Characterization of Two Malaria Parasite Organelle Translation Elongation Factor G Proteins: The Likely Targets of the Anti-Malarial Fusidic Acid

Malaria parasites harbour two organelles with bacteria-like metabolic processes that are the targets of many anti-bacterial drugs. One such drug is fusidic acid, which inhibits the translation component elongation factor G. The response of P. falciparum to fusidic acid was characterised using extended SYBR-Green based drug trials. This revealed that fusidic acid kills in vitro cultured P. falciparum parasites by immediately blocking parasite development. Two bacterial-type protein translation elongation factor G genes are identified as likely targets of fusidic acid. Sequence analysis suggests that these proteins function in the mitochondria and apicoplast and both should be sensitive to fusidic acid. Microscopic examination of protein-reporter fusions confirm the prediction that one elongation factor G is a component of parasite mitochondria whereas the second is a component of the relict plastid or apicoplast. The presence of two putative targets for a single inhibitory compound emphasizes the potential of elongation factor G as a drug target in malaria

Targeting Hepatitis B Virus with Zinc Finger Nucleases

Despite an existing effective vaccine, hepatitis B virus (HBV) remains a major public health concern. There are effective suppressive therapies for HBV, but they remain expensive and inaccessible to many, and not all patients respond well. Furthermore, HBV can persist as genomic covalently closed circular DNA (cccDNA) that remains in hepatocytes even during otherwise effective therapy and facilitates rebound in patients after treatment has stopped. Therefore, the need for an effective treatment that targets active and persistent HBV infections remains. As a novel approach to treat HBV, we have targeted the HBV genome for disruption to prevent viral reactivation and replication. We generated 3 zinc finger nucleases (ZFNs) that target sequences within the HBV polymerase, core and X genes. Upon the formation of ZFN-induced DNA double strand breaks (DSB), imprecise repair by non-homologous end joining leads to mutations that inactivate HBV genes. We delivered HBV-specific ZFNs using self-complementary adeno-associated virus (scAAV) vectors and tested their anti-HBV activity in HepAD38 cells. HBV-ZFNs efficiently disrupted HBV target sites by inducing site-specific mutations. Cytotoxicity was seen with one of the ZFNs. scAAV-mediated delivery of a ZFN targeting HBV polymerase resulted in complete inhibition of HBV DNA replication and production of infectious HBV virions in HepAD38 cells. This effect was sustained for at least 2 weeks following only a single treatment. Furthermore, high specificity was observed for all ZFNs, as negligible off-target cleavage was seen via high-throughput sequencing of 7 closely matched potential off-target sites. These results show that HBV-targeted ZFNs can efficiently inhibit active HBV replication and suppress the cellular template for HBV persistence, making them promising candidates for eradication therapy

Fibrinogen and hemoglobin predict near future cardiovascular events in asymptomatic individuals

10.1038/s41598-021-84046-7Scientific Reports1114605-complete