**5. Conclusion**

196 Liver Tumors

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

Doxorubicin, as a chemotherapy drug, has been commonly used in the treatment of hepatocellular carcinoma, but is thwarted by a key obstacle-cumulative cardiotoxicity. To improve its anti-cancer activity and reduce its toxicity, a number of doxorubicin formations such as pegylated liposomal doxorubicin, doxorubicin-eluting-bead has been performed in liver cancer clinical trials recently. Among them, anthracycline-DNA complexes as therapeutic reagents have been used for the treatment of liver cancer for a long history, due its easy-fabrication, no change in the structure of the drug molecules and its anti-cancer activity (Trouet, 1979a). Clinical trials were also carried out to evaluate their enhancement in the performances in effectiveness and toxicology compared to free anthracycline (Trouet, 1979b). However, except a few of *in vitro* reports about successfully reducing the toxicity of the anthracycline by using the complexes, there is no convincing results demonstrating the advantages of using the complexes instead of free anthracycline (Dorr et al., 1991). One of the most important reasons underlies the ineffectiveness of the intercalation between DNA and anthracycline is that there is abundant Dnase in circulation and body fluids which digests un-protected DNA molecular with extremely high efficiency (Paik & Kim, 1970). The anthracycline-DNA complexes were destroyed soon after their injection into the body. The anthracycline in complexes was released very quickly and did not change their

Protection of DNA from destroying by Dnase is necessary for gene delivery technology (Tranchant et al., 2004). Non-viral gene delivery practices take the advantage of using cationic polymers to combine DNA, which prevents the exposure of DNA molecules to Dnases (Wagner, 2004). DNA complexed with cationic polymers could totally resist the degradation of Dnase I or serum Dnases in reported *in vivo* and *in vitro* tests (Paleos et al., 2009). These researches instruct us cationic polymers could be used to combine the intercalation of anthracycline-DNA to form an anthracycline-DNA-cationic polymer ternary complex. Such a complex can avoid the unexpected early release of anthracycline into serum by protecting DNA from the digestion of Dnases. In our experimets, cationic gelatin was used to combine DNA-DOX intercalation. This simple process generated regular nanocomplexes that could be observed. It is difficult to release DOX in short time using Dnase I alone to digest the complex, which suggested that DNA is well protected by cationic gelatin. Additionally, in the present study, synthesized GC oligonucleotide was used to substitute the natural DNA fragments, which increased the drug-loading capacity of DNA. Moreover, the combination between DOX and polyGC is more stable than DOX-natural DNA intercalation fragments which may release DOX when combined by some kind of cationic polymer. No DOX release was observed in the study when DOX-polyGC was

A lot of cationic polymers are used as gene carriers, such as polyether imide (PEI), polylysine (PLL), cationic polysaccharides and other polycation materials (Bodley et al.,

within 5 weeks while CPX2 rescued 90% of them in more than 4 weeks.

**4. Discussion** 

pharmaceutical properties any more.

combined by cationic gelatin.

In our present study, a PH-/enzyme responsive complex composed by self-assemble of doxorubicin, CpG DNA fragments, cationic gelatin and a PH-sensitive alginate was developed. This complex increased the accumulation of doxorubicin in tumor, reduced its deposition in heart and could specifically release doxorubicin in tumor sites, which resulted in the enhanced anti-cancer activity and decreased cardiotoxicity of doxorubicin. When tested in animal model of implanted tumor, the complex exhibited high effective in preventing the growth of the tumors and dramatically alleviated toxicity compared to free doxorubicin. All results suggest that this easy- manufactured, cost-effective nanoscale formulation of doxorubicin hold great promise to be used to clinical practices.
