**4.3 BRAF**

BRAF is a proto-oncogene that encodes a serine/threonine protein kinase that is a downstream effector protein of RAS and it transduces the signal through the mitogenactivated protein kinase pathway, which promotes cell proliferation and survival. Phosphorylation of downstream mediators MEK1 and MEK2 occurs on activation of BRAF, which subsequently activates ERK1 and ERK2, which take part in the regulation of growth-regulating proteins such as c-JUN and ELK1. BRAF-activating mutations lead to an increase in the kinase activity that exhibits transforming activity in vitro.

BRAF mutations are most commonly noted in melanoma, and they can also occur in about 3% cases of NSCLC. The mutations seen in NSCLC differ from those *Perspective Chapter: Molecular Pathology of Lung Cancer DOI: http://dx.doi.org/10.5772/intechopen.109598*

in melanoma and colorectal carcinoma. In lung adenocarcinoma, V600E mutations in exon 15 are most commonly seen accounting for 50% of BRAF mutations, which are followed by G469A in exon 11 and D594G in exon 15. In NSCLC BRAF mutations occur either in the kinase domain (such as V600E, D594G, and L596R) or in the G-loop of the activation domain of the gene (such as G465V and G468A). BRAF and EGFR mutations are mutually exclusive. Non-V600E BRAF mutations are observed in current or former smokers, while V600E mutations are more common in female never smokers. BRAF mutations act as an important therapeutic target in NSCLC [14, 21, 30, 31].

#### **4.4 MEK**

MEK1 (also known as MAPK1) is a serine–threonine kinase that acts as an important downstream target of RAS activation. MEK1 further activates MAPK2 and MAPK3 that are downstream of BRAF. In lung adenocarcinoma, somatic mutations of MEK1 are reported rarely and they are mostly activating mutation in exon 2 [27, 32].

#### **4.5 Met**

MET is located on chromosome 7q21-q31 and acts as a protooncogene that encodes a membrane tyrosine kinase receptor that is also known by the name hepatocyte growth factor receptor. Hepatocyte growth factor binds to its ligand, there occurs a sequence of events which are homodimerization of receptor, activation of kinase, and downstream signaling through PI3K/AKT/RAS/RAF/MEK/MAPK and c-SRC kinase pathways. Gene amplification of MET is observed in NSCLC. MET copy number is increased more commonly in SCC than ADC and is mutually exclusive with KRAS mutations. Amplification of MET results in overexpression of MET protein and thus activation of downstream signaling pathways. MET amplification is a known mechanism of secondary EGFR-TKI resistance. In this scenario, the amplification of MET drives and maintains the PI3K/ AKT pathway bypassing the EGFR blockade by TKIs, which suggests concomitant MET inhibition may be a way of overcoming TKI resistance [33–36].

#### **4.6 HER-2**

The human epidermal growth factor receptor 2 (HER2/ERBB2) is a part of the ERBB family of receptors which encodes a membrane-bound receptor tyrosine kinase. Unlike other members of ERBB receptors, it does not bind directly to the ligand but can form heterodimers with other ligand-bound members of the receptor family. On activation, there occurs signaling through PI3K, MAPK, and JAK/STAT pathways. HER2 activation is seen in only a few cases of lung cancers with 20% of the case showing overexpression, gene amplification in 2%, and activating mutations in 1.6–4% of NSCLC. HER2 mutations are most commonly observed in adenocarcinoma and this mutation occurs mostly in tumors that are wild type for EGFR and KRAS and they are observed in the female gender, Asian ethnicity, and non-smokers, which is similar to the clinical profile of EGFR mutant tumors [37–41].

#### **4.7 ROS1**

ROS1, a proto-oncogene, belongs to receptor tyrosine kinase of the insulin family receptor and is located on chromosome 6q22. The rearrangement was initially described in glioblastoma involving the ROS1-FIG gene fusion.

More recently, ROS1 fusions were identified as potential driver mutations in an NSCLC cell line (HCC78; SLC34A2-ROS1) and an NSCLC patient sample (CD74- ROS1). These fusions result in tyrosine kinase activation, although the details of the downstream signaling transduced by ROS1 fusion are not fully understood yet.

Patients with ROS1 rearrangements are significantly younger, more likely to be never smokers, and overrepresented in the Asian race. Also, ROS-positive lung cancer patients are associated with sensitivity toward TKIs, specifically crizotinib, with a patient demonstrating prompt and durable complete response to therapy [42–44].

#### **4.8 Ret**

It is a proto-oncogene that encodes for receptor tyrosine kinase and is located on chromosome 10q11.2. The chromosomal rearrangements involving RET gene and kinesin family 5B (KIF5B) & coiled-coil domain containing-6 (CCDC6) resulting in KIF5B-RET and CCDC6-RET fusion genes have been identified in 70 to 90% and 10 to 25% of lung tumors, respectively.

As a result of chromosomal rearrangements, there is overexpression of RET protein. The RET fusion is found in 1–2% of NSCLC, particularly in younger, nonsmoking patients with adenocarcinoma histology, and is associated with increased risk of brain metastases and patients with RET fusions show minimal response to immunotherapy [43–46].

#### **4.9 ALK**

ALK encodes for receptor tyrosine kinase. Mutation of the ALK gene is associated with 4% of unselected NSCLC approximately. EML4-ALK fusion gene that is found in a subset of lung cancers occurs because of chromosomal rearrangements forming EML4– ALK fusion gene that is involved in the activation and upregulation of RAS/RAF1/ MAP2K1/MAPK1 pathway that is involved in cell proliferation as well as cell survival.

Lung cancers associated with ALK rearrangements are more common in young patients with male dominance who are more commonly non-smokers or light smokers. These tumors show typical histology characterized by the presence of mucin as well as the solid pattern of tumor growth comprising of signet cells in the Western population or acinar growth pattern in Asian patients. They are commonly diagnosed at an advanced stage of clinical presentation. The response rate to chemotherapy as well as the overall survival rate in patients with ALK rearrangement is prompt and comparable [42, 47–50].

#### **4.10 DDR2**

The DDR2 gene is a tyrosine kinase receptor, present on the long arm of chromosome 1 (1q23.3). It is involved in cell proliferation, survival, and migration by promoting matrix metalloproteinase expression. DDR2 mutation is associated with 3.8% of cases of squamous cell carcinoma. DDA2 mutations have also been observed in a few cases of NSCLC. It acts as an oncogene and its over expression promotes cell survival and proliferation in SCC lungs [51–53].

#### **4.11 FGFR**

FGFR belongs to the receptor tyrosine kinase family and is one of the most promising predictive biomarkers in lung cancer. There is overexpression and gain of function *Perspective Chapter: Molecular Pathology of Lung Cancer DOI: http://dx.doi.org/10.5772/intechopen.109598*

mutation of FGFR in lung cancer. Fibroblast growth factors bind with FGFR causing the activation and upregulation of JAK–STAT/MAPK/PI3K-AKT pathway causing cell proliferation, angiogenesis, differentiation, and survival. Multiple FGFR aberrations are present in Sq-NSCLC tumors—alterations (mutations and fusions), amplification, and mRNA/protein over-expression—but their predictive potential is unclear. FGFR1 amplification is seen in 22% of lung squamous cell carcinoma patients. Mutations are more common in non-smokers than smokers, with more advanced TNM stages. FGF mutations in small-cell carcinoma lung is associated with poor prognosis [54–56].

#### **5. Tumor suppressor genes**

#### **5.1 TP53**

TP53 (Tumor protein 53) is a tumor suppressor gene located on chromosome 17p13. It encodes a protein containing DNA binding, transcriptional activation, and oligomerization domains. The encoded protein regulates the expression of certain target genes involved in cell cycle arrest, DNA repair, or metabolic changes. It also regulates the expression of genes engaged in promoting growth arrest in the G1 phase or cell death in response to genotoxic stress. P53 is thus known as "the guardian of the genome." P53 prevents the damaged cells from undergoing mitosis. On entering the G2 phase, p53 blocks cells at the G2 checkpoint, by inhibiting the cyclin-dependent kinase required to enter mitosis. This ability of p53 to inhibit cellular proliferation or induce apoptosis is suppressed by the HDM2 protein product. There is proteosomedependent degradation and downregulation of p53 expression caused by HDM2 protein. Also, p53 itself causes the activation of HDM2 protein by binding directly with HDM2 protein resulting in the upregulation of HDM2. Thus, p53 downregulates its own expression, and as a result of which, p53 levels in normal cells are merely detectable because of this autoregulatory mechanism. Damaged DNA inducesTP53 activation leading to cell cycle arrest by inducing the expression of cyclin-dependent

**Figure 4.** *TP53 pathway.*

kinase inhibitors, which may cause DNA repair or apoptosis (**Figure 4**). One of the most common abnormalities noted in lung cancer is the inactivation of TP53 with a hemizygous deletion of 17p13, containing the locus of TP53, which accounts for 90% of mutations of small-cell carcinomas and about 65% of NSCLC. Missense mutations involving TP53 have been known to be associated with 80–100% of small-cell lung carcinomas. In NSCLC mutations or proteins, accumulation has been known to occur more commonly in SCC than in ADC. Also, mutations of TP53 were found in at least 81% of SCCs in a meta-analysis by the cancer genome atlas (TCGA). Exposure to tobacco and smoking has also been noted to be associated with the varied nature of the mutation spectrum as smoking-associated cancers have a higher propensity of G to T transversions compared to G to C transversions due to the presence of polycarbonates associated with tobacco smoke. Also, G to A transitions at CpG dinucleotides are more commonly seen in never smokers. Mutations of EGFR and KRAS can also occur in association with TP53, and also, loss of function mutation of TP53 has been associated with poor response to treatment [57–64].

#### **5.2 LKB1 (STK11)**

The LKB1 is a tumor suppressor gene located on chromosome 19p13.3. It was thought to be involved as the causative agent behind Peutz-Jeghers syndrome through a germline-inactivating mutation. LKB1 mutation is typically rare in most types of cancer, except pancreatic cancer and NSCLC. It was found that LKB1 possesses inactivating mutations in NSCLC tumors. Inactivation of LKB1 is known to be more common in adenocarcinomas than in squamous cell carcinomas at a rate of 34 and 19%, respectively. LKB1 in lung cancer may be inhibited by a variety of somatic mutations or deletions that produce truncated proteins causing inactivation of LKB1in about 11–30% of lung ADC. It is thought to be the third commonest genetic aberration in lung ADC after TP53 and KRAS. There are few studies in the literature supporting the association between LKB1 mutations and a history of smoking in men. Also, a correlation with KRAS mutations has been reported [65–67].

#### **5.3 PTEN**

PTEN, a tumor suppressor gene, is located on the long arm of chromosome 10 and encodes for lipid and protein phosphatases. This phosphatase causes the dephosphorylation of PIP3 as a result of which there is downregulation in the expression of PI3K/ AKT/mTOR signaling pathway. Thus, the loss of function mutation of PTEN results in unrestricted upregulation and activation of signaling pathways causing uncontrolled cell proliferation, resulting in tumor mass. Some lung cancers are associated with PTEN mutations or deletions. PTEN mutations are more common in smokers than non-smokers, present in 10.2% cases of SCC lung as compared to 1.7% cases of adenocarcinoma lung. Overall, PTEN mutations are seen in 5% cases of NSCLC, in which 75% cases of NSCLC show reduced protein expression [68, 69].

#### **5.4 PI3K**

The PI3K/mTOR/AKT pathway is an important signal transduction pathway involved in the regulation of cell proliferation, differentiation, survival, adhesion, and motility. Both NSCLC and small-cell carcinoma have been known to be associated with alterations in this pathway. A variety of membrane tyrosine kinase receptors

*Perspective Chapter: Molecular Pathology of Lung Cancer DOI: http://dx.doi.org/10.5772/intechopen.109598*

including EGFR, vascular endothelial growth factor receptors, insulin-like growth factor receptors, HER2, and platelet-derived growth factor receptors has been known to activate this pathway. Activation of tyrosine kinase receptors results in the phosphorylation of PIP2 (phosphatidylinositol 4, 5-bisphosphate) to PIP3 (phosphatidyl inositol 3,4,5-triphosphate) with the help of enzymes phospho inositol 3 kinase (PI3K). PI3K and mTOR further cause phosphorylation of PIP3. Downstream regulation of AKT is caused by mTOR, which is a serine/threonine kinase. Activation and upregulation of AKT result in cell proliferation and survival. Also, upregulation of other pathways like RAF/MAPK/RAS results in direct activation of PIP3 [70–73].
