Developing high throughput measurement of thiopurine in DNA by mass spectrometry

通过质谱法开发 DNA 中硫嘌呤的高通量测量

基本信息

批准号：
9909135
负责人：
W. Andy Tao
金额：
$ 30万
依托单位：
TYMORA ANALYTICAL OPERATIONS, LLC
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-02 至 2023-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9909135
关键词：
6-Mercaptopurine Acute Lymphocytic Leukemia Adopted Adverse reactions Antimetabolites Autoimmune Diseases Biological Cell Death Chemicals Child Childhood Leukemia Clinical DNA DNA biosynthesis Data Detection Disease Dose Dose-Limiting Erythrocytes Funding Genetic Genetic Variation Genotype Hematologic Neoplasms Hematopoietic Inflammatory Bowel Diseases Inherited Knockout Mice Lead Leukocytes Mass Spectrum Analysis Measurement Measures Methods Mismatch Repair Modernization Monitor Morbidity - disease rate Non-Malignant Nucleotides Patients Pharmaceutical Preparations Pharmacogenetics Pharmacology Phase Phenotype Positioning Attribute Reaction Reagent Regimen Reporting Reproducibility Research Personnel Risk Sampling Sensitivity and Specificity Small Business Technology Transfer Research Solid Specificity Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization Stable Isotope Labeling Sulfhydryl Compounds TPMT gene Techniques Testing Therapeutic Index Thioguanine Thiopurine Methyltransferase Deficiency Toxic effect Treatment Efficacy Treatment Failure Validation Variant White Blood Cell Count procedure Yang analog analytical tool antitumor drug base cohort cytotoxicity dose individualization improved individual patient individualized medicine ionization leukemia/lymphoma mortality mouse model novel nucleobase outcome prediction patient population personalized medicine precision medicine prototype response thiopurine treatment response

项目摘要

PROJECT SUMMARY Thiopurines such as mercaptopurine are S-substituted antimetabolites used as functional analogs to natural nucleobase precursors. They are highly effective against hematologic malignancies (leukemia and lymphoma) and autoimmune diseases (inflammatory bowel diseases [IBD]), with an estimated patient population >350,000 in the US. However, the cytotoxicity of thiopurines depends on their conversion into 6-thioguanine (TG) nucleotides (TGN), which are incorporated into DNA, causing cell death by post-replicative DNA mismatch repair. They have narrow therapeutic indexes with dose-limiting hematopoietic toxicity whereas low-responders are undertreated with standard dosing. Therefore, there is enormous clinical benefit from preemptively identifying patients at risk of thiopurine toxicity and individualizing therapy to mitigate it. We propose here a high throughput method based on matrix-assisted laser desorption ionization (MALDI) mass spectrometry (MS) for quantifying the pharmacological endpoint of thiopurines, TG incorporated in chromosomal DNA (DNA-TG) of white blood cells, and examine its specificity and accuracy in standard and clinical samples. To test this strategy, we propose three specific aims: Aim #1. Chemical derivatization, isolation and detection of 6-thioguanine (6-TG) by MALDI-MS. Aim #2. Examination of linear range, sensitivity, specificity and reproducibility of the method. Aim #3. Validation of high throughput 6-TG detection with biological samples. With the aim to individualize thiopurine therapy through reducing adverse reactions and obtaining optimum therapeutic efficacy for patients on thiopurine treatment, the high throughput technique facilitates continuous monitoring of thiopurine in DNA before and during the therapy. This highly translational project will likely have high impact on a large number of patients who are under chemo-treatment or thiopurine therapy for non-malignant conditions (e.g., IBD).

项目摘要硫嘌呤（如胃嘌呤）是S取代的抗代谢物，用作功能类似物天然核碱酶前体。它们对血液学恶性肿瘤非常有效（白血病和淋巴瘤）和自身免疫性疾病（炎症性肠病[IBD]），有一个在美国，估计患者人群> 350,000。但是，硫嘌呤的细胞毒性取决于在转化为6-硫代氨酸（TG）核苷酸（TGN）时，该核苷酸（TGN）掺入DNA中，通过复制后DNA不匹配修复引起细胞死亡。他们的治疗指数狭窄用限制剂量的造血毒性，而低响应者则未使用标准给药。因此，先发制人的患者有巨大的临床益处硫嘌呤毒性和个性化治疗以减轻它。我们在这里提出了高吞吐量基于基质辅助激光解吸电离（MALDI）质谱（MS）的方法量化硫嘌呤的药理终点，TG掺入染色体DNA中（DNA-TG）白细胞，并检查其在标准和临床上的特异性和准确性样品。为了测试此策略，我们提出了三个具体目标：目标＃1。化学衍生化， MALDI-MS分离和检测6-硫代氨酸（6-TG）。目标＃2。线性范围的检查，该方法的灵敏度，特异性和可重复性。目标＃3。验证高吞吐量6-TG 用生物样品检测。旨在通过减少硫嘌呤疗法个性化不良反应并获得对硫嘌呤治疗患者的最佳治疗功效，高通量技术促进了前后DNA中硫嘌呤的连续监测在治疗期间。这个高度翻译的项目可能会对大量接受化学治疗或硫嘌呤治疗的患者（例如 IBD）。