Objectives

CD66b positive tumor-infiltrating neutrophils (TINs) are key immunity cells in the tumor microenvironment (TME). However, their relationship with clinicopathological features, immune checkpoints (ICs), and prognostic value remains undetermined in lung adenocarcinoma (LUAD). In this study, we aimed to characterize the infiltration by TINs and the prognostic significance in patients with surgically resected LUAD.

Materials and methods

Expression of CD66b and ICs, including PD-L1, PD-1, CTLA4, LAG3, TIM3, TIGIT, VISTA, and BTLA, in both cancer cell and tumor-infiltrating lymphocytes (TILs) were estimated by immunohistochemistry in resected LUAD. The associations between CD66b expression and clinicopathological characteristics in patient prognoses were analyzed. We also verified results in another cohort from 85 patients with untreated LUAD and further analyzed the correlation between CD66b expression and EGFR and KRAS mutation status in addition to the rearrangement of the anaplastic lymphoma receptor tyrosine kinase gene (ALK).

Results

A total of 240 patients were included in this study. CD66b expression was observed in 87 (36.2%) samples. ICs including PD-L1, PD-1, CTLA4, LAG3, TIM3, TIGIT, VISTA, and BTLA were observed in percentages that ranged from 23.8% to 59.4%. Positive CD66b expression significantly correlated with smoking history (p=0.029), pathological stage (p=0.040), and the positive expression of LAG-3 (p<0.001), PD-1 (p=0.008), CTLA-4 (p=0.013), TIM-3 (p =0.025), TIGIT (p=0.002), PD-L1 in TILs (p=0.015), and PD-L1 in tumor cells (p=0.010). CD66b positivity was significantly associated with worse recurrence-free survival (RFS) (hazard ratio, HR, 1.687; 95% confidence interval, CI, 1.058–2.690, p=0.028) and overall survival (OS) (HR, 1.667; 95% CI, 1.097–2.534, p=0.017). Subgroup analysis revealed that the CD66b+/LAG-3+ group had the worst RFS (5-year rate: 39.5%,) and OS (5-year rate: 53.7%,), while the CD66b−/LAG-3− group had the best RFS (5-year rate: 65.6%) and OS (5-year rate: 78.8%). The p value in analysis of RFS and OS was 0.005 and 0.008, respectively. In the verification set, high expression of CD66b was also significantly correlated with the positive expression of LAG-3 (p<0.001), PD-1 (p=0.002), CTLA-4 (p=0.034), TIM-3 (p=0.049), PD-L1 in TILs (p=0.003), and PD-L1 in tumor cells (p=0.045). There was no correlation between CD66b expression and positive TIGIT expression (p=0.077), EGFR mutation (p= 0.223), KRAS mutation (p= 0.151), and ALK fusion (p=0.310).

Conclusion

CD66b had a relatively high positive expression rate and special clinicopathological features in patients with LUAD. CD66b+ TINs were related to the expression of ICs and associated with poor prognoses in LUAD. A combination of CD66b and ICs, especially LAG-3 could further stratify patients into different groups with distinct prognoses.

中文翻译：

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CD66b 的特征及其与免疫检查点的关系及其对手术切除肺腺癌预后的协同影响

目标

CD66b 阳性肿瘤浸润中性粒细胞 (TIN) 是肿瘤微环境 (TME) 中的关键免疫细胞。然而，它们与临床病理特征、免疫检查点 (IC) 和预后价值的关系在肺腺癌 (LUAD) 中仍未确定。在这项研究中，我们旨在表征 TIN 的浸润和手术切除的 LUAD 患者的预后意义。

材料和方法

CD66b 和 IC 的表达，包括 PD-L1、PD-1、CTLA4、LAG3、TIM3、TIGIT、VISTA 和 BTLA，在癌细胞和肿瘤浸润淋巴细胞 (TIL) 中的表达通过免疫组织化学在切除的 LUAD 中进行评估。分析了 CD66b 表达与患者预后中临床病理特征之间的关联。我们还验证了来自 85 名未经治疗的 LUAD 患者的另一个队列的结果，并进一步分析了 CD66b 表达与 EGFR 和 KRAS 突变状态之间的相关性，以及间变性淋巴瘤受体酪氨酸激酶基因 (ALK) 的重排。

结果

本研究共纳入 240 名患者。在 87 个 (36.2%) 样品中观察到 CD66b 表达。包括 PD-L1、PD-1、CTLA4、LAG3、TIM3、TIGIT、VISTA 和 BTLA 在内的 IC 被观察到的百分比范围从 23.8% 到 59.4%。CD66b阳性表达与吸烟史（p =0.029）、病理分期（p =0.040）、LAG-3（p <0.001）、PD-1（p =0.008）、CTLA-4（p）阳性表达显着相关=0.013)、TIM-3 ( p =0.025)、TIGIT ( p =0.002)、TILs 中的 PD-L1 ( p =0.015) 和肿瘤细胞中的 PD-L1 ( p=0.010）。CD66b 阳性与较差的无复发生存 (RFS)（风险比，HR，1.687；95% 置信区间，CI，1.058–2.690，p = 0.028）和总生存（OS）（HR，1.667；95%）显着相关CI，1.097–2.534，p = 0.017）。亚组分析显示，CD66b+/LAG-3+ 组的 RFS（5 年率：39.5%）和 OS（5 年率：53.7%）最差，而 CD66b-/LAG-3- 组最好的 RFS（5 年利率：65.6%）和 OS（5 年利率：78.8%）。RFS 和 OS 分析中的p值分别为 0.005 和 0.008。在验证集中，CD66b的高表达也与LAG-3（p <0.001）、PD-1（p =0.002）、CTLA-4（p）的阳性表达显着相关。=0.034)、TIM-3 ( p =0.049)、TIL 中的 PD-L1 ( p =0.003) 和肿瘤细胞中的 PD-L1 ( p =0.045)。CD66b表达与TIGIT阳性表达（p =0.077）、EGFR突变（p =0.223）、KRAS突变（p =0.151）和ALK融合（p =0.310）之间无相关性。

结论

CD66b在LUAD患者中具有较高的阳性表达率和特殊的临床病理特征。CD66b+ TIN 与 IC 的表达有关，并与 LUAD 的不良预后相关。CD66b 和 IC，尤其是 LAG-3 的组合可以进一步将患者分为具有不同预后的不同组。