 AD Atrophin 1

Friedreich’s ataxia (FRDA1) 9q21.11 GAA 7–22/200–900 AR Frataxin

Abbreviations: AD, autosomal dominant; AR, autosomal recessive; XR, X-linked recessive.

3656 PART 16 Genes, the Environment, and Disease

event separates them (Figs. 466-6); and (2) the odds of a crossover, or

recombination event, between two linked genes is proportional to the

distance that separates them. Thus, genes that are farther apart are

more likely to undergo a recombination event than genes that are very

close together. The detection of chromosomal loci that segregate with

a disease by linkage can be used to identify the gene responsible for

the disease (positional cloning) and to predict the odds of disease gene

transmission in genetic counseling.

Polymorphic variants are essential for linkage studies because they

provide a means to distinguish the maternal and paternal chromosomes in an individual. On average, 1 out of every 1000 bp varies from

one person to the next. Although this degree of variation seems low

(99.9% identical), it means that >3 million sequence differences exist

between any two unrelated individuals and the probability that the

sequence at such loci will differ on the two homologous chromosomes

is high (often >70–90%). These sequence variations include variable

number of tandem repeats (VNTRs), short tandem repeats (STRs),

and SNPs. Most STRs, also called polymorphic microsatellite markers,

consist of di-, tri-, or tetranucleotide repeats that can be characterized

readily using the polymerase chain reaction (PCR). Characterization

of SNPs, using DNA chips or beads, permits comprehensive analyses

of genetic variation, linkage, and association studies. Although these

sequence variations often have no apparent functional consequences,

they provide much of the basis for variation in genetic traits.

In order to identify a chromosomal locus that segregates with a

disease, it is necessary to characterize polymorphic DNA markers from

affected and unaffected individuals of one or several pedigrees. One

can then assess whether certain marker alleles cosegregate with the

disease. Markers that are closest to the disease gene are less likely to

undergo recombination events and therefore receive a higher linkage

score. Linkage is expressed as an lod (logarithm of odds) score—the

ratio of the probability that the disease and marker loci are linked

rather than unlinked. Lod scores of +3 (1000:1) are generally accepted

as supporting linkage, whereas a score of –2 is consistent with the

absence of linkage.

ALLELIC ASSOCIATION, LINKAGE DISEQUILIBRIUM, AND HAPLOTYPES Allelic association refers to a situation in which the frequency

of an allele is significantly increased or decreased in individuals

affected by a particular disease in comparison to controls. Linkage and

TABLE 466-6 Genes and Loci Involved in Mono- and Polygenic Forms of Diabetes

DISORDER GENES OR SUSCEPTIBILITY LOCUS CHROMOSOMAL LOCATION OTHER FACTORS

Monogenic permanent neonatal

diabetes mellitus

KCNJ11 (inwardly rectifying potassium channelKir6.2) 11p15.1 AD

GCK (glucokinase) 7p13 AR

INS (insulin) 11p15.5 AR, hyperproinsulinemia

ABCC8 (ATP-binding cassette, subfamily c, member 8;

sulfonylurea receptor)

11p15.1 AD or AR

GLIS3 (GLIS family zinc finger protein 3) 9p24.2 AR, diabetes, congenital

hypothyroidism

Maturity-onset diabetes of the young

(MODY): monogenic forms of diabetes

mellitus

MODY 1 HNF4α (hepatocyte nuclear factor 4α) 20q13.12 AD inheritance

MODY 2 GCK (glucokinase) 7p13

MODY 3 HNF1α (hepatocyte nuclear factor 1α) 12q24.31

MODY 4 IPF1 (insulin receptor substrate) 13q12.2

MODY 5 (renal cysts, diabetes) HNF1β (hepatocyte nuclear factor 1β) 17q12

MODY 6 NeuroD1 (neurogenic differentiation factor 1) 2q31.3

MODY 7 KLF1 (Kruppel-like factor 1) 19p13.13

MODY 8 CEL (carboxyl ester lipase) 9q34.13

MODY 9 PAX4 (paired box transcription factor 4) 7q32.1

MODY 10 INS (insulin) 11p15.5

MODY 11 BLK (B-lymphocyte-specific tyrosine kinase) 8p23.1

Diabetes mellitus type 2; loci and

genes linked and/or associated with

susceptibility for diabetes mellitus

type 2

Genes and loci identified by linkage/association studies Heavily influenced by diet,

energy expenditure, obesity

PPARG, KCNJ11/ABCC8, TCF7L2, HNF1B, WFS1, SLC30A8,

FTO, HHEX, IGF2BP2, CDKN2A/B, CDKAL1, TSPAN8, ADAMTs9,

CDC123/CAMK1D, JAZF1, NOTCH2, THADA, KCNQ1, DUSP8,

MTNR1B, IRS1, SPRY2, SRR, ZFAND6, GCK, KLF14, TP53INP1,

PROX1, PRC1, BCL11A, ZBED3, RBMS1, HNF1A, DGKB/TMEM195,

CCND2, C2CD4A/C2CD4B, PTPRD, ARAP1/CENTD2, HMGA2,

TLE4/CHCHD9, ADCY5, UBE2E2, DUSP9, GCKR, COBLL1/GRB14,

HMG20A, VPS26A, ST6GAL1, AP3S2, HNF4A, BCL2, LAMA1, GIPR,

MC4R, TLE1, KCNK16, ANK1, KLHDC5, ZMIZ1, PSMD6 , FITM2/

R3HDML/HNF4A, CILP2, ANKRD55, GLIS3, PEPD, GCC1/PAX4,

ZFAND3, MAEA, BCAR1, RBM43/RND3 , MACF1, RASGRP1, GRK5,

TMEM163, SGCG, LPP, FAF1, TMEM154, MPHOSPH9, ARL15,

POU5F1/TCF19, SSR1/RREB1, HLA-B, INS-IGF2, GPSM1, LEP,

SLC16A13, PAM/PPIP5K2, SLC16A11, CCDC63, C12orf51, CCND2,

HNF1A, TBC1D4, CCDC85A, INAFM2, ASB3, FAM60A, ATP8B2,

MIR4686, MTMR3, DMRTA1, SLC35D3, GLP2R, GIP, MAP3K11,

PLEKHA1, HSD17B12, NRXN3, CMIP, ZZEF1, MNX1, ABO, ACSL1,

HLA-DQA1

Abbreviations: AD, autosomal dominant; AR, autosomal recessive; MODY, maturity-onset diabetes of the young.

Principles of Human Genetics

3657CHAPTER 466

Rare alleles

Mendelian disease

Low-frequency

variants with

intermediate effect

Typical:

Common variants

with low effect on

complex disease

Rare:

Common variants

with high effect on

complex disease

Rare variants

with small effect:

difficult to identify

Low frequency

Allele frequency

Very rare Common

Effect size

50

3.0

1.5

1.1

0.001 0.005 0.05

High

Intermediate

Modest

Low

Rare

FIGURE 466-15 Relationship between allele frequency and effect size in monogenic and polygenic disorders. In classic Mendelian disorders, the allele frequency is

typically low but has a high impact (single-gene disorder). This contrasts with polygenic disorders that require the combination of multiple low-impact alleles that are

frequently quite common in the general population.

association differ in several aspects. Genetic linkage is demonstrable in

families or sibships. Association studies, on the other hand, compare

a population of affected individuals with a control population. Association studies can be performed as case-control studies that include

unrelated affected individuals and matched controls or as family-based

studies that compare the frequencies of alleles transmitted or not transmitted to affected children.

Allelic association studies are particularly useful for identifying

susceptibility genes in complex diseases. When alleles at two loci occur

more frequently in combination than would be predicted (based on

known allele frequencies and recombination fractions), they are said

to be in linkage disequilibrium. Evidence for linkage disequilibrium can

be helpful in mapping disease genes because it suggests that the two

loci are tightly linked.

Detecting the genetic factors contributing to the pathogenesis of

common complex disorders is challenging. In many instances, these

are low-penetrance alleles (e.g., variations that individually have a

subtle effect on disease development, and they can only be identified

by unbiased GWAS) (Catalog of Published Genome-Wide Association Studies; Table 466-1) (Fig. 466-16). Most variants occur in

noncoding or regulatory sequences but do not alter protein structure.

The analysis of complex disorders is further complicated by ethnic

differences in disease prevalence, differences in allele frequencies in

known susceptibility genes among different populations, locus and

allelic heterogeneity, gene-gene and gene-environment interactions,

and the possibility of phenocopies. Catalogues of human variation and

genotype data (HapMap, International Genome Sample Resource) have

greatly facilitated GWAS for the characterization of complex disorders.

Adjacent SNPs are inherited together as blocks, and these blocks can

be identified by genotyping selected marker SNPs, so-called Tag SNPs,

thereby reducing cost and workload (Fig. 466-4). The availability of

this information permits the characterization of a limited number of

SNPs to identify the set of haplotypes present in an individual (e.g.,

in cases and controls). This, in turn, permits performing GWAS by

searching for associations of certain haplotypes with a disease phenotype of interest, an essential step for unraveling the genetic factors

contributing to complex disorders.

POPULATION GENETICS In population genetics, the focus changes

from alterations in an individual’s genome to the distribution pattern of

different genotypes in the population. In a case where there are only two

alleles, A and a, the frequency of the genotypes will be p2

 + 2pq + q2

 = 1,

with p2

 corresponding to the frequency of AA, 2pq to the frequency of

Aa, and q2

 to aa. When the frequency of an allele is known, the frequency

of the genotype can be calculated. Alternatively, one can determine an

allele frequency if the genotype frequency has been determined.

Allele frequencies vary among ethnic groups and geographic

regions. For example, heterozygous mutations in the CFTR gene are

relatively common in populations of European origin but are rare in

the African population. Allele frequencies may vary because certain

allelic variants confer a selective advantage. For example, heterozygotes

for the sickle cell mutation, which is particularly common in West

Africa, are more resistant to malaria infection because the erythrocytes

of heterozygotes provide a less favorable environment for Plasmodium

parasites. Although homozygosity for the sickle cell mutation is associated with severe anemia and sickle crises, heterozygotes have a higher

probability of survival because of the reduced morbidity and mortality

from malaria; this phenomenon has led to an increased frequency of

the mutant allele. Recessive conditions are more prevalent in geographically isolated populations because of the more restricted gene pool.

APPROACH TO THE PATIENT

Inherited Disorders

For the practicing clinician, the family history remains an essential

step in recognizing the possibility of a hereditary predisposition

to disease. When taking the history, it is useful to draw a detailed

pedigree of the first-degree relatives (e.g., parents, siblings, and

children), because they share 50% of genes with the patient. Standard symbols for pedigrees are depicted in Fig. 466-11. The family

history should include information about ethnic background, age,

health status, and deaths, including infants. Next, the physician

should explore whether there is a family history of the same or

related illnesses to the current problem. An inquiry focused on

commonly occurring disorders such as cancers, heart disease, and

diabetes mellitus should follow. Because of the possibility of agedependent expressivity and penetrance, the family history will need

intermittent updating. If the findings suggest a genetic disorder,

the clinician should assess whether some of the patient’s relatives

may be at risk of carrying or transmitting the disease. In this circumstance, it is useful to confirm and extend the pedigree based on

input from several family members. This information may form the

3658 PART 16 Genes, the Environment, and Disease

Candidate gene

Linkage

GWAS

Exome sequencing

Genome sequencing

Targeted sequencing

2000

PPARG

1.00

1.10

1.20

1.30

1.40

1.50

1.60

1.70

1.80

1.90

2.00

2003

KCNJ11 2006

TCF7L2

4,96

10,30

Approximate allelic effect size

2017

SLC35D3

GLP2R

GIP

MAP3K11

PLEKHA1

HSD17B12

NRXN3

CMIP

ZZEF1

MNX1

ABO

ACSL1

HLA-DQA1

2017

SLC35D3

GLP2R

GIP

MAP3K11

PLEKHA1

HSD17B12

NRXN3

CMIP

ZZEF1

MNX1

ABO

ACSL1

HLA-DQA1

2016

CCDC85A

INAFM2

ASB3

FAM60A

ATP8B2

MIR4686

MTMR3

DMRTA1

2014

LLPP

FAF1

TMEM154

MPHOSPH9

ARL15

POU5F1/TCF19

SSR1/RREB1

HLA-B

INS-IGF2

GPSM1

LEP

SLC16A13

PAM/PPIP5K2

SLC16A11

CCDC63

C12orf51

CCND2

HNF1A

TBC1D4

2013

MACF1

RASGRP1

GRK5

TMEM163

SGCG

2012

BCL2

LAMA1

GIPR

MC4R

TLE1

KCNK16

ANK1

KLHDC5

ZMIZ1

PSMD6

FITM2/R3HDML/HNF4A

CILP2

ANKRD55

GLIS3

PEPD

GCC1/PAX4

ZFAND3

MAEA

BCAR1

RBM43/RND3

2011

COBLL1/GRB14

HMG20A

VPS26A

ST6GAL1

AP3S2

HNF4A

2010

SPRY2

SRR

ZFAND6

GCK

KLF14

TP53INP1

PROX1

PRC1

BCL11A

ZBED3

RBMS1

HNF1A

DGKB/TMEM195

CCND2

C2CD4A/C2CD4B

PTPRD

ARAP1/CENTD2)

HMGA2

TLE4/CHCHD9)

ADCY5

UBE2E2

DUSP9

GCKR

2009

DUSP8

MTNR1B

IRS1

2008

TSPAN8

ADAMTs9

CDC123

JAZF1

NOTCH2

THADA

KCNQ1

2007

HNF1B

WFS1

SLC30A8

FTO

HHEX

IGF2BP2

CDKN2A/B

CDKAL1

FIGURE 466-16 Loci and genes associated with diabetes mellitus type 2. Loci are listed by year of identification, and the color indicates discovery method. Gene names

indicate the locus and do not necessarily imply that the gene itself is causally involved. Approximate allelic effect sizes were either derived from the discovery cohort or, if

available, from the DIAGRAM (Diabetes Genetics Replication and Meta-analysis consortium) European ancestry meta-analysis and the Asian ancestry meta-analysis. Gene

names that are underlined denote identification in population isolates. (The data have been graciously provided by Dr. Miriam Udler and Dr. Jose Florez, Harvard Medical

School, Boston.)

basis for genetic counseling, carrier detection, early intervention,

and disease prevention in relatives of the index patient (Chap. 467).

In instances where a diagnosis at the molecular level may be relevant, it is important to identify an appropriate laboratory that can

perform the appropriate test. Genetic testing is available for a large

number of monogenic disorders through commercial laboratories.

For uncommon disorders, the test may only be performed in a specialized research laboratory. Approved laboratories offering testing for

inherited disorders can be identified in continuously updated online

resources (e.g., Genetic Testing Registry; Table 466-1). If genetic testing is considered, the patient and the family should be counseled

about the potential implications of positive results, including psychological distress and the possibility of discrimination. The patient

or caretakers should be informed about the meaning of a negative

result, technical limitations, and the possibility of false-negative and

inconclusive results. For these reasons, genetic testing should only

be performed after obtaining informed consent. Published ethical

guidelines address the specific aspects that should be considered

when testing children and adolescents. Genetic testing should usually be limited to situations in which the results may have an impact

on medical management.

IDENTIFYING THE DISEASE-CAUSING GENE

Precision medicine aims to enhance the quality of medical care

through the use of genotypic analysis (DNA testing) to identify

genetic predisposition to disease, to select more specific pharmacotherapy, and to design individualized medical care based

on genotype. Genotype can be deduced by analysis of protein

Principles of Human Genetics

3659CHAPTER 466

(e.g., hemoglobin, apoprotein E), mRNA, or DNA. Many (pathogenic) variants can be readily identified by DNA analyses; technical

advances in RNA sequencing now add increasing depth to genetic

and genomic investigations (e.g., for the detection of gene fusions

or aberrant gene expression patterns).

DNA testing is performed by mutational analysis or linkage

studies in individuals at risk for a genetic disorder known to be

present in a family. Mass screening programs require tests of high

sensitivity and specificity to be cost-effective. The benefits and risks

of screening newborns with genomic sequencing remain uncertain,

and the potential impact on surveillance, preventative health care,

and personalized treatment options is currently being explored

(BabySeq Project). Prerequisites for the success of genetic screening

programs include the following: that the disorder is potentially

serious; that it can be influenced at a presymptomatic stage by

changes in behavior, diet, and/or pharmaceutical manipulations;

and that the screening does not result in any harm or discrimination. Screening in Jewish populations for the autosomal recessive

neurodegenerative storage disease Tay-Sachs has reduced the number of affected individuals. In contrast, screening for sickle cell trait/

disease in African Americans has led to unanticipated problems

of discrimination by health insurers and employers. Mass screening programs harbor additional potential problems. For example,

screening for the most common genetic alteration in cystic fibrosis,

the ΔF508 mutation with a frequency of ~70% in northern Europe,

is feasible and seems to be effective. One has to keep in mind,

however, that there is pronounced allelic heterogeneity and that the

CFTR gene can be affected by ~2000 other mutations. The search

for these less common mutations would substantially increase

costs but not the effectiveness of the screening program as a whole.

Next-generation genome sequencing permits comprehensive and

cost-effective mutational analyses after selective enrichment of

candidate genes. For example, tests that sequence all the common

genes causing hereditary deafness or hereditary pheochromocytomas are commercially available. Occupational screening programs

aim to detect individuals with increased risk for certain professional

activities (e.g., α1

 antitrypsin deficiency and smoke or dust exposure). Integrating genomic data into electronic medical records is

evolving and can provide significant decision support at the point of

care, for example, by providing the clinician with genomic data and

decision algorithms for the prescription of drugs that are subject to

pharmacogenetic influences.

Mutational Analyses DNA sequence analysis is widely used as a

diagnostic tool and has significantly enhanced diagnostic accuracy.

It is used for determining carrier status and for prenatal testing in

monogenic disorders. Numerous techniques, discussed in previous

versions of this chapter, are available for the detection of mutations.

Analyses of large alterations in the genome are possible using classic

methods such as karyotype analysis, cytogenetics, fluorescent in

situ hybridization (FISH), and array- or bead-based techniques that

search for multiple single exon deletions or duplications.

More discrete sequence alterations rely heavily on the use of

PCR, which allows rapid gene amplification and analysis. Moreover,

PCR makes it possible to perform genetic testing and mutational

analysis with small amounts of DNA extracted from leukocytes or

even from single cells, buccal cells, or hair roots. DNA sequencing

can be performed directly on PCR products. Sequencing of the

whole genome or exome, of selected chromosomes, or of numerous

candidate genes in a single run is now possible with NGS platforms

and has transformed the characterization of patients with rare

disorders and advanced malignancies. Analysis of cell-free DNA

(cfDNA; also referred to as “liquid biopsy”) present in body fluids

is playing a growing role for minimally invasive diagnostics and

disease monitoring. Genomic tests are also widely used for the

detection of pathogens and for the identification of viral or bacterial

sequence variations.

The integration of genomic tests into clinical medicine is associated with a number of ongoing challenges related to costs, variable

sensitivities of the tests, bioinformatics analyses, storage and sharing of data, and the difficulty of interpreting all genetic variants

identified with comprehensive testing. The discovery of incidental

(or secondary) findings that are unrelated to the indication for the

sequencing analysis, but indicators of other disorders of potential

relevance for patient care can pose a difficult ethical dilemma. It can

TABLE 466-7 Genetic Approaches for Identifying Disease Genes

METHOD

INDICATIONS AND

ADVANTAGES LIMITATIONS

Linkage Studies

Classical linkage analysis

(parametric methods)

Analysis of monogenic

traits

Difficult to collect large

informative pedigrees

Suitable for genome scan Difficult to obtain

sufficient statistical

power for complex traits

Control population not

required

Useful for multifactorial

disorders in isolated

populations

Allele-sharing methods

(nonparametric methods)

Suitable for identification

of susceptibility genes

in polygenic and

multifactorial disorders

Difficult to collect

sufficient number of

subjects

Affected sib and relative

pair analyses

Suitable for genome scan Difficult to obtain

sufficient statistical

power for complex traits

Sib pair analysis Control population

not required if allele

frequencies are known

Reduced power

compared to classical

linkage, but not sensitive

to specification of

genetic mode

Statistical power can be

increased by including

parents and relatives

Association Studies

Case-control studies Suitable for identification

of susceptibility genes

in polygenic and

multifactorial disorders

Requires large sample

size and matched control

population

Linkage disequilibrium Suitable for testing

specific allelic variants of

known candidate loci

False-positive results in

the absence of suitable

control population

Transmission

disequilibrium test (TDT)

Facilitated by

comprehensive catalogs

of genotypes and

variants

Candidate gene

approach does not permit

detection of novel genes

and pathways

Whole-genome

association studies

Does not necessarily

need relatives

Susceptibility genes can

vary among different

populations

Next-Generations Sequencing Technologies

Whole exome or genome

sequencing

Unbiased approach,

analysis can be

performed without

reference sequences

from parents or siblings

Requires appropriate

bioinformatics, may have

low sensitivity if CNV

analysis is not included,

detects numerous VUS,

can lead to the detection

of unrelated deleterious

alleles

Targeted sequencing of

gene panels

Captures multiple

candidate genes and

loci with hybridization

techniques followed by

deep sequencing

Permits analyses of

multiple candidate

genes in parallel;

facilitates molecular

characterization of

disorders with locus

heterogeneity

Abbreviations: CNV, copy number variation; VUS, variants of unknown significance.

3660 PART 16 Genes, the Environment, and Disease

Patient with (advanced) cancer

DNA and RNA extraction

Bioinformatics

Tumor board

Tumor biopsy: Somatic analysis

Peripheral cells: Germline analysis

Therapy integrating genetic

and genomic information

Treatment based

on pathophysiology

Genetic counseling

Testing of other family members

Targeted sequencing

Deep sequencing of DNA

Deep sequencing Deep sequencing of RNA (RNAseq)

(Linkage analysis and

sequencing of linked region)

Determine functional

properties of identified

mutations in vitro and in vivo

Characterization of phenotype

Familial or sporadic genetic disorder

Gene unknown Gene known or

candidate genes

Pedigree analysis

Mutational analysis

FIGURE 466-17 Approach to genetic disease.

lead to the detection of undiagnosed medically actionable genetic

conditions but can also reveal deleterious mutations that cannot

be influenced, as numerous sequence variants are of unknown

significance.

A general algorithm for the approach to mutational analysis in

patients with a suspected genetic disorder and (advanced) malignancies is outlined in Fig. 466-17. The importance of a detailed

characterization of the clinical phenotype cannot be overemphasized. This is the step where one should also consider the possibility

of genetic heterogeneity and phenocopies. If obvious candidate

genes are suggested by the phenotype, they can be analyzed directly.

After identification of a mutation, it is essential to demonstrate that

it segregates with the phenotype. The functional characterization of

novel mutations is labor intensive and may require analyses in vitro

or in transgenic models in order to document the relevance of the

genetic alteration.

Prenatal diagnosis of numerous genetic diseases in instances with

a high risk for certain disorders is possible by direct DNA analysis.

Amniocentesis involves the removal of a small amount of amniotic

fluid, usually at 16 weeks of gestation. Cells can be collected and

submitted for karyotype analyses, FISH, and mutational analysis

of selected genes (Table 466-4). The main indications for amniocentesis include advanced maternal age (>35 years), presence of an

abnormality of the fetus on ultrasound examination, an abnormal

serum “quad” test (α fetoprotein, β human chorionic gonadotropin,

inhibin-A, and unconjugated estriol), a family history of chromosomal abnormalities, or a Mendelian disorder amenable to genetic

testing. Prenatal diagnosis can also be performed by chorionic

villus sampling (CVS), in which a small amount of the chorion is

removed by a transcervical or transabdominal biopsy. Chromosomes and DNA obtained from these cells can be submitted for

cytogenetic and mutational analyses. CVS can be performed earlier

in gestation (weeks 9–12) than amniocentesis, an aspect that may

be of relevance when termination of pregnancy is a consideration.

Later in pregnancy, beginning at ~18 weeks of gestation, percutaneous umbilical blood sampling (PUBS; cordocentesis) permits

collection of fetal blood for analysis. Prenatal cfDNA allows DNA

analyses from the mother and fetus from a maternal blood sample

to screen for certain chromosomal abnormalities and fetal sex.

These approaches enable screening for clinically relevant and deleterious alleles inherited from the parents, as well as for de novo

germline mutations, and they have the potential to identify genetic

disorders in the prenatal setting.

In combination with in vitro fertilization (IVF) techniques, it is

possible to perform genetic diagnoses in a single cell removed from

the four- to eight-cell embryo or to analyze the first polar body

from an oocyte. Preconceptual diagnosis thereby avoids therapeutic

abortions but is costly and labor intensive. It should be emphasized that excluding a specific disorder by any of these approaches

is never equivalent to the assurance of having a normal child.

Postnatal indications for cytogenetic analyses in infants or children include multiple congenital anomalies, suspicion of a known

cytogenetic syndrome, developmental delay, dysmorphic features,

autism, short stature, and disorders of sexual development, among

others (Table 466-4).

Mutations in certain cancer susceptibility genes such as BRCA1

and BRCA2 may identify individuals with an increased risk for the

development of malignancies and result in risk-reducing interventions. The detection of cytogenetic alterations and mutations is an

important diagnostic and prognostic tool in leukemias, and it has

also transformed the management of solid tumors. In addition to

providing diagnostic information, mutational analysis can inform

the choice of targeted therapies (“actionable mutations”) and characterize the mutational load, which is emerging as an important

indicator of sensitivity to immune checkpoint inhibitors, and can

be used for surveillance.

The demonstration of the presence or absence of mutations

and polymorphisms is also relevant for the rapidly evolving field

of pharmacogenomics, including the identification of differences

in drug treatment response or metabolism as a function of genetic

background

Gene therapy through the introduction of a normal gene or the

ability to make site-specific modifications to the human genome

has, so far, limited clinical application. However, several gene

Principles of Human Genetics

3661CHAPTER 466

transfer methods have now been approved for clinical use, for example, for the treatment of Leber congenital amaurosis, B-cell acute

lymphoblastic leukemia, spinal muscular atrophy, and hereditary

transthyretin-mediated amyloidosis. Moreover, CRISPR-Cas9 genome

editing is currently being investigated in clinical trials as a novel therapeutic approach for selected genetic disorders (Chap. 470).

ETHICAL ISSUES

Determination of the association of genetic defects with disease,

comprehensive data of an individual’s genome, and studies of

genetic variation raise many ethical and legal issues. Genetic information is generally regarded as sensitive information that should

not be readily accessible without explicit consent (genetic privacy).

The disclosure of genetic information may risk possible discrimination by insurers or employers. The scientific components of the

Human Genome Project have been paralleled by efforts to examine

ethical, social, and legal implications. An important milestone

emerging from these endeavors is the Genetic Information Nondiscrimination Act (GINA), signed into law in 2008, which aims

to protect asymptomatic individuals against the misuse of genetic

information for health insurance and employment. It does not,

however, protect the symptomatic individual. Provisions of the U.S.

Patient Protection and Affordable Care Act, effective in 2014, have,

in part, filled this gap and prohibit exclusion from, or termination

of, health insurance based on personal health status. Potential

threats to the maintenance of genetic privacy consist of the emerging integration of genomic data into electronic medical records,

compelled disclosures of health records, and direct-to-consumer

genetic testing.

It is widely accepted that identifying disease-causing genes can

lead to improvements in diagnosis, treatment, and prevention.

However, the information gleaned from genotypic results can have

quite different impacts, depending on the availability of strategies

to modify the course of disease. For example, the identification of

mutations that cause MEN 2 or hemochromatosis allows specific

interventions for affected family members. On the other hand, at

present, the identification of an Alzheimer’s or Huntington’s disease

gene does not currently alter therapy and outcomes. Most genetic

disorders are likely to fall into an intermediate category where the

opportunity for prevention or treatment is significant but limited.

However, the progress in this area is unpredictable, as underscored

by the finding that angiotensin II receptor blockers appear to slow

disease progression in Marfan’s syndrome. Genetic test results can

generate anxiety in affected individuals and family members. Comprehensive sequence analyses are particularly challenging because

most individuals can be expected to harbor several serious recessive gene mutations. Moreover, the sensitivity of comprehensive

sequence analyses is not always greater, for example, if CNV analysis is not integrated but can be associated with higher costs. Genetic

manipulation and patient selection for gene therapy approaches

have aroused ethical controversy and safety concerns that remain

unresolved.

The impact of genetic testing on health care costs remains

unclear. It does vary among disorders and depends on the availability of effective therapeutic modalities, and there are significant

differences among health care systems. A significant problem arises

from the marketing of genetic testing directly to consumers by

commercial companies. The validity of these tests is, in part, not

well defined, and there are persisting concerns about the lack of

appropriate regulatory oversight, the accuracy and confidentiality

of genetic information, the availability of counseling, and the handling of these results.

Many issues raised by the genome project are familiar, in principle, to medical practitioners. For example, an asymptomatic

patient with increased low-density lipoprotein (LDL) cholesterol,

high blood pressure, or a strong family history of early myocardial

infarction is known to be at increased risk of coronary heart disease.

In such cases, it is clear that the identification of risk factors and

an appropriate intervention are beneficial. Likewise, patients with

phenylketonuria, cystic fibrosis, or sickle cell anemia are often identified as having a genetic disease early in life. These precedents can

be helpful for adapting policies that relate to genetic information.

One confounding aspect of the rapid expansion of information

is that our ability to make clinical decisions often lags behind initial

insights into genetic mechanisms of disease. For example, when

genes that predispose to breast cancer such as BRCA1 are described,

they generate tremendous public interest in the potential to predict

disease, but many years of clinical research are still required to rigorously establish genotype and phenotype correlations.

Genomics may contribute to improvements in global health by

providing a better understanding of pathogens and diagnostics and

through contributions to drug development. There is, however,

persisting concern about the development of a “genomics divide”

because of the costs associated with these developments and uncertainty as to whether these advances will be accessible to the populations of developing countries faced with pressing health needs

associated with poverty, infectious diseases, and the relative lack of

essential infrastructure. The World Health Organization’s “Human

Genomics in Global Health Initiative” aims to address these issues

and inequities surrounding genomic medicine.

Whether related to informed consent, participation in research,

or the management of a genetic disorder that affects an individual

or his or her family, there is a great need for more information

about fundamental principles of genetics. The pervasive nature of

the role of genetics in medicine makes it important for physicians

and other health care professionals to become more informed about

genetics and to provide advice and counseling in conjunction with

trained genetic counselors (Chap. 467). Therefore, the application

of screening and prevention strategies does require continuing

patient and physician education, changes in health care financing,

and legislation to protect patient’s rights.

Acknowledgment

Selected sections and Table 466-4 have been integrated from the chapter

on Chromosome Disorders by Dr. Nancy B. Spinner and Dr. Laura K.

Conlin, published in the 19th edition of Harrison’s Principles in Internal

Medicine. The data and concept for Figure 466-16 have been graciously

provided by Dr. Miriam Udler and Dr. Jose Florez, Massachusetts General Hospital and Harvard Medical School, Boston.

■ FURTHER READING

Anastasia L et al: Genomic medicine for undiagnosed diseases.

Lancet 394:533, 2019.

Ashley EA: Towards precision medicine. Nat Rev Genet 17:507, 2016.

Corcoran RB, Chabner BA: Application of cell-free DNA analysis to

cancer treatment. N Engl J Med 379:1754, 2018.

Doudna JA: The promise and challenge of therapeutic genome editing. Nature 578:229, 2020.

Green ED et al: Strategic vision for improving human health at the

forefront of genomics. Nature 586:683, 2020.

High KA, Roncarolo MG: Gene therapy. N Engl J Med 381:455,

2019.

Jameson JL, Longo DL: Precision medicine—personalized, problematic, and promising. N Engl J Med 372:2229, 2015.

Jarvik GP, Evans JP: Mastering genomic terminology. Genet Med

19:491, 2017.

Karczewski KJ et al: The mutational constraint spectrum quantified

from variation in 141,456 humans. Nature 581:434, 2020.

Khera AV et al: Genome-wide polygenic scores for common diseases

identify individuals with risk equivalent to monogenic mutations.

Nat Genet 50:1219, 2018.

Priestley P et al: Pan-cancer whole-genome analyses of metastatic

solid tumours. Nature 575:210, 2019.

Richards S et al: ACMG Laboratory Quality Assurance Committee.

Standards and guidelines for the interpretation of sequence variants:

3662 PART 16 Genes, the Environment, and Disease

A joint consensus recommendation of the American College of

Medical Genetics and Genomics and the Association for Molecular

Pathology. Genet Med 17:405, 2015.

Splinter K et al: Effect of genetic diagnosis on patients with previously

undiagnosed disease. N Engl J Med 379:2131, 2018.

Stuart T, Satija R: Integrative single-cell analysis. Nat Rev Genet

20:257, 2019.

Wauters A, Van Hoyweghen I: Global trends on fears and concerns

of genetic discrimination: A systematic literature review. J Hum

Genet 61:275, 2016.

World Health Organization: Genomics and World Health: Report

of the Advisory Committee on Health Research. 1-254, 2002.

APPLICATIONS OF MOLECULAR GENETICS

IN CLINICAL MEDICINE

Genetic testing for inherited abnormalities associated with disease

risk is increasingly used in the practice of clinical medicine. Germline alterations include chromosomal abnormalities, specific gene

mutations (also called pathogenic variants) with autosomal dominant

or recessive patterns of transmission (Chap. 466), and single nucleotide polymorphisms (SNPs) with small relative risks associated with

disease. Germline alterations are responsible for disorders beyond

classic Mendelian conditions with genetic susceptibility to common

adult-onset diseases such as asthma, hypertension, diabetes mellitus,

macular degeneration, and a number of types of cancer. For many of

these diseases, there is a complex interplay of genes (often multiple)

and environmental factors that affect lifetime risk, age of onset, disease

severity, and treatment options.

The expansion of human genetic knowledge is changing our

understanding of pathophysiology and influencing our classification of diseases. Awareness of genetic etiology can have an impact

on clinical management, including prevention and screening for or

treatment of a range of diseases. Primary care physicians are relied

upon to help patients navigate testing and treatment options. Consequently, they must understand the genetic basis for a large number

of genetically influenced diseases, incorporate personal and family

history to determine the risk for a specific pathogenic variant, and be

positioned to provide counseling. Even if patients are seen by genetic

specialists who assess genetic risk and coordinate testing, primary

care providers should offer information to their patients regarding the

indications, limitations, risks, and benefits of genetic counseling and

testing. They must also be prepared to offer risk-based management

following genetic risk assessment. Given the pace of genetics, this is

an increasingly difficult task. The field of clinical genetics has rapidly

transitioned from single-gene testing to multigene panel testing, with

techniques such as whole exome and genome sequencing on the horizon, increasing the complexity of test selection and interpretation, as

well as patient education and medical decision-making.

COMMON ADULT-ONSET GENETIC

DISORDERS

■ INHERITANCE PATTERNS

Adult-onset hereditary diseases follow multiple patterns of inheritance. Some are autosomal dominant conditions. These include many

467 The Practice of Genetics

in Clinical Medicine

Susan M. Domchek, J. Larry Jameson,

Susan Miesfeldt

common cancer susceptibility syndromes such as hereditary breast

and ovarian cancer (due to germline BRCA1 and BRCA2 pathogenic

variants) and Lynch syndrome (caused by germline mutations in the

mismatch repair genes MLH1, MSH2, MSH6, and PMS2). In both

of these examples, inherited pathogenic variants are associated with

a high penetrance (lifetime risk) of cancer, although penetrance is

incomplete (risk is not 100%). In other conditions, although there is

autosomal dominant transmission, penetrance is lower, thereby making the disorders more difficult to recognize. For example, germline

mutations in CHEK2 increase the risk of breast cancer but with a moderate lifetime risk in the range of 20–30%, as opposed to 50–70% for

mutations in BRCA1 or BRCA2. Other adult-onset hereditary diseases

are transmitted in an autosomal recessive fashion where two mutant

alleles are necessary to cause full expression of disease. Examples

include hemochromatosis and MUTYH-associated polyposis. There

are more pediatric-onset autosomal recessive disorders, such as lysosomal storage diseases and cystic fibrosis.

The genetic risk for many adult-onset disorders is multifactorial.

Risk can be conferred by genetic factors at a number of loci (polygenic),

which individually have very small effects (usually with relative risks of

<1.5). These risk loci (generally SNPs) combine with other genes and

environmental factors in ways that are not well understood. Despite

our incomplete understanding of gene-environment interactions, data

suggest that a healthy lifestyle can mitigate risk associated with elevated

polygenic risk diseases, such as cardiovascular disease. SNP panels are

available to assess risk of disease, but the optimal way of using this

information in the clinical setting to improve patient outcomes remains

uncertain.

Many diseases have multiple patterns of inheritance, adding to the

complexity of evaluating patients and families for these conditions.

For example, colon cancer can be associated with a single germline

mutation in a mismatch repair gene (Lynch syndrome; autosomal

dominant), biallelic mutations in MUTYH (autosomal recessive), or

multiple SNPs (polygenic). Many more individuals will have SNP risk

alleles than germline pathogenic variants in high-penetrance genes,

but cumulative lifetime risk of colon cancer related to the former is

modest, whereas the risk related to the latter is significant. Personal

and family histories provide important insights into the possible mode

of inheritance.

■ FAMILY HISTORY

When two or more first-degree relatives are affected with asthma,

cardiovascular disease, type 2 diabetes, breast cancer, colon cancer, or

melanoma, the relative risk for disease among close relatives ranges

from two- to fivefold, underscoring the importance of family history

for these prevalent disorders. In most situations, the key to assessing

the inherited risk for common adult-onset diseases is the collection

and interpretation of a detailed personal and family medical history in

conjunction with a directed physical examination.

Family history should be recorded in the form of a pedigree, conveying health-related data on first- and second-degree relatives. When

such pedigrees suggest inherited disease, they should be expanded to

include additional family members. The determination of risk for an

asymptomatic individual will vary depending on the size of the family,

the number of unaffected relatives, the types of diagnoses, and the ages

of disease onset. For example, a woman with two first-degree relatives

with breast cancer is at greater risk for a specific Mendelian disorder

if she has a total of 3 female first-degree relatives (with only 1 unaffected) than if she has a total of 10 female first-degree relatives (with

8 unaffected). Factors such as adoption and limited family structure

(few women in a family or multiple early deaths unrelated to the target

disease) should to be taken into consideration in the interpretation of a

pedigree. Additional considerations include young age of disease onset

(e.g., a 30-year-old nonsmoking woman with a myocardial infarction),

unusual diseases (e.g., male breast cancer or medullary thyroid cancer),

and the finding of multiple potentially related diseases in an individual

(e.g., a woman with a history of both colon and endometrial cancer).

Some adult-onset diseases are more prevalent in certain ethnic groups.

The Practice of Genetics in Clinical Medicine

3663CHAPTER 467

GENETIC TESTING FOR ADULT-ONSET

DISORDERS

A critical first step before initiating genetic testing is to ensure that the

correct clinical diagnosis has been made, whether it is based on family

history, characteristic physical findings, pathology, or biochemical

testing. Such careful clinical assessment can define the phenotype.

In the traditional model of genetic testing, testing is directed initially

toward the most probable genes (determined by the phenotype), which

prevents unnecessary testing. Many disorders exhibit the feature of

locus heterogeneity, which refers to the fact that mutations in different genes can cause phenotypically similar disorders. For example,

osteogenesis imperfecta (Chap. 413), long QT syndrome (Chap. 252),

muscular dystrophy (Chap. 449), and hereditary predisposition to

breast (Chap. 79) or colon (Chap. 81) cancer can each be caused by

pathogenic variants in a number of distinct genes. The patterns of

disease transmission, disease risk, clinical course, and treatment may

differ significantly depending on the specific gene affected. Historically, the choice of which gene to test has been determined by unique

clinical and family history features and the relative prevalence of

candidate genetic disorders. However, rapid changes in genetic testing

techniques, as discussed below, are impacting this paradigm. It is now

technically and financially feasible to sequence many genes (or even the

whole exome) at one time. The incorporation of expanded testing for

germline mutations has rapidly evolved both within the clinic as well

as through direct-to-consumer marketing of genetic and genomic tests.

■ METHODOLOGIC APPROACHES TO GENETIC

TESTING

Genetic testing is regulated and performed in much the same way as

other specialized laboratory tests. In the United States, genetic testing laboratories are Clinical Laboratory Improvement Amendments

(CLIA) approved to ensure that they meet quality and proficiency

standards. A useful information source for various genetic tests is www

.genetests.org. It should be noted that many tests need to be ordered

through specialized laboratories.

Genetic testing is performed largely by DNA sequence analysis for

mutations, although genotype can also be deduced through the study

of RNA or protein (e.g., apolipoprotein E, hemoglobin S, and immunohistochemistry). For example, universal Lynch syndrome screening

of colorectal and uterine cancers via immunohistochemical analysis for

absence of expression of mismatch repair proteins is recommended by

the National Comprehensive Cancer Network. The determination of

DNA sequence alterations relies heavily on the use of polymerase chain

For instance, 2.5% of individuals of Ashkenazi Jewish ancestry carry

one of three founder mutations in BRCA1 and BRCA2. Factor V Leiden

mutations are much more common in Caucasians than in Africans or

Asians.

Additional variables that should be documented are nonhereditary

risk factors among those with disease (such as cigarette smoking and

myocardial infarction; asbestos exposure and lung disease; and mantle

radiation and breast cancer). Significant associated environmental

exposures or lifestyle factors decrease the likelihood of a specific

genetic disorder. In contrast, the absence of nonhereditary risk factors typically associated with a disease raises concern about a genetic

association. A personal or family history of deep-vein thrombosis in

the absence of known environmental or medical risk factors suggests

a hereditary thrombotic disorder. The physical examination may

also provide important clues about the risk for a specific inherited

disorder. A patient presenting with xanthomas at a young age should

prompt consideration of familial hypercholesterolemia. The presence

of trichilemmomas in a woman with breast cancer raises concern for

Cowden syndrome, associated with PTEN mutations.

Recall of family history is often inaccurate. This is especially so

when the history is remote and families lose contact or separate geographically. It can be helpful to ask patients to fill out family history

forms before or after their visits, because this provides them with an

opportunity to contact relatives. Ideally, this information should be

embedded in electronic health records and updated intermittently.

Attempts should be made to confirm the illnesses reported in the

family history before making important and, in certain circumstances,

irreversible management decisions. This process is often labor intensive and ideally involves interviews of additional family members or

review of medical records (including pathology reports) and death

certificates.

Although many inherited disorders will be suggested by the clustering of relatives with the same or related conditions, it is important to

note that disease penetrance is incomplete for most genetic disorders.

As a result, the pedigree obtained in such families may not exhibit a

clear Mendelian inheritance pattern because not all family members

carrying the disease-associated alleles will manifest clinical evidence

of the condition. Furthermore, genes associated with some of these

disorders often exhibit variable disease expression. For example, the

breast cancer–associated gene BRCA2 can predispose to several different malignancies in the same family, including cancers of the breast,

ovary, pancreas, skin (melanoma), and prostate. For common diseases

such as breast cancer, some family members without the susceptibility

allele (or genotype) may develop breast cancer (or phenotype) sporadically. Such phenocopies represent another confounding variable in the

pedigree analysis.

Some of the aforementioned features of the family history are

illustrated in Fig. 467-1. In this example, the proband (the individual

serving as the starting point for genetic assessment in a family), a

36-year-old woman (IV-1), has a strong history of breast and ovarian

cancer on the paternal side of her family. The early age of onset and the

co-occurrence of breast and ovarian cancer in this family suggest the

possibility of an inherited mutation in BRCA1 or BRCA2, associated

with the hereditary breast and ovarian cancer syndrome. However,

without genetic testing, it is unclear whether her father harbors such a

pathogenic variant and transmitted it to her. After appropriate genetic

counseling of the proband and her family, the most informative and

cost-effective approach to DNA analysis in this family is to test the

cancer-affected 42-year-old living cousin for the presence of a BRCA1

or BRCA2 mutation. If a pathogenic variant is found, then it is possible

to test for this particular alteration in other family members, if they

so desire. In the example shown, if the proband’s cousin has a BRCA1

pathogenic variant, testing for this alteration (single site testing) would

be an option for the proband’s father. If he tests positive, there is a 50:50

probability that the mutation was transmitted to her, and genetic testing can be used to establish the absence or presence of this alteration.

In contrast, if he tests negative for the known familial BRCA1 mutation

(a true-negative result), the proband and her brother are not at risk for

having inherited this variant from their father.

I

II

III

IV

V

Symbol key

Breast cancer

52

Breast

ca 44

46

Ovarian

ca 43

Ovarian cancer

2

40

Ovarian

ca 38

42

Breast

ca 38

24

Pneumonia

56

36

62

69

Breast

ca 44

55

Ovarian

ca 54

10 62

Accident

6

40

5 2

2

FIGURE 467-1 A 36-year-old woman (arrow) seeks consultation because of her

family history of cancer. The patient expresses concern that the multiple cancers in

her relatives imply an inherited predisposition to develop cancer. The family history

is recorded, and records of the patient’s relatives confirm the reported diagnoses.

3664 PART 16 Genes, the Environment, and Disease

Traditional approach to genetic testing Genetic testing in the era of

next-generation sequencing

Patient identified by clinical

history of familial disorder

Pretest counseling for risks and

benefits of analysis of specific

genes and informed consent

Mutational

analysis

Posttest counseling and

treatment implications for

patient and family members

Pretest counseling for risks and

benefits of analysis of multiple

genes (or whole exome) and

informed consent

Mutational analysis

Interpretation of the findings by

a physician in the context of the

individual’s personal and family

medical history

Posttest counseling and

treatment implications for

patient and family members

Return to primary care

physician for follow-up

Return to primary care

physician for follow-up

If negative

consider next

most likely genes

FIGURE 467-2 Approach to genetic testing.

reaction (PCR), which allows rapid amplification and analysis of the

gene of interest. In addition, PCR enables genetic testing on minimal

amounts of DNA extracted from a wide range of tissue sources including leukocytes (obtained from blood), leukocytes and mucosal epithelial cells (obtained via saliva or buccal swabs), and tumors (obtained

from biopsies or archival tissues). Amplified DNA can be analyzed

directly by DNA sequencing, or it can be hybridized to DNA chips

or blots to detect the presence of normal and altered DNA sequences.

Direct DNA sequencing is frequently used for determination of hereditary disease susceptibility and prenatal diagnosis. Analyses of large

alterations (e.g., deletions, duplications, rearrangements, translocations) of the genome are possible using cytogenetics, fluorescent in

situ hybridization (FISH), Southern blotting, or multiplex ligationdependent probe amplification (MLPA).

Massively parallel sequencing (also called next-generation sequencing) has significantly altered the approach to genetic testing for adultonset hereditary susceptibility disorder. This technology encompasses

high-throughput approaches to DNA analysis that can reliably examine many genes at one time. Technically, this involves sequencing of

millions of small fragments of DNA in parallel. Through bioinformatics, these fragments are pieced together by mapping the individual sequence reads to the human reference genome, a very different

process than traditional Sanger sequencing, which is time-consuming

and expensive.

Multiplex panels for inherited susceptibility are commercially available and include testing of a number of genes that have been associated with the condition of interest. For example, panels are available

for Brugada syndrome, hypertrophic cardiomyopathy, and CharcotMarie-Tooth neuropathy. For many syndromes, this type of panel testing may make sense. However, in other situations, the clinical utility

of panel testing is evolving and may be dependent on the particular

composition of the panel. Currently available breast cancer susceptibility panels contain close to 30 genes, with larger multicancer panels

available. Some of the genes included in the larger multicancer panels

have no known association with breast cancer or have only a modest

associated risk, and the clinical utility is uncertain. An additional

problem of sequencing many genes, rather than focusing on leading

candidate genes, is the identification of one or

more variants of uncertain significance (VUS),

discussed below, or an unexpected yet clinically

relevant result.

Whole exome sequencing (WES) is also now

commercially available, although largely used

in individuals with syndromes unexplained

by traditional genetic testing. As cost declines,

WES may be more widely used. Whole genome

sequencing is also commercially available.

Although it may be quite feasible to sequence the

entire genome, there are many issues in doing so,

including the daunting task of analyzing the vast

amount of data generated. Other issues include

(1) the optimal way in which to obtain informed

consent, (2) interpretation of frequent sequence

variations of uncertain significance, (3) interpretation of alterations in genes with unclear

relevance to specific human pathology, and (4)

management of unexpected but clinically significant genetic findings.

Testing strategies are evolving as a result of

these new genetic testing platforms. As the costs

of multiplex gene panels and WES continue to

fall and as interpretation and understanding

of the clinical relevance of such test results

improve, there has been a shift to more extensive panel-based genetic testing in the clinic. For

example, in the past, a 30-year-old woman with

breast cancer but no family history of cancer and

no syndromic features would undergo BRCA1/2

testing and would be offered TP53 testing in

light of her early-onset disease (notably, a reasonable number of individuals offered TP53 testing for Li-Fraumeni syndrome in the past

declined because mutations are associated with extremely high cancer

risks—including childhood cancers—in multiple organs and means

to mitigate risk are evolving). Without features consistent with other

high-risk, breast cancer–related conditions such as Cowden syndrome,

the woman would not have been routinely offered PTEN analysis

(associated with Cowden syndrome) or testing for other breast cancer–

associated genes including PALB2, CHEK2, and ATM. It is now possible to synchronously analyze all of the aforementioned genes, along

with genes such as BRIP, RAD51C, and RAD51D (which are associated

with moderate ovarian cancer risk but unclear risk for other cancers,

including breast cancer), for a nominally higher cost than BRCA1/2

testing alone. Concerns about such panels include appropriate consent

strategies related to unclear findings, including one or more VUSs,

unanticipated results, and the uncertain clinical utility of some of the

genes included on the panel (Fig. 467-2).

■ DIRECT-TO-CONSUMER GENETIC TESTING

Historically, genetic testing occurred entirely, or predominantly, in the

context of clinical care. However, technologic advances combined with

shifts in social norms have created an increasingly complex landscape

for genetic information, and patients come to providers with a wide

range of information.

Clinician-directed genetic testing is based on the individual’s personal or family medical history. Results are disclosed by the clinician

with a management plan formulated based on results, within the context of the individual’s personal and family medical history. Consumerdirected genetic testing differs in that the individual electively requests

the tests to be performed by a direct-to-consumer (DTC) company and

submits the sample directly. DTC tests are analyzed through proprietary technology, with results communicated directly to the consumer.

DTC testing can include assays for disease risk or disease carrier status; however, they often place a greater emphasis on ancestry or physical traits and characteristics, such as obesity, nutrition, taste, or hair

loss. From a clinical utility standpoint, such testing companies most

often examine sets of SNPs, common “hot spots” in genes associated

The Practice of Genetics in Clinical Medicine

3665CHAPTER 467

with specific diseases; however, SNP analysis does not provide a definitive risk/no risk answer. In contrast, clinical genetic testing thoroughly

interrogates specific genes known be associated with an individual

disease or disease risk. Quality standards between DTC and clinical

laboratories vary greatly. In the United States, clinical genetic testing

labs meet established strict quality standards discernible as CLIA certification and College of American Pathologists (CAP) accreditation.

Some DTC labs are granted U.S. Food and Drug Administration (FDA)

approval for the tests offered but are not CLIA/CAP certified. The

FDA mandates that “results obtained from the tests should not be used

for diagnosis or to inform treatment decisions. Users should consult

a health care professional with questions or concerns about results.”

Recommendations from the American College of Medical Genetics

and Genomics are summarized in Table 467-1.

A third, hybrid model has been developed by commercial genetic

testing laboratories, particularly focused on analysis for cancer and

cardiovascular disease risk. Here, after an individual initiates the testing process, either their own physician or a physician provided by the

company orders the test. Many of these tests are similar to those offered

to a patient in the clinical setting; however, the order and results remain

separate from the medical record unless integrated by the patient’s primary or specialist physician or other health care provider.

Limitations to the accuracy and interpretation of genetic testing

exist. In addition to technical errors, genetics tests are sometimes

designed to detect only the most common pathogenic variants. In addition, genetic testing has evolved over time. For example, it was not possible to obtain commercially available, comprehensive, large genomic

rearrangement testing for BRCA1 and BRCA2 until 2006. Therefore, a

negative result must be qualified by the possibility that the individual

may have a variant that was not detectable in the test. In addition,

the individual may have a pathogenic variant in another untested

cancer-associated gene or in a gene not yet reported to be associated

with elevated disease risk. As such, unless there is a known mutation

in the family, a negative result in an individual with a suggestive

personal or family history is typically classified as an uninformative

negative. In this circumstance, medical management decisions should

be based on personal and family history. For example, a woman with

a strong family history of breast cancer who receives an uninformative

negative panel-based genetic test result may still be eligible for highrisk care, including consideration of MRI-based breast cancer screening and the option of chemoprevention, in addition to close clinical

surveillance and mammograms.

The finding of a VUS is another limitation to genetic testing. A VUS

(also termed unclassified variant) is a sequence variation in a gene where

the effect of the alteration on the function of the protein is not known.

Many of these variants are single nucleotide substitutions (also called

missense mutations) that result in a single amino acid change. Although

many VUSs will ultimately be reclassified as benign variants, some will

prove to be functionally important. As more genes are sequenced (e.g.,

in a multiplex panel or through WES), the percentage of individuals

found to have one or more VUSs increases significantly. The finding

of a VUS is difficult for patients and providers alike and complicates

decisions regarding medical management. In this setting, until there is

further reclassification of the variant, ongoing screening, surveillance,

and care is typically determined based on personal and family history.

Clinical utility is an important consideration because genetic testing

for susceptibility to chronic diseases is increasingly integrated into the

practice of medicine. In some situations, there is proven clinical utility

to genetic testing with significant evidence-based changes in medical management options and recommendations based on results. For

example, there is clear evidence that risk-reducing bilateral salpingooophorectomy benefits women with a documented BRCA1/2 mutation,

relative to both ovarian and breast cancer–related risk. However, in many

cases, the discovery of disease-associated genes has outpaced studies that

assess how such information should be used in the clinical management

of the patient and family. This is particularly true for moderate- and

low-penetrance pathogenic variants. Therefore, predictive genetic testing should be approached with caution and offered to individuals who

have been adequately counseled and have provided informed consent.

Predictive genetic testing falls into two distinct categories. Presymptomatic testing applies to diseases where a specific genetic alteration

is associated with a near 100% likelihood of developing disease. In

contrast, predisposition testing predicts a risk for disease that is <100%.

For example, presymptomatic testing is available for those at risk for

Huntington’s disease, whereas predisposition testing is considered for

those at risk for hereditary colon cancer. It is important to note that

for the majority of adult-onset disorders, testing is only predictive.

Test results cannot reveal with confidence whether, when, or how the

disease will manifest itself. For example, not everyone with the apolipoprotein ε4 allele will develop Alzheimer’s disease, and individuals

without this genetic marker can still develop the disorder.

The optimal testing strategy for a family is to initiate testing in a

(target) disease-affected family member first. Identification of a mutation can direct the testing of other at-risk family members (whether

symptomatic or not). In the absence of additional familial or environmental risk factors, individuals who test negative for the mutation

found in the affected family member can be informed that they are at

general population risk for that particular disease. Furthermore, they

can be reassured that they are not at risk for passing the mutation on

to their children. On the other hand, asymptomatic family members

who test positive for the known mutation must be informed that they

are at increased risk for disease development and for transmitting the

alteration to their children.

Pretest counseling and education are important, as is an assessment

of the patient’s ability to understand and cope with test results. Genetic

testing has implications for entire families, and thus, individuals

interested in pursuing genetic testing must consider how test results

might impact their relationships with relatives, partners, spouses, and

children. In families with a known genetic mutation, those who test

positive must consider the impact of their carrier status on their present and future lifestyles; those who test negative may manifest survivor

guilt. Parents who are found to have a disease-associated mutation

often express considerable anxiety and despair as they address the issue

TABLE 467-1 Summary of American College of Medical Genetics and

Genomics Position Statement on Direct-to-Consumer Genetic Testing

The clinical laboratory

must:

• Be accredited by the CLIA (Clinical Laboratory

Improvement Amendments) program, the state,

and/or other applicable accrediting agencies

with this accreditation indicated on test result

reports

Appropriately trained

professionals should order,

interpret, and disclose test

result to avoid:

• Inadequate or lack of informed consent

• Testing without appropriate indications

• Selection of inadequate testing methods or

incorrect test

• Misinterpretation of results leading to improper

clinical management

The individual undergoing

testing should be

adequately informed of:

• What the test can or cannot determine about

their health or health risk

• The potential for receiving results that neither

confirm nor exclude the possibility of disease

• Potential unexpected results, unrelated to the

indication for testing

• The potential impact of a given genetic test

result on relatives

Professionals will review

and provide information

about:

• The validity and utility of a genetic test, including

the limitations of a given test

• Privacy issues to include review of who will

have access to test results, the processes in

place to protect results, and disposition of DNA

samples once analysis is complete

• Personal or family implications for life, long-term

care, or disability insurance

• Whether data generated will be shared with

third parties

Source: Adapted from ACMG Board of Directors: Direct-to-consumer genetic

testing: A revised position statement of the American College of Medical Genetics

and Genomics. Genet Med 18:207, 2016.

3666 PART 16 Genes, the Environment, and Disease

of risk to their children. In addition, some individuals consider options

such as preimplantation genetic testing (PGT) in their reproductive

decision-making.

When a condition does not manifest until adulthood, clinicians and

parents are faced with the question of whether at-risk children should

be offered genetic testing and, if so, at what age. Although the matter is

debated, several professional organizations have cautioned that genetic

testing for adult-onset disorders should not be offered to children.

Many of these conditions have no known interventions in childhood

to prevent disease; consequently, such information can pose significant

psychosocial risk to the child. In addition, there is concern that testing

during childhood violates a child’s right to make an informed decision

regarding testing upon reaching adulthood. On the other hand, testing

should be offered in childhood for disorders that may manifest early in

life, especially when management options are available. For example,

children with multiple endocrine neoplasia 2 (MEN 2) may develop

medullary thyroid cancer early in life and should be considered for

prophylactic thyroidectomy (Chap. 388). Similarly, children with

familial adenomatous polyposis (FAP) due to a mutation in APC may

develop polyps in their teens with progression to invasive cancer in the

twenties, and therefore, colonoscopy screening is started between the

ages of 10 and 15 years (Chap. 81).

■ INFORMED CONSENT

Informed consent for genetic testing begins with education and counseling. The patient should understand the risks, benefits, and limitations of genetic testing, as well as the potential implications of test

results. Informed consent should include a written document, drafted

clearly and concisely in a language and format that is understandable

to the patient. Because molecular genetic testing of an asymptomatic

individual often allows prediction of future risk, the patient should

understand all potential long-term medical, psychological, and social

implications of testing. There have long been concerns about the potential for genetic discrimination. The Genetic Information Nondiscrimination Act (GINA) was passed in 2008 and provides some protections

related to job and health insurance discrimination. It is important to

explore with patients the potential impact of genetic test results on

future health as well as disability and life insurance coverage. Patients

should understand that alternatives remain available if they decide not

to pursue genetic testing, including the option of delaying testing to a

later date. The option of DNA banking should be presented so that samples are readily available for future use by family members, if needed.

■ FOLLOW-UP CARE AFTER TESTING

Depending on the nature of the genetic disorder, posttest interventions

may include (1) cautious surveillance and awareness; (2) specific medical interventions such as enhanced screening, chemoprevention, or

risk-reducing surgery; (3) risk avoidance; and (4) referral to support

services. For example, patients with known deleterious mutations in

BRCA1 or BRCA2 are strongly encouraged to undergo risk-reducing

salpingo-oophorectomy at an appropriate age and are offered intensive

breast cancer screening as well as the option of risk-reducing mastectomy. In addition, such women may wish to take chemoprevention

with tamoxifen, raloxifene, or exemestane. Those with more limited

medical management and prevention options, such as patients with

Huntington’s disease, should be offered continued follow-up and

supportive services, including physical and occupational therapy and

social services or support groups as indicated. Specific interventions

will change as research continues to enhance our understanding of the

medical management of these genetic conditions and more is learned

about the functions of the gene products involved.

Individuals who test negative for a mutation in a disease-associated

gene identified in an affected family member must be reminded that

they may still be at risk for the disease. This is of particular importance

for common diseases such as diabetes mellitus, cancer, and coronary

artery disease. For example, a woman who finds that she does not carry

the disease-associated mutation in BRCA1 previously discovered in the

family should be reminded that she still requires the same breast cancer

screening recommended for the general population.

GENETIC COUNSELING AND EDUCATION

Genetic counseling is the process of helping people understand and adapt

to the medical, psychological, and familial implications of genetic contributions to disease. This process integrates the following: interpretation

of family and medical histories to assess the chance of disease occurrence or recurrence; education about the natural history of the condition, inheritance pattern, testing, management, prevention, support

resources, and research; and counseling to promote informed choices

in view of risk assessment, family goals, and ethical and religious values. Genetic counseling should be distinguished from genetic testing

and risk-based medical screening and care, though genetic counselors

are involved in these later issues.

Genetic risk assessment is complex and often involves elements

of uncertainty. Genetic counseling can be useful in a wide range of

situations (Table 467-2). The roles of the genetic counselor include

the following:

1. Gather and document a detailed family history.

2. Educate patients about general genetic principles related to disease

risk, both for themselves and for others in the family.

3. Assess and enhance the patient’s ability to cope with the genetic

information offered.

4. Discuss how nongenetic factors may relate to the ultimate expression of disease.

5. Address medical management issues.

6. Assist in determining the role of genetic testing for the individual

and the family.

7. Ensure the patient is aware of the indications, process, risks, benefits, and limitations of the various genetic testing options.

8. Assist the patient, family, and referring physician in the interpretation of the test results.

9. Ensure that the patient has the resources necessary to alert relatives

to their risk, particularly in the face of a positive genetic test result.

10. Address the reproductive implications of a positive genetic test

result, including the risk for a recessive disorder as well as discussion of reproductive options, including gamete donation or preimplantation genetic testing (PGT).

11. Refer the patient and other at-risk family members for additional

medical and support services, if necessary.

The principles of voluntary and informed decision-making and

protection of the individual’s privacy and confidentiality are core principles in the practice of genetic counseling. Genetic counseling is generally offered in a nondirective, noncoercive manner, wherein patients

learn to understand how their values factor into a particular medical

decision. Nondirective counseling is particularly appropriate when

there are no data demonstrating a clear benefit associated with a particular intervention or when an intervention is considered experimental.

For example, nondirective genetic counseling is used when a person is

deciding whether to undergo genetic testing for Huntington’s disease.

At this time, there is no clear benefit (in terms of medical outcome) to

an at-risk individual undergoing genetic testing for this disease because

its course cannot be altered by therapeutic interventions. However,

testing can have an important impact on the individual’s perception of

advanced care planning and his or her interpersonal relationships and

plans for childbearing. Therefore, the decision to pursue testing rests

on the individual’s belief system and values. On the other hand, a more

TABLE 467-2 Indications for Genetic Counseling

Advanced maternal age (>35 years)

Consanguinity

Previous history of a child with birth defects or a genetic disorder

Personal or family history suggestive of a genetic disorder

High-risk ethnic groups

Documented genetic alteration in a family member

Mutation analysis of tumors

Ultrasound or prenatal testing suggesting a genetic disorder

The Practice of Genetics in Clinical Medicine

3667CHAPTER 467

directive approach is appropriate when a condition can be treated. In

a family with FAP, colon cancer screening and prophylactic colectomy

should be recommended for known APC mutation carriers. The counselor and clinician following this family must ensure that the at-risk

family members have access to the resources necessary to adhere to

these recommendations.

Genetic education is central to an individual’s ability to make an

informed decision regarding testing options and treatment. An adequate knowledge of patterns of inheritance will allow patients to understand the probability of disease risk for themselves and other family

members. It is also important to impart the concepts of disease penetrance and expression. For most complex adult-onset genetic disorders,

asymptomatic patients should be advised that a positive test result

does not always translate into future disease development. In addition,

the role of nongenetic factors, such as environmental exposures and

lifestyle, must be discussed in the context of multifactorial disease risk

and disease prevention. Finally, patients should understand the natural

history of the disease as well as the potential options for intervention,

including screening, prevention, and in certain circumstances, pharmacologic treatment or prophylactic surgery.

THERAPEUTIC INTERVENTIONS BASED ON

GENETIC RISK FOR DISEASE

Specific treatments are available for a number of genetic disorders.

Strategies for the development of therapeutic interventions have a long

history in childhood metabolic diseases; however, these principles have

been applied in the diagnosis and management of adult-onset diseases

as well (Table 467-3). Hereditary hemochromatosis is usually caused

TABLE 467-3 Examples of Genetic Testing and Possible Interventions

GENETIC DISORDER INHERITANCE GENES INTERVENTIONS

Oncologic

Lynch syndrome (HNPCC) AD MLH1, MSH2, MSH6, PMS2 Early endoscopic screening; risk-reducing surgery

Familial adenomatous polyposis AD APC Early and frequent endoscopy; prophylactic colectomy;

chemoprevention

Hereditary breast and ovarian

cancer

AD BRCA1, BRCA2 Risk-reducing salpingo-oophorectomy; intensified breast

surveillance including breast MRI; risk-reducing mastectomy

Hereditary diffuse gastric cancer AD CDH1 Prophylactic gastrectomy; enhanced breast cancer surveillance

Hematologic

Factor V Leiden AD F5 Avoidance of thrombogenic risk factors

Hemophilia A XL F8 Factor VIII replacement

Hemophilia B XL F9 Factor IX replacement

Glucose-6-phosphate

dehydrogenase deficiency

XL G6PD Avoidance of oxidant drugs and certain foods

Cardiovascular

Hypertrophic cardiomyopathy AD >10 genes including MYBPC3, MYH7,

TNNT2, TPM1

Echocardiographic screening; pharmacologic intervention;

myomectomy

Long QT syndrome AD, AR >10 genes including KCNQ1, SCN5A,

KCNE1, KCNE2

Electrocardiographic screening; pharmacologic intervention;

implantable cardiac defibrillator devices

Marfan’s syndrome AD FBN1 Echocardiographic screening; prophylactic beta blockers; aortic

valve replacement as indicated

Gastrointestinal

Familial Mediterranean fever AR MEFV Colchicine

Hemochromatosis AR HFE Phlebotomy

Pulmonary

α1

 Antitrypsin deficiency AR SERPINA1 Avoidance of smoking and occupational and environmental toxins

Cystic fibrosis AR CFTR Chest physiotherapy; agents to promote airway secretion

clearance; CFTR modulators; lung transplantation

Endocrine

Neurohypophyseal diabetes

insipidus

AD AVP Replace vasopressin

Familial hypocalciuric

hypercalcemia

AD CASR Avoidance of parathyroidectomy; calcimimetics

Multiple endocrine neoplasia

type 2

AD RET Prophylactic thyroidectomy; screening for pheochromocytoma and

hyperparathyroidism

Renal

Polycystic kidney disease AD, AR PKD1, PKD2, PKHD1 Prevention of hypertension; prevention of urinary tract infections;

kidney transplantation

Nephrogenic diabetes insipidus XL, AR AVPR2, AQP2 Fluid replacement; thiazides with or without amiloride

Neurologic

Malignant hyperthermia AD RYR1, CACNA1S Avoidance of precipitating anesthetics

Hyperkalemic periodic paralysis AD SCN4A Diet rich in calcium and low in potassium; thiazides or

acetazolamide

Duchenne’s and Becker’s

muscular dystrophy

XL DMD Corticosteroids; physical therapy

Wilson’s disease AR ATP7B Zinc, trientine

Abbreviations: AD, autosomal dominant; AR, autosomal recessive; HNPCC, hereditary nonpolyposis colorectal cancer; MRI, magnetic resonance imaging; XL, X-linked.

3668 PART 16 Genes, the Environment, and Disease

by mutations in HFE (although other genes have been less commonly

associated) and manifests as a syndrome of iron overload, which can

lead to liver disease, skin pigmentation, diabetes mellitus, arthropathy,

impotence in males, and cardiac issues (Chap. 414). When identified

early, the disorder can be managed effectively with therapeutic phlebotomy. Therefore, when the diagnosis of hemochromatosis has been

made in a proband, it is important to counsel other family members in

order to minimize the impact of the disorder.

Preventative measures and therapeutic interventions are not

restricted to metabolic disorders. Identification of familial forms of

long QT syndrome, associated with ventricular arrhythmias, allows

early electrocardiographic testing and the use of prophylactic antiarrhythmic therapy, overdrive pacemakers, or defibrillators. Individuals

with familial hypertrophic cardiomyopathy can be screened by ultrasound, treated with beta blockers or other drugs, and counseled about

the importance of avoiding strenuous exercise and dehydration. Those

with Marfan’s syndrome can be treated with beta blockers or angiotensin II receptor blockers and monitored for the development of aortic

aneurysms.

The field of pharmacogenetics identifies genes that alter drug

metabolism or confer susceptibility to toxic drug reactions. Pharmacogenetics seeks to individualize drug therapy in an attempt to improve

treatment outcomes and reduce toxicity. Examples include thiopurine

methyltransferase (TPMT) deficiency, dihydropyrimidine dehydrogenase deficiency, malignant hyperthermia, and glucose-6-phosphate

deficiency. Despite successes in this area, it is not always clear how to

incorporate pharmacogenetics into clinical care. For example, although

there is an association with CYP2C6 and VKORC1 genotypes and warfarin dosing, there is no evidence that incorporating genotyping into

clinical practice improves patient outcomes compared with clinical

algorithms.

The identification of germline abnormalities that increase the risk

of specific types of cancer is rapidly changing clinical management.

Identifying family members with mutations that predispose to FAP or

Lynch syndrome leads to recommendations of early cancer screening

and prophylactic surgery, as well as consideration of chemoprevention

and attention to healthy lifestyle habits. Similar principles apply to

familial forms of melanoma as well as cancers of the breast, ovary, and

thyroid. In addition to increased screening and prophylactic surgery,

the identification of germline mutations associated with cancer may

also lead to the development of targeted therapeutics, for example,

the FDA has approved multiple poly-ADP ribose polymerase (PARP)

inhibitors for BRCA1/2-associated breast, ovarian, pancreatic, and

prostate cancers.

Although the role of genetic testing in the clinical setting continues

to evolve, such testing holds the promise of allowing early and more

targeted interventions that can reduce morbidity and mortality. Rapid

technologic advances are changing the ways in which genetic testing is

performed. As genetic testing becomes less expensive and technically

easier to perform, it is anticipated that there will be an expansion of

its use. This will present challenges but also opportunities. It is critical

that physicians and other health care professionals keep current with

advances in genetic medicine in order to facilitate appropriate referral

for genetic counseling and judicious use of genetic testing, as well as

to provide state-of-the-art, evidence-based care for affected or at-risk

patients and their relatives.

■ FURTHER READING

ACMG Board of Directors: Direct-to-consumer genetic testing:

A revised position statement of the American College of Medical

Genetics and Genomics. Genet Med 18:207, 2016.

Anya ER et al: Genetic information, non-discrimination, and privacy

protections in genetic counseling practice. J Genet Couns 23:891,

2014.

Artin MG et al: Cases in precision medicine: When patients present with direct-to-consumer genetic test results. Ann Intern Med

170:643, 2019.

Clayton EW: Ethical, legal, and social implications of genomic medicine. N Engl J Med 349:562, 2003.

FDA News Release. FDA allows marketing of first direct-toconsumer tests that provide genetic risk information for certain conditions. Available at https://www.fda.gov/NewsEvents/Newsroom/Press

Announcements/ucm551185.htm. Accessed November 18, 2020.

Hampel H et al: A practice guideline from the American College of

Medical Genetics and Genomics and the National Society of Genetic

Counselors: Referral indications for cancer predisposition assessment. Genet Med 17:70, 2015.

Resta R et al: A new definition of genetic counseling: National Society

of Genetic Counselors’ Task Force report. J Genet Couns 15:77, 2006.

Robson ME et al: American Society of Clinical Oncology policy statement update: Genetic and genomic testing for cancer susceptibility. J

Clin Oncol 33:3660, 2015.

Splinter K et al: Effect of genetic diagnosis on patients with previously

undiagnosed disease. N Engl J Med 379:2131, 2018.

Mitochondria are cytoplasmic organelles whose major function is

to generate ATP by the process of oxidative phosphorylation under

aerobic conditions. This process is mediated by the respiratory electron transport chain (ETC) multiprotein enzyme complexes I–V and

the two electron carriers, coenzyme Q10 (CoQ10) and cytochrome c,

located in the inner mitochondrial membrane. Other cellular processes

to which mitochondria make a major contribution include apoptosis

(programmed cell death) and additional cell type–specific functions

(Table 468-1). The efficiency of the mitochondrial ETC in ATP production is the major determinant of overall body energy balance and

thermogenesis. In addition, mitochondria are the predominant source

of reactive oxygen species (ROS), whose rate of production is a delicate

balance between health and disease and relates to the coupling of ATP

production to oxygen consumption. Given the centrality of oxidative

phosphorylation to the normal activities of almost all cells, it is not

surprising that mitochondrial dysfunction can affect almost any organ

system (Fig. 468-1). Until recently, it was thought that disruption of

energy production was the source of the pathophysiology in those with

mitochondrial dysfunction, but recent evidence suggests that free radical production and the redox state of the mitochondria may play a role

468 Mitochondrial DNA

and Heritable Traits and

Diseases

Karl L. Skorecki, Bruce H. Cohen

TABLE 468-1 Functions of Mitochondria

All Cells and Tissues

Oxidative phosphorylation

Free radical production

Calcium homeostasis

Apoptosis (programmed cell death)

Tissue- or Cell-Specific

Cholesterol metabolism

Amino and organic acid metabolism

Fatty acid beta oxidation

Sex steroid synthesis

Heme synthesis

Hepatic ammonia detoxification

Neurotransmitter metabolism

Mitochondrial DNA and Heritable Traits and Diseases

3669CHAPTER 468

and 467. However, mutations in POLG

can disrupt the endonuclease function of

polγ, resulting in somatic mutations in

the mtDNA that endure with future replication. Unless this mutation occurs and

repopulates in an oocyte, it is not heritable. This dual nuclear and mitochondrial

genetic control of mitochondrial function results in unique and diagnostically challenging patterns of inheritance.

The current chapter focuses on heritable traits and diseases related to the

mtDNA component of the dual genetic

control of mitochondrial function. The

reader is referred to Chaps. 449 and

466 for consideration of mitochondrial

disease originating from mutations in

the nuclear genome. The former include

(1) disorders due to mutations in nuclear

genes directly encoding structural components or assembly factors of the oxidative phosphorylation complexes, (2)

disorders due to mutations in nuclear

genes encoding proteins indirectly

related to oxidative phosphorylation, (3)

mtDNA depletion syndromes (MDSs)

characterized by a reduction of mtDNA

copy number in affected tissues without mutations or rearrangements in

the mtDNA, and (4) disorders due to

mutations in nuclear genes that disrupt normal mitochondrial dynamics

(biosynthesis, mitophagy, fission, and

fusion).

The classic physical structure of the

mitochondria is that of a thread-like

organelle, which under fixed conditions,

such as observed with immunohistochemical stains or electron microscopy, has a submarine shape and

measures about 1 μm in length. However, in the living state, mitochondrial shape is highly variable based on the cell type and manifests a complex and ever-changing syncytial form, with continuous

appearance and disappearance of budding structures (representing

mitochondrial fission) and reorganization of separate mitochondria

(representing mitochondrial fusion). Although we often think of the

mitochondrial number in an individual cell, in fact, the more accurate

concept in a living cell is probably mitochondrial volume.

Although the presence of mitochondria has been known for

>150 years, the first knowledge of their respiratory function was proposed ~100 years ago, and the initial description of an illness linked

to mitochondrial dysfunction was only made in 1962. The presence

of mtDNA was noted in the 1960s, and it was not until 1988 when the

first mutations in the mtDNA causing human illness were described.

These included the demonstration of a large-scale mtDNA deletion

ATP

Oxidative

phosphorylation

Nuclear Subunits

DNA

Nuclear DNA

Brain

Seizures

Myoclonus

Ataxia

Stroke

Dementia

Migraine

Skeletal muscle

Weakness

Fatigue

Myopathy

Neuropathy

Heart

Conduction disorder

Wolff-Parkinson-White

syndrome

Cardiomyopathy

Eye

Optic neuropathy

Ophthalmoplegia

Retinopathy

Blood

Pearson's syndrome

Inner ear

Sensorineural

Colon hearing loss

Pseudo obstruction

Liver

Hepatopathy

Kidney

Fanconi's syndrome

Glomerulopathy

Pancreas

Diabetes mellitus

Mitochondrial

DNA

FIGURE 468-1 Dual genetic control and multiple organ system manifestations of mitochondrial disease. (From DR

Johns: Mitochondrial DNA and disease. N Engl J Med 333:638, 1995. Copyright © 1995, Massachusetts Medical Society.

Reprinted with permission from Massachusetts Medical Society.)

as well. Thus, physicians in many disciplines might encounter patients

with mitochondrial diseases and should be aware of their existence and

characteristics.

The integrated activity of an estimated 1500 gene products is

required for normal mitochondrial biogenesis, function, maintenance,

and integrity. Aside from the 37 genes that comprise the mitochondrial

DNA (mtDNA) molecule, the remaining 1400+ gene products are

encoded by nuclear genes (referred to as nDNA) and thus follow the

rules and patterns of nuclear genomic inheritance (Chap. 466). These

nuclear-encoded proteins are synthesized in the cell cytoplasm and

imported to their location of activity within the mitochondria through

a complex biochemical process. This process includes unfolding of the

nuclear-encoded protein, attachment to a chaperone protein that shuttles it through a specific channel to a specific mitochondrial location,

and detachment from the chaperone followed by assembly with other

mtDNA- and nDNA-encoded proteins. In addition, the mitochondria contain their own small genome consisting of numerous copies

(polyploidy) per mitochondrion of a circular, double-strand mtDNA

molecule comprising 16,569 nucleotides. This mtDNA sequence

(also known as the “mitogenome”) might represent the remnants of

endosymbiotic prokaryotes from which mitochondria are thought to

have originated. The mtDNA sequence contains a total of 37 genes,

of which 13 encode mitochondrial protein components of the ETC

(Fig. 468-2). The remaining 22 tRNA- and 2 rRNA-encoding genes are

mitochondria-specific and dedicated to the process of translating the

13 mtDNA-encoded proteins. The mtDNA itself replicates constantly,

independent of cell division, and requires its own unique polymerase,

referred to as polymerase gamma (polγ), which is encoded by the

nuclear gene POLG, disorders of which are discussed in Chaps. 449

I

II

III

FIGURE 468-2 Maternal inheritance of mitochondrial DNA (mtDNA) disorders and

heritable traits. Affected women (filled circles) transmit the trait to their children.

Affected men (filled squares) do not transmit the trait to any of their offspring.