TUMOR MARKERS&MEDICAL

GENETICS
•One minimally invasive method for early detection and
therapeutic monitoring of cancer involves the measurement
of tumor markers.
•The pathogenesis of the disease has been associated
historically with (1) chemical carcinogens (diet, fumes, skin
contacts), (2) radiation, (3) viruses, and (4) heredity.

* Mechanistically, all of these act by causing DNA mutations,

culminating in the loss of growth control with increased
cellular proliferation and decreased cellular differentiation.
•Since neoplastictransformation is associated with altered
gene expression, which in turn affects the production of
enzymes, hormones, receptors, proteins and metabolites that
are released into the circulatory system, (1)measurement of
these substances aids in the diagnosis and characterization of
the disease.

* Similarly, (2) the DNA content of a tumor cell can be

examined for mutations, the absence of suppressor
genesthat would prevent tumor formation, and the presence
of oncogenesthat turn on tumor growth and ploidy, which is
an indicator of growth rate and aggressiveness.
•This group of substances has come to be known as tumor
markers.

> Characteristics of an Ideal Tumor Marker:

•A tumor markeris a substance synthesized by the host in
response to a tumor that can be used to detect the presence
of the tumor.
•An ideal tumor marker has the following characteristics:
Specificityfor cancer. The substance should be produced
only by the tumor
Sensitivityfor cancer. A very small tumor growth or
metastasis produces measurable amounts of marker.
The amount of marker produced correlates well with the
tumor load.
There should be a consistent concentration of the tumor
marker in patients with stable disease.
The half-lifeof the marker needs to be short enough so that
when production drops, the level of marker falls off rapidly,
producing low or undetectable concentrations in patients
who are in remission.
Levels of the marker (initial and serial) should have
prognostic value.
•The assay for the marker should be analytically sensitive,
specific, accurate, precise, easy to perform, inexpensive, and
rapid.
•Unfortunately, there are no ideal tumor markers.
> Uses of Tumor Markers;
1) Screening for disease in asymptomatic population
2) Diagnosis of disease in symptomatic patients
3) Aid in clinical staging
4) Measurement of tumor burden
5) Therapeutic monitoring and selection
6) Detection of recurrence of disease (relapse).
7) Prognostic indicator
•Tumor markers have limited use in diagnosis because of the
overlap between values obtained in benign and malignant
disease.
•The primary use of tumor markers is to monitor therapy in
patients who have been diagnosed as having a malignant
tumor.

> Biological Sources:

•In general, these are found and measured in body fluids and
tissues.
. Benign tumors are generally well differentiated.
*The cells in a benign tumor are similar to the cells of the
corresponding normal tissue, and therefore the tumor
markers produced are typically the products found in the
normal tissue.
•They may be found in increased amounts in the circulation,
depending on the size of the tumor.
•Malignant tumors may produce substances associated with
a normal cell or they may be different.
•When a normal cell is transformed into a tumor cell, gene
expression changes.
•The affected cell may lose its ability to synthesize some
specific cell product, or it may manufacture greatly increased
amounts.

*The cell may be less specialized than the tissue it evolved

from and assume the characteristics of the less well-
differentiated cells of the embryo, synthesizing proteins (on-
cofetal) found in the embryo but not in a normal adult.
* Tumors are assumed to be unicellular in origin.
•After the cell is transformed, it loses growth control and
begins to divide rapidly.
•The cells lose contact inhibition and invade the primary site.
•They then invade the adjacent organs and blood and lymph
systems, which may carry the cells to distant organs.
•As this invasion process takes place, new proteins can also
be used as markers.
1) Receptors (estrogen, progesterone, androgen, and
corticosteroid)
2) Proteins (immunoglobulins, glycoproteins,
carcinoembryonicproteins, or oncofetalantigens)
3) Genetic markers (oncogenesand suppressor genes)
4) Other markers (sialicacid conjugates, polyamines, and
amino acids
5) Enzymes
•Patterns of commonly measured chemical analytesmay
serve as possible markers of neoplasticdisease and often
serve as a clue to the origin of the disease:
ENZYMES
•Enzymes may have been the first markers used in screening
for cancer.
•An elevated level of any one of them leads to a more
extensive search for disease, although certainly this does not
always lead to the discovery of a malignant disease.
•Other enzymes are more specific for cancer.
•Elevated levels of the prostatic isoenzyme(PAP) of acid
phosphatasehave been used to diagnose and monitor
prostate cancer but may not necessarily be found when
malignancy is present, especially in the early stages of
disease.
•It is sometimes used in conjunction with PSA to discriminate
between patients who have extracapsular disease and those
whose disease is encapsulated’

•ACP is not a reliable screening or early disease diagnostic

tool.

Acid Phosphatase(ACP)
•Alkaline Phosphatase

*Quantitationof total phosphatase(ALP) is useful in the

diagnosis of primary bone and hepatic malignancies and
metastases to these organs.
* Organ-specific ALP isoenzymeshave been identified (bone,
kidney, liver, small intestine, and placenta [Regan
isoenzyme]), and often it is necessary to identify which
isoenzymesare present before ALP can be used as a tumor
marker.
* Frequently, one needs to differentiate the bone from the
liver fraction.
•The isoenzymesmay then be quantitated.
•Elevations of ALP correlate well with osteoblasticactivity
and be used to monitor therapy and look for recurrence in
patients with osteosarcomaor metastatic prostate cancer.
•CreatineKinase

* The BB isoenzymeof creatinekinase(CK-BB, CK-1),

commonly known as the brain fraction, is normally not
observed in serum due to the blood-brain barrier.
•CK-BB is the fetal form of the enzyme and as such is
associated with many malignant diseases, especially those of
epithelial cell origin.
•CK-BB measured in cerebrospinal fluid (CSF) may be used in
estimating lesion size in brain tumors.
•CK-BB isoenzymesare identified and quantitatedusing
electrophoresis.
•Lactate Dehydrogenase
•Lactate dehydrogenase(LD) is a nonspecific marker.
•Elevated serum LD-1has been described in germ cell tumors.
•Similarly, increased serum concentrations of LD-3are
observed in patients with leukemiasand lymphomas, whereas
the LD-5isoenzymeis frequently elevated in patients with
colon, breast, lung, liver, stomach cancers, and a variety of
adenocarcinomas such as that of prostate.
•Neuron-Specific Enolase
•Neuron-specific enolase(NSE) is a specific marker for tumors
associated with the neuroendocrinesystem.
•Serum and/or CSF elevations of this marker are observed.
•NSE may be quantitatedby RIA and the levels correlate well
with the tumor burden
•Amylase, Lipase, Trypsin, and Ribonuclease

Amylase (AMS), lipase (LPS), trypsin(TPS), and

ribonuclease(RNase) are enzymes associated with the
pancreas.
•5’Nucleotidase and Gamma Glutamyltransferase
•5’Nucleotidase (5’NT) and gamma glutamyltransferase(GGT)
are both more sensitive markers for liver cancer than ALP.
•However, neither of them is specific for cancer.
•They may both be elevated in cases of cirrhosis and GGT is
elevated in a large percentage of pancreatic cancers.
•Terminal DeoxynucleotidylTransferase
•It is quantitatedusing an immunoassay (RIA or EIA) and is
used to predict prognosis and responsiveness to drugs as well
as to help classify the leukemias.

•Collagenaseand CathepsinD
•Collagenase and cathepsinD are part of a larger class of
compounds known as proteases.
•Synthesized and secreted by malignant cells, these
compounds are responsible for degrading the extracellular
matrixand thus permitting their invasion and metastasis.
•Both can be quantitatedin extracts from surgically removed
solid tumors.
•The results correlate well with the 5-year survival time for
patients and hence are used in developing a prognosis.
•Collagenaseis measured in bone malignancies and
carhepsinD is often quantitatedin breastcancer patients.
•Histaminases
•Histaminases or diamineoxidase(DAO) has been used as a
tumor marker in cases of medullarythyroid cancer to confirm
a high level of calcitonin, which is normally found in this
disease.
•Muramidase
•Muramidaseor lysozymeis sometimes used in the
monitoring of monocyticand myelomonocyticleukemias.
•The synthesis and secretion of these hormones can cause
severe symptoms and even life-threatening effects.
•

•The paraneoplasticendocrine syndromes, which result from

the excessive hormonal production, are often the first
indication of malignancy and the quantitationof the
hormone(s) in question can support or establish a diagnosis.

HORMONES
* Hormones may be secreted in abnormally increased
concentrations by tumors of the endocrine glands normally
responsible for their production (eutopicproduction) or by
tumors of other organs that normally do not produce the
hormone (ectopic production).
•It is important to note that benign as well as malignant
tumors secrete excessive amounts of hormone.
•A tumor may secrete multiple hormones, some of which
may be synergisticand some antagonistic to each other.
•A tumor may secrete intact hormones, hormone precursors,
fragments, and subunits of hormones.

•Hormones are regularly employed to monitor a patient

following surgery and other therapeutic modalities but they
have not been use to screen an asymptomatic population.
RECEPTORS
•Cell receptors are protein structures located on external cell
membranesand within the cell.
•The function of the receptor is to recognize and bind a
specific ligandsuch as a hormone or neurotransmitter.
•The ligand-receptor complex then initiates a biologic
response.
•The production of receptors is regulated in part by
ligandconcentration.
•Because malignancy causes changes in receptor function and
quantity, receptors can be used as tumor markers.
•Estrogen and Progesterone Receptors
•Estrogen (ER) and progesterone (PgR) receptors are assayed
clinically to determine which breast cancer patients may
respond to endocrine therapy.
•If tumor growth is stimulated by estrogen, an
antiestrogensuch as tamoxifenmay be used to limit the
growth of the tumor.
•Estrogen and progesterone receptors have also been used to
predict the response of patients with endometrial cancer to
hormonal therapy.
•Cell receptor studies are done on a tissue sample obtained
from tumor biopsy or excision.
•OTHER RECEPTORS
•Androgen (AR) and progesterone (PgR) receptors have been
measured to predict the outcome of antiandrogentherapy in
prostate cancer patients.
•Similarly, glucocorticoidreceptors as a prognostic indicator
are being evaluated in cases of lymphoblastic leukemia.
•Epidermal growth factor receptor (EGFR)is a glycoprotein
known to bind epidermal growth factor (EGF) and
transforming growth factor alpha (TGF-α).
•The absenceof this receptor correlates well with a good
response to tamoxifen.
•High levels of the receptor seem to indicate a poor
prognosis in terms of relapse and patient’s survival.
•Lamininreceptor is another potential marker for breast
cancer.
•Lamininis a major constituent of basement membrane.
•Lamininreceptors allow malignant cells to attach to
basement membranes, which are then dissolved, and the
cells destroyed as cancer invades other tissue.
•Malignant cells have more unoccupied lamininreceptors
than do normal cells.
PROTEINS
•Many glycoproteinscan be classified as oncofetalantigens or
as carbohydrate markers.
•Oncofetalproteins (oncofetalantigens) are proteins that are
normally expressed by the fetus but not by the healthy adult.
•Since these proteins are synthesized in high concentration by
tumor cells in cancer patients, they were named
oncofetalantigens or proteins.
•Historically, alpha-fetoproteinand carcinoembryonicantigen
were the first of these proteins discovered.
•Alpha-Fetoprotein
•Alpha-fetoprotein (AFP), so named because it migrates
electrophoretically between alpha1and albumin, is the most
abundant protein present during fetal development.
•The level gradually drops until at the age of about 1 year,
AFP is at normal adult levels (0 to 15 ng/mL).
•AFP is associated with various germ cell tumors and
hepatoma,
•It is useful in characterization and staging of disease and in
monitoring therapy.
•AFP is a good marker for primary hepatoma because it is
elevated in approximately 60-80% of the cases.
•It shows potential as a screening tool for Down’s syndrome.

* In adults, high blood levels (over 500 nanograms/milliliter)

of AFP are seen in only three situations:
· HCC (HepatocellularCA)· Germ cell tumors (cancer of the
testes and ovaries) · Metastatic cancer in the liver (originating
in other organs)
•Additionally, it is elevated in alcoholism, inflammation of
the bowel, cystic fibrosis, and in heavy cigarette smokers.
•Despite the nonspecificityof CEA, it is useful in establishing
prognosis and in monitoring therapy.
•CEA measured in body fluids other than serum (ascetic fluid,
fluid from cyst, urine, or lavagefrom any cavity) may also aid
in diagnosis.

•CarcinoembryonicAntigen
•Carcinoembryonicantigen (CEA) is one of the older
oncofetalproteins still in use.
* It is not specific for the colon but is found in a variety of
malignant and nonmalignant conditions such as breast, GI,
lung, ovary, pancreas, and prostate cancers.
•The carbohydrate tumor markers are glycoproteinsdefined
by their antigenic property.
•They are cell surface antigens or secreted antigens that are
expressed by tumor cells, and against which a monoclonal
antibody or antibodies has been produced.
•These cancer antigens or epitopesare generally more specific
than natural markers, and are defined by the monoclonal
antibody system used to quantitatethem.
•The carbohydratetumor markers can be subdivided into
mucinsand blood group antigens.

CARBOHYDRATES
•Examples of mucin markers are CA15-3 and CA125, whereas
CA19-9 and CA72-4 and examples of blood group antigen
markers.
•It should be noted, however, that patients who genetically
do not express a particular blood group antigen will test
negativefor certain tumor antigens even though they have
the tumor.
•Additionally, a given cancer epitopemay be present in more
than one type of cancer and two or more epitopesmay be
expressed by a single tumor.

* The use of mouse monoclonal antibodies in cancer therapy

has led to the development of a subset of patients who have
human antimouseantibodies (HAMA), which interfere with
mouse-derived monoclonal antibody immunoassays.
•Prostate-Specific Antigen
•The function of PSA is to cause liquefaction of seminal
coagulum.
•Because of its tissue specificity, it became the first tumor
marker recommended for screening for prostate cancer in
older men.
•The serum level is roughly proportional to tumor volume
and should return to normal within a month after radical
prostatectomy. Serum levels are sensitive to recurrence of
disease.
•Increased PSA density (>0.15) and increased PSA velocity
(>0.75 ng/mL/y) correlate better with prostatic cancer than
with BPH.
•CA15-3
•CA15-3 is a glycoprotein mucinpresent on mammary
epithelium.

•Assays for CA15-3 are not specific for breast cancer because
they can be elevated in a variety of carcinomas.
•However, they are sensitive for breast cancer and are
currently used to monitor patients following surgery.
•CA549, CA27.29, breast cancer mucin(BCM), and mucin-like
carcinoma-associated antigen (MCA) are newer breast cancer
markers.
•CA19-9
•CA19-9 is a mucin-like oligosaccharide characterized by
antibody first developed from mice immunized to a human
colon cancer cell line (SW-1116).
•It has a high specificity for pancreatic cancerbut also
recognizes other gastrointestinal cancers.
•CA19-9 is used clinically to monitor therapy and to predict
disease recurrence.
•Because CA19-9 is related to the Lewis substance, Lewis-
negative patients will not produce CA19-9.
•CA125
•CA125 is a glycoprotein defined by the monoclonal antibody
OC125. It is a good marker for ovarian carcinomas.
•It has limited use in diagnosis but has been used in
conjunction with CEA to characterize ovarian tumors.
•The highest levels are associated with ovarian cancer.
•Ovarian cancer antigen (OCA) is thought to be similarly
useful.
•CA72-4
•Immunometricassays for CA72-4 are marketed for use in the
detection of all forms of GI cancer, but especially gastric
cancer, which does not react especially well with other
markers.
•SquamousCell Carcinoma Antigen
•Squamouscell carcinoma antigen (SCC) is one of 14
subfractionsof tumor-associated antigen (TA4).
•Fifty-eight percent (58%) of squamous cell carcinomas
(head, neck, ling, esophagus) show elevated levels of SCC.
•CYFRA 21-1
•Cyfra21-1 is a marker for non-small cell lung cancer
•P-Glycoprotein
•P-glycoprotein is found in cell membranes of drug-resistant
cells. It is theorized that P-glycoprotein is active in
transporting the drugs out of the cells.
•It is normally found in kidney, liver, adrenal, and GI tract cells.
•P-glycoprotein can be measured using a monoclonal
antibody, C219.
•Ferritin
Ferritinis the major iron storage protein and is found in
most cells.
Only a small percentage of total body ferritinis found in
plasma where it binds and transports iron (the major iron
transport protein is transferrin).
It is frequently assayed as an indicator of iron status since
the plasma or serum ferritin level is directly proportional to
the body iron stores.
Ferritinis elevated in any disease that causes a profound
disturbance in iron metabolism and erythropoiesis.
•It may be elevated in hepatitis and aplastricanemia, but it is
also elevated in leukemias, lymphomas, myeloma,
neuroblastoma, gastric, colon, pancreatic, lung and breast
cancers, and in melanoma.
•Beta2-Microglobulin

*Beta2-microglobulin (B2M) is an antigen found on the

surface of all nucleated cells.
•It is a subunit of human leukocyte antigen (HLA) and is
elevated in all diseases associated with rapid cell turnover.
•It is used as a marker for leukemias, lymphoma, and multiple
myeloma and it correlates well with B lymphocyte activity.
•It may also be elevated in patients suffering from human
immunodeficiency virus (HIV) infection.
•C-Peptide
•C-peptide is the connecting peptide between the A and B
chain of proinsulin.
•It is dissociated from these chains to produce active insulin.
•C-peptide is elevated in patients with increased endogenous
insulin production, for example, insulinoma.
•C-peptide is not elevated in patients receiving exogenous
insulin.
•Some patients with beta cell pancreatic tumors produce
fluctuating levels of insulin but their C-peptide levels of
insulin are uniformly high.
•This permits early diagnosis of recurrent disease.
•Immunoglobulins
•The immunoglobulinshave been used as markers in multiple
myeloma and in Waldenstrom’smacroglobulinemiafor many
years.
•They are quantitatedand characterized using electrophoresis
and immunoelectrophoresis(monoclonal bands) and by using
turbidimetricmethods with specific antisera.
•BenceJones protein (urinary free immunoglobulin light
chains) are also present in patients with these cancers.
•Thyroglobulin
•Thyroglobulin(Tg) is a large glycoprotein located in the
thyroid where it serves as a precursor to thyroxine(T4) and
triiodothyronine(T3).
•Thyroglobulinis elevated in patients with follicular and
papillary thyroid carcinoma and thyroid adenoma as well as
in a variety of other thyroid conditions.
•Since thyroglobulin is not elevated in patients with
medullarythyroid carcinoma, it serves to help discriminate
between this and other hyperthyroid diseases.
•Tissue Polypeptide antigen
•Tissue polypeptide antigen (TPA) is an oncofetalprotein
related to the cytokeratins.

*It is synthesized during mitosis and therefore is a useful

marker of cellular proliferation.
Quantitationof this marker has proved useful in monitoring
and staging because it correlates well with the tumor burden.
MEDICAL GENETICS &
Genetic DISORDERs
> Clinical Laboratory Medicine -Focused on 2 Subspecialties of
Genetics:
a) human genetics –the study of heredity in man
b) medical genetics –the study of human genetic
variation of medical significance.
1959 = cytogenetic studies were first utilized in clinical
laboratory studies, making a major advancement in clinical
diagnosis:

a) the ability to detect changes in chromosome structure,

b) direct correlation of this changes to disease & phenotypic
anomalies in individuals.
> Many of tests have become the “gold standard” for
diagnosis.
There is a growing number of disease genes being
cloned… there is increasing emphasis on developing
molecular direct mutation analyses.

These analytical studies work well when a diagnosis

is known or suspected…

Clinical applications for cytogenetic analysis =

range from prenatal diagnosis to cancer evaluation.

Scientists called cytogeneticistscan recognize and
identify many of these gross chromosomal
abnormalities by examining chromosomes through a
microscope.

Cytogeneticistsuse three things to tell chromosomes

apart:
· chromosome size· the position of the centromere·
characteristic banding patterns of alternating light
and dark bands (caused by staining the chromosomes
with dyes)
The parts of a chromosome:
>Telomere –The ends of the chromosome>
Centromere–The primary constriction of the
chromosome;It also divides the chromosome into a
short arm (p) and a long arm (q)> Chromatid–A single
molecule of DNA
CHROMOSOME BANDS LABELING:
The long arm and short arm are labeled q (for
queue) and p (for petit), respectively.

At the lowest resolution, only a few major bands

can be distinguished, which are labeled q1, q2, q3; p1,
p2, p3, etc., counting from the centromere.

Higher resolution reveals sub-bands, labeled q11,

q12, q13, etc, numbered from the centromereout
toward the telomere.
Sub-sub-bands identified by even higher resolution
are labeled q11.1, q11.2, q11.3, etc.
KARYOTYPE
= the representation of entire metaphase
chromosomes in a cell, arranged in order of size.
Autosomesare further divided into seven groups: A to
G
For example, the cytogenetic maplocation of a
genetermed CFTR (cystic fibrosis
transmembraneconductance regulator) is 7q31.2,
which indicates it is on chromosome 7, q arm, band 3,
sub-band 1, and sub-sub-band 2.

> The ends of the chromosomes are labeled pteland

qtel. For example, the notation 7qtel refers to the
telomere (the end) of the long arm of chromosome 7.
Normal male karyotype
Normal female karyotype
MEASURES OF VARIATION
"Genetic variation among individual humans occurs
on many different scales, ranging from gross
alterations in the human karyotypeto single
nucleotidechanges.
1) SINGLE NUCLEOTIDE POLYMORPHISMS
> Nucleotide diversityis based on single mutations
called single nucleotide polymorphisms(SNPs).
> The nucleotide diversity between humans is about
0.1%, which is 1 difference per 1,000 base pairs.
A difference of 1 in 1,000 nucleotidesbetween two
humans chosen at random, amounts to
approximately 3 million nucleotide differences since
the human genome has about 3 billion nucleotides.

Most of these SNPs are neutralbut some are

functional and influence phenotypicdifferences
between humans through alleles.

It is estimated that a total of 10 million SNPs exist in

the human population of which at least 1% are
functional (see International HapMapProject).
2) COPY NUMBER VARIATION
In full sequences of an individual's genome, the analysis
of diploid sequences, has shown that non-SNP variation
accounts for much more human genetic variation than
single nucleotide diversity.

This non-SNP variation includes copy number

variationand results from 1)deletions, 2)inversions,
3)insertionsand 4)duplications.

When copy number variation is included, human to

human genetic variation is estimated to be at least 0.5%
(99.5% similarity).
Copy number variations are inherited but can also arise
during development.
1.a) AUTOSOMAL DOMINANT INHERITANCE
AutosomalDominant Traits are those in which a single
copy of an allele is enough for the trait to be
expressed or shown in the phenotype of the animal.
Dominant conditions are those that are expressed in
heterozygotes.
1.b) AUTOSOMAL RECESSIVE INHERITANCE
Autosomalrecessive traits require that
the Individual have 2 copies of the trait to
express the phenotype.
The genes for autosomal-recessive traits are also
located on the autosomes, but the mutant, disease-
causing alleles are recessive to the normal alleles;
thus, if one normal allele is present, it is usually
sufficient to prevent any expression of the disease.
3) MITOCHONDRIAL INHERITANCE
-refers to the additional genes in cell’s mitochondria.
-Because mitochondria are almost exclusively passed
from parent to child in the egg and not in the sperm,
a hallmark of mitochondrial inheritance is
transmission from an affected woman to all of her
children.
All daughters of an affected father will also be
affected but none of his sons will be affected (unless
the mother is also affected).

In addition, the mother of an affected son is also

affected (but not necessarily the other way round).

> X-linked dominant inheritance works differently

depending upon whether the mother (left image) or
father (right image) is the carrier of a gene that
causes a disease or disorder.

expressed in

(1)males (who are necessarily

hemizygousfor the gene mutation because they have

only one X chromosome); and
(2) females who arehomozygousfor the gene
mutation
> Carrier females do not usually express
thephenotype.
Normal placenta
-maternal side
Normal placenta
-fetal side
Hydatidiformmole
a 3N complement may be derived from dispermy, the fertilization of a
haploid egg by two sperm, and this generally results in a partial
hydatidiformmole.
A ring chromosome is a chromosome whose arms have
fused together to form a ring.


Ring chromosomes may form in cells following genetic
damage by mutagens like radiation, they may also arise
spontaneously during development.

> Symptoms seen in patients carrying ring chromosomes are

more likely to be caused by the deletion of genes in the
telomericregions of affected chromosomes, rather than by
the formation of a ring structure itself.
> Complex rearrangements including segmental
microdeletionsand microduplicationshave been seen in
numerous ring chromosomes providing important clues
regarding the mechanisms of their formation.
Disorders arising from the formation of a ring chromosome
include ring chromosome 20 syndromewhere a ring formed
by one copy of chromosome 20 is associated with epilepsy;
ring chromosome 14 and ring chromosome 13 syndrome are
associated with mental retardationand dysmorphicfacial
features; ring chromosome 15 is associated with mental
retardation, dwarfismand microcephaly.

Ring formation of an X-chromosome causes Turner

syndrome.
dwarfism
microcephaly
Translocationsare rearrangements involving two or more
nonhomologouschromosomes.

Each chromosome breaks off and the pieces exchange

places, establishing two (or more derivative chromosomes.
As with inversions, the majority of translocations are
balanced, and only when the important structural gene is
disrupted does the translocation have clinical ramifications.
Again, the primary danger of a balanced translocation is
that carriers are at increased risk for chromosomally
abnormal livebornchildren.
In the first meiotic division, translocatedchromosomes
assume a cross-shaped structure in order for the alleles to
pair properly.
At anaphase, the altered change in chromosome
complement can be noted directly.
MonosomyX becomes 45, X and gain of a sex chromosome
would be 47, XXY, 47XXX, or 47,XYY.
On the other hand , if the sex chromosome change is
acquired as in some cancer cell lines , a +or –indicating this
would be required (i.e. 45, X-Y for a male whose Y
chromosome has been lost as a result of his disease.
Examples :
Deletion: 46,XX,del(4)(p15)
= Terminal deletion of the short arm of 4 at band 15
Duplication:46,XX, dup(11)(q23)
= Terminal duplication of the long arm of 11 at band 23
Translocation:46,XY,t(4;9)(q21.2;p22)
= Translocation between 4 and 9
Inversion: 46,XY,inv(9)(p11q21.1)
= Pericentricinversion in 9 between bands p11 & q21.1
•Cytogenetic abnormalities may be found in apparently
“normal” individuals as well as in patients with phenotypic
anomalies or with a diagnosed genetic disorder.
•Diagnosis may occur at any stage of life. When the same set
of features is seen in several unrelated individuals, it may be
possible to establish a syndrome.
•The characteristics associated with a syndrome are assumed
to have a common basis, which often can be shown to be a
specific chromosome abnormality.

•However, although a syndrome is defined by a certain set of

characters, there is variability in affected individuals and not
all will show an identical phenotype.
PRENATAL CYTOGENETICS
•Studies have shown that 1 to 13 conceptuseshas a
chromosomal abnormality, but of these, only 6 per1000 are
liveborn, indicating most errors are recognized and
eliminated.
•For example, of all 45,X conceptions, 95% will spontaneously
terminate.
•The most common chromosome abnormalities detected in
prenatal testing are the liveborntrisomiesand the sex
chromosome aneuploidies.
•Because these are usually the result of a meiotic
nondisjuctionerror, there is a very low risk of recurrence.
POSTNATAL
Approximately 0.6% of newborns have a chromosome
abnormality.

If the child dies shortly after birth, cytogenetic analysis may
provide information critical in understanding the demise.
CHILDHOOD AND ADULT
•A common misconception about genetic disorders is that
because they are inherited, the diagnosis will be obvious at
birth.
•In fact, the full clinical presentation of many disorders takes
time to develop and may not be fully expressed until later in
life.

•In addition to considering the full range of cytogenetic

possibilities, molecular and biochemical options must also be
taken into account.
CANCER GENETICS
Another area of medicine in which cytogeneticsis becoming
increasingly important is oncology.

>Excellent clinical data exist for leukemiasand lymphomas,

including specific chromosome rearrangements that are
directly associated with tumorigenesis.
> If treatment is successful, most chromosome abnormalities
are no longer evident in the bone marrow and as long as the
karyotypeappears to be “normal,” the patient would be said
to be in remission.
•However, if the treatment does not eliminate the aberrant
cell line completely, remission may be merely an interlude in
which the disease-causing cells are suppressed to such low
levels that they are not detectable by routine
karyotypeanalysis.
•Then at relapse, the same chromosome anomalies will
reappear and may be accompanied by additional
abnormalities and/or more complex cell lines, finding
consistent with disease progression.
•An increase in complexity over time is known as
karyotypeevolution and, in general, poor prognosis and
severity of disease is directly correlated to the number and
type of chromosome abnormalities seen.
CHROMOSOMAL ANEUPLOIDY SYNDROMES
AutosomalAneuploidies
> The most common cause of mental retardation is trisomy21
or Down syndrome.
This disorder has a birth incidence of 1 in 700 and is
characterized by hypotonia, flat facies, slanted
palpebralfissures, small ears, protruding tongue, transverse
palmarcrease, heart defects, and hypogonadism.

> Approximately 92.5% of all Down syndrome individuals

have 47 chromosomes including three copies of chromosome
21 resulting from a nondisjuctionerror in meiosis.
However, just under 3% of patients tend to express a
milder phenotype and have been shown to be mosaicswith
two cell lines that may include a 46, XX or 46, XY line.
In addition, about 5% of Down syndrome patients have only
46 chromosomes, since the extra 21 is part of a
Robertsonianor other translocation.

> A child with a translocation is often indicative of the

presence of a translocation carrier parent, so, in these cases,
karyotypeanalysis of both parents is generally recommended
to determine if the couple is at increase risk of having
additional Down syndrome childrenin future pregnancies.
•The other two liveborntrisomiesare trisomy13 and
trisomy18. Trisomy13, Patausyndrome, has a birth incidence
of 1 in 4000 to 1 in 10,000.

•Approximately 1 in 8000 newborns are diagnosed with

trisomy18 or Edwards syndrome.
Down syndrome
Patausyndrome
Edwards syndrome
Sex Chromosome Aneuploidies
•The aberrant karyotypesof individuals with three X
chromosomes [47,XXX females] or with one X and two Y
chromosomes [47 XYY males] often go undetected
throughout life.

•Some individuals have generalized learning difficulties.

•Klinefeltersyndrome (47,XXY) males tend to be tall and thin

with relatively long legs.
•The most common reason for referral is
postpubertalhypogonadism; female-like breast development;
and infertility due to small testicles, hyalinizedtesticular
tubules, and azoospermia.

•The most commonly recognized sex chromosome

aneuploidypresenting with a female phenotype is Turner
syndrome: 45,X (Fig. 62-23B)

•The single X chromosome is usually maternal in origin,

indicating that a paternal meiotic nondisjunctionis the most
common source of error.
•The phenotype for Turner syndrome is highly variable.

•Affected individuals are typically short (,4’11”) with

gonadaldysgenesisand learning difficulties.
•In general, however, most livebornTurner syndrome
patients can live productive lives, and mildly affected
individuals may not know that they are affected with the
disorder until puberty.
Turner syndrome
Klinefeltersyndrome
•Wolf-Hirschhornsyndrome also known as 4p-syndrome, is
due to an apparently terminal deletion of the short arm of
chromosome 4 [del(4)(p16)] (Fig 62-14).
•The characteristic facial appearance has been likened to
‘Greek warrior’s helmet’ due to the arched eyebrows and
long nose.

STRUCTURAL CHROMOSOMAL ANOMALIES

•Cri du chat or 5p-syndrome [del(5)(p15)] is characterized by
a distinctive high-pitched, cat-like cry in infancy.
Wolf-Hirschhornsyndrome
Cri du chat
MICRODELETION SYNDROMES AND CONTIGOUS GENE
SYNDROMES
It is a very small deletion, usually only a fraction of a single
chromosome band.

Microdeletionscan be confused with molecular deletions,

so it is important to recognize that although
microdeletionsare small, they are significantly larger (>500
kb) than the typical molecular deletion (1 base pair [bp] to
several hundred bp, which can only be resolved by molecular
technology.

> Certain syndromes are actually due to deletions that

encompass portions of several adjacent, unrelated genes.
The size of the deletion and number of genes affected may
vary such that the phenotype expressed may differ
significantly between individuals.

>These syndromes are collectively called contiguous gene

syndromesand represent a subset of the larger category of
microdeletionsyndromes
Miller-Diekersyndromeclearly demonstrates the overlap
between microdeletionand contiguous gene syndromes.

> The disorder has been associated with a microdeletionof

the distal short arm of chromosome 17 (17p13.3).
> Cardinal clinical features are lissencephaly(smooth brain)
and craniofacial anomalies.
The best known of the microdeletionsyndromes are Prader-
Willisyndrome (PWS) andAngelmansyndromes (AS).
They both share the same interstitial deletion of the
proximal long arm of chromosome 15 [del(15)(q11.2q11.2)],
but the presentation is significantly different.
Prader-Willipatients are small and hypotonic at birth, but
change within the first year of the life & begin to gain weight
rapidly.
If not placed on a controlled diet, they can become quite
obese due to overeating.










The disorder is characterized by cardiac abnormalities,
hypertension, hoarse voice, premature aging of the skin, and
behavioral anomalies.

> Velocardiofacialsyndromeis possibly the most common

microdeletionsyndrome in humans, but is often not
recognized due to a broad spectrum of clinical features and a
presentation that can be quite mild.
OTHER CYTOGENETIC PHENOMENAL FRAGILE X SYNDROME
Fragile X syndrome
= the second leading cause of mental retardation and is the
primary cause of inherited mental retardation.
The mutation was identified as an amplification of a
trinucleotiderepeat sequence.

Breakage Syndromes
Chromosomal breakage syndromes (table 62-7) are a set of
autosomalrecessive disorders that were originally grouped
together due to the common finding of chromosome
instability or fragility. Inability to repair the DNA can lead to:
(1)breakage or increased recombination that can be
characterized by chromosome instability

(2)additional mutations and defects in the DNA sequence

that can lead to cancer.
Williams syndrome
Velocardiofacialsyndrome
Fragile X syndrome
Breakage Syndromes
(AmbrasSyndrome) = gene MAP2K6
•GENETIC MARKERS
•Direct analysis of nuclear DNA is useful in determining
diagnosis (benign versus malignant), determining prognosis
(survival time), monitoring patients for increase aneuplasia,
and predicting or evaluating therapeutic response
•Normal nonproliferatingcells (in G1 or G0 cell cycle phase)
are diploid, whereas DNA doubles during the synthesis phase
(S phase) of proliferation and remains doubled (cell cycle G2)
until mitosis (M phase).
•Cancer cells may have an abnormally increased amount of
DNA (aneuploidy) owing to increased number of
chromosomes, increase DNA per chromosome, and minutes
(small pieces of DNA from viruses).
•Additionally, cancer cells may have an abnormally high
percentage of their cells that are in S phase.
•Both the ploidy(DNA) could be determined by flow cell
cytometry, giving a measure of tumor aggressiveness.
•There are two types of genetic tumor markers: (1)
oncogenes, and (2) anti-oncogenes(suppressor genes).
•The oncogenesare derived from normal cellular genes
(proto-oncogenes).

* The oncogenesare activated by mutational events that

cause increased transcription of growth and proliferation,
promoting protein products or products which antagonize
(suppress) normal cellular aphyperthyroid
ptosis.
•Mutations include: insertions, deletions, translocations,
inversions, and point mutations.
•Many oncogenesare of viral origin and many human
oncogenesare associated with hematologic malignancies but
a few are associated with solid tumors.

* The suppressor genes code for normal protein products that

down regulate (control) cellular proliferation.
* The loss of these genes, and consequently their protein
products, allows continuous transcription of growth,
proliferation, and metastasis, promoting protein products.
•The c-erbB-2 gene(HER-2/neu) codes for a
transmembranereceptor with protein kinaseactivity.

•The protein product functions as a growth factor receptor

and the over expression of the gene is strongly associated
with breast and ovarian cancer.

•Assays exist for both the gene and the protein product, and
results correlate well with prognosis.

* The c-mycgene codes for a protein product (p62) that binds

to DNA and regulates transcription.
ONCOGENES
•Located on Chromosome 1, N-ras(“neuroblastomaRAS viral
(v-ras) oncogenehomolog”).
•Overexpressionof the gene is associated with small cell
carcinoma of the lung, breast carcinoma, gastric and other GI
cancers, and promyelocyticleukemia.

•Translocation of the gene is seen in Burkitt’slymphoma and

a variety of others B and T cell lymphomas.

* The Philadelphia chromosome is the result of a

translocation of the ablgene from the distal portion of
chromosome 9 to the bcrregion of chromosome 22.
•The result of this abnormal gene splicing is to produce an
abl-like tyrosine kinaseprotein product that reacts faster and
with a wider group of substrates.

•The c-abl/bcrgene is involved in signal transduction and

associated with chronic myelogenousleukemia.

•The N-myconcogenewas first observed in brain cancer.

•It is the result of abnormal DNA amplification and its protein

product functions as a transcription regulator.

•It is associated with neuroendocrinetumors.

•SUPPRESSOR GENES
•The DCC gene is located on chromosome 18 and encodes a
protein that is similar to cell adhesion proteins.
•It is responsible for down regulating proliferation in
gastrointestinal cells.
•The loss of this gene causes late stage colon cancer.
•The gene for p53, located on chromosome 17, codes for a
protein that regulates transition into S phase of the cell cycle.
•Mutation or loss of this gene leads to continuous cellular
proliferation.
•Abnormal or absent p53 gene and its product are associated
with several cancers.
•The NFI gene is located on chromosome 17 and acts as a
rasGTpaseactivator.
•The loss of this gene causes neurofibromas, sarcoma, and
glioma.
•The RB gene was first described in patients with inherited
retinoblastoma.
•The gene is located on chromosome 13 and the protein
product (pRB) is an important transcription inhibitor and
regulates transition from G1 to S phase.
•The majority of familial (inherited) breast cancer patients
have a mutation or loss of one of these genes.

•OTHER MARKERS

•Lipid-associated Sialicacid

* Sialicacid is a family of acylatedderivatives of

neuraminicacid usually found on the terminal end of the
carbohydrate portion of glycoprotein or glycolipidin cell
membranes.
•The carbohydrate portion may influence cell-to-cell
interaction, affecting cohesion, adherency, and antigenicity.
•These characteristics change following malignant
transformation of a cell.
•It is used for monitoring patient response to therapy and
recurrence of disease.
•Hydroxyproline
•Hydroxyproline(Hyp) is an amino acid that is elevated in
patient’s with bone metastases.
•It is a marker of invasion and metastasis and is measured in
urine using HPLC.
•Polyamines and acetylated polyamines
•Polyamines (putrescine, spermidineand spermine) are
stabilizing agents that associate with cell membranes and
nucleic acids.
•The polyamines are metabolic products produced by all
proliferation cells and acetylated in the liver.
•Since their concentration in urine appears to parallel the
rate of proliferation, they have been used to monitor therapy
and recurrence of disease.
•Blood Cell Surface Antigens
•Antibodies to lymphoid and myeloid cell surface antigens
together with flow cell cytometrycan be used to confirm the
diagnosis of leukemiasand lymphomas and to discriminate
between different subtypes.
•This is an aid in diagnosis and gives reliable prognostic
information.
•This is because one of the two RB gene alleles is absent in
the germ line, and therefore it only takes the loss of one (the
other) allele to produce a tumor.

•The WT1 gene is located on chromosome 11 and codes for a

nuclear protein that acts as a transcription factor.

•Loss of this gene causes Wilm’stumor (nephroblastoma), a

childhood kidney cancer.

The BRCA1 and BRCA2 suppressor genes are currently under

investigation as markers of genetic susceptibility for breast
cancer.
“For whoever wants to save their life will lose it, but
whoever loses their life for me will find it, says the
Lord .”Matthew 16:25NIV