Fig. 1. Identifying when RDSuptake switches to RDSall, that is, the tipping point. (A) Extensive inhibition of CLmet + bile can lead to a significant increase in the systemic AUC for three theoretical victim drugs that have RDSuptake (i.e., CLmet + bile > > CLsef) in the absence of DDI. When inhibition of CLmet + bile eventually violates the condition required for RDSuptake, the RDSuptake switches to RDSall. An AUCR ≥1.25 was observed when CLmet + bile was inhibited ≥84%, ≥98%, and ≥99.8% for CLmet + bile = 1, 10, 100 liters/h, respectively. For all three victim drugs, however, the CLmet + bile value after such inhibition was similar (0.2 liters/min), as was the CLmet + bile/CLsef ratio (= 2). Simulations were performed as follows: CLsin = 1xQh, CLmet + bile = 1, 10, 100 liters/h, CLsef = 0.1 liters/min. (B) The systemic AUC (in the absence of any DDI) of a theoretical drug remains unchanged when the CLmet + bile/CLsef ratio remains fixed (blue bars) but not when the CLmet + bile/CLsef ratio is varied (yellow bars), even though the absolute value of CLmet + bile and CLsef is varied in both scenarios. This trend was observed irrespective of the value of CLsin (Supplementary Fig. 1). Furthermore, this trend is true for when CLmet + bile > CLsef or CLmet + bile < CLsef (also refer to Supplementary Fig. 1). Thus, the CLmet + bile/CLsef ratio, irrespective of the magnitude of the absolute values of these clearances, is important for establishing the RDS and henceforth when the RDS switches from uptake to all hepatobiliary clearances. Simulations were performed as follows: CLsin = 0.25×Qh, and the other input clearance values for scenarios A–E are shown in the table provided. (C) Since the RDS depends on the CLmet + bile/CLsef ratio, we define the tipping point as the CLmet + bile/CLsef ratio at which RDSuptake switches to RDSall. Similarly to (A), the RDSuptake switch to RDSall is represented by an AUCR of 1.25 and the decrease in CLmet + bile/CLsef ratio is akin to inhibition of CLmet + bile when CLsef is kept constant. As shown by the gray arrows, the tipping point for a low-, mid-, and high-ER drug is 3.2, 2, and 0.8, respectively. For example, if the CLmet + bile/CLsef ratio for the low ER drug is above the tipping point (i.e., CLmet + bile/CLsef > 3.2), then the drug will have RDSuptake; therefore, DDIs from CLsin but not CLmet + bile should be expected; however, inhibition of CLmet + bile that makes the CLmet + bile/CLsef ratio lower than the tipping point (i.e., CLmet + bile/CLsef < 3.2) will lead to a significant increase in the systemic AUC. Crossing the tipping point is indicative of the RDSuptake switch to RDSall. Simulations were performed as follows: systemic AUC were simulated for CLsin = 0.25×, 1×, 4 × Qh (representing low, mid, and high ER, respectively) and CLmet + bile/CLsef ratios from 1 to 10 and then normalized to a control simulation where the CLmet + bile/CLsef ratio was set to 1000 (i.e., RDSuptake).