Effect of hydroxychloroquine and azithromycin on cardiac repolarization in iPSC cardiomyocytes
Data from our screen of hERG potency showed only a subtle (hydroxychloroquine), or non-significant (chloroquine), additive effect of azithromycin (Supplementary figure 2). In order to measure the effects of the combination of drugs directly on cardiac repolarisation, as well as to assess whether there are any potentially proarrhythmic effects that manifest with longer term exposure, we measured field potential duration, a surrogate of the QT interval, in hiPSC derived cardiomyocytes from three normal patients. Representative field potentials for hydroxychloroquine alone (Figure 3A) and in combination with azithromycin (Figure 3B) are shown for (i) baseline; (ii), acutely after drug addition; (iii) 24 hrs and (iv) 48 hrs after drug addition. Summary data for all three lines over time is presented in Figure 3C (individual technical replicates for each cell line are presented in Supplementary Figure 4). There was no significant difference in the extent of prolongation of repolarisation between hydroxychloroquine alone, or in combination with azithromycin (Repeated measures two-way ANOVA; p = 0.109). However, there was a significant time dependent effect (Repeated measures two-way ANOVA; p = 0.027), indicating an accumulation of block with longer term exposure to the drugs. Additionally, hydroxychloroquine and the combination of hydroxychloroquine and azithromycin caused cessation of beating or loss of capture in multiple wells (Table 2). This is consistent with previous reports showing that profound drug-induced disruption of repolarisation, corresponding to ~ 75 % or more block of IKr, can result in cessation of beating or occurrence of arrhythmia (23). Next, we assessed the effect of kalaemic variation on prolongation of repolarisation by hydroxychloroquine. 10 µM hydroxychloroquine caused significantly more prolongation in hypokalaemia (3 mM K+, 7.7 ± 1.1 % prolongation (SEM; n=16)) than normokalaemia (5.4 mM K+, 3.3 ± 0.5 % prolongation (SEM; n=17)) demonstrating that the hypokalaemia-dependent increase in potency of hydroxychloroquine against hERG, as measured in our Syncropatch screens (Figure 2), manifests in a more severe prolongation of cardiac repolarisation.