Abstract C78: ATR-FANCD2 dependent JAK-STAT pathway up-regulates Procaspase-8 and Bax in Coralyne-UVA induced cancer cell apoptosis.

Abstract

Abstract Previously we have shown a synergism between the coralyne, a protoberberine molecule and UVA light i.e., CUVA treatment in photosensitizing multiple cancer cell lines. Mechanistically, the CUVA treatment dramatically enhanced DNA damage and apoptosis in cancer cells through the activation of ATM-CHK2 mediated S-phase arrest1. Here we explored the putative molecular machinery responsible for CUVA induced cancer cell apoptosis. In order to carry out this mechanistic study, we have used basic cell culture techniques along with flow cytometry based applications, immunoblotting, siRNA technique and chromatin immunoprecipitation etc. Our result showed that CUVA treatment causes a rapid and irreversible lysosomal membrane permeabilization and cytochrome C release from mitochondria. Further, we demonstrate that the perturbation in lysosomal and mitochondrial function is associated with rapid up-regulation of BAX as well as procaspase-8 and consequent cleavage of BID protein. The CUVA treatment stimulated the tyrosine phosphorylation of JAK2 and STAT1. STAT1 tyrosine phosphorylation was inhibited by a JAK2 inhibitor, AG490 as well as short-interfering RNA (siRNA) induced silencing of JAK2. Binding of tyrosine-phosphorylated STAT1 to the promoter region of procaspase-8 and its consequent expression was inhibited by AG490, suggesting that CUVA treatment up-regulates procaspase-8 expression via the JAK2/STAT1 pathway. Alternatively, CUVA treatment also increased STAT1 phosphorylation at serine 727 residue, which was inhibited by p38 inhibitor (SB203580), ATR inhibitor and also in siRNA mediated silencing of ATR. In addition, CUVA treatment mediated ATR activation facilitates translocation of serine-phosphorylated STAT1 to the Bax promoter region promoting its expression, which was decreased by SB203580 and ATR siRNA technique. Together these findings suggest that ATR mediates CUVA-induced Bax expression through p38. Silencing of JAK2 or p38 inhibition blocked procaspase-8 expression, whereas only p38 inhibition decreased Bax expression. Furthermore, inhibition of both JAK2 and p38 reduced CUVA-induced apoptosis whereas only p38 inhibition partially blocked apoptosis. Intriguingly, ATR mediated FANCD2 activation is also found to be a key in differentially regulating JAK-STAT pathway to induce apoptotic cell death in response to CUVA treatment. Thus our data suggests that the CUVA treatment may represent a novel mechanism-based cancer therapeutics for targeting ATR-FANCD2 pathway induced activation of JAK-STAT pathway in cancer cell apoptosis. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C78. Citation Format: Rahul Bhattacharyya, Mrityunjay Tyagi, Sandip K. Bandyopadhyay, Subrata Chattopadhyay, Birija Sankar Patro. ATR-FANCD2 dependent JAK-STAT pathway up-regulates Procaspase-8 and Bax in Coralyne-UVA induced cancer cell apoptosis. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C78.