Chromosomal instability, aneuploidy, and loss of heterozygosity

There are several types of genetic damage that con-
tribute to lung cancer pathogenesis: (i) changes in
chromosome number; (ii) changes in chromosome
structure; (iii) allelic alterations and loss of het-
erozygosity (LOH); and (iv) sequence alterations in
the form of point mutations or small amplifications
or deletions [17]. The first three types of genetic
damage fall under the rubric of genomic instability
and can occur anywhere in the genome, whereas
the final type involves mutations in protein coding
sequences. In this section, we will discuss genomic
instability in the context of chromosomal instabil-
ity, aneuploidy, and loss of heterozygosity. In the
next section, we will discuss the genes frequently
affected by mutational events in human lung can-
cer, and howknowledge of their functionwill trans-
late into novel, effective therapeutics.While we dis-
cuss genomic instability and loss or gain of gene
function in different sections, it is important to re-
alize that these factors are not mutually exclusive
and both contribute to cellular transformation in
complex and cooperative ways. The consequences
of specific alterations in DNA sequence, be they
large-scale translocations or single-pointmutations,
are rarely binary events; rather, it is the accrued ef-
fects of multiple, sequential genetic alterations over
time that gives each tumor its idiosyncratic clinical
course and outcome.
It has been argued that the term genetic insta-
bility properly refers to the rate at which genetic al-
terations occur [17]. Vogelstein and others correctly
argue that the rate of genetic change cannot be in-
ferred from the extant alterations in a given sam-
ple, but rather should be determined experimen-
tally. As a result, here we will distinguish between
the terms genomic instability and genetic instability,
and use the termgenomic instability to refer only to
the fact of alterations in chromosome number (ane-
uploidy) or gross alterations in chromosome struc-
ture through translocation, amplification, and dele-
tion (chromosomal instability). Genomic instability
can involve LOH, particularly in the context of tu-
mor suppressor genes. In this case, one allele has a
mutation or epigenetic change inactivating one al-
lelewhile the otherwild-type allele is lost alongwith
many other genes leaving the cell with a completely
inactive tumor suppressor gene. This commonly oc-
curs in the case of the well-known TSGs p53, p16,
and RB.
LOH refers to the loss of one allele of a given lo-
cus, but says nothing about the number of copies
of that locus. This distinction is important be-
cause tumor cells frequently duplicate their chro-
mosome complement on a background of LOH
such that one parental allele of a chromosome
is lost, but the other is duplicated. The net ef-
fect is that daughter cells are hemizygous for
a given allele, but retain a normal karyotype
for that particular chromosome. The mechanisms
that cause genomic instability include exposure
to carcinogens, hypoxia, hypomethylation of het-
erochromatic DNA, loss of mitotic checkpoint con-
trols, defective DNA repair, and telomere shortening
[18–20].
Karyotypic studies were the first to shed light on
the genetic complexity of cancer pathogenesis, and
one of the first observations was that cancer cells of-
ten exhibit significant aneuploidy. Solid tumors fre-
quently undergo genome duplication early in their
evolution, and many malignancies exhibit a hy-
potetraploid genotype. Genome duplication occurs
during mitosis, and may involve centrosome am-
plification and the formation of multipolar spindles
prior to cytokinesis [21]. Genome duplication prob-
ably occurs in normal cells, but functional mitotic
checkpoints and sentinel DNA damage response
proteins such as p53 and ATM detect aberrant spin-
dle formation and either induce apoptosis or repair
the damage. In preneoplastic cells with mutations
in p53 or other crucial genes, this type of damage
can go undetected.
Karyotypic studies also yielded the first informa-
tion about large genetic alterations in lung cancer.
A major step to achieve lung cancer chromosome
analysis occurred with the ability to grow lung
cancer cells in tissue culture, which allowed prepa-
ration of cancer cell metaphases for analysis [22].
Indeed, karyotypic studieswhere the first to demon-
strate genetic similarities and differences between
NSCLC and SCLC [23]. Frequent sites of chromoso-
mal losses in SCLC include 3p, 5q, 13q, and 17p.
These occur together with double minutes asso-
ciated with amplification of the myelocytomatosis
viral oncogene homolog (MYC), particularly the c-
Myc, family of genes. In NSCLCs, deletions of 3p,
9q, and 17p; +7, i(5)(p10), and i(8)(q10) are com-
mon [23]. Molecular cytogenetic methods includ-
ing array-based comparative genomic hybridization
(CGH), microsatellite marker analysis, and single-
nucleotide polymorphism (SNP) studies have con-
firmed and extended earlier work. CGH analysis
incorporates whole genome-scale analyses with
relatively high-resolution quantitative information
and revealed gains in 5p, 1q24, and Xq26, and dele-
tions in 22q12.1–13.1, 10q26, and 16p11.2 [23].
Comparative genomic studies led to the finding
that nearly all SCLCs and many NSCLCs suffer LOHon chromosome 3p, suggesting the presence of one
or more tumor suppressor genes in this chromo-
some region. Although LOH by itself is not sufficient
to indicate the presence of a tumor suppressor lo-
cus, subsequent, high-resolution analyses showed
that in some SCLCs, several genes in the minimally
deleted 3p21.3 and 3p14.2were deleted on both the
maternal and paternal alleles and thus completely
gone from the cancer cell genome, a so-called ho-
mozygous deletion [24]. Homozygous deletions are
rare in cancer cell genomes, and are taken as a
strong indication that a tumor suppressor gene ex-
ists in the affected region. Other homozygous dele-
tions common in lung cancer occur on 9p21 and
17p13. These loci turned out to include the tumor
suppressor genes p16 and p53, respectively. Subse-
quentwork showed the 3p14.2 region to include the
TSG fragile histidine triad, FHIT, while the 3p21.3
region encodes several closely linked TSGs includ-
ing RASSF1A, FUS1, NPRG2, 101F6, SEMA3B, and
SEMA3F [24,25].
Another common type of genomic instability in
lung cancer primarily affects short repetitive se-
quences of DNA, which are called microsatellites.
These microsatellites, while polymorphic can un-
dergo tumor-specific (compared to normal DNA
from the same patient) alterations in length as a
result of insertion or deletion of the repeating units.
Tumors vary significantly in the rate of microsatel-
lite instability (MSI), which may be due to differ-
ences in extant DNA repair pathways as is the case
in colon cancer [17]. MSI can be measured by us-
ing a series of microsatellite markers in polymerase
chain reaction (PCR)-based assays. The overall fre-
quencies of MSI from 13 studies are 35%for SCLCs
and 22% for NSCLCs [26]. However, it remains to
be determined whether MSI is a cause or corollary
of lung tumorigenesis.