83 0.06 13.8 86 ± 3 16 2 42.5 24 1.5 0.04 37.5 89 ± 3 19 3 21 29 0.8 0.07 11.4 85 ± 3 15 aBuffer solution: 10 mM HEPES, 200 mM

KCl, 3 mM EDTA and 0.01% P20 surfactant with the final pH adjusted to 7.4. bHuman telomeric sequence 5′-d[AGGG(TTAGGG)3]-3′. c5′-d[CGA3T3C(CT)2GA3T3CG]-3′ hairpin sequence. dThermal stability data for h-Tel (anti-parallel) determined by CD in the absence and presence of compounds. eTm for unaligned h-Tel = 70 ± 3. MI-503 Ligand redesign to minimize off target effects The potent hERG inhibition compromised the acceptability of 1 as a clinical candidate, despite this agent having many of the attributes of an ideal pharmaceutical [28]. Two strategies have been adopted in an attempt to minimize the hERG interaction: (i) sterically masking the (delocalized) positive charge on the acridinium cation by increasing the size of the substituent at position 13 as in compound 8; and (ii) evaluating compounds 2 and 3 as prototypes of two series of isomeric pentacyclic acridinium salts of the same chemotype as 1. hERG tail Selleckchem Staurosporine current inhibition was used as a marker of potential off-target liabilities. The prototypic agent 1 potently inhibited hERG by 100% at 10 μM (IC50 0.2 μM) (Table  1); inhibition of hERG was reduced to 43% at 10 μM (IC50 3.7 μM) in the 2-acetylaminoquinoacridinium iodide 2 and to 18% by 13-ethyl

homologue 8, while the least potent hERG inhibitor (IC50 18 μM) was the 3-acetylamino isomer 3, a 90-fold improvement over 1. The marked improvement of 8 over 1, was paralled by a >10-fold reduction in the on-target

effect against the h-Tel DNA sequence as measured by surface plasmon resonance (see below) suggesting that increasing the size of the onium head was not a fruitful developmental approach, for these reason the compound 8 was excluded from further studies. The interaction with β2-adrenergic receptor was determined by a binding assay of 1, 2 and 3 to the transgenic β2-adrenegic receptor expressed on the surface of CHO cells. Inhibition of receptor was reported as inhibition of control specific binding (100 – (measured specific binding/control specific binding) × 100) obtained in the presence of Urocanase the test compounds. A decay of 75% and 70% of receptor inhibition is observed comparing 1 to 2 and 3 compounds respectively (Table  1). These results indicate that potential toxicities in this chemotype, as predicted by hERG and β2-adrenergic receptor interactions, can be addressed by suitable molecular modification. On target-effects: ligand-quadruplex interactions The Surface Plasmon Resonance (SPR) technique is a powerful tool to compare binding affinities for G-quadruplex binding agents [11, 29]. When the h-Tel DNA sequence comprising 5′-d[AGGG(TTAGGG)3]-3′ is immobilised on a sensor chip surface, binding of drug elicits a refractive index change at the surface, and hence the refractive light angle at which SPR is observed.