Nerve growth factor inhibition with tanezumab influences weight-bearing and subsequent cartilage damage in the rat medial meniscal tear model.

OBJECTIVE: To investigate whether the effects of nerve growth factor (NGF) inhibition with tanezumab on rats with medial meniscal tear (MMT) effectively model rapidly progressive osteoarthritis (RPOA) observed in clinical trials. METHODS: Male Lewis rats underwent MMT surgery and were treated weekly with tanezumab (0.1, 1 or 10 mg/kg), isotype control or vehicle for 7, 14 or 28 days. Gait deficiency was measured to assess weight-bearing on the operated limb. Joint damage was assessed via histopathology. A second arm, delayed onset of treatment (starting 3-8 weeks after MMT surgery) was used to control for analgesia early in the disease process. A third arm, mid-tibial amputation, evaluated the dependency of the model on weight-bearing. RESULTS: Gait deficiency in untreated rats was present 3-7 days after MMT surgery, with a return to normal weight-bearing by days 14-28. Prophylactic treatment with tanezumab prevented gait deficiency and resulted in more severe cartilage damage. When onset of treatment with tanezumab was delayed to 3-8 weeks after MMT surgery, there was no increase in cartilage damage. Mid-tibial amputation completely prevented cartilage damage in untreated MMT rats. CONCLUSIONS: These data suggest that analgesia due to NGF inhibition during the acute injury phase is responsible for increased voluntary weight-bearing and subsequent cartilage damage in the rat MMT model. This model failed to replicate the hypotrophic bone response observed in tanezumab-treated patients with RPOA.

Intrathecal delivery of frataxin mRNA encapsulated in lipid nanoparticles to dorsal root ganglia as a potential therapeutic for Friedreich's ataxia.

In Friedreich's ataxia (FRDA) patients, diminished frataxin (FXN) in sensory neurons is thought to yield the predominant pathology associated with disease. In this study, we demonstrate successful usage of RNA transcript therapy (RTT) as an exogenous human FXN supplementation strategy in vitro and in vivo, specifically to dorsal root ganglia (DRG). Initially, 293 T cells were transfected with codon optimized human FXN mRNA, which was translated to yield FXN protein. Importantly, FXN was rapidly processed into the mature functional form of FXN (mFXN). Next, FXN mRNA, in the form of lipid nanoparticles (LNPs), was administered intravenously in adult mice. Examination of liver homogenates demonstrated efficient FXN LNP uptake in hepatocytes and revealed that the mitochondrial maturation machinery had efficiently processed all FXN protein to mFXN in ~24 h in vivo. Remarkably, greater than 50% mFXN protein derived from LNPs was detected seven days after intravenous administration of FXN LNPs, suggesting that the half-life of mFXN in vivo exceeds one week. Moreover, when FXN LNPs were delivered by intrathecal administration, we detected recombinant human FXN protein in DRG. These observations provide the first demonstration that RTT can be used for the delivery of therapeutic mRNA to DRG.

14-deoxyandrographolide as a platelet activating factor antagonist in bovine neutrophils.

14-Deoxyandrographolide (14-DAP) is a labdane diterpene isolated from Andrographis paniculata with previously reported calcium channel blocking activity. Its potential platelet activating factor (PAF) antagonistic activity in bovine neutrophils was assessed. 14-DAP, in concentrations between 10-100 microM, reduced the extracellular acidification rate and the intracellular alkalinization in a dose-dependent manner. In addition, 14-DAP reduced PAF-induced calcium flux in the presence of extracellular calcium, and tyrosine phosphorylation of a 44 kDa protein corresponding to the MAPK(ERK1). However, 14-DAP reduced the 3H-PAF binding with a Ki of 7.8 x 10 (- 9)M, and a Hill slope of 0.63, suggesting that there is more than one binding site for 14-DAP. We concluded that 14-DAP is an effective antagonist of PAF-mediated processes in bovine neutrophils, probably by virtue of its calcium channel blocking property.

Effect of 14-deoxyandrographolide on calcium-mediated rat uterine smooth muscle contractility.

In this study, the effect of 14-deoxyandrographolide (14-DAP) on calcium channel-dependent rat uterine smooth muscle contraction was evaluated. Using a tissue bath preparation, 14-DAP was able to reduce the contractile response to 0.3 and 3.0 mm of CaCl(2), with an IC(50) of 1.24 +/- 0.23 x 10(-5) m and 5.94 +/- 0.29 x 10(-5) m, respectively. 14-DAP shifted the CaCl(2) cumulative dose response curve to the right, increasing the EC(50) from 2.08 +/- 0.20 x 10(-4) m to 4.22 +/- 0.22 x 10(-4) m (5 micrometer 14-DAP) and 2.5 +/- 1.0 x 10(-3) m (50 micrometer 14-DAP). In order to determine if 14-DAP had any effect on intracellular calcium, the relaxant response to 14-DAP following contraction by oxytocin, PGF(2alpha) and vanadate in Ca(+2)-free solution was compared with that of isoproterenol and phenylbutazone. While isoproterenol and phenylbutazone relaxed the smooth muscle in a dose-dependent manner, 14-DAP did not have any effect on either the oxytocin, PGF(2alpha) or vanadate-induced smooth muscle contraction. Based on these data, it appears that 14-DAP is an uterine smooth muscle relaxant which produces a selective blockade of voltage operated calcium channels.