Abstract 164: Oncogenic PKC-ι/PKC-ζ/14-3-3 and PKC-ι/PKC-ζ/Smad2/3 signaling cascades are crucial for increased invasiveness of Neuroblastoma cells

Abstract

Abstract Neuroblastoma (NB) is a the pediatric cancer of the early nerve cells in which solid tumors originate from the peripheral sympathetic nervous system and as such can occur primarily anywhere along the sympathetic chain with development on the adrenal gland being most common. The expression of the disease is highly variable as in some cases lead to spontaneously regression without any medical intervention whereas in other cases the tumors become highly metastatic and therapeutically resistant. Our results demonstrated that atypical protein kinase C-iota and zeta (aPKC-ι/ζ) levels were overexpressed in NB cells (BE-2C and BE-M17) and tissues. The present study shows the downstream effects PKC-ι specific inhibitor 5-amino-1-(2,3-dihydroxy-4-(hydroxymethyl)cyclopentyl)-1H-imidazole-4-carboxamide (ICA-1S) and a PKC-ζ specific inhibitor 8-hydroxy-1,3,6-naphthalenetrisulfonic acid (ζ-Stat) on aPKCs, 14-3-3, p-14-3-3, Akt1, NF-κB p65, Smad2/3 and p-Smad2/3. Decreased Vimentin and increased E-cadherin levels indicated the downregulation of epithelial-mesenchymal transition (EMT). Result also indicated that SNAIL1 transcriptional activity was downregulated as a result of aPKC diminution. SNAIL1 is known to induce EMT. 14-3-3 proteins have the ability to bind a multitude of functionally diverse signaling proteins, including kinases, phosphatases, and transmembrane receptors. Our immunoprecipitation (IP) data suggested that both aPKCs associate with 14-3-3 and Smad2/3. Our data also indicated that both 14-3-3 and Smad2/3 levels were significantly decreased as a result of aPKC inhibition. In addition, pIκB, NFκB p65, p-14-3-3, pAkt1, levels were significantly decreased. Our data indicates that 14-3-3 acts as a central key regulator in aPKC driven cell cycle progression in BE-2C and BE-M17 cells. In addition, we found that Smad2/3 plays a vital role in upregulation of aPKC driven Vimentin dynamics in NB cell lines in relation to 14-3-3. In-vitro migration/invasion assays, IP, immunofluorescence, real time qPCR and Western blot techniques were used to further analyze role of aPKCs in NB malignancy. Data confirmed that aPKC attenuation triggers multiple pro-apoptotic signaling cascades through downregulation of NFκB, Akt, and Smad pathways through downregulation of 14-3-3. In-vivo experiments on murine models are being conducted using ICA-1S and ζ-Stat. Excised tumors will be analyzed for pathways observed in in-vitro experiments. Overall results suggest that aPKCs are crucial for increased invasiveness in NB. Results suggested that aPKC can be targeted to develop customized, tailored therapies for NB which merit further research. Citation Format: Sloan Breedy. Oncogenic PKC-ι/PKC-ζ/14-3-3 and PKC-ι/PKC-ζ/Smad2/3 signaling cascades are crucial for increased invasiveness of Neuroblastoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 164.