**4. Clinical trials**

Local and systemic drug administrations are the main routes of drug delivery in current clinical trials *in vivo*. Local drug administration depends on the type of target tissue, including intravitreal injection for ophthalomological diseases; intranasal administration for respiratory diseases caused by respiratory syncytial virus; intracranial injection for central nervous system diseases; intratumor injection for carcinogenic genes; and intramuscular injection for muscle tissue diseases such as rheumatoid arthritis. Drugs delivered by local administration can directly act on the target organ, thus reducing the probability of degradation of siRNA by nuclease and the dosage of siRNA. Systemic drug administration mainly depends on intraperitoneal and intravenous injection. Systemic drug administration often needs more siRNA to enable the target organ or tissue to obtain an efficacious drug concentration.

A lot of studies have demonstrated that RNAi has substantial potential for tumor treatment. Several RNAi agents have entered clinical trials in the world (Table 1). In May 2008, RNAi agent CALAA-01 for the treatment of solid tumors, developed by Calando Pharmaceuticals, was approved by the Food and Drug Administration (FDA) of the United States for Phase I clinical trial. CALAA-01 is a complex of siRNA and cyclodextrin-transferrin-adamant-PEG nanoparticles, in which human transferrin can specifically interact with transferrin receptor

RNA Interference for Tumor Therapy 75

spreading to the lungs in mice. In addition, Atu027 could apparently inhibit the growth of transplanted prostate cancer in nude mice, where the tumor size was about 50% that of the control group; the number of metastatic lymph nodes was only 50% that of the control group; and the lymphatic density in the tumor area was also decreased markedly, though there was no significant difference in vascular density in the tumor area between the study and control groups. In a toxicity experiment, the researchers injected different doses (0.3, 1.0 and 3.0mg/kg) of Atu027 to different groups of cynomolgus monkeys at a 4-day interval, and no significant toxic and adverse effects were observed. Phase I clinical trial with Atu027

The understanding of signaling pathways participating in the development and progression of malignant tumors and the development of specific target RNAi technique have offered hope to conduct individualized regimens for tumor treatment. However, current RNAi therapeutic approaches are immature and many problems need to be tackled, such as targeting and off-target effects, drug delivery systems, drug administrations and safety. As siRNA with a sequence of only 20-nucleotides could induce a tremendous effect of gene silencing, the sequence of siRNA must be selected accurately in the process of designing siRNA sequences to avoid the off-target effect. Additionally, a mRNA target sequence may hide in the mRNA secondary structure or folding region, making it impossible for siRNA to silence the mRNA. It is therefore necessary to conduct experiments repeatedly to select a suitable target site. Before RNAi medical treatment can be used clinically, it is necessary to seek an efficient delivery system and rational drug administration. In addition, as nucleotide sequences used for RNAi have no cell targetability and are likely to degrade, it is necessary to design and synthesize a drug delivery system that is non- or low toxic to normal cells and

This work was financially supported by the Key Scientific Project of Shanghai Putuo District(PTKW10-B01), the Young Program of Shanghai health bureau(2009Y127), Young

Aleku, M.; Schulz, P.; Keil, O.; Santel, A.; Schaeper, U.; Dieckhoff, B.; Janke, O.; Endruschat,

Alnylam Pharmaceuticals. (August 2011).Dose Escalation trial to evaluate the safety,

J.; Durieux, B.; Roder, N.; Loffler, K.; Lange, C.; Fechtner, M.; Mopert, K.; Fisch, G.; Dames, S.; Arnold, W.; Jochims, K.; Giese, K.; Wiedenmann, B.; Scholz, A.; Kaufmann, J.(2008). Atu027, a liposomal small interfering RNA formulation targeting protein kinase N3, inhibits cancer progression, *Cancer Research.*, Vol.68,

tolerability, pharmacokinetics and pharmacodynamics of intravenous ALN-VSP02

backbone promotion plan of Shanghai society of Nuclear Medicine (2009-NM-07).

No.23, (December 2008), pp.9788-9798, ISSN0008-5472.

mainly showed safety, tolerance and pharmacokinetics.

can deliver siRNA to tumor cells efficiently.

**6. Acknowledgment** 

**7. References** 

**5. Conclusion** 

that is overexpressed on the tumor cell membrane, thereby triggering the entry of the nanoparticles into tumor cells through receptor-mediated endocytosis. The target gene is ribonucleotide reductase subunit 2 (RRM2). After systemic administration by i.v. injection, siRNA efficiently inhibited tumor growth by down-regulating RRM2 expression (Davis,2009). It was also found that the levels of RRM2 mRNA and RRM2 protein were both decreased in the tumor. To detect the efficiency of RNAi, researchers found that there existed large amounts of RRM2 mRNA degradation products at tumor site containing the nanoparticles(Davis *et al,* 2010). This is the first clinical trial on RNAi medical treatment of malignant tumors.


Table 1. A summary of current siRNA agent for clinical trials

Alnylam Pharmaceuticals (Alnylam Pharmaceuticals,2011a,2011b) conducted Phase I clinical trial by intravenous administration of ALN-VSP02 for the treatment of HCC, where the siRNA therapeutic was enveloped in liposome. A total of 58 patients were recruited in the study and administered with the therapeutic at an interval of two weeks. The therapeutic agent targets kinesin spindle protein (KSP) and VEGF that are highly expressed in tumor cells. KSP plays a key role in cell mitosis and is the dynein necessary for bipolar spindle formation and maintenance. Inhibition of KSP would arrest the cell cycle. VEGF promotes angiogenesis, and therefore inhibition of VEGF would suppress tumor growth. Currently, Phase I clinical trial with ALN-VSP02 has been done, and Phase II clinical trial is to be started in 2012.

Recently, Silence Therapeutics has started Phase I clinical trial on Atu027(Aleku et al, 2008; Santel et al, 2010). Atu027 is a lipid-based siRNA complex containing target protein kinase N3 (PKN3), which participates in tumor occurrence and plays an important role in tumor metastasis. Atu027 can inhibit tumor progression by silencing the expression of PKN3. Preclinical experiments demonstrated that Atu027 could prevent breast cancer cells from spreading to the lungs in mice. In addition, Atu027 could apparently inhibit the growth of transplanted prostate cancer in nude mice, where the tumor size was about 50% that of the control group; the number of metastatic lymph nodes was only 50% that of the control group; and the lymphatic density in the tumor area was also decreased markedly, though there was no significant difference in vascular density in the tumor area between the study and control groups. In a toxicity experiment, the researchers injected different doses (0.3, 1.0 and 3.0mg/kg) of Atu027 to different groups of cynomolgus monkeys at a 4-day interval, and no significant toxic and adverse effects were observed. Phase I clinical trial with Atu027 mainly showed safety, tolerance and pharmacokinetics.
