Genotoxic impurities and analysis

Genotoxic impurities and analysis

1. Genotoxic Impurities and Analysis 22/05/2020 Prepared By : Mahendra G S M-Pharm, Pharmaceutical Chemistry JSSCP, MYSURU

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5. Single-base alteration Depurination Deamination of cytosine to uracil Deamination of adenine to hypoxanthine Alkylation of base Insertion or deletion of nucleotide Base-analog incorporation Two-base alteration UV light–induced thymine-thymine (pyrimidine) dimer Chain breaks Ionizing radiation Radioactive disintegration of backbone element Oxidative free radical formation Cross-linkage Between bases in same or opposite strands Between DNA and protein molecules ( eg , histones ) Types of DNA damage 22/05/2020

6. Single-base alteration 22/05/2020

7. Two-base alteration Pyrimidine Dimers 22/05/2020

8. Keto – Enol Tautomerization 22/05/2020

9. Point mutations: The replacement of one base pair by another results in point mutation. They are of two sub-types. Transitions: In this case, a purine (or a pyrimidine) is replaced by another. Transversions : These are characterized by replacement of a purine by a pyrimidine or vice versa. 22/05/2020

10. Types of DNA damage 22/05/2020

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12. Types of Genetic Damage Mutagenic (not all DNA damage results in mutation) Causing change in the genetic information Senescence –irreversible dormancy Apoptosis –programmed cell death unregulated cell division Clastogenic Causing change in the chromosome Structural (breaks) Numerical (aneuploidy) –changes in the number in a cell Carcinogenic Causing or facilitating propagation of cancer Sufficient mutagenic and/or clastogenic activity = carcinogenicity 22/05/2020

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14. ICH Q3 Impurities Q3A Drug Substances– 1995 (step 4), R1 (2002), R2 (2006) Q3B Drug Products– 1996 (step 4), R1 (2003), R2 (2006) Q3C Residual Solvents– 1997, R1-5 (2002, 2005, 2009, 2011) Most ICH guidelines on impurities in drug substances and drug products are >15 years old 22/05/2020

15. Filling the Gaps ICH Q3D Elemental Impurities – Step 4 (Dec. 2014) - USP , Establishment of Allowable Concentration of Genotoxic Impurities in Drug Substance and Product 2005, PhRMA Position Paper EMA, Guideline on the limit of Genotoxic Impurities 2006 FDA Draft: Genotoxic and Carcinogenic Impurities in Drug Substance and Products: Recommended Approaches and Acceptable Limits -2008 ( allined with EMA) ICH M7 – Genotoxic Impurities – Step 4 (June 2014) - changes from EMA and FDA guidance 22/05/2020

16. ICH M7 – Genotoxic Impurities Filling the ICH Q3 A/B gap for “impurities that are expected to be unusually potent , producing toxic or pharmacological effects at a level not more than (≤) the identification threshold .” Identification of "unusually potent" impurities not described and No threshold of concern given EMA guideline and FDA draft guidance (earlier to ICH M7 guidelines followed for GTIs) Threshold of Toxicological Concern (TTC), 1.5 µg/day 22/05/2020

17. Assessing Impurities – ICH M7 All impurities (actual and potential), where the structures are known, should be evaluated for mutagenic potential. 22/05/2020

18. Classifying impurities – PhRMA paper recommendation, adopted in M7 22/05/2020

19. Alerting Structures – examples 22/05/2020

20. Assessing Impurities – ICH M7 Is an impurity potentially genotoxic? Search databases and literature for carcinogenicity and bacterial mutagenicity data in order to classify impurity as Class 1, 2, or 5 When data are not available Use Structure-Activity Relationships (SAR) that focus on bacterial mutagenicity predictions. This could lead to a classification into Class 3, 4, or 5. 22/05/2020

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23. AMES TEST The Ames test is a biological assay to assess the mutagenic potential of chemical compounds. Negative result classifies compound as normal ICH impurity and overrides a positive in silico prediction for genotoxicity 22/05/2020

24. Procedure for AMES TEST The mutagenicity of a substance is proportional to the number of colonies observed. 22/05/2020

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29. 22/05/2020 Inductively coupled plasma mass spectrometry ( ICP - MS ) –detection capability is at concentrations as low as one part in 10 15

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31. 22/05/2020 Inductively coupled plasma atomic emission spectroscopy ( ICP - AES ), also referred to as inductively coupled plasma optical emission spectrometry ( ICP - OES ), is an analytical technique used for the detection of trace metals.

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