New Insights on the Role of Anti-PD-L1 and Anti-CTLA-4 mAbs on Different Lymphocytes Subpopulations in TNBC

Abstract

Antibody-based cancer immunotherapy includes monoclonals against immune checkpoints (ICs), to modulate specific T cell responses against cancer. NK cells are a newly emerging target for immune checkpoint receptor inhibition in cancer immunotherapy, as ICs are also expressed on NK cells in various cancers. The latter cells are becoming attractive targets for cancer immunotherapy, as they are effector cells similar to CTLs, exerting natural cytotoxicity against primary tumor cells and metastasis, and they are able to distinguish tumor cells from healthy ones, leading to more specific anti-tumor cytotoxicity and reduced off-target effects. Thus, we decided to test the effects on isolated NK cells and T cell subpopulations of novel immunomodulatory mAbs, recently generated in our lab, in comparison with those in clinical use, such as ipilimumab and atezolizumab. Interestingly, we found that the novel anti-CTLA-4 (ID-1) and anti-PD-L1 (PD-L1_1) antibodies are able to induce NK cell activation and exert anti-tumor effects on TNBC cells co-cultured with NK cells more efficiently than the clinically validated ones, either when used as single agents or in combinatorial treatments. On the other hand, ipilimumab was found to be more effective in activating T cells with respect to ID-1. These findings indicate that antibodies targeting different epitopes can have differential effects on different lymphocytes subpopulations and that novel combinations of mAbs could be suitable for therapeutic approaches aimed at activating not only T cells but also NK cells, especially for tumors lacking MHC.

Triple Negative Breast Cancer Treatment Options and Limitations: Future Outlook

“…While immunotherapy does not have a robust response in general to breast cancer patients, it has been shown that a subset of TNBCs has high TMB and high TILs, like those observed in melanoma or lung cancer, which may be treated with immune checkpoint inhibitors. Consequently, developing immunotherapies that target TNBC has become possible due to its immunogenic nature [ 111 ]. Recently, the FDA approved atezolizumab combined with nab-paclitaxel for treating metastatic and unresectable programmed cell death ligand 1 (PD-L1)-positive TNBC [ 112 ].…”

A Comparison of the Antitumor Efficacy of Novel Multi-Specific Tribodies with Combinations of Approved Immunomodulatory Antibodies

“…Human peripheral blood mononuclear cells (PBMCs) were isolated from the blood of healthy donors using a Greiner Leucosep ® tube (Sigma-Aldrich, St. Louis, MO, USA) following the manufacturer's instructions, as previously reported [16,19,[23][24][25], and frozen in 90% FBS plus 10% dimethyl sulfoxide (DMSO) solution until use. Cryopreserved cell vials were gently thawed using RPMI 1640 medium supplemented with 1% L-glutamine, 1% CTLWash™ (Immunospot by Cellular Technology Limited, Shaker Heights, Cleveland, OH, USA) and 100 U/mL Benzonase (Merck Millipore, Darmdstadt, Germany) and washed via centrifugation.…”

“…As negative controls, the co-cultured cells were analyzed in the absence of treatments or in the presence of a human-unrelated IgG used at the same concentration. The cell supernatants were collected at the end of incubation, and the level of lactate dehydrogenase (LDH) released by the cells was evaluated as a marker of tumor cell lysis and expressed as a percentage with respect to the max lysis obtained after treatment with 10% Triton X-100, following the manufacturer's recommendations for the LDH detection kit (Thermofisher Scientific, Rockford, IL, USA) [18,19].…”

“…These two novel human constructs should be endowed with more potent immunomodulatory activity than conventional monospecific mAbs, since they incorporate two different immune checkpoint binding domains, combine the binding specificities and biological properties of two different immunomodulatory mAbs in one single molecule, and, indeed, have already shown more potent antitumor effects with respect to their parental immunomodulatory antibodies [20]. Here, we decided to compare them to the clinically validated and used anti-LAG-3 (Relatlimab), anti-PD-L1 (Atezolizumab), and anti-PD-1 mAbs (Pembrolizumab), alone or in combination, for the treatment of different types of tumors [13][14][15][17][18][19][27][28][29].…”

“…These antibodies showed different intriguing properties: they were able to bind to their targets with comparable or even higher affinity than the clinically validated mAbs, and they were found to be capable of more efficiently activating human peripheral blood mononuclear cells (hPBMCs), resulting in increased cytokine secretion. Indeed, when the antibodies were used in co-cultures of hPBMCs with tumor cells, they showed more potent tumor cell lysis when compared to the mAbs approved for clinical use, and these effects were enhanced when the antibodies were used in combinatorial treatments [16][17][18][19].…”