以人體循環中的長鏈非編碼核糖核酸作為巴金森氏症的風險評估和疾病進展的生物標記物

Abstract

巴金森氏症為最常見的神經退化性疾病之一，除了動作遲緩障礙外，也會有非運動症狀，其中認知功能減退，甚至演變為巴金森失智症，對病患預後影響甚鉅。在現今人口老化的社會，其照護對社會成本產生重大的負擔。因此，找出具有反映疾病病程和預測巴金森病患是否會合併失智症的生物標記物，以及釐清此病症之致病機轉，實為重要的臨床課題。 長鏈非編碼核糖核酸(long non-coding RNA,以下稱lncRNA)為長度超過200個鹼基的非編碼核糖核酸，在發展中的腦部以及成人的腦部都被大量的表現。過去的研究顯示，lncRNA具有調控基因的功能。在巴金森氏症患者的腦脊髓液中的lncRNA表現量和健康對照組不同，另一研究發現巴金森氏症患者在接受深腦電刺激術之後lncRNA的表現型態變化會出現在腦部的杏仁核和基底核(巴金森氏症病變腦區)中，暗示了lncRNA在血液中的表現可能可以反映出巴金森氏症患者的嚴重程度。但礙於腦脊髓液的取得為一侵襲性高的檢查，加上lncRNA可透過血腦屏障至周邊血液中，因此我們假設血漿中相關的lncRNA片段將可以協助臨床醫師評估病患運動症狀和認知功能退化，作為非侵襲性周邊血液指標。 我們的研究的第一部分目標為尋找與巴金森氏症風險相關之lncRNA。我們收納了19位早發型(發病年齡小於50歲)巴金森氏症患者、20位典型(發病年齡晚於50歲)巴金森氏症患者以及40位年齡與性別對照之健康受試者。我們利用次世代RNA定序法，比較巴金森氏症患者和健康對照組受試者的血漿中lncRNA的表現形態差異，以找出與疾病風險相關之lncRNA。結果顯示血漿中之lncRNA的表現模式可用以分辨健康受試者和巴金森氏症患者，其準確率在早發型巴金森氏症患者和典型巴金森氏症患者分別可達68.29%和80.00%。因為lncRNA往往是透過順式調控來達到附基因調控，故我們分析了在健康受試者和巴金森氏症患者身上表達最顯著不同之20個lncRNA附近之mRNA的表現，發現在早發型巴金森氏症患者和典型巴金森氏症患者中，分別有25%和35%的lncRNA附近之mRNA呈現順性的變化。 我們研究的第二部分為藉由逐年臨床追蹤病患動作與評估認知功能，希望能找出與巴金森症認知功能退化相關的疾病進展lncRNA指標。67位典型巴金森氏症患者在經過約一年之追蹤後，有13位(19.4%)患者產生認知功能退化(簡短智能測驗分數MMSE下降大於等於兩分)。經由比較有產生認知功能退化和未產生認知功能退化的患者的lncRNA表現，我們發現有6個lncRNA的表現在此兩群患者間有顯著的不同 (lnc-ZNF582-3, lnc-LYPLAL1-5, lnc-RGS21-1, lnc-TRIM69-2, lnc-SLC40A1-3, lnc-C14orf135-3)。存活分析進一步發現其中兩個lncRNA(lnc-RGS21-1, lnc-TRIM69-2)可用以預測巴金森氏症患者產生認知功能退化的風險。 綜上所述，我們的結果顯示血漿中的lncRNA可以部份反映巴金森氏症疾病罹病風險與疾病進程的退化，尤其部分標的lncRNA可以做為預測巴金森症患者認知功能減退的指標。我們的實驗結果仍需將來的動物或是細胞功能性實驗以驗證這些lncRNA在巴金森症致病機轉之角色。

Parkinson’s disease (PD) is the second most common neurodegenerative disorder, characterized by symptoms of resting tremor, rigidity, bradykinesia and postural instability. The current mainstay treatment is supplement with dopaminergic medications, and there is no disease-modifying therapy available for PD yet. Therefore, there is an unmet need to develop biomarkers that could predict the risk and reflect disease progression in patients with PD. Many of the transcription output of human genome is composed with non-coding RNAs. Long non-coding RNA (lncRNA) is defined as non-coding RNA with molecules greater than 200 nucleotides, and are highly expressed in the adult and developing brain. Many lncRNA-related dysfunction has been found to play critical roles in neurodegenerative disorders. However, to our knowledge, there are no studies design to analyze the significance of plasma levels of lncRNAs in patients with PD till date. In this study, we aim to determine the circulating lncRNA expression profiles in patients with PD, and to test the hypothesis that the lncRNA expression signature could be used to predict the progression of non-motor symptoms in patients with PD, as well as to explore pathogenic mechanisms via which lncRNAs contributing to PD. We first enrolled 19 young-onset PD patients (onset age at less than 50 years, YPD), 20 late-onset PD patients (onset age at order than 50 years, OPD) and 41 age/gender matched control subjects. The plasma levels of lncRNAs were examined to determine the specific lncRNA expression signature for PD. LncRNAs could be identified from plasma of young onset and late onset PD patients with an accuracy rate of 68.29% and 80.00%, respectively. For lncRNAs could exert its epigenetic regulation through cis-regulation, we also analysis expression pattern of its nearby mRNA for the twenty lncRNAs with the most significant different expression pattern between PD patients and controls. 25% and 35% of lncRNAs with significant differential expression pattern in OPD group and YPD group, respectively, showed cis-change with its nearby mRNA. And we followed Mini-Mental State Examination (MMSE) in an OPD patient cohort for around 1 year and to determine the specific lncRNAs expression pattern that could be use as prognostic biomarkers in cognition performance in PD patients. Among these 67 OPD patients, 13(19.4%) with MMSE deterioration ≧2 points after 1 year follow up. While comparing the lncRNA expression pattern between PD patients with and without dementia, we found that the expression of 6 lncRNAs (lnc-ZNF582-3, lnc-LYPLAL1-5, lnc-RGS21-1, lnc-TRIM69-2, lnc-SLC40A1-3, lnc-C14orf135-3) were significantly different between these two groups. The result of survival analysis showed that 2 of these afore mentioned lncRNAs (lnc-RGS21-1, lnc-TRIM69-2) could predict the risk of dementia in PD patients. In conclusion, our results demonstrated the feasibility of specific plasma lncRNA as a surrogate biomarker for risk and cognitive progression of PD. Further functional studies are warranted to validate the role of these lncRNAs in the pathogenesis of PD.