Abstract 2208: Rare neoplastic subpopulation predicts relapse in nasopharyngeal carcinoma
Authors: Wu Ka Chun , Yu Ken , Su Nelson , Lau Jeffrey Yan Ho , Lo Anthony , Kam Yvonne Ngar Woon , Tsang Raymond , Chung Joseph , Ho Joshua , Che Chi Ming , Lee Victor Ho Fun 
Published: June 2023

Abstract
Nasopharyngeal carcinoma (NPC), with the highest incidence and prevalence in Southern China including Hong Kong, originates from the nasopharyngeal epithelium and frequently metastasises to cervical lymph nodes and distal organs such as liver. Radiation therapy with/without concurrent chemoradiation is the most common first-line treatment. Although most patients respond to the treatment, about 30-50% of treated patients still relapse. Bulk sequencing uncovered numerous disease-associated genes, but still there is no effective targeted therapy or immunotherapy to prevent relapse. Tools for early relapse prediction at initial diagnosis still remain unavailable.
Recently, single-cell RNA sequencing (scRNA-seq) technology has emerged as a sensitive long-sought solution to low resolution in bulk seqeuncing. We launched the largest and most comprehensive scRNA-seq NPC study in the world. 55 patients were recruited. Matched biopsies from normal adjacent tumour, primary tumour and lymph node malignancy sites were harvested and 70488 epithelial cells had been successfully scRNA-seq profiled. Unsupervised clustering identified 9 different epithelial subclusters. Copy number variation (CNV) analysis identified neoplastic cells with wide-spread CNV aberrations consistent with previous bulk genomic findings. For the first time, we had also attempted to unfold single nucleotide variation (SNV) of neoplastic cells from scRNA-seq using modified Mutect2 pipeline. Our study spanned 4 years and 5 patients were reported relapsed. Encouragingly, survival analysis identified rare neoplastic subcluster 1 as the only neoplastic subcluster positively correlated with relapse. Abundance of neoplastic subcluster 1 biomarker-positive cells were verified to be elevated in FFPE primary tumours of relapsed patients collected at initial diagnosis. Deconvolution analysis of a publicly available bulk RNA sequencing dataset revealed only cells harbouring neoplastic subcluster 1 expression signature tended to positively correlate with relapse. Interestingly, cross-tissue abundance analysis identified neoplastic subcluster 1 was positively correlated with preferential colonialization at cervical lymph node. Single-cell gene-set enrichment analysis (scGSEA) suggested neoplastic subcluster 1 possessed higher metastasis and treatment resistance potential. For the first time, we had also investigated the cell-cell communications among epithelial cells and identified interaction pathways implicated in self-renewal and tumorigenic capacity of cancer-initiating cells to be associated with neoplastic subcluster 1 and relapse.
To conclude, we successfully discovered a previously overlooked rare neoplastic subpopulation in NPC that may (1) possess higher metastasis and treatment resistance capacity, and (2) play a regulatory role in tumourigenesis, and its abundance at initial diagnosis could predict relapse.
Citation Format: Ka Chun Wu, Ken Yu, Nelson Su, Jeffrey Yan Ho Lau, Anthony Lo, Yvonne Ngar Woon Kam, Raymond Tsang, Joseph Chung, Joshua Ho, Chi Ming Che, Victor Ho Fun Lee. Rare neoplastic subpopulation predicts relapse in nasopharyngeal carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2208.