Laboratory of Medical Oncology, Research Institutes of Medical Science, The Catholic University of Korea, Seoul, Republic of Korea.
Int J Oncol. 2013 Aug;43(2):383-93. doi: 10.3892/ijo.2013.1953. Epub 2013 May 24.
Patients with HPV-positive oropharyngeal cancer show better tumor response to radiation or chemotherapy than patients with HPV-negative cancer. HPV oncoprotein E6 binds and degrades a typically wild-type p53 protein product. However, HPV16 infection and p53 mutation infrequently coexist in a subset of HNSCCs. The purpose of this study was to investigate the mechanisms through which tumor biology and molecular genetic mechanisms change when two HPV-negative, p53-mutated oropharyngeal cell lines (YD8, non-disruptive p53 mutation; YD10B, disruptive p53 mutation) derived from patients with a history of heavy smoking are transfected with HPV E6 and E7 oncogenes in vitro. Transfection with HPV E6 and E7 oncogenes in YD8, reduced the abundance of proteins encoded by tumor suppressor genes, such as p-p53 and p-Rb. Cell proliferative activity was increased in the cells transfected with E6E7 compared to cells transfected with vector alone (P=0.09), whereas the invasiveness of E6E7-transfected cells was significantly reduced (P=0.02). cDNA microarray of the transfected cells with E6E7 showed significant changes in mRNA expression in several signaling pathways, including focal adhesion, JAK-STAT signaling pathway, cell cycle and p53 signaling pathway. Regarding the qPCR array for the p53 signaling pathway, the mRNA expression of STAT1 was remarkably upregulated by 6.47-fold (P<0.05); in contrast, IGF-1R was significantly downregulated by 2.40-fold in the YD8-vector compared toYD8-E6E7 (P<0.01). Finally, data collected from these two array experiments enabled us to select two genes, STAT1 and IGF-1R, for further study. In immunohistochemical study, nuclear STAT1 expression was slightly higher in HPV-positive compared to HPV-negative oropharyngeal tumors (P=0.18); however, cytoplasmic STAT1 was significantly lower in HPV-positive cases (P=0.03). IGF-1R expression levels were remarkably lower in HPV-positive compared to HPV-negative cases (P=0.01). Our data suggest that upregulated STAT1 and interferon signals by HPV16 E6 and E7 genes may play a major role in the relatively favorable prognosis for HPV-positive oropharyngeal squamous cell carcinoma cases with non-disruptive p53 mutations.
HPV 阳性口咽癌患者对放化疗的肿瘤反应优于 HPV 阴性癌症患者。HPV 癌蛋白 E6 结合并降解通常为野生型 p53 蛋白产物。然而，HPV16 感染和 p53 突变在一组 HNSCC 中很少共存。本研究的目的是研究当两个 HPV 阴性、p53 突变的口咽癌细胞系（YD8，非破坏性 p53 突变；YD10B，破坏性 p53 突变）源自有吸烟史的患者时，体外转染 HPV E6 和 E7 癌基因时肿瘤生物学和分子遗传机制如何发生变化。在 YD8 中转染 HPV E6 和 E7 癌基因降低了肿瘤抑制基因如 p-p53 和 p-Rb 的编码蛋白的丰度。与单独转染载体的细胞相比，转染 E6E7 的细胞的细胞增殖活性增加（P=0.09），而 E6E7 转染细胞的侵袭性显著降低（P=0.02）。用 E6E7 转染的细胞的 cDNA 微阵列显示在几个信号通路中，包括粘着斑、JAK-STAT 信号通路、细胞周期和 p53 信号通路，mRNA 表达发生显著变化。关于 p53 信号通路的 qPCR 阵列，STAT1 的 mRNA 表达显著上调 6.47 倍（P<0.05）；相比之下，在 YD8-载体中 IGF-1R 的表达显著下调 2.40 倍，而在 YD8-E6E7 中（P<0.01）。最后，从这两个阵列实验中收集的数据使我们能够选择两个基因，STAT1 和 IGF-1R，进行进一步研究。在免疫组织化学研究中，HPV 阳性口咽肿瘤的核 STAT1 表达略高于 HPV 阴性肿瘤（P=0.18）；然而，HPV 阳性病例的细胞质 STAT1 显著降低（P=0.03）。与 HPV 阴性病例相比，IGF-1R 的表达水平明显降低（P=0.01）。我们的数据表明，HPV16 E6 和 E7 基因上调的 STAT1 和干扰素信号可能在非破坏性 p53 突变的 HPV 阳性口咽鳞状细胞癌病例中发挥相对有利的预后作用。
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