Dual miRNA Targeting Restricts Host Range and Attenuates Neurovirulence of Flaviviruses

Mosquito-borne flaviviruses are among the most significant arboviral pathogens worldwide. Vaccinations and mosquito population control programs remain the most reliable means for flavivirus disease prevention, and live attenuated viruses remain one of the most attractive flavivirus vaccine platforms. Some live attenuated viruses are capable of infecting principle mosquito vectors, as demonstrated in the laboratory, which in combination with their intrinsic genetic instability could potentially lead to a vaccine virus reversion back to wild-type in nature, followed by introduction and dissemination of potentially dangerous viral strains into new geographic locations. To mitigate this risk we developed a microRNA-targeting approach that selectively restricts replication of flavivirus in the mosquito host. Introduction of sequences complementary to a mosquito-specific mir-184 and mir-275 miRNAs individually or in combination into the 3’NCR and/or ORF region resulted in selective restriction of dengue type 4 virus (DEN4) replication in mosquito cell lines and adult Aedes mosquitos. Moreover a combined targeting of DEN4 genome with mosquito-specific and vertebrate CNS-specific mir-124 miRNA can silence viral replication in two evolutionally distant biological systems: mosquitoes and mouse brains. Thus, this approach can reinforce the safety of newly developed or existing vaccines for use in humans and could provide an additional level of biosafety for laboratories using viruses with altered pathogenic or transmissibility characteristics.     

The Macrophage Inhibitor CNI-1493 Blocks Metastasis in a Mouse Model of Ewing Sarcoma through Inhibition of Extravasation

Metastatic Ewing Sarcoma carries a poor prognosis, and novel therapeutics to prevent and treat metastatic disease are greatly needed. Recent evidence demonstrates that tumor-associated macrophages in Ewing Sarcoma are associated with more advanced disease. While some macrophage phenotypes (M1) exhibit anti-tumor activity, distinct phenotypes (M2) may contribute to malignant progression and metastasis. In this study, we show that M2 macrophages promote Ewing Sarcoma invasion and extravasation, pointing to a potential target of anti-metastatic therapy. CNI-1493 is a selective inhibitor of macrophage function and has shown to be safe in clinical trials as an anti-inflammatory agent. In a xenograft mouse model of metastatic Ewing Sarcoma, CNI-1493 treatment dramatically reduces metastatic tumor burden. Furthermore, metastases in treated animals have a less invasive morphology. We show in vitro that CNI-1493 decreases M2-stimulated Ewing Sarcoma tumor cell invasion and extravasation, offering a functional mechanism through which CNI-1493 attenuates metastasis. These data indicate that CNI-1493 may be a safe and effective adjuvant agent for the prevention and treatment of metastatic Ewing Sarcoma.     

Nickel chloride inhibits the DNA repair of UV-treated but not methyl methanesulfonate-treated chinese hamster ovary cells

Nickel, a human carcinogen, has been shown to enhance the cytotoxicity, mutagenicity, and sister-chromatid exchanges (SCE) induced by ultraviolet (UV) light but not by methyl methanesulfonate (MMS). To verify that the cocytotoxicity and cogenotoxicity of nickel are correlated with its inhibition on DNA repair, the effects of nickel on the DNA repair induced by UV and by MMS have been investigated. Our analyses of DNA repair of single-strand breaks by alkaline elution and alkaline sucrose sedimentation indicate that nickel inhibited the DNA repair in UV-treated, but not in MMS-treated cells. Therefore, the inhibition of DNA repair seems to play an important role in the cocytotoxicity and comutagenicity of nickel. However, the inhibition of DNA repair seems not to play a decisive role in enhancing SCE, because we have previously shown that arsenite inhibits the UV-induced DNA repair, but has no enhancing effect on the UV-induced SCE. Our results also show that nickel had obvious inhibitory effects on DNA ligation and postreplication repair, but had no apparent effect on nucleotide excision and DNA polymerization in the UV repair. The results of the DNA ligation inhibition by nickel in UV but not in MMS repair suggest that different ligases are used in the DNA repair of UV- and MMS-induced damages.