**5. Mouse models for squamous cell lung cancer (SqCLC)**

So far genomic alterations in SqCLC have not been comprehensively characterized. The Cancer Genome Atlas group recently profiled 178 lung squamous cell carcinomas to provide a comprehensive view of genomic and epigenomic alterations (Hammerman *et al.*, 2012). They showed that the SqCLC is characterized by hundreds of exonic mutations, genomic re‐ arrangements, and gene copy number alterations. In addition to *TP53* mutations found in nearly all specimens, loss-of-function mutations were found in the *HLA-A* class I gene. In addition, *Nuclear factor (erythroid-derived 2)-like 2*, *Kelch-like ECH-associated protein 1*, *Squamous differentiation*, and *Phosphatidylinositol-3-OH kinase pathway* genes were frequently altered. *CDKN2A* and *RB1* genes were inactivated in as many as 72% of SqCLC cases. This compre‐ hensive study identified a potential therapeutic target in most tumors, offering new avenues of investigation for the treatment of human SqCLC (Hammerman *et al.*, 2012).

Although squamous cell carcinoma is a common type of lung cancer causing nearly 400,000 deaths per year worldwide, there is no established gene-engineered mouse model for squa‐ mous cell carcinoma of the lung. Human lung SqCLC is closely linked with smoking and shows a distinct order of pre-malignant changes in the bronchial epithelium from hyperpla‐ sia, metaplasia, dysplasia and carcinoma *in situ* to invasive and metastatic SqCLC (Brambilla *et al*., 2000). A better understanding of the cell of origin that give rise to SqCLC and identifi‐ cation of unique genetic alterations that are specific to lung squamous cell carcinoma as re‐ ported by the comprehensive study might help to create SqCLC mouse models. One important issue that should be taken into account is that normal human or mouse lungs do not contain squamous epithelium. Mice do not smoke, so only under pathological condi‐ tions does squamous metaplasia accompanied by high expression levels of keratins occur in the airway epithelium (Wistuba *et al*., 2002, 2003). Only a few mouse models reported the onset of SqCLC, mostly after carcinogen application. For instance, intratracheal intubation of methyl carbamate (Jetten *et al*., 1992) or extensive topical application of *N*-nitroso-com‐ pounds (Nettesheim *et al*., 1971; Rehm *et al*., 1991) caused SqCLC in mice. Wang *et al.* (2004) treated eight different inbred strains of mice with N-nitroso-tris-chloroethylurea by skin painting and found that this chemical induced SqCLCs in five strains (SWR, Swiss, A/J, BALB/c, and FVB), but not in the others (AKR, 129/svJ, and C57BL/6). Besides, specific loci for SqCLC susceptibility have been identified through linkage analyses in several mice strains (Wang *et al.*, 2004), using 6,128 markers in publically available databases. Three markers (*D1Mit169, D3Mit178*, and *D18Mit91*) were found significantly associated with sus‐ ceptibility to SqCLC. Interestingly, none of these sites overlapped with the major suscepti‐ bility loci associated with lung adenoma/adenocarcinomas in mice indicating that different sets of genes are responsible for SqCLC and AdCA. Their model can be used in determining genetic modifiers that contribute to susceptibility or resistance to SqCLC development.

healthy volunteers with high sensitivity, specificity (Hennessey *et al.,* 2012). Upon further testing on additional 130 subjects, this miRNA pair predicted NSCLC with a specificity of 84%, sensitivity of 100%. These data provide evidence that serum miRNAs have the poten‐

So far genomic alterations in SqCLC have not been comprehensively characterized. The Cancer Genome Atlas group recently profiled 178 lung squamous cell carcinomas to provide a comprehensive view of genomic and epigenomic alterations (Hammerman *et al.*, 2012). They showed that the SqCLC is characterized by hundreds of exonic mutations, genomic re‐ arrangements, and gene copy number alterations. In addition to *TP53* mutations found in nearly all specimens, loss-of-function mutations were found in the *HLA-A* class I gene. In addition, *Nuclear factor (erythroid-derived 2)-like 2*, *Kelch-like ECH-associated protein 1*, *Squamous differentiation*, and *Phosphatidylinositol-3-OH kinase pathway* genes were frequently altered. *CDKN2A* and *RB1* genes were inactivated in as many as 72% of SqCLC cases. This compre‐ hensive study identified a potential therapeutic target in most tumors, offering new avenues

Although squamous cell carcinoma is a common type of lung cancer causing nearly 400,000 deaths per year worldwide, there is no established gene-engineered mouse model for squa‐ mous cell carcinoma of the lung. Human lung SqCLC is closely linked with smoking and shows a distinct order of pre-malignant changes in the bronchial epithelium from hyperpla‐ sia, metaplasia, dysplasia and carcinoma *in situ* to invasive and metastatic SqCLC (Brambilla *et al*., 2000). A better understanding of the cell of origin that give rise to SqCLC and identifi‐ cation of unique genetic alterations that are specific to lung squamous cell carcinoma as re‐ ported by the comprehensive study might help to create SqCLC mouse models. One important issue that should be taken into account is that normal human or mouse lungs do not contain squamous epithelium. Mice do not smoke, so only under pathological condi‐ tions does squamous metaplasia accompanied by high expression levels of keratins occur in the airway epithelium (Wistuba *et al*., 2002, 2003). Only a few mouse models reported the onset of SqCLC, mostly after carcinogen application. For instance, intratracheal intubation of methyl carbamate (Jetten *et al*., 1992) or extensive topical application of *N*-nitroso-com‐ pounds (Nettesheim *et al*., 1971; Rehm *et al*., 1991) caused SqCLC in mice. Wang *et al.* (2004) treated eight different inbred strains of mice with N-nitroso-tris-chloroethylurea by skin painting and found that this chemical induced SqCLCs in five strains (SWR, Swiss, A/J, BALB/c, and FVB), but not in the others (AKR, 129/svJ, and C57BL/6). Besides, specific loci for SqCLC susceptibility have been identified through linkage analyses in several mice strains (Wang *et al.*, 2004), using 6,128 markers in publically available databases. Three markers (*D1Mit169, D3Mit178*, and *D18Mit91*) were found significantly associated with sus‐ ceptibility to SqCLC. Interestingly, none of these sites overlapped with the major suscepti‐ bility loci associated with lung adenoma/adenocarcinomas in mice indicating that different

tial to be sensitive, cost-effective biomarkers for the early detection of NSCLC.

of investigation for the treatment of human SqCLC (Hammerman *et al.*, 2012).

**5. Mouse models for squamous cell lung cancer (SqCLC)**

46 Oncogenesis, Inflammatory and Parasitic Tropical Diseases of the Lung

The other group tried to induce SqCLC through constitutive expression of human K14 by creating *CC10-hK14* mice (Dakir *et al*., 2008). Although hK14 is highly expressed in bronchial epithelium, only precursor lesions varying from hyperplasia to squamous metaplasia were observed (Dakir *et al*., 2008). Clearly, the increased K14 expression and onset of squamous cell metaplasia alone was not sufficient to generate fully developed SqCLC. As far as trans‐ genic/knockout mice models are concerned, only the *LSLKrasG12D;Lkb1flox/flox* somatic mouse model has been able to generate advanced SqCLC. By using a somatically activatable mu‐ tant *Kras*-driven model of mouse lung cancer (*K-rasLA*), Ji *et al.* (2007) compared the role of Lkb1 to other tumor suppressors in lung cancer. Although *Kras* mutation cooperated with loss of *p53* or *Ink4a/Arf* in this system, the strongest cooperation was seen with homozygous inactivation of *Lkb1*. *Lkb1*-deficient tumors demonstrated shorter latency, an expanded histo‐ logical spectrum (adeno-, squamous, and large-cell carcinoma) and more frequent metasta‐ sis as compared to tumors lacking *p53* or *Ink4a/Arf*. Interestingly up to 60% of *Lkb1* deficient lung tumors had squamous or mixed squamous histology (Ji *et al*., 2007), which has not been reported in other mouse lung cancer models. Pulmonary tumorigenesis was also accelerated by hemizygous inactivation of *Lkb1*, confirming its haplo-insufficiency. Consistent with these findings, inactivation of *LKB1* was found in 34% and 19% of 144 human lung adeno‐ carcinomas and squamous cell carcinomas, respectively. They also identified a variety of metastasis-promoting genes, such as *NEDD9*, *VEGFC* and *CD24*, as targets of LKB1 repres‐ sion in lung cancer. These studies established LKB1 as a critical barrier to prevent lung carci‐ nogenesis, controlling initiation, differentiation and metastasis (Ji *et al*., 2007).
