Abstract 4559: Novel T-cell targets for HPV16-associated oropharyngeal cancer immune therapy
Authors: Krishna Sri , Parikh Falguni , Ulrich Peaches , Yang Shanshan , Clark Amelia , Kim-Schulze Seunghee , Posner Marshall , Park Jin , Sikora Andrew , Anderson Karen S. 
Published: July 2017

Background: Human papillomavirus subtype 16 (HPV16) is the primary cause of an increasing number of oropharyngeal cancers (HPVOPC) worldwide. Despite effective prophylactic vaccines, low vaccine uptake and high incidence of HPV16 infections provides a strong rationale for the development of novel targeted immune therapies against HPVOPC. We sought to 1) Identify novel CD8+ T-cell (CTL) epitopes from HPV16 antigens; 2) Understand mechanisms of immune resistance in HPVOPC to enhance T-cell based immune therapies.

Methods: We developed a comprehensive bioinformatic strategy to predict potential CTL epitopes for 10 HLA class I alleles representing common HLA-supertypes (with &gt;90% global coverage) against HPV16 antigens E2, E6 and E7. T-cell reactivity against candidate epitopes was assessed by short term ex vivo T-cell cultures from healthy donors (n=15) and HPVOPC patients (n=18). We data mined publicly available OPC transcriptomes (University of Michigan (UM-OPC) cohort, n=36) for gene expression analysis.

Results: IFN-gamma Elispot assays using 58 candidate epitopes from HPV16 E2, E6 and E7 revealed that &gt;70% of HPVOPC patients had either E2 or E6-specific CTL reactivity. E2-CTL reactivity was &gt;2-fold higher in HPVOPC patients than in healthy controls (mean = 68.75 vs. 24.6 SFUs/106 cells, P = 0.027), while no difference in E6- CTL responses were observed between patients and controls. In contrast, E7-specific CTL responses were low to undetectable. We have further identified 7 novel E2 and 3 novel E6 CTL-epitopes, and confirmed immunogenicity of previously described E2, E6 and E7 epitopes. RNAseq analysis of the UM-OPC dataset indicated that 78% of HPV16+ OPCs expressed E2 (P&lt;0.05), while E6 and E7 was expressed in all HPVOPCs (P&lt;0.01) demonstrating tumor antigen expression. Unsupervised hierarchical clustering of immune transcriptomes revealed that HPVOPCs have higher expression of immune-regulatory ligands than HPV-negative OPC tumors, indicating high immune infiltration and possible immune-editing. We identified high expression of several key immune exhaustion genes such as IDO-1, TIM-3 and LAG-3, previously undescribed in HPVOPC.

Conclusion: HPV16-E2 represents a novel antigen target for HPVOPC immune therapy, as demonstrated by T-cell reactivity and tumor antigen expression. Several novel and previously defined E2, E6 and E7 CTL have been confirmed for T-cell reactivity in HPVOPC patients. Transcriptomic analysis of OPCs indicate extensive immune dysfunction in HPVOPCs that are distinct from non-HPV OPCs. The T-cell epitopes identified in this study in combination with blockade of HPVOPC-specific checkpoints may be useful for targeted immunotherapy.

Citation Format: Sri Krishna, Falguni Parikh, Peaches Ulrich, Shanshan Yang, Amelia Clark, Seunghee Kim-Schulze, Marshall Posner, Jin Park, Andrew Sikora, Karen S. Anderson. Novel T-cell targets for HPV16-associated oropharyngeal cancer immune therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4559. doi:10.1158/1538-7445.AM2017-4559