Abstract IA06: Driver mutations and cell-of-origin as critical factors determining the phenotypic characteristics of thoracic tumor subtypes
Authors: Berns Anton , Semenova Ekaterina , Ferone Giustina , Vries Hilda de , Badhai Jitendra , Kwon Min-chul , Sutherland Kate , Bombardelli Lorenzo , Bhaskaran Rajith , Song Ji-Ying 
Published: May 2018

Abstract
We have generated mouse models for specific lung tumors and mesothelioma. These were based on the conditional somatic (in)activation of tumor suppressor genes and oncogenes. By using viruses driving Cre recombinase from specific promoters, we could achieve both sporadic and cell-type specific switching of the conditional alleles allowing us to address the importance of the target cell in the development of specific tumor subtypes. We made the following observations.
Small cell lung cancer (SCLC) development was fully dependent on the inactivation of Rb in conjunction with loss of p53. This neuroendocrine tumor could be most efficiently induced by targeting neuroendocrine cells using an Ad5-CGRP-Cre virus. However, when Rb and p53 were inactivated in combination with overexpression of L-myc, a gene frequently found amplified in both human and mouse, we noted additional tumors with a more peripheral location (more precisely at the bronchiolar-alveolar junction region) and with a less aggressive phenotype, but only when these tumors were induced by an Ad5-CMV-Cre virus. Since the latter tumors were strongly positive for neuroendocrine markers including CGRP, this indicates that the latter tumor subtype originated from a cell with no or very low CGRP expression and therefore different from the neuroendocrine cell targeted by Ad5-CGRP-Cre. The cell type giving rise to this tumor is not yet defined. Nevertheless, the different phenotypic characteristics of this tumor—that shows many of the features of SCLC—likely also have consequences for its prognosis and response to therapy.
In testing a number of different combinations of lesions to induce squamous cell carcinoma (SCC), we found that development of SCC strongly depended on the overexpression of Sox2. Biallelic inactivation of Pten and Cdkn2a;Cdkn2b;p19Arf in combination with Sox2 overexpression promoted SCC at high penetrance but after a relatively long latency period. Phenotypically mouse SCC closely resembled human SCC, showing a very similar immunophenotyped, indicating that this is largely commanded by the tumor subtype. Interestingly, SCC could be induced at the same extend by activating the aforementioned genetic lesions in Basal, Club, and Alveolar type II cells (AT2). When targeting the latter two cell subtypes, they went through a dedifferentiation/transdifferentiation process with the concomitant loss and gain of transient cell markers; e.g., when targeting AT2 cells, the cells would lose SPC expression, transiently express the Club marker CC10, to acquire subsequently the biomarker of SCC, K5, and p63. Histopathology showed a process of transdifferentiation of the alveolar cells to Club cells and eventually to neoplastic squamous cells. Although the cell-of-origin did influence the extent of progression of individual tumor nodules (more small lesions upon induction of transformation of AT2 cells), the phenotypic characteristics of the tumors were very similar and could not be distinguished on basis of RNA expression.
In the case of adenocarcinomas developing in LSL-KrasG12D;p53f/f mice, the cell-of-origin did again influence the tumor phenotype. When Club cells were targeted, the cells would lose the Club cell specific marker expression (CC10 and Sox2) and acquire pronounced SPC expression while such changes were not seen upon targeting AT2 cells. They retained their SPC expression. However, the tumors induced in Club cells showed a more pronounced papillary phenotype as well as a more aggressive growth pattern as compared to tumors induced in AT2 cells.
A similar situation was encountered in mesothelioma. Although this tumor is less stringently dependent on the specific lesions, loss of Nf2 in conjunction with p53 pathway inactivation and Cdkn2a loss is an effective route to malignant mesothelioma (MM). MM development can be further accelerated by BAP1 loss resulting in an autochthonous model that is as fast as tumor graft models. Some of the advantageous and disadvantages of each will be discussed.
MM can manifest as three major subtypes: epithelioid, sarcomatoid, and biphasic. Each of those can be induced by the same set of mutations (loss-of-function of Nf2, p53, and Cdkn2a). The tumor subtype that arises depends on the cell-of-origin. The three subtypes exhibit relative stable phenotypes with the phenotypic appearance of the biphasic tumor depending on the microenvironmental cues. Changing those makes this subtype either more epithelioid or sarcomatoid.
The examples described show that both the combination of genetic lesion and the cell-of-origin are critical factors defining tumor characteristics. In some cases, targeting different cell types with the same lesions gives rise to seemingly indistinguishable tumors (e.g., SCC), whereas in other cases the same genetic lesions cause tumor features that are strongly influenced by the cell-of-origin (as is the case in MM and SCLC), emphasizing that the cell-of-origin might be an important factor that should be taken into account when designing intervention strategies within otherwise highly similar tumors.
Citation Format: Anton Berns, Ekaterina Semenova, Giustina Ferone, Hilda de Vries, Jitendra Badhai, Min-chul Kwon, Kate Sutherland, Lorenzo Bombardelli, Rajith Bhaskaran, Ji-Ying Song. Driver mutations and cell-of-origin as critical factors determining the phenotypic characteristics of thoracic tumor subtypes [abstract]. In: Proceedings of the AACR Special Conference: Advances in Modeling Cancer in Mice: Technology, Biology, and Beyond; 2017 Sep 24-27; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(10 Suppl):Abstract nr IA06.