Complement-Dependent Cytotoxicity (CDC)

Complement System

The complement system is a protein response system with a precise regulatory mechanism. It is not only an important part of the innate immune defense system, but also one of the important mechanisms by which antibodies exert their immune effects.

Fig.1 Complement activates the classical pathway.¹

The classical pathway of complement activation consists of three main steps: recognition, activation, and membrane attack. The identification units include C1q, C1r, and C1s. Activation units include C2, C3, C4. Membrane attack complex units include C5, C6, C7, C8 and C9.

When the antibody recognizes and binds to the cell surface antigen, the structure of its crystallizable Fc segment will be significantly changed, and when the six antigen-bound IgG molecules interact with each other to form a hexamer structure on the cell surface, the binding site of the C1q molecule can be exposed. This facilitates the fixation of the CH4 domain of IgM and the CH2 domain of IgG3, IgG1, and IgG2 (to a lesser extent) than in the order of magnitude.

C1q, as the first protein of the complement system, has "six long arms" and is able to grasp the six Fc regions separately. In addition, it also carries C1r and C1s proteases, which can further cause downstream cascade reactions, and eventually form a membrane attack complex (MAC) on the surface of the target cell, which destroys the local phospholipid bilayer structure of the cell membrane and forms a "leaky plaque", or forms a hydrophilic pore across the membrane, which allows water, ions, and soluble small molecules to flow freely through this pore, and eventually leads to cell swelling, lysis, and death.

Complement Regulatory Proteins

A variety of complement regulatory proteins are present in cell membranes and plasma. They play a role in the regulation of complement function by controlling key steps such as enzyme activity and MAC assembly during the cascade. The main membrane-bound regulatory proteins associated with complement activation are: CD46 (membrane cofactor protein), CD55 (decay acceleration factor, DAF), and CD59 (protective protein). Among them, CD46 acts as a cofactor for factor I, promoting the degradation of C3b to the inactivated form of iC3b. CD55 prevents CDC by inhibiting C3 convertase by preventing the production of C3b. There is also evidence that CD55 can inhibit T cell-mediated immune responses. CD59 can prevent MAC assembly by binding to C7, C8, or C9. In addition to membrane-bound complement regulatory proteins, sialic acid molecules on the cell surface are also involved in the regulation of CDC effects. In addition, there are soluble molecules involved in the regulation of cellular complement function.

Table 1. Common complement regulatory proteins.²

Factors Influencing the CDC Effect

• Antibody subtype

Generally speaking, the strength of the CDC effect (the ability to activate C1q): IgM> IgG3> IgG1> IgG2, IgG4 do not have the ability to activate the complement classical pathway. But condensed IgA, IgG4, and IgE can activate complement through alternative pathways.

• Expression level of target cell antigen

Target cells need to express enough antigen to bind to the antibody and activate complement. When antigen expression levels are low, complement cannot be activated.

• Complement regulatory proteins

Some of the proteins present on the surface or inside target cells and in serum can modulate complement activity, which in turn affects CDC effects, as shown in Table 2 for the various common complement inhibitory molecules.

• Antigen-antibody binding orientation

The alignment of antigen-antibody and the arrangement of adjacent antibodies can significantly affect the role of CDC.

• Complement source

Treatment Strategies

At present, monoclonal antibodies against CDC effects have been used in some hematologic tumors and solid tumors, such as antibodies targeting CD20, CD19, and CD22 for B-cell lymphoma and EpCAM antibodies for solid tumors. Activation of the complement pathway is generally beneficial for immunotherapy, but in many cases, particularly in patients with autoimmune diseases and cancer, the complement pathway is often deficient. Current and future therapeutic strategies for monoclonal antibody-mediated CDC effects include:

Fig.2 Strategies for drug development for the complement system.¹

• Additional serum supplementation as a source of complement to compensate for congenital and/or acquired complement deficiencies, thereby enhancing complement-mediated CDC effects.
• Perform single-point mutations (e.g., E345K, E430G, etc.) of the Fc segment of IgG to enhance the Fc-Fc interaction. Or replace the IgG1 isoform with the IgG3 isoform for better immobilization of the C1q molecule.
• Development of bispecific antibodies that simultaneously target tumor antigens and inhibit complement-regulatory proteins.
• Development of factors for the neutralization of soluble and cellular complement regulatory proteins.

References

1. Bordron, Anne.; et al. "Complement system: a neglected pathway in immunotherapy." Clinical reviews in allergy & immunology. 58,2 (2020): 155-171.
2. Cummings, K.L.; et al. "Role of complement in immune regulation and its exploitation by virus." Viral immunology. 20,4 (2007): 505-24.