Table 2.

Increasing complexity of measurements for assessing new compounds in vitro

Target	Parameters	Effects
Receptor (R)	$L+R\underset{k_{\text{off}}}{\overset{k_{\text{on}}}{\rightleftarrows }}LR\to \text{conformational change}$	Multiple effects possible: on, off, partial, etc
Enzyme (E)	Reversible $L+E\underset{k_{\text{off}}}{\overset{k_{\text{on}}}{\rightleftarrows }}\text{LE}$ Irreversible $L+E\underset{k_{\text{off}}}{\overset{k_{\text{on}}}{\rightleftarrows }}\text{LE}\stackrel{k_{\text{inact}}}{\to }L-E$	Decreases product, prevents depletion of substrate but can be out-competed by substrate Inactivated enzyme; recovery depends on normal turnover of enzyme (minutes to days)
Protein or DNA (M)	$L+M\underset{k_{\text{off}}}{\overset{k_{\text{on}}}{\rightleftarrows }}LM$	Ligand and target often at same concentration in vitro. Outcome of binding best measured downstream
Cell target (T)	$L_{\text{out}}\stackrel{\text{cell uptake}}{\to }{L}_{\text{in}}\stackrel{\text{metabolism?}}{\to }\frac{\text{Organelle uptake}}{\text{Other binding}}\to L+T\underset{k_{\text{off}}}{\overset{k_{\text{on}}}{\rightleftarrows }}\text{LT}$	Concentration at target site unknown Major effects often easy to assess (e.g., cell death) but mechanism can be obscure