Using the KrasLA1/+; P53R172H?G/+ (KP) mouse model, which simulates NSCLC, we compared cell lines derived from non-metastatic versus highly metastatic tumors. This identified upregulation of Msi2, an RNA-binding protein that regulates mRNA translation, as one of the most consistent features of metastatic cells. In an initial probe of 123 primary human NSCLC specimens, it has been found that Msi2 is significantly elevated in tumors versus normal lung epithelium, suggesting relevance to NSCLC in patients. Msi2 knockdown in four independent murine and human metastatic NSCLC cell lines decreased invasion in vitro, with preliminary confirmation of reduced metastasis in vivo. Candidate pathway analysis and reverse-phase protein array (RPPA) screening identified EMT-associated proteins including the TGF-? receptor Type I (TGF-?RI), the Notch inhibitor Numb, fibronectin (FN1), and claudin-7, as strongly regulated by Msi2. The investigators hypothesize that Msi2 regulation of these proteins is critical for its role in invasion and metastasis, and provides essential support for Notch- and TGF-?RI dependent oncogenic signaling in a subset of metastatic NSCLC. The objective is to explore these mechanisms in vitro, determine whether Msi2 expression regulates the response to drugs targeting Notch and TGF-?RI, and determine whether the relationships we have identified predict pathway activity in human tumors.
Aim 1 will validate functional significance of Msi2 dependent signaling effectors, focusing on interconnection between NUMB/Notch, TGF-?, and novel targets such as the cell junction regulator claudin-7. This work will emphasize the role of Msi2 expression on response to ?- secretase (Notch-targeting) and TGF-?RI inhibitors, and will combine in vitro analysis of signaling and drug response with xenograft analysis.
Aim 2 will establish if expression of Msi2 expression correlates with Notch, TGF-?, CLDN7, and E-cadherin, and clinical data in cancer patients. In complementary experiments, the researchers will use a conditional 129S/Sv-Krastm3Tyj/J; Trp53tm1Brn/J; Msi2-/- mouse model to determine the action of Msi2 at discrete stages of lung tumorigenesis, including early growth, invasion, and metastasis.
A functional role for Msi2 has not previously been identified in NSCLC, so this work will provide an entirely novel source of insight into NSCLC progression and invasion. I believe that characterization of Msi2 will lead to better understanding of what drives lung cancer, and help development of future individualized treatment strategies for lung cancer patients, particularly if Msi2 expression conditions therapeutic response to ?- secretase and TGF-? receptor Type I kinase (TGF-?RI) inhibitors. We would also develop the first in vivo model of specific loss of Msi2 in the lung and perform initial evaluation of the consequences of Msi2 loss on tumor formation.
