Phase I Trial ofU racil-Tegafur (UFT) plus Oral Leucovorin: 28-Day Schedule

UFT [Taiho Pharmaceutical Co. Ltd., Tokyo, Japan; (BMS-200604), Bristol-Myers Squibb, Princeton, NJ], a fluorouracil prodrug, is an oral 4:1 molar concentration of uracil plus tegafur. This study examined the dose-limiting toxic effects and maximum tolerated dose of UFT plus leucovorin administered for 28 consecutive days followed by a 7-day rest period. A course of therapy was repeated every 35 days. UFT dose levels examined were 200 mg/m²/day, with planned escalations to 250, 300, 350, and 400 mg/m²/day; the leucovorin dose remained at 150 mg/day. Three patients were initially enrolled at each UFT dose level. The total daily doses of both UFT and leucovorin were divided into three doses administered every 8 hr. Diarrhea became the dose-limiting toxicity at 400 mg/m²/day UFT, with grade 3 diarrhea noted in 2 of the 3 patients receiving that dose. To further define a phase II UFT starting dose, 3 additional patients were entered at the 350 mg/m² level; 3 of the 6 patients treated at this level developed grade 3 nonhematological toxic effects. No partial or complete responses were observed. The recommended phase II UFT starting dose is 300 mg/m²/day plus 150 mg/day leucovorin. Since neutropenia, significant mucositis, and “hand-foot syndrome“ were not observed with UFT plus leucovorin, the toxicity profile of this regimen appears favorable compared with that of intravenous regimens of fluorouracil plus leucovorin. This phase I trial of UFT served as the basis for a phase II trial, current phase III trials, and a national adjuvant therapy trial of UFT for high-risk colon cancer patients.     

骨髓间充质干细胞促进大鼠IgA肾病修复

目的 观察骨髓间充质干细胞（MSC）对大鼠IgA肾病有无修复作用，并探讨其可能的机制。 方法 SD大鼠随机分为MSC注射组、生理盐水（NS）组及健康对照组。前两组以牛血清白蛋白(BSA)+葡萄球菌肠毒素B（SEB）＋皮下注射四氯化碳（CCl4）的改良法建立IgA肾病模型。体外连续培养SD大鼠MSC并通过流式细胞仪和成骨成脂细胞诱导分化鉴定MSC，用5-溴脱氧尿嘧啶核苷（BrdU）体外标记培养的MSC。移植后1周及4周分别观察3组的体质量、尿蛋白量（24 h）、肾功能、肾脏病理变化、IgA荧光沉积变化；ELISA法检测尿中的MCP-1、TGF-β1量；RT-PCR法检测肾组织中MCP-1、TGF-β1 mRNA的表达情况；免疫组化观察细胞因子及BrdU标记的MSC在肾组织中的分布情况。 结果 移植后1周，MSC组尿蛋白量（24 h）（36.86±4.78） mg，Scr（53.50± 6.28） μmol/L；NS组尿蛋白量（24 h）(66.98±5.86) mg，Scr (82.50±8.36) μmol/L，两组差异有统计学意义（均P < 0.05）；同时，MSC组MCP-1、TGF-β1在尿中的含量及肾脏中表达均显著低于NS组（均P < 0.05）。移植后4周，MSC组体质量、肾脏病理变化、IgA荧光沉积与NS组差异有统计学意义；MCP-1、TGF-β1在尿中的含量及肾脏中的表达与健康对照组差异无统计学意义。随时间延长，BrdU标记的MSC在肾组织中分布却逐渐减少。 结论 MSC输注可促进大鼠IgA肾病的修复，其作用机制可能并不完全是依赖于MSC的直接分化，而是通过调节肾组织中细胞因子的分泌和（或）其他的功能进行修复。     

Circumvention of 5-Fluorouracil Resistance in Human Stomach Cancer Cells by Uracil Phosphoribosyltransferase Gene Transduction

A human stomach cancer cell line with acquired resistance to 5-fluorouracil (5-FU), NUGC-3/5FU/L, has been found to possess reduced ability to convert 5-FU into active metabolites. We attempted in vitro gene therapy for this 5-FU-resistant cell line. NUGC-3 and NUGC-3/5FU/L cells were infected with recombinant adenovirus (Ad) containing Escherichia coli uracil phosphoribosyltransferase ( UPRT ) gene driven by CAG promoter (CA), AdCA-UPRT, and changes in their 5-FU metabolism and sensitivity were investigated. Activities of orotate phosphoribosyltransferase increased from 10.2 and 1.56 (nmol/mg protein/30 min) in the uninfected cells of NUGC-3 and NUGC-3/5FU/L to 216 and 237, respectively, after the transfection of UPRT gene. The 5-FU nucleotide level in the acid-insoluble fraction increased from 7.32 to 15.9 (pmol/mg protein) in NUGC-3 cells on infection with AdCA-UPRT, and in NUGC-3/5FU/L cells it increased from 1.91 to 21.4. The 50% growth-inhibition concentration (IC50) was 12.7 μmol/liter for NUGC-3 and much higher than 100 μmol/liter for NUGC-3/5FU/L, indicating over 8-fold resistance. NUGC-3/5FU/L transfected with the UPRT gene showed very high sensitivity to 5-FU with an IC₅₀ of 3.2 μmol/liter. The high resistance in this metabolic activation-deficient cell line was thus completely reversed by transduction of an exogenous gene coding for a 5-FU-anabolizing enzyme.     

采用十二烷基硫酸钠抑制PCR产物中残留尿苷酶

目的:研究抑制PCR产物中残留尿苷酶(uracil DNA glycosylase,UDG)活性的试剂.方法:采用DNA-EIA技术检测在全dU-DNA扩增物中含有稳定剂和不含稳定剂的样品,置室温24 h和37 ℃ 48 h的DNA杂交百分率.结果:(1)在PCR产物中加入稳定剂A液和B液置室温24 h,取5 μl直接杂交,其杂交百分率为61.470%和68.996%,对照组为99.283%.(2)在30 μl PCR产物中加入稳定剂后,并加1×PCR缓冲液稀释至70 μl,置37 ℃ 48 h取10 μl样品杂交,5 μl和2.5 μl A液组杂交百分率为71.092%和67.691%,对照组为68.020%和63.906%.5 μl和2.5 μl B液组杂交百分率为91.333%和80.307%,对照组为63.906%和68.020%.结论:B液可抑制PCR产物残留UDG活性,对保护全dU-PCR产物起重要作用.     

Microwave-assisted synthesis and biological evaluation of novel uracil derivatives inhibiting human thymidine phosphorylase

New 5-chloro-6-substituted-uracil derivatives have been prepared by microwave assisted-synthesis and tested in vitro as thymidine phosphorylase inhibitors. One of these compounds showed potent inhibitory activity, with an IC50 value in the submicromolar range. The biological activity of the new compounds is discussed in terms of structure-activity relationship.     

Clinical implications of dihydropyrimidine dehydrogenase (DPD) activity in 5-FU-based chemotherapy: mutations in the <Emphasis Type="Italic">DPD</Emphasis> gene,and DPD inhibitory fluoropyrimidines

Dihydropyrimidine dehydrogenase (DPD) is the rate-limiting enzyme in the degradation of pyrimidine bases. DPD is also responsible for the degradation of 5-fluorouracil (5-FU), which is the most frequently prescribed anticancer drug for the treatment of malignancies of the gastrointestinal tract. DPD could influence the antitumor effect and the adverse effects of 5-FU. High intratumoral DPD activity markedly decreases the cytotoxic effect of 5-FU. More than 80% of administered 5-FU is detoxified and excreted as F--alanine in urine. In 5-FU-based chemotherapy, escape from the degradation catalyzed by DPD is important. Recently, the dihydropyrimidine dehydrogenase gene (DPYD) was isolated, and its physical map and exon-intron organization were determined. To date, many DPYD variant alleles associated with a lack of DPD activity have been identified. In 5-FU-based cancer chemotherapy, severe toxicities were observed at higher rates in patients who were heterozygous for a mutant DPYD allele, compared with toxicities in patients who were homozygous for the wild DPYD allele. Furthermore, the adverse effects of 5-FU are often lethal for patients homozygous for the mutant DPYD allele. The apparently high prevalence of the DPYD mutation associated with lack of DPD activity in the normal population warrants genetic screening for the presence of these mutations in cancer patients before the administration of 5-FU. DPD inhibitory fluoropyrimidines (DIFs), including uracil plus tegafur (UFT) and tegafur plus 5-chloro-2,4-dihydroxypyridine plus potassium oxonate, in a molar ratio of 1:0.4:1 (TS-1), have recently been used in clinical settings. DIFs should provide chemotherapy that improves both quality of life and duration of survival.     

Timing Effects of Uracil-induced Urolithiasis on Amplification of Second-stage Promotion in Rat Bladder Carcinogenesis

The post-initiation enhancing activities of the non-genotoxic agent NaHCO₃ and the genotoxic agent N -ethyl-] N -(4-hydroxybuty])nitrosainine (EHBN) in combination with uracil-induced urolithiasis were investigated in a rat bladder carcinogenesis model. Animals were treated with 0.05% N -butyl- N -(4-hydroxybutyl)nitrosamine (BBN) for 4 weeks, and then 3% uracil was given for 3 weeks in the early (weeks 4-7), middle (weeks 8 11) or late (weeks 12–15) post-initiation phase. In addition, administration of 3% NaHCO₃, 20 ppm EHBN or no chemical supplement was performed for the 13 weeks when the rats were not receiving BBN or uracil. NaHCO₃ in sequential combination with early and middle stages uracil treatment strongly enhanced tumorigenesis in the urinary bladder, while EHBN treatment amplified lesion development at the middle stage only of uracil treatment. DNA synthesis and associated epithelial surface alterations observed by scanning electron microscopy tended to be increased in the NaHCO₃ and EHBN groups without BBN initiation, independently of uracil treatment timing. The present results demonstrated that uracil-induced urolithiasis during the middle post-initiation phase is highly active in enhancing bladder tumor development under the influence of a promoter or carcinogen.     

CX-659S: a novel diaminouracil derivative that has antioxidative and acute anti-inflammatory activities

We investigated the antioxidative activities and the effects on acute inflammation in mice of a novel diaminouracil derivative, CX-659S ((S)-6-amino-5-(6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxamido)-3-methyl-1-phenyl-2,4(1H,3H)-pyrimidinedione). CX-659S showed potent scavenging activities against the hydroxyl radical and peroxynitrite and inhibited lipid peroxidation in rat brain homogenates in vitro. Topically applied CX-659S dose-dependently inhibited arachidonic acid- and 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced ear edema in mice. Consistent with its antioxidative properties in vitro, CX-659S dramatically attenuated the accumulation of lipid peroxides in the mouse ear elicited by repeated application of TPA. Previously, we reported the effectiveness of CX-659S against contact hypersensitivity reactions in both mouse and guinea pig models. These present results further suggest the therapeutic potential of CX-659S for acute skin inflammation that may involve oxidative tissue damage.     

mdr1启动子调控CD::UPP基因对紫杉醇耐药卵巢癌细胞的杀伤作用

目的探讨腺病毒介导的mdr1启动子调控胞嘧啶脱氨酶尿嘧啶磷酸核糖转移酶(CDUPP)融合基因联合5-氟胞嘧啶(5-FC)对紫杉醇耐药卵巢癌细胞的特异性杀伤作用.方法扩增、纯化含有mdr1-CDUPP基因的重组腺病毒,转染人卵巢癌紫杉醇耐药细胞株A2780/Taxol和亲本细胞株A2780,RT-PCR检测mdr1和CDUPP基因的表达水平;之后加入5-FC,MTT法检测细胞抑制情况及旁观者效应,并观察腺病毒转染后裸鼠移植瘤的生长情况.结果mdr1和CDUPP基因在A2780/Taxol细胞中可稳定表达,转染后A2780/Taxol组的细胞生长明显低于A2780组;转基因的A2780/Taxol细胞联合5-FC后可通过旁观者效应杀伤周围未转基因的耐药细胞;耐药组移植瘤生长明显受到抑制,肿瘤体积为(569.10±187.93)mm3,对照组肿瘤体积为(2 111.98±230.82)mm3,差异有统计学意义(P＜0.01).结论mdr1启动子可调控CDUPP基因特异性表达并特异性杀伤紫杉醇耐药卵巢癌细胞.     

HPLC法测定各沪产地龙和广地龙中次黄嘌呤、黄嘌呤、尿嘧啶和尿苷的含量

目的 测定各种沪产地龙与广地龙中次黄嘌呤、黄嘌呤、尿嘧啶、尿苷的含量,建立地龙药材质量评价的方法.方法 用0.9％生理盐水超声提取地龙,SORBAX SB-Aq色谱柱(250 mm×4.6mm,5μm,Aglient Co.,Ltd),流动相:5 mmol/L磷酸二氢钾(pH 2.9),流速1 mL/min,检测波长254 nm,柱温30℃,进样量10μL,采用HPLC法,同时测定次黄嘌呤、黄嘌呤、尿嘧啶、尿苷4种核苷类成分的含量.结果 次黄嘌呤的线性范围为0.500 0～100.00 μg (r=0.999 9),平均回收率99.37％,RSD=1.36％(n=6);黄嘌呤的线性范围为0.500 0～100.00 μg(r=0.993 1),平均回收率91.57％,RSD=1.40％(n=6);尿嘧啶的线性范围为0.500 0～100.00 μg (r=0.999 9),平均回收率95.31％,RSD=1.64％(n=6);尿苷的线性范围为0.500 0～100.00 μg(r=0.999 9),平均回收率100.21％,RSD=1.98％(n=6).结论 该方法重复性、回收率好,可用于测定地龙与广地龙中次黄嘌呤等4种核苷类成分的含量.