Supplementary Material for: Clinical Impact of KIR2DS3 and KIR2DL3 Genes in Neuroblastoma Patients

Objective: Neuroblastoma is a common fatal tumor of childhood. Natural killer (NK) cells can exert direct cytotoxicity on tumor cells. The killer immunoglobulin-like receptor (KIR) family of NK cell receptors is involved in activation/inhibition of NK cells. In the KIR gene cluster, six of them (3DS1, 2DS1–5) encode receptors triggering activation, while seven of them (3DL1–3, 2DL1–3, 2DL5) encode receptors triggering inhibition. We aimed to assess the distribution of genetic polymorphisms of KIRs on the clinical course of neuroblastoma and provide guidance on potential therapeutic options. Methods: Our study group included 50 neuroblastoma patients and 100 healthy children as controls. Twenty-eight patients were boys, and twenty-two were girls; median age was 36 months. Fourteen patients had stage 1, 2, 3, or 4S disease, and 36 patients had stage 4 disease. Isolated DNA from the peripheral blood was amplified for sequence-specific oligonucleotide probe analysis of 16 KIR genes. The Fisher’s exact test was used to evaluate the variation of KIR gene distribution. Results: All patients had a lower frequency of KIR2DS3 compared to the control group (p = 0.005). Evaluation of individual KIR genes/genotypes in patients with early stages (stage 1, 2, 3, and 4S) versus stage 4 disease revealed that the frequency of KIR2DS3 was increased in early stages (p = 0.023). Inhibitory KIR2DL3 was increased in the patient group compared to controls (p = 0.038). Furthermore, the frequency of KIR2DL3 was higher in stage 4 neuroblastoma patients compared to the patients with early stages (p = 0.023). Conclusion: Our data suggest a role for KIR2DS3 and KIR2DL3 in development of neuroblastoma. Thus, modulation of KIR2SD3 and/or KIR2DL3 expression or function might present a novel therapeutic strategy for neuroblastoma.

研究目的：神经母细胞瘤是儿童常见的致死性肿瘤。自然杀伤（NK，Natural Killer）细胞可对肿瘤细胞产生直接细胞毒性作用。NK细胞表面的杀伤细胞免疫球蛋白样受体（KIR，killer immunoglobulin-like receptor）家族参与调控NK细胞的激活与抑制过程。在KIR基因簇中，6个基因（3DS1、2DS1–5）编码具有激活功能的受体，另有7个基因（3DL1–3、2DL1–3、2DL5）编码具有抑制功能的受体。本研究旨在评估KIR基因多态性的分布特征与神经母细胞瘤临床病程的关联，并为潜在治疗方案提供指导。

研究方法：本研究纳入50例神经母细胞瘤患者与100例健康儿童作为对照。患者中男性28例，女性22例，中位年龄为36个月。其中14例患者处于1、2、3或4S期，36例患者处于4期。从外周血中提取基因组DNA，采用序列特异性寡核苷酸探针分析法对16个KIR基因进行扩增检测。采用Fisher精确检验评估KIR基因分布的差异。

研究结果：与对照组相比，所有受试患者的KIR2DS3基因携带频率均显著降低（p=0.005）。对比早期（1、2、3及4S期）与4期患者的单个KIR基因/基因型分布，发现早期患者的KIR2DS3基因携带频率更高（p=0.023）。患者组的抑制性受体KIR2DL3基因携带频率较对照组升高（p=0.038）。进一步分析显示，4期神经母细胞瘤患者的KIR2DL3基因携带频率高于早期患者（p=0.023）。

研究结论：本研究数据表明KIR2DS3与KIR2DL3在神经母细胞瘤的发生发展中发挥作用。因此，调控KIR2SD3和/或KIR2DL3的表达或功能或可成为神经母细胞瘤的新型治疗策略。