长链非编码RNA OMRUL调控ABCB1介导骨肉瘤多药耐药发生、发展的分子机制研究

Osteosarcoma is the most common primary malignant bone tumor in adolescent. Chemotherapy is its most important adjuvant therapy, however, the emergence of multi-drug resistance has greatly impeded the treatment of osteosarcoma. Long non-coding RNA (lncRNA) can be widely involved in the regulation of gene expression, which could mediate tumor development, drug resistance and other procedures. Through microarray and PCR validation, our previous study found that lncRNA OMRUL highly upregulated in three doxorubicin-resistant osteosarcoma cell lines and simultaneously we detected lncRNA OMRUL in 120 cases of human osteosarcoma samples, which showed the expression of lncRNA OMRUL in chemotherapy resistance group was significantly higher than chemotherapy sensitive group. By bioinformatic analysis, we initially confirmed that lncRNA OMRUL tightly correlated with classical muti-drug resistance related ABCB1 gene. Then by use of the genetic intervention technique, we proved that the expression of lncRNA OMRUL and ABCB1 was positively relevant and closely associated with chemosensitivity of osteosarcoma cells. By use of RIP and mass spectrometry techniques, we expect to further validate the direct interaction between lncRNA OMRUL and ABCB1 and find possible intermediate protein, miRNA etc. to consummate the mechanisms of osteosarcoma durg-resistance in terms of lncRNA, which may provide a new potential therapy target for multidrug-resistant osteosarcoma.

骨肉瘤是青少年最常见的原发性恶性骨肿瘤。化疗是其最重要的辅助疗法，然而多药耐药的出现极大地阻碍了其治疗。长链非编码RNA（lncRNA）可广泛参与基因表达的调控，从而介导肿瘤发展、耐药等过程。我们前期通过芯片筛选及PCR验证发现在三种阿霉素抵抗的骨肉瘤细胞系中高度表达上调的lncRNA OMRUL，同时检测120例人骨肉瘤标本，结果显示lncRNA OMRUL在化疗抵抗组的表达明显高于化疗敏感组。通过生物信息学分析，初步确认lncRNA OMRUL与经典耐药基因ABCB1高度相关。然后通过基因干预技术，证明了lncRNA OMRUL与 ABCB1的表达正相关且与细胞化疗敏感性密切相关。我们预计采用RIP及质谱分析等技术，进一步验证lncRNA OMRUL 与ABCB1的直接相互作用关系并寻找可能的中间蛋白、miRNA等，从lncRNA的角度完善耐药机制，为多药耐药骨肉瘤的治疗提供新的靶点。

骨肉瘤是青少年最常见的原发性恶性骨肿瘤。化疗是其最重要的辅助疗法，然而多药耐药的出现极大地阻碍了其治疗。长链非编码RNA（lncRNA）是近年来备受关注的一类调控分子，它可在转录前、转录和转录后水平广泛参与基因表达的调控，从而介导肿瘤发展、耐药等过程。我们前期通过芯片筛选及PCR验证发现在三种阿霉素抵抗的骨肉瘤细胞系中高度表达上调的lncRNA ODRUL，其在骨肉瘤组织标本中同样高表达，与患者的不良预后密切相关；功能研究发现lncRNA ODRUL可以体内、外促进骨肉瘤细胞对于阿霉素的抵抗性、促进细胞增殖、克隆形成、侵袭和迁移以及裸鼠成瘤能力；分子机制研究发现主要定位于细胞质中的lncRNA ODRUL作为一种反义lncRNA，可以通过与其共轭基因FOXC2的mRNA所形成的局部RNA-RNA双链结构保护FOXC2的mRNA免受RNA酶的降解，从而促进转录因子FOXC2的高表达，进而促进ABCB1的高表达，导致骨肉瘤细胞多药耐药的发生、发展，同时研究发现lncRNA ODRUL可以通过内源竞争性RNA（ceRNA）调控机制，吸附miR-3182，从而解除其对基质金属蛋白酶-2（MMP-2）的表达抑制，促进MMP-2的高表达，进而促进骨肉瘤的侵袭和迁移。总之，我们的研究结果表明，高表达的lncRNA ODRUL与骨肉瘤的多药耐药以及侵袭、迁移的发生、发展密切相关，可能是骨肉瘤诊断、预后判断的潜在生物标志物以及治疗新靶点。