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Dec 1, 2009 (Vol. 29, No. 21)

China Takes Aim at RNAi

Government Agencies Are Funding Many of the Novel Research Strategies Currently Under Development

  • Click Image To Enlarge +
    Scientists at Sirnaomics are evaluating a silica-based siRNA drug-delivery system.

    China-based companies, others with facilities or collaborative ventures in China, and academic groups all committed to advancing RNA-interference (RNAi) research and therapeutic drug development were well-represented at Select Biosciences’ “RNAi Asia” meeting held recently in Kunshan, China. Most of the siRNA work going on in China is being funded by the government, with industry only playing a small role at present, according to Patrick Lu, Ph.D., president and CEO of Sirnaomics.

    Some local governments—such as the Suzhou-Kunshan region where the conference took place and which promotes itself as the RNAi Valley—“want to use RNAi as an entry point into the biopharmaceutical arena and are putting a lot of effort and money into RNAi research,” Dr. Lu said.

    Ling Qin, Ph.D., professor of orthopedic translational research in the department of orthopedics and traumatology at the Chinese University of Hong Kong, described his group’s work using siRNA to treat musculoskeletal disorders and, in particular, osteoporosis. The group is developing a bone anabolic siRNA therapeutic targeting severe postmenopausal osteoporosis.

    Osteoporosis is an attractive target for RNAi because genes associated with the regulation of bone formation and resorption have been identified, and siRNA molecules tend to accumulate in the bone marrow after systemic administration, Ge Zhang, Ph.D., a research assistant professor in Dr. Qin’s laboratory, told GEN.

    The incidence of osteoporotic fracture has doubled over the past three decades in Hong Kong. While various drugs are available to prevent and treat postmenopausal osteoporosis, except for parathyroid hormone (PTH) they all act by inhibiting bone resorption to prevent further bone loss; they do not stimulate the formation of new bone. Dr. Zhang emphasized the unmet medical need for bone anabolic therapies that do not promote bone resorption, as is the case with PTH.

    “Our collaborators identified casein kinase-2 interacting protein-1 (CKIP-1) as a newly discovered intracellular negative regulator gene of bone formation without stimulating bone resorption,” he said. Dr.  Qin’s group has shown that the bone volume/total volume ratio and trabecular bone thickness and number are higher and the trabecular space is lower in female CKIP-1 knockout mice compared to wild type.

    Furthermore, silencing of CKIP-1 in mice caused no abnormalities in major organs such as the liver or kidney. The group has screened a series of siRNA sequences using rat osteoblast cells and monkey and human osteoblasts in culture and has identified a cross-species siRNA with high knockdown efficiency, according to Tang Tao, Ph.D., a postdoctoral research fellow in Dr. Qin’s group.

    Dr. Zhang identified the main challenge in developing a systemic siRNA drug as the need to deliver relatively large doses, increasing the cost of treatment, and the risk for harmful side effects. The group is developing a bone-targeted delivery system to enhance the tissue selectivity and knockdown efficiency of systemically administered CKIP-1 siRNA. They have also demonstrated that linking the siRNA to polyethylenimine increases its accumulation in the bone and bone marrow compared to delivery of free CKIP-1 siRNA. The next step is to evaluate the drug’s pharmacokinetics and dose-effect following systemic delivery in an aged, osteoporotic rat model.

  • Screening

    Click Image To Enlarge +
    The application of an EdU-labelling method in HCS assays (RiboBio)

    Guangzhou, China-based RiboBio is using an EdU assay for siRNA screening. RiboBio offers functional genomics and drug target validation services and produces natural and unnatural oligonucleotides for RNAi applications. The company collaborated with the Guangzhou Institute of Biomedicine and Health to develop high-throughput siRNA synthesis and RNAi-screening platforms.

    Bill Zhang, Ph.D., head of science and technology at RiboBio, provided an example of how the company screens its library of 3,000 human gene siRNAs for their effects on cell viability/cell proliferation using the Click-iT® EdU (5-ethynyl-2´-deoxyuridine) assay technology, an alternative to the BrdU assay for detecting and quantifying newly synthesized DNA that does not require DNA denaturation.

    The EdU becomes incorporated into the growing DNA chain, and when exposed to a fluorescent azide (Cell-Light™ Apollo-azide) emits a fluorescent signal that indicates active cell division. The EdU staining protocol takes less than 2.5 hours, according to Dr. Zhang. Combining EdU with other dyes in the same experiment allows for multiplexing and simultaneous evaluation for cell-cycle proliferation, apoptosis, and phenotype, as well as cytotoxicity and cell-signaling pathway activation.

    RiboBio’s screening campaign identified groups of siRNAs that either enhance or inhibit cell proliferation, leading to the description of 13 genes required for cell cycle progression.

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