Addition of an increasing concentration of tetrabutylammonium (TBA) chloride to a 1.5 m M solution of compound 16⋅PF6 in [D6]DMSO induced progressive shifts in the H NMR signals for protons 14, 15 and c, which is consistent with fast-exchange complexation of the chloride anion within the [2]catenane's interlocked binding domain.
For this purpose 10−2 M solution of each reactant containing (1) and N H compound and 5 × 10−3 M of compound (2) were prepared in methanol solvent.
After 24 h, they were treated with 0.5 μ M of compound.
After 24 h, they were treated with 0.5 μ M of compound for a further 24 h.
In contrast, TAT-S-S-Life significantly affects platelet viability; even at relatively low concentrations (1 2 μ m) of compound, more than 50%% of cells are incorrectly spread on fibrinogen.
The biological activity of compound 1 was assessed in a multiparametric phenotypic assay on non-transformed human olfactory neurosphere-derived cells (hONS), which model functional aspects of Parkinson's disease.[ 7] The hONS cells were cultured in 384-well plates to 1350 cells per well, and treated with 1 μ m of compound 1 for 24 h.
An additional single-point analysis of the inhibitory activity of 10 μ M of compounds YA's 1, 2, 3 and 4 against p38, HGK, Aurora A, and ROCKs 1 and 2 confirmed that YA2 had the greatest activity against ROCK's 1 and 2. YAs1, 3 and 4 all have significant activity against p38, HGK and Aurora A, but show less activity against ROCKs than YA2.
These assays yielded dissociation constants for Ubc13 of 4.4×10−12 M for compound Ia and of 4.68×10−7 M for compound IIa (Figure 3C).
SPR determinations yielded a IC50 of 1.0×10−11 M for compound Ia, and of 1.1×10−6 M for compound IIa (Fig. 3B), indicating a significantly more effective inhibition of the Ubc13-Uev1 interaction by compound Ia.
The HREI-MS of compound 3 showed the M+ at m/z 304.2113, corresponding to the formula C20H32O2 (Calcd. 304.2115).
