The hereditary spastic paraplegias are an expanding and heterogeneous group of disorders characterized by spasticity in the lower limbs. Plasma biomarkers are needed to guide the genetic testing of spastic paraplegia. Spastic paraplegia type 5 (SPG5) is an autosomal recessive spastic paraplegia due to mutations in CYP7B1, which encodes a cytochrome P450 7α-hydroxylase implicated in cholesterol and bile acids metabolism. We developed a method based on ultra-performance liquid chromatography electrospray tandem mass spectrometry to validate two plasma 25-hydroxycholesterol (25-OHC) and 27-hydroxycholesterol (27-OHC) as diagnostic biomarkers in a cohort of 21 patients with SPG5. For 14 patients, SPG5 was initially suspected on the basis of genetic analysis, and then confirmed by increased plasma 25-OHC, 27-OHC and their ratio to total cholesterol. For seven patients, the diagnosis was initially based on elevated plasma oxysterol levels and confirmed by the identification of two causal CYP7B1 mutations. The receiver operating characteristic curves analysis showed that 25-OHC, 27-OHC and their ratio to total cholesterol discriminated between SPG5 patients and healthy controls with 100% sensitivity and specificity. Taking advantage of the robustness of these plasma oxysterols, we then conducted a phase II therapeutic trial in 12 patients and tested whether candidate molecules (atorvastatin, chenodeoxycholic acid and resveratrol) can lower plasma oxysterols and improve bile acids profile. The trial consisted of a three-period, three-treatment crossover study and the six different sequences of three treatments were randomized. Using a linear mixed effect regression model with a random intercept, we observed that atorvastatin decreased moderately plasma 27-OHC (∼30%, P < 0.001) but did not change 27-OHC to total cholesterol ratio or 25-OHC levels. We also found an abnormal bile acids profile in SPG5 patients, with significantly decreased total serum bile acids associated with a relative decrease of ursodeoxycholic and lithocholic acids compared to deoxycholic acid. Treatment with chenodeoxycholic acid restored bile acids profile in SPG5 patients. Therefore, the combination of atorvastatin and chenodeoxycholic acid may be worth considering for the treatment of SPG5 patients but the neurological benefit of these metabolic interventions remains to be evaluated in phase III therapeutic trials using clinical, imaging and/or electrophysiological outcome measures with sufficient effect sizes. Overall, our study indicates that plasma 25-OHC and 27-OHC are robust diagnostic biomarkers of SPG5 and shall be used as first-line investigations in any patient with unexplained spastic paraplegia.

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血浆氧固醇：用于5型痉挛性截瘫的诊断和治疗的生物标志物

遗传性痉挛性截瘫是一种以下肢痉挛为特征的扩大的异质性疾病。需要血浆生物标志物来指导痉挛性截瘫的基因检测。痉挛性截瘫5型（SPG5）是由于CYP7B1突变引起的常染色体隐性性痉挛性截瘫，它编码涉及胆固醇和胆汁酸代谢的细胞色素P4507α-羟化酶。我们开发了一种基于超高效液相色谱电喷雾串联质谱的方法，以验证两个血浆25-羟基胆固醇（25-OHC）和27-羟基胆固醇（27-OHC）作为21名SPG5患者的诊断生物标志物。对于14例患者，最初是根据基因分析怀疑是SPG5，然后通过血浆25-OHC，27-OHC及其与总胆固醇的比率升高来确认。对于7例患者，诊断最初是基于血浆中的氧固醇水平升高，并通过鉴定两个因果CYP7B1来确定突变。接受者的工作特征曲线分析表明，在SPG5患者和健康对照组中，25-OHC，27-OHC及其与总胆固醇的比例具有100％的敏感性和特异性。利用这些血浆氧固醇的坚固性，我们随后在12位患者中进行了II期治疗试验，并测试了候选分子（阿托伐他汀，鹅去氧胆酸和白藜芦醇）是否可以降低血浆氧固醇和改善胆汁酸谱。该试验由三期，三疗程交叉研究组成，对三种疗程的六个不同序列进行了随机分组。使用具有随机截距的线性混合效应回归模型，我们观察到阿托伐他汀可适度降低血浆27-OHC（〜30％，P<0.001），但并未改变27-OHC与总胆固醇之比或25-OHC的水平。我们还发现SPG5患者的胆汁酸分布异常，与脱氧胆酸相比，总血清胆汁酸显着降低，而熊去氧胆酸和石胆酸相对减少。鹅去氧胆酸治疗可恢复SPG5患者的胆汁酸谱。因此，阿托伐他汀和鹅去氧胆酸的组合可能值得SPG5患者的治疗，但是这些代谢干预措施的神经学益处尚需通过临床，影像学和/或电生理结果措施在III期治疗试验中进行评估，并且要有足够的疗效。全面的，