Abstract

Background

Vulvar High-grade Squamous Intraepithelial Lesion (vHSIL) is predominantly induced by high-risk Human Papilloma Virus type 16 (HPV16). In two independent trials, therapeutic vaccination against the oncoproteins of HPV16 with synthetic long peptides (SLP) resulted in vHSIL regression in about half of the patients after 12 months. Several studies have shown that the immune microenvironment influences therapy outcome. Therefore, a thorough investigation of the vHSIL immune microenvironment before and after SLP vaccination was performed, and its impact on clinical response was studied.

Methods

Two novel multiplex immunofluorescence panels were designed for formalin-fixed paraffin-embedded tissue, one for T cells (CD3, CD8, FoxP3, Tim3, Tbet, PD-1, DAPI) and one for myeloid cells (CD14, CD33, CD68, CD163, CD11c, PD-L1, DAPI). Pre- and 3 months post-vaccination biopsies of 29 patients and 27 healthy vulva excisions were stained, scanned with the Vectra multispectral imaging system, and automatically phenotyped and counted with inForm advanced image analysis software.

Results

A pre-existing pro-inflammatory TME, marked by high numbers of CD4 and CD8 T cells expressing Tbet and/or PD-1 as well as CD14+ inflammatory macrophages, is a strong predictor for good clinical response. A clear stepwise increase in pre-vaccination infiltrating Tbet+, CD4+, CD8+ T cells and CD14+ macrophages, and decrease in Foxp3+ Tregs was observed as response increased from non to partial to complete response. Moreover, the pre-vaccination immune microenvironment of complete responders resembled healthy vulva. Vaccination further increased infiltrating CD4+ and Tbet+ T cells and CD14+ macrophages and decreased FoxP3+ Tregs in the complete and partial responders, but not in the non responders.

Conclusion

Clinical responsiveness to therapeutic HPV16 SLP vaccination requires a pre-existing inflamed type 1 immune contexture in vHSIL. Hence, only patients with an inflamed TME should be selected for monotherapy by therapeutic vaccination, since this strategy is incapable of creating an inflamed TME in patients where this is absent.

Legal entity responsible for the study

The authors.

Funding

Leiden University Medical Center.

Disclosure

S.H. van der Burg: Advisory / Consultancy: ISA Pharmaceuticals. All other authors have declared no conflicts of interest.

中文翻译：

---

3O预先存在的炎症性免疫微环境预测外阴高级别鳞状上皮内病变对治疗性HPV16肽疫苗的临床反应和免疫反应

抽象的

背景

外阴高级别鳞状上皮内病变（vHSIL）主要由高危型人乳头瘤病毒16型（HPV16）诱导。在两项独立试验中，用合成长肽（SLP）对HPV16癌蛋白进行的治疗性疫苗接种导致12个月后约一半的患者出现vHSIL下降。几项研究表明，免疫微环境会影响治疗效果。因此，对SLP疫苗接种前后的vHSIL免疫微环境进行了彻底的调查，并研究了其对临床反应的影响。

方法

为福尔马林固定石蜡包埋的组织设计了两种新型的多重免疫荧光板，一种用于T细胞（CD3，CD8，FoxP3，Tim3，Tbet，PD-1，DAPI），一种用于髓样细胞（CD14，CD33，CD68，CD163） ，CD11c，PD-L1，DAPI）。接种前和接种后3个月对29例患者和27例健康的外阴切除活组织进行染色，使用Vectra多光谱成像系统进行扫描，并使用inForm先进的图像分析软件自动进行表型分析和计数。

结果

以高表达Tbet和/或PD-1的CD4和CD8 T细胞以及CD14 +炎性巨噬细胞为特征的预先存在的促炎性TME是良好临床反应的有力预测指标。观察到随着反应从非反应到部分反应到完全反应的增加，浸润前的Tbet +，CD4 +，CD8 + T细胞和CD14 +巨噬细胞明显逐步增加，而Foxp3 + Tregs减少。此外，完全应答者的疫苗接种前免疫微环境类似于健康的外阴。在完全和部分反应者中，疫苗接种进一步增加了浸润性CD4 +和Tbet + T细胞以及CD14 +巨噬细胞的表达，并减少了FoxP3 + Treg，但在非反应者中没有。

结论

对治疗性HPV16 SLP疫苗的临床反应要求在vHSIL中预先存在发炎的1型免疫情况。因此，只有具有TME炎症的患者才应选择通过治疗性疫苗进行单药治疗，因为这种策略无法在不存在TME的患者中产生TME炎症。

负责研究的法人实体

作者。

资金

莱顿大学医学中心。

揭露

SH van der Burg：咨询/顾问：ISA Pharmaceuticals。所有其他作者都声明没有利益冲突。