研究Homoharringtonine與Apicidin組合於高危險性神經母細胞瘤之治療效果

Abstract

神經母細胞瘤是交感神經系統中最常見的兒童顱外腫瘤。某些遺傳因子改變顯示了預測的疾病發展情況，對於臨床上高風險的過程，例如，在約20％的神經母細胞瘤中發現的，擴增表現的V-myc禽骨髓細胞病變 (V-myc avian myelocytomatosis)致癌基因的神經母細胞瘤衍生同源物(MYCN)。現在而言，晚期的神經母細胞瘤患者仍然難以治癒，目前的治療方案只能將5年生存率提高到50%。我們藉由基因表現差異的分析，在高風險的神經母細胞瘤中找到幾個與MYCN基因擴增表現有關的基因。另外，我們利用美國國家衛生研究院成立的綜合網絡的細胞表現資料庫 (LINCS)中，大量的小分子干擾物與多個細胞株交互作用的資料，我們推斷出的“擾動影響基因”關係。我們提出了一個治療的策略，針對藥物對於基因表現，通過這個方式搜索，我們可以找到疾病基因表現“相反”效應的藥物和藥物組合。基於這些分析結果，我們找到蛋白質合成抑制劑homoharringtonine和組蛋白脫乙酰酶（HDAC）抑制劑apicidin，我們預測這兩個藥物在單獨或組合治療下，可能會抵制擴增MYCN影響的作用。單一給藥時，我們證實了兩種藥物在奈級 (nano)的莫爾濃度 (nM)下，在MYCN基因擴增的神經母細胞瘤細胞系中有細胞毒性。此外，這些藥物的聯合治療也有顯著的協同作用。為了測試生物體內的治療效果，我們通過皮下注射MYCN擴增的神經母細胞瘤細胞系SK-N-BE（2）C，植入異種移植小鼠模型。跟之前在細胞測試的結果相同的，我們發現單獨使用homoharringtonine或apicidin可明顯降低腫瘤體積，並增加神經母細胞瘤小鼠模型的存活率，並且在聯合藥物治療的小鼠中，也有進一步改善。我們將進行更多的功能性研究，來探討這些藥物的精確抗腫瘤機制和神經母細胞瘤的組合。我們的研究結果證明了，以基因表現差異的治療藥物開發方法，並找到對於高危險性神經母細胞瘤患者，有希望的候選藥物。

Neuroblastoma is the most ordinary extracranial childhood tumor of the sympathetic nervous system. Some genetic alterations have been shown to be prognostic of high-risk clinical courses, such as the amplification of V-myc avian myelocytomatosis viral oncogene neuroblastoma derived homolog (MYCN), which was found in approximately 20% of neuroblastomas. The patients with advanced neuroblastoma were still hard to cure, and 5-year survival could only be improved to 50% under the current therapeutic regimen. Here, through a gene expression-based integrative analysis, we uncovered several genes associated with the aggressive biology by MYCN amplification in high-risk neuroblastoma. Additionally, using a myriad of perturbation profiles across multiple cell lines from Library of Integrated Network-based Cellular Signatures (LINCS), we inferred recurrent ‘perturbation−affected gene’ relationships. We proposed an expression-based therapeutic discovery strategy by searching for drugs and drug combinations that can achieve the ‘reversal’ effect given a disease profile. Based on these analytic results, we identified a protein synthesis inhibitor homoharringtonine and a histone deacetylase (HDAC) inhibitor apicidin that, alone or in combination, were predicted to reverse the effect mediated by MYCN amplification. We confirmed the cytotoxicity of the two drugs at nanomolar concentrations in MYCN-amplified neuroblastoma cell lines when given individually. Furthermore, the combination treatment of these drugs was strongly synergistic. To test the therapeutic efficacy in vivo, we used the xenograft mouse model via subcutaneous injection of the MYCN-amplified neuroblastoma cell line SK-N-BE(2)C. Consistently, we found that either homoharringtonine or apicidin alone significantly reduced tumor volume and increased survival in this neuroblastoma mouse model, and such responses were further improved in mice with the combined treatment. We will perform more functional studies to interrogate the exact antineoplastic mechanisms for these drugs and the combination in neuroblastoma. Together, our results demonstrate the efficacy of the expression-based therapeutic discovery and identify promising drug candidates for high-risk neuroblastoma patients.