视网膜母细胞瘤中使用房水液体活检进行核酸定量的多中心分析：对临床检测的意义
A Multicenter Analysis of Nucleic Acid Quantification Using Aqueous Humor Liquid Biopsy in Retinoblastoma: Implications for Clinical Testing.
作者信息
Im Deborah H, Pike Sarah, Reid Mark W, Peng Chen-Ching, Sirivolu Shreya, Grossniklaus Hans E, Hubbard G Baker, Skalet Alison H, Bellsmith Kellyn N, Shields Carol L, Lally Sara E, Stacey Andrew W, Reiser Bibiana J, Nagiel Aaron, Shah Rachana, Xu Liya, Berry Jesse L
机构信息
Department of Surgery, Division of Ophthalmology, The Vision Center at Children's Hospital Los Angeles, Los Angeles, California.
USC Roski Eye Institute, Keck School of Medicine of the University of Southern California, Los Angeles, California.
出版信息
Ophthalmol Sci. 2023 Feb 19;3(3):100289. doi: 10.1016/j.xops.2023.100289. eCollection 2023 Sep.
PURPOSE
Retinoblastoma (RB) is most often diagnosed with clinical features and not diagnosed with tumor biopsy. This study describes tumor-derived analyte concentrations from aqueous humor (AH) liquid biopsy and its use in clinical assays.
DESIGN
Case series study.
PARTICIPANTS
Sixty-two RB eyes from 55 children and 14 control eyes from 12 children from 4 medical centers.
METHODS
This study included 128 RB AH samples including: diagnostic (DX) samples, samples from eyes undergoing treatment (TX), samples after completing treatment (END), and during bevacizumab injection for radiation therapy after completing RB treatment (BEV). Fourteen-control AH were analyzed for unprocessed analytes (double-stranded DNA [dsDNA], single-stranded DNA [ssDNA], micro-RNA [miRNA], RNA, and protein) with Qubit fluorescence assays. Double-stranded DNA from 2 RB AH samples underwent low-pass whole-genome sequencing to detect somatic copy number alterations. Logistic regression was used to predict disease burden given analyte concentrations.
MAIN OUTCOME MEASURES
Unprocessed analyte (dsDNA, ssDNA, miRNA, RNA and protein) concentrations.
RESULTS
Results revealed dsDNA, ssDNA, miRNA, and proteins, but not RNA, were quantifiable in most samples (up to 98%) with Qubit fluorescence assays. Median dsDNA concentration was significantly higher in DX (3.08 ng/μl) compared to TX (0.18 ng/μl; < 0.0001) at an order of 17 times greater and 20 times greater than END samples (0.15 ng/μl; = 0.001). Using logistic regression, nucleic acid concentrations were useful in predicting higher versus lower RB disease burden. Retinoblastoma somatic copy number alterations were identified in a TX, but not in a BEV sample, indicating the correlation with RB activity.
CONCLUSIONS
Aqueous humor liquid biopsy in RB is a high-yield source of dsDNA, ssDNA, miRNA, and protein. Diagnostic samples are most useful for RB 1 gene mutational analyses. Genomic analysis may be more informative of tumor activity status than quantification alone and can be performed even with smaller analyte concentrations obtained from TX samples.
FINANCIAL DISCLOSURES
Proprietary or commercial disclosure may be found after the references.
目的
视网膜母细胞瘤（RB）大多通过临床特征进行诊断，而非肿瘤活检。本研究描述了房水（AH）液体活检中肿瘤衍生分析物的浓度及其在临床检测中的应用。
设计
病例系列研究。
参与者
来自4个医疗中心的55名儿童的62只RB眼以及12名儿童的14只对照眼。
方法
本研究纳入了128份RB房水样本，包括：诊断（DX）样本、正在接受治疗（TX）的眼睛样本、完成治疗后（END）的样本以及RB治疗完成后在进行贝伐单抗注射以辅助放疗期间（BEV）的样本。使用Qubit荧光测定法对14份对照房水样本的未处理分析物（双链DNA [dsDNA]、单链DNA [ssDNA]、微小RNA [miRNA]、RNA和蛋白质）进行分析。对2份RB房水样本的双链DNA进行低通量全基因组测序以检测体细胞拷贝数改变。使用逻辑回归根据分析物浓度预测疾病负担。
主要观察指标
未处理分析物（dsDNA、ssDNA、miRNA、RNA和蛋白质）的浓度。
结果
结果显示，在大多数样本（高达98%）中，使用Qubit荧光测定法可对dsDNA、ssDNA、miRNA和蛋白质进行定量，但无法对RNA进行定量。DX样本中的双链DNA中位数浓度（3.08 ng/μl）显著高于TX样本（0.18 ng/μl；<0.0001），约为TX样本的17倍，是END样本（0.15 ng/μl；=0.001）的20倍。使用逻辑回归分析，核酸浓度有助于预测RB疾病负担的高低。在一个TX样本中检测到视网膜母细胞瘤体细胞拷贝数改变，但在BEV样本中未检测到，这表明其与RB活性相关。
结论
RB的房水液体活检是dsDNA、ssDNA、miRNA和蛋白质的高产来源。诊断样本对RB1基因突变分析最为有用。基因组分析可能比单独定量更能提供有关肿瘤活性状态的信息，即使从TX样本中获得的分析物浓度较小也可进行分析。
财务披露
专有或商业披露信息可在参考文献之后找到。
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