Biophysical and stability characterization of antibody-drug conjugates | Malvern Panalytical

Date recorded: September 13 2018

Duration: 34 minutes 35 seconds

Antibody-Drug Conjugates (ADCs) are a rapidly growing class of oncology therapeutics wherein a potent cytotoxic drug is conjugated to the antibody molecule. The impact of drug conjugation on the antibody's conformation needs to be evaluated. Additionally, the drug conjugation can also affect various physical properties of the antibody molecule which in turn can affect its stability and aggregation propensity. This presentation summarizes results from our study on aggregation of a lysine-conjugated ADC and its unconjugated mAb under accelerated heat and agitation stresses. Additionally, we also highlight spectroscopic and DSC based analytical approaches that can be used to detect subtle differences between the ADC and mAb.

Table of contents

1. Welcome

00:12

2. Introduction

00:52

3. Physical Stability and Aggregation of Antibody-Drug Conjugates

00:07

4. Outline

00:41

5. Therapeutic proteins offer treatments for growing number of diseases

00:35

6. Protein aggregation

00:43

7. Antibody-Drug Conjugates: A New Class of Therapeutic Agents

01:01

8. Current Conjugation Strategies

01:19

9. Challenges specific to antibody-drug conjugates

01:19

10. Outline

00:07

11. Types of reactions for synthesizing trastuzumab-DM1 (T-DM1)

01:28

12. Difference in intact mass analyses between T-DM1 vs Syn-ADC

01:43

13. Sample preparation

00:35

14. Effects of drug and linker conjugation on higher order structure of mAb

02:09

15. Thermal Stability using Differential Scanning Calorimetry

01:07

16. Heating-rate dependence of DSC thermograms

00:30

17. Apparent Tm Versus DSC Heating Race

00:49

18. Energy of Activation (Ea) for Heating Induced Unfolding/Aggregation Reaction

00:22

19. Plot of 1 𝑇 𝑚 vs ln 𝑣 𝑇 𝑚 2 and Energies of Activation

00:35

20. Heating induced aggregation using Dynamic Light Scattering

00:46

21. Tertiary and secondary structural changes for thermally stressed TmAb and T-DM1

00:55

22. Apparent solubility of ADCs and unconjugated mAb

00:38

23. PEG precipitation assay in PBS buffer, pH 7.4

00:49

24. Summary

00:05

25. Outline

00:15

26. Chemical denaturation of TmAb and T-DM1 to assess conformational stabilities

00:29

27. Colloidal Stability of proteins

00:54

28. Measurement of colloidal stability using light scattering technique

00:48

29. Colloidal stability of TmAb and T-DM1

01:28

30. Outline

00:11

31. Accelerated stress conditions

00:25

32. Aggregation of TmAb and T-DM1 During Isothermal Storage

00:39

33. Size exclusion chromatography for TmAb and T-DM1 samples stored at 50°C

01:17

34. Effects of isothermal incubation on sub-visible particles in mAb and ADCFlow-imaging microscopy

00:24

35. Effects of Isothermal Incubation on Sub-Visible Particles in mAb and ADC Resonant Mass Measurements

01:07

36. Agitation induced aggregation of TmAb and T-DM1

00:43

37. Aggregate Characterization Using SEC

01:17

38. Aggregate Characterization Using SEC

00:25

39. Micro-Particle Characterization Using Flow Imaging Microscopy

00:58

40. Micro-Particle Size Distribution

00:05

41. Sub-visible Particle Characterization Using Resonant Mass Measurement

00:27

42. Summary of results

00:42

43. Conclusions

00:42

44. Acknowledgements

00:24

45. Thank you

00:08

46. Thank you for your attentioncraig.sagar@malvernpanalytical.com

01:20

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