Exploring the role of poly-LacNAc and galectins in cancer immunotherapy
2National Glycoengineering Research Center, Shandong University
Galectins are a family of immune checkpoint soluble proteins that bind b-galactoside-containing glycans on and within cells. Galectin-1 and galectin-3 are widely expressed in tumor microenvironments, and have been shown to promote cancer growth and metastasis. Thus, galectin interaction with cancer cells is a new pathway for immune checkpoint regulation, equivalent to PD-1/PD-L1. Both galectin-1 and galectin-3 bind to poly N-acetyllactosamine [poly-LacNAc; (Galb1–4GlcNac)n]. However, while galectin-3 recognize internal LacNAc within poly-LacNAc, galectin-1 binds to terminal LacNAc and its binding is blocked by capping the glycan with sialic acids. We hypothesize that antibodies targeting tumor-associated poly-LacNAc glycans can inhibit galectin-glycan interactions, thereby hold great potential for cancer treatment. To generate antibodies against poly-LacNAc epitopes, we first conjugated the antigenic glycans to Keyhole limpet hemocyanin (KLH) carrier protein, with or without terminal sialic acids (Neu5Ac or Neu5Gc). Neu5Gc-deficient Cmah-KO mice were then immunized with Neu5Gca2-3(LacNAc)3-KLH, Neu5Aca2-3(LacNAc)3-KLH or KLH only (control group). Blood samples were collected weekly and examined by glycan microarrays. Mice immunized with Neu5Gca2-3(LacNAc)3-KLH or Neu5Aca2-3(LacNAc)3-KLH groups had a diverse and high polyclonal antibody responses against a variety of (LacNAc)3/4/5 glycans, in contrast to the control group. In particular, there was a higher serum binding to (LacNAc)3 than to (LacNAc)4/5. Furthermore, antibody responses against (LacNAc)3/4/5 glycans remained rather high even eight weeks after the last boost immunization. Altogether, this immunization regimen resulted in diverse, high and sustained polyclonal antibody responses against various poly-LacNAc glycans, and some of these antibodies could potentially inhibit galectin-glycan interactions. Such potent antibodies could open up new opportunities of immune checkpoint regulation, as novel cancer therapies.