Abstract
PD-1/PD-L1 blockade has revolutionized the field of immunooncology. Despite the relative success, the response rate to anti-PD-1 therapy requires further improvements. Our aim was to explore the enhancement of T-cell function by using novel PD-1-blocking proteins and compare with clinically approved monoclonal antibodies (mAbs). We isolated T-cells from the ascites and tumor of 17 patients with advanced epithelial ovarian cancer (EOC) and analyzed the effects using the mAbs nivolumab and pembrolizumab and two novel engineered ankyrin repeat proteins (DARPin® proteins). PD-1 blockade with either mAb or DARPin® molecule significantly increased the release of IFN-γ, granzyme B, IL-2, and TNF-α, demonstrating successful reinvigoration. The monovalent DARPin® protein was less effective compared to its bivalent equivalent, demonstrating that bivalency brings an additional benefit to PD-1 blockade. Overall, we found a higher fold increase of lymphokine secretion in response to the PD-1 blockade by tumor-derived T-cells; however, the absolute amounts were significantly lower compared to the release from ascites-derived T-cells. Our results demonstrate that PD-1 blockade can only partially reinvigorate functionally suppressed T-cells from EOC patients. This warrants further investigation preferably in combination with other therapeutics. The study provides an early pilot proof-of-concept for the potential use of DARPin® proteins as eligible alternative scaffold proteins to block PD-1.
Highlights
Many human solid tumors are known to contain tumorinfiltrating lymphocytes (TILs) [1,2,3,4]; tumor cells are able to avoid elimination through various escape mechanisms including functional suppression of T-cells
This exhausted state is associated with the expression of several surface co-inhibitory receptors, which can be expressed on TILs in various combinations and suppress cell functions by interacting with ligands on tumor cells and other suppressive cell types [5,6,7,8]
A median of 66.5% of CD8+ TILs and 32.2% of CD8 + ascites-derived T-cells expressed PD-1, compared to Tcells isolated from the blood of healthy controls (HC, median 0.8%) and patient blood (4.0%) (Figure 1(a))
Summary
Many human solid tumors are known to contain tumorinfiltrating lymphocytes (TILs) [1,2,3,4]; tumor cells are able to avoid elimination through various escape mechanisms including functional suppression of T-cells. Impaired TIL functionality is characterized by defective cytotoxic activity, diminished cytokine secretion, and failure to proliferate in Journal of Immunology Research response to stimulation [5]. This exhausted state is associated with the expression of several surface co-inhibitory receptors, which can be expressed on TILs in various combinations and suppress cell functions by interacting with ligands on tumor cells and other suppressive cell types [5,6,7,8]. There are still many aspects of this strategy requiring additional optimization including (a) identification of benefitting patient groups, (b) finding relevant combination therapies to improve clinical efficacy, (c) optimizing treatment schedules, (d) identification of biomarkers, (e) limiting adverse events, and (f) managing acquired resistance [12, 13]
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