Rituxan/Mabthera (Rituximab) Biosimilars

Rituxan/Mabthera Biosimilars

Rituxan/Mabthera (rituximab) is a chimeric human/murine monoclonal antibody targeting the CD20 antigen expressed on the surface of B cells. Rituxan is used therapeutically in the treatment of hematological cancers and autoimmune disorders.

A number of Rituxan biosimilars have been launched across the world, signifying the importance of this critical therapeutic. However, comprehensively demonstrating the comparability of your biosimilar to the innovator product can be a barrier to achieving regulatory approval.

Sartorius has developed a suite of pre-qualified assays to assess the biosimilarity of your product. Our chemical, structural, binding, and functional assays are designed to meet regulatory requirements, providing you with the detailed analysis needed to simplify and accelerate biosimilar development.

Simplify Your Biosimilar Characterization

Sartorius offers a panel of ready-to-use assays allowing you to characterize your Rituxan biosimilars in detail. 

  • Uncover valuable insights into your biosimilar 

  • Accelerate development times 

  • Meet regulatory requirements 

  • Boost confidence in your product 

  • Limit the risks associated with biosimilar development

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Characteristics of Rituxan

Rituxan accomplishes its clinical effects by binding and destroying B cells, which are central to the function of the adaptive immune system. Rituxan is effective against both normal and malignant B cells, meaning it is an effective therapy for conditions caused by aberrant B cells, overproduction of B cells, or overactive B cells. 

Therefore, Rituxan is an effective therapeutic for blood cancers like B cell lymphoma or leukemia and certain autoimmune conditions, such as rheumatoid arthritis and some types of vasculitis, that have not responded to other treatments. 

Rituxan mediates its activity through the interaction with the protein CD20, found on the surface of B cells. Rituxan binds this protein with high affinity and mediates several different outcomes that lead to the destruction of B cells. These include inhibition of cell proliferation, induction of apoptosis, and recruitment of the effector cells of the immune system. 

Both the function of CD20 and the mechanisms of action of Rituxan are not fully understood. However, Rituxan is known to mediate antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC), two crucial immune pathways through which targeted therapies trigger the death of the cells they have marked.  

Sartorius can support your Rituxan biosimilar project with our comprehensive selection of analysis platforms. Our off-the-shelf characterization solutions were designed to meet regulatory requirements for comparability. 

  • Physicochemical and structural assays reveal detailed insights into the composition of your biosimilar. 

  • Binding assays quantify and evaluate the binding of your biosimilar to CD20 and other components of the immune system. 

  • Our bioassays reveal the functional characteristics of your Rituxan molecule and determine its effect on immune pathways.  

Assay Types

Physicochemical and structural characterization is a crucial step in determining the potential biological activity, stability, and safety of your biosimilar.   

Our versatile platform of physicochemical methods provides the basis for your comparability study.  With options suited for clone selection through to formal comparability, we can build detailed insights into the properties of your biosimilar.    

The combination of our off-the-shelf physicochemical and structural analyses and our binding and bioassays allows you to evaluate the structure-function relationship and ticks the regulators’ box for orthogonal comparability. 

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Rituxan has a number of different mechanisms of action. Therefore, a range of bioassays are required to fully characterize Rituxan biosimilars and demonstrate comparability. 

ADCC and CDC  

The principle of both Rituxan ADCC and CDC assays is that the relevant target cells, typically Raji, Daudi, or Ramos cells, expressing the CD20 protein are prepared in cell plates using a procedure designed to optimize receptor expression. A dilution series of Rituxan is then added to the plate and binds the target cells.  

Engineered cells stably expressing the FcgRIIIa receptor (Rituxan ADCC bioassay) or complement (Rituxan CDC bioassay) are then added to the target cells. These cells will bind to Rituxan (which is in turn bound to the CD20 protein on the target cell surface), resulting in target cell lysis (CDC) or a luminescent signal (ADCC). 

Antibody-dependent cellular phagocytosis (ADCP) 

By targeting CD20, Rituxan signals B cell depletion by recruiting the full range of Fc-Fc gamma receptor interaction activities. 

ADCP recruits macrophages, monocytes, neutrophils, and dendritic cells, which attach and kill antigen-expressing target cells. Once the target cell has been recognized, Fc-gamma Receptor I (FcγRI/CD64), Fc-gamma Receptor IIa (FcγRIIa/CD32a), and Fc-gamma Receptor IIIa (FcγRIIIa/CD16a) expressed on the effector cells binds the projected Fc region. This binding results in the activation of intracellular signaling pathways leading to the phagocytic destruction of the target cell. 

Our ADCP assay utilizes a report gene approach to report the ADCP activity of your Rixutan biosimilar, which avoids the obstacles and variability evident in a classical assay.  

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Rituxan exerts its biological function by binding to CD20. Therefore, efficient antigen-binding represents one of the key quality attributes of a Rituxan biosimilar. 

CD20 binding assays can be difficult to perform as the CD20 protein is unstable when not expressed on the cell membrane. To overcome this challenge, we use a cell-based binding assay combined with flow cytometry using the iQue Screener, a high throughput, multiplex instrument, complemented by analysis on the meso scale discovery (MSD) platform.  

Fc Characterization

Rituxan must also interact with immune components to provide its clinical benefits. Our full range of Fc binding assays have been extensively qualified with Rituxan innovator material. We can support testing of: 

  • Fc-Gamma Receptor I (FcRI)  

  • Fc-Gamma Receptor IIa (both R and H variants) (FcRIIa)  

  • Fc-Gamma Receptor IIb (FcRIIb)  

  • Fc-Gamma Receptor IIIa (V) (FcRIIIa V)  

  • Fc-Gamma Receptor IIIa (F) (FcRIIIa F)  

  • Fc-Gamma Receptor IIIb (FcRIIIb) 

  • FcRn 

  • C1q 

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