Coronary Computed Tomographic Prediction Rule for Time-Efficient Guidewire Crossing Through Chronic Total Occlusion: Insights From the CT-RECTOR Multicenter Registry (Computed Tomography Registry of Chronic Total Occlusion Revascularization)

ObjectivesThis study sought to establish a coronary computed tomography angiography prediction rule for grading chronic total occlusion (CTO) difficulty for percutaneous coronary intervention (PCI).BackgroundThe uncertainty of procedural outcome remains the strongest barrier to PCI in CTO.MethodsData from 4 centers involving 240 consecutive CTO lesions with pre-procedural coronary computed tomography angiography were analyzed. Successful guidewire (GW) crossing ≤30 min was set as an endpoint to eliminate operator bias. The CT-RECTOR (Computed Tomography Registry of Chronic Total Occlusion Revascularization) score was developed by assigning 1 point for each independent predictor, and then summing all points accrued. Continuous distribution of scores was used to stratify CTO into 4 difficulty groups: easy (score 0); intermediate (score 1); difficult (score 2); and very difficult (score ≥3). Discriminatory performance was tested by 10-fold cross-validation and compared with the angiographic J-CTO (Multicenter CTO Registry of Japan) score.ResultsStudy endpoint was achieved in 55% of cases. Multivariable analysis yielded multiple occlusions, blunt stump, severe calcification, bending, duration of CTO ≥12 months, and previously failed PCI as independent predictors for GW crossing. The probability of successful GW crossing ≤30 min for each group (from easy to very difficult) was 95%, 88%, 57%, and 22%, respectively. Areas under receiver-operator characteristic curves for the CT-RECTOR and J-CTO scores were 0.83 and 0.71, respectively (p < 0.001). Both the original model fit and 10-fold cross-validation correctly classified 77.3% of lesions.ConclusionsThe CT-RECTOR score represents a simple and accurate noninvasive tool for predicting time-efficient GW crossing that may aid in grading CTO difficulty before PCI. (Computed Tomography Angiography Prediction Score for Percutaneous Revascularization for Chronic Total Occlusions [CT-RECTOR]; NCT02022878)     

Anti-inflammatory and anti-hyperalgesic effect of all-trans retinoic acid in carrageenan-induced paw edema in Wistar rats: Involvement of peroxisome proliferator-activated receptor-β/δ receptors

Objective:

In this study, we investigated the role of peroxisome proliferator-activated receptors (PPAR)-β/δ receptors in carrageenan-induced inflammation and in the anti-inflammatory effects of all-trans retinoic acid (ATRA).

Materials and Methods:

The λ-carrageenan (0.1 ml of 1% w/v) was injected into intra-plantar (i.pl.) region of the hind paw to produce acute inflammation. Paw volume was measured by using the mercury plethysmography. Further, mechanical and thermal hyperalgesia (TH) were assessed by using the dynamic plantar aesthesiometer and plantar test apparatus, respectively. In addition, markers of oxido-nitrosative stress were assessed spectrophotometrically in the hind paw tissue 5 h post-carrageenan.

Results:

An i.pl injection of carrageenan has produced a marked mechanical hyperalgesia (MH) and TH in ipsilateral paw, which was associated with significant elevated oxido-nitrosative stress. Treatment with ATRA (5 mg/kg/p.o/4 days) and GW0742, a selective PPAR-β/δ receptor agonist (0.1 mg/kg/i.p/4 days), significantly decreased the paw volume, mechanical and TH as compared to vehicle control. Administration of GSK0660, selective PPAR-β/δ receptor antagonist, at a dose of (0.3 mg/kg/i.p/4 days), did not produce a significant effect on carrageenan-induced paw edema, MH and TH. However, co-administration of GSK0660 (0.3 mg/kg/i.p/4 days) along with both ATRA (5 mg/kg/p.o/4 days) and GW0742 (0.1 mg/kg/i.p/4 days), significantly reverse the decreased paw edema, MH, and TH. These observed ameliorative effects on inflammatory pain symptoms are correlated with the extent of reduction of oxido-nitrosative stress.

Conclusion:

From above findings, it can be concluded that ATRA exerts anti-inflammatory and anti-hyperalgesic effect, possibly through activation of PPAR-β/δ and subsequent reduction of oxido-nitrosative stress.     

Clinical and serological features of patients with autoantibodies to GW/P bodies

GW bodies (GWBs) are unique cytoplasmic structures involved in messenger RNA (mRNA) processing and RNA interference (RNAi). GWBs contain mRNA, components of the RNA-induced silencing complex (RISC), microRNA (miRNA), Argonaute proteins, the Ge-1/Hedls protein and other enzymes involving mRNA degradation. The objective of this study was to identify the target GWB autoantigens reactive with 55 sera from patients with anti-GWB autoantibodies and to identify clinical features associated with these antibodies. Analysis by addressable laser bead immunoassay (ALBIA) and immunoprecipitation of recombinant proteins indicated that autoantibodies in this cohort of anti-GWB sera were directed against Ge-1/Hedls (58%), GW182 (40%) and Ago2 (16%). GWB autoantibodies targeted epitopes that included the N-terminus of Ago2 and the nuclear localization signal (NLS) containing region of Ge-1/Hedls. Clinical data were available on 42 patients of which 39 were female and the mean age was 61 years. The most common clinical presentations were neurological symptoms (i.e. ataxia, motor and sensory neuropathy) (33%), Sj?gren's syndrome (SjS) (31%) and the remainder had a variety of other diagnoses that included systemic lupus erythematosus (SLE), rheumatoid arthritis (RA) and primary biliary cirrhosis (PBC). Moreover, 44% of patients with anti-GWB antibodies had reactivity to Ro52. These studies indicate that Ge-1 is a common target of anti-GWB sera and the majority of patients in a GWB cohort had SjS and neurological disease.     

GW4064激活法尼酯X受体抑制结肠癌细胞生长浸润及基质细胞衍生因子1的表达

目的 探讨法尼酯X受体（FXR）特异性激动剂GW4064抑制结肠癌细胞生长浸润的机制。方法 体外培养人结肠癌细胞系HT-29,用浓度为0、0.1、1、3、5、7、10 μmol/L的GW4064分别处理HT-29细胞72 h,采用MTT法检测细胞活力;用浓度为0、1、5 μmol/L的GW4064分别处理HT-29细胞24 h,用相差显微镜观察细胞形态;用浓度为0、1 μmol/L的GW4064分别处理HT-29细胞24 h,采用细胞划痕实验检测细胞浸润的变化;用浓度为0、1、5 μmol/L的GW4064分别处理HT-29细胞24 h,采用PCR检测FXR mRNA和基质细胞衍生因子1（SDF-1）mRNA表达的变化;用浓度为0、1、5、7 μmol/L的GW4064分别处理24 h,采用ELISA法检测细胞培养上清液中SDF-1表达的变化。于裸鼠皮下接种HT-29细胞建立裸鼠成瘤模型,灌胃给予GW4064或溶剂DMSO,16 d后检测肿瘤生长情况及瘤体中FXR与SDF-1 mRNA的表达。结果 GW4064处理HT-29细胞后,细胞生长受到抑制,且呈剂量依赖性,1、3、5、7、10 μmol/L GW4064组HT-29细胞的生长活力与对照组（0 μmol/L）相比差异均有统计学意义（P均<0.05）。用相差显微镜观察可见GW4064处理HT-29细胞后,细胞收缩变圆,从瘦长的细胞向表皮样细胞改变。细胞划痕实验结果显示GW4064处理HT-29细胞后,细胞迁移距离变短,与对照组（0 μmol/L）相比差异有统计学意义（P<0.05）。PCR检测结果显示GW4064处理后FXR mRNA表达呈剂量依赖性增加,而SDF-1 mRNA表达则相反。ELISA检测结果显示细胞培养上清液中SDF-1的表达量随着GW4064浓度的增加而逐渐下降,1、5、7 μmol/L GW4064组与对照组（0 μmol/L）相比差异均有统计学意义（P均<0.05）。给予GW4064后,荷瘤小鼠肿瘤体积变小,与对照组（给予DMSO）相比差异有统计学意义（P<0.05）,并且瘤体内FXR mRNA表达增加、SDF-1 mRNA表达减少。结论 FXR激活后抑制了结肠癌细胞生长浸润,同时抑制了结肠癌细胞SDF-1的表达分泌。     

PPARγ配体罗格列酮及其激动剂GW9662对脂肪细胞因子表达的影响

目的：脂肪组织是一个内分泌器官已逐渐得到了肯定,它能分泌多种信号分子如：脂联素和抵抗素。过氧化物酶体增殖物激活受体（peroxisome pro-liferator activated receptorγ,PPARγ）在脂肪组织高水平表达,胰岛素增敏剂-噻唑烷二酮类药物是它的选择性激动剂,噻唑烷二酮类药物如罗格列酮的胰岛素增敏作用部分是通过激活PPARγ调节脂联素（胰岛素增敏分子）和抵抗素（涉及胰岛素抵抗）表达介导的。但现在不同研究发现PPARγ激动剂对抵抗素的表达调控方向存在矛盾,我们的问题是当抵抗素表达增加的情况下脂联素的表达还能否上调。方法：用3T3-L1细胞株作为研究模型,分别用溶媒对照、罗格列酮（10μmol/L）、GW9662（5μmol/L）或罗格列酮＋GW9662作用细胞,然后检测脂联素和抵抗素 mRNA表达变化情况。结果：与对照组相比,罗格列酮分别增加脂联素和抵抗素 mRNA水平1.77和1.66倍,其差异具有统计学意义（P〈0.05） ;重要的是GW9662也增加脂联素水平（1.57倍,P〈0.05）但对抵抗素无影响。罗格列酮和GW9662两者合用时,仍上调adiponectin mRNA水平（对照组的1.87倍,P〈0.05） ,抵抗素的增加与罗格列酮单用比弱下降（对照组的1.31倍,P〈0.05）。结论：本研究为PPARγ激动剂（罗格列酮）和拮抗剂（GW9662）都上调脂联素的转录提供了新的证据,两者合用时GW9662不阻断罗格列酮诱导的脂联素上调作用。综合这些数据提示噻唑烷二酮类药上调脂联素的机制可能不依赖于PPARγ。并且, GW9662在增加脂联素水平的同时不上调抵抗素水平的特性进一步支持PPARγ拮抗剂用于临床治疗胰岛素抵抗的可能性。降低抵抗素表达可能不是罗格列酮胰岛素增敏作用的重要机制。我们的结果为将来研究噻唑烷二酮类药物对人脂肪细胞因子表达在剂量和时间上提供了一定的基础。     

新型抗糖尿病药GW409544的合成工艺改进

目的改进新型抗糖尿病药物GW409544的合成工艺.方法以丙酰乙酸甲酯和L-酪氨酸为起始原料,经酯化、胺基保护、溴代、环合、还原、成醚、脱保护、水解、缩合成盐9步反应合成GW409544钙盐(1).结果与结论合成GW409544钙盐,总产率为9.3%,其结构经核磁(1H-NMR)和质谱(MS)确证.改进了中间体2-(5-甲基-2-苯基-1,3-噁唑-4-基)-乙酸甲酯(7)、(2S)-2-叔丁氧羰基胺基-3-{4-[2-(5-甲基-2-苯基-1,3-噁唑-4-基)-乙氧基]-苯基}-丙酸甲酯(9)的合成工艺.     

Phase I pharmacokinetic studies evaluating single and multiple doses of oral GW572016, a dual EGFR-ErbB2 inhibitor, in healthy subjects

GW572016 is a dual EGFR-ErbB2 inhibitor that has promise as an anticancer agent. Two phase I studies were conducted to determine the safety, tolerability and pharmacokinetics of single and multiple doses given to healthy subjects. The single dose study evaluated two groups of eight subjects in an ascending dose, 4-way cross-over, while the multiple dose study evaluated twenty-seven healthy volunteers in an ascending dose, double-blind, randomized, placebo-controlled, staggered parallel design. No serious adverse events were seen in either study. The most common adverse events for subjects receiving GW572016 were headache, diarrhea, rash, cold symptoms, gastrointestinal symptoms, and elevated LFTs, which were similar between treatment and placebo groups. Absorption of single doses of GW572016 was slightly delayed, with median t(lag) of 15 minutes (range 0-90 minutes) and achieved peak serum concentrations at a median of three hours (range 1.5-6 hours) post-dose. Serum concentrations after multiple doses of GW572016 demonstrated no significant accumulation at the 25 mg dose, and approximately 50% accumulation at the 100 mg and 175 mg doses, achieving steady state in six to seven days. A modest time-dependent increase in serum concentrations also was detected with multiple doses of GW572016. Single and multiple oral doses of GW572016 were well tolerated in healthy subjects, and resulted in dose-related systemic exposure of GW572016.     

Effects of antifolates on the binding of 5-fluoro-2'-deoxyuridine monophosphate to thymidylate synthase

Folate based inhibitors of thymidylate synthase (TS) might facilitate binding of 5-fluoro-2'-deoxyuridine-5'-monophosphate (FdUMP) to TS similar to the natural reduced folate 5,10-methylenetetrahydrofolate (CH(2)-H(4)-folate). We studied the lipophilic, non-polyglutamatable antifolates Nolatrexed (NTX) and AG331 and antifolates, that can have a polyglutamate side chain like the natural folate CH(2)-H(4)-folate; GW1843U89, Raltitrexed (RTX) and Multi-targetted antifolate (MTA) and pentaglutamates (RTX-Glu(5) and MTA-Glu(5)). The capacity of these compounds to facilitate the binding of [(3)H]FdUMP to Lactobacillus casei TS and an ammoniumsulphate precipitate of human TS was investigated. Only NTX, RTX-Glu(5) and MTA-Glu(5) facilitated FdUMP binding to L. casei TS and their dissociation constant K(d) (0.2-0.7 microM) was low compared to CH(2)-H(4)-folate (2.0 microM). The small lipophilic molecule NTX was favorable to the larger AG331. Polyglutamylation, as indicated by the difference in effect of RTX vs. RTX-Glu(5) and MTA vs. MTA-Glu(5), seems to be important for a classical antifolate to facilitate binding of FdUMP to bacterial TS. Effects of antifolates on FdUMP binding to human TS were different. At a low concentration (0.05 microM) NTX, RTX-Glu(5) and MTA-Glu(5) facilitated 3-5 times higher binding of [(3)H]FdUMP to TS than CH(2)-H(4)-folate. At higher concentrations (0.3-5 microM) of NTX, RTX-Glu(5) and MTA-Glu(5) the FdUMP binding decreased. The complex remained stable in the absence of (anti)folate for at least 24hr. The K(d) values of the antifolates for human TS varied from 19 to 387 nM, while the K(d) of CH(2)-H(4)-folate for human TS was 351 nM. The Hill coefficients, which indicated the type of cooperativity of the antifolates in the binding of FdUMP to TS were positive (0.58-0.99) at low concentrations (<0.3 microM) and negative (-0.35 to -0.81) at concentrations >0.3 microM except for GW1843U89, which only showed negative cooperativity (-1.70). It was shown with [(14)C]NTX that when the binding of FdUMP decreased at high NTX concentrations, the binding of NTX to TS still increased. This also held for the natural substrate dUMP. The negative cooperativity of the antifolates was clearly concentration dependent. The difference between human and L. casei TS in the FdUMP binding assays with antifolates can possibly be explained by interaction of the two subunits of human TS, which was absent in L. casei TS. The binding of antifolates to one of the two subunits induced a conformational change of the other subunit. This change no longer allowed the binding of FdUMP or dUMP at the active site. In conclusion this study showed that antifolates enhanced the binding of FdUMP to TS, especially at low antifolate concentrations, that are also clinically achievable, e.g. in human plasma.