Supplementary Information

Structural and functional comparability study of anti-CD20 monoclonal antibody with

reference product

Authors: Sanjay Kumar Singh, Santosh Pokalwar, Sandip Bose, Shivika Gupta, Suhani

Almal, Ranjit Sudhakar Ranbhor*

Affiliation: Sun Pharmaceutical Industries Limited, Tandalja, Vadodara – 390 012 India

*Corresponding author email: ranjit.ranbhor@sunpharma.com

Figure S1: A) SDS-PAGE analysis of reference product and SB-02 product under reduced condition. Lane 1: Molecular weight marker (250-10 kDa), Lane 2: reference product, Lane 3: Reference product, Lane 4: SB-02 B): SDS-PAGE analysis of reference product and SB-02 under non-reduced condition. Lane 1: Molecular weight marker (250-10 kDa), Lane 2: reference product, Lane 3: reference product, Lane 4: SB-02.

Figure S2: Light chain amino acid sequence comparison between SB-02 and reference product.

Figure S3: Heavy chain amino acid sequence comparison between SB-02 and reference product.

Figure S4: Representative intact mass profile of reference product and SB-02.

Figure S5: Representative subunit mass profile of heavy chain of reference product and SB-02.

Figure S6: Representative subunit mass profile of light chain of reference product and SB-02.

Figure S7: Representative subunit mass profile of de-glycosylated heavy chain of reference product and SB-02.

Figure S8: Representative total ion chromatogram (TIC) profile of reference product and SB-02.

Figure S9: Representative peptide finger print profile of reference product and SB-02 after trypsin digestion

Figure S10: Representative peptide finger print profile of reference product and SB-02 after Lys-C digestion

Figure S11: Representative UV profile of reference product and SB-02.

Figure S12: Representative far UV CD spectrum of reference product and SB-02.

Figure S13: Representative near UV CD spectrum of reference product and SB-02.

Figure S14: Representative fluorescence profile of reference product and SB-02.

Figure S15: Representative FTIR spectrum of reference product and SB-02.

Figure S16: Representative XIC profile of S-S linked peptides observed in SB-02 and reference product after digestion with trypsin in non-reducing conditions

Figure S17: Free thiol content of reference product and SB-02.

Figure S18: Representative cIEF profile of reference product and SB-02.

Figure S19: Representative cation exchange chromatographic profile of reference product and SB-02.

Figure S20: Representative CZE profile of reference product and SB-02.

Figure S21: Low molecular weight impurity content of reference product and SB-02.

Figure S22: Non glycosylated heavy chain impurity content of reference product and SB-02.

Figure S23: Oxidized content of reference product and SB-02.

Figure S24: Oligomer content of reference product and SB-02 by SEC-MALS.

Figure S25: Glycated content of reference product and SB-02.

Figure S26: Representative CD20 binding profile for reference product and SB-02.

Table S1: C1q binding assay by surface plasmon resonance.

C1qTest Sample KD (μM)

US reference product 3.56US reference product 3.26EU reference product 1.43EU reference product 3.15Indian reference product 1.59Indian reference product 2.18Indian reference product 2.53Indian reference product 1.70SB-02 2.89SB-02 2.83