Abstract 176: Thymoquinone attenuates tumor development in a mouse model of Lynch syndrome.
Authors: Kortüm Benedikt , Campregher Christoph , Pinter Matthias , Lang Michaela , Evstatiev Rayko , Khare Vineeta , Kucherlapati Melanie H. , Edelmann Winfried , Gasche Christoph 
Published: August 2014

Background: Hereditary nonpolyposis colorectal cancer (HNPCC, Lynch syndrome) is caused by germline mutations of DNA mismatch repair proteins leading to microsatellite instability (MSI). Nigella sativa oil has known antitumorigenic effects and was identified in an in vitro screen for compounds that reduce microsatellite instability (C. Campregher, unpublished data). Here we used a mouse model for HNPCC (Villin-Cre Msh2loxP/loxP) to test the effect of thymoquinone (TQ), the active ingredient of Nigella sativa oil, on intestinal tumor development.

Methods: 60 mice (20 per group) were treated for 10 months with chow containing TQ-low (37.5 mg/kg diet), TQ-high (375 mg/kg diet) or regular mouse chow. Serial tissue sections of paraffin embedded Swiss rolls of the entire intestine were H&E stained and analyzed for tumor multiplicity, incidence and size. As tumor multiplicity in these mice is generally low (1-2 tumors/mouse) serial sections were prepared from the upper-, middle- and lower part of the paraffin block. Tumor size was scored as small, medium or large according to its presence on 1, 2 or all 3 sections of the block. DNA was isolated from microdissected normal intestinal and tumor tissue and tested for MSI by fragment analysis using a panel of six markers (Kabbarah 2003). Data were quantitated as nucleotide shift per marker (NSPM) and compared to germline DNA from mouse tails.

Results: Treatment with TQ-low and TQ-high resulted in a significant reduction of tumor incidence from 94.4% to 57.9% (p&lt;0.01) and 55.0% (p&lt;0.006), respectively. TQ-low and TQ-high also reduced tumor multiplicity from 3.1 to 1.4 (p=0.003) and to 0.9 (p&lt;0.001), respectively. The number of small and medium sized tumors was reduced in both treatment groups. The number of large tumors was not affected. Fragment analysis demonstrated global MSI in normal intestinal epithelial cells (on average 1.8 NSPM). TQ-low and TQ-high reduced the global MSI in normal intestinal epithelial cells to 0.9 and 0.9 NSPM, respectively. Tumor DNA displayed generally higher MSI than normal intestinal epithelial cells (2.7, 2.5, and 1.5 NSPM for control mice, TQ-low, and TQ-high, respectively) and was only reduced by TQ-high.

Conclusion: In this mouse model of HNPCC TQ reduces the tumor incidence and tumor multiplicity by increasing replication fidelity in the Msh2-deficient intestinal epithelial cells. We propose TQ as a candidate compound for prevention of colorectal cancer in carriers of HNPCC mutations.

Citation Format: Benedikt Kortüm, Christoph Campregher, Matthias Pinter, Michaela Lang, Rayko Evstatiev, Vineeta Khare, Melanie H. Kucherlapati, Winfried Edelmann, Christoph Gasche. Thymoquinone attenuates tumor development in a mouse model of Lynch syndrome. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 176. doi:10.1158/1538-7445.AM2013-176