Carboxylesterase catalyzed 18O-labeling of carboxylic acid and its potential application in LC-MS/MS based quantification of drug metabolites

LC-MS quantification of drug metabolites is sometimes impeded by the availability of internal standards that often requires customized synthesis and/or extensive purification. Although isotopically labeled internal standards are considered ideal for LC-MS/MS based quantification, de novo synthesis using costly isotope-enriched starting materials makes it impractical for early stage of drug discovery. Therefore, quick access to these isotope-enriched compounds without chemical derivatization and purification will greatly facilitate LC-MS/MS based quantification. Herein, we report a novel ¹⁸O-labeling technique using metabolizing enzyme carboxylesterase (CES) and its potential application in metabolites quantification study. Substrates of CES typically undergo a two-step oxygen exchange with H₂¹⁸O in the presence of the enzyme, generating singly- and doubly-¹⁸O-labeled carboxylic acids; however, unexpected hydrolytic behavior was observed for three of the test compounds – indomethacin, piperacillin and clopidogrel. These unusual observations led to the discovery of several novel hydrolytic mechanisms. Finally, when used as internal standard for LC-MS/MS based quantification, these in situ labeled compounds generated accurate quantitation comparable to the conventional standard curve method. The preliminary results suggest that this method has potential to eliminate laborious chemical synthesis of isotope-labeled internal standards for carboxylic acid-containing compounds, and can be developed to facilitate quantitative analysis in early-stage drug discovery.     

通心络胶囊联合阿托伐他汀及氯吡格雷治疗冠心病心绞痛临床研究

目的:观察通心络胶囊联合阿托伐他汀、氯吡格雷治疗冠心病心绞痛的临床疗效。方法:选取84例冠心病心绞痛患者,按随机数字表法分为观察组与对照组,对照组40例给予阿托伐他汀联合氯吡格雷治疗,观察组44例在对照组基础上给予通心络胶囊治疗,3个月后比较2组临床疗效。结果:观察组总有效率(93.18%)显著高于对照组(70.00%),差异有统计学意义(P<0.05)。治疗前,2组心绞痛的持续时间、心绞痛发作次数、心功能指标、血管内皮功能指标比较,差异无统计学意义(P>0.05)。治疗后,2组心绞痛持续时间、心绞痛发作次数较治疗前明显降低,观察组心绞痛持续时间、心绞痛发作次数显著低于对照组(P<0.05);2组心功能指标包括心输出量(CO)、心博出量(SV)、心脏指数(CI)及射血分数(EF)均较治疗前明显上升,且观察组各项心功能指标明显高于对照组(P<0.05);2组内皮功能指标内皮素(ET)、血栓素B2(TXB2)水平较治疗前明显降低,一氧化氮(NO)水平较治疗前明显上升(P<0.05),观察组ET、TXB2水平明显低于对照组,NO水平明显高于对照组(P<0.05)。结论:对冠心病心绞痛患者给予通心络胶囊联合阿托伐他汀、氯吡格雷治疗疗效显著,可有效降低心绞痛持续时间和发作次数,改善患者心功能和内皮功能,值得临床推广应用。     

PON1Q192R基因多态性与冠心病患者使用氯吡格雷疗效的关系

目的：探讨不同民族PON1Q192R等位基因变异频率以及PON1Q192R基因多态性与冠心病患者氯吡格雷抵抗（CR）的关联性。方法：纳入新疆医科大学第二附属医院2020年1月至2020年12月使用氯吡格雷且诊断为冠心病的患者127例,患者每日服用氯吡格雷75 mg,5 d后用血栓弹力图测定相关参数,并进行CYP2C19*2、CYP2C19*3、CYP2C19*17和PON1Q192R基因分型检测。结果：不同民族之间PON1Q192R基因突变频率差异具有统计学意义（P <0.05）,患者的血栓弹力图参数结果显示,PON1Q192R基突变并不会显著降低氯吡格雷疗效。结论：PON1Q192R基因突变频率与冠心病患者氯吡格雷反应的相关性还需要进一步研究。     

服用氯吡格雷的老年髋部骨折患者尽早手术的安全性研究

目的探讨服用氯吡格雷的老年髋部骨折患者尽早手术的安全性。方法回顾性分析2016年11月至2017年4月期间北京积水潭医院创伤骨科连续收治的242例老年髋部骨折患者资料。将服用氯吡格雷且停药4 d以内手术患者纳入研究组,共20例,年龄中位数为80(77,81)岁;男6例,女14例。手术方式:股骨颈骨折空心钉固定2例,股骨颈骨折股骨头置换11例,股骨转子间骨折内固定7例。将未服用抗凝、抗栓药且手术方式与研究组一致的患者纳入对照组(222例)。比较两组患者的入院至手术时间、手术时间、术中出血量、围手术期输血量和出血相关并发症等,分析尽早手术的安全性。结果两组患者术前一般资料比较差异均无统计学意义(P>0.05),具有可比性。研究组患者的入院至手术时间为42.5(36.3,48.0)h,与对照组患者[43.0(28.0,61.0)]比较差异无统计学意义(P=0.870)。研究组患者末次服用氯吡格雷至手术时间为55.0(30.5,71.0)h。两组患者的术前血红蛋白、手术时间、术中出血量、输血比例、输血量、切口引流放置比例、引流量等方面比较差异均无统计学意义(P>0.05)。研究组患者的全身麻醉比例(45.0%,9/20)显著高于对照组患者(18.5%,41/222),差异有统计学意义(P=0.012)。研究组椎管内麻醉患者均未发现椎管内出血的相关并发症。术后3个月内对照组2例患者出现切口血肿、继发感染。结论服用氯吡格雷的老年髋部骨折患者,停用氯吡格雷后可以尽早手术。     

氯吡格雷联合阿司匹林、溶栓治疗急性ST段抬高型心肌梗死的疗效观察

目的观察常规溶栓联合氯吡格雷治疗对急性ST段抬高型心肌梗死近期心脏事件的影响。方法符合入选条件的患者随机分为常规溶栓治疗组(34例)和氯吡格雷组(36例,在常规溶栓治疗的基础上即刻300mg,随后每日1次75mg,连用7d)。结果7d后氯吡格雷组复合终点(13.90%)与常规治疗组(29.41%)比较差异有统计学意义(P<0.05),而主要的副作用两组之间比较差异无统计学意义(P>0.05)。结论氯吡格雷对急性ST段抬高型心肌梗死患者的近期(7d)心脏事件是安全有效的。     

Thrombolytic therapy-associated acute myocardial infarction in patients with acute ischemic stroke: A treatment dilemma

Acute myocardial infarction (AMI) is uncommon in the acute phase of acute ischemic stroke (AIS) and occurs in approximately 1% of the population. Here, we report a paradoxical case of AMI during tissue plasminogen activator (t-PA) infusion for AIS. We review and analyze the previously reported cases. We found that only patients with AMI which occurred after thrombolytic therapy for AIS who received an adequate combination of anticoagulation plus percutaneous coronary intervention survived their events. Several mechanisms have been proposed for the development of AMI after thrombolytic therapy. These mechanisms include fragmented intra-cardiac thrombus, intensified platelet aggregation that may lead to an increased potential for intra-cardiac thrombus formation, and a reduction in clot-associated plasminogen that may lead to a paradoxical hypercoagulable state of the coronary arteries. Currently, there is no consensus regarding this specific scenario. We propose that the therapeutic benefit and the potential risk of hemorrhagic complications should be further investigated and individualized. In patients who receive thrombolytic therapy for AIS and who then develop post-thrombolytic AMI, we suggest that the maximum treatment for the subsequent AMI be instituted promptly to avoid short-term mortality.     

Switching from Clopidogrel to Prasugrel in patients undergoing PCI: A meta-analytic overview

Despite the demonstrated benefits of Prasugrel, a new generation thienopyridine, in the prevention of thrombotic complications after percutaneous coronary interventions (PCI) for Acute Coronary Syndromes (ACS), its use is still precluded to those many patients arriving to the cath lab pre-treated with Clopidogrel. Conclusive data on the strategy of switching from Clopidogrel to Prasugrel are still missing, therefore we aimed to perform a meta-analysis of current studies evaluating the safety and efficacy of switching from Clopidogrel to Prasugrel (PS) as compared to a standard thienopyridine therapy with Clopidogrel or Prasugrel in patients undergoing PCI. Literature archives and main scientific sessions’ abstracts were scanned for studies comparing a switching strategy from Clopidogrel to Prasugrel vs. Prasugrel or Clopidogrel. Primary efficacy endpoint was overall mortality. Secondary endpoints were: non-fatal myocardial infarction and definite/probable stent thrombosis. Safety endpoint was the rate of major bleedings according to a per-protocol definition. A total of 12 studies, involving 3956 patients, were included. Among them, 1396 patients (35.3%), received Prasugrel after a Clopidogrel treatment (PS), while 2560 (64.7%) received either Prasugrel or Clopidogrel. The switch from Clopidogrel to Prasugrel was in the majority of the studies periprocedural. The mortality was numerically lower, but not statistically significant, in the PS group as compared with patients who did not switch (1.7% vs. 3.8%, OR [95% CI]?=?0.68 [0.40,1.15], p?=?0.15, p_het?=?0.61), without any relationship with patients’ risk profile (r?=??0.68 [?2.09, 0.73], p?=?0.35). Similar results were obtained for secondary efficacy endpoints and at sensitivity analysis in the majority of subgroups evaluated. Moreover, the PS strategy did not increase major bleedings as compared with standard therapy (1.4% vs. 2.5%, OR [95% CI?=?0.70 [0.39, 1.25], p?=?0.23, p_het?=?0.6). The present meta-analysis confirms that, among patients undergoing PCI, switching from Clopidogrel to Prasugrel may be safely performed and therefore should be encouraged among patients eligible to Prasugrel.     

低剂量氯吡格雷在冠状动脉再狭窄家猪模型中应用的初步探讨

目的评价低剂量氯吡格雷在冠状动脉再狭窄家猪模型中的应用效果。方法选取健康家猪6头,抽取静脉血测定ADP(Adenosine Diphosphate,二磷酸腺苷)诱导的血小板聚集率作为基础值,然后开始喂食氯吡格雷(25mg/d)和常规剂量的阿司匹林,开始给药后第4天行介入手术构建冠状动脉过度扩张再狭窄模型并行支架植入术,同时在给药后第4、8﹑14天再次抽血测定ADP诱导的血小板聚集率,评价氯吡格雷对血小板聚集率的抑制效果,记录术后压迫止血情况及术中、术后的安全性,14天时处死动物并血管取材评价支架内血栓情况。采用SPSS10.0进行统计学分析。结果家猪平均体重为(23.1±4.4)Kg,ADP诱导的血小板聚集率基础值为(43.08±5.06)%。用药后第4天,血小板聚集率值较基础值明显下降[(29.05±9.26)%:(43.08±5.06)%,P=0.008],用药后8天血小板聚集率值进一步下降,用药后14天,血小板聚集率值趋于稳定,达到并维持抗血小板聚集的效果,术中和术后24小时均未发生冠状动脉事件,14天时血管取材病理切片未见支架内血栓形成。结论在冠状动脉再狭窄家猪模型中,低剂量氯吡格雷可以达到抑制ADP诱导的血小板聚集的效果,同时低剂量氯吡格雷联合常规剂量的阿司匹林可以保证冠状动脉再狭窄家猪模型的安全性,并降低实验费用。     

冠状动脉支架患者氯吡格雷抵抗现象探讨

目的 ①评价冠状动脉支架(CS)置入患者氯吡格雷抵抗的发生情况;②观察CS术后1个月复发缺血事件与氯吡格雷抵抗的关系.方法 ①所有入选患者于服氯吡格雷前、服药后24 h及1个月分别测定ADP(25 μmol/L)诱导的血小板聚集率,计算血小板聚集抑制率(△A),△A≤10%(包括负值)时考虑存在氯吡格雷抵抗;②观察CS术后1个月内复发缺血事件的情况.结果 ①共入选患者42例,其中△A 24 h≤10%者18例(43%),△A1m≤10%者13例(31%);②在1个月的随访中,无一例患者因心血管疾病死亡或再发心肌梗死,△A1m≤10%的患者有1例发生脑梗死,1例复发心绞痛,缺血事件发生率为15.4%,△A1m＞10%的患者未发现缺血事件.结论 ①一部分CS患者存在氯吡格雷抵抗,测定血小板聚集率有助于发现这部分患者;②氯吡格雷抵抗患者CS术后1个月内复发缺血事件增加.     

替格瑞洛在急性冠脉综合征患者介入治疗中的近期疗效评价

目的 探讨替格瑞洛对急性冠脉综合征(acute coronary syndrome,ACS)患者介入治疗近期临床疗效的影响.方法 将入选的拟行冠状动脉支架植人术(percutaneous coronary intervention,PCI)的ACS 283例患者随机分为对照组(氯吡格雷+常规治疗)和试验组(替格瑞洛+常规治疗).检测两组PCI治疗前后TIMI血流分级、血小板聚集率和血小板计数;随访30 d,记录心血管不良事件(MACE)发生率;观察治疗后出血并发症的发生率.结果 两组患者PCI术前TIMI血流分级比较,差异无统计学意义(P＞0.05),术后试验组TIMI血流3级例数明显高于对照组(P＜0.05);PCI术后试验组血小板聚集率为0.27±0.08,明显低于对照组的0.37±0.09(P＜0.05);术后随访30 d,与对照组比较,试验组MACE发生率明显低于对照组(P＜0.05);而对照组与试验组出血并发症发生率无明显差异(P＞0.05).结论 对于行PCI的ACS患者,替格瑞洛能有效改善患者术后TIMI血流分级,降低血小板聚集率,改善近期临床疗效,且不增加出血风险.