Abstract B03: Suppression of lung adenocarcinoma growth and metastasis by stromal Hedgehog pathway activation

Abstract

Abstract Aberrant activation of the Hedgehog (Hh) signaling pathway drives the tumor growth of Gorlin-type cancers, but its role in other epithelial cancers, including non-small cell lung cancer (NSCLC), has been controversial. We used human lung adenocarcinoma (LAD) cell lines and autochthonous lung adenocarcinoma mouse models to elucidate the pathway’s role in lung cancer pathology. SHH and IHH is expressed heterogeneously among >35 tested LAD cell lines, although mostly in mutant Kras cells. SHH secreted from lung cancer cells activated the pathway in co-cultured fibroblasts in a paracrine, rather than autocrine, manner. Treatment with KAAD-cyclopamine, an Hh pathway antagonist, inhibited the growth of high SHH-expressing tumor cells when co-cultured with NIH-3T3 embryonic fibroblasts but not tumor cells alone, further reinforcing the paracrine nature of Hh pathway activation. However, the growth effect due to fibroblast pathway inhibition was diminished when co-cultured with normal lung fibroblasts. Analysis of potential Hh pathway target genes suggest that normal lung fibroblasts secrete ligands that induce a differentiation program in epithelial cells rather than ligands that induce mitogenic programs from NIH-3T3 fibroblasts. In KrasLSL-G12D/+;Trp53fl/fl; Shhfl/fl \(KPS) mice, genetic loss of SHH, surprisingly, had no effect on survival compared to KrasLSL-G12D/+;Trp53fl/fl;Shh+/+ (KP) mice. However, treatment with 5E1, an anti-SHH/IHH blocking antibody, 2 weeks after tumor induction in KP mice resulted in significantly worse survival with increased metastases compared to KP mice treated with control IgG1 antibody. Furthermore, stromal Hh pathway abrogation by 5E1 led to increased tumor burden and decreased tumor vessel density compared to control mice. To account for the survival differences seen between KP mice with genetic SHH loss and those treated with 5E1, we analyzed KP tumors and found unexpectedly high levels of Ihh mRNA, compared to Shh mRNA, that have not been reported previously in normal or neoplastic murine lungs. Preliminary results using an in vivo CRISPR/Cas9 (pSECC) system to genetically ablate Ihh in KrasLSL-G12D/+;Trp53fl/fl;Rosa26LSL-fluc/+ mice showed significant enhancement of tumor growth compared to control mice treated with sgGFP. These results suggest that IHH, rather than SHH, activates a tumor-suppressive program in lung stroma. These results correlate to human survival data of LAD with low lhh mRNA expression (KMPLotter). We are in the process of isolating and examining murine stromal cells from KP mice to identify the secreted stromal factors that limit tumor growth and metastases. Thus, stromal Hh pathway activation, in response to IHH secreted from lung cancer epithelia, suppresses LAD growth and metastases, increases tumor vessel growth, and prolongs survival. Citation Format: Sahba Kasiri, Baozhi Chen, Alexandra Wilson, Annika Reczek, Ummay Marriam, Zhiqun Zeng, Luc Girard, James Kim. Suppression of lung adenocarcinoma growth and metastasis by stromal Hedgehog pathway activation [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr B03.