AKR1C3 Overexpression Mediates Methotrexate Resistance in Choriocarcinoma Cells

Background: Chemotherapy is typically used to treat choriocarcinoma, but a small proportion of tumors develop resistance to chemotherapy. Similarly, methotrexate (MTX) is a first-line chemotherapy used to treat choriocarcinoma; although ~30% of patients are drug-resistant for MTX mono-therapy. Thus, we sought to elucidate the mechanism of chemotherapeutic-resistance of MTX.Methods: RNA interference technology, colony formation, and MTT assays were used to investigate the role of aldo-keto reductase family 1, member C3 (AKR1C3) in MTX resistance in choriocarcinoma cells.Results: AKR1C3 expression was higher in JeG-3R cells compared to JeG-3 cells and targeted inhibition of AKR1C3 expression with shRNA suppresses growth of choriocarcinoma cells as measured by colony formation and MTT assays. Overexpression of AKR1C3 increased chemotherapeutic resistance in JeG-3 cells. Furthermore, AKR1C3 silencing increases sensitivity to MTX in JeG-3R choriocarcinoma cells. Increasing MTX sensitivity spears to be related to DNA damage induction by increased reactive oxygen species (ROS), apoptosis, and cell cycle arrest.Conclusions: Data show that AKR1C3 is critical to the development of methotrexate resistance in choriocarcinoma and suggest that AKR1C3 may potentially serve as a therapeutic marker for this disease.     

Clinical characteristics and risk factors for low dose methotrexate toxicity: A cohort of 28 patients

Objective

Low dose (10–25 mg/week) methotrexate is widely used for the management of systemic inflammatory diseases, and is considered to be relatively safe. Toxicity due to low dose MTX has been reported but is poorly characterized. We describe the clinical features, risk factors, and outcomes of low dose MTX toxicity in a large case series at our center.

Patients and methods

We conducted a retrospective case series of all adult (> 18 years) patients hospitalized at Sheba Medical Center, between 2005 and 2012 for low dose MTX toxicity.

Results

We identified 28 patients (age: 70.4 ± 13.7 years, range: 33–88; 20 (71%) females) hospitalized for low dose MTX toxicity. Indications for MTX therapy included: rheumatoid arthritis (39.2%), psoriasis ± arthritis (21.5%), polymyalgia rheumatica (10.8%) and other inflammatory conditions (28.5%). Pancytopenia was the most common manifestation of low dose MTX toxicity detected in 78.5% of the patients. Potential risk factors included acute renal failure, hypoalbuminemia, concurrent use of drugs known to interact with MTX, and dose errors. Serum MTX concentrations (n = 20, mean 0.04 ± 0.07 μg/mL range: 0–0.3) did not correlate with the degree of either neutropenia (r = − 0.36; p = 0.18) or thrombocytopenia (r = 0.44; p = 0.10). Seven (25%) patients died, all from pancytopenia followed by sepsis. Serum MTX concentrations did not differ between the patients who died from MTX toxicity (n = 6; mean: 0.05 ± 0.04 μg/mL) and those who survived the toxicity (n = 14 mean 0.04 ± 0.08; p = 0.45).

Conclusions

Low-dose MTX toxicity can be life threatening, mainly due to myelosuppression. There is no rationale for MTX therapeutic drug monitoring in the setting of low-dose toxicity.     

Methotrexate patient education: A quality improvement study

Objective . To determine patients' knowledge of the safe use and toxicity of methotrexate (MTX) and to define educational interventions implemented by a rheumatology nurse that improved patients' understanding of MTX therapy. Methods . One hundred eighty-three patients from a university-based rheumatology clinic who were taking MTX completed an initial knowledge questionnaire concerning the proper use and possible toxicity of MTX. Following completion, a nurse reviewed the correct answers with each patient and provided written information on MTX. One hundred thirty-eight of these patients completed a followup questionnaire at the next visit or by mail. The questionnaires were analyzed, and a total MTX knowledge score was calculated. Results . MTX knowledge improved significantly between questionnaires; mean total score (±SD) increased from 7.32 ± 3.99 to 10.23 ± 3.29 (P < 0.001). After accounting for a person's initial questionnaire score, the addition of a supplemental “MTX pocketcard” was associated with a higher score on the followup questionnaire (adjusted odds ratio [OR] = 2.37; 95% confidence interval [CI] 1.14, 4.95; P = 0.021). Patients over age 55 were 4 times more likely to have a poorer score compared with patients under age 45 (adjusted OR = 0.23; 95% CI 0.07, 0.73; P = 0.013). Conclusion . Knowledge of the toxicity and safe use of MTX was significantly improved by a patient education program utilizing a rheumatology nurse. Older individuals appear to be at higher risk for knowledge deficits. A supplemental MTX pocket-card proved to be a simple but beneficial addition to our MTX educational program.     

甲氨蝶呤皮下注射治疗应用与展望

【摘要】 研究表明,甲氨蝶呤皮下注射较口服途径可以获得更快、更好的吸收和更高的血药浓度以及较低的生物利用度变异度。在类风湿关节炎患者和银屑病患者,甲氨蝶呤皮下注射的疗效显著优于口服途径,并且胃肠道不良反应发生率更低。对于甲氨蝶呤口服疗效欠佳或者不耐受的患者,改为皮下注射后仍然可以获得较好的治疗反应。甲氨蝶呤皮下注射可以作为生物制剂使用前的治疗选择,可节省大量费用。总之,甲氨蝶呤皮下注射具有较好的临床应用前景。     

病理证实的原发性中枢神经系统淋巴瘤40例临床分析

目的探讨原发性中枢神经系统淋巴瘤（PCNSL）的临床与影像学特点，治疗与预后。方法对40例病理证实的PCNSL的临床与影像学表现、治疗和预后情况进行回顾性分析。结果PCNSL好发于50～60岁男性，急性或亚急性起病。肿瘤单发者占75．0％，多发者占25．0％，多位于脑脊液循环通路附近，即深部结构近脑室（57．5％）或靠近皮质的浅表部位（32．5％）。病理均为B细胞型，有特征性的“血管周围淋巴套”和“星空现象”。临床不具有特异性。头颅MRI多为T1低信号，T2高信号，界限清晰，水肿明显，显著均匀强化占67．6％，厚壁环形强化占13．5％。采用大剂量甲氨蝶呤（HDMTX）化疗者，存活率（75．0％）较未采用者（48．1％）高。平均生存时间14．8个月，复发后69．2％的患者存活不足6个月。结论PCNSL临床表现无特异性，确诊需靠穿刺活体组织检查。治疗首选HDMTX为基础的综合化疗，预后差。     

Low-Dose Cyclosporin A with Short-Term Methotrexate for Graft-versus-Host Disease Prophylaxis in Allogeneic Bone Marrow Transplantation from Human Leukocyte Antigen—Identical Siblings: A Prospective Phase II Study in Japanese Patients

We hypothesized that reducing the dosage of prophylaxis for graft-versus-host disease (GVHD) would reduce the risk of relapse and toxicity after bone marrow transplantation (BMT) from human leukocyte antigen (HLA)-identical siblings. In a prospective phase II trial, 21 patients with leukemia and myelodysplastic syndrome underwent BMT from HLA-identical siblings and received GVHD prophylaxis consisting of low-dose (1.5 mg/kg per day) cyclosporin A (CSP) with short-term methotrexate (MTX) treatment. This low-dose group was compared with a group of retrospective control patients (n = 22) who received a standard CSP dosage (3.0 mg/kg per day) and MTX. One patient died of transplantation-related causes within 100 days. The regimen-related toxicity was quite tolerable. Although acute GVHD of grades II to III was more frequent in the low-dose group (47.6%) than in the control group (22.7%), the increase in acute GVHD did not significantly contribute to morbidity or mortality. There were no differences between the groups in the incidence and severity of chronic GVHD. The probabilities of relapse and survival of the groups were similar according to the risk for relapse at the time of transplantation. A prospective randomized study is required to determine whether low-dose or standard-dose CSP in combination with MTX is optimal for Japanese patients who undergo allogeneic BMT from HLA-identical siblings.     

去除第11天甲氨蝶呤方案预防同胞相合供体造血干细胞移植后急性移植物抗宿主病

目的:探讨去除第11天甲氨蝶呤(methotrexate,MTX)急性移植物抗宿主病(acute graft-versus-host disease,a GVHD)预防方案在同胞相合供体异基因外周血造血干细胞移植(allogeneic peripheral blood stem cell transplantation,allo-PBSCT)中的疗效和安全性。方法 :回顾性分析2015年1月至2017年8月接受同胞相合alloPBSCT治疗的32例患者临床资料,GVHD预防方案为环孢素(Cs A)联合MTX 15 mg/m2第1天,10 mg/m2第3、第6天。结果:中位随访时间14(7,30)个月,32例患者移植后全部造血重建。可评估患者中性粒细胞和血小板中位植入时间分别为13(11,18)d和16.5(14,36)d。a GVHD总体发生率为40.6%,Ⅱ~Ⅳ度a GVHD为25.0%。慢性GVHD(c GVHD)总体发生率为48.3%,其中轻度13.8%,中度20.7%,重度17.2%。移植后30 d内口腔黏膜炎的发生率为40.6%,Ⅲ~Ⅳ度口腔黏膜炎的发生率为18.8%。相比采用标准MTX预防方案的研究结果 ,重度口腔黏膜炎发生率显著降低(P     

Clinical outcomes of patients treated for cervical pregnancy with or without methotrexate

The objective of this study is to describe the clinical outcomes of patients treated for cervical pregnancy with or without methotrexate (MTX) and to evaluate the effects of MTX in the treatment of cervical pregnancy. Between January 1993 and February 2000, 31 patients were diagnosed with cervical pregnancy. Twenty-two patients were treated with MTX chemotherapy and nine patients were treated with surgical procedures without MTX treatment. In the non-MTX treatment group, three patients underwent total abdominal hysterectomy, five required adjuvant procedures to control the bleeding during dilatation and curettage (D&C) and only one patient was treated with a simple D&C. In the MTX treatment group, fourteen (63.6%) patients were treated with only MTX and eight (36.4%) cases underwent concomitant procedures (simple curettage, curettage and Foley catheter tamponade, cervical cerclage, ligation of the descending branches of uterine arteries, or ligation of hypogastric arteries). The uterus was preserved in all cases and three women delivered healthy babies in their subsequent pregnancy. In conclusion, early diagnosis, appropriate MTX regimen in combination of necessary adjuvant conservative procedures could contribute to successful treatment with preservation of the uterus and future reproductive ability.     

云克和甲氨蝶呤联合治疗类风湿关节炎的疗效及安全性观察

目的　观察云克 (99锝 亚甲基二膦酸盐 )和甲氨蝶呤联合治疗类风湿关节炎 (RA)的疗效及其安全性。方法　选取类风湿关节活动期患者 90例。随机分为两组 ,其中治疗组 (云克 甲氨蝶呤 ) 4 5例 ,对照组 (甲氨蝶呤 柳氮磺吡啶 氯喹 ) 4 5例 ,两组均接受相应药物治疗 12周。观察临床症状改善、实验室指标变化及不良反应。结果　 4周时治疗组显效率为 6 2 % ,有效率为 2 2 % ,对照组分别为 2 7%、2 0 % ,两组差异有显著性意义 (P <0 0 0 1)。治疗组在其他各项临床指标的改善值上均高于对照组 (P <0 0 5 ) ;12周时治疗组显效率为 6 0 % ,有效率为 36 % ,对照组分别 4 4 %、16 % ,两组差异无显著性意义 (P >0 0 5 )。在其他各项临床指标的改善值上两组比较差异无显著性意义 (P >0 0 5 )。不良反应两组总体比较差异无显著性意义。结论　云克和甲氨蝶呤联合治疗类风湿关节炎起效快 ,副作用少 ,12周时疗效与对照组相当。     

实时荧光定量PCR法检测MTX对映体耐药A549细胞株及白血病患者骨髓细胞中FPGS mRNA的表达

目的 建立实时荧光定量PCR检测肿瘤细胞中FPGS mRNA表达的方法,研究MTX对映体[L-(+)-MTX和D-(-)-MTX]耐药细胞株中FPGS的基因表达差异,并用于观察白血病患者MTX治疗耐药前后FPGS mRNA表达水平的变化.方法 用SYBR Green Ⅰ为荧光染料,以β-actin作参照,建立检测FPGS mRNA的实时荧光定量PCR方法.根据Ct值、标准曲线相关系数、斜率、重复性曲线、熔解曲线、扩增效率曲线等进行方法学评价.并用该方法检测MTX对映体耐药细胞株及应用MTX耐药的14例白血病患者骨髓细胞中FPGS mRNA的表达.结果 建立的标准曲线Ct值与模板浓度有良好的线性关系,FPGS和β-actin标准曲线相关系数分别为0.996 8和0.998 7,斜率分别为-3.595和-3.740,批内CV为1.27%～2.95%,批间CV为3.82%;熔解曲线均呈单个特异峰,扩增效率相似(斜率为0.021 7);L-(+)-MTX耐药细胞和D-(-)-MTX耐药细胞中FPGS mRNA相对含量分别为(3.51±0.66)和(0.16±0.01),A549亲本细胞(S)中FPGS mRNA相对含量为(1.00±0.31),差异有统计学意义(F=64.45,P＜0.01);L-(+)-MTX耐药细胞和D-(-)-MTX耐药细胞间FPGS mRNA相对含量差异有统计学意义(q=9.29,P＜0.01).白血病患者应用MTX治疗耐药后,FPGS mRNA表达水平为(0.35±0.04),用药前为(1.00±0.44),差异有统计学意义(t=8.83,P＜0.01).结论 建立的实时荧光定量PCR检测FPGS mRNA的方法重复性好、特异度高,可用于FPGSmRNA含量分析.MTX诱导耐药后细胞株及白血病患者骨髓细胞中FPGS mRNA表达发生了变化,MTX 2种对映体形式在细胞耐药机制中可能发挥了不同的作用.