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Baseline input values are literature values (Heuser et al., 1993; Van Dyke, 1993; Gambale et al., 1994; Sonawane et al., 2002; Alberts, 2008) representing physiological lysosomes and are in agreement with previously published model (Grabe and Oster, 2001; Ishida et al., 2013). *estimated intrinsic surface potentials for inner CWin) and outer CW out) leaflets of the lysosomal membrane accounted for 

when modeling membrane transporter mediated dynamic lysomal and cytoplasmic ion concentration at the membrane surface (Grabe and Oster, 2001). 

[0159] Simulating lysosomal CFZ-HCl accumulation. To simulate the dose-dependent lysosomal CFZ-HCl accumu› lation, the rates of proton and chloride sequestration by CFZ were adjusted to either 0.01 or 0.1 picomoles/cell/day, which equal to 1.2x10- 21 moles/lysosome/day and 1.2x10- 20 moles/lysosome/day, respectively, as calculated by assum› ing there are approximately 100 lysosomes in a cell. Inter› changeably from here onwards, these rates; 0.01 and 0.1 picomoles/cell/day, are referred to as I-fold (lx) and 10-fold (lOx) CFZ-HCl accumulation, respectively. 

[0160] Simulating the inhibition oflysosomal proteins and cytoplasmic chloride. In order to study the role ofV-ATPase, CLC7, and cytoplasmic chloride concentration on the lyso› somal accumulation of CFZ-HCl, the total number of active V-ATPase and CLC7 molecules per lysosome, as well as the cytoplasmic chloride concentration were varied, while fixing the values of other lysosomal parameters at their baseline physiological input values in the presence and absence of lysosomal CFZ-HCl accumulation. More specifically, to simulate the simultaneous inhibition of V-ATPase and CLC7, the total number of CLC7 molecules per lysosome was varied from O to 5000 in geometric interval of 2.09 while one simulation at a time the total number ofV-ATPase molecules per lysosome was manually varied from O to 300 in arbitrary intervals. Moreover, to simulate the simultane› ous inhibition ofV-ATPase and cytoplasmic chloride, cyto› plasmic chloride concentration was varied from O to 10 mM in arithmetic intervals of 6.67xl 0-4 while one simulation at a time the total number of V-ATPase molecules per lyso› some was manually varied from O to 300 in arbitrary intervals. 

[0161] Using the aforementioned ranges of the adjustable lysosomal parameters, the corresponding inhibition range of 0 to 100% was calculated; where 0% represents no change from respective physiological baseline input value whereas 100% represents maximum change from respective physi› ological baseline input value. The inhibition range was calculated as follows by comparing its corresponding input value from the aforementioned given range (Adjusted Input Value) with its respective physiological input value (Base› line Input Value). 

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Calculating the effect of CFZ-HCl accumulation on lyso› somal ion homeostasis. The set of differential equations detailed earlier (equations 27-29) was solved by numerical integration in Berkeley Madonna (BM) using Rosenbrock stiff solver as numerical integrator. Upon performing para› metric simulations in the absence and presence ofCFZ-HCl accumulating at either 1 x or 1 Ox, along with different ranges of lysosomal parameters such as V-ATPase, CLC7, and cytoplasmic chloride (as elaborated earlier), final lysosomal pH, chloride and membrane potential were chosen as read› out values. The readout values in the absence of CFZ-HCl were subtracted from the ones in the presence of CFZ-HCl to determine the effect oflysosomal CFZ-HCl accumulation on lysosomal ion homeostasis represented by the changes in lysosomal pH, Cl, and membrane potential. 

[0162] Calculating the effects of V-ATPase, CLC7, and cytoplasmic chloride on the physiological lysosomal accu› mulation of CFZ-HCI. Time-plot simulations were per› formed to obtain final lysosomal pH, chloride, and mem› brane potential values (specifically chosen because they are direct indicators of lysosomal ion homeostasis and physiol› ogy) while adjusting all of the lysosomal parameters to their respective baseline values in the absence of CFZ-HCl accu› mulation. This allowed us to obtain physiological lysosomal pH of 4.53, chloride concentration of 224.6 mM, and membrane potential of 0.79 m V, which from here onwards thes values are referred to as "physiological baseline readout values" as they are associated with no changes to physi› ological baseline lysosomal parameters and contents. 

[0163] Moreover, in the presence ofCFZ-HCl accumulat› ing at 1 x and 1 Ox, final lysosomal pH, Cl and membrane potential values were chosen as readout values while per› forming parametric simulation runs in the presence of simul› taneous V-ATPase-CLC7 and V-ATPase-cytoplasmic chlo› ride inhibitions. These values were then subtracted from the physiological baseline readout values to calculate the effects of the simultaneous lysosomal parameter inhibitions on dose-dependent lysosomal CFZ-HCl accumulation. 

[0164] Steady state and mass balance check. For all of the aforementioned simulations, the final readout values were confirmed that they were steady state values by performing the simulation for >24 hrs. Moreover, mass balance was maintained in all of the simulations in the presence as well as absence ofCFZ-HCl accumulation as long as physiologi- 

