Background: Immune therapies rely on successful activation of systemic immunity as well as expansion of the T-cell clones for tumor regression and successful therapeutic outcomes. PEGylated IL-10 (pegilodecakin or AM0010) monotherapy has been reported to achieve 25% objective tumor responses (ORR) in intermediate to poor risk renal cell cancer (RCC) in median 4th line of treatment (LOT) (range 1-8) (Naing A et al, JCO 2016). Here we report the immunological underpinnings of pegilodecakin induced tumor responses alone and in combination with anti-PD-1. Methods: Samples were collected post written consent from patients enrolled in a multi-basket trial (NCT0200944) and were analyzed in accordance with the IRB. Patients on AM0010 alone administered daily self-injection of pegilodecakin at 20 µg/kg, SC. Patients in the pembrolizumab + pegilodecakin received in addition pembrolizumab at 2mg/kg IV, Q3W. Normal healthy volunteers (NCT03267732) received a single (day 1) and multiple doses (days 4-9) of 5 µg/kg or 10 µg/kg AM0010 SC. Systemic and cellular antitumor immune responses were assessed by serum cytokine analysis (luminex) and by PBMC flow cytometry respectively. Additionally, immune fluorescence (IF) or immunohistochemistry was performed on formaldehyde fixed archival, pretreatment biopsies and on treatment biopsies CD8, granzyme B, phospho-STAT-3 or LAG-3, T-bet/CD3 and HLA-A. T-cell clones were quantified by TCR deep sequencing (Adaptive biotechnology) from DNA isolated using EDTA blood samples. Results: Pegilodecakin treatment induced a systemic anti-tumor immune cytokine response biased towards Th1 & Th2 cytokines (IFNg, IL-18, TNFa, IL-3, IL-4) along with IL-7. Tuppressive (TGFb) and Th17 cytokines (IL-23, IL-17) were reduced. Cytotoxic effector molecules (Granzyme B, FasL, lymphotoxinB) were increased in the serum. PBMC analysis by flow cytometry in pre- and post-treatment samples show invigoration of the exhausted, T-cells, with increased proliferative index among CD8+ T-cells expressing LAG3 and PD1 throughout pegilodecakin treatment. The increase of PD-1+ Lag-3+ Ki-67+ CD8+ T-cells correlates with objective response to AM0010. On-treatment biopsies showed that pegilodecakin increased GzmB+, Phospho-Stat3+, Lag-3+ CD8+ T-cells in the tumor. T-cell clonal analysis by TCR sequencing on PBMCs from patients during pegilodecakin treatment showed expansion of several hundred previously undetected T-cell clones per patient. Expansion of these T-cell clones in the blood correlated with tumor response, with patients with objective response showing increased number of novel clones as compared to patients with progressive disease. Conclusion: We report that pegilodecakin treatment induced a systemic Th1 immune activation with reduction of Th17 related cytokines. We further report the hallmarks of CD8+ T-cell immunity in these cancer patients, including the systemic elevation of IFNg and GranzymeB levels, expansion and activation of CD8+ TILs, and the proliferation and invigoration and expansion of PD-1+/Lag-3+ CD8+ T-cell sub-set. In addition, pegilodecakin treatment led to the expansion of T-cell clones that were undetectable pretreatment. Clinically, expansion of these novel T-cell clones during pegilodecakin treatment correlated with achievement of objective response.

Citation Format: Martin Oft, Aung Naing, Jeffrey R. Infante, Kyriakos P. Papadopoulos, Ivan H. Chan, Cong Shen, Navneet P. Ratti, Karen A. Autio, Deborah J. Wong, Manish R. Patel, Patrick A. Ott, Gerald S. Falchook, Shubham Pant, Annie Hung, John B. Mumm, Matthew Adamow, Scott McCauley, Rakesh Verma, Phillip Wong, Peter VanVlasselaer, Joseph Leveque, Nizar M. Tannir. PEGylated IL-10 (pegilodecakin) induces systemic immune activation, CD8+ T-cell invigoration and polyclonal T-cell expansion in cancer patients [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A016.