Comparative proteomic analysis of HPV(+) oropharyngeal squamous cell carcinoma recurrence

The global rise of HPV(+) oropharyngeal squamous cell carcinoma (OPSCC) has generated considerable interest underlying its etiology and management. Despite an overall decline in head and neck malignancies, the incidence in OPSCC has by contrast sharply risen, with HPV(+) subtypes now comprising 80% of all OPSCC.1,2 Relative to HPV(-) OPSCC, HPV(+) patients are more responsive to chemoradiation and harbor a 52% risk-of-death reduction.3 Despite this distinct outcome, treatment regimens remain the same for all OPSCC subtypes (smoking-driven and virus-driven), rather than an adaptive approach to what most consider distinct diseases. The short-term effects (e.g., mucositis, odynophagia) and long-term toxicities (e.g., xerostomia, dysphagia, ototoxicity) from treatment substantially affect quality of life, and rival the impact of the cancer itself. Recently published as well as ongoing trials are actively examining deintensification approaches2,4-9, with the goal of diminishing treatment sequelae for HPV(+) subtypes. While deintensification may decrease chemoradiation-related toxicities, it nonetheless may also undertreat a meaningful percentage of HPV(+) patients who may then recur. In examining national trials, 19% of HPV(+) patients had disease progression after therapy, with a two-year survival rate of 60%.10 Current methods to ascertain HPV status involve p16 staining. However, on comparison with gold standard E6/E7 expression by qPCR, p16 harbored a 15% false positivity rate11, suggesting limited utility as a sole biomarker for deintensification. Improved molecular stratification would greatly enhance the clinician’s ability to precisely tailor treatment while minimizing the risk of jeopardizing outcomes. One approach encompasses in-depth proteomic profiling of HPV(+) OPSCC to reveal distinct protein expression profiles and delineate clinically relevant upstream pathways. In turn, these proteomic differences may distinguish higher-risk disease (cases predisposed to recurrence that may benefit from treatment intensification) from lower-risk phenotypes (cases whose treatment response is sufficiently robust to warrant deintensification). Here, two HPV(+) OPSCC cohorts stratified by treatment response are compared via a hybrid data dependent acquisition/data independent acquisition (DDA/DIA) approach via mass spectrometry. We focused on detection of low-abundance proteins to highlight proteomic signatures that can be potentially exploited for treatment stratification.

全球范围内HPV阳性（human papillomavirus-positive, HPV）口咽鳞状细胞癌（oropharyngeal squamous cell carcinoma, OPSCC）的发病率持续攀升，引发学界对其病因及诊疗策略的广泛关注。尽管头颈部恶性肿瘤整体呈下降趋势，但OPSCC的发病率却逆势大幅上涨，目前HPV阳性亚型已占全部OPSCC病例的80%[1,2]。与HPV阴性（HPV(-)）OPSCC患者相比，HPV阳性患者对放化疗（chemoradiation）的响应更佳，死亡风险可降低52%[3]。尽管预后特征如此迥异，但当前所有OPSCC亚型（吸烟驱动型与病毒驱动型）的治疗方案均保持统一，并未针对这类被大多数学者视为两类截然不同的独立疾病，采用个体化适配的诊疗策略。
治疗引发的短期不良反应（如黏膜炎（mucositis）、吞咽痛（odynophagia））与长期毒性反应（如口干症（xerostomia）、吞咽困难（dysphagia）、耳毒性（ototoxicity））会严重影响患者的生存质量，其影响程度甚至不亚于肿瘤本身。近期发表及正在进行的多项临床试验正积极探索降强度治疗（deintensification）策略[2,4-9]，以期减轻HPV阳性亚型患者的治疗后遗症。尽管降强度治疗或许能够降低放化疗相关毒性，但也有可能对相当比例的HPV阳性患者治疗不足，进而导致疾病复发。据一项全国性临床试验数据显示，19%的HPV阳性患者在治疗后出现疾病进展，两年生存率仅为60%[10]。
当前用于确定HPV感染状态的检测方法主要为p16染色。然而，与以实时定量聚合酶链反应（quantitative polymerase chain reaction, qPCR）检测E6/E7表达这一金标准相比，p16检测的假阳性率高达15%[11]，这表明单独将其作为降强度治疗筛选的生物标志物存在一定局限性。
更为精准的分子分层技术将极大提升临床医生精准定制治疗方案的能力，同时最大限度降低因治疗不当危及患者预后的风险。其中一种思路是对HPV阳性OPSCC开展深度蛋白质组学分析，以揭示其独特的蛋白质表达谱，并厘清与临床相关的上游信号通路。据此，这些蛋白质组学差异或可区分高风险疾病亚型（即易复发、需强化治疗的病例）与低风险表型（即治疗响应足够良好、可接受降强度治疗的病例）。
本研究通过质谱分析（mass spectrometry），采用数据依赖采集/数据非依赖采集（data dependent acquisition/data independent acquisition, DDA/DIA）混合策略，对两个基于治疗响应分层的HPV阳性OPSCC队列进行了比较分析。本研究重点关注低丰度蛋白质的检测，以期挖掘可用于治疗分层的潜在蛋白质组学特征。