Abstract 4168: Tumor cell stimulator of interferon genes (STING) activation primes NK cell therapy in mesothelioma
Authors: Knelson Erik H. , Ivanova Elena , Tarannum Mubin , Campisi Marco , Lizotte Patrick H. , Booker Matthew A. , Meisenheimer Brittany , Chen Minyue , Spicer Nathaniel , Obua Bonje , Vo Ha V. , Albertson Thomas P. , Vivero Marina , Tolstorukov Michael Y. , Romee Rizwan , Paweletz Cloud P. , Bueno Raphael , Barbie David A. 
Published: June 2022

Abstract
Background: Activation of the anti-viral STING pathway promotes antitumor immunity but STING agonists have yet to achieve clinical success. Understanding their mechanism of action in human tumors is key to developing STING-based therapies and combinations. Higher tumor STING expression correlates with better response to treatment and multiple cancers have been shown to silence STING and the downstream interferon response to avoid immune detection, demonstrating an important role for tumor cell STING in antitumor immunity. Recent mouse studies have explored the complex interplay of STING activation in the tumor immune microenvironment (TIME), identifying unexpected T-cell toxicity and novel effector mechanisms including NK cells. Yet how STING agonists impact the human TIME has not been carefully examined and could inform development of novel therapeutic combinations. Here we address this question using human tumors cultured ex vivo to maintain the TIME, dissecting STING agonist response in malignant pleural mesothelioma (MPM), an inflamed cancer with high STING expression.
Methods: We performed STING immunohistochemistry on 300 thoracic tumor specimens, followed by functional ex vivo studies of STING agonist response in human mesothelioma explants, measuring cell death and cytokine production. Dynamic single cell RNA sequencing was performed after treatment with a STING agonist. Primary human T-cells and NK cells were used to study STING agonist toxicity in the context of STING cycling and patient derived organotypic tumor spheroids (PDOTS) were treated for six days with STING agonists +/- NK and anti-mesothelin chimeric antigen receptor (CAR)-NK cell therapies.
Results: Amongst thoracic cancers, MPM robustly expressed tumor cell STING and was responsive to STING agonist treatment ex vivo. We observed tumor cell death &gt;20%, p&lt;0.05 in 12/35 MPM PDOTS after STING agonist treatment. Cell death was prevented by CD8 neutralizing antibody co-treatment. Dynamic single-cell RNA sequencing of MPM explants treated with a STING agonist unveiled CXCR3 chemokine activation primarily in tumor cells and cancer associated fibroblasts, coupled with an interferon stimulated gene program. However, STING agonism was also toxic to T-cells. In contrast, primary NK cells manifested rapid STING turnover and resisted STING agonist-induced toxicity. STING agonists enhanced NK and especially anti-mesothelin CAR-NK cell migration and killing, improving therapeutic activity in MPM PDOTS.
Conclusions: MPM, an inflamed cancer type with marginal response to immune checkpoint blockade, demonstrated high tumor cell STING expression and response to STING agonists in combination with NK cell therapies ex vivo. These studies reveal the fundamental importance of using human tumor samples to assess innate and cellular immune therapies, identifying STING agonism as an effective strategy to prime NK cell therapy.
Citation Format: Erik H. Knelson, Elena Ivanova, Mubin Tarannum, Marco Campisi, Patrick H. Lizotte, Matthew A. Booker, Brittany Meisenheimer, Minyue Chen, Nathaniel Spicer, Bonje Obua, Ha V. Vo, Thomas P. Albertson, Marina Vivero, Michael Y. Tolstorukov, Rizwan Romee, Cloud P. Paweletz, Raphael Bueno, David A. Barbie. Tumor cell stimulator of interferon genes (STING) activation primes NK cell therapy in mesothelioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4168.