Figure 4.

Rhamnose-functionalized CB2 induces CDC against B cell lymphoma. (A) Model for complement activation. CB2-Rha-thFF03 recruits endogenous anti-Rha Abs to lymphoma cells via its Rha moieties. Anti-Rha Abs activate the complement cascade, leading to cell lysis. Magnified: The thFF03 peptide contains six lysines for the chemical conjugation of Rha₃. (B) Illustration of the conjugation approach. First, several Rha₃ moieties are attached to the lysine side chains of the thFF03 peptide (1). Then, the sortase reaction is used to couple rhamnosylated thFF03 to the C-terminus of CB2 (2). (C) Glycan array confirming the presence of anti-Rha Abs in human serum. Note that Rha₃ is recognized by more IgGs isolated from human serum than Rha monomers. (D) Complement-dependent cytotoxicity assay. Note that CB2-Rha-thFF03 shows increased cytotoxicity compared to CB2. Antihuman IgG Ab was included as a positive control. Cells were incubated with mixtures of CB2 or CB2-Rha-thFF03 (0.4 mg/mL = 24 μM) and human Abs (0.2 mg/mL) for 1 h at RT, followed by incubation with rabbit complement for 2 h at 37 °C. Values are normalized to the positive control (100%) and represent mean ± SEM (N = 3), (*) p < 0.05, (**) p < 0.01, (***) p < 0.001.