PDF(Pubmed)
Abstract：
BACKGROUND: Recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC) generally has a poor prognosis for patients with limited treatment options. While incorporating immune checkpoint inhibitors (ICIs) has now become the standard of care, the efficacy is variable, with only a subset of patients responding. The complexity of the tumor microenvironment (TME) and the role of tertiary lymphoid structures (TLS) have emerged as critical determinants for immunotherapeutic response.
METHODS: In this study, we analyzed two independently collected R/M HNSCC patient tissue cohorts to better understand the role of TLS in response to ICIs. Utilizing a multi-omics approach, we first performed targeted proteomic profiling using the Nanostring GeoMx Digital Spatial Profiler to quantify immune-related protein expression with spatial resolution. This was further characterized by spatially resolved whole transcriptome profiling of TLSs and germinal centers (GCs). Deeper single-cell resolved proteomic profiling of the TLSs was performed using the Akoya Biosciences Phenocycler Fusion platform.
RESULTS: Our proteomic analysis revealed the presence of T lymphocyte markers, including CD3, CD45, and CD8, expressing cells and upregulation of immune checkpoint marker PD-L1 within tumor compartments of patients responsive to ICIs, indicative of \'hot tumor\' phenotypes. We also observed the presence of antigen-presenting cells marked by expression of CD40, CD68, CD11c, and CD163 with upregulation of antigen-presentation marker HLA-DR, in patients responding to ICIs. Transcriptome analysis of TLS and GCs uncovered a marked elevation in the expression of genes related to immune modulation, diverse immune cell recruitment, and a potent interferon response within the TLS structure. Notably, the distribution of TLS-tumor distance was found to be significantly different across response groups (H = 9.28, p = 0.026). The proximity of TLSs to tumor cells was found to be a critical indicator of ICI response, implying that patients with TLSs located further from tumor cells have worse outcomes.
CONCLUSIONS: The study underscores the multifaceted role of TLSs in modulating the immunogenic landscape of the TME in R/M HNSCC, likely influencing the efficacy of ICIs. Spatially resolved multi-omics approaches offer valuable insights into potential biomarkers for ICI response and highlight the importance of profiling the TME complexity when developing therapeutic strategies and patient stratification.
METHODS: In this study, we analyzed two independently collected R/M HNSCC patient tissue cohorts to better understand the role of TLS in response to ICIs. Utilizing a multi-omics approach, we first performed targeted proteomic profiling using the Nanostring GeoMx Digital Spatial Profiler to quantify immune-related protein expression with spatial resolution. This was further characterized by spatially resolved whole transcriptome profiling of TLSs and germinal centers (GCs). Deeper single-cell resolved proteomic profiling of the TLSs was performed using the Akoya Biosciences Phenocycler Fusion platform.
RESULTS: Our proteomic analysis revealed the presence of T lymphocyte markers, including CD3, CD45, and CD8, expressing cells and upregulation of immune checkpoint marker PD-L1 within tumor compartments of patients responsive to ICIs, indicative of \'hot tumor\' phenotypes. We also observed the presence of antigen-presenting cells marked by expression of CD40, CD68, CD11c, and CD163 with upregulation of antigen-presentation marker HLA-DR, in patients responding to ICIs. Transcriptome analysis of TLS and GCs uncovered a marked elevation in the expression of genes related to immune modulation, diverse immune cell recruitment, and a potent interferon response within the TLS structure. Notably, the distribution of TLS-tumor distance was found to be significantly different across response groups (H = 9.28, p = 0.026). The proximity of TLSs to tumor cells was found to be a critical indicator of ICI response, implying that patients with TLSs located further from tumor cells have worse outcomes.
CONCLUSIONS: The study underscores the multifaceted role of TLSs in modulating the immunogenic landscape of the TME in R/M HNSCC, likely influencing the efficacy of ICIs. Spatially resolved multi-omics approaches offer valuable insights into potential biomarkers for ICI response and highlight the importance of profiling the TME complexity when developing therapeutic strategies and patient stratification.
摘要：
背景：复发/转移性头颈部鳞状细胞癌（R/MHNSCC）通常对治疗选择有限的患者预后不良。虽然纳入免疫检查点抑制剂（ICIs）现在已成为护理标准，功效是可变的，只有一部分患者有反应。肿瘤微环境（TME）的复杂性和三级淋巴结构（TLS）的作用已成为免疫治疗反应的关键决定因素。
方法：在本研究中，我们分析了两个独立收集的R/MHNSCC患者组织队列,以更好地了解TLS在ICIs应答中的作用.利用多组学方法，我们首先使用NanostringGeoMxDigitalSpatialProfiler进行了靶向蛋白质组学分析,以空间分辨率定量免疫相关蛋白质表达.这进一步通过TLS和生发中心（GC）的空间分辨全转录组分析来表征。使用AkoyaBiosciencesPhenocycloer融合平台进行TLS的更深层的单细胞解析蛋白质组学分析。
结果：我们的蛋白质组学分析显示存在T淋巴细胞标志物，包括CD3，CD45和CD8，表达细胞和免疫检查点标志物PD-L1在对ICIs有反应的患者肿瘤区室内的上调，指示\'热肿瘤\'表型。我们还观察到了以CD40、CD68、CD11c、和CD163与抗原提呈标记HLA-DR的上调，对ICIs有反应的患者。TLS和GC的转录组分析发现与免疫调节相关的基因表达显著升高，不同的免疫细胞募集，和TLS结构内有效的干扰素反应。值得注意的是,发现TLS-肿瘤距离的分布在应答组之间存在显著差异(H=9.28,p=0.026).发现TLS与肿瘤细胞的接近度是ICI反应的关键指标，这意味着TLS位于远离肿瘤细胞的患者的预后较差。
结论：该研究强调了TLS在调节R/MHNSCC中TME的免疫原性景观中的多方面作用，可能影响ICIs的疗效。空间分辨的多组学方法为ICI反应的潜在生物标志物提供了有价值的见解，并强调了在制定治疗策略和患者分层时分析TME复杂性的重要性。
方法：在本研究中，我们分析了两个独立收集的R/MHNSCC患者组织队列,以更好地了解TLS在ICIs应答中的作用.利用多组学方法，我们首先使用NanostringGeoMxDigitalSpatialProfiler进行了靶向蛋白质组学分析,以空间分辨率定量免疫相关蛋白质表达.这进一步通过TLS和生发中心（GC）的空间分辨全转录组分析来表征。使用AkoyaBiosciencesPhenocycloer融合平台进行TLS的更深层的单细胞解析蛋白质组学分析。
结果：我们的蛋白质组学分析显示存在T淋巴细胞标志物，包括CD3，CD45和CD8，表达细胞和免疫检查点标志物PD-L1在对ICIs有反应的患者肿瘤区室内的上调，指示\'热肿瘤\'表型。我们还观察到了以CD40、CD68、CD11c、和CD163与抗原提呈标记HLA-DR的上调，对ICIs有反应的患者。TLS和GC的转录组分析发现与免疫调节相关的基因表达显著升高，不同的免疫细胞募集，和TLS结构内有效的干扰素反应。值得注意的是,发现TLS-肿瘤距离的分布在应答组之间存在显著差异(H=9.28,p=0.026).发现TLS与肿瘤细胞的接近度是ICI反应的关键指标，这意味着TLS位于远离肿瘤细胞的患者的预后较差。
结论：该研究强调了TLS在调节R/MHNSCC中TME的免疫原性景观中的多方面作用，可能影响ICIs的疗效。空间分辨的多组学方法为ICI反应的潜在生物标志物提供了有价值的见解，并强调了在制定治疗策略和患者分层时分析TME复杂性的重要性。
