Abstract A78: Antibody profiling of melanoma patients using a custom protein microarray platform
Authors: Duarte Jessica , Peyper Janique , Woods Katherine , Tsao Simon Chang-Hao , Cebon Jonathan , Blackburn Jonathan 
Published: March 2017

Abstract
There is substantial evidence that the mainly tumor-restricted aberrant expression of the highly immunogenic cancer-testis (CT) antigens across several different cancer types makes them attractive cancer diagnostic and predictive biomarkers. Therefore, we aimed to measure differences in CT antigen-specific antibody repertoires between melanoma patient samples, and assess whether these could identify novel diagnostic or prognostic biomarkers, which could aid in the detection and management of cancer. In order to detect and quantify CT-antigen-specific antibodies circulating in the serum or plasma of cancer patients, we co-developed a novel CT antigen microarray platform. This tool represents a high-throughput means of profiling antibody repertoires against over 100 cancer-restricted antigens in a highly reproducible manner with high sensitivity (detection limit: 10-100pg/ml) and specificity. In addition, it provides a significant advantage over tissue-based approaches, due to the ease of sample access without the need for invasive procedures. We carried out a retrospective serological study of antibody titers across a large cohort of eighty-eight mainly advanced malignant melanoma patients (8% unknown stage, 2% stage I, 10% stage II, 40% stage III and 40% stage IV) undergoing a variety of distinct cancer treatments (surgery, chemotherapy, radiotherapy, immunotherapy or none), using our recently developed and validated novel CT antigen microarray platform. We successfully identified abundant antibody titers towards two leading CT antigens, NY-ESO-1 and CTAG2 - a highly homologous pair - expressed across 50% (n = 44/88) and 45% (n = 40/88) of all patients, respectively, with reported diagnostic and clinical efficacy predictive biomarker potential. Furthermore, the overlap in antibody titers of this pair, most likely due to a shared immunogenic epitope, validated our array platform, showing consistency amongst our obtained results across patients. Additional abundant titers were found across fifteen further antigens, including several members of the melanoma antigen (MAGE) family, such as MAGEB1 (n = 30/88, 34%), MAGEB6 (n = 29/88, 33%), MAGEB5 (n = 26/88, 29%), MAGEA3 (n = 26/88, 29%) and MAGEA11 (n = 24/88, 27%). Abundant antibody titers towards at least one of these leading seventeen cancer antigens were detected across 82% of this cohort (n = 72/88), thus emphasizing the potential uses of these as diagnostic or prognostic biomarkers, warranting further larger studies. Furthermore, healthy individuals showed no detectable cancer-antigen-specific antibody titers, therefore highlighting the specificity of this tool in a cancer setting. In conclusion, we showed that our novel protein microarray platform represents a sensitive, high-throughput and customizable means to detect and quantify the presence of large panels of cancer-specific human antibodies in serum, obtaining consistently robust, high quality and reproducible data, and demonstrating its potential feasibility and inferred biological significance.
Citation Format: Jessica Duarte, Janique Peyper, Katherine Woods, Simon Chang-Hao Tsao, Jonathan Cebon, Jonathan Blackburn. Antibody profiling of melanoma patients using a custom protein microarray platform. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr A78.