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n e w e ng l a n d j o u r na l


m e dic i n e

review article
Molecular Origins of Cancer

Lung Cancer
Roy S. Herbst, M.D., Ph.D., John V. Heymach, M.D., Ph.D.,
and Scott M. Lippman, M.D.


ung cancer is the leading cause of cancer deaths in the united
States1 and worldwide. The two major forms of lung cancer are non–smallcell lung cancer (about 85% of all lung cancers) and small-cell lung cancer
(about 15%). Despite advances in early detection and standard treatment, non–
small-cell lung cancer is often diagnosed at an advanced stage and has a poor
prognosis. The treatment and prevention of lung cancer are major unmet needs that
can probably be improved by a better understanding of the molecular origins and
evolution of the disease.
Non–small-cell lung cancer can be divided into three major histologic subtypes:
squamous-cell carcinoma, adenocarcinoma, and large-cell lung cancer. Smoking
causes all types of lung cancer but is most strongly linked with small-cell lung
cancer and squamous-cell carcinoma; adenocarcinoma is the most common type
in patients who have never smoked (Fig. 12-8). This review will focus on major
recent advances in the molecular study of the origins and biology of squamous-cell
carcinoma and adenocarcinoma, since they constitute the vast majority of diagnosed
lung cancers (Table 15,8-14). These advances have been facilitated by the development
of molecular techniques and biomarkers for defining cancer risk, prognosis, and
optimal therapy aimed at personalized prevention and treatment of lung cancer.

From the Departments of Thoracic/Head
and Neck Medical Oncology (R.S.H.,
J.V.H., S.M.L.), Cancer Biology (R.S.H.,
J.V.H.), and Clinical Cancer Prevention
(S.M.L.), University of Texas M.D. Anderson Cancer Center, Houston. Address reprint requests to Dr. Lippman at the Department of Thoracic/Head and Neck
Medical Oncology, Unit 432, M.D. Anderson Cancer Center, 1515 Holcombe Blvd.,
Houston, TX 77030, or at slippman@
Drs. Herbst and Heymach contributed
equally to this article.
N Engl J Med 2008;359:1367-80.
Copyright © 2008 Massachusetts Medical Society.

Mol ecul a r Or igins
Host Susceptibility

Epidemiologic studies showing an association between family history and an increased risk of lung cancer provided the first evidence of host susceptibility (Fig. 1).
Lung-cancer susceptibility and risk also are increased in inherited cancer syndromes
caused by rare germ-line mutations in p53,15 retinoblastoma,16 and other genes,17
as well as a germ-line mutation in the epidermal growth factor receptor (EGFR)
gene.18 More recently, three large genomewide association studies identified an association between single-nucleotide polymorphism (SNP) variation at 15q24–15q25.1
and susceptibility to lung cancer. The region of the SNP variation was recently
linked to lung carcinogenesis and includes two genes encoding subunits of the
nicotinic acetylcholine receptor alpha, which is regulated by nicotine exposure.19-22
Lung-cancer susceptibility and risk also increase with reduced DNA repair capacity (particularly when accompanied by exposure to tobacco smoke)23 that results, for example, from germ-line alterations in nucleotide excision repair genes
such as ERCC1.24 Increased expression of DNA synthesis and repair genes, including RRM1 (the regulatory subunit of ribonucleotide reductase) and ERCC1, in non–
small-cell lung cancer correlates with a better prognosis overall but no benefit
from platinum-based chemotherapy.25,26 Table 1 presents gene abnormalities involved in the development of different histologic types of lung cancer.

n engl j med 359;13  september 25, 2008

The New England Journal of Medicine
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Copyright © 2008 Massachusetts Medical Society. All rights reserved.


These changes can persist long term cancer dysregulated ME changes can result in proliferation and apoptosis) to produce premalignant changes. Additional Schwartz in earliest-stage cancer angiogenesis. to smoking — including genetic.g. COLOR FIGURE Environmental factors. including those of mutations in p53. . All rights reserved.. and HER2. Copyright © 2008 Massachusetts Medical Society. 1368 n engl j med 359. invasion and early-stage cancer. and mutations (e. including dysplasia and clonal patches. and advanced cancer and metastasis. also occur in advanced disease. proliferation.13  www. and genetic susceptibility interact to influence carcinogenesis. in p53 and epidermal growth factor receptor [EGFR]).g. For personal use only.6.  september 25.7 Premalignant patches contain clones and subclones (inset). No other uses without permission. 1 Fig # mutations.g.. and sensitivity to treatment through predictive markers. 2014. Smoking-related patches Issue date 9/25/08 and primary cancers (usually squamous-cell carcinoma and small-cell lung cancer) most often develop in the central airway.5 Many molecular changes DE Artist Knoper of heterozygosity. Molecular markers can signify risk (in people without cancer). Molecular Evolution of Lung Cancer.g.8 Most tumors that are not related to smoking are adenocarcinomas and develop in the peripheral airways. epigenetic changes Dysregulated pathways.Draft Herbst from tobacco smoke.4. reflected in the discolored smoking-related lungs) initially occurs in the form of genetic and Author epigenetic changes (e.. Factors that are unrelated 2 Tissue 11 injury9/4/08 (e.. 2008 The New England Journal of Medicine Downloaded from on November 18. and viral (e. human papillomavirus) factors — have been suggested. angiogenesis Metastatic spread Definitive local therapy with or without adjuvant therapy Systemic therapy with or without radiation therapy Prognostic. apoptosis Invasion.4 and eventually lead to aberrant pathway activation and cellular function (e.nejm.The Normal n e w e ng l a n d j o u r na l Precancer of m e dic i n e Early-stage cancer Advanced cancer Smoking-related Clonal patch Tobacco smoke Host susceptibility Unknown factors Not smoking-related Molecular– cellular features Common germ-line genetic variations Treatment approaches Prevention Roles for molecular markers Cancer risk Tissue injury Clonal patches Genetic.g. hormonal. predictive Predictive Figure 1. and promoter methylation) and global transcriptome changes (e. Such stage-specific markers can span the course of disease from its early stages through its late stages. ways). inflammation and apoptosis pathMolecular evolution of lung Title 3. They also can help define mechanisms of resistance to therapy. KRAS. Lung cancers unrelated and related to reset Please check carefully smoking have strikingly different molecular profiles. such as tobacco smoke. prognosis (outcome independent of treatment).. loss of heterozygosity.g. which can involve lossAUTHOR PLEASE NOTE: Figure has been redrawn and type has been microsatellite instability.

All rights reserved. is transforming in ­fibroblasts and occurs in adenocarcinoma but not in other types of non–small-cell lung cancer or other nonlung cancers. The percentages are higher in primary tumors from patients with metastatic disease. Copyright © 2008 Massachusetts Medical Society.g.Molecular Origins of Cancer Table 1.31 Early events in the development of non–smallcell lung cancer include loss of heterozygosity at chromosomal region 3p21. a fragile histidine triad n engl j med 359. proinflammatory interleukin-8 [IL8] and some DNA-repair genes) occur in nonmalignant lung tissue of smokers and patients with lung  september 25.* Abnormality Non–Small-Cell Lung Cancer Squamous-Cell Carcinoma Small-Cell Lung Cancer Adenocarcinoma Precursor Lesion Known (dysplasia) Genetic change Probable (atypical adenomatous hyperplasia) Possible (neuroendocrine field)† p53 mutation KRAS mutation (atypical adenomatous hyperplasia in smokers).27-29 These changes probably precede epithelial clonal evolution.3.30 The size and number of subclones in a clonal patch may contribute to the cancer on November 18. 3p14. § The percentages include increased gene copy numbers from amplification or polysomy and represent percentages from resected cancers. a member of the Ras association domain family. No other uses without permission. Clonal Evolution Changes in certain genes (e. The incidence of 5% is substantially lower than that of 30 to 40% for the detection in squamous-cell carcinoma or adenocarcinoma by immunohistochemical analysis or other techniques.2 (FHIT. 1). Patches of clon- ally related cells. For personal use only.13  www. 2014. have been mapped in the lung. 1369 . consisting of parts of EML4 and ALK. ‖ The anaplastic lymphoma kinase (ALK) fusion gene (involving chromosome 2p). a finding consistent with diffuse tissue injury. ‡ Variations are based in part on smoking profiles.nejm. EGFR kinase domain mutation (in nonsmokers) Overexpression of c-MET KRAS mutation Very rare 10 to 30%‡ Very rare BRAF mutation 3% 2% Very rare Cancer EGFR Very rare 10 to 40%‡ Very rare Amplification§ Kinase domain mutation 30% 15% Very rare Variant III mutation 5%¶ Very rare Very rare Very rare 4% Very rare HER2 Kinase domain mutation Amplification 2% 6% Not known Very rare 7% Not known Mutation 12% 14% 13% Amplification 21% 20% Not known TITF-1 amplification 15% 15% Very rare 60 to 70% 50 to 70%‡ 75% 19% 34% Very rare Mutation 2% 2% Very rare Amplification 33% 6% 4% ALK fusion‖ MET p53 mutation LKB1 mutation PIK3CA * Non–small-cell lung cancer includes squamous-cell carcinoma and adenocarcinoma. an important element of the molecular origins of lung and other cancers (Fig. Increased copy numbers have been reported in squamous dysplastic lesions but not in adenocarcinoma precursors.000 to 360. 2008 The New England Journal of Medicine Downloaded from nejm. and FUS1). ¶ Genomic EGFR variant III mutations have been detected only in lung squamous-cell carcinoma.3 (site of RASSF1A. and these tumors are sensitive preclinically to irreversible EGFR tyrosine kinase inhibitors. Genetic Abnormalities Specific in the Lung to Non–Small-Cell Lung Cancer and Small-Cell Lung Cancer. or clonal patches containing 40.000 cells. † Neuroendocrine fields have been detected only in tissue surrounding tumors and have been characterized by extremely high rates of allelic loss and by c-MET overexpression (Salgia R: personal communication).

TSC2 denotes tuberous sclerosis complex 2. survival. 2008 The New England Journal of Medicine Downloaded from nejm... kinase-independent functions.The n e w e ng l a n d j o u r na l of m e dic i n e Ligand binding and dimerization Hypoxia Other receptor tyrosine kinases (e. death-associated protein kinase (DAPK). cellular effects Proliferation Invasion Metastasis Resistance to apoptosis Angiogenesis COLOR FIGURE Figure 2. and 17p13 (p53). All rights reserved.g. Copyright © 2008 Massachusetts Medical Society. which is involved in energy sensing and cellular  september 25.13  www. such as insulin-like growth factor 1 receptor (IGF-1R) and cMET. chromosome 3p deletions are much more extensive in squamouscell carcinoma). Mutations in the EGFR kinase domain occur early in the development of adenocarcinoma that is generally unrelated to smoking. These findings suggest that in the future molecular analyses of surgical margins may help identify patients most likely to benefit from adjuvant therapy. and mammalian target of rapamycin (mTOR). These pathways also may be modulated drive lung cancer development.nejm. Epidermal Growth Factor Receptor (EGFR) Cell-Signaling Pathways. Draft 8 9/4/08 EGFR activates several major downstream signaling pathways. loss of hetero­ zygosity.g. metaplasia and dysplasia) are p16 and FHIT (frequently and very early) and O-6methylguanine-DNA methyltransferase (MGMT). c-Myc amplification. and RASSF1A (less frequently or rarely and in advanced precancers). which may have an 2 Fig # in turn effect on proliferation. IGF-1R. No other uses without permission.34 Clonal patches with methylation of promoter regions of genes (epigenetic changes). growth and by progression by other receptor tyrosine kinases.35 and may be associated with a 1370 9/25/08 greater risk of recurrence and second primary tumors. p53 mutation.32. and the ME LKB1–amp-activated protein kinase (AMPK) pathway. metastatic spread.. .33. Akt. For personal use only. and microsatellite instability can occur in normal tissue surrounding non–small-cell lung tumors5. However.g. such Artist Knoper 46 as maintaining glucose transport. Loss-of-hetero­ zygosity patterns in squamous-cell carcinoma and adenocarcinoma differ (e. Please check carefully Issue date gene). and KRAS mutations occur early in the development of smoking-related on November 18. in n engl j med 359.34. Methylated genes in premalignant squamouscell lung lesions (e.g.30. including Ras–Raf–Mek and the pathway consisting Author Herbst of phosphoinositide 3-kinase (PI3K). invasiveness. 2014. c-Met) VEGF EGFR LKB1 AMPK PI3K TSC2 HIF-1α Akt mTOR Ras Raf Mek Gene transcription. have been reported.. in normal lung tissue in approximately 50% of smokers)36 exemplifies differences with that in the development of adenocarcinoma. EGFR mutation.32 All these genes are tumor-suppressor genes. Most DE Schwartz of these functions depend on signaling through the kinase domain. 9p21 (p16). and VEGF vasAUTHOR PLEASE NOTE: Figure has been redrawn and type has been reset cular endothelial growth factor.35-41 The early methylation of p16 in the development of squamous-cell lung cancer (e. and tumor angiogenesisTitle throughEpidermal pathways growth that factor are receptor cell signaling pathways either dependent on or independent of the hypoxia inducible factor (HIF).

43 The KRAS. Pten. such as erlotinib and gefitinib. cetuximab (combined with chemotherapy). phosphoinositide 3-kinase (PI3K). Several groups of investigators independently identified somatic mutations in the kinase domain of EGFR in lung adenocarcinoma in approximately 10% of specimens from patients in the United States and in 30 to 50% of specimens from patients in Asia49 (Fig.41 Agents that reverse epigenetic changes have shown promise in a mouse model of lung carcinogenesis and are being tested in humans with lung cancer. were identified recently in studies in mice. high-grade atypical adenomatous hyperplasia). For personal use only. 3). In contrast. 1371 . are Knoper located within the catalytic domain and result in Artist constitutive EGFR activaAUTHOR PLEASE NOTE: Figure sensitivity has been redrawn and has been reset tion. which occur predominantly in adenocarcinoma of the lung.40.g. angiogenesis.g. 2008 The New England Journal of Medicine Downloaded from nejm. apoptosis. EGFR point mutationsME (e. methylated p16)38 and the recurrence of lung cancer (methylated ASC-TMS1. Mol ecul a r E volu t ion EGFR Family EGFR regulates important tumorigenic processes.46 Clinical trials of the EGFR tyrosine kinase inhibitor erlotinib for second-line or thirdline treatment of such tumors and of the monoclonal antibody against EGFR. nonsmokers and are tightly associated with sensitivity to the EGFR tyrosine kinase inhibitors gefitinib and erlotinib and so appear to explain most of the dramatic responses to these agents.2.13  www. for treatment of previously untreated.42 Bronchoalveolar stem cells. EGFR mutations are associated with an improved prognosis in non–small-cell lung cancer.5.nejm.45 The potential role of bronchoalveolar stem cells and other tumorigenic stem-cell populations in the development and prognosis of human lung cancer and its resistance to drugs is an important area of future investigation. EGFR mutations that were discovered during clinical trials led to extensive studies of the roles of these mutations and EGFR amplification in the pathogenesis of the disease and its prognosis and sensitivity to treatment. Effect of Deletions and Mutations in the Epidermal Growth Factor 6 Targeting. and invasion (Fig. including proliferation. with the phosphate derived Title Deletions and mutations of epidermal growth factor from ATP bound within the kinase catalytic domain. is frequently overexpressed in the development and progression of non–small-cell lung cancer.48 validated EGFR as a molecular target for therapy.50‑52 More than 80% of these mutations in lung cancer involve in-frame deletions within exon 19 or the L858R mutant within exon 21. L858R) and exon 19 DE Schwartz deletions. which may be precursors of lung adenocarcinoma. All rights reserved.Molecular Origins of Cancer which p16 methylation occurs very rarely and only late in precursors (e.34.53. and cyclindependent kinase pathways have been implicated in the proliferation of these stem cells.. advanced disease47. EGFR mutations have affecting lung cancer transforming potential in preclinical lung models and candevelopment occur early huand drugin targeting man lung carcinogenesis. along with its ­ligands. is associated with recurrence after resection.37 Methylation markers in sputum are associated with the risk of lung cancer (e.. which is associated with lung-cancer risk n engl j med 359. 2). The mutations occur with increased frequency in women and Extracellular domain Ligand-binding pocket EGFR variant III del (del exons 2–7) Squamous-cell carcinoma Transmembrane domain Intracellular domain Exon 19 del Adenocarcinoma ATP-binding pocket Catalytic domain T790M Adenocarcinoma L858R (exon 21 del) Adenocarcinoma P Phosphorylation COLOR FIGURE Figure 3. including p16 in stage I non–small-cell lung cancer.44. These mutations are associated with increased totypeEGFR tyPlease check carefully rosine kinase inhibitors. 8/28/08 Receptor Gene (EGFR) on Disease Development andDraft Drug Ligand binding to the EGFR extracellular domainAuthor results 3Herbst in receptor hoFig # modimerization and tyrosine phosphorylation. Copyright © 2008 Massachusetts Medical Society. Recent data show that promoter methylation of various genes. EGFR variant III mutant deletions occur in the extracellular domain and are associated with squamous-cell cancer. and. 2014. No other uses without on November 18. mutaIssue date 9/25/08 tions in T790M (an amino acid located within the ATP binding site of the EGFR kinase domain) are associated with acquired resistance to these drugs..54 EGFR amplification is detected in dysplasia (especially of a high grade). even when treated with cytotoxic  september 25.g.39 also called PYCARD).

78.66-70 EGFR mutations. 1). marks a poor prognosis. The vast majority of patients who have an initial response to erlotinib and gefitinib eventually have a relapse. and the frequency of such amplification is 5 to 10%. Preclinical and clinical data suggest that EGFR mutations are early events in the development of non–small-cell lung cancer.65 The epithelial-to-mesenchymal transition is a program of cell development involving loss of cell adhesion.58 Recent studies have identified EGFR T790M mutations (in exon 20) in tumors before drug treatment59 and in tumors of patients who had a relapse after therapy with standard reversible EGFR tyrosine kinase inhibitors.60 Amplification of the met proto-oncogene (MET).13 Mutations of the region that encodes the tyrosine kinase domain of EGFR have been detected in specimens of atypical adenomatous hyperplasia from Asian patients with no history of smoking. HER2 mutations are associated with resistance to such inhibitors but also with sensitivity to HER2-targeted therapy. pathological.64. Copyright © 2008 Massachusetts Medical Society. with a possible association with male sex and smoking. repressed E-cadherin expression.68 Deinduction 1372 of m e dic i n e of mutant EGFR expression led to regression of tumors. nonsmoking status.58. atypical adenomatous hyperplasia. The frequency of such mutations is less than 5%.13  www. virtually mutually exclusive of mutations in the EGFR and HER2 kinase domains. All rights reserved.71 These mutations also occur in normal epithelium within and adjacent to tumors with EGFR tyrosine kinase mutations (a localized-field effect. a change associated with tumor progression and metastasis. KRAS mutations.58. and increased cell mobility. and Asian background in patients with adenocarcinoma.56 The epidemiologic links highlight three factors — whether the patient is a nonsmoker. suggesting the need for persistent mutant EGFR activity for continued tumor survival. .61-63 Other proposed resistance mechanisms include activation of other receptor tyro­ sine kinases.49.79 A dis- n engl j med 359. was followed by lesions resembling bronchioalveolar carcinoma at 5 to 6 weeks of age and invasive adenocarcinomas at 8 to 10 weeks. and 20 and L858R. possibly reflect­ ing stem-cell expansion) and before EGFR amplification. 2014.2. HER2 kinase domain mutations (in-frame insertions in exon 20) and EGFR kinase domain mutations have similar associations with female sex. No other uses without permission.79 Most KRAS mutations in lung adenocarcinoma are smoking-related G→T transversions (substitutions of a purine for a pyrimidine) and affect exon 12 (in 90% of patients) or exon 13. 19. However. and female — that are associated independently and collective­ ly with an improved response to EGFR tyro­sine kinase inhibitors.57 The binding kinetics of the mutant EGFR appear to be altered by the T790M mutation (Fig.75 HER3 kinase domain mutations have not been detected in patients with non–small-cell lung cancer.73 HER2 amplification is associated with sensitivity to inhibitors of the EGFR tyrosine kinase74. which is considered to be a precursor lesion of peripheral adenocarcinoma. Lung tumors also developed in transgenic mice with lung-specific expression of EGFR variant III mutation (in-frame deletion of exons 2–7 from the extracellular domain) (Fig. in transgenic mice with lung-specific expression of exon 19 deletion or the L858R mutation. and associated with resistance to EGFR inhibitors (tyrosine kinase inhibitors and cetuximab) and chemotherapy. 2). another major mechanism of acquired resistance to EGFR tyrosine kinase on November 18. Irreversible EGFR inhibitors suppress T790M-mutant tumor cells in vitro and are promising treatments for T790M-mutant tumors.57. erlotinib appears to prolong survival in virtually all subgroups of patients with non–small-cell lung cancer. sex-related. can transform fibroblasts and lung epithelial cells. and is associated with a poor prognosis but also with sensitivity to EGFR inhibitors.55. and molecular factors between smokers and lifelong nonsmokers in whom lung cancer develops (Fig.76 Mutations in the HER4 kinase domain were found in 2 to 3% of Asian patients with this disease. including those involving exons 18. 3). Asian. 2008 The New England Journal of Medicine Downloaded from nejm.72 HER2 mutations and amplification have been identified in patients with lung adenocarcinoma.48 There are major differences in clinical. For personal use only. 3). such as insulin-like growth factor 1 receptor (which can bypass EGFR to activate critical downstream signaling pathways [Fig. Activating KRAS mutations are limited to non–small-cell lung cancer (predominantly adenocarcinomas).org  september 25. and the epithelial-to-mesenchy­ mal transition. 2]).The n e w e ng l a n d j o u r na l when detected in the sputum of smokers.67 Furthermore.77 Ras–Raf–Mek The Ras–Raf–Mek pathway is involved in signaling downstream from EGFR and in other pathways leading to the growth of cancer cells and tumor progression (Fig.

especially squamous-cell carcinoma. For personal use only.13  www. in contrast to  september 25.87 The presence of LKB1 mutations alone (i. much lower rates of LKB1 mutation (<5%) were found in adenocarcinomas from Asian patients.g. Copyright © 2008 Massachusetts Medical Society.3 region was the most common focal event in a high-resolution analysis of gene copy numbers in human lung adenocarcinoma. its functional significance is unclear. the antitumor effect of mTOR inhibition requires the tumor microenvironment.Molecular Origins of Cancer tinct KRAS mutational profile consisting of G→A transition mutations was recently detected in patients with adenocarcinoma who had never smoked.3 region caused increased growth of the cells. No other uses without permission.nejm. An mTOR inhibitor can block malignant progression of atypical adenomatous hyper­ plasia lesions in the KRas mouse model. 2008 The New England Journal of Medicine Downloaded from nejm. In vitro.91 LKB1 mutations are associated with smoking and with KRAS mutations and are virtually exclusive of EGFR mutations. and several cellcycle genes in the tumor specimens. vascular endothelial growth factor (VEGF). also called NKX2-1) in the 14q13. without KRas mutations) was not associated with the development of lung cancer in mice.89 LKB1 mutations (including point mutations and deletions) were found in 34% of adenocarcinomas and 19% of squamouscell carcinomas from 144 human specimens of non–small-cell lung cancer.84 Since mTOR drives tumorigenesis in part through macrophages.. is activated early in lung carcinogenesis.93 suggesting that these three genes may work cooperatively in the pathogenesis of lung cancer in which there is amplification at 14q13.91 PI3K–Akt–mTOR TITF1 The pathway consisting of PI3K. as detected on fluorescence in situ hybridization.VEGF levels in bronchial epithelial cells of smoktion in early tumorigenesis.82 tiation. and mammalian target of rapamycin (mTOR). There is mutation or amplification of PIK3CA.81 BRAF mutations have also been detected in non–small-cell lung cancer9 and may be an early event in lung tumorigenesis. Low levels of LKB1 protein were associated with high grades of dysplasia in atypical adeno­ matous hyperplasia lesions.79 Transversions (smokers) and transitions (nonsmokers) also have been reported for p53 mutations in lung adenocarcinoma. 2014. transfection of immortalized normal human lung epithelial cells with at least two of the three genes TITF1. a prominent component of the tumor microenvironment. Myc. detectable in the preinvasive lesions of atypical adenomatous hyperplasia and bronchoalveolar carcinoma33) that precedes smoking-related lung adenocarcinoma. and the development of metastases.2 KRAS mutations appear to be an early event (e. Further evidence supporting this gene’s role in the pathogenesis of lung cancer comes from transgenic mice bearing a mutated KRAS and in which multifocal atypical adenomatous hyperplasia and adenocarcinoma develop. Akt.86.87 LKB1 is thought to func. As a result. the investigators also identified amplification in regions containing KRAS.e.83 Akt is also overexpressed in bronchial dysplasia.94 LKB1 LKB1 (also called STK11) is frequently mutated in non–small-cell lung tumors and is thought to act as a tumor-suppressor gene through interactions with p53 and CDC42. modulating the activity of AMPK (a multifunctional protein kinase) and other possible mechanisms that are just begin. which encodes the PI3K catalytic subunit. subsequent differen.80 MET activation occurs early in KRAS-induced carcinogenesis in this model.. All rights reserved. Recent data indicate that squamous-cell carcinoma also exhibits TITF1 amplification. but not TITF1 protein. which is downstream of EGFR.88 Results in transgenic KRas-mutant mice in which LKB1 was inactivated suggest that the gene plays a role in the differentiation and invasive behavior of such tumors. in association with increased PI3K activity and Akt expression.85 Amplification of thyroid transcription factor 1 (TITF1.90. Inhibition of Akt can induce apoptosis of human premalignant and malignant lung cells and prevent lung carcinogenesis in an animal model.ers increase in association with the progression n engl j med 359. NKX2-8. They generally mark a poor prognosis. in a subgroup of non–small-cell lung tumors.Angiogenesis ning to be on November 18.92 This study used an array with the capacity to genotype many SNPs. and PAX-9 in the 14q13. suggesting that LKB1 has an early role in the development of premalignant lesions in the lung. TITF1 encodes a lineage-specific transcription factor that is essential for the formation of cells lining lung alveoli (type II pneumocytes). 1373 .87 However.3 (detected by high-resolution comparative genomic hybridization array). on November 18. Emerging high-throughput techniques for assessing genomic DNA. controlled trials will need to validate these signatures and establish whether the patients with stage IA tumors who were identified as being at high risk will benefit from adjuvant therapy. Phase 3 testing of both approaches in patients with platinum-resistant disease is ongoing. including the profiling of genes and proteins. 1).The n e w e ng l a n d j o u r na l of bronchial dysplasia from low grade to high grade. One such phase 3 trial. coordinated by the Cancer and Leukemia Group B. is approved and under final review.115 In a recent analysis of 672 invasion-associated genes from 125 frozen specimens of early-stage tumors. No other uses without permission. alterations in gene copy number. In two validation cohorts.109 and with reduced recurrence-free or overall survival of patients have been identified.107 had promising results. Copyright © 2008 Massachusetts Medical Society. Randomized phase 2 trials of dual inhibition with bevacizumab plus erlotinib105 or the VEGF receptor–EGFR tyrosine kinase inhibitor vandetanib (combined with chemotherapy)106. 4). Microarray techniques that profile the expressions of tens of thousands of genes simultaneously can measure this tumor heterogeneity at a global level. It will evaluate a large predictive set of metagenes (or subgroups of gene-expression profiles consisting of 25 to 200 genes) in patients n engl j med 359. and carcinoma in situ are associated with increased microvessel density. and epigenetic changes.110-113 Combined clinical and molecular information provides better indications of cancer risk114 and prognosis. The Cancer Genome Atlas is a large-scale project designed to provide a comprehensive profile of human tumors according to their gene mutations.nejm. Gene Profiling Tumor molecular heterogeneity is a major reason that patients with non–small-cell lung cancer with a similar clinical stage and tumor histology can have dramatically different clinical outcomes and responses to treatment. multiplicity. and genomic signatures far outstrips the modest improvement in treatments that are based on these molecular advances. microRNA.103 Interactions between the VEGF and EGFR pathways and an association between acquired resistance to EGFR blockade and increased VEGF expression in preclinical models104 led to the hypothesis that dual blockade of VEGF and EGFR might be more effective than either approach alone.101 In addition to hypoxia. which led the Food and Drug Administration (FDA) to approve the VEGF monoclonal antibody bevacizumab in combination with standard chemotherapy for previously untreated. Formidable obstacles to developing effective markers include tumor heterogeneity. metaplasia. nodal status. mol ecul a r profil ing technical advances Molecular profiling. Progress in the identification of markers. and microenvironmental effects. 1374 of m e dic i n e and redundancy of tumor-cell signaling networks involving genetic. or other clinical measures) and predicted the outcome in patients with stage IA tumors. mutations.96 Factors associated with increased tumor angiogenesis correlate with the development and prognosis of lung cancer. methylation.13  www. another recently developed gene-expression profile (metagene) predicted clinical outcome with an accuracy of 72% and 79% (greater than that for tumor stage. advanced non–smallcell lung cancer. tumor diameter. VEGF has recently been validated as a therapeutic target on the basis of the results of a phase 3 trial. such as hypoxia-induced factor (HIF) 1α and 2α. VEGF and other angiogenic factors are also regulated by EGFR through HIFdependent and independent mechanisms102 and by oncogenes such as KRAS and p53. Gene-expression profiles that are associated with subtypes of non– small-cell lung cancer108. the highly complex interplay between the environment and host and the complexity.97-99 Circulating VEGF levels may predict the clinical benefit of VEGF inhibitors in patients with this disease. Squamous-cell carcinoma of the lung will be one of the first tumors profiled by this atlas. messenger RNA (mRNA). All rights reserved. and a distinctive pattern known as angiogenic squamous dysplasia can occur. For personal use only.112 Randomized. Many angiogenic factors are regulated at least in part through the hypoxia-regulated pathways. and protein or phosphoprotein signaling networks should help address these obstacles (Fig. epigenetic.111 microarray and reverse-transcriptase– polymerase-chain-reaction (RT-PCR) analyses identified a molecular signature of five genes as an independent predictor of relapse-free and overall survival. 2008 The New England Journal of Medicine Downloaded from nejm. 2014.95 Bronchial  september 25. . to guide treatment may improve the clinical outcome in patients with non– small-cell lung cancer (Fig.

g. 1375 . gene-expression signatures have been developed for cisplatin and pemetrexed on the basis of in vitro sensitivity. No other uses without permission. For personal use only. epiTitle gene-expression.g. but emerging techniques permit global analyses of the genomic. angiogenic factors Circulating DNA or RNA Circulating cells (e.g.. Molecular profiling approaches for development of personalized genetic. the cisplatin in vitro signature predicted the likelihood of response. local environment Proteins IHC Proteomics Figure 4. and multiplex beads) to assess intracellular AUTHOR PLEASE NOTE: sue and cytokines and angiogenic factors in blood.. Predictive markers identify groups of patients who are likely to have increased sensitivity Author Herbst a given therapy. All rights reserved. 4). bisulfite sequencing. HER2 for breast cancer). lymphocytes) Figure has been redrawn and type has been reset Please check carefully and the tumor and local environment (e.g. a critical step in personalizing treatment. These methods therapy include single-nucleotide polymorphism (SNP) arrays to assess genomic alterations.. For the majority of patients with advanced or metastatic non–small-cell lung cancer.116. Molecular-Profiling Approaches to the Development of Personalized Therapy.113 MicroRNA has recently emerged as an important regulator of gene expression. reverse-phase protein arrays. All markers that are involved in profiling lung cancerCOLOR canFIGURE apply to the 9/4/08 to Draft 6 or resistance tumor or its local environment. and therapy Tumor sensitivity to therapy Personalized medicine Gene expression Expression arrays MicroRNA profiling PCR-based approaches Tumor.13  www. microarrays for assessing gene expression or microRNA levels. It has been traditional to assess individual genetic orFigprotein prognostic or 4 # predictive markers (e. Many exciting potential predictive markers have been developed in vitro and need validation in tumor samples and clinical trials. Recently developed in vitro profiles predicting the sensitivity of tumors to EGFR inhibitors and other therapies have yet to be assessed clinically. including markers in blood and in tumor or nonmalignant lung tissue. andME methylation-specific polymerase chain reaction (PCR) to assess epigenetic changes. a putative regulator of EGFR that is located on chromosome 3p. DE Schwartzand proteomArtist signaling Knoper in tumor tisic methods (such as mass spectroscopy. 2014.. circulating tumor cells and tumor-derived cytokines) (red arrows).116 Loss of microRNA-128b.. SNP arrays) Blood-based profiling Proteomics on November 18. with stage IA tumors who are undergoing adjuvant chemotherapy. has been shown to correlate with the n engl j med 359. 2008 The New England Journal of Medicine Downloaded from nejm.Molecular Origins of Cancer Host Germ-line genetic profiling (e.113 For example.nejm. Blood-based profiling includes markers derived from the host (  september 25.g. High-throughput analyses have shown that microRNA expression is commonly deregulated in lung and other cancers. IHC denotes immunohisIssue date 9/25/08 tochemical analysis. tumor. investigators recently identified a five-microRNA signature that is associated with treatment outcome. Host profiling involves innate characteristics of the cancer patient. endothelial. tumor.117 Using real-time RT-PCR. the most important potential effect of molecular markers is likely to be in predicting the response to specific therapies with the goal of “personalizing” treatment (Fig. lymphocytes) Genetic or epigenetic Mutation Deletion Amplification Translocation Methylation Interactions involving the host. and protein profiles of the host (innate). Copyright © 2008 Massachusetts Medical Society.

and levels of this DNA are associated with a poor progno­ sis. C onclusions The molecular origins of lung cancer lie in complex interactions between the environment and host genetic susceptibility. including deregulated signaling pathways. Copyright © 2008 Massachusetts Medical Society. which are potential targets for chemoprevention and therapy. 2014.132 and treatment sensitivity133. allowing repeated measurements during treatment without the need for tumor tissue. All rights reserved. Furthermore. Such techniques are feasible in small amounts of tumor tissue. and proteomic profil­ ing92. epigenetic.19-21 prognosis. and KRAS and EGFR mutations have also been detected in the blood of patients with lung can­ cer.123 These studies suggest that blood profiling may provide useful information about genetic changes in tumors that could ultimately help detect lung cancer and guide therapy.125.124 Furthermore. Protein Profiling Profiling of genomic and mRNA expression provides an incomplete picture of the heterogeneity of non–small-cell lung cancer.134 and somatic genetic alterations that occur in lung adenocarcinomas14. Levels of mRNA do not always correlate with protein levels and do not provide information on protein–protein interactions or post-translational modifications such as phosphorylation that may be critical for regulating protein activity.126. Immunohistochemical analysis remains the most widely applied method for assessing individual proteins and may be useful for estimating prognosis and predicting the response to ther­ apy. geneexpression.128 and patients with better outcomes from those with worse outcomes after treatment with EGFR tyrosine kinase inhib­ itors.131 could revolutionize clinical approaches across the spectrum of lung-cancer types and subtypes by identifying practical molecular markers of risk (in precancer). have the potential to view signal transduction networks more globally than is possible with immunohistochemical analysis.3.125 Proteomic signatures for prog- 1376 of m e dic i n e nosis and predicting the response to chemotherapy or EGFR inhibitors have been developed in tumors and cell lines.nejm. Lung cancer then evolves through genetic and epigenetic changes.117 Studies suggest that information about tumorspecific genetic and epigenetic changes also may be obtained from the blood of patients with lung cancer.127 Proteomic profiling from blood is also under study. Such techniques appear to be more sensitive than those for capturing circulating DNA.114. .130. on November 18. Circulating DNA can be detected in the plasma and serum of such patients.29 Molecular targeted research has produced the recently FDA-approved EGFR and VEGF inhibitors erlotinib and bevacizumab.118.26 Emerging high-throughput proteomic techniques.28.120-122 New techniques for capturing circulating tumor cells allow the detection of EGFRactivating mutations and the drug-resistance allele T790M. protein-based profiling is likely to be essential in understanding the complexity of protein signaling networks and developing molecular signatures that predict a response to therapy. This approach is being used in developing predictive markers in non–small-cell lung cancer. No other uses without permission. such as mass spectrometry and protein microarrays.25. For personal use only.129 New techniques also permit the multiplex analysis of dozens of cytokines and angiogenic factors in small amounts of serum or plasma. promoter methylation. which have modestly improved the outcome in patients with non–smallcell lung cancer. blood. and treatment sensitivity (in early-stage and advancedstage cancer). Although promising.and tissuebased proteomic approaches remain investigational and await prospective testing and validation in large.103 Molecular profiling of the type described in this review has begun in clini- n engl j med 359.119 Tumor-specific DNA alterations (such as loss of heterozygosity). randomized trials before they can be applied clinically. a decline in the number of circulating tumor cells was associated with tumor response on radiography.48. Emerging techniques for genomic.The n e w e ng l a n d j o u r na l response to EGFR inhibition in patients with lung cancer. most targeted therapeutic agents are designed to inhibit the activity of proteins such as tyrosine kinases. Genomewide and other molecular assessments are helping elucidate germ-line variations that may contribute to lung cancer risk.13  www.50-52 and in high-risk lung tissue associated with tumors or in smokers. 2008 The New England Journal of Medicine Downloaded from nejm. Serum mass spectrometry profiles can distinguish patients with non–small-cell lung cancer from normal controls124. early detection and prognosis (in early-stage cancer).org  september 25.

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