IND Enabling Studies II: Toxicity and CMC

Which specific toxicities are mechanistic studies used to determine if potential drugs might cause?

• membrane integrity

• mitochondrial toxicity (MitoTox)

• oxidative stress

• inflammation

How do mutation assays for genotoxicity work?

Ames test first → secondary follow-up tests performed in mammalian cells as needed

• HPRT assay

• Mouse lymphoma TK assay (MLA)

What are the two “in vivo” toxicity assays?

acute and chronic toxicity

acute toxicity

single dose studies over a broad concentration range to identify dose-limiting toxicities and specific organ toxicities

maximum tolerated dose (MTD)

highest dose that doesn’t cause unacceptable side effects

chronic toxicity

lower dose levels tested over longer period of time; usually daily administration for 14-28 days but dose and length can depend

For both acute and chronic tests:

animals are sacrificed and their tissues are studied for damage; often involves at least two different species (rodent + non-rodent); tested through at least two routes of administration

lethal dose 50 (LD₅₀)

dose required to kill 50% of animal group

effective dose 50 (ED₅₀)

dose required to produce desired effect in 50% of animals

therapeutic index

calculated as the ratio of LD₅₀ divided by ED₅₀

safety pharmacology

series of “in vitro” and “in vivo” studies designed to identify any undesired effects of potential drug on key organs

What tests would be run on the cardiovascular system for safety pharmacology?

• blood pressure

• heart rate

• ECGs

• ventricular vs pulmonary pressure

• echocardiography

• cardiac ion channel assessments

What tests would be run on the respiratory system for safety pharmacology?

• respiratory rate

• hemoglobin oxygen saturation

• tidal and minute volumes

What tests would be run on the CNS for safety pharmacology?

• functional observation battery

• modified Irwin screen

• body temperature

• large animal neurological evaluations

Why do we want to reduce reliance on animal models for data?

• don’t always do the best job of predicting human efficacy/toxicity

• expensive and time consuming

• can’t replicate individual genetics or specific human disease parameters

• ethical concerns

organ-on-a-chip

microfluidic devices lined with living human cells that simulate the structure and function of human organs

• single or multiple organs

• specific stimuli to determine effects of certain conditions

Why do we use organ-on-a-chip?

to understand metabolism and toxicity (liver), cardiotoxicity, and lung absorption/toxicity

3D cell culture models

spheroids or organoids; readily mimics natural environment of an organ and can be used to test both efficacy and toxicity

physiologically based pharmacokinetic (PBPK) models

math-based models systems designed to predict ADME/PK properties of a small molecule

• utilize known info about human body and parameters of drug to predict ADME/PK

chemistry, manufacturing and controls (CMC)

series of studies to ensure that final drug product maintains the same structure, quality, and consistency across each batch prepared prior to dosing humans

API analytical characterization studies

clearly demonstrate that the API is the correct structure and identify any impurities during the manufacturing process

analysis of formulated dosage form

identifies what the components and stability is of the final dosage form

manufacturing process and development and validation

ensures that the process complies with good manufacturing practices (GMP), is scalable, and will consistently produce the drug

API structural characterization

full suite of analytical techniques (NMR/IR/MS) to clearly demonstrate the structure of the API

impurity profile

list of any organic/inorganic impurities generated from either the API synthesis or generated from the formulated drug

synthetic/process route

helps identify potential impurities while also ensuring safety and reproducibility

solid form ID

process used and final identification of API solid form for dosing

Name the solid forms that an API can make:

• crystals (polymorphs)

• non-crystals (amorphous solids)

• salts (charged)

• hydrates

formulated drug analysis

provides full characterization of the API, excipients, contain-closure system, and stability studies

container-closure system

combo of packaging components for final drug; primary (vials/bottles) and secondary (boxes/labels)

real-time stability

how stable the formulated drug is over time for the recommended shelf life under ideal conditions

accelerated stability

elevate normal conditions to determine potential stability issues (temperature/humidity)

forced degradation

a measure of stability and byproducts associated with harsh conditions (high heat/light/pH)

good manufacturing practices (GMP)

specific parameters enforced by FDA to guarantee safety/quality of drugs through consistent production across different batches