A Peptide of SPARC Interferes with the Interaction between Caspase8 and Bcl2 to Resensitize Chemoresistant Tumors and Enhance Their Regression In Vivo Mahbuba Rahman1,2, Annie P. K. Chan1,2, Isabella T. Tai1,2* 1 Division of Gastroenterology, University of British Columbia, Vancouver, British Columbia, Canada, 2 Genome Sciences Centre, British Columbia Cancer Agency, Vancouver, British Columbia, Canada Abstract SPARC, a matricellular protein with tumor suppressor properties in certain human cancers, was initially identified in a genome-wide analysis of differentially expressed genes in chemotherapy resistance. Its exciting new role as a potential chemosensitizer arises from its ability to augment the apoptotic cascade, although the exact mechanisms are unclear. This study further examines the mechanism by which SPARC may be promoting apoptosis and identifies a smaller peptide analogue with greater chemosensitizing and tumor-regressing properties than the native protein. We examined the possibility that the apoptosis-enhancing activity of SPARC could reside within one of its three biological domains (N- terminus (NT), the follistatin-like (FS), or extracellular (EC) domains), and identified the N-terminus as the region with its chemosensitizing properties. These results were not only confirmed by studies utilizing stable cell lines overexpressing the different domains of SPARC, but as well, with a synthetic 51-aa peptide spanning the NT-domain. It revealed that the NT- domain induced a significantly greater reduction in cell viability than SPARC, and that it enhanced the apoptotic cascade via its activation of caspase 8. Moreover, in chemotherapy resistant human colon, breast and pancreatic cancer cells, its chemosensitizing properties also depended on its ability to dissociate Bcl2 from caspase 8. These observations translated to clinically significant findings in that, in-vivo, mouse tumor xenografts overexpressing the NT-domain of SPARC had significantly greater sensitivity to chemotherapy and tumor regression, even when compared to the highly-sensitive SPARC- overexpressing tumors. Our results identified an interplay between the NT-domain, Bcl2 and caspase 8 that helps augment apoptosis and as a consequence, a tumor’s response to therapy. This NT-domain of SPARC and its 51-aa peptide are highly efficacious in modulating and enhancing apoptosis, thereby conferring greater chemosensitivity to resistant tumors. Our findings provide additional insight into mechanisms involved in chemotherapy resistance and a potential novel therapeutic that specifically targets this devastating phenomenon. Citation: Rahman M, Chan APK, Tai IT (2011) A Peptide of SPARC Interferes with the Interaction between Caspase8 and Bcl2 to Resensitize Chemoresistant Tumors and Enhance Their Regression In Vivo. PLoS ONE 6(11): e26390. doi:10.1371/journal.pone.0026390 Editor: Pan-Chyr Yang, National Taiwan University Hospital, Taiwan Received June 24, 2011; Accepted September 26, 2011; Published November 1, 2011 Copyright: 2011 Rahman et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was funded by the Canadian Institutes of Health Research (CIHR, MOP 82881) (www.cihr-irsc.gc.ca), and the Michael Smith Foundation for Health Research (MSFHR) (www.msfhr.org). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: itai@bcgsc.ca Introduction Many pathological conditions arise because of abnormal regulation in cellular activities, such as apoptosis, that disrupt the fine balance between cell survival and death. This dysregu- lation can contribute to cancer initiation, progression, and even influence a tumor’s response to chemotherapy. SPARC (secreted protein and rich in cysteine), a matricellular protein found to be underexpressed in certain cancers, has emerged as a multifunc- tional protein capable of inhibiting the growth of neuroblastomas [1], leukemia [2], pancreatic [3], colorectal [4] and ovarian cancers [5]. Its pro-apoptotic activity in ovarian, pancreatic, lung and colorectal cancers (CRC) [4,6,7], is also thought to enhance chemotherapeutic response and reverse drug resistance [4,8]. Recent studies revealed that the recruitment and propagation of the apoptotic cascade involved the interaction between the N- terminus of caspase 8 and SPARC [8]. In this study, the mechanisms involved in SPARC-mediated apoptosis are further examined, with a specific focus on identifying a region within SPARC that may be responsible for promoting apoptosis. This is based on reports that the three structural domains of SPARC contribute to this protein’s multi-functional yet distinct biological properties (Fig. 1A): (1) N-terminus (NT), (2) follistatin-like (FS), and (3) the extracellular C-terminus (EC) domains [9,10]. For example, the N-terminus contributes to its cell spreading properties [11], the follistatin-like domain contains cysteine-rich residues, and has been shown to inhibit endolethial cell migration [12,13], while the C-terminus contains the extracellular Ca2+- binding module [14] and may have anti-angiogenic properties [11,13,15]. Our current study demonstrates that the pro-apoptotic activity of SPARC is confined to a specific region of the protein, and that a PLoS ONE | www.plosone.org 1 November 2011 | Volume 6 | Issue 11 | e26390