The genetic disease cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR), a Cl- channel with complex regulation that plays a central role in transepithelial salt and water movement. The commonest CF mutation, F508del-CFTR disrupts CFTR-mediated transepithelial ion transport by preventing the delivery of CFTR protein to the apical membrane of epithelial cells and perturbing channel gating. In Portugal, the second most common CF mutation is A561E-CFTR [1]. Like F508del-CFTR, A561E-CFTR is located in the first nucleotide-binding domain of CFTR, where it disrupts CFTR processing and trafficking [1]. To restore function to CF mutants, small molecules have been developed that rescue the cell surface expression (termed CFTR correctors) and function (termed CFTR potentiators) of CF mutants. In this study, we investigated the rescue of A561E-CFTR by the 2-quinazolinyl-4-aminopyrimidinone CFTR corrector corr-3a [2] and the tetrahydrobenzothiophene CFTR potentiator ΔF508act-02 [3]. We used baby hamster kidney (BHK) cells expressing high levels of A561E-CFTR and the iodide efflux assay to study the activity of a large number of CFTR Cl- channels in intact cells. As controls, we studied BHK cells expressing wild-type human CFTR (WT-CFTR) and the F508del-CFTR mutation (F508del-CFTR). We measured iodide efflux as described in Schmidt et al [4]. At 37 °C, the cAMP agonist forskolin (10 μM) and the CFTR potentiator genistein (50 μM) stimulated a large transient efflux of iodide from WT-CFTR cells, but did not have any effect on either A561E-CFTR or F508del-CFTR cells. Incubation of BHK cells expressing F508del- or A561E-CFTR at 27 °C for 24 hrs restored CFTR-mediated iodide efflux to both CF mutants. Although less efficacious than low temperature correction, incubation of BHK cells with the CFTR corrector corr-3a (20 μM) at 37 °C for 24 hrs rescued both F508del- and A561E-CFTR function. Moreover, after low temperature correction, the CFTR potentiator ΔF508act-02 (10 μM) enhanced the Cl- channel activity of both A561E- and F508del-CFTR cells in the presence of forskolin (10 μM) (p < 0.05, Student’s unpaired t-test vs. vehicle), albeit much less potently than genistein (50 μM). We interpreted our data to suggest that the function of the CF mutant A561E is enhanced by small-molecules which rescue F508del-CFTR. However, the pharmacology of A561E-CFTR requires further investigation to identify small-molecules suitable for therapy development. * L Bugeja and R Warner contributed equally to this work.