GENE FUSIONS IN LUNG CANCER: RARE OR UNDETECTED?

Gene fusions encoding chimeric oncoproteins usually result
from chromosomal structural rearrangements such as translocations,
inversions, and insertions. When the coding regions
of two genes are juxtaposed, the chimeric transcript produces
a novel protein with an altered function. Despite the numerous
chromosomal rearrangements detected by karyotyping
techniques, only occasional activation of oncogenes has been
found as recurrent event in carcinomas through fusions. 229,230
Thyroid carcinomas exhibit the largest number of reported
gene fusions, most of them involving the RET and BRAF genes.
Aggressive midline and mucoepidermoid carcinomas have
one described gene fusion each ( BRD4-NUT and MECT1-
MAML2 , respectively), while fusions involving the TFE3 gene
in kidney carcinoma and ETV6-NTRK3 and ODZ4-NRG1 in
breast carcinoma were reported in very rare patients ( 1%).
The recent discovery of gene fusions in a large proportion
of prostate carcinomas brought new excitement to the field.
These fusions are generated by translocations or interstitial deletions,
but were identified by advanced technical bioinformatics
approaches for analyses of gene expression rather than by
cytogenetic approaches. 231 The prostate cancer fusions occur
between TMPRSS2 , a prostate-specific, strongly androgenregulated
gene, and the genes of the ETS transcription factor
family ( ERG , ETV1 , and ETV4 ) 232,233 or between ETV1 and
other partners. 234 The fusion of the five prime untranslated region
(5´ UTR) of TMPRSS2 to ERG through an intronic deletion
is the prevalent event in prostate cancer. 231 Investigation
of the prognostic impact of the presence of TMPRSS2-ERG
fusion in prostate carcinoma has generated conflicting results.
A significant association with specific death or development of
metastases is supported by some studies, 235,236 whereas longer
progression-free survival in patients treated by prostatectomy
was reported by other. 237
More recently, the fusion of the ALK (anaplastic lymphoma
kinase) and EML4 (echinoderm microtubule-associated
proteinlike 4) genes, separated by 12 Mb in the short arm of
chromosome 2 and oriented in opposite 5 to 3 directions,
has been identified in NSCLC patients of Japanese origin. 238
Soon after that publication, other reports have confirmed the
occurrence of the EML4-ALK gene fusion, in four distinct
variant forms, in Asian and white lung cancer patients. 239–243
All variant forms of the EML4-ALK fusion gene possess
prominent transforming activity. The fusion creates a chimeric
protein with sequences of EML4 replacing the extracellular
and transmembrane domains of ALK , which results in
constitutive dimerization of the ALK kinase domain and consequent
increase in its catalytic activity. 238 The frequency of
NSCLC carrying this fusion is reportedly low, ranging from
1.5% to 2.6% in whites 242,244 and from 2.6% to 6.7% in
Asians. 238,241–243,245 The frequency of EML4-ALK was associated
with adenocarcinoma histology and limited smoking history
( 10 pack-years). 242 The EML4-ALK fusion was found
in patients who had exon 19 deletion in the EGFR gene but
not in patients with KRAS or BRAF mutations. 242 The prognostic
implication of this fusion in lung cancer is not explored
yet, but the overexpression of ALK in the tumors carrying the
fusion may qualify them for treatment with inhibitors of ALK
kinase, as demonstrated in in vitro models. 242
The detection of gene fusions with relevant role in cancer
causation and progression has been hampered by the challenge
of molecularly identifying cytogenetically cryptic rearrangements.
The TMPRSS2 and ETV gene fusions in prostate cancer
and EML4-ALK in lung cancer were identified by cutting-edge
investigative tools. The ongoing advances in the development
of more sophisticated tools for analyses of the molecular data
already available are expected to substantially increase the detection
of intracellular targets of fusions in a near future.
MICRORNAS IN LUNG CANCER
MicroRNAs are a recently identified class of highly conserved,
endogenous, short noncoding RNAs of 21 to 25 nucleotides
that regulate gene expression in a sequence-specific manner.
These molecules work posttranscriptionally by binding to complementary
sequences in the three prime untranslated region
(3'UTR) of target messenger RNAs (mRNAs), which may lead
to repression of protein translation and downregulation of protein
expression. 246–248 Expression patterns and function of microRNAs
in normal cells are not completely understood. Some
microRNAs are located within introns of pre-mRNAs and are
likely transcribed together with the cognate protein-coding
genes, 246,249,250 whereas others are clustered and transcribed as
multicistronic primary transcripts. 251,252 Although the precise
functions have not yet been characterized for most of the detected
microRNAs, it is known that each individual microRNA
can target numerous transcripts, 253,254 whereas each single gene
can be targeted by numerous microRNAs. 250,255–258 This new
mechanism of gene regulation provides an alternative biologic
explanation for the impact of chromosomal loss or gain in one
area of the genome on the expression of genes mapped in another
part of the genome.
Expression of microRNAs is emerging as an important
area in cancer biology because of the evidence that they are
essential regulators of various physiologic and developmental
processes 249 and are altered in human cancer. 259,260 Most importantly,
signatures of microRNA expression can define molecular
subsets of tumors 259,261 and predict outcome. 260,262,263
There are already scores of studies in microRNAs and
lung cancer with relevant results. For instance, they have been
shown to hold a prognostic effect. Lung adenocarcinoma patients
with high expression of either MIRN155 (21q21.3),
MIRN1 , MIRN106A , MIRN 93 , or MIRN21 and low expression
of either MIRNLET7A2 , MIRNLET7A , or MIRN145
were found to have a significantly worse prognosis 262 and
shortened postoperative survival in NSCLC. 263 Interestingly,
underexpression of two microRNAs mapped at 3p was recently
found to be associated with overexpression of RAS and EGFR
in lung cancer. Loss of these microRNAs would be equivalent
to the loss of a tumor suppressor gene because they downregulate
the expression of the target genes. RAS was determined to
be downregulated by the MIRNLET7G (miRNA let-7g) gene,
which resides on chromosome 3p21.2. 264,265 Expression levels
of MIRNLET7G were on average 30% lower in NSCLC
samples than in normal adjacent tissues 264 and reduction
of tumor growth was observed in tumor xenographs when
overexpression of MIRNLET7G was induced from lentiviral
vectors. 265 The gene MIRN128-2 (miRNA 128b) mapped
at 3p22 was predicted to target EGFR , which is frequently
overexpressed in lung cancer. This finding was hypothesized
to provide a functional link between two common molecular
phenomena in lung cancer, the loss of 3p, and the deregulation
of EGFR . 266 Further exploration of this potential link led to
molecular evidence that MIRN128-2 directly regulates EGFR
and, most importantly, studies in clinical specimens showed
that loss of this miRNA gene was associated with significantly
better disease control and longer survival in NSCLC treated
with gefitinib, an EGFR TKI. In short, MIRN128-1 loss had
similar impact in the sensitivity to the EGFR inhibitors as
EGFR gene gain.
Under the same premises, overexpressed microRNAs are
expected to serve as oncogenes and there are examples of this
role in lung cancer, one of which involves the MIRN17-92
cluster. This gene cluster comprises seven distinct microRNAs
residing in intron 3 of the MIRHG1 gene at 13q31.3 and was
shown to be markedly overexpressed and occasionally amplified
in lung cancer. 267 The predicted targets for the microRNA
cluster comprise a large number of genes, including the tumor
suppressors PTEN and RB2 . 255 Therefore, amplification of the
MIRN17-92 cluster offers the molecular conditions for suppression
of PTEN and RB2 .
Studies focusing on the functional role of microRNA in
cancer, including lung cancer, have been expanded dramatically
lately. Ultimately this knowledge is expected to contribute
not only to the better understanding of cell growth and
differentiation but also to the development of novel therapeutics
and translational tools such as biomarkers for assessment
of risk for disease, early diagnosis, and selection of patients for
treatment with specific agents.

Lung cancer is a challenging disease to patients and their families,
to physicians, and researchers. Lung cancers are characterized
by an extremely diverse collection of genomic alterations,
of which a proportion of unknown dimension is still concealed.
However, numerous pathogenetically important changes have
already been detected in substantial fraction of patients and
translated into a system for detection and determination of the
prognosis of the disease. Specific genomic profiles have supported
the development of new treatment strategies and a growing
use of customized therapy regimens using molecular targeted
or chemotherapeutic agents. These aspects will be discussed in
more details in other chapters. Despite the apparent caveat that
each of the customized therapies is likely to benefit only a small
subset of lung cancer patients, the high incidence of this disease
worldwide ultimately guarantees that the benefit will impact a
large number of patients. Therefore, it remains critical to improve
the characterization of emerging genomic profiles and to
discover new subsets of lung cancer patients. The achievement
of these goals is dependent on new and important insights into
the molecular pathways that underlie lung tumor development.