Abstract 5002: Investigating the role of NF- κB signaling and immune checkpoint blockade therapy in melanoma
Authors: Wells Keith , Hintzsche Jennifer , Amato Carol M. , Tobin Richard , Vorwald Victoria , McCarter Martin , Shellman Yiqun , Tan Aik Choon , Robinson William 
Published: August 2019

Abstract
Immune checkpoint blockade (ICB) treatments, such as anti-PD1, improves survival in patients with metastatic melanoma. Despite durable response rates, 40-50% of patients show no benefit. Implicated in acquired resistance is the downregulation of antigen presentation, via dysfunction of the interferon-receptor pathway. However, little is known about intrinsic response mechanisms to ICB. The NF-κB-pathway is important for immune response as a key transcription factor for several antigen presentation proteins and cytokine/chemokine production. This study explored NF-κB signaling in patients with favorable, or unfavorable responses to anti-PD1 therapy. Fifty-two patients with metastatic melanoma were grouped into complete or partial responders (CR/PR; n = 6 and 15, respectively), and stable or progressive disease (SD/PD; n = 16 and 15, respectively). Consistent with other studies, tumor mutational burden assessed by whole exome sequencing (WES) was higher in the CR/PR group (p = 0.03). WES analysis also showed an enrichment of variants in NF-κB-pathway related genes within the CR/PR group (14/21, 67%) compared with SD/PD patients (6/31, 19%), p = 0.001. Notably, the hotspot variants G34E/R and G41E in NFKBIE, a negative regulator of NF-κB, were found exclusively in patients with complete or partial response to ICB (5/21, 24%). RNA-seq analysis of 19 tumors showed that the TNFα/NF-κB signaling pathway is upregulated in the CR/PR group, and highly expressed genes included CD83, HLA-DQB, and NFATC1. Therefore we hypothesized that when activated, particularly within the melanoma cells, this pathway contributes to an anti-tumor immune response. Using HEK293 cells, we demonstrated NFKBIEG34E increased NF-κB activity compared with the wildtype, upon TNFα stimulation. Consistently, flow cytometry also show an increase of targets of NF-κB transcription such as CD83, MHC class II, and PDL1. Similarly, the melanoma cell line A375, which harbors the NFKBIEG34E mutation, showed increases in protein expression when stimulated with TNFα and INFγ. Furthermore, progression free survival increased in patients that had tumors with an NF-κB-pathway variant compared with patients that were pathway wildtype (p = 0.03). Patients harboring tumors with NF-κB-pathway variants also had a greater response by RECIST criteria compared with wildtype. Taken together, our results suggest that an increase of pro-immunogenic NF-κB signaling in the tumor may contribute to the favorable response to ICB, highlighting a role for CD83 in the tumor microenvironment that merits further examination.
Citation Format: Keith Wells, Jennifer Hintzsche, Carol M. Amato, Richard Tobin, Victoria Vorwald, Martin McCarter, Yiqun Shellman, Aik Choon Tan, William Robinson. Investigating the role of NF- κB signaling and immune checkpoint blockade therapy in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5002.