Molecular and traditional chemotherapy: A united front against prostate cancer

Castrate resistant prostate cancer (CRPC) is essentially incurable. Recently though, chemotherapy demonstrated a survival benefit (∼2 months) in the treatment of CRPC. While this was a landmark finding, suboptimal efficacy and systemic toxicities at the therapeutic doses warranted further development. Smart combination therapies, acting through multiple mechanisms to target the heterogeneous cell populations of PC and with potential for reduction in individual dosing, need to be developed. In that, targeted molecular chemotherapy has generated significant interest with the potential for localized treatment to generate systemic efficacy. This can be further enhanced through the use of oncolytic conditionally replicative adenoviruses (CRAds) to deliver molecular chemotherapy. The prospects of chemotherapy and molecular-chemotherapy as single and as components of combination therapies are discussed.     

嘌呤核苷酸补偿对海洛因依赖大鼠脑组织中TPH和GTPCH基因表达的影响

目的研究嘌呤核苷酸补偿对海洛因依赖大鼠脑组织中色氨酸羟化酶(tryptophan hydroxylase,TPH)、GTP环水化酶1(GTP cyclohydrolase1,GTPCH)基因表达的影响。方法剂量递增腹腔注射海洛因,建立海洛因依赖大鼠模型,50只正常雄性Wistar大鼠随机分为对照组(C)、海洛因组(H)、海洛因补偿AMP+GMP组(HAG)、海洛因补偿AMP组(HA)、海洛因补偿GMP组(HG),每组10只。Real-timePCR法测定各组大鼠脑组织中TPH和GTPCH1基因的表达。结果与C组比较,H组大鼠脑组织中TPH基因表达下降71%(P0.05);与H组比较,HAG,HA和HG组大鼠脑组织中TPH基因表达分别升高60%,100%和150%(P0.05)。与C组比较,H组大鼠脑组织中,GTPCH基因表达下降43%(P0.05);与H组比较,HAG,HA和HG组大鼠脑组织中GTPCH基因表达均有所升高,但差异不显著(P0.05)。结论海洛因给药使大鼠脑组织中TPH,GTPCH基因表达下调;嘌呤核苷酸补偿可以减弱海洛因对TPH和GTPCH基因表达的影响。     

Abnormal development of hypoxanthine–guanine phosphoribosyltransferase-deficient CNS neuroblastoma

Lesch–Nyhan syndrome encompasses a host of neurological symptoms, caused by a deficiency of the purine salvage enzyme, hypoxanthine–guanine phosphoribosyltransferase (HGPRT). How the absence of this enzymes activity affects development of the nervous system is unknown. In this study, we examined the ability of N2aTG, a HGPRT-deficient neuroblastoma and its HGPRT-positive counterpart to proliferate and differentiate at various densities. In summary, N2aTG cells proliferated less and differentiated more than N2a cells, with the former cells exhibiting enhanced sensitivity to the effects of low-density culture. Given the homogeneity of this neuroblastoma cell line and its use in studies of neuronal development, the present study indicates that N2aTG cells may prove a suitable in vitro model for the study of non-dopaminergic neuronal development in Lesch–Nyhan syndrome.     

不同器官保存液对大鼠肝脏缺血再灌注损伤时嘌呤核苷磷酸酶活性和透明质酸吸收率的影响

目的 探讨大鼠肝脏低温保存及常温缺血再灌注过程中在不同的保存液中嘌呤核苷磷酸酶(PNP)活性和透明质酸(HA)吸收率的变化.方法 将大鼠肝脏在三种不同保存液中低温保存16 h和24 h后,用37℃Krebs-Henseleit液连续循环灌注90 min,分别于不同灌注时间检测灌洗液中PNP活性和外源性透明质酸的吸收率的变化.结果 经过16 h的低温保存后,再灌注60 min前,HTK保存的肝脏中PNP明显高于uw和Celsior;60 min后HTK和Celsior保存的肝脏中PNP明显高于UW;经过24 h的低温保存后,再灌注15 min后,HTK保存的肝脏中PNP明显高于Celsior,而Celsior又明显高于UW.低温保存16 h后,再灌注时,3种保存液保存的肝脏对外源性透明质酸的吸收率均为负值,表明肝窦内皮细胞受到一定程度的损伤;保存24 h者,UW液保存肝脏外源性透明质酸的吸收率明显高于Celsior液和HTK液.结论 随着低温保存和再灌注时间的延长,大鼠肝脏中PNP活性逐渐增高,而外源性透明质酸的吸收率下降;二者可作为评价肝脏缺血再灌注损伤的指标.     

咬合干扰致大鼠咬肌能量代谢产物含量变化

目的：观察咬合干扰后不同时间大鼠咬肌能量代谢产物腺嘌呤核苷三磷酸（adenosine triphosphate,ATP）、腺嘌呤核苷二磷酸（adenosine diphosphate,ADP）、次黄嘌呤核苷酸（inosine monophosphate,IMP）、磷酸肌酸、肌酸、乳酸及pH水平的变化,分析咬合干扰对咀嚼肌能量代谢的影响。方法：选用雄性Sprague-Dawley大鼠（220~250 g）50只,随机分为实验组（40只）和对照组（10只）,实验组于右上第一磨牙粘固0.4 mm厚金属冠建立咬合干扰,并分别维持3、7、10、14 d（每个时间点各10只）, 对照组不施加咬合干扰。各组大鼠全麻下取双侧咬肌组织,其中5只大鼠样本加入0.4 mol/L高氯酸（10 mL/g）充分匀浆,离心、过滤后采用高效液相色谱分析ATP、ADP、IMP、磷酸肌酸、肌酸及乳酸含量,另外5只大鼠样本加入含5 mmol/L碘醋酸钠的匀浆液（10 mL/g）,充分匀浆后在37 ℃恒温水浴环境中利用pH计测试pH值。结果：与对照组相比,大鼠双侧咬肌ATP含量在咬合干扰3 d ［右侧：（5.36±0.13） μmol/g,左侧：（5.77±0.25） μmol/g］ 升高（P<0.05）,7、10和14 d没有显著改变;大鼠双侧咬肌IMP［右侧：（0.21±0.03） μmol/g,左侧：（0.19±0.03） μmol/g］、肌酸［右侧：（24.76±2.94） μmol/g,左侧：（27.75±2.23） μmol/g］含量在咬合干扰7 d升高（P<0.05）,3、10和14 d没有显著改变;大鼠双侧咬肌磷酸肌酸含量在咬合干扰7、10和14 d降低［右侧分别为：（10.70±0.71） μmol/g、（11.57±0.52） μmol/g、（10.74±1.39） μmol/g,左侧分别为：（10.05±0.57） μmol/g、（10.75±1.12） μmol/g、（10.61±1.15） μmol/g, P<0.05］,3 d没有显著改变;大鼠双侧咬肌ADP、乳酸含量及pH水平在咬合干扰后各时间点均没有显著改变（P>0.05）。结论：咬合干扰导致大鼠咀嚼肌能量代谢产物含量改变,可能与咬合干扰诱发咀嚼肌疼痛、功能紊乱、肌纤维构筑改变等病理过程相关。     

Cloning and sequencing of a human cDNA coding for a multifunctional polypeptide of the purine pathway by complementation of the ade2-101 mutant in Saccharomyces cerevisiae

Summary A HeLa cell cDNA library on a yeast expression vector was used to complement auxotrophic markers of Saccharomyces cerevisiae. Clones complementing the ade2-101 mutation harbor a 1.5 kb poly(A)⁺ tailed insert with a 425 amino acid open reading frame hybridizing with two human mRNAs of 1.5 kb and 3.1 kb. Its 5 half is homologous to Bacillus subtilis SAICAR synthetase (E.C.6.3.2.6.) and its 3 terminal half corresponds to the catalytic subunit of Escherichia coli and B. subtilis AIR carboxylase (E.C.4.1.1.21). In agreement with these homologies, pADE2H1 clones complement both ade1 and ade2 mutants of S. cerevisiae, as was also recently reported for a 3.1 kb cDNA isolated from human hepatocytes.     

Long-Term Outcomes of Hairy Cell Leukemia Treated With Purine Analogs: A Comparison With the General Population

Hairy cell leukemia (HCL) is a rare hematologic malignancy with high response rates and long progression-free survival (PFS) after treatment with purine nucleoside analogs (PNAs; Pentostatin/Cladribine). However, treatment is not curative, and subsequent treatment at relapse is often required. Rechallenge with a purine analog is commonly implemented despite limited data regarding the efficacy of this approach. We retrospectively analyzed 61 consecutive patients with HCL diagnosed between 1995 and 2013 at Cleveland Clinic. Median follow-up was 72 months (3-193). Cladribine as first-line therapy was administered to 59 patients (97%). Overall response rate (ORR) was 97%, with 78% of patients achieving complete remission (CR). PFS after response was significantly improved for patients who achieved CR compared with those with a partial remission (PR) (5-year PFS 71% vs. 39%, respectively [P = .004]). Of the 19 patients who relapsed, 12 received PNAs as second-line treatment with an ORR (83%) comparable to what these patients had with first-line treatment (ORR 92%). Overall survival of all 61 patients was excellent and superior to that of age-, sex-, and race-matched controls from the general population, possibly due to selection bias. In an analysis of a larger cohort of unselected patients in the Surveillance, Epidemiology, and End Results (SEER) database, we found that mortality rates for patients with HCL were similar to those of the general population approximately 5 years after diagnosis. These data confirm the excellent prognosis for patients with HCL after first- and second-line PNA therapy.     

嘌呤核苷磷酸化酶/氟达拉滨(PNP/fludarabine)自杀基因系统对人肝癌细胞系BEL7402体外杀伤效应的研究

背景:肿瘤细胞内表达的自杀基因可将无毒性的前药转化为高毒性代谢产物以杀伤肿瘤细胞,嘌呤核苷磷酸化酶/氟达拉滨(PNP/fludarabine)自杀基因系统具有目前已知最强的旁观者效应,成为基因治疗领域中的研究热点.目的:研究PNP/fludarabine自杀基因系统对人肝癌细胞系BEL7402的体外杀伤效应.方法:应用聚合酶链反应(PCR)从大肠杆菌K12菌株基因组DNA扩增PNP基因,通过8个甘氨酸的接头克隆至真核表达载体pEGFP-M.将pEGFP-N-PNP以Lipofectamine 2000转染BEL7402细胞,G418筛选,直至出现抗性克隆,命名为BEL7402/PNP.应用逆转录(RT)-PCR和RNA印迹法(Northern blotting)鉴定PNP基因的表达.分别应用四甲基偶氮唑蓝(MTT)比色法和Annexin V-FITC凋亡检测试剂盒检测PNP/fludarabine自杀基因系统对BEL7402细胞的细胞毒效应、旁观者效应和诱导凋亡作用.结果:RT-PCR和RNA印迹法证实转染的PNP基因能在BEL7402和BEL7402/PNP细胞中表达.低浓度fludarabine对经PNP基因转染的BEL7402细胞具有明显的细胞毒效应;旁观者效应检测显示,10%的BEL7402/PNP细胞与90%的BEL7402细胞混合,经低浓度fludarabine处理,90%的细胞可被杀死;细胞凋亡检测显示,PNP/fludarabine自杀基因系统对BEL7402细胞具有明显的诱导凋亡作用.结论:PNP/fludarabine自杀基因系统对人肝癌细胞系BEL7402有明显的杀伤作用和旁观者效应.     

Effects of differentiation-inducing agents on purine nucleotide metabolism in an ovarian cancer cell line

The effects of the differentiation-inducing agents sodium butyrate (NaOBt), dimethylsulfoxide (DMSO) and mycophenolic acid (MA), on purine nucleotide metabolism, was studied in an ovarian carcinoma cell line (GZL-8). Exposure to these agents inhibited cell proliferation, but did not affect cell viability. Three hours following exposure, NaOBt and DMSO moderately decelerated purine synthesis de novo, but MA accelerated it three-fold, this being associated with a two-fold increase in the excretion of hypoxanthine and xanthine into the incubation medium. NaOBt and DMSO did not affect the cellular nucleotide content, but MA caused a 73% decrease in GTP content and about a 50% increase in the cellular content of UTP. The following alterations in cellular enzyme activity were observed 72 h following exposure: NaOBt decreased the activity of hypoxanthine-guanine phosphoribosyltransferase and increased the activity of IMP and of IMP 5-nucleotidases, DMSO increased the activity of IMP 5-nucleotidase, and MA increased the activity of the two nucleotidases. The results suggest that, in the carcinoma cell line studied, the differentiation process induced by NaOBt and DMSO may be associated with a general shift in the direction of purine metabolism from anabolism to catabolism, whereas that induced by MA is associated with a specific decrease in the production of GTP.Abbreviations DMSO dimethylsulfoxide - MA mycophenolic acid - PRibPP 5-phosphoribosyl 1-pyrophosphate - NaOBt sodium butyrate - HGPRT hypoxanthine-guanine phosphoribosyltransferase     

李小娟教授辨治急性期痛风性关节炎

李小娟教授认为古人对痛风认识无外乎素体亏虚、伤于湿、伤于热。急性期痛风性关节炎病因,既有外因又有内因、内外因夹杂。急则治其标,缓则治其本,以调和营卫,清热祛湿为治则随证加减。选用《金匮要略.中风历节病》桂枝芍药知母汤为主方辨证加减以调和营卫,清热祛湿。湿热偏盛者加入四妙散;关节红肿热痛较重者据肺主皮毛之理,酌加泻白散,"诸痛痒疮,皆属于心",灼热疼痛之时可用清心火之药物,清心除烦泻火解毒,如黄连、连翘之类,以缓解疮疡之红肿热痛。若有瘀血者加入全虫、赤芍、伸筋草等活血通络之品。