Anti-FECH monoclonal antibody Made to order

Edwardsiella tarda is a Gram-negative bacterial pathogen that can infect a wide range of freshwater and marine fish. One salient feature of E. tarda is the ability to survive and replicate in various host cells. In this study, we observed that E. tarda replicated robustly in the zebrafish cell line ZF4, and that E tarda-infected cells exhibited no detectable signs of apoptosis. Global transcriptome analysis and quantitative real-time RT-PCR revealed that E. tarda infection generally significantly downregulated pro-apoptotic genes and upregulated anti-apoptotic genes. To investigate the role of apoptosis in E. tarda infection, two upregulated anti-apoptotic genes (Fech and Prx3) and two downregulated pro-apoptotic genes (Brms1a and Ivns1a) were overexpressed in zebrafish. Subsequent infection study showed that Fech and Prx3 overexpression significantly promoted E. tarda dissemination in and colonization of fish tissues, while Brms1a and Ivns1a overexpression significantly reduced E tarda dissemination and colonization. Consistently, when Fech and Prx3 were knocked down in zebrafish, E. tarda infection was significantly inhibited, whereas Brms1a and Ivns1a knockdown significantly enhanced E. tarda infection. These results indicate for the first time that E. tarda prevents apoptosis in teleost as a strategy for intracellular survival, and that some putative apoptotic genes of teleost function in the apoptosis pathway probably in a manner similar to that in mammalian systems.

Erythropoietic protoporphyria (EPP) is a hereditary disease characterized by a deficiency in ferrochelatase (FECH) activity. FECH activity is responsible for the accumulation of protoporphyrin IX (PPIX). Without etiopathogenic treatment, EPP manifests as severe photosensitivity. 95% of affected individuals present a hypomorphic FECH allele trans to a loss-of-function (LOF) FECH mutation, resulting in a reduction in FECH activity in erythroblasts below a critical threshold. The hypomorphic allele promotes the use of a cryptic acceptor splice site, generating an aberrant FECH mRNA, which is responsible for the reduced level of wild-type FECH mRNA and, ultimately, FECH activity. We have previously identified an antisense oligonucleotide (AON), AON-V1 (V1), that redirects splicing to the physiological acceptor site and reduces the accumulation of PPIX. Here, we developed a specific strategy that uses transferrin receptor 1 (TRF1) as a Trojan horse to deliver V1 to erythroid progenitors. We designed a bifunctional peptide (P-1-9R) including a TFR1-targeting peptide coupled to a nine-arginine cell-penetrating peptide (CPP) that facilitates the release of the AON from TFR1 in endosomal vesicles. We demonstrated that the P-1-9R/V1 nanocomplex promotes the efficient and prolonged redirection of splicing towards the physiological splice site and subsequent normalization of WT FECH mRNA and protein levels. Finally, the P-1-9R/V1 nanocomplex increases WT FECH mRNA production and significantly decreases PPIX accumulation in primary cultures of differentiating erythroid progenitors from an overt EPP-affected individual. P-1-9R is a method designed to target erythroid progenitors and represents a potentially powerful tool for the in vivo delivery of therapeutic DNA in many erythroid disorders.