Metformin, cimetidine, and ibuprofen separately exhibit immunomodulatory and anti-tumorigenic effects. Herein, the impacts of metformin alone and in combination with cimetidine/ibuprofen on some Th1- and regulatory T (Treg) cell-related parameters were evaluated using a breast cancer (BC) model. For establishing the BC model, four groups of Balb/c mice were challenged with the carcinoma cell line. After 11-30 days post-induction, they were treated intraperitoneally (with metformin (200 mg/kg), "metformin plus cimetidine (20 mg/kg)"; "metformin plus ibuprofen (20 mg/kg)", or with all three drugs in mentioned doses. Untreated BC and without tumor mice were enrolled as control groups. On day 31, splenic Th1 and Treg cell frequencies, serum interferon-gamma (IFN-γ), and transforming growth factor-beta (TGF-β) concentration, and intra-tumoral T-bet, TGF-β, and forkhead box protein P3 (FOXP3) expression were measured; using flow cytometry, enzyme-linked immunosorbent assay (ELISA), and real-time-PCR, respectively. Treatment of the BC mice with metformin alone and in combination with cimetidine and/or ibuprofen enhanced the frequency of Th1 cells, and IFN-γ concentration, while it resulted in a decrease in the frequency of Treg cells, serum TGF-β concentration, and the expression of FOXP3 and TGF-β compared with un-treated BC mice. FOXP3 expression in the metformin-treated group was lower in mice who received combination therapy. Survival rate and body weight were increased, while tumor size and spleen index were reduced in mice treated with metformin alone and its combination with cimetidine and/or ibuprofen. No remarkable differences were found between metformin-treated mice and those who received combination therapies regarding Th1 and Treg cell percentages, TGF-β expression, body weight, tumor size, and spleen index. The benefits of combinational therapy may be largely attributed to metformin. Immunotherapeutic potentials of metformin in cancers need further considerations.  

Objectives The potent anti-tumorigenic effects were attributed to ginger and there are some reports regarding the anti-cancer and immunomodulatory properties ginger-derived components. This study aimed to investigate the effects of zingerone on some immune-related parameters in an animal model of breast cancer. Methods The breast cancer was established in female BALB/c mice using a carcinogenic 4T1 cell line. At day 10 after cancer induction, tumor-bearing mice were divided into five groups and treated intraperitoneal (daily from days 11–30) with saline or zingerone (at doses 10, 20, 50 and 100 mg/kg/day). The mice were sacrificed on day 31 and the number of splenic Th1- and Treg cells, the expression of IFN-γ and TGF-β in the blood mononuclear cells, the antibody production against sheep red blood cell (SRBC) were determined using flow cytometry, real time-PCR and a standard hemagglutination assay, respectively. Results Zingerone at doses 50 and 100 mg/kg enhanced the number of splenic Th1 cells (p<0.03 and 0.007, respectively); at doses 10, 20, 50 and 100 mg/kg reduced the number of splenic Treg cells (p<0.02, 0.01, and 0.01, respectively), at doses 50 and 100 mg/kg enhanced the expression of IFN-γ (p<0.03), at doses 50 and 100 mg/kg reduced the expression of TGF-β, at doses 50 mg/kg reduced the titer of anti-SRBC antibody (p<0.05). Conclusions Zingerone improve the T cell-mediated and antibody responses in a mouse model of breast cancer. The immunotherapeutic potentials of zingerone in cancers need more considerations.

Cimetidine and ibuprofen exhibit immunomodulatory effects as an antagonist of histamine H2 receptor, and a cyclooxygenase inhibitor, respectively. Here, the effects of cimetidine and ibuprofen on some effector T cell-related parameters were investigated using a breast cancer (BC) model. BC was established in Balb/c mice using the 4T1 cell line. On day 10 after tumor induction, the BC-bearing mice were classified into four groups and treated with PBS, cimetidine (20 mg/kg), ibuprofen (20 mg/kg) or a combination of "cimetidine + ibuprofen" via intraperitoneal injection (daily from days 11 to 30). The mice were sacrificed on day 31 and the frequency of splenic Th1 and Treg cells, plasma IFN-γ and TGF-β levels, and intra-tumoral T-bet, GATA3, FOXP3 and RORγt expressions were detected using flowcytometry, ELISA and real-time-PCR, respectively. In untreated cancerous mice, the percentage of splenic Th1 cells and plasma IFN-γ levels were lower (P<0.003 and P<0.01, respectively), whereas the percentage of splenic Treg cells and plasma TGF-β levels were higher than in healthy mice (P<0.04 and P<0.005, respectively). Treatment of BC-bearing mice with cimetidine, ibuprofen or both drugs promoted the frequency of Th1 cells (P<0.05, P<0.007 and P<0.005, respectively) as well as IFN-γ levels (P<0.004, P<0.0001 and P<0.03, respectively), while reduced the frequencies of Treg cells (P<0.02, P<0.03 and P<0.01, respectively), TGF-β levels (P<0.006, P<0.02 and P<0.002, respectively), intra-tumoral expression of FOXP3 (P<0.006, P<0.005 and P<0.005, respectively), and intra-tumoral expression of RORγt (P<0.04, P<0.03 and P<0.05, respectively) compared with untreated BC mice. The "cimetidine + ibuprofen"-treated mice displayed greater T-bet expression than the un-treated mice (P<0.006). Cimetidine and/or ibuprofen-treated BC-bearing mice exhibited reduced intra-tumoral expression of GATA3 compared with the untreated BC mice, but the differences were not significant. Cimetidine and ibuprofen correct some effector T cell-related parameters in cancerous mice. Immunotherapeutic potentials cimetidine and ibuprofen in cancers need investigations.