Microenvironmental effects limit efficacy of thymoquinone treatment in a mouse model of ovarian cancer

Abstract

BackgroundOvarian cancer is the most lethal gynecologic malignancy, with limited treatment options for chemoresistant disease. An important link between inflammation and peritoneal spread of ovarian cancer is NF-κB signaling. Thymoquinone (TQ) exerts multiple anti-tumorigenic cellular effects, including NF-κB inhibition. We aimed to investigate the therapeutic potential of TQ in an established murine syngeneic model of ovarian cancer.MethodsID8-NGL mouse ovarian cancer cells stably expressing an NF-κB reporter transgene were injected intra-peritoneally into C57BL/6 mice, and mice were treated with TQ or vehicle for 10 or 30 days. TQ was combined with the macrophage depleting drug, liposomal clodronate, in selected experiments. Effects on peritoneal tumor burden were measured by volume of ascites, number of peritoneal implants and mesenteric tumor mass. NF-κB reporter activity and markers of proliferation and apoptosis were measured in tumors and in confirmatory in vitro experiments. Protein or mRNA expression of M1 (anti-tumor) and M2 (pro-tumor) macrophage markers, and soluble cytokine profiles, were examined from harvested ascites fluid, peritoneal lavages and/or tumor sections. 2-tailed Mann–Whitney tests were used for measuring differences between groups in in vivo experiments.ResultsConsistent with its effects in vitro, TQ reduced proliferation and increased apoptosis in ID8-NGL tumors after 10 and 30 day treatment. Prolonged TQ treatment did not significantly alter tumor number or mass compared to vehicle, but rather exerted an overall deleterious effect by stimulating ascites formation. Increased ascites was accompanied by elevated NF-κB activity in tumors and macrophages, increased pro-tumor M2 macrophages and expression of pro-tumorigenic soluble factors such as VEGF in ascites fluid, and increased tumor infiltration of M2 macrophages. In contrast, a 10 day exposure to TQ produced no ascites, and reduced tumor NF-κB activity, M2 macrophages and soluble VEGF levels. Peritoneal macrophage depletion by clodronate significantly reduced tumor burden. However, TQ-stimulated ascites was further enhanced by co-treatment with clodronate, with macrophages present overwhelmingly of the M2 phenotype.ConclusionsOur findings show that pro-tumorigenic microenvironmental effects limited the efficacy of TQ in a syngeneic mouse model of ovarian cancer, and provide caution regarding its potential use in clinical trials in ovarian cancer patients.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0463-5) contains supplementary material, which is available to authorized users.

Anti-cancer properties and mechanisms of action of thymoquinone, the major active ingredient ofNigella sativa

“…This was manifested through stimulation of ascites formation accompanied by elevated NF-κB activity in tumors and macrophages. Importantly, this was contrary to the results of a 10-day TQ treatment, where down-regulation of NF-κB activity was reported along with reduction in tumor mass (Wilson et al, 2015).…”

“…Collectively, the inhibitory effect that TQ exerts on NF-B activity is another mechanism of action that underlies the anti-cancer potential of TQ and TQ-containing oil and extracts of N. sativa. With that being said, a recent study provides caution regarding the potential use of TQ in clinical trials (Wilson et al, 2015). It highlights the effect of prolonged TQ-treatment (>30 days) as a deleterious factor for its efficacy ACCEPTED MANUSCRIPT…”

Bipolar Tumor-Associated Macrophages in Ovarian Cancer as Targets for Therapy

“…An unexpected observation made by our group revealed that inhibition of NF-κB in a syngeneic mouse model of ovarian cancer, increased pro-tumorigenic M2 macrophages, which promoted ascites, an increased expression of pro-tumorigenic soluble factors (such as VEGF in ascites fluid), and an infiltration of more M2 macrophages into the TME [ 48 , 72 ]. These results suggest that the activation of NF-κB in TAMs, not tumor cells, could be a viable therapeutic strategy.…”

Thymoquinone is knocking at the door of clinical trial

“…In this occasion, TQ diminished the expression of iNOS, TNF-α, COX-2, NF-κB and MPO activities [3]. Moreover, TQ is active against breast cancer [83][84][85], ovarian adenocarcinoma [86,87], polycystic ovarian carcinoma [88], cervical squamous carcinoma and prostate [3] cancer. A research findings suggesting TQ at 50 mg/kg/day (p.o.)…”