Osteosarcoma (OS) is an aggressive bone malignancy with a poor prognosis. One putative proto-oncogene in OS is SKP2, encoding a substrate recognition factor of the SCF E3 ubiquitin ligase. We previously demonstrated that SKP2 knockout in murine OS improved survival and delayed tumorigenesis. Here we performed RNA-sequencing (RNA-seq) on tumors from a transgenic OS mouse model with conditional Trp53 and Rb1 knockouts in the osteoblast lineage (“DKO”: Osx1-Cre;Rb1lox/lox;p53lox/lox) and a triple-knockout model with additional Skp2 germline knockout (“TKO”: Osx1-Cre;Rb1lox/lox;p53lox/lox;SKP2-/-), followed by qPCR and immunohistochemistry validation. To investigate the clinical implications of our results, we analyzed a human OS patient cohort (“NCI-TARGET OS”) with RNA-seq and clinical data. We found large differences in gene expression after SKP2 knockout. Surprisingly, we observed increased expression of genes related to immune microenvironment infiltration in TKO tumors, especially the signature genes for macrophages and to a lesser extent, T cells, B cells and vascular cells. We also uncovered a set of relevant transcription factors that may mediate these changes. In OS patient cohorts, high expression of genes upregulated in TKO was correlated with favorable overall survival, which was largely explained by the macrophage gene signatures. This relationship was further supported by our finding that SKP2 expression was negatively correlated with macrophage infiltration in the NCI-TARGET OS and the TCGA Sarcoma cohorts. Overall, our findings indicate that SKP2 may mediate immune exclusion from the OS tumor microenvironment, suggesting that SKP2 modulation in OS may induce anti-tumor immune activation.

中文翻译：

---

Rb1/p53 缺陷型骨肉瘤小鼠模型中 SKP2 敲除可诱导免疫浸润并驱动转录程序，预后良好

骨肉瘤（OS）是一种侵袭性骨恶性肿瘤，预后较差。OS 中一种假定的原癌基因是 SKP2，编码 SCF E3 泛素连接酶的底物识别因子。我们之前证明，小鼠 OS 中的 SKP2 敲除可提高生存率并延缓肿瘤发生。在这里，我们对来自转基因 OS 小鼠模型的肿瘤进行了 RNA 测序 (RNA-seq)，其中条件性敲除成骨细胞谱系中的 Trp53 和 Rb1（“DKO”：Osx1-Cre；Rb1lox/lox；p53lox/lox）和三重基因组。具有额外 Skp2 种系敲除的敲除模型（“TKO”：Osx1-Cre；Rb1lox/lox；p53lox/lox；SKP2-/-），然后进行 qPCR 和免疫组织化学验证。为了研究我们结果的临床意义，我们利用 RNA-seq 和临床数据分析了人类 OS 患者队列（“NCI-TARGET OS”）。我们发现 SKP2 敲除后基因表达存在很大差异。令人惊讶的是，我们观察到 TKO 肿瘤中与免疫微环境浸润相关的基因表达增加，特别是巨噬细胞的特征基因，以及较小程度的 T 细胞、B 细胞和血管细胞。我们还发现了一组可能介导这些变化的相关转录因子。在 OS 患者队列中，TKO 中上调基因的高表达与良好的总体生存率相关，这在很大程度上可以通过巨噬细胞基因特征来解释。我们的发现进一步支持了这种关系，即 SKP2 表达与 NCI-TARGET OS 和 TCGA 肉瘤队列中的巨噬细胞浸润呈负相关。总体而言，我们的研究结果表明 SKP2 可能介导 OS 肿瘤微环境中的免疫排斥，这表明 OS 中的 SKP2 调节可能诱导抗肿瘤免疫激活。