Inhibitory effect of vitamin C plus vitamin K3 on tumor growth and metastasis of Lewis lung carcinoma xenografted in C57BL/6 mice and the protective effect of vitamin C against cisplatin toxicity

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Overall abstract: Lung cancer, a highly metastatic cancers and difficultly diagnosis in an early stage, is one of the most common cause of death for cancer patients death in worldwide. Metastasizing cells must first disseminate from the primary tumor, invade the surrounding tissue, intravasate and extravasate the circulatory system, arrest, initiate angiogenesis and colonize distant site. Therefore, development of novel therapeutic strategy for treatment lung cancer is important. Vitamin C in combination with vitamin K3 (vit CK3) has been shown to inhibit tumor growth and metastasis in vivo, but the mechanism of action is poorly understood. In the first part thesis of, C57BL/6 mice were implanted (s.c.) with Lewis lung carcinoma (LLC) for 9 days before injection (i.p.) with low- (100 mg vit C/kg + 1 mg vit K3/kg) and high- dose (1000 mg vit C/kg + 10 mg vit K3/kg) vit CK3 twice a week for additional 28 days. As expected, vit CK3 or cisplatin (6 mg /kg, as positive control) significantly and dose-dependently inhibited tumor growth and lung metastasis in LLC-bearing mice. Vit CK3 restored the body weight of tumor-bearing mice to the level of tumor-free mice. Vit CK3 significantly decreased activities of plasma metalloproteinase (MMP)-2, -9 and urokinase plasminogen activator (uPA). In lung tissues, Vit CK3 1) increased protein expression of tissue inhibitor of metalloproteinase-1 (TIMP-1), TIMP-2, non-metastatic protein 23 homolog 1 (nm23-H1) and plasminogen activator inhibitor-1 (PAI-1); 2) reduced protein expression of MMP-2 and MMP-9; 3) inhibited the proliferating cell nuclear antigen (PCNA). These results demonstrate that vitamin CK3 inhibits primary tumor growth and exhibits anti-metastastic potential in vivo through attenuated tumor invasion and proliferation. Platiunm complexes have been shown to inhibit the tumor growth in animals and in human. Among these platinum complexs, cisplatin was the first found to be anti-cancer drugs, but produced strong nephrotoxicity. In the second part of this thesis, we further investigated the role of vit C on nephrotoxicity and oxdative damage caused by cisplatin and the additive effect of vit C in combination with cisplatin. Similarly, we used the same animal model from first part of thesis. Herein, C57BL/6 mice were implanted (s.c.) with Lewis lung carcinoma (LLC) for 9 days before injection (i.p.) with low (200 mg/kg), high-dose (1000 mg/kg) vitamin C in the present or absence of cisplatin (5 mg/kg) twice a week for additional 28 days. Results revealed that vitamin C or cisplatin alone significantly inhibited tumor growth, whereas the inhibitory effect of vitamin C in combination with cisplatin did not exhibit synergistic effect. In addition, we found that vitamin C in combination with cisplatin reduce the nephrotoxicity and oxidative damage caused by cisplatin, as evidenced by decreasing blood urea nitrogen (BUN) and creatinine in plasma and decreasing TBARS, carbonyls and GSH/GSSG ratio in liver and kidney tissues. In summary, we demonstrate that vitamin CK3 inhibits primary tumor growth and exhibits anti-metastastic potential in vivo in first part of thesis, and the results suggest that this effect is related to attenuation of tumor invasion and proliferation. Besides, in second part of this thesis, we demonstrate that vitamin C in combination with cisplatin inhibits primary tumor growth, and reduced the nephrotoxicity and oxidative damage caused by cisplatin in vivo. chapter 1: Vitamin C in combination with vitamin K3 (vit CK3) has been shown to inhibit tumor growth and lung metastasis in vivo, but the mechanism of action is poorly understood. Herein, C57BL/6 mice were implanted (s.c.) with Lewis lung carcinoma (LLC) for 9 d before injection (i.p.) with low- (100 mg vit C/kg + 1 mg vit K3/kg), high-dose (1000 mg vit C/kg + 10 mg vit K3/kg) vit CK3 twice a week for additional 28 days. As expected, vit CK3 or cisplatin (6 mg /kg, as positive control) significantly and dose-dependently inhibited tumor growth and lung metastasis in LLC-bearing mice. Vit CK3 restored the body weight of tumor-bearing mice to the level of tumor-free mice. Vit CK3 significantly decreased activities of plasma metalloproteinase (MMP)-2, -9 and urokinase plasminogen activator (uPA). In lung tissues, Vit CK3 1) increased protein expression of tissue inhibitor of metalloproteinase-1 (TIMP-1, TIMP-2), non-metastatic protein 23 homolog 1 (nm23-H1) and plasminogen activator inhibitor-1 (PAI-1); 2) reduced protein expression of MMP-2 and MMP-9; 3) inhibited the proliferating cell nuclear antigen (PCNA). These results demonstrate that vitamin CK3 inhibits primary tumor growth and exhibits anti-metastastic potential in vivo through attenuated tumor invasion and proliferation. chapter 2: Platiunm complexes have been shown to inhibit the tumor growth in animals and in human. Among these platinum complexs, cisplatin was the first found to be anti-cancer drugs, but produced strong nephrotoxicity. Vitamin C (vit C), an important water soluble vitamin and antioxidant, has been shown to protect cell membranes and DNA integrity against free radicals attack to avoid the progress of chronic disease, such as cardiovascular disease and cancer. However, little is known whether vit C enhanced the anticancer effect and reduces the nephrotoxicity and oxidative damage caused by cisplatin in tumor-xenografted mice. Herein, C57BL/6 mice were implanted (s.c.) with Lewis lung carcinoma (LLC) for 9 days before administration (i.p.) with low (200 mg/kg), high-dose (1000 mg/kg) vit C in the present or absence of cisplatin (5 mg/kg) twice a week for additional 28 days. Results reveal that vit C or cisplatin alone significantly inhibited tumor growth whereas vit C in combination with cisplatin did not exhibit synergistic effect. In addition, vit C ameliorated the nephrotoxicity and oxidative damage caused by cisplatin, as evidenced by decreased blood urea nitrogen (BUN) and creatinine in plasma and decreased TBARS, carbonyls and GSH/GSSG ratios in liver and kidney tissues. In summary, the present study demonstrates that vit C inhibits tumor growth and reduces the nephrotoxicity and oxidative damage in LLC-bearing mice induced by cisplatin.