Inhibition of RNA Viruses In Vitro and in Rift Valley Fever-Infected Mice by Didemnins A and B

Abstract

were dissolved in ethanol, reconstituted with Hanks balanced salt solution, and buffered to pH 7.2 with 10 mM HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) plus 2% (vol/vol) heat-inactivated fetal bovine serum (4% ethanol, final concentration). Drug solutions (0.3 ml) of appropriate concentration (0 to 2.5 ,ig/ml for didemnin A and 0 to 0.5 ,ug/ml for didemnin B) were added to 24-well tissue culture trays (Falcon). Virus (0.1 ml, 50 to 200 PFU) or medium (for tissue culture toxicity controls) was then added. After incubating the trays for 60 min to allow adsorption of the virus, 0. Didemnins A and B were both found to exhibit significant activity against RVF (median inhibition dose [ID50] for didemnins A and B was 1.37 and 0.04 .Ig/ml, respectively), Venezuelan equine encephalomyelitis virus (ID50, 0.43 and 0.08 gjg/ml, respectively), and yellow fever virus (ID50, 0.4 and 0.08 ,ug/ml, respectively). A concentration of 0.1 ,ug of didemnin B per ml inhibited plaque formation by these three viruses by more than 80%o. Didemnin A, on the other hand, was less efficient, requiring a 25-fold increase in concentration to achieve the same level of virus plaque inhibition. Both didemnins A and B showed mild cytotoxicity in Vero-76 cells as judged by phase-contrast microscopy at concentrations greater than 5 and 0.5 Fg/ml (5.3 and 0.45 M), respectively, consisting of increased vacuolation with moderate cell rounding but no detachment. As compared with the other three test viruses, Pichinde virus (ID50 for didemnins A and B was 2.9 and 0.22 ,ug/ml, respectively), a representative arenavirus, was less sensitive to the in vitro antiviral effect of both didemnins.Studies were also conducted with RVF-infected mice. Female Swiss Webster mice, 3 to 4 weeks of age, were challenged subcutaneously with approximately 250 PFU of RVF per 0.1 ml.

Natural products as probes of cell biology: 20 years of didemnin research

“…Both the A and B congeners showed comparable in vitro inhibitory effects against the viruses causing RVF, Venezuelan equine encephalomyelitis, and yellow fever. The inhibition was observed at concentrations similar to those initially reported by Rinehart et al, with didemnin B consistently showing a roughly tenfold potency enhancement relative to didemnin A. Additionally, Canonico et al 27 reported the ®rst results showing in vivo ef®cacy of didemnins. At a dose of 0.25 mg/kg/day of didemnin B, mice infected with RVF virus had a 90% survival rate, although some unspeci®ed fatal drug-related deaths were observed.…”

“…Canonico et al 27 reported that didemnins A and B were able to protect mice against lethal doses of the Rift Valley fever (RVF) virus. Reportedly, there existed no effective therapy for this virulent human pathogen.…”

Biomedical Potential of Marine Natural Products

“…In addition to their antitumor activity, they exhibited in vitro and in vivo antiviral properties. Didemnin B significantly reduced the yields of DNA and RNA viruses by 104-105 at 0.5 IJ.g/ml, and in in vivo tests protected over 70% of a population of mice from lethal vaginal doses of herpes simplex type 2 virus (intravaginal application three times per day) (Rinehart et al, 1983), and a lethal subcutaneous challenge of Rift Valley fever virus with a 90% survival rate at 0.25 mg/kg (Canonico et al, 1982).…”

Fate of tritiated didemnin B in mice: excretion and tissue concentrations after an intraperitoneal dose

“…Iodine (30 mg, 0.24 mmol) in CH 2 Cl 2 (1.2 ml) was added to didemnin B (33 mg, 0.03 mmol) and dry silver trifluoroacetate (26.4 mg, 0.12 mmol) in dry methylene chloride (1.2 ml) [7]; stirred overnight; diluted with CH 2 Cl 2 ; the silver iodide was filtered; the filtrate was washed with 5% aqueous Na 2 Iododidemnin B (20 mg, 0.018 mmol) in dimethylformamide (0.6 ml) was reacted with 5% Pd/C (20 mg) and PtO 2 (7.0 mg) in a flask connected to a tritium source for 3 days; the mixture was diluted with ethanol (3 ml); filtered; and solvent removed via vacuum. The sample was purified on an ODS-HPLC with methanol/ H 2 O (83/17).…”