Antitumoral and antimetastatic effects of metronomic chemotherapy with cyclophosphamide combined with celecoxib on murine mammary adenocarcinomas

Abstract

Purpose: Metronomic chemotherapy (MCT) refers to the chronic and equally spaced administration of low doses of different chemotherapy drugs, without extended interruptions. Previously, we demonstrated the antitumor effect of MCT with cyclophosphamide (Cy) in a mouse mammary adenocarcinoma model. Herein, we investigated the therapeutic efficacy of metronomic Cy combined with celecoxib (Cel) in two murine mammary adenocarcinoma models.

Methods: Mice were s.c. challenged with M-234 p or M-406 mammary tumors and from day 5 or 8 on, respectively, treated with: (I) no treatment (controls); (II) Cy in the drinking water (25-30 mg/kg body weight/day); (III) Cel (30 mg/kg p.o.), five times/week; (IV) treated as II + III. Mice challenged i.v. with M-234 p or M-406 tumor cells received, on day 3, the same treatments.

Results: We found that MCT with Cy plus Cel inhibited tumor growth decreased lung metastases, and increased the median survival time, in both tumor models, having very low toxicity. MCT with Cy combined with Cel was more effective than each monotherapy.

Conclusions: The therapeutic benefits of combined MCT with cyclophosphamide plus celecoxib on mammary adenocarcinomas together with its very low toxicity profile warrant further study in an attempt to make the translation into the clinic.

Fig. 1

Tumor growth and overall survival: On day 5 (M-234p) and day 8 (M-406) when tumor volume reached 100–140 mm³, animals were distributed in four groups and treated as indicated in “Materials and methods” section. Tumor growth assessment: (M-234p: N = 7/group; M-406: N = 5 or 6/group), data for each time-point are mean ± SEM. a Day 35: control vs. Cy + Cel (P < 0.01); day 39: Cy + Cel vs. Cy (P < 0.01), vs. Cel (P < 0.05). b Day 19: control vs. Cy (P < 0.05) and vs. Cy + Cel (P < 0.01) (ANOVA and Tukey’s multiple comparison test). Overall survival: N = 6 and N = 5 or 6/group for M-234p and M-406, respectively. c Control vs. Cy + Cel: P < 0.001 and vs. Cy: P < 0.05; Cy and Cel vs. Cy + Cel P < 0.05. d Control vs. Cy + Cel: P < 0.01 and vs. Cy P < 0.05; Cel vs. Cy + Cel P < 0.01 (Logrank/Mantel-Cox Test)

Fig. 2

Antimetastatic effect: adult Balb/C and CBi mice were injected i.v. with 5 × 10⁵ M-234p cells or 2 × 10⁵ M-406 cells, respectively. On day 3, the animals were distributed in four groups and treated as indicated in “Materials and methods” Section. Metastatic foci: a control vs. Cy + Cel P < 0.01. b Control vs. Cy + Cel P < 0.01. (Kruskal–Wallis and Dunns multiple comparison test). Metastasis size:c: control vs. Cy + Cel P < 0.001. d Control vs. Cy + Cel P < 0.001 (ANOVA and Tukey’s multiple comparison test). Lungs weight: e control vs. Cy + Cel P < 0.05. f Control vs. Cy + Cel P < 0.001 (ANOVA and Tukey’s multiple comparison test)

Fig. 3

Overall survival in experimental metastatic assay: a control vs. Cy + Cel P < 0.0001; b control vs. Cy + Cel P < 0.0001 (Logrank/Mantel-Cox test)

Fig. 4

Evolution of body weight: data for each time-point are the percentage of initial body weight, and mean ± SEM. a M-234p s.c., b M-406 s.c., c M-234p i.v., d M-406 i.v. No significant body weight losses were detected in either tumor model during treatment

Fig. 5

Serologic parameters: Data for each time-point and group are mean ± SEM serum concentration of: urea (a and b), creatinine (c and d), SGOT (e and f), SGPT (g and h). g Day 25: basal vs. control P < 0.05 and vs. Cel P < 0.01; Cel vs. Cy + Cel P < 0.01. (ANOVA and Tukey’s multiple comparison test)

Fig. 6

Total leukocytes count: a Day 25: control vs. Cy and vs. Cy + Cel P < 0.05; Cy + Cel (day 25) vs. Cy + Cel (day 56) P < 0.01. b Day 20: control vs. Cy and vs. Cy + Cel P < 0.05. (Kruskal–Wallis and Dunns multiple comparison test)

Fig. 7

VEGF serum concentration: blood samples were taken on day 0 and on days 30 and 20 from mice bearing M-234p (a) and M-406 (b) mammary adenocarcinomas, respectively. Data for each time-point and group are mean ± SEM serum concentration a basal vs. control P < 0.001; control vs. Cy and vs. Cy + Cel P < 0.01. b Basal vs. control P < 0.001; control vs. Cel P < 0.01; control vs. Cy and vs. Cy + Cel P < 0.001. (ANOVA and Tukey’s multiple comparison test)