Abstract CT018: Intratumor and peripheral Treg modulation as a pharmacodynamic biomarker of the GITR agonist antibody TRX-518 in the first in-human trial
Authors: Zappasodi Roberta , Li Yanyun , Abu-Akeel Mohsen , Qi Jingjing , Wong Philip , Sirard Cynthia , Postow Michael , Schaer David A. , Newman Walter , Koon Henry , Velcheti Vamsidhar , Callahan Margaret K. , Wolchok Jedd D. , Merghoub Taha 
Published: July 2017

Abstract
Background: The tumor necrosis factor receptor GITR is expressed at high levels on regulatory T cells (Tregs) and up-regulated on activated T cells. Evidence from preclinical studies has shown the dual activity of GITR stimulation to counteract Tregs functionally and quantitatively and enhance T-cell effector functions, thus providing the rationale to develop GITR agonist antibodies for cancer immunotherapy. These agents have demonstrated potent therapeutic effects associated with both Treg reduction and modulation in tumor-bearing mice. Here we investigate Treg modulation as a biomarker of activity of the fully humanized agonist anti-GITR antibody TRX-518 in advanced solid cancer patients and the potential mechanisms underlying this effect. Methods: Patients were accrued to 9 cohorts (up to 6 patients/cohort) to receive a single dose of TRX518 (dose range: 0.0001-8 mg/kg). Pharmacodynamic analyses included flow cytometric evaluation of frequency and phenotype of circulating T cells at different time points up to 12 weeks after treatment. Relevant changes observed with these analyses were monitored in pre- and post-treatment tumor biopsies by immunofluorescence staining. Effects of GITR stimulation with TRX-518 on human Tregs were studied in in vitro cultures with CD4+ T cells from healthy donors. Results: Analysis of peripheral blood mononuclear cells from 37 advanced refractory solid cancer patients treated with &gt;=0.005 mg/kg TRX-518 (cohorts 3-9) revealed frequent reduction in circulating Tregs after treatment with TRX-518. Proliferation potential of Tregs was not altered by TRX-518, as indicated by their persistent Ki67 expression. We then asked whether a subset of Tregs was preferentially affected and found that GITR+ Tregs and CD45RA-Foxp3hi effector Tregs (eTregs) were specifically targeted by TRX-518 in a dose-dependent manner. Accordingly, we observed that eTregs express GITR at higher levels than naïve CD45RA+Foxp3+ Tregs. In a subset of patients for whom pre- and post-treatment tumor biopsies were available (n=8; 3 melanoma, 2 lung, 2 colorectal and 1 bladder cancer patients), we tested whether intra-tumor Tregs, which are typically enriched in eTregs, were also affected. Noteworthy, we observed that changes in intra-tumor and circulating Tregs after TRX-518 were positively correlated. As a potential mechanism responsible for this effect, our initial in vitro analyses indicate that GITR stimulation with TRX-518 destabilizes Treg phenotype by down-regulating Foxp3 and enhancing expression of the prototype Th1-lineage transcription factor T-bet. This suggests that TRX-518 may favor Treg to effector-T-cell conversion, thus explaining the effect of Treg loss observed in patients. Conclusions: Circulating Treg reduction is a potential pharmacodynamic biomarker of TRX518 biological activity. This parameter may allow predictive correlation with changes in intratumoral Treg infiltration. We plan to further investigate this effect and its relevance for the association with clinical responses in our recently opened TRX518 multi-dose study. Clinical trial information: NCT01239134.
Citation Format: Roberta Zappasodi, Yanyun Li, Mohsen Abu-Akeel, Jingjing Qi, Philip Wong, Cynthia Sirard, Michael Postow, David A. Schaer, Walter Newman, Henry Koon, Vamsidhar Velcheti, Margaret K. Callahan, Jedd D. Wolchok, Taha Merghoub. Intratumor and peripheral Treg modulation as a pharmacodynamic biomarker of the GITR agonist antibody TRX-518 in the first in-human trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr CT018. doi:10.1158/1538-7445.AM2017-CT018