**3.5 Anti-cancer activity of CPX1 and CPX2**

194 Liver Tumors

Fig. 5. Toxicology analysis. A) Body weight changes of animals received i.v. injections of saline, free DOX, CPX1 or CPX2 at a dose of 20 mg/kg body weight; B) Mortality caused by free DOX, CPX1 or CPX2 at different doses; C) H & E stained heart, liver and lung tissue sections of

Treatment with free doxorubicin caused massive myocardial degeneration (Figure 5C), which resulted in the increase of the plasma level of CK and LDH (Table 1) and heart failure which induced significant pulmonary congestion (Figure 5C). This toxicity was greatly reduced when DOX was delivered by CPX1 and CPX2 (Figure 5C and Table 1). CPX1 showed more hepatotoxicity than free doxorubicin at the same dose, which resulted in the necrosis of hepatocytes (Figure 5C) and increase in plasma levels of ALT (Table 1) while CPX2 did not cause this toxicity. CPX2 also reduced the nephrotoxicity of doxorubicin

animal treated by 20 mg/kg body weight free DOX or delivered by CPX1 and CPX2.

which was demonstrated by the decrease of BUN level (Table 1).

The anti-tumor activity was evaluated in a mouse orthotopic allograft liver cancer model using a hepatoma cell line, Hepa 1-6. Figure 6A shows the photograph of tumors separated from animals 16 days after they received free DOX, CPX1, CPX2 and saline in vein. From the results of histological analyze of tumor sections (Figure 6B), massive cancer cell remissions could be observed from the sections of tumors treated with CPX1 and CPX2. CPX2 exhibited the most efficiency on prevention of cancer cell's expansion.

Fig. 6. Anti-tumor activity. A) Representative Tumors separated from animals received intravenous injections of saline, free DOX, CPX1 or CPX2. B) Sections of tumor tissues taken from animal received intravenous injections of saline, free DOX, CPX1 or CPX2. C) Mean weights of tumors separated from animals received of saline, free DOX, CPX1 or CPX2 intravenously. D) Survive rates of animals implanted tumors received treatments with saline, free DOX, CPX1 or CPX2 in more than 4 weeks. \*p<0.05 compared with saline treated group.

The mean weight of the tumors isolated from the liver was shown in figure 6C. CPX2 exhibited more than 1 times higher anti-tumor activity than free DOX. Results of the examination of animal survive shown in Figure 6D. Animals bearing tumors and received saline as control died within 4 weeks. Free doxorubicin kept 30% animals from death within 5 weeks while CPX2 rescued 90% of them in more than 4 weeks.
