Immune Checkpoint inhibitors (ICIs) are used in a type of immunotherapy that harnesses the power of the body’s own immune system against cancer. The immune system has checkpoints that prevent it from attacking healthy cells. Cancer cells can hide from the immune system by using these checkpoints. ICIs block these checkpoints so that the immune system can recognize cancer cells and attack them. One strategy involves blocking two or even three of these immune checkpoints. This technology has benefits but it also has significant toxicity profiles. Research involves studying these technologies in vitro.
Researchers in Poland used monoclonal antibodies to block these checkpoints. Examples of checkpoints used were V-type immunoglobulin suppressor of T-cell activation (VISTA), cytotoxic T-lymphocyte antigen 4 (CTLA-4), programmed cell death 1 (PD-1), and programmed cell death ligand 1 (PD-L1).
Authors used a certain type of monoclonal antibody called a ‘bispecific antibody’ in this research. These are engineered to bind to two different antigens simultaneously. They combine the capabilities of two different monoclonal antibodies into a single molecule.
Bispecific antibodies (bsAbs) have many advantages including: a) immune cell recruitment and better cytotoxicity; b) superior protein blocking power ie they can block proteins from multiple pathways all at the same time, c) they can penetrate tissues that monospecific antibodies cannot, d) they form synapses and activate immune cells, e) they can facilitate receptor clustering, and f) they may engage tumor cells with immune cells, according to the authors.
Researchers designed these bsAbs using three different formats: symmetric, asymmetric, and single-chain variable fragment (scFV). Recall that an symmetric antibody has two identical "halves," meaning it can only bind to a single type of target antigen. These have the characteristic Y shape. An asymmetric antibody is an engineered bispecific antibody that can bind to two different target antigens. Lastely, ScFV is the smallest functional unit of an antibody, it’s a small building block you can use to construct more complex antibody formats.
The binding and blocking properties of these bispecific antibodies (bsAbs) and their efficacy compared to monotherapy and combination therapy were determined using cancer cell lines in vitro including endometrial (RL95-2), pancreatic (PANC-1) and breast cancer (BT-20) cell lines.
Authors saw a higher level of tumor cell lysis using bispecific antibodies compared to monospecific antibodies. The high level of tumor cell lysis and increased expression of pro-inflammatory cytokines induced by the ScFV-based bispecific antibodies suggests a novel approach.
Authors used our gene selector G418 Disulfate after transfection via electroporation in the generation of stable cell lines during this research.
Reference
Bielski et al (2025) The bispecific antibody targeting VISTA and PD-L1 shows enhanced tumor inhibitory activity in pancreatic, endometrial and breast cancers compared to mono- and combination immune checkpoint blockade. Front. Immunol. 16: 1486799 Link