IC50

Understanding IC50

IC50 represents the midpoint of an inhibition curve:

Activity (%)
100 |___
    |   \
 75 |    \
    |     \
 50 |------*-------- IC50 point
    |       \
 25 |        \
    |         \___
  0 |________________
       [Inhibitor concentration]

Key Points

• Lower IC50 = More potent inhibitor (less drug needed)
• Measured in concentration units (nM, uM, ng/mL)
• Specific to the target being inhibited (enzyme, receptor, pathway)
• Dependent on assay conditions (substrate concentration, incubation time)

IC50 in Context

Related Inhibition Measures

Measure	Definition	Advantage
IC50	Concentration for 50% inhibition	Easy to measure
Ki	Inhibition constant	Independent of substrate
IC90	Concentration for 90% inhibition	More relevant for clinical efficacy
Selectivity ratio	IC50 target / IC50 off-target	Measures specificity

IC50 vs Ki

• IC50 depends on assay conditions (substrate concentration)
• Ki is an intrinsic property of the inhibitor-enzyme pair
• Cheng-Prusoff equation relates them: Ki = IC50 / (1 + [S]/Km)

Types of Inhibition

Competitive Inhibitors

Characteristic	Effect
Binds active site	Competes with substrate
IC50 behavior	Increases with substrate concentration
Can be overcome	Yes, with high substrate

Non-competitive Inhibitors

Characteristic	Effect
Binds allosteric site	Doesn’t compete with substrate
IC50 behavior	Independent of substrate concentration
Can be overcome	No

IC50 in Peptide Research

DPP-4 Inhibitors

DPP-4 enzyme rapidly degrades GLP-1 and GIP. Inhibitors protect these incretins:

Inhibitor	IC50	Notes
Sitagliptin	~18 nM	First approved DPP-4 inhibitor
Vildagliptin	~3 nM	More potent
Saxagliptin	~1 nM	Most potent
Linagliptin	~1 nM	Highly potent

Clinical implication: All achieve near-complete DPP-4 inhibition at therapeutic doses despite potency differences.

ACE Inhibitors

Angiotensin-converting enzyme inhibitors for cardiovascular disease:

Inhibitor	IC50 Range	Notes
Enalaprilat	~1-5 nM	Active form of enalapril
Lisinopril	~1-2 nM	No prodrug conversion needed
Captopril	~5-20 nM	First ACE inhibitor

Selectivity and IC50

Calculating Selectivity

Selectivity ratio = IC50 (off-target) / IC50 (target)

Drug	Target IC50	Off-target IC50	Selectivity
Selective	1 nM	1000 nM	1000x
Non-selective	1 nM	10 nM	10x

Higher selectivity ratios indicate less off-target activity and potentially fewer side effects.

Multi-target Inhibitors

Some drugs intentionally inhibit multiple targets:

Target	IC50	Therapeutic Role
Target A	5 nM	Primary effect
Target B	10 nM	Secondary benefit
Target C	500 nM	Minimal activity

Practical Considerations

What IC50 Doesn’t Tell You

• In vivo efficacy - Tissue penetration and metabolism matter
• Duration of effect - Binding kinetics (on/off rates)
• Mechanism - Competitive vs non-competitive
• Reversibility - Covalent vs non-covalent binding

From IC50 to Clinical Dose

IC50 → Required tissue concentration → Account for PK → Clinical dose
        (usually 5-10x IC50            (absorption,
         for near-complete              distribution,
         inhibition)                    metabolism)

Frequently Asked Questions

Why do some drugs need higher concentrations than their IC50?

IC50 represents 50% inhibition. For therapeutic effect, you often need 90-99% inhibition, requiring concentrations 10-100x higher than IC50 depending on the steepness of the inhibition curve.

Can IC50 change?

IC50 is measured under specific assay conditions. It can appear to change with different substrate concentrations (competitive inhibitors), temperature, pH, or assay methodology. Intrinsic Ki is more consistent.

How does resistance affect IC50?

In infectious disease and oncology, resistance mutations often increase IC50 dramatically. A 10-fold increase in IC50 may render previously effective concentrations inadequate, requiring higher doses or alternative drugs.