toxicological studies of impurities and degradation...

Toxicological studies of impurities and

degradation products: in silico Methodologies as a safety assessment tool

Azeddine Elhajouji, Novartis Institutes for Biomedical Research, Basel, Switzerland

22-Jun-2016

3 | V Simpósio SINDUSFARMA - IPS/FIP - ANVISA| Azeddine Elhajouji| 22-Jun-2016 | Impurities and degradation products

Impurities, solvents and heavy metals in drugs… Definitions (according to ICH)

Impurity Accompanying substance that is not identical with the structure of the active ingredient (by-products, degradation products, residual solvents, etc.)

By-product Starting materials, intermediates, reagents, products of side reactions

Degradation product

Substance formed by chemical reactions of drug substance or by-products during processing and/or storage

Impurity profile

Description of identified/unidentified impurities in the drug substance

Qualification

of impurities

The process of acquiring & evaluating data that establishes the biological safety of an impurity or an impurity profile at the level(s) specified


Impurities in General

• Impurities in drug substances and drug products give no benefit to patients and if not tightly controlled, may be hazardous

• Impurities must be reduced to levels ‘As Low As Reasonably Practicable’ (ALARP)

• Limits should be based on toxicological reasoning in congruence with analytical and technical capabilities


Genotoxic Impurities (GTIs)

• Genotoxic impurities (i.e., potentially carcinogenic impurities with a genotoxic mechanism of action) deserve special attention in pharmaceuticals

• Impurities in pharmaceuticals require higher safety margins and lower limits than most other types of impurities

• Accepted modern toxicological risk assessment implies that the risk stemming from exposure to genotoxic carcinogens is a function of dose and length of exposure


Impurities, solvents and heavy metals in drugs… Which guidance to follow?

• ICH Q 3 A (R2) Impurities in New Drug Substances CPMP/ICH/2737/99 Rev. 2 Aug 2002

• ICH Q 3 B (R2) Impurities in New Drug Products CPMP/ICH/2738/99 Aug 2003

• ICH Q 3 C (R3) Impurities: Residual Solvents CPMP/ICH/283/95 Sep 1997, Mar 1998

• ICH Q 3 D Guideline for Elemental Impurities EMA/CHMP/ICH/353369/2013, Aug 2015

• Guideline on the limits of genotoxic impurities CPMP/SWP/5199/02, CHMP/QWP/251344/ Jan 2007

• Genotoxic and Carcinogenic Impurities in Drug Substances & Products: Recommended Approaches US FDA, CDER, guideline, Dec 2008

• ICH M7 Assessment and Control of DNA Reactive (Mutagenic) Impurities in Pharmaceuticals to Limit Potential Carcinogenic Risk, Jun 2014


The good old days are over…..

• Analytical capabilities are continuously improving

• Example: DS containing a GTI at 20-40 ppm

– 1990 not detectable with available method (LoD 50 ppm): no toxicological assessment requested

– 2016 easily quantifiable: Is this level acceptable? Toxicological assessment imperative

• New: detailed process analysis for potential GTIs!

Why a guideline for genotoxic impurities?

ICH Q3A does indicate that „lower thresholds for reporting, identification & qualification can be appropriate if the impurity is unusually toxic“ but no guidance is given on what unusual toxicity is & how to handle it

Guideline emphasizes the “No-threshold-dogma“ for genotoxicants

“.... for genotoxic carcinogens ....

• there is no discernable threshold and

• any level of exposure carries a risk.“

NOEL + Uncertainty Factors = Safe Dose NO OPTION!



Exceptions from the „dogma“?

• Threshold effects for genotoxins increasingly acknowledged, particularly for those acting on non-DNA targets

• For “threshold“ impurities “Permitted Daily Exposure“ (PDE) can be calculated according to ICH Q3C (“Residual Solvents“) using N(L)OEL/UF approach

However: in practice data providing sufficient evidence for a threshold will be rarely available


Assessment of acceptable limits for genotoxic impurities without evidence for thresholded MoA

Proposal: Generic threshold below which hazard to human health is considered negligible!

Threshold of Toxicological Concern (TTC):

Based on an analysis of the potencies (TD50) of over 700 chemical carcinogens from the Gold et al. carcinogenic potency database (CPDB) it is estimated that for most carcinogens an intake level of less than 1.5 µg/day (=TTC) would give rise to less than 10-5 risk of cancer (“virtually safe dose“)


TTC translates into ppm impurity in drug

0.1

1

10

100

1000

Daily Dose of Drug [g]

Lim

it o

f Im

purity

[ppm

]

5 0.5 0.05 0.005

Limit of Impurity [ppm] = 1.5 µg

Daily Dose of Drug [g]

0.15 1.5

0.1 ppm

1 ppm

10 ppm

Analytical

control:

impractical?

technically

challenging

readily

achievable


Classification of mutagenic impurities ICH M7 classes

Class Definition Proposed action for control

1 Known mutagenic

Known carcinogens

Control at or below compound specific

acceptable limit

2 Known mutagens (Ames positive)

Unknown carcinogenic potential (no rodent

carcinogenicity data)

Control at or below acceptable limit (LTL /

TTC)

3 Alerting structure (in-silico), unrelated to

the structure of the drug substance;

No mutagenicity data

Control at or below acceptable limit

(appropriate TTC) or conduct bacterial

mutagenicity assay

If mutagenic: class 2

If not mutagenic: class 5

4 Alerting structure (in-silico), same alert in

the drug substance or related Ames

negative intermediate,

No mutagenicity data

Treat as non-mutagenic impurity

according to ICH Q3A/B

5 No structural alert or alerting structure with

sufficient data to demonstrate lack of

mutagenicity

Treat as non-mutagenic impurity

according to ICH Q3A/B

Cohort of concern Individual control strategy


Count colonies

Determine ratio treatment group/solvent control

+/- 0,5 ml S9 mix

0.1 ml test compound

2 ml minimal agar (0.05 mM histidine)

Incubate for about 72 hours

Overnight culture of bacteria

Histidine-free bottom agar

0.1 ml Bacteria (108 cells)

+/- Pre-incubation

The Salmonella mutagenicity (“Ames”) test


Evaluation of drug substance synthetic route First step of identifying GTIs

In silico tox evaluation of synthetic route, raw materials and intermediates

Class 1 and 2: Known mutagens/

carcinogens

Class 3: Intermediates with

structural alerts

Class 4 and 5: Intermediates API related or without

structural alerts

Ames test: 5 strains

Positive Negative

Class 1: Calculate PDE Class 2: Limit according to TTC

Limit according to ICHQ3A


Compound-specific risk assessment Mutagenic carcinogens (ICH M7 Class 1)

• Compound-specific risk assessments to derive acceptable intakes (AI) should be applied instead of TTC-based acceptable intakes where sufficient carcinogenicity data exists

• For known mutagenic carcinogen, a compound-specific acceptable intake can be calculated based on carcinogenic potency and linear extrapolation as default approach


Compound-specific risk assessment Methods

• Linear mode of action and calculation of acceptable intake (AI) (standard method) – Selection of studies (consider quality), usually from CPDB

– Selection of tumor and site, route of administration, human relevance of tumor

– AI = TD50 / 50’000 x 50 kg

• Published regulatory limits • Non-linear (threshold) mode of action and calculation of

permissible daily exposure (PDE) – Clear evidence for thresholded mechanism of action

• Acceptable limit based on exposure in the environment, e.g. intake from food


Addendum to ICH M7 guideline

Compound specific AIs (lifelong, all patient populations, all administration routes; with exceptions) • Acrylonitrile: 6 μg/day • Aniline: 720 μg/day • Benzyl chloride: 41 ug/day • Bis(chloromethyl)ether: 4 ng/day • p-Chloroaniline: 34 μg/day • 1-Chloro-4-nitrobenzene: 117 μg/day • p-Cresidine: 45 μg/day • Dimethylcarbamyl chloride: 5 μg/day; inhalation: 0.6 μg/day • Dimethyl sulfate: 1.5 μg/day • Ethyl chloride: 1’810 μg/day • Glycidol: 4 μg/day • Hydrazine: 42 μg/day; inhalation: 0.2 μg/day • Hydrogen peroxide: 6’960 μg/day • Hydroxylamine: 2 μg/day • Methyl chloride: 1’360 μg/day


Evaluation of DS byproducts, degradants... Continuous step of identifying GTIs

• During whole development process for all drug substances or product related impurities this basic process is applied:

• Evaluate for structural alerts/ literature data

• Evaluate a molecule as GTI or carcinogen (genotoxic or non-genotoxic carcinogen) or conventional impurity

• Apply limits (TTC, PDE) or qualify (following ICH Q3C guideline for limits of impurities in new drug substances) if necessary

In silico systems QSAR approaches

• Why use Quantitative Structure Activity Relationship (QSAR) approaches for safety evaluation?

• Mining of knowledge stored in huge public and proprietary data bases allows to profit from experiences of thousands of laboratories

• Possibility to predict safety issues without before test material synthesis and wet lab experiments

• Various commercial software packages available that allow to add proprietary databases and which considerably improves predictions


(Q)SAR systems: Overview

• Commercial systems –MCase

–DEREK

–TOPKAT

–OncoLogic

–etc

• Company specific systems

• Software packages to develop QSAR packages –OASIS

–etc 20 | V Simpósio SINDUSFARMA - IPS/FIP - ANVISA| Azeddine Elhajouji| 22-Jun-2016 | Impurities and degradation products

• Computer expert systems for the prediction of toxicological hazard - Rule based, reflecting the current state of knowledge of the relationship between chemical structure and biological activity - Every rule covers one endpoint (carcinogenicity, mutagenicity, skin sensitisation and more) - Toxophores of a compound are identified, highlighted and explained - Brief statement about the hazard is given

DEREK v. 9.0.0:

List of all alerts detected

Description of alerting

substructure

References

Comment

(explanations, mechanism of

action)

(Q)SAR systems DEREK (Lhasa Ltd.)


• Predicts toxicity on the basis of discrete structural fragments found to be

statistically relevant for a specific biological activity (biophores)

• Presence of biophores previously found in a number of active compounds

(represent in the database) is indicative of potential activity

• Generates also information on deactivating fragments (biophobes) and unknown

functionalities (coverage of the molecule)

QSAR systems MCASE (MultiCASE Inc.)

2-10 atom

fragmentation Alerting fragments

for rat tumorigenicity

NH2

O

N

CH2

O

N

O

N

O

O

…. and many more

N

Databases used

- Rodent carcinogenicity

(FDA + non-proprietary)

- Ames mutagenicity a)

- Skin sensitization a)

- Human liver toxicity

(FDA modules)

- hERG channel interaction a)

a) in-house data


“Super-mutagen” Prominent structural alerts


A snapshot from a computational toxicology report

Structural alert and MCASE biophore

Compound has a high likelihood to show genotoxicity

Synthesize the impurity

Test the impurity in an Ames test to verify or reject the in silico prediction

Synthesis scheme evaluation


Summary

• Pharmaceutical industries have to optimize the synthesis of the drugs to minimize and/or eliminate the impurities

• Limits for impurities and degradation products are regulated in the respective ICH guidelines. Specification and qualification needs input from Expert Toxicologists

• Genotoxic impurities are limited according to the TTC concept (Guideline for genotoxic impurities: ICH M7)

• (Q)SAR methods are highly sensitive tools to identify genotoxic impurities via determination of structural alerts

• The careful control of the levels of impurities in drugs is very important, because impurities may be hazardous to patients without giving them any benefit

• GMP principles must be followed for the production of drug substance to ensure the tight control of potential impurities

Thank you!

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