Abstract 426: Autoantibodies: A promising prognostic tool for immunotherapy response in advanced non-small cell lung cancer
Authors: Tarhoni Imad , Fhied Cristina , Pergande Melissa , Kollipara Revathi , Wakefield Connor J. , Gallo Katherine , Tangri Apoorva , Batus Marta , Fidler Mary Jo , Bonomi Philip , Borgia Jeffrey A. 
Published: August 2019

Abstract
Background: Immune-checkpoint blockade has revolutionized cancer therapy in advanced non-small cell lung cancer (NSCLC). Tissue expression of programmed death protein ligand (PD-L1) remains the gold standard for patient stratification, however, the limited performance of this marker encourages investigations for improved molecular diagnostics. The objective of this study is to identify and evaluate the role of neoantigen-associated autoantibodies to predict the clinical response to anti-PD-1/-L1 in advanced stage NSCLC.
Method: Lung adenocarcinoma A549 and H358 cell lysate proteins were resolved via 2-dimensional electrophoresis, electroblotted onto nitrocellulose, and immunoprobed with pooled, pretreatment sera (n= 4/ group) derived from patients with advanced NSCLC who received PD-1/-L1 directed immunotherapy. These patients have documented disease progression within 12 weeks (“rapid progression”) or demonstrated radiographical stable disease/progression after the first 180 days of therapy (“late progression”). Immunoreactive spots were detected with an HRP-conjugated, anti-human IgG secondary antibody with digital densitometry. A 4-fold cutoff threshold in expression was used to prioritize spots for identification via tandem mass spectrometry. From A549 cells, recombinant proteins were selected for STIP-1, annexin A2, HSPA8, and GAPDH. These proteins were then analyzed via immunoblotting methods using sera from each indicated group (n=4 per group). In addition, identified proteins from H358 cells include FH, HSP70B, IMPDH2, NY ESO-1, PGAM-1, and vimentin. Recombinant versions of a selection of autoantigens identified in this manner were commercially acquired and used to develop custom Luminex immunobead assays to quantitatively assess autoantibody production in individual patient sera (rapid progressors, n=14; late progressors, n=18). Values were statistically compared via Mann-Whitney test.
Results: Series of differentially expressed autoantigens predictive of clinical response to PD-1/-L1 directed immunotherapy were identified. Western blots of neoantigens identified from A549 cells; STIP-1, annexin A2, HSPA8, and GAPDH were significantly able to distinguish between response groups (p-value &lt; 0.001). Six H358 targets resulting from the custom bead-based immunoassay development; FH, HSP70B, IMPDH2, NY-ESO-1, PGAM-1, and vimentin were also able to distinguish between groups (p-values of 0.01, 0.01, 0.022, 0.005, 0.034, and 0.027, respectively).
Conclusion: Our study demonstrates that serum autoantibodies have great promise to serve as a robust tool to prognosticate response for patients receiving PD-1/-L1 directed immunotherapy and potentially aid current treatment selection methods. Additional targets are currently being developed into multiplexed immunobead assays for evaluation across larger cohorts of patients.
Citation Format: Imad Tarhoni, Cristina Fhied, Melissa Pergande, Revathi Kollipara, Connor J. Wakefield, Katherine Gallo, Apoorva Tangri, Marta Batus, Mary Jo Fidler, Philip Bonomi, Jeffrey A. Borgia. Autoantibodies: A promising prognostic tool for immunotherapy response in advanced non-small cell lung cancer [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 426.