Successful use of the new immune-suppressor sirolimus in IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome)

IPEX (immune-dysregulation, polyendocrinopathy, enteropathy, X-linked) syndrome is an autoimmune disorder with an often lethal outcome in spite of immunosuppressive therapy. We report the successful use of sirolimus in 3 patients with IPEX. The efficacy of sirolimus is probably due to its different mode of action compared to calcineurin-dependent agents.     

Tsc2 is not a critical target of Akt during normal Drosophila development

Signaling by insulin and target of rapamycin are both required for cell growth, but their interrelationships remain poorly defined. It was reported that Akt, an essential component of the insulin pathway, stimulates growth by phosphorylating and inhibiting tuberous sclerosis complex 2 (TSC2). Here we evaluate this model genetically in Drosophila by engineering Tsc2 mutants in which the Akt phosphorylation sites are changed to nonphosphorylatable or phospho-mimicking residues. Strikingly, such mutants completely rescue the lethality and cell growth defects of Tsc2-null mutants. Taken together, our data suggest that Tsc2 is not a critical substrate of Akt in normal Drosophila development.     

Pharmacokinetics of rapamycin-eluting stents in miniswine coronary model

Background The results of clinical trials of rapamycin-eluting stents reduce restenosis have been quite promising. The main purpose of this study was to characterize the in vivo pharmacokinetics of high dose rapamycin (Rapa)-eluting stents in a miniswine coronary model.Methods Ten miniswines underwent placement of 18 high dose Rapa-eluting stents in the left anterior descending and right coronary arteries. At the planned times of the 1.5th, 12th, 24th hour, 3th, 7th and 28th day, the animals (n=1, 1, 2, 2, 2, and 2, respectively) were euthanized after completion of coronary angiography. Blood samples were obtained at 0, 10, 20, 30 minutes; 1, 2, 6, 24 hours; and 3, 7, 28 days to determine systemic Rapa levels. Rapa levels in whole blood, arterial wall, heart, renal and liver tissues were determined by high-performance liquid chromatography/mass spectroscopy.Results Peak whole blood concentration (Cmax), time to peak concentration (tmax), elimination half-life (t1/2β), area under the curve (AUC), and apparent systemic clearance (Cl/F) were (10.91±1.28) ng/ml, (2.0±0.2) hours, (7.25±0.63) hours, (1.15±0.11) ng·h·ml（-1）, and (180±12) ml·h（-1）·kg（-1）, respectively. More than 95% Rapa detected is localized in the coronary artery surrounding the stent and heart.Conclusion Stent-based delivery of Rapa via a copolymer stent is feasible and safe. This strategy holds promise for the prevention of stent restenosis.     

Estrogen and rapamycin effects on cell cycle progression in T47D breast cancer cells

The contribution of estrogen (and progesterone) in driving cell cycle progression of hormone dependent breast cancer cells is well documented, however, the roles of the various relevant signal transduction pathways remain unclear. The immunosuppressant rapamycin is a potent inhibitor of cell cycle progression and has been used to define signal transduction pathways. In this study we have determined rapamycin's effects on cell cycle progression in estrogen dependent breast cancer cells using a novel method of inducing S-phase. In this method estradiol-17- alone induced S-phase without mitogen support. In our studies the T47D cells were quite sensitive to estradiol-17-, with half-maximal induction in the picomolar range, indicating that the estrogen can induce S-phase in the absence of mitogens such as insulin. The estrogen response does not seem to be particularly specific because estriol estrone and estradiol-17--BSA were about as effective as estradiol-17-. R5020, a progestin also induced S-Phase, while rapamycin blocked steroid driven transition of cells from G₁ to S-phase.     

雷帕霉素对肝脏缺血再灌注损伤修复的影响

目的 应用小鼠肝脏缺血损伤模型研究雷帕霉素对损伤肝脏修复的延迟作用并初步探讨其机制.方法 建立Balb/c小鼠肝左中叶缺血损伤模型.将实验小鼠随机分为4组,A组(n=16):雷帕霉素手术组;B组(n=16):生理盐水手术组;C组(n=12):雷帕霉素假手术组;D组(n=12):生理盐水假手术组.给药后第4天对各组小鼠行手术及假手术处理.术后第1天、第4天分批处死各组动物,取出损伤肝组织作组织病理学榆测;免疫组织化学方法检测组织内增殖细胞核抗原(PCNA)及细胞周期蛋白D1(Cyclin D,)的表达.结果 ①在术后各检测时间点,A组小鼠肝组织病理学变化明显重于B组,表现为肝细胞有明显肿胀变性,肝板结构紊乱,肝窦结构不清晰,伴有肝细胞坏死;C组及D组未见明显组织病理损伤.②在各时间点.A组小鼠肝组织中PCNA阳性率显著低于B组;而且Cyelin D1的阳性率也低于B组,差异有统计学意义(均P<0.05).C组及D组未见明显PCNA及Cyclin D1的表达.结论 雷帕霉素阻抑了缺血再灌注损伤肝脏的修复过程,而对正常肝脏无明显损伤作用.     

基因多态性对器官移植中免疫抑制剂疗效影响的研究进展

环孢素、他克莫司、雷帕霉素和霉酚酸酯都是广泛用于器官移植患者的免疫抑制剂，他们具有治疗指数窄、个体差异大的特点。环孢素、他克莫司和雷帕霉素主要通过肝脏和小肠的CYP3A代谢；同时他们又是药物转运体的底物。霉酚酸酯是通过葡萄糖醛酸转移酶代谢。不同个体间药物代谢酶和转运体活性的差异，可能是造成不同器官移植患者对以上药物药代动力学差异的主要原因；而遗传因素即编码药物代谢酶和转运体基因序列的差异，可能是其产生活性差异的分子机制。因此，从编码药物代谢酶和转运体的基因入手，可能会为器官移植患者提供最优的治疗方案。     

Human Herpesvirus-8 (HHV-8)-Associated Primary Effusion Lymphoma in two Renal Transplant Recipients Receiving Rapamycin

The Akt/mammalian target of rapamycin (mTOR) signaling cascade has been demonstrated to be constitutively activated in several malignancies, including Kaposi sarcoma (KS) and human herpesvirus-8 (HHV-8)-associated primary effusion lymphoma (PEL). In organ transplant recipients, therapeutic change from cyclosporin to the mTOR inhibitor rapamycin can lead to regression of KS lesions. Recent experiments using PEL cell lines and murine xenograft PEL models suggested that rapamycin could inhibit the growth of PEL cells. In the present report, we describe the cases of two HIV-1-negative males of African origin who underwent renal transplantation and developed PEL while receiving rapamycin as immunosuppressive treatment. Both patients were retrospectively found to be HHV-8 seropositive before renal transplantation. The present case report suggests that rapamycin may not protect HHV-8-infected renal transplant recipients from occurrence of PEL or progression of pre-existing PEL.     

CD4+CD25+调节性T细胞的体外扩增

 【目的】 探讨有效的体外培养扩增CD4+CD25+Treg的方法。【方法】 收集健康志愿者外周血5例，免疫磁珠法分选出CD4+CD25+Treg。将实验分为三组，第一组在CD4+CD25+Treg中加入100 nM rapamycin (1μg/ml)和包被了CD3/CD28单抗的磁珠培养3周，第8 天开始加入1000U/mL的IL-2，第二组培养条件除不加rapamycin外其余与第一组相同，第三组培养细胞为CD4+CD25-T细胞，培养条件同第一组。培养第7、14、21天收集细胞计数，第21天收集细胞行流式细胞术三标法检测CD4、CD25、FOXP3的表达。MTT法检测体外扩增的CD4+CD25+Treg对自体和异体的CD4+T细胞增殖的抑制作用。 【结果】 三组细胞均有明显的扩增，加rapamycin培养的CD4+CD25+Treg细胞的扩增率比不加rapamycin扩增倍数低，但细胞的纯度明显增加。CD4+CD25-T细胞组在培养第一周扩增迅速，从第二周开始增殖减慢，CD4+CD25+ Treg 细胞的比例较培养前无明显的差别。三组至第三周时扩增倍数分别为 89.4+20.53、338.4+77.64 、215.6+54.58（F=484.655,p<0.0001），细胞的纯度分别为84.32+4.62%、34.04+5.78%、0.68+0.39% (F=24.660,p<0.0001)。体外抑制实验显示体外扩增的CD4+CD25+T细胞对自体和异体的CD4+T细胞增殖均有明显的抑制作用，并且随着效靶比浓度的降低而降低。在CD4+T/Treg比例为8：1、4：1、2：1、1：1时对自体CD4+T细胞的抑制率分别为9.65%、16.43%、28.75%、55.34%，对异体CD4+T细胞的抑制率分别为6.43%、14.72%、26.47%、50.25%，而且在各浓度梯度其抑制作用在自体和异体之间均无明显的差异。结论 我们采用rapamycin+CD3/CD28单抗标记的磁珠+IL-2在体外成功扩增了CD4+CD25+Treg，其扩增的倍数为89.4+20.53，具有免疫抑制功能，有望进一步应用于临床。     

Sirolimus-associated interstitial pneumonitis in solid organ transplant recipients

Sirolimus is a potent immunosuppressive agent used with increasing frequency in solid organ transplantation (SOT). However, it has been associated with rare but devastating pulmonary toxicity. We describe a case of pulmonary toxicity associated with the use of sirolimus in a 64-yr-old heart transplant recipient. We also review all reported cases of sirolimus-associated lung toxicity among SOT recipients in an effort to better understand the pathophysiology, risk factors, and outcomes of this rare but serious complication. A total of 64 cases have been reported since January 2000 including the present case. These consisted of 52 kidney, four lung, three liver, three heart, one heart-lung and one islet cell transplants. In most cases, patients presented with a constellation of symptoms consisting of fever, dyspnea, fatigue, cough, and occasionally hemoptysis. Although the risk factors for this association have not been clearly established, high dose, late exposure to the drug and male gender have been noticed among most. In almost all of the reported cases, sirolimus was added later in the course of immunosuppressive therapy, usually in an effort to attenuate the nephrotoxic effects of a previous regimen containing a calcineurin inhibitor. There were three deaths (4.8%) among 62 patients with known status at follow up; all deaths were among heart transplant recipients. Most patients (95%) resolved their clinical and radiographic findings with discontinuation or dose-reduction of the drug. Sirolimus-induced pulmonary toxicity is a rare but serious entity that should be considered in the differential diagnosis of a transplant recipient presenting with respiratory compromise. Dose-reduction or discontinuation of the drug can be life saving.