Page 18 - Shimadzu Journal vol.9 Issue2

Biopharmaceutical

Liquid Chromatograph Mass Spectrometer LCMS™-9030

Characterization of Control and Stress

Induced Samples of Trastuzumab Biosimilar

Amita Puranik , Ratnesh Jain , Deepti Bhandarkar , Pratap Rasam , Prajakta Dandekar , Tian hua Wang 4
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1
1
3
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1 Department of Chemical Engineering, Institute of Chemical Technology, Mumbai
2 Shimadzu Analytical (India) Pvt. Ltd.
3 Department of Pharmaceutical Science and Technology, Institute of Chemical Technology, Mumbai
4 Shimadzu (Asia Pacific) Pte. Ltd.
User Benefits
deamidation are the most commonly observed Post-Translational
・ In-depth peptide mapping and PTM analysis methodology is described
for the analysis of control and stress induced samples of mAb which Modifications (PTMs). Early identification of these prone sites en-
can be useful in identification of sites susceptible to oxidation and ables antibody engineering to eliminate liability of leading candi-
deamidation modifications
dates to such modifications while maintaining binding activity.
・ Excellent and stable mass accuracy, comprehensive fragmentation
pattern and high sensitivity offered by LCMS-9030 helped obtain Bottom-up approach which is essentially peptide mapping helps
good sequence coverage, identification of PTMs and sites susceptible in determination of primary amino acid sequence and identifica-
for modifications
tion of site–specific PTMs. In this application note, methodology
for identification of sites susceptible to oxidation and deamidation
is described by analyzing the control and artificial chemically stress
Introduction induced (forced degradation) samples of mAb.

Monoclonal antibodies (mAbs) are a major class of biopharmaceu-
ticals covering a large panel of diseases, from cancer to asthma, Experimental
including central nervous system disorders, infectious diseases
and cardiovascular diseases . Throughout manufacturing, storage, Trastuzumab biosimilar sample (2 mg/mL) was incubated in water
1
transportation, and administration, mAbs are subjected to biophys- with pH 9 (adjusted using tris base solution) for 7 days at 37°C to
ical and biochemical stress from multiple sources, which may lead induce deamidation. Similarly, 10 µL of 30% H O solution was
2 2
to their degradation via aggregation, fragmentation, and chemical added to 2 mg/mL of trastuzumab sample and incubated for 1 hour
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modifications, such as oxidation, deamidation, or isomerization . at 37°C to induce oxidation. Oxidation reaction was quenched by
Among these undesirable degradation products, oxidation and adding pinch of methionine.

Shimadzu Journal vol.9 Issue2 51