Abstract B25: ASK1 is involved in tumor lung metastasis through regulation of platelet functions

Apoptosis signal-regulating kinase 1 (ASK1) is a MAP3K in the JNK and p38 MAPK pathways and responds to various stresses. Accumulating evidence indicates that ASK1 plays important roles in tumorigenesis by regulating apoptosis and inflammation. However, little is known about its role in tumor metastasis. Here we examined the roles of ASK1 in lung metastasis by intravenous injection of Lewis lung carcinoma cells constitutively expressing luciferase (3LL-Luc2). 14 days after injection of 3LL-Luc2 cells, ASK1 -/- mice showed remarkably reduced number of tumor nodules and decreased luciferase activity of the lung lysates than wild-type mice. Taken together with the data obtained by in vivo imaging system ( IVIS ), ASK1 deficiency appeared to result in the attenuation of lung metastasis in this model. Hematogenous tumor metastasis is known to consist of multiple steps such as invasion, intravasation, extravasation and colonization. We next measured the time-course-dependent luciferase activity of the lung lysates after injection from 10 minutes to 7 days in order to investigate which stages of tumor metastasis ASK1 is involved in. ASK1 -/- mice had markedly lower luciferase activity of the lung lysates than wild-type mice as early as 3 hours after injection. Moreover, ASK1 phosphorylation in lungs of wild-type mice was accelerated at that point; hence ASK1 is suggested to be involved in the very early stage called seeding stage of lung metastasis, which is the time point prior to the extravasation of tumor cells. Myeloid cells such as platelets and innate immune cells are known to participate in the seeding stage and we next investigated whether ASK1 in myeloid cells is involved in tumor metastasis by analyzing bone marrow chimeric mice. Wild-type mice with bone marrow from ASK1 -/- mice showed significant decrease in luciferase activity of the lung lysates compared to wild-type mice with bone marrow from wild-type mice, indicating that ASK1 in myeloid cells plays an important role in tumor metastasis. Among various myeloid cell types, we first focused on platelets. Platelets adhere and aggregate to surround tumor cells in the seeding stage and are known to positively regulate hematogenous tumor metastasis. Transcriptional analysis of platelets revealed that the mRNA expression of some genes related to platelet functions was reduced in ASK1 -/- mice. In addition, analysis by western blot revealed that platelets of ASK1 -/- mice exhibit markedly reduced phosphorylation of JNK and p38, both of which are well-known downstream signaling molecules of ASK1 and also are indicated to participate in platelet functions. Moreover, ASK1 -/- mice showed bleeding tendency in tail bleeding assay. These data collectively indicate that ASK1 deficiency may attenuate platelet functions. Defect in platelet functions of ASK1 -/- mice may result in attenuated interaction between platelets and tumor cells. In conclusion, ASK1 deficiency is suggested to attenuate lung metastasis through impaired platelet functions. Citation Format: Miki Kamiyama, Isao Naguro, Hidenori Ichijo. ASK1 is involved in tumor lung metastasis through regulation of platelet functions. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr B25. doi:10.1158/1538-7445.CHTME14-B25