(18)F and (18)FDG PET imaging of osteosarcoma to non-invasively monitor in situ changes in cellular proliferation and bone differentiation upon MYC inactivation

Osteosarcoma is one of the most common pediatric cancers. Accurate imaging of osteosarcoma is important for proper clinical staging of the disease and monitoring of the tumor's response to therapy. The MYC oncogene has been commonly implicated in the pathogenesis of human osteosarcoma. Previously, we have described a conditional transgenic mouse model of MYC-induced osteosarcoma. These tumors are highly invasive and are frequently associated with pulmonary metastases. In our model, upon MYC inactivation osteosarcomas lose their neoplastic properties, undergo proliferative arrest and differentiate into mature bone. We reasoned that we could use our model system to develop noninvasive imaging modalities to interrogate the consequences of MYC inactivation on tumor cell biology in situ. We performed positron emission tomography (PET) combining the use of both (18)F-fluorodeoxyglucose ((18)FDG) and (18)F-flouride ((18)F) to detect metabolic activity and bone mineralization/remodeling. We found that upon MYC inactivation, tumors exhibited a slight reduction in uptake of (18)FDG and a significant increase in the uptake of (18)F along with associated histological changes. Thus, these cells have apparently lost their neoplastic properties based upon both examination of their histology and biologic activity. However, these tumors continue to accumulate (18)FDG at levels significantly elevated compared to normal bone. Therefore, PET can be used to distinguish normal bone cells from tumors that have undergone differentiation upon oncogene inactivation. In addition, we found that (18)F is a highly sensitive tracer for detection of pulmonary metastasis. Collectively, we conclude that combined modality PET/CT imaging incorporating both (18)FDG and (18)F is a highly sensitive means to non-invasively measure osteosarcoma growth and the therapeutic response, as well as to detect tumor cells that have undergone differentiation upon oncogene inactivation.     

Amplification of PVT1 contributes to the pathophysiology of ovarian and breast cancer.

PURPOSE: This study was designed to elucidate the role of amplification at 8q24 in the pathophysiology of ovarian and breast cancer because increased copy number at this locus is one of the most frequent genomic abnormalities in these cancers. EXPERIMENTAL DESIGN: To accomplish this, we assessed the association of amplification at 8q24 with outcome in ovarian cancers using fluorescence in situ hybridization to tissue microarrays and measured responses of ovarian and breast cancer cell lines to specific small interfering RNAs against the oncogene MYC and a putative noncoding RNA, PVT1, both of which map to 8q24. RESULTS: Amplification of 8q24 was associated with significantly reduced survival duration. In addition, small interfering RNA-mediated reduction in either PVT1 or MYC expression inhibited proliferation in breast and ovarian cancer cell lines in which they were both amplified and overexpressed but not in lines in which they were not amplified/overexpressed. Inhibition of PVT1 expression also induced a strong apoptotic response in cell lines in which it was overexpressed but not in lines in which it was not amplified/overexpressed. Inhibition of MYC, on the other hand, did not induce an apoptotic response in cell lines in which MYC was amplified and overexpressed. CONCLUSIONS: These results suggest that MYC and PVT1 contribute independently to ovarian and breast pathogenesis when overexpressed because of genomic abnormalities. They also suggest that PVT1-mediated inhibition of apoptosis may explain why amplification of 8q24 is associated with reduced survival duration in patients treated with agents that act through apoptotic mechanisms.     

Interaction between MYC and MCL1 in the genesis and outcome of non-small-cell lung cancer

MYC exerts both positive and negative functions in cancer cells, such that its procancerous effects are unmasked only after its anticancer effects are blocked. Here we used multiple mouse models of lung adenocarcinoma to identify genetic events that can cooperate with MYC activation to promote the genesis of non-small-cell lung cancer (NSCLC), the most common form of lung cancer in humans. MYC overexpression targeted to pulmonary alveolar cells was sufficient to induce lung adenomas and carcinomas. Tumorigenesis was assisted by either spontaneous mutations in Kras or experimental introduction of activated RAS, but investigations revealed that additional events were required to circumvent apoptosis, one of the most significant negative functions exerted by MYC. We determined that overexpression of the antiapoptotic protein MCL1 was sufficient to circumvent apoptosis in this setting. Previous clinical studies have indicated that prognosis of human NSCLC is not associated with MCL1, despite its overexpression in many NSCLCs. In reexamining the prognostic value in this setting, we found that MCL1 overexpression does correlate with poor patient survival, but only when accompanied by MYC overexpression. Our findings therefore produce a convergence of mouse and human results that explain how MCL1 can block an important negative consequence of MYC overexpression in both experimental models and clinical cases of NSCLC.     

肺癌组织中bax、bcl-xL、bcl-xS基因表达的研究

目的探讨bax、bcl—xL、bcl—xS3个基因在非小细胞肺癌组织及癌旁组织中的表达。方法以凝胶上rRNA条带的亮度为依据对后续分析用的起始RNA浓度进行调整；以组成型表达的beta—actin基因为外对照，对两类组织来源的起始RNA量进行监测；基因克隆及序列分析验证所用引物的可靠性；随后采用逆转录-聚合酶链式反应(RT—PCR)的方法对各基因在两类组织中的表达谱进行对比分析。结果在两类组织来源的起始RNA量基本一致的前提下，3个基因在成对组织中的表达谱存在明显差异，bax和bcl—xS在肺癌组织及癌旁组织中均有表达，bcl—xL基因在肺癌组织中的表达量较高。结论肺癌组织中也存在．bax和bcl—xS等促凋亡基因的转录。     

Conditionally MYC: insights from novel transgenic models

MYC was one of the first oncogenes identified to be associated with chromosomal aberrations and one of the most common oncogenes involved in the pathogenesis of cancer. However, until recently it was not clear if MYC would be a good target for the treatment of cancer. New conditional transgenic models have been used to demonstrate that even the brief inactivation of MYC can reverse tumorigenesis. Here we review results from recent experimental model systems, which demonstrate that the inactivation of MYC may be a specific and effective treatment for many types of cancer.     

MYC overexpression and poor prognosis in sporadic breast cancer with BRCA1 deficiency

Breast cancer is a complex disease; the molecular mechanisms involved in sporadic breast carcinogenesis remain to be elucidated. The present study aimed to explore the deficiency of breast cancer susceptibility gene 1 (BRCA1), including protein loss expression, promoter hypermethylation and gene copy deletion, its correlationship with other tumor markers expression (TP53, MYC, etc.), and clinical significance in sporadic breast cancer. BRCA1 protein expression was negative in 226 of 374 (60.4 %) cases of this study. Cases negative for BRCA1 protein were more often with pathological tumor–node–metastasis stage III, positive for lymph node metastasis and MYC overexpression than BRCA1-positive tumors. BRCA1 hypermethylation was detected in 16.4 % (31 of 189) breast cancers, which correlated with BRCA1 negative, ER negative, MYC overexpression, and triple-negative phenotype. In addition, the percentage of cells with BRCA1 gene copy deletion was significantly increased in BRCA1-methylated tumors. Kaplan–Meier survival analysis showed that patients with BRCA1-negative expression showed a worse overall survival (OS) than those with BRCA1-positive expression, and patients with BRCA1-methylated tumors had a significantly worse disease-free survival than did patients with unmethylated tumors. Furthermore, BRCA1 hypermethylation showed an inverse association with OS in LN-positive or p53-negative subgroup patients. Importantly, uni- and multivariate Cox regression analyses revealed that BRCA1 was an independent prognostic indicator of OS in sporadic breast cancer. Thus, we found MYC overexpression and poor prognosis in sporadic breast cancer with BRCA1 deficiency. The targeting of BRCA1 deficiency in combination with MYC–pathways inhibitors may provide a promising strategy for sporadic breast cancer care, the triple-negative subtype in particular.     

四种乳腺癌相关基因联合检测对乳腺癌诊断的临床应用研究

目的:探讨核内原癌基因(nuclear oncogene,MYC)、B细胞淋巴瘤/白血病-2基因(B cell lymphoma/leukemia 2 gene,Bcl-2)、细胞周期蛋白D1（Ccnd1）和细胞角蛋白19(cytokeratin 19,CK19)基因表达水平在乳腺癌诊断和治疗监测中的应用。方法:建立荧光定量聚合酶链反应(FQ-PCR)法,并以GAPDH为内对照测定34名健康女性体检者、55例良性乳腺疾病患者和91例乳腺癌患者外周血中MYC,Ccnd1,Bcl-2和CK19的表达量。结果:MYC,Ccnd1,Bcl-2和CK19表达水平与GAPDH的比值在乳腺癌组显著高于健康女性体检者和良性乳腺疾病患者（P<0.05）。它们在正常对照组和良性乳腺疾病组间差异无统计学意义（P>0.05）。四组GAPDH差异无统计学意义（P>0.05）。MYC,Ccnd1,Bcl-2与CK19联合检测的灵敏度高于单个基因的检测。结论:FQ-PCR技术可以快速定量检测MYC,Ccnd1,Bcl-2和CK19 mRNA,四者联合检测可有效提高对乳腺癌诊断的灵敏度。     

MYC is amplified in BRCA1-associated breast cancers.

PURPOSE: Germ-line mutations in the BRCA1 tumor suppressor gene predispose to early onset breast cancers with a distinct phenotype characterized by high tumor grade, aneuploidy, high proliferation rate, and estrogen receptor-negativity. The molecular mechanisms and cooperative oncogenes contributing to multistep tumor progression in cells lacking BRCA1 are not well defined. To examine whether C-MYC (MYC), a transforming oncogene associated with genetic instability, contributes to multistep tumor progression in BRCA1-associated breast cancer, we have analyzed tumors from women with hereditary BRCA1-mutated and sporadic breast cancers. EXPERIMENTAL DESIGN: We performed fluorescence in situ hybridization using a MYC:CEP8 assay on formalin-fixed paraffin-embedded tumor tissues from 40 women with known deleterious germ-line BRCA1 mutations and 62 sporadic cases, including 20 cases with hypermethylation of the BRCA1 gene promoter. RESULTS: We observed a MYC:CEP8 amplification ratio >/=2 in 21 of 40 (53%) BRCA1-mutated tumors compared with 14 of 62 (23%) sporadic tumors (P = 0.003). Of the 14 sporadic cases with MYC amplification, 8 (57%) were BRCA1-methylated. In total, MYC amplification was found in a significantly higher proportion of tumors with BRCA1 dysfunction (29 of 60, 48% versus 6 of 42, 14%; P = 0.0003). In a multivariable regression model controlling for age, tumor size, and estrogen receptor status, BRCA1-mutated tumors demonstrated significantly greater mean MYC:CEP8 ratio than sporadic tumors (P = 0.02). CONCLUSIONS: Our data indicate that MYC oncogene amplification contributes to tumor progression in BRCA1-associated breast cancers. Thus, we conclude that the aggressive histopathological features of BRCA1-associated tumors are in part due to dysregulated MYC activity.     

The long noncoding RNA LINC01207 promotes proliferation of lung adenocarcinoma

Lung adenocarcinoma (LAD) and lung squamous cell cancer (LSCC) are two most common histological types of lung cancer, while they differ in many aspects. Recent evidence shows that long non-coding RNAs (lncRNAs) play an important role in the process of cancer initiation and progression. Thus, characterization of LAD and LSCC associated lncRNAs may help understand the difference between LAD and LSCC. Here, we analyzed three sets of RNA-seq data, including LAD RNA-seq data from TCGA project. We identified a novel lncRNA, long intergenic non-protein coding RNA 1207 (LINC01207) which was significantly up-regulated in LAD tissues compared with paired non-tumor tissues (5.78 fold increase, P<0.05), while there was no significant differences between LSCC tissues and adjacent non-tumor tissues. The expression level of LINC01207 was associated with TNM stage of LAD patients, and higher LINC01207 level indicated advanced TNM stage (P<0.05) and shorter survival (HR=2.53, P<0.05). By small interfering RNA (siRNA) mediated knockdown of LINC01207, we determined the biological function of LINC01207 in A549 cell line. After knockdown of LINC01207, cell proliferation ability was inhibited. Further analysis showed that after silence of LINC01207, the percentage of apoptotic cells significantly increased. By RNA immunoprecipitation and Chromatin immunoprecipitation assay, we demonstrated that LINC01207 could bind with EZH2 and mediated trimethylation of histone 3 lysine 27 at the promoter region of Bad, an important pro-apoptotic gene. Finally, we developed xenograft tumor models in nude mice and xenograft tumors derived from A549 cells transfected with siRNA-LINC01207 had significantly lower tumor weight and smaller tumor volume. In summary, the novel lncRNA, LINC01207 is specifically up-regulated in LAD but not in LSCC; and LINC01207 could promote LAD cell growth both in vivo and in vitro.     

Single-nucleotide polymorphism rs4142441 and MYC co-modulated long non-coding RNA OSER1-AS1 suppresses non-small cell lung cancer by sequestering ELAVL1

Single-nucleotide polymorphisms (SNP) and long non-coding RNAs (lncRNAs) have been involved in the process of lung cancer. Following clues given by lung cancer risk-associated SNP, we aimed to find novel functional lncRNAs as candidate targets in lung cancer. We identified a lncRNA Oxidative Stress Responsive Serine Rich 1 Antisense RNA 1 (OSER1-AS1) through a lung cancer risk-associated SNP rs4142441. OSER1-AS1 was down-regulated in tumor tissue and its low expression was significantly associated with poor overall survival among non-smokers in non-small cell lung cancer (NSCLC) patients. Gain- and loss-of-function studies showed that OSER1-AS1 acted as a tumor suppressor by inhibiting lung cancer cell growth, migration and invasion in vitro. Xenograft tumor assays and a metastasis mouse model confirmed that OSER1-AS1 suppressed tumor growth and metastasis in vivo. The promoter of OSER1-AS1 was repressed by MYC, and the 3′-end of OSER1-AS1 was competitively targeted by microRNA hsa-miR-17-5p and RNA-binding protein ELAVL1. Our results indicated that OSER1-AS1 exerted tumor-suppressive functions by acting as an ELAVL1 decoy to keep it away from its target mRNAs. Our findings characterized OSER1-AS1 as a new tumor-suppressive lncRNA in NSCLC, suggesting that OSER1-AS1 may be suitable as a potential biomarker for prognosis, and a potential target for treatment.     

MYC和MUC1 mRNA联合检测在乳腺癌诊断中的应用

目的:建立逆转录聚合酶链反应(RT-PCR)检测乳腺癌核内原癌基因(nuclear oncogene,MYC)和黏蛋白1(Mucin1,MUC1)基因表达水平,探讨其联合检测在乳腺癌诊断和治疗监测中的应用。方法:建立RT-PCR反应,并以β2-微球蛋白为内对照测定30例健康女性体检者、52例良性乳腺肿瘤和103例乳腺癌外周血中MYC和MUC1的表达量。结果:外周血中MYC和MUC1在乳腺癌组表达率显著高于良性乳腺疾病组及健康体检组,而良性乳腺疾病组和健康体检组的表达率无差异。MUC1和MYC联合检测的特异性显著高于单一基因。结论:RT-PCR联合检测外周血MUC1和MYC基因表达,可提高对乳腺癌诊断的特异性,具有标本来源容易、操作简便和价格便宜等优点,可有效辅助诊断乳腺癌。