Despite pronounced associations of major histocompatibility complex (MHC) regions with nasopharyngeal carcinoma (NPC), causal variants underlying NPC pathogenesis remain elusive. Our large-scale comprehensive MHC region deep sequencing study of 5689 Hong Kong Chinese identifies eight independent NPC-associated signals and provides mechanistic insight for disrupted transcription factor binding, altering target gene transcription. Two novel protective variants, rs2517664 (Trs2517664 = 4.6%, P = 6.38 × 10−21) and rs117495548 (Grs117495548 = 3.0%, P = 4.53 × 10−13), map near TRIM31 and TRIM39/TRIM39-RPP21; multiple independent protective signals map near HLA-B including a previously unreported variant, rs2523589 (P = 1.77 × 10−36). The rare HLA-B*07:05 allele (OR < 0.015, P = 5.83 × 10−21) is absent in NPC, but present in controls. The most prevalent haplotype lacks seven independent protective alleles (OR = 1.56) and the one with additional Asian-specific susceptibility rs9391681 allele (OR = 2.66) significantly increased NPC risk. Importantly, this study provides new evidence implicating two non-human leukocyte antigen (HLA) genes, E3 ubiquitin ligases, TRIM31 and TRIM39, impacting innate immune responses, with NPC risk reduction, independent of classical HLA class I/II alleles.

Background Nasopharyngeal carcinoma (NPC) is an important cancer in Hong Kong. We aim to utilise liquid biopsies for serial monitoring of disseminated NPC in patients to compare with PET-CT imaging in detection of minimal residual disease. Method Prospective serial monitoring of liquid biopsies was performed for 21 metastatic patients. Circulating tumour cell (CTC) enrichment and characterisation was performed using a sized-based microfluidics CTC chip, enumerating by immunofluorescence staining, and using target-capture sequencing to determine blood mutation load. PET-CT scans were used to monitor NPC patients throughout their treatment according to EORTC guidelines. Results The longitudinal molecular analysis of CTCs by enumeration or NGS mutational profiling findings provide supplementary information to the plasma EBV assay for disease progression for good responders. Strikingly, post-treatment CTC findings detected positive findings in 75% (6/8) of metastatic NPC patients showing complete response by imaging, thereby demonstrating more sensitive CTC detection of minimal residual disease. Positive baseline, post-treatment CTC, and longitudinal change of CTCs significantly associated with poorer progression-free survival by the Kaplan–Meier analysis. Conclusions We show the potential usefulness of application of serial analysis in metastatic NPC of liquid biopsy CTCs, as a novel more sensitive biomarker for minimal residual disease, when compared with imaging.

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Background: Esophageal squamous cell carcinoma (ESCC) is a lethal and aggressive malignancy that is a threat worldwide. It is of significant prevalence in regions of Asia including Iran, Korea, Japan and especially China. Epidemiological studies of ESCC reveal genetic variations as a risk-factor to ESCC incidence. Genomic profiling of ESCC primary tumors have already identified candidate genes involved in different cellular processes such as cell cycle and apoptosis. Data from our whole exome sequencing study of primary tumors and lymph node metastases also reveal mutations involved in these pathways; however, we also uncovered an interesting candidate involved in differentiation namely, ZNF750. ZNF750 has been reported to play important roles in other squamous cell carcinomas, and as a candidate tumor suppressor gene in ESCC. Methods and Results: We exogenously upregulated ZNF750 expression by lentiviral system and used this model to investigate ZNF750 function in ESCC. By MTT 2D cell proliferation assay, we showed that in vitro cell proliferation in the ZNF750 group was hindered compared to controls. In addition, we set up 2D and 3D colony formation assays and found that the ZNF750 groups produced fewer colonies in 2D culture plates and produced significantly smaller colonies, when embedded in Matrigel. All of these findings are consistent with previous studies. We also studied the potential functional role of ZNF750 in 3D invasion/migration. Transwell permeable inserts were used to support a thick Matrigel culture with a nutrient gradient. Cells were embedded in Matrigel and moved along the gradient within the gel. After prolonged incubation (&gt;2 weeks), location and size of the colonies were examined by confocal microscopy scanning. Cells with upregulated ZNF750 were found to exhibit greater mobility compared to the control cells. We also subjected ZNF750 and control groups to cisplatin to test for chemoresistance in ESCC. We found that ZNF750 expression increases chemoresistance, as opposed to the control cells. Conclusion: From these results, ZNF750 expression inhibits cell proliferation, while enhancing cell mobility and drug resistance, all of which are characteristics of cells undergoing epithelial-mesenchymal transition (EMT). We, therefore, hypothesize that ZNF750 functions in ESSC beyond that of a tumor suppressor, but as a putative EMT regulator. Acknowledgements: Research Grants Council Collaborative Research Fund grant number 106150246 and Asian Cancer Research Fund to MLL. Citation Format: Sheyne SA Choi, Valen Zhuoyou Yu, Josephine Mun-Yee Ko, Wei Dai, Maria L. Lung. Functional characterization of ZNF750 in esophageal squamous cell carcinoma [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 174.

Nasopharyngeal carcinoma (NPC) is a malignant tumor that emerges from the epithelium of the nasopharynx, the upper part of the throat behind the nose. It has remarkable ethnic and geographical distribution. It is extremely common in Southern China including Hong Kong. Multiple factors including host genetics, environmental factors, and Epstein-Barr virus (EBV) infection contribute to NPC pathogenesis. NPC diagnosis usually occurs in people in their upper 40s and early 50s, which is close to 20 years younger than for many of the other common cancers. Familial NPC accounts for about 10% of the cases. The risk of the individuals with first-degree relatives having NPC is dramatically increased compared to those without a family history, strongly suggesting the importance of host genetics in NPC development. Recently, we used the whole-exome sequencing (WES) approach to characterize the genetic variants in the coding regions in the family-history positive (FH+) and early age onset (EAO) NPC patients. Gene-level burden tests suggested a link between multiple deleterious variants at MST1R and NPC risk, particularly in the EAO patients with disease onset at or younger than 201. In addition, we identified multiple rare deleterious variants in a number of candidate genes in at least 5% of the FH+ or EAO cases. To further investigate the role of these potential genetic susceptibility genes, we used a NGS-based targeted sequencing approach to examine the germline rare deleterious variants with minor allele frequency (MAF)&lt;0.001 in 17 genes selected from WES study in 1269 controls and 1242 cases including 334 FH+ and 904 sporadic cases. These genes are involved in DNA repair, immune response, and epigenetic regulator. Both filtering strategy and gene-level burden tests were used to identify the genetic susceptibility genes in NPC. Multiple loss-of-function (LOF) germline mutations of a critical DNA repair gene, ATM, was identified in FH+ cases. This gene plays an important role in the formation of the replication compartment during lytic replication of EBV in nasopharyngeal epithelial cells. In addition, we discovered that the rare deleterious variants of MST1R in the intracellular part may play a role in the pathogenesis in both FH+ and sporadic cases. Our study highlights the importance of innate immunity and DNA repair in NPC genetic susceptibility. Reference: 1. Dai W, Zheng H, Cheung AK, et al. Whole-exome sequencing identifies MST1R as a genetic susceptibility gene in nasopharyngeal carcinoma. Proc Natl Acad Sci U S A 2016;113:3317-22. Citation Format: Wei Dai, Hong Zheng, Josephine M.Y. Ko, Arthur K.L. Cheung, Maria Li Lung. Elucidation of genetic susceptibility genes using next-generation sequencing (NGS) approach in nasopharyngeal carcinoma [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; April 23-25, 2017; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(23_Suppl):Abstract nr 14.