Molecular Recalibration of PD-1+ Antigen-Specific T Cells from Blood and Liver.

Itziar Otano,Harsimran Singh,Antonio Bertoletti,Brian R Davidson,Shao-An Xue,Andrea Pavesi,Hans J Stauss,Anna Schurich,Frederick A Vargas,Patrick T.F Kennedy,Zhi M.D Tan,Mala K Maini,Giuseppe Fusai,Navjyot Hansi,Jia Y.J Aw,Francis Robertson,David Escors

doi:10.1016/j.ymthe.2018.08.013

Abstract

Checkpoint inhibitors and adoptive cell therapy provide promising options for treating solid cancers such as HBV-related HCC, but they have limitations. We tested the potential to combine advantages of each approach, genetically reprogramming T cells specific for viral tumor antigens to overcome exhaustion by down-modulating the co-inhibitory receptor PD-1. We developed a novel lentiviral transduction protocol to achieve preferential targeting of endogenous or TCR-redirected, antigen-specific CD8 T cells for shRNA knockdown of PD-1 and tested functional consequences for antitumor immunity. Antigen-specific and intrahepatic CD8 T cells transduced with lentiviral (LV)-shPD-1 consistently had a marked reduction in PD-1 compared to those transduced with a control lentiviral vector. PD-1 knockdown of human T cells rescued antitumor effector function and promoted killing of hepatoma cells in a 3D microdevice recapitulating the pro-inflammatory PD-L1hi liver microenvironment. However, upon repetitive stimulation, PD-1 knockdown drove T cell senescence and induction of other co-inhibitory pathways. We provide the proof of principle that T cells with endogenous or genetically engineered specificity for HBV-associated HCC viral antigens can be targeted for functional genetic editing. We show that PD-1 knockdown enhances immediate tumor killing but is limited by compensatory engagement of alternative co-inhibitory and senescence program upon repetitive stimulation.

Highlights

CD8 T cells are critical for immune control of persistent viral infections and cancer
Pre-stimulation of isolated CD8 T cells with CMV peptide NLV for 24 hr was successful in focusing vesicular stomatitis virus G-protein (VSV-g) pseudotyped lentiviral transduction on the responding population (Figure 1A)
We have shown that the pro-inflammatory cytokines IFN-g, tumor necrosis factor alpha (TNF-a), and IL-2 enhance the capacity of TCR-transfected T cells to lyse tumor target cells;[28] by contrast, the addition of autologous monocytes mimics the PD-L1hi environment of the liver.[29]

Summary

Introduction

CD8 T cells are critical for immune control of persistent viral infections and cancer. A therapeutic response to antibody-mediated checkpoint blockade requires the tumor to have a relatively high mutation burden and a pre-existing lymphocytic infiltrate.[4,5,6] The use of blocking monoclonal antibodies means that effects are of limited duration and require repeated dosing, with its associated problems. Cells expressing PD-1 will potentially be affected, resulting in the unleashing of bystander and autoreactive T cell specificities and a substantial risk of autoimmune disease.[7] Regulatory populations such as Tregs can express high levels of PD-1, so PD-1 blockade can expand regulatory T cells (Tregs), which will tend to counteract the boosting of effector T cells.[8]

Methods

Results

Conclusion