Development of a method for the direct analysis of peptide AM336 in monkey cerebrospinal fluid using liquid chromatography/electrospray ionization mass spectrometry with a mixed-function column.

A liquid chromatography/mass spectrometry (LC/MS) analytical procedure, using a single column for sample clean-up, enrichment and separation, has been developed for the determination of the peptide AM336 in monkey cerebrospinal fluid (CSF). CSF samples were injected and analyzed using a polymer-coated mixed-function high-performance liquid chromatography (HPLC) column with gradient elution and application of a timed valve-switching event. The mass spectrometer was operated in the positive electrospray ionization (ESI(+)) mode with single ion recording (SIR) at m/z 920. The method was validated, yielding calibration curves with correlation coefficients greater than 0.9892. Assay precision and accuracy were evaluated by direct injection of AM336-fortified CSF samples at three concentration levels. Analyzed concentrations ranged from 99.93 to 113.1% of their respective theoretical concentrations with coefficients of variation below 9.0%. An evaluation of the signal-to-noise (S/N) ratio for a 200 ng/mL calibration standard, considered to be the lower limit of quantitation (LLOQ), resulted in an estimated limit of detection (LOD) of 31.2 ng/mL. Preliminary data suggest the possibility of using this method to analyze AM336 also in plasma samples, pending the successful outcome of additional investigations.

The effect of leukaemia inhibitory factor on SOD1 G93A murine amyotrophic lateral sclerosis.

Before potential therapeutic strategies for the treatment of amyotrophic lateral sclerosis (ALS) can be advanced to human clinical trials, there is a need to assess them in an animal model that best resembles the disease process. SOD1 G93A mice have close resemblance to familial ALS (fALS) and have been used in this study to evaluate the therapeutic potential of leukaemia inhibitory factor (LIF). LIF action was investigated by assessing three delivery methods: (1) daily subcutaneous injection; (2) through LIF rods placed adjacent to hind limb skeletal muscle and (3) continuous intrathecal infusion. The effect on disease progression was assessed by semi-quantitative and quantitative functional measurements, and histologically on the survival of motor neurons and number of reactive astrocytes. The results show that LIF had no beneficial effects when administered using the three methods of drug delivery. These results suggest that further evaluation of LIF in this transgenic model is required to fully characterize its' therapeutic potential.

Behavioural and anatomical effects of systemically administered leukemia inhibitory factor in the SOD1(G93A G1H) mouse model of familial amyotrophic lateral sclerosis.

We investigated the anatomical and behavioural effects of daily intraperitoneal injection of 25 microg/kg of LIF in the SOD1(G93A G1H) mouse model of familial ALS. We found some subtle beneficial behavioural changes in LIF treated mice. These included later onset of clinical disease in females as determined by clinical scoring; better grip strength in males; and delayed development of motor impairment in males as determined by the rotarod test. However, we found no significant rescue of motoneurons or prolongation of survival as a result of this systemic dose of LIF in these mice.