Compensatory Paracrine Mechanisms That Define The Urothelial Response to Injury in Partial Bladder Outlet Obstruction

Diseases and conditions affecting the lower urinary tract are a leading cause of dysfunctional sexual health, incontinence, infection, and kidney failure. The growth, differentiation, and repair of the bladder's epithelial lining are regulated, in part, by fibroblast growth factor (FGF)-7 and -10 via a paracrine cascade originating in the mesenchyme (lamina propria) and targeting the receptor for FGF-7 and -10 within the transitional epithelium (urothelium). The FGF-7 gene is located at the 15q15-q21.1 locus on chromosome 15 and four exons generate a 3.852-kb mRNA. Five duplicated FGF-7 gene sequences that localized to chromosome 9 were predicted not to generate functional protein products, thus validating the use of FGF-7-null mice as an experimental model. Recombinant FGF-7 and -10 induced proliferation of human urothelial cells in vitro and transitional epithelium of wild-type and FGF-7-null mice in vivo.To determine the extent that induction of urothelial cell proliferation during the bladder response to injury is dependent on FGF-7, an animal model of partial bladder outlet obstruction was developed. Unbiased stereology was used to measure the percentage of proliferating urothelial cells between obstructed groups of wild-type and FGF-7-null mice. The stereological analysis indicated that a statistical significant difference did not exist between the two groups, suggesting that FGF-7 is not essential for urothelial cell proliferation in response to partial outlet obstruction. In contrast, a significant increase in FGF-10 expression was observed in the obstructed FGF-7-null group, indicating that the compensatory pathway that functions in this model results in urothelial repair

Safety and Pharmacokinetic Profiles of Phosphorodiamidate Morpholino Oligomers with Activity against Ebola Virus and Marburg Virus: Results of Two Single-Ascending-Dose Studies

Two identical single-ascending-dose studies evaluated the safety and pharmacokinetics (PK) of AVI-6002 and AVI-6003, two experimental combinations of phosphorodiamidate morpholino oligomers with positive charges (PMOplus) that target viral mRNA encoding Ebola virus and Marburg virus proteins, respectively. Both AVI-6002 and AVI-6003 were found to suppress disease in virus-infected nonhuman primates in previous studies. AVI-6002 (a combination of AVI-7537 and AVI-7539) or AVI-6003 (a combination of AVI-7287 and AVI-7288) were administered as sequential intravenous (i.v.) infusions of a 1:1 fixed dose ratio of the two subcomponents. In each study, 30 healthy male and female subjects between 18 and 50 years of age were enrolled in six-dose escalation cohorts of five subjects each and received a single i.v. infusion of active study drug (0.005, 0.05, 0.5, 1.5, 3, and 4.5 mg/kg per component) or placebo in a 4:1 ratio. Both AVI-6002 and AVI-6003 were safe and well tolerated at the doses studied. A maximum tolerated dose was not observed in either study. The four chemically similar PMOplus components exhibited generally similar PK profiles. The mean peak plasma concentration and area under the concentration-time curve values of the four components exhibited dose-proportional PK. The estimated plasma half-life of all four components was 2 to 5 h. The safety of the two combinations and the PK of the four components were similar, regardless of the target RNA sequence.Keywords: Marburg virus, Ebola virus, Single-ascending-dose studies, Phosphorodiamidate morpholino oligomer

A Single Phosphorodiamidate Morpholino Oligomer Targeting VP24 Protects Rhesus Monkeys against Lethal Ebola Virus Infection

Ebola viruses (EBOV) cause severe disease in humans and nonhuman primates with high mortality rates and continue to emerge in new geographic locations, including several countries in West Africa, the site of a large ongoing outbreak. Phosphorodiamidate morpholino oligomers (PMOs) are synthetic antisense molecules that are able to target mRNAs in a sequence-specific fashion and suppress translation through steric hindrance. We previously showed that the use of PMOs targeting a combination of VP35 and VP24 protected rhesus monkeys from lethal EBOV infection. Surprisingly, the present study revealed that a PMOplus compound targeting VP24 alone was sufficient to confer protection from lethal EBOV infection but that a PMOplus targeting VP35 alone resulted in no protection. This study further substantiates recent data demonstrating that VP24 may be a key virulence factor encoded by EBOV and suggests that VP24 is a promising target for the development of effective anti-EBOV countermeasures. IMPORTANCE: Several West African countries are currently being ravaged by an outbreak of Ebola virus (EBOV) that has become a major epidemic affecting not only these African countries but also Europe and the United States. A better understanding of the mechanism of virulence of EBOV is important for the development of effective treatments, as no licensed treatments or vaccines for EBOV disease are currently available. This study of phosphorodiamidate morpholino oligomers (PMOs) targeting the mRNAs of two different EBOV proteins, alone and in combination, demonstrated that targeting a single protein was effective at conferring a significant survival benefit in an EBOV lethal primate model. Future development of PMOs with efficacy against EBOV will be simplified if only one PMO is required instead of a combination, particularly in terms of regulatory approval

AVI-7288 for Marburg Virus in Nonhuman Primates and Humans

BACKGROUND: AVI-7288 is a phosphorodiamidate morpholino oligomer with positive charges that targets the viral messenger RNA that encodes Marburg virus (MARV) nucleoprotein. Its safety in humans is undetermined. METHODS: We assessed the efficacy of AVI-7288 in a series of studies involving a lethal challenge with MARV in nonhuman primates. The safety of AVI-7288 was evaluated in a randomized, multiple-ascending-dose study in which 40 healthy humans (8 humans per dose group) received 14 once-daily infusions of AVI-7288 (1 mg, 4 mg, 8 mg, 12 mg, or 16 mg per kilogram of body weight) or placebo, in a 3: 1 ratio. We estimated the protective dose in humans by comparing pharmacokinetic variables in infected nonhuman primates, uninfected nonhuman primates, and uninfected humans. RESULTS: Survival in infected nonhuman primates was dose-dependent, with survival rates of 0%, 30%, 59%, 87%, 100%, and 100% among monkeys treated with 0 mg, 3.75 mg, 7.5 mg, 15 mg, 20 mg, and 30 mg of AVI-7288 per kilogram, respectively (P < 0.001 with the use of the log-rank test for the comparison of survival across groups). No safety concern was identified at doses up to 16 mg per kilogram per day in humans. No serious adverse events were reported. Drug exposure (the area under the curve) was dose-dependent in both nonhuman primates and humans; drug clearance was independent of dose but was higher in nonhuman primates than in humans. The protective dose in humans was initially estimated, on the basis of exposure, to be 9.6 mg per kilogram per day (95% confidence interval, 6.6 to 12.5) for 14 days. Monte Carlo simulations supported a dose of 11 mg per kilogram per day to match the geometric mean protective exposure in nonhuman primates. CONCLUSIONS: This study shows that, on the basis of efficacy in nonhuman primates and pharmacokinetic data in humans, AVI-7288 has potential as postexposure prophylaxis for MARV infection in humans

IversenPatrickEnvirMoleToxAVI-7288MarburgVirus.pdf

BACKGROUND: AVI-7288 is a phosphorodiamidate morpholino oligomer with positive charges that targets the viral messenger RNA that encodes Marburg virus (MARV) nucleoprotein. Its safety in humans is undetermined. METHODS: We assessed the efficacy of AVI-7288 in a series of studies involving a lethal challenge with MARV in nonhuman primates. The safety of AVI-7288 was evaluated in a randomized, multiple-ascending-dose study in which 40 healthy humans (8 humans per dose group) received 14 once-daily infusions of AVI-7288 (1 mg, 4 mg, 8 mg, 12 mg, or 16 mg per kilogram of body weight) or placebo, in a 3: 1 ratio. We estimated the protective dose in humans by comparing pharmacokinetic variables in infected nonhuman primates, uninfected nonhuman primates, and uninfected humans. RESULTS: Survival in infected nonhuman primates was dose-dependent, with survival rates of 0%, 30%, 59%, 87%, 100%, and 100% among monkeys treated with 0 mg, 3.75 mg, 7.5 mg, 15 mg, 20 mg, and 30 mg of AVI-7288 per kilogram, respectively (P < 0.001 with the use of the log-rank test for the comparison of survival across groups). No safety concern was identified at doses up to 16 mg per kilogram per day in humans. No serious adverse events were reported. Drug exposure (the area under the curve) was dose-dependent in both nonhuman primates and humans; drug clearance was independent of dose but was higher in nonhuman primates than in humans. The protective dose in humans was initially estimated, on the basis of exposure, to be 9.6 mg per kilogram per day (95% confidence interval, 6.6 to 12.5) for 14 days. Monte Carlo simulations supported a dose of 11 mg per kilogram per day to match the geometric mean protective exposure in nonhuman primates. CONCLUSIONS: This study shows that, on the basis of efficacy in nonhuman primates and pharmacokinetic data in humans, AVI-7288 has potential as postexposure prophylaxis for MARV infection in humans

IversenPatrickEnvirMoleToxAVI-7288MarburgVirusSupplementaryProtocol.pdf

BACKGROUND: AVI-7288 is a phosphorodiamidate morpholino oligomer with positive charges that targets the viral messenger RNA that encodes Marburg virus (MARV) nucleoprotein. Its safety in humans is undetermined. METHODS: We assessed the efficacy of AVI-7288 in a series of studies involving a lethal challenge with MARV in nonhuman primates. The safety of AVI-7288 was evaluated in a randomized, multiple-ascending-dose study in which 40 healthy humans (8 humans per dose group) received 14 once-daily infusions of AVI-7288 (1 mg, 4 mg, 8 mg, 12 mg, or 16 mg per kilogram of body weight) or placebo, in a 3: 1 ratio. We estimated the protective dose in humans by comparing pharmacokinetic variables in infected nonhuman primates, uninfected nonhuman primates, and uninfected humans. RESULTS: Survival in infected nonhuman primates was dose-dependent, with survival rates of 0%, 30%, 59%, 87%, 100%, and 100% among monkeys treated with 0 mg, 3.75 mg, 7.5 mg, 15 mg, 20 mg, and 30 mg of AVI-7288 per kilogram, respectively (P < 0.001 with the use of the log-rank test for the comparison of survival across groups). No safety concern was identified at doses up to 16 mg per kilogram per day in humans. No serious adverse events were reported. Drug exposure (the area under the curve) was dose-dependent in both nonhuman primates and humans; drug clearance was independent of dose but was higher in nonhuman primates than in humans. The protective dose in humans was initially estimated, on the basis of exposure, to be 9.6 mg per kilogram per day (95% confidence interval, 6.6 to 12.5) for 14 days. Monte Carlo simulations supported a dose of 11 mg per kilogram per day to match the geometric mean protective exposure in nonhuman primates. CONCLUSIONS: This study shows that, on the basis of efficacy in nonhuman primates and pharmacokinetic data in humans, AVI-7288 has potential as postexposure prophylaxis for MARV infection in humans

IversenPatrickEnvirMoleToxAVI-7288MarburgVirusSupplementaryAppendix.pdf

BACKGROUND: AVI-7288 is a phosphorodiamidate morpholino oligomer with positive charges that targets the viral messenger RNA that encodes Marburg virus (MARV) nucleoprotein. Its safety in humans is undetermined. METHODS: We assessed the efficacy of AVI-7288 in a series of studies involving a lethal challenge with MARV in nonhuman primates. The safety of AVI-7288 was evaluated in a randomized, multiple-ascending-dose study in which 40 healthy humans (8 humans per dose group) received 14 once-daily infusions of AVI-7288 (1 mg, 4 mg, 8 mg, 12 mg, or 16 mg per kilogram of body weight) or placebo, in a 3: 1 ratio. We estimated the protective dose in humans by comparing pharmacokinetic variables in infected nonhuman primates, uninfected nonhuman primates, and uninfected humans. RESULTS: Survival in infected nonhuman primates was dose-dependent, with survival rates of 0%, 30%, 59%, 87%, 100%, and 100% among monkeys treated with 0 mg, 3.75 mg, 7.5 mg, 15 mg, 20 mg, and 30 mg of AVI-7288 per kilogram, respectively (P < 0.001 with the use of the log-rank test for the comparison of survival across groups). No safety concern was identified at doses up to 16 mg per kilogram per day in humans. No serious adverse events were reported. Drug exposure (the area under the curve) was dose-dependent in both nonhuman primates and humans; drug clearance was independent of dose but was higher in nonhuman primates than in humans. The protective dose in humans was initially estimated, on the basis of exposure, to be 9.6 mg per kilogram per day (95% confidence interval, 6.6 to 12.5) for 14 days. Monte Carlo simulations supported a dose of 11 mg per kilogram per day to match the geometric mean protective exposure in nonhuman primates. CONCLUSIONS: This study shows that, on the basis of efficacy in nonhuman primates and pharmacokinetic data in humans, AVI-7288 has potential as postexposure prophylaxis for MARV infection in humans

IversenPatrickEnvirMoleToxAVI-7288MarburgVirusSlideSet.pptx

BACKGROUND: AVI-7288 is a phosphorodiamidate morpholino oligomer with positive charges that targets the viral messenger RNA that encodes Marburg virus (MARV) nucleoprotein. Its safety in humans is undetermined. METHODS: We assessed the efficacy of AVI-7288 in a series of studies involving a lethal challenge with MARV in nonhuman primates. The safety of AVI-7288 was evaluated in a randomized, multiple-ascending-dose study in which 40 healthy humans (8 humans per dose group) received 14 once-daily infusions of AVI-7288 (1 mg, 4 mg, 8 mg, 12 mg, or 16 mg per kilogram of body weight) or placebo, in a 3: 1 ratio. We estimated the protective dose in humans by comparing pharmacokinetic variables in infected nonhuman primates, uninfected nonhuman primates, and uninfected humans. RESULTS: Survival in infected nonhuman primates was dose-dependent, with survival rates of 0%, 30%, 59%, 87%, 100%, and 100% among monkeys treated with 0 mg, 3.75 mg, 7.5 mg, 15 mg, 20 mg, and 30 mg of AVI-7288 per kilogram, respectively (P < 0.001 with the use of the log-rank test for the comparison of survival across groups). No safety concern was identified at doses up to 16 mg per kilogram per day in humans. No serious adverse events were reported. Drug exposure (the area under the curve) was dose-dependent in both nonhuman primates and humans; drug clearance was independent of dose but was higher in nonhuman primates than in humans. The protective dose in humans was initially estimated, on the basis of exposure, to be 9.6 mg per kilogram per day (95% confidence interval, 6.6 to 12.5) for 14 days. Monte Carlo simulations supported a dose of 11 mg per kilogram per day to match the geometric mean protective exposure in nonhuman primates. CONCLUSIONS: This study shows that, on the basis of efficacy in nonhuman primates and pharmacokinetic data in humans, AVI-7288 has potential as postexposure prophylaxis for MARV infection in humans

IversenPatrickEnvirMoleToxAVI-7288MarburgVirusSupplementaryDisclosures.pdf

BACKGROUND: AVI-7288 is a phosphorodiamidate morpholino oligomer with positive charges that targets the viral messenger RNA that encodes Marburg virus (MARV) nucleoprotein. Its safety in humans is undetermined. METHODS: We assessed the efficacy of AVI-7288 in a series of studies involving a lethal challenge with MARV in nonhuman primates. The safety of AVI-7288 was evaluated in a randomized, multiple-ascending-dose study in which 40 healthy humans (8 humans per dose group) received 14 once-daily infusions of AVI-7288 (1 mg, 4 mg, 8 mg, 12 mg, or 16 mg per kilogram of body weight) or placebo, in a 3: 1 ratio. We estimated the protective dose in humans by comparing pharmacokinetic variables in infected nonhuman primates, uninfected nonhuman primates, and uninfected humans. RESULTS: Survival in infected nonhuman primates was dose-dependent, with survival rates of 0%, 30%, 59%, 87%, 100%, and 100% among monkeys treated with 0 mg, 3.75 mg, 7.5 mg, 15 mg, 20 mg, and 30 mg of AVI-7288 per kilogram, respectively (P < 0.001 with the use of the log-rank test for the comparison of survival across groups). No safety concern was identified at doses up to 16 mg per kilogram per day in humans. No serious adverse events were reported. Drug exposure (the area under the curve) was dose-dependent in both nonhuman primates and humans; drug clearance was independent of dose but was higher in nonhuman primates than in humans. The protective dose in humans was initially estimated, on the basis of exposure, to be 9.6 mg per kilogram per day (95% confidence interval, 6.6 to 12.5) for 14 days. Monte Carlo simulations supported a dose of 11 mg per kilogram per day to match the geometric mean protective exposure in nonhuman primates. CONCLUSIONS: This study shows that, on the basis of efficacy in nonhuman primates and pharmacokinetic data in humans, AVI-7288 has potential as postexposure prophylaxis for MARV infection in humans